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12th France‐Japan DDS Symposium 1

12th

France-Japan Drug Delivery Systems

Symposium

“Recent Achievements and Further

Challenges in Drug Delivery Research”

October 9-12, 2016

Abbaye des Vaux de Cernay, France


PREFACE

The 12th France‐Japan DDS symposium will be held in the Vaux de Cernay, 50 Km south of Paris. In addition of being a wonderful site, the ancient abbey turned into a welcoming place for seminar and symposium. This area is also a cluster of excellence for medical sciences and several renowned research centers and companies are located quite close to this location. This environment together with the charm of the surroundings should be an occasion of inspiration to this joint meeting.

French and Japanese people are sharing similar interest for culture, art, erudition, cooking and science. But even if French and Japanese cultures and food may be very different, their scientific language and accomplishments are the same, especially in the drug delivery field. This common interest for research in a major pharmaceutical field explains the vitality and longevity of this French‐Japanese DDS symposium, which takes place for the 12th time during the last 24 years period. The organization of such an event is only possible, thanks to the generosity of both French and Japanese sponsors which are here warmly acknowledged.

The very appealing program of the symposium covers all the different aspects that has been involved during the recent year in the development of novel DDS: material chemistry, physical chemistry, biological applications and translation into clinics of major discoveries. Our symposium is a perfect example of the tremendous evolution of this scientific field.

We do hope you will enjoy the symposium. While not very far from Paris, the capital, the Vaux de Cernay is a perfect place for increased scientific interactions and connection.

We wish you a marvelous stay!

Pr. Patrick Couvreur Pr. Elias Fattal University of Paris‐Sud


ACKNOWLEDGEMENTS The organizers wish to thank warmly the following Institutions and Companies for their support:

GATTEFOSSE

SANTEN FRANCE

ROQUETTE

CAPSUGEL


IPSEN

ASTELLAS PHARMA INC.

THE NAGAI FOUNDATION TOKYO


CONFERENCE PROGRAM

Sunday October 9, 2016

16h30 Pick‐up participants at Hyatt CDG airport, Terminal 2, Terminal 1 19h00 Arrival at Vaux de Cernay 20h00 Welcome Dinner

Monday October 10, 2016 8h15 Welcome address Hideyoshi HARASHIMA and Elias FATTAL Hokkaido University, Japan and Paris‐Sud University, France Chairs: MIGNET Nathalie

KOGURE Kentaro 8h45 ISHIDA Tatsuhiro – Tokushima University, Japan

Development of a RNAi‐Based Anticancer Drug 9h15 BARTHELEMY Philippe – University of Bordeaux, France

Nucleic Acid based Supramolecular Systems: a Route to Hydrogels for Regenerative Medicine 9h45 HARASHIMA Hideyoshi ‐ Hokkaido University, Japan

Multifunctional Envelope‐Type Nano Device for Gene Delivery: Concept and Application for Nanomedicine

10h15 FATTAL Elias ‐ Paris‐Sud University, France Targeted and Local Delivery of Nucleic Acids

10h45 Break

Chairs: RICHARD Joël MARUYAMA Kazuo 11h15 OKADA Hiroaki – Okada DDS Research Institute, Japan

Cytoplasm‐Responsive Delivery Systems for siRNA using TAT‐Like Peptide Nanomicelles 11h45 SASAKI Shigeki – Kyushu University, Japan

The Pseudo‐Cytidine Derivative Enables the Formation of Triplex DNA with Multiple CG Inversion Sites to Inhibit Transcription

12h15 KOGURE Kentaro – University of Tokushima, Japan Effective Cytoplasmic Delivery of Macromolecules by Faint Electric Treatment

12h45 Lunch

Chairs: JANNIN Vincent IMAI Teruko 14h15 BENAMEUR Hassan – CAPSUGEL, France enTRinsicTM Drug Delievry Technology for Live Biotherapeutics (Microbiomes) 14h45 KIKUCHI Hiroshi – Tsukuba Research Laboratories, Eisai Co., Ltd., Japan

Start of Commission for Species Difference Problem on Formulation Design 15h15 RICHARD Joël – IPSEN, France

Delivery of Peptides by Non‐Invasive Routes : Focus on Successful Oral Technologies Progressing in the Clinic and Future Challenges

15h45 KONDO Hiromu – Pharmaceutical Research and Technology Labs, Astellas Pharma Inc., Japan Role of exploratory clinical study for oral DDS formulation development


16h15 Break Chairs: BENAMEUR Hassan KATAOKA Kazunori 16h45 JANNIN Vincent – GATTEFOSSE S.A.S., France

Lipid‐Based Formulations for Enhanced Oral Bioavailability of BCS Class II Drugs 17h15 LEFEVRE Philippe – ROQUETTE, France

Comprehension of mannitol behavior in high speed rotary Press tableting using simulator 17h45 MARUYAMA Kazuo – Teikyo University, Japan

Development of a New Lipid Bubble for Ultrasound Theranostics 18h15 MIGNET Nathalie – Paris Descartes University, France

Conception of contrast Agents or nanotherapeutics with the Help of Imaging Modalities

20h00 Dinner

Tuesday October 11, 2016 Chairs : LECOMMANDOUX Sébastien HARASHIMA Hideyoshi 8h45 COUVREUR Patrick ‐ Paris‐Sud University, France

To Pass or not to Pass the Blood Brain Barrier 9h15 KATAOKA Kazunori – Innovation Center of nanoMedicine and the University of Tokyo, Japan

Self‐assembled Supramolecular nanosystems for Smart Targeted Therapy of Intractable Diseases 9h45 GAZEAU Florence ‐ Paris Diderot University, France

Modulation of the tumor microenvironment by nanotherapy 10h15 AKIYOSHI Kazunari – Kyoto University, Japan

Development of new bio‐nanotransporters for biologics DDS

10h45 Break

Chairs : ISHIDA Tatsuhiro SIEPMANN Juergen 11h15 PENG Ling – Aix‐Marseille University, France

Charm of Dendrimer nanotechnology in Cancer Therapy 11h45 YAMASHITA Fumiyoshi ‐ Kyoto University, Japan

Use of Informatics Technologies for the Prediction of Pharmacokinetic Behaviors 12h15 IMAI Teruko – Kumamoto University, Japan

Catalytic Properties of Esterase in Several Organs and Design of Prodrug and Soft Drug

12h45 Lunch

15h00 Social Programme: Visit at the Château de Breteuil 20h00 Gourmet Dinner


Wednesday October 12, 2016 Chairs : COUVREUR Patrick AKASHI Mitsuru 8h45 LECOMMANDOUX Sébastien

Ecole Nationale Supérieure de Chimie et de Physique de Bordeaux, France Bioactive glycopolypeptide self‐assembled biohybrid nanomaterials

9h15 TSUKAMOTO Ikuko & KONISHI Ryoji – Kagawa University, Japan Promising Capabilities of an Adenosine Analogue, COA‐CI

9h45 DOURLAT Jennifer – SANTEN FRANCE Rationale for the use of cationic emulsions in ophthalmology : from bench to bedside 10h15 OZEKI Tetsuya – Nagoya City University, Japan

Formulation Design of Composite Nano‐Particles for Inhalation by Using a Two‐Solution Mixing type Spray Nozzle

10h45 Break

Chairs: PENG Ling OZEKI Tetsuya 11h15 AKASHI Mitsuru – Osaka University, Japan

Development of Vascularized 3D Tissue Models using Layer‐by‐Layer Technique and Application for DDS Research

11h45 SIEPMANN Juergen – University of Lille, France Simulation of controlled drug delivery systems in vitro and in vivo

12h15 YAMAMOTO Hiromitsu – Aichi Gakuin University, Japan Design of Polymeric Nanoparticle and Micelle for Treatment of Biofilm Infection Disease

12h45 Lunch

14h15 BOISSIERE Cédric –Pierre & Marie Curie University, France Title to be announced 14h45 MAKINO Yuji – Musashino University, Japan

X Ray Micro CT Imaging of Penetrated Microneedles 15h30 Departure for CDG by Bus


ABSTRACTS


A DEVELOPMENT OF RNAi‐BASED ANTICANCER DRUG

Tatsuhiro ISHIDA, Ph.D.

Institute of Biomedical Sciences, Tokushima University

KEY WORDS: RNAi, shRNA, liposomal DDS, cancer treatment ABSTRACT : During about two decades, a number of therapeutic drugs focused to RNA interference (RNAi) have been developed, however these drugs have been still unsuccessful due to various problems such as targeted disease, drug delivery system, dosing route, stability in the body and so on. To resolve these problems, we focused to developing new drug delivery system using cationic liposome and short‐hairpin RNAi molecule for thymidylate synthase (TS shRNA), an important rate‐limiting enzyme of DNA biosynthesis in cancer. To avoid a rapid degradation in the blood and immunological incidence of TS shRNA conjugated with liposome (named as DFP‐10825), intraperitoneal and/or intrathoracic administration of DFP‐10825 were applied to evaluate the antitumor activity on disseminated ascetic ovarian, gastric and pancreatic cancer models and/or malignant mesothelioma xenografts implanted into intrathoracic cavity. Locally injected DFP‐10825 resulted in potent antitumor activity and prolonged survival on these cancer models with relative long‐time retention of DFP‐10825 in the cavity. In addition, the combination of DFP‐10825 (i.p.) and paclitaxel (i.p.), down‐regulator of TS, or pemetrexed (i.p.), one of TS inhibitors, showed augmented antitumor efficacy and further prolonged survival in those cancer models. DFP‐10825 is a stable freeze‐dried preparation and capable of being prepared in a large amount at once. Our novel trial for RNAi‐based anticancer therapy will have a clinical benefit to treat patients with disseminated peritoneal cancers. We cooperate with a biotechnology venture company, Delta‐Fly Pharma and plan to enter a clinical trial within 1.5 years. REFERENCES : 1) Abu Lila AS, Kato C, Fukushima M, Huang CL, Wada H, Ishida T., Downregulation of thymidylate

synthase by RNAi molecules enhances the antitumor effect of pemetrexed in an orthotopic malignant mesothelioma xenograft mouse model. Int J Oncol, 48, 1399‐1407 (2016)

2) Ando H, Kobayashi S, Abu Lila AS, Essam Eldin N, Kato C, Shimizu T, Ukawa M, Kawazoe K, Ishida T,

Advanced therapeutic approach for the treatment of malignant pleural mesothelioma via the

intrapleural administration of liposomal pemetrexed. J Control Release, 220, 29‐36 (2015)

3) Hashimoto Y, Abu Lila AS, Shimizu T, Ishida T, Kiwada H, B cell‐intrinsic toll‐like receptor 7 is

responsible for the enhanced anti‐PEG IgM production following injection of siRNA‐containing

PEGylated lipoplex in mice. J Control Release, 184, 1‐8 (2014)

4) Kadota K, Huang CL, Liu D, Yokomise H, Haba R, Wada H., Combined therapy with a thymidylate

synthase‐inhibiting vector and S‐1 has effective antitumor activity against 5‐FU‐resistant tumors. Int J

Oncol, 38, 355‐363 (2011)


NUCLEIC ACID BASED SUPRAMOLECULAR SYSTEMS: A ROUTE TO HYDROGELS FOR REGENERATIVE MEDICINE

Philippe BARTHELEMY1,2,3

1University of Bordeaux, ARNA laboratory, F‐33000 Bordeaux, France.

2INSERM, U1212, ARNA laboratory

3UMR CNRS 5320

KEY WORDS : Amphiphiles, Biocompatible scaffolds, Low Molecular Weight Gels (LMWG), stem cells, Tissue engineering, supramolecular chemistry.

ABSTRACT : The combination of nucleic acids chemistry (e.g., nucleoside, nucleotides, oligonucleotides) with supramolecular principles provides an efficient and powerful approach to prepare well‐defined systems with tunable physico‐chemical properties and functions. We are currently developing biocompatible gels resulting from the self‐assembly of nucleic acids derivatives for i) drug delivery applications, and ii) tissue engineering. This communication will present novel “smart” nucleic acid based nanosystems derivatives developed in our lab [1]. REFERENCES :

1) K. Oumzil, M. A. Ramin, C. Lorenzato, A. Hemadou, J. Laroche, M. J. Jacobin‐Valat, S. Mornet, C.‐E. Roy, T. Kauss, K. Gaudin, G. Clofent‐Sanchez, and P. Barthélémy (2016) Bioconjugate Chemistry 27 (3), pp. 569‐575

2) J. A. Kaplan, P. Barthélémy and M. W. Grinstaff, Chem. Commun., (2016), 52, pp. 5860‐5863 3) Latxague, L., Ramin, M.A., Appavoo, A., Berto, P., Maisani, M., Ehret, C., Chassande, O., and

Barthélémy, P. (2015) Angewandte Chemie, 54 (15), pp. 4517‐4521. 4) Patwa A. ; Labille, J. ; Bottero JY. ; Thiéry, A.; and Barthélémy, P. (2015) Chem. Commun. 51 (13), pp.

2547‐2550 5) Oumzil, K. Benizri, S. Tonelli, G. Staedel, C. Appavoo, A. Chaffanet, M. Navailles L. and Barthélémy

P. (2015) ChemMedChem 10 (11), pp. 1797‐1801 6) Luvino, D. ; Khiati, S. ; Oumzil, K. ; Rocchi, P. ; Camplo, M. ; and Barthélémy P. (2013) J. Control.

Release, 172, 954–961 7) Khiati, S., Luvino, D., Oumzil, K., Chauffert, B., Camplo, M. and Barthélémy P. (2011) ACS Nano, 5

(11), pp 8649–8655. 8) Patwa, A., Gissot, A., Bestel, I. and Barthélémy, P. (2011) Chem. Soc. Reviews, 40, 5844–5854

ACKNOWLEDGEMENTS : The authors acknowledge financial support from the French National Agency (ANR) and the LABEX AMADEUS.


MULTIFUNCTIONAL ENVELOPE‐TYPE NANO DEVICE FOR GENE DELIVERY: CONCEPT AND APPLICATION FOR NANOMEDICINE

Hideyoshi HARASHIMA

Faculty of Pharmaceutical Sciences, Hokkaido University Sapporo City, Hokkaido, JAPAN

E‐mail: [email protected]

KEY WORDS : MEND, targeting, pH‐responsive cationic lipid, mitochondria

Programmed Packaging: Recently, we developed a multifunctional envelope‐type nano device (MEND) as a novel non‐viral gene delivery system based on a new packaging concept termed “Programmed Packaging”. Programmed Packaging was proposed to develop a rational non‐viral gene delivery system equipped with various functional devices, including ligands for specific receptors, pH‐sensitive fusogenic peptides for endosomal escape, and a nuclear localization signal (NLS) for enhanced nuclear delivery, to overcome several barriers in the process of gene delivery to the nucleus of target cells [1‐3].

R8‐MEND: Octaarginine (R8) is known as a cell penetrating peptide and we have developed R8‐MEND for gene delivery system. During the course of development, R8‐MEND has been applied to bladder cancer vaccine by encapsulating BCG‐CWS (cell wall skeleton). In a preliminary trial, BCG‐R8MEND can exerted significant antitumor activity in the rat bladder cancer model. However, it was found that BCG‐CWS is difficult to encapsulate into R8‐MEND due to a serious problem in formulation, we had to find a new packaging method of BCG‐CWS. Finally, we succeeded to establish a new formulation method as named LEEL (liposomes evaporated via the emulsified lipid) method, which can make a homogenous nanoparticles in diameter and can exert antitumor activity in mouse tumor model [4].

In vivo application of MEND: In order to apply MEND via a systemic administration, it is important to control surface of MEND. Here we introduced pH‐responsive cationic lipid to control biodistribution as well as intracellular trafficking. A newly designed YSK05 can respond to endosomal pH to induce efficient escape from

endosome while neutral surface charge in blood circulation. The YSK‐MEND can induce gene silencing in hepatocytes at a dose of 0.06 mg/kg. YSK‐MEND has been applied to HCV therapy in collaboration with Dr. Kohara and we have successfully sent siRNA to chimeric mice carrying human hepatocytes infected with HCV and produced gene silencing as well as antivirus effects [5]. As an active targeting to adipose vasculature, we have developed Prohibitin Targeted Nano Particles (PTNP) using a peptide ligand (KGGRAKD). PTNP can be recognized by the prohibitin which is up‐regulated on the surface of adipose endothelial cells and efficiently internalized and released encapsulated Cytochrom C to induce apoptosis. As a result, a remarkable antiobesic effect was induced without producing a significant toxicity [6]. This strategy will imply a new strategy for antiobesic drugs.

MITO‐Porter: A variety of human diseases, including neurodegenerative disorders, diabetes, cancer and inherited mitochondrial disease, are associated with mitochondrial dysfunction. Therefore, this organelle is a promising therapeutic drug target in mitochondrial therapy. To achieve such an innovative therapy, it is necessary to deliver therapeutic agents into mitochondria in living cells. Recently, we proposed a new concept of mitochondrial delivery, a MITO‐Porter, a liposome‐based carrier system that introduces macromolecular cargos into mitochondria via membrane fusions. We screened for fusogenic lipid compositions in isolated mitochondria using a FRET principle. Highly fusogenic lipid compositions were identified for MITO‐Porter such as DOPE/SM/STR‐R8 or DOPE/PA/STR‐R8. To improve MITO‐Porter for higher performance, MITO‐Porter was coated with endosome‐fusogenic envelopes and named as Dual Function (DF)‐MITO‐Porter [7‐8]. DF‐MITO‐Porter could efficiently delivered macromolecules to the matrix of mitochondria in living cells. Mitochondrial gene therapy with MITO‐Porter will also be discussed.


TARGETED AND LOCAL DELIVERY OF NUCLEIC ACIDS

Elias FATTAL, Juliette VERGNAUD, Nicolas TSAPIS, Hervé HILLAIREAU

University Paris‐Sud, Institut Galien Paris‐Sud, School of Pharmacy, Châtenay‐Malabry, France

KEY WORDS : Liposomes, aptamer, dendrimers, siRNA, lungs INTRODUCTION Since its discovery, the mechanism of RNA interference has become the method of choice for silencing specific gene expression in mammalian cells. This possibility of controlling disease‐associated genes makes RNA interference an attractive choice for therapeutics in cancer, inflammatory and autoimmune diseases as well as dominant genetic disorders and viral infections (1). Despite the great potential applications for systemic and local delivery, small interfering RNAs (siRNAs) still face pharmacokinetic and cell penetration limitations. They are also susceptible to degradation by nucleases in biological fluids. Our strategy is therefore to design smart carriers for systemic and local delivery of siRNAs in order to circumvent these drawbacks (1). We have first developed targeted delivery systems for systemic delivery which will be illustrated by the design of anti‐CD44 aptamer‐functionalized liposomes. We have secondly started to study the feasibility of lung delivery of siRNA using dendrimers. TARGETED GENE SILENCING IN CANCER CELLS USING ANTI‐CD44 APTAMER‐FUNCTIONALIZED LIPOSOMES CD44, a transmembrane glycoprotein that exists in different isoforms, plays an important role on cell – cell / cell – matrix interaction, cell adhesion and migration and signal transduction from the extracellular to the intracellular compartment. Moreover, it was clearly shown that many cancer types over‐express this receptor which is also known to be a major biomarker of cancer stem cells (2). Aptamer are nucleic acids that adopt a 3D conformation which allows them to bind as efficiently as antibodies (without the drawback of the latest) to receptor proteins. They are designed by the SELEX technique for systematic evolution of ligands by exponential enrichment. Apt1, a 2'‐F‐pyrimidine‐containing RNA aptamer, was selected against CD44 (3). Apt1 was successfully functionalized to the surface of PEGylated liposomes (Lip) using a thiol‐maleimide click reaction, resulting in ~140 nm liposomes (4) and the whole displayed a high affinity to CD44 protein as well as a greater internalization within CD44 positive cells. The system is supposed to get internalized by cancer cells through CD44‐mediated endocytosis and release liposomal content in the cytoplasm. siRNA was efficiently loaded in such liposomes after complexation with protamine. The efficacy of Apt1‐Lip‐siRNA liposomes in knocking down Luc2 expression was tested in MDA‐MB‐231‐Luc2‐GFP breast cancer cells in vitro and showed higher inhibition to Luc2 expression compared to Lip‐siRNA alone and to mere protamine/siRNA polyplexes. Finally, an orthotopic xenograft model of human breast cancer was developed by implanting the MDA‐MB‐231‐Luc2‐GFP cells into the mammary fat pad of female nude mice. After 3 daily intravenous injections, a significant knockdown of Luc expression in tumors was observed for liposomes loaded with anti‐Luc siRNA. Noteworthy, while non‐functionnalized PEGylated liposomes did silence Luc in tumors, Apt1‐conjuated liposomes entailed the more intense and prolonged response. siRNA THERAPY FOR TREATMENT OF LUNG INFLAMMATION USING PHOSPHOROUS CATIONIC DENDRIMERS AS TRANSFECTION AGENTS Chronic Obstructive Pulmonary Disease (COPD) is characterized by a persistent inflammation in the lungs and is currently the third leading cause of death globally with no cure available. Current medicinal treatment for COPD includes anti‐inflammatory drugs and bronchodilators, which are relatively unspecific in their action and are administered systemically resulting in increased drug exposure and undesirable side effects. A siRNA‐based treatment may provide a more specific therapy in which the inflammatory pathways occurring in COPD could be targeted. Of the many cytokines involved in the COPD inflammation process, TNF‐α is believed to play a central role in the recruitment of inflammatory cells and soliciting other cytokines. It can thus be considered as a good target for a siRNA‐based therapy against COPD. Two phosphorus‐based dendrimers with either pyrrolidinium or morpholinium groups as terminal amines were synthesized and characterized. Their performance as transfection agents for lung delivery of siRNA


directed against TNF‐α for the treatment of lung inflammation was assessed in vitro in cell cultures and in vivo in a murine model of acute lung inflammation. The dendrimers with pyrrolidinium and morpholinium surface groups, exhibited very different characteristics and performances when complexed with siRNA. Dendrimer‐siRNA nanocomplexes (dendriplexes) containing pyrrolidinium surface groups demonstrated better complexation, higher cell uptake and better transfection efficacy in cells The better performance of pyrrolidium dendriplexes can be explained by higher pKa leading to a stronger siRNA complexation and protection and a better cell uptake. The better efficacy was confirmed in vivo for pyrrolidinium dendriplexes with up to 80% TNF‐α inhibition in a dose dependent manner after direct lung delivery. CONCLUSIONS In this work, we demonstrate a successful conjugation of anti‐CD44 aptamer to the surface of liposomes. Such liposomes show an enhanced affinity to the CD44 protein and CD44‐expressing cancer cells. Finally, these liposomes loaded with an anti‐Luc siRNA achieve an efficient gene silencing in vivo to orthotopic breast tumors, showing a promising potency of Apt1 decoration for selective gene delivery nanocarriers. We have also demonstrated that local delivery of siRNA using dendrimers can after inhalation target alveolar macrophages and block the expression of inflammatory cytokines. REFERENCES 1. Fattal E, Barratt G. Nanotechnologies and controlled release systems for the delivery of antisense oligonucleotides and small interfering RNA. Br J Pharmacol. 2009; 157(2):179‐194. 2. Leite Nascimento et al. Lipid based nanosystems for CD44 targeting in cancer treatment: recent significant advances, ongoing challenges and unmet needs. Nanomedicine 2016; 11(14):1865‐1887. 3. Ababneh et al., In vitro selection of modified RNA aptamers against CD44 cancer stem cell marker. Nucleic Acids Ther 2013; 23(6):401‐407. 4. Alshaer et al., Functionalizing Liposomes with anti‐CD44 Aptamer for Selective Targeting of Cancer Cells. Bioconjugate Chem. 2015; 26(7);1307‐1313.


CYTOPLASM‐RESPONSIVE DELIVERY SYSTEMS FOR siRNA USING TAT‐LIKE PEPTIDE NANOMICELLES

Hiroaki OKADA

Okada DDS Research Institute, Inc.

Tokyo University of Pharmacy & Life Sciences

KEY WORDS : Cytoplasm‐responsive nanocarriers, Gene transfection, Cell‐penetrating peptide, Local and systemic delivery, siRNA therapy, Anti‐tumor effects

1. Introduction. On 21 June, 2016, the NIH’s Recombinant DNA Research Advisory Committee (RAC) approved a proposal of the University of Pennsylvania to use CRISPR‐Cas9 system to help augment cancer therapies that rely on enlisting a patient’s T cells by knocking out PD‐1. This first trial in people could begin as early as end of the year. Among the new and excellent biologic treatments, gene therapy (Glybera®, AAV vector, UniQure) and cell therapy using bone marrow stem cells (Prochymal®, Osiris Therapeutics) have been the world’s first to receive approval in 2012. Futhermore, in 2015, FDA approved ImlygicTM (Amgen) as first oncolytic viral therapy in the USA, which is an engineered herpesvirus (GM‐CSF, not prolified in nomal cells) infecting tumour cells to destroy by stimulating the immune system to direct an attack against malignant cells in the body. On 27 May, 2016, StrimvelisTM (GSK) (autologous CD34+ cells transduced to express adenosine deaminase), the first ex‐vivo stem cell gene therapy to treat patients with ADA‐SCID (Severe Combined Immunodeficiency due to ADA deficiency) by only administered once, was excitingly approved in the European. This is the start of a new chapter in the treatment of genetic diseases. New biomaterials, nucleotides, oligonucleotides and cells are expected to be developed using an advanced drug delivery system (DDS) techniques. Herein, I have described a none‐viral vector, cytoplasm‐responsive nanomicelles conjugated with artificial cell‐penetrating peptides (CPP), for siRNA. 2. Cytoplasm‐responsive nanocarriers. We used a novel drug discovery approach using nucleotides and prepared formulations, including vaginal DNA vaccine [1] and siRNA medicines by depot formulations of anti‐VEGF [2], anti‐cFLIP, anti‐RAF1, and anti‐Int6, mucosal preparations of anti‐NF‐�B for atopic dermatitis [3], asthma, and hay fever, and local or systemic injectable preparation using CPPs [4‐7]. In this study, we have described the cytosol‐responsive nanomicelles of biodegradable block polymers conjugated with CPPs for systemic injection of siRNA to treat a cancer. Modified Tat peptide and artificial CPP of CH2R4H2C as a transcellular carrier of siRNA were synthesized. The guanidine of Arg (R) triggers endocytosis of the micelles, His (H) enhances the endosomal escape by a protom sponge effect, and Cys (C) rigidly condenses siRNA by disulfide bonding as well as ionic interaction with Arg. Because the concentration of glutathione (GSH) is low (0.5–10 �M) in the blood and high (0.5–10 mM) in the cytoplasm and nucleus, disulfide‐linked complexes of siRNA with cationic nanomicelles are very stable in the blood, but easily cleaved within cells, rapidly releasing the nucleotides (Fig. 1A). A new cytoplasm responsive gene carrier, stearoyl‐CH2R4H2C (OK‐102), can form rigid coplexes with nucleotides by the disulfide linkages, and OK‐102/siVEGF (100 nm �) showed highly efficient cellular endosomal uptake and sequence‐specific silencing effects of VEGF secretion in S‐180 sarcoma cells, exerting strong suppresion of tumor growth by intratumor injection. To avoid the RES uptake, we synthesized amphiphilic block copolymers of MPEG and poly(�‐caprolactone) conjugated with CPP analog, MPEG‐PCL‐CH2R4H2C (OK‐103), and determined their transfection ability and antitumor activity. Results of the SYBR Green exclusion assay indicated the OK‐103/siVEGF complexes can be effectively condensed and that the carrier has suitably high compaction ability to provide a physical shield for siRNA enough to protect them from emzyme attack. These complexes significantly suppressed VEGF secretion from S‐180 cells and showed no any cytotoxicity. Thus, we observed strong regression of tumor growth in tumor‐bearing mice by systemic administration (Fig. 1B).


3. Conclusion. STR‐CH2R4H2C (OK‐102) was used to promote the delivery of siRNA and pDNA for suppressing tumor by intratumor injection, treating atopic dermatitis by dermal application, asthma by intratracheal inhalation (not publised) and DNA vaccine via needleless injector. A novel diblock‐copolymer conjugated with a functional CPP, MPEG‐PCL‐CH2R4H2C (OK‐103), safely promotes the delivery of siRNA into tumor cells after systemic administration. REFERENCES :

1) Kanazawa T, Tamura T, Okada H et al. Needle‐free intravaginal DNA vaccination using a stearoyl oligopeptide carrier promotes local gene expression and immune responses. Int J Pharm, 447, 70‐74 (2013)

2) Murata N, Takashima Y, Toyoshima K, Yamamoto M, Okada H. Anti‐tumor effects of anti‐VEGF siRNA encapsulated with PLGA microspheres in mice. J Control Release, 126, 246‐254 (2008)

3) Uchida T, Kanazawa T, Okada H, et al. Therapeutic effects on atopic dermatitis by anti‐RelA siRNA combined with functional peptides, Tat and AT1002. J Pharmacol Exp Ther, 338, 443‐450 (2011)

4) Tanaka K, Kanazawa T, Ogawa T, Okada H. Disulfide crosslinked stearoyl carrier peptides containing arginine and histidine enhance siRNA uptake and gene silencing. Int J Pharm, 398, 219‐224 (2010)

5) Tanaka K, Kanazawa T, Okada H, et al. Cytoplasm‐responsive nanocarrieres conjugated with a functional cell‐penetrating peptide for systemic siRNA delivery. Int J Pharm, 455, 40‐47 (2013)

6) Okada H, Ogawa T, Tanaka K, Kanazawa T, Takashima Y, Cytoplasm‐responsive delivery systems for siRNA using cell‐penetrating peptide nanomicelles. J Drug Del Sci Tech, 24, 3‐11 (2014)

7) Okada H, Targeted siRNA therapy using cytoplasm‐responsive nanocarriers and cell‐penetrating peptides. J Pharm Inv, 44, 505‐516 (2014)

ACKNOWLEDGEMENTS : We thank Dr. Tsunehiko Fukuda (Nagahama Institute of Bio‐Science and Technology), and Mr. Takaya Ogawa, Dr. Ko Tanaka, Dr. Takanori Kanazawa, Dr. Yuuki Takashima, and many graduate students (School of Pharmacy, Tokyo University of Pharmacy and Life Sciences) for their excellent collaboration.


THE PSEUDO‐CYTIDINE DERIVATIVE ENABLES THE FORMATION OF TRIPLEX DNA WITH MULTIPLE CG INVERSION SITES TO INHIBIT TRANSCRIPTION

Hidenori OKAMURA, Yosuke TANIGUCHI, Shigeki SASAKI

GRADUATE SCHOOL OF PHARMACEUTICAL SCIENCES

KYUSHU UNIVERSITY

KEY WORDS : TRIPLEX DNA • PSEUDOCYTIDINE • INVERSION SITE • ANTIGENE •HTERT GENE ABSTRACT : The sequence‐specific triplex formation against duplex DNA offers a potential basis for genome targeting technology. In an antiparallel triplex DNA, the sequence‐specificity is established via the formation of specific base triplets (G‐GC, A‐AT and T‐AT) between a triplex forming oligonucleotide (TFO) and a duplex DNA, however, there is no natural nucleoside which can selectively recognize the inverted CG and TA base pair. Therefore, the recognition of the CG and TA inversion sites to form a stable triplex DNA has been a long‐standing goal for the triplex forming technology.

In order to contribute this field, we first developed W‐shaped nucleoside analogues (WNA), bicyclic nucleoside analogs having a phenyl ring as a stacking part and a nucleobase as a recognition unit.1‐7 The WNA analogs exhibited useful property for stabilizing triplexes having the TA inversion sites in an antiparallel triplex geometry. We incorporated WNA‐�T into the TFOs with the targeting sequences of the bcl‐2 or survivin oncogene, and successfully demonstrated antitumor effect based on the antigene strategy.8

We recently developed iso‐dC9,10 and pseudo‐dC (�dC) derivatives for selective recognition of the CG inversion site (Figure 1). The aminopyridine‐bearing �dC derivatives showed a high selectivity and affinity toward the CG base pair without being affected by neighboring bases. Remarkably, the TFO incorporating four 3‐methyl‐2‐aminopyridinyl‐�dC (MeAP‐�dC) formed the stable triplex with the promoter sequence of EGFR gene containing four CG inversion sites, to which the natural TFO did not form the triplex DNA (Figure 2). The MeAP‐�dC containing TFO designed to target hTERT effectively inhibited its transcription in human cancer cells. Thus, MeAP‐�dC is expected to be a new candidate of triplex‐formation‐oligonucleotides for a wide variety of genome‐targeting applications.

Figure 1. Design of iso‐dC and pseudo‐dC derivatives for selective recognition of the CG inversion site.


Figure 2. Evaluation of triplex formation with the promoter sequences (EG1) of the EGFR gene. REFERENCES 1) S. Sasaki, H. Yamauchi, F. Nagatsugi, R. Takahashi, Y. Taniguchi and M. Maeda, Tetrahedron Lett., 2001,

42, 6915–6918. 2) S. Sasaki, Y. Taniguchi, R. Takahashi, Y. Senko, K. Kodama, F. Nagatsugi and M. Maeda, J. Am. Chem.

Soc., 2004, 126, 516–528. 3) Y. Taniguchi, A. Nakamura, Y. Senko, K. Kodama, F. Nagatsugi and S. Sasaki, Nucleosides Nucleotides

and Nucleic Acids, 2005, 24, 823–827. 4) Y. Taniguchi, A. Nakamura, Y. Senko, F. Nagatsugi and S. Sasaki, J. Org. Chem., 2006, 71, 2115–2122. 5) Y. Taniguchi, M. Togo, E. Aoki, Y. Uchida and S. Sasaki, Tetrahedron, 2008, 64, 7164–7170. 6) Y. Taniguchi, Y. Uchida, T. Takaki, E. Aoki and S. Sasaki, Bioorg. Med. Chem., 2009, 17, 6803–6810. 7) E. Aoki, Y. Taniguchi, Y. Wada and S. Sasaki, ChemBioChem, 2012, 13, 1152–1160. 8) Y.Taniguchi, S.Sasaki, Org. Biomol. Chem., 2012, 10, 8336‐8341. 9) H. Okamura, Y. Taniguchi, S. Sasaki, Org. Biomol. Chem. 2013, 11, 3918. 10) H. Okamura, Y. Taniguchi, S. Sasaki, ChemBioChem 2014, 15, 2374. ACKNOWLEDGEMENTS This study was supported by a Grant‐in‐Aid for Scientific Research (B) (Grant Numbers 15H04633 for S.S.), a Grand‐in‐Aid for Young Scientific (A) (Grant Number 24689006 for Y.T.), a Challenging Exploratory Research (Grant Number 26670056 for Y.T.), a Grant‐in‐Aid for JSPS Fellows (Grant Number 13J04516 for H.O.) from the Japan Society for the Promotion of Science (JSPS) and Platform for Drug Discovery, Informatics, and Structural Life Science from the Ministry of Education, Culture, Sports, Science and Technology, Japan.


EFFECTIVE CYTOPLASMIC DELIVERY OF MACROMOLECULES BY FAINT ELECTRIC TREATMENT

1Kentaro KOGURE, 1Kouki FUJIKAWA, 2Hasan MAHADI, 2Susumu HAMA

1UNIVERSITY OF TOKUSHIMA, INSTITUTE OF HEALTH BIOSCIENCES

2KYOTO PHARMACEUTICAL UNIVERSITY

KEY WORDS : ELECTRIC TREATMENT, TRANSDERMAL DELIVERY ABSTRACT : Functional macromolecules, such as siRNA, are expected as the ideal drugs to induce immune response and suppress specific genes for therapy of skin disorders and cancer. However, delivery of the macromolecules into skin is difficult due to large molecular weight and high hydrophilicity. Therefore, since iontophoresis is known to accelerate transdermal permeation of charged molecules by applying a faint electricity to the skin, we paid attention to iontophoresis as an ideal technology for transdermal delivery, and attempted transdermal delivery of macromolecules. siRNA is expected as novel nucleic acid medicines. Thus, we examined iontophoresis of naked siRNA on rat skin in vivo. Naked siRNA effectively accumulated in the skin after iontophoresis. In a rat model of atopic dermatitis, skin was sensitized with ovalbumin to stimulate IL‐10 mRNA expression. Iontophoretic delivery of anti‐IL‐10 siRNA significantly reduced (73%) the level of IL‐10 mRNA1). From this result, it was suggested that siRNA was delivered into not only the skin, but also cytoplasm of target cells. Then, we analyzed the intracellular delivery of naked anti‐luciferase siRNA by faint electric treatment (ET), and found that the luciferase activity of cells expressing luciferase was reduced by in vitro ET like in vivo iontophoresis. Cellular uptake of fluorescent‐label siRNA was increased by ET, while low temperature exposure, macropinocytosis inhibitor amiloride and caveolae‐mediated endocytosis inhibitor filipin significantly prevented siRNA uptake. These results indicate that the cellular uptake mechanism involved endocytosis. In addition, voltage sensitive fluorescent dye DiBAC4 (3) penetration was increased by ET, and the transient receptor potential channel inhibitor SKF96365 reduced siRNA uptake, suggesting that faint ET reduced membrane potentials by changing intracellular ion levels. Moreover, to analyze cytoplasmic delivery, we used in‐stem molecular beacon (ISMB), which fluoresces upon binding to target mRNA in the cytoplasm. Surprisingly, cytoplasmic ISMB fluorescence appeared rapidly and homogeneously after ET, indicating that cytoplasmic delivery is markedly enhanced by ET.2)

In conclusion, we successfully delivered macromolecules into the skin and cytoplasm for induction of

functionalities. This system is expected to serve as a noninvasive approach for transdermal delivery of

various functional macromolecules.

REFERENCES : 1) Kajimoto K, Yamamoto M, Watanabe M, Kigasawa K, Kanamura K, Harashima H, Kogure K. Int J Pharm.

403, 57‐65 (2011). 2) Hasan M, Nishimoto A, Ohgita T, Hama S, Kashida H, Asanuma H, Kogure K. J Control Release. 228, 20‐

25 (2016).


enTRinsic™ DRUG DELIVERY TECHNOLOGY FOR LIVE BIOTHERAPEUTICS (Microbiomes)

Dr. Hassan BENAMEUR

Sr. Director Pharmaceutical Sciences, Capsugel R&D Strasbourg,

Parc d’Innovation, 180 rue Tobias Stimmer, 67412 Illkirch‐Graffenstaden, France

KEY WORDS: Enteric drug delivery capsule / microbiome / intestinal delivery ABSTRACT:

There is a growing need to develop drug delivery systems for the oral delivery of compounds highly sensitive to acidity and enzymes, like peptides, proteins, nucleotides, live organisms. Over the last decade, microbiome became a promising discovery field that aims at developing new therapeutic strategies using commensal bacteria strains. Indeed, the human body hosts more than 100 trillion microorganisms, principally in the whole gut, but they also live in other mucosal surfaces (e.g. mouth, skin). These microorganisms are a key component of our physiology: they participate to several beneficial functions (e.g. digestion of food, prevention of our body from pathogens) but can also be involved in diseases when the equilibrium is broken. Microbiome applications show a great diversity and aim at targeting local inflammatory gastro‐intestinal diseases like Inflammatory Bowel Syndrome and Crohn’s disease, and systemic diseases as well (e.g. diabetes, asthma). However, oral delivery of such strains is challenging as they are sensitive to stomach acidity and an enteric protection is therefore needed to ensure the delivery of viable cells in the intestine. Also, these strains often show a certain level of sensitivity to water, temperature, oxygen, which often increase the complexity of the development strategies. For example, standard formulation approaches, like solvent or water‐based coating of solid dosage forms using enteric polymers resulted in stability issues due to exposure to heat and/or humidity. In addition, such microorganisms are mainly formulated as freeze‐dried powders that frequently show stability challenges due to their hygroscopicity. enTRinsic™, a new intrinsically enteric capsule drug delivery system, was developed to meet customers demand for an oral dosage form that provides the required level of protection from stomach acid degradation, fully delivers the strain cells in the intestine, maintains the strain viability over long‐term storage and simplifies the overall manufacturing process.

enTRinsic™ drug delivery system is made by conventional hard capsules pin‐dipping process, using an enteric polymer formulation. The resulting capsules meet the same appearance and dimensions standard than regular hard capsules but provide a full enteric protection (compliance demonstrated with both Eur. Ph. and USP pharmacopeial disintegration and dissolution specifications for gastro‐resistant dosage forms). Therefore, avoids all the stability issues associated with conventional coating process and ensure higher viable microbiome strain content in the capsules. The reduced amount of residual water in enTRinsic™ (< 7%) also improves the compatibility with freeze‐dried formulations containing microbiome strains. enTRinsic™ technology is currently used by different biopharmaceutical companies for the product development of numerous new live biotherapeutic products, including various long‐term stability studies and clinical trials. In a recent customer Phase I trial, whilst the primary objective was safety and tolerability, the data trends towards the effectiveness of the treatment, with an overall improvements in symptoms and an increase of the strain levels in microbiome samples taken from the volunteers. Several stability programs are on‐going for various microbiome strains, using


enTRinsic™ filled with freeze‐dried formulations. Three studies testing different strains and different packaging types already demonstrated good product viability after 12 months at 5°C, with a maximum of one Log drop in the number of viable cells per capsules. It therefore showed the ability

enTRinsic™ drug delivery technology, in addition to provide an effective enteric protection from stomach acidity, is also able to successfully deliver a viable single live biotherapeutic strain to the gut and to significantly improve the viability over long‐term storage of challenging products such as freeze‐dried live biotherapeutic strains. Additional on‐going stability and clinical studies will confirm these unique features. REFERENCES:

M. Dave, et al., The human gut microbiome: current knowledge, challenges, and future directions, Translational Research 160 (2012).

M. Mimee, et al., Microbiomes therapeutics – Advances and challenges, Adv. Drug Deliv. Rev. (2016).

R. Martin, et al., Role of commensal and probiotic bacteria in human health: a focus on inflammatory bowel disease, Microbial Cell Factories 12 (2013).

C. Reiff, et al., Inflammatory bowel disease, gut bacteria and probiotic therapy, Int. J. Med. Microbio. 300 (2010).

J.J Ross, et al., Considerations in the development of Live Biotherapeutic Products for clinical use, Curr. Issues Mol. Biol. 10 (2008).

A.K. Anal, et al., Recent advances in microencapsulation of probiotics for industrial applications and targeted delivery, Trends in Food Sci. & Technol 18 (2007).

H. Benameur, Enteric Capsule Drug Delivery Technology –Achieving Protection Without Coating Drug Development & Delivery Vol 15 No 5 (2015)

ACKNOWLEDGEMENTS: Dr Sivert Aurélien Head pharmaceutical products and Pharmaceutical Sciences team for their work


START OF ‘COMMISSION FOR SPECIES DIFFERENCE PROBLEM ON FORMULATION DESIGN’

SUPPORTED BY THE ACADEMY OF PHARMACEUTICAL SCIENCE & TECHNOLOGY, JAPAN (APSTJ)

Hiroshi KIKUCHI1,2,3,4

1Executive Director, Tsukuba Research Laboratories, Eisai Co., Ltd. Tokodai 5‐1‐3, Tsukuba‐shi, Ibaraki 300‐2635, Japan

2Adjunct Professor, Faculty of Pharmaceutical Sciences, The University of Tokyo 3Affiliate Professor, Graduate School of Pharmaceutical Sciences, Kyushu University

4Adjunct Professor, Graduate School of Engineering, Kyoto University

KEY WORDS: species difference, formulation design, animals, humans, systematization

ABSTRACT :

Various animal experiments in vivo are performed in order to confirm biodistribution, efficacy, and safety of the final formulations of the drugs in addition to designing and screening of the formulations. Results of such animal experiments are very significant, however, there are some problems attributable to species difference for evaluation and analysis of those animal experiment results. In this lecture, the author will show some data as examples and explain species difference problems (Table 1‐3, Fig.1‐6). Because the systematization of the evaluation method of DDS formulations in animals has been considered to be very valuable, he proposes two kinds of solutions. One is collecting the various kinds of animal data by using the commercialized DDS medicines whose clinical data are already obtained. Another is establishing an organization like consortiums where the participants can exchange their own information related to species difference. As the latter case, ‘Commission for Species Difference Problem on Formulation Design’ supported by The Academy of Pharmaceutical Science & Technology, Japan (APSTJ) and six Japanese pharmaceutical companies will be briefly introduced, which has just started this year. In this commission, the experiences and/or information concerning species difference observed in R&D of DDS formulations are provided, shared and discussed with participants.

REFERENCES:

1) H. Kikuchi, Importance of dose number and absorption test in formulation optimization: an industrial case, Oral Delivery of Poorly Soluble Actives – From Drug Discovery to Marketed Products, Capsugel Library, Tokyo (2003), pp. 63‐71．

2) T. Suzuki, M. Ichihara, K. Hyodo, E. Yamamoto, T. Ishida, H. Kiwada, H. Ishihara and H. Kikuchi, Accelerated blood clearance of PEGylated liposomes containing doxorubicin upon repeated administration to dogs, Int. J. Pharm. 436, 636‐643 (2012).

3) T. Suzuki, M. Ichihara, K. Hyodo, E. Yamamoto, T. Ishida, H. Kiwada, H. Kikuchi and H. Ishihara, Influence of dose and animal species on accelerated blood clearance of PEGylated liposomal doxorubicin, Int. J. Pharm. 476, 205‐212 (2014).


EFF


DELIVERY OF PEPTIDES BY NON‐INVASIVE ROUTES: FOCUS ON SUCCESSFUL ORAL TECHNOLOGIES PROGRESSING IN THE CLINIC

AND FUTURE CHALLENGES

Joël RICHARD

IPSEN – Peptides Development ‐ 20, Rue Ethé Virton – 28100 Dreux, FRANCE

KEY WORDS : Non‐invasive peptide delivery – Transdermal peptide delivery ‐ Oral peptide delivery – Oral bioavailability – Permeation enhancer – Clinical studies

ABSTRACT : Most of large molecules like proteins and peptides are administered chronically through the parenteral route, sometimes for long periods of treatment. For this reason, there is a real interest in developing and bringing to the market formulations that can be administered through a non‐invasive route, either the transdermal, oral, buccal, or nasal and pulmonary. The purpose is really to focus on the patient and improve compliance, treatment adherence and patient comfort. Many progresses have been made in the recent years in the field of non‐invasive routes, most of them significantly progressing in the Clinic. For instance, transdermal delivery of large molecules progressed very well. The most successful technology for this route was the micro‐needle technology that went in clinical Phase III for vaccines and Phase II for a PTH analogue, showing a very high absorption of the peptide. Oral and buccal routes were investigated and strongly progressed as well. Buccal insulin progressed in late stage development for treatment of type 2 diabetes and went to the market in some countries. Different types of technologies were developed for this purpose, like non‐ionic surfactant‐based gels, pelleted nanoparticles, phospholipid vesicles, buccal muco‐adhesive films, sprayed micelle solutions. As large molecules like proteins and peptides generally show a very low ability to cross the membranes, one of the main challenges is the development of new technologies that improve permeability of these molecules across the epithelial membranes. These can be permeation enhancers like acylated amino‐acids or functionalized nanoparticles that behave as active carriers of the molecules through various mechanisms. Protection of the proteins in the gastro‐intestinal (GI) tract until they reach the absorption site is also a key success factor for these new delivery technologies. Hence the importance of having enteric coated oral forms and enzyme inhibitors in the formulation to prevent degradation before absorption. Generally speaking, the development of non‐invasive routes of administration requires the development of a full arsenal of stabilizing excipients and technologies to avoid degradation in “aggressive” media (e.g. GI fluid) and permeation enhancement technologies to increase crossing of the membranes. Degradation and poor permeability are actually the key challenges for oral delivery of large molecules like proteins and peptides. For this reason, most oral delivery technologies intended to deliver peptides are designed to: ‐ protect orally delivered proteins from detrimental enzymatic activity within the gastrointestinal tract, and ‐ enhance their absorption across the intestinal wall. Usually, the active peptide is formulated in a capsule or tablet that bears an enteric coating (made of pH‐sensitive acrylic and/or cellulosic polymers) which is not dissolved nor destroyed in the most acidic segments of the gut, as well as specialized protease inhibitors (like citric acid beads or soy bean trypsin inhibitors) that avoid enzymatic degradation. Drug bioavailability is then increased by an absorption enhancer that facilitates peptide passing across the intestinal barrier. As already mentioned, acylated amino‐acids were among the first and quite popular for making physical complexes with proteins and improving their transport through the epithelial cells. Acylcarnitine was also successfully used as well as bile salts and fatty acids. Many other permeation enhancers are under development. The interest of muco‐adhesive polymer‐based formulations was also shown based on the increase of residence time of the drug in the area of absorption close to the membrane.


ROLE OF EXPLORATORY CLINICAL STUDY FOR ORAL DDS FORMULATION

DEVELOPMENT

Hiromu KONDO

Pharmaceutical Research and Technology Labs, Astellas Pharma Inc.

180, Ozumi, Yaizu‐shi, Shizuoka 425‐0072, JAPAN

KEY WORDS : Oral controlled release formulation/gamma scintigraphy/gastrointestinal transit/in vitro‐in vivo correlation

ABSTRACT : Oral controlled release formulations release drugs at intended rate with traveling through the gastrointestinal tract. It is meaningful to understand the characteristics of the gastrointestinal transit in order to design oral controlled release formulation. Though several in vitro test methods with mimic human gastrointestinal tract conditions have been reported, in vitro‐in vivo correlation is still under discussion. The gastrointestinal tract conditions vary according to food intake, exercise, dosing time, and so on. Therefore, it is difficult to capture the gastrointestinal tract as a specific condition. Nevertheless, to understand the characteristics of the human gastrointestinal transit under various conditions is useful to establish appropriate in vitro test methods. A formulation prepared with a model drug could be administered to collect blood samples, which make it to estimate and evaluate the gastrointestinal transit property in human. But, this method would evaluate the gastrointestinal transit property indirectly. And, absorption profile of a model drug might complicate to realize the gastrointestinal transit property. In contrast, the method to monitor the travel manner of the formulation with a small amount of radioisotope from outside the body by scintigraphic camera non‐invasively is valuable for the evaluation of the site and the situation of formulation in the gastrointestinal tract in human1). Radioisotope with gamma ray enables to obtain information of inside the body from outside the body because of its strong permeability. Astellas Pharma Inc. has researched and developed several unique oral controlled release formulations. Oral Controlled Absorption System, OCAS2), is a matrix type tablet which is composed of water soluble additives such as polyethylene glycol, which can form enough gel in the tablet within a short time. Also, it is composed of polyethylene oxide as a gel forming polymer, which can intertangle strongly each other. As a result, OCAS tablet has enough strength against mechanical stress caused by the gastrointestinal motility. OCAS shows pseudo zero order drug release profile. In application of OCAS technology, it is important to understand gastrointestinal transit property of OCAS in order to design and develop OCAS formulation. Gamma scintigraphy method could evaluate not only gastrointestinal transit of OCAS formulation but also the effect of food intake and/or administration timing on the gastrointestinal transit. Erosion profile of OCAS in human has also been investigated. The results of these evaluations indicated that the OCAS could work in vivo as designed in vitro. In vitro‐in vivo correlation of OCAS can be confirmed and therefore makes it possible to develop OCAS formulation effectively. Products with OCAS technology are launched in more than 100 countries worldwide at the present.


LIPID‐BASED FORMULATIONS FOR ENHANCED ORAL BIOAVAILABILITY OF BCS CLASS II DRUGS

Vincent JANNIN

Gattefossé S.A.S.

KEY WORDS: lipids, solubility, self‐emulsifying drug delivery system, digestion, coculture ABSTRACT : Lipid‐based formulations (LBFs) can be effective drug delivery systems for poorly water‐soluble chemical entities, provided they are designed with careful selection of the excipients, based on their role in the delivery system and in relation to the API properties (1). Among the key considerations in the selection step must be the series of mechanisms underlining oral bioavailability enhancement. The primary factor leading to increased bioavailability by LBFs is the administration of the drug in a pre‐dissolved state thus avoiding the solid‐to‐liquid phase transition process. Moreover, the interactions between the LBF and the endogenous lipids such as bile salts, phospholipids and dietary lipids will help maintain the drug in solution within colloidal structures (2). Next, lipids and lipid metabolites (3‐5) can overcome other bioavailability limiting factors such as mitigation of intestinal efflux transporters, fluidization of the intestinal epithelium or promotion of lymphatic transport, thus minimizing the impact of first pass metabolism. The fate of a drug formulated in a LBF is hence dependent on the ability of the formulation components to keep the drug in solution during the initial dispersion and/or digestion processes (6). These systems should therefore be formulated by the wise selection of excipients and appropriate combination of oils, water‐soluble surfactants, water‐insoluble surfactants, and hydrophilic cosolvents based on their intended function: some excipients are needed to dissolve the drug within the dosage form; some to promote the self‐emulsification (7‐8), and others to generate the metabolites that maintain drug solubility in the gastro‐intestinal colloidal phases. Additionally, certain excipients may be added to promote the drug passage across the intestinal epithelium. To better appreciate the impact of lipid‐based excipients on drug absorption, an in vitro goblet cell/absorptive cell‐based coculture with a continuous mucus layer on top of the cells was developed (9). It protects the cells from the surfactants and enzymes contained in the biorelevant media and LBFs (10). Moreover, standardized in vitro tests of dispersion and digestion are now available to help predict the in vivo performance of LBFs (11). The protocol to be used for developing LBFs is presented and exemplified by case studies with model drugs (12). REFERENCES :

1) Jannin, V., Musakhanian, J., Marchaud, D. 2008. Approaches for the development of solid and semi solid lipid‐based formulations. Advanced Drug Delivery Reviews, 60 (6) 734‐746.

2) Fernandez, S., Jannin, V., Chevrier, S., Chavant, Y., Demarne, F., Carrière, F. 2013. In vitro digestion of the self‐emulsifying lipid excipient Labrasol® by gastrointestinal lipases and influence of its colloidal structure on lipolysis rate. Pharmaceutical Research, 30(12) 3077‐3087.

3) Bakala N’Goma, J.C., Amara, S., Dridi, K, Jannin, V., Carrière, F. 2012. Understanding lipid digestion in the GI tract for effective drug delivery. Therapeutic Delivery, 3(1) 105‐124.

4) Fernandez, S., Jannin, V., Rodier, J.D., Ritter, N., Mahler, B., Carrière, F. 2007. Comparative study of digestive lipases performance on the self emulsifying excipient Labrasol®, medium chain glycerides and PEG esters. Biochimica et Biophysica Acta ‐ Molecular and Cell Biology of Lipids, 1771 (5), 633‐640.

5) Fernandez, S., Rodier, J.D., Ritter, N., Mahler, B., Demarne, F., Carrière, F., Jannin, V. 2008. Lipolysis of the semi‐solid self emulsifying excipient Gelucire® 44/14 by digestive lipases. Biochimica et Biophysica Acta ‐ Molecular and Cell Biology of Lipids, 1781 (8), 367‐375.


6) Fernandez, S., Chevrier, S., Ritter, N., Mahler, B., Demarne, F., Carrière, F., Jannin, V. 2009. In vitro gastrointestinal lipolysis of four formulations of piroxicam and cinnarizine with the self emulsifying excipients Labrasol® and Gelucire® 44/14. Pharmaceutical Research, 26 (8) 1901‐1910.

7) Chambin, O., Karbowiak, T., Djebili, L., Jannin, V., Champion, D., Pourcelot, Y., Cayot, P. 2009. Influence of drug polarity upon the solid‐state structure and release properties of self‐emulsifying drug delivery systems in relation with water affinity. Colloids and Surfaces B: Biointerfaces, 71 (1) 73‐78.

8) Chamieh, J., Davanier, F., Jannin, V., Demarne, F., Cottet, H. 2015. Size characterization of micelles and microemulsions by Taylor Dispersion Analysis. International Journal of Pharmaceutics. 492(1‐2) 46‐54.

9) Béduneau, A., Tempesta, C., Fimbel, S., Pellequer, Y., Jannin, V., Demarne, F., Lamprecht, A. 2014. A tunable Caco‐2/HT29‐MTX co‐culture model mimicking variable permeabilities of the human intestine obtained by a new seeding procedure. European Journal of Pharmaceutics and Biopharmaceutics, 87 (2) 290‐298.

10) Antoine, D., Pellequer, Y., Tempesta, C., Lorscheidt, S., Kettel, B., Tamadon, L., Jannin, V., et al. 2015. Biorelevant media resistant co‐culture model mimicking permeability of human intestine. International Journal of Pharmaceutics. 481(1‐2) 27‐36.

11) Williams, H., Sassene, P., Kleberg, K., Bakala N’Goma, J.C., Calderone, M., Jannin, V., Igonin, A., et al. 2012. Towards the establishment of standardized in vitro tests for lipid‐based formulations: 1) Method parameterisation and comparison of in vitro digestion profiles across a range of representative formulations. Journal of Pharmaceutical Sciences, 101(9) 3360‐3380.

12) Jannin, V., Chevrier, S., Michenaud, M., Dumont, C., Belotti, S., Chavant, Y., Demarne, F. 2015. Development of Self Emulsifying Lipid Formulations of BCS class II drugs with low to medium lipophilicity. International Journal of Pharmaceutics. 495(1) 385‐392.


COMPREHENSION OF MANNITOL BEHAVIOR IN HIGH SPEED ROTARY PRESS TABLETING USING SIMULATOR

Philippe LEFEVRE, Nicolas TARLIER

ROQUETTE, Customer Technical Service, Pharma Global Business Unit, 1 rue de la Haute Loge, 62136 Lestrem, France

KEY WORDS: Tablet, Mannitol, Compression, Simulator, Stylcam,

ABSTRACT: Mannitol is widely used as pharmaceutical excipient in solid dosage form. The non‐hygroscopic properties and inherent chemical structure of mannitol make it a very inert and stable excipient that is preferred when processing moisture sensitive products. Textured mannitol powders are being increasingly used as fillers for tablets produced by direct compaction (Bolhuis et al., 2009). These textured mannitol powders are processed to obtain good flowability, blending and compactibility properties which are essential requirements for directly compressible filler. The behavior of mannitol in tableting has always been difficult to apprehend and literature is conflicting on the nature of mannitol deformation mechanisms. Robert et al described non textured mannitol powder as being a moderately hard and ductile material (Roberts and Rowe, 1987). Since, there have been many studies reporting non‐textured and textured mannitol as having a ductile deformation mechanism and therefore classified the mannitol as a ductile / moderately hard material (Bolhuis et al., 2009; Juppo et al., 1995; Ohrem et al., 2014; Westermarck et al., 1998; Yoshinari et al., 2003). Recently however, non‐textured mannitol was found to be more a fragmentary material (Klevan et al., 2010; Nordström et al., 2012). One probable explanation for this confusion is that mannitol behavior changes with the way the compression is done. Past studies were mainly done on nonsymmetrical single punch or hydraulic presses, which do not reflect intended use conditions. We anticipate that compression kinetic was the key point when tableting mannitol. A rotary tablet press simulator STYLCAM™ was used to characterize the deformation behavior of textured mannitol in conditions of high speed rotary press tableting. Compression simulators are described as able to mimic industrial compression on rotary presses. They apply a symmetrical compression (with or without pre‐compression) with kinetics usually observed on industrial rotary press. They are fully instrumented and allow formulators to realize studies in real production conditions while acquiring a set of data enabling a scientific approach. More, having only one set of punches, studies on simulator require a very low amount of powder and short time for a full evaluation. So series of studies are performed whereas they are too long or too expensive to be done on high speed rotary press. STYLCAM™ 200R rotary press simulator was used to:

Evaluate and characterize the behavior of a textured mannitol powder in simulated industrial compaction conditions, which to our knowledge has not been performed yet. We investigated Magnesium Stearate (MgSt) lubricant effect on the compaction and deformation behavior of textured mannitol powder. Heckel modeling, strain sensitivity rate and Walker modeling were then applied to determine deformation mechanisms.

Comparison with a single punch press (nonsymmetrical compression) to identify the limits of that type of press when working on mannitol

Determine what tableting conditions or parameters are detrimental to the performance of mannitol in direct compression

Solve issues experienced during production of formulated tablets The results of this work show that it was possible to characterize the deformation behavior of textured mannitol in conditions of industrial tableting using an instrumented rotary tablet press simulator. Compressibility characterization by Heckel and Walker modeling showed that the mean yield pressure and


the W* coefficient were typically more fragmentary than ductile. Furthermore the strain rate sensitivity close to 0 is a characteristic of brittle material. We thus can confirm that under industrial tableting conditions, the studied textured mannitol behaves more like a fragmentary material than the previously claimed plastic deformation (Ohrem et al., 2014). The textured mannitol compaction behavior was shown to be unaffected by MgSt lubricant concentration or mixing time which is typical of fragmentary deformation. Contrary to single punch nonsymmetrical tablet press, compression simulators are able to apply a pre‐compression step. This pre‐compression is compulsory to avoid capping at high compression force or high compression speed when using highly concave punches. With flat punches it allows to increase production yield as capping troubles start at higher speed. On STYLCAM™ simulator data were acquired using pre‐compression and compression profiles similar to the ones obtained without pre‐compression were observed. So rotary press simulator gives access to data which are not accessible using a single punch nonsymmetrical tablet press when pre‐compression is required. Some common problems encountered during tablets production were studied on compression simulator, as well as the impact of tooling characteristics. Troubles experienced on formulated tablets production were investigated and the proposed solutions were successfully applied on high speed tableting press. Rotary tablet press simulators turn out to be efficient and unescapable tool for studying scientifically the industrial tableting and notably when using powder with a complex behavior as mannitol. A rotary press simulator is not a real rotary press. Notably, there is no possibility to study the fouling and the effect of temperature during long time tableting. Nevertheless up to now the data obtained from STYLCAM™ simulator and consequently the solutions proposed were always confirmed on industrial rotary press production. REFERENCES : Bolhuis, G.K., Rexwinkel, E.G., Zuurman, K., 2009. Polyols as filler‐binders for disintegrating tablets prepared by direct compaction. Drug Dev. Ind. Pharm. 35, 671–677. Juppo, A.M., Kervinen, L., Yliruusi, J., Kristoffersson, E., 1995. Compression of lactose, glucose and mannitol granules. J. Pharm. Pharmacol. 47, 543–549. Klevan, I., Nordström, J., Tho, I., Alderborn, G., 2010. A statistical approach to evaluate the potential use of compression parameters for classification of pharmaceutical powder materials. Eur. J. Pharm. Biopharm. 75, 425–435. Nordström, J., Klevan, I., Alderborn, G., 2012. A protocol for the classification of powder compression characteristics. Eur. J. Pharm. Biopharm. 80, 209–216. Ohrem, H.L., Schornick, E., Kalivoda, A., Ognibene, R., 2014. Why is mannitol becoming more and more popular as a pharmaceutical excipient in solid dosage forms? Pharm. Dev. Technol. 19, 257–262. Roberts, R.J., Rowe, R.C., 1987. The compaction of pharmaceutical and other model materials ‐ A pragmatic approach. Chem. Eng. Sci. 42, 903–911. Westermarck, S., Juppo, A.M., Kervinen, L., Yliruusi, J., 1998. Pore structure and surface area of mannitol powder, granules and tablets determined with mercury porosimetry and nitrogen adsorption. Eur. J. Pharm. Biopharm. 46, 61–68. Yoshinari, T., Forbes, R.T., York, P., Kawashima, Y., 2003. The improved compaction properties of mannitol after a moisture‐induced polymorphic transition. Int. J. Pharm. 258, 121–131. ACKNOWLEDGEMENTS: Nicolas TARLIER for his 3 years thesis on the compaction on mannitol, Pf. BATAILLE and the team of the Pharmaceutical Technology Department, University of Montpellier


DEVELOPMENT OF A NEW LIPID BUBBLE FOR ULTRASOUND THERANOSTICS

Kazuo MARUYAMA1, Johan UNGA1, Daiki OMATA1,2,3, Yusuke ODA1, Mutsumi SUGII1,

Hitoshi URUGA1, Ryo SUZUKI1 1Teikyo University, Faculty of Pharma Sciences, 2JSPS Research Fellow,

3Utrecht University, Faculty of Science

KEY WORDS : microbubble, ultrasound, theranostics,

【Purpose】 To develop a novel bubble formulation for ultrasound (US) imaging and therapy with small particle size and a good stability and test the formulation as US imaging contrast agent and for gene delivery in vitro and in vivo.

【Methods】 Lipid‐stabilized bubbles were prepared by homogenization of a lipid dispersion in the presence of perfluoropropane gas. Different phospholipid compositions were tested and evaluated. After bubble formation the bubbles were freeze‐dried so that a dry sample containing bubbles was formed. After re‐constitution of the samples they were analyzed for size, gas content and US signal intensity. The bubbles were also evaluated as contrast agents in vivo, and for US activated gene delivery of luciferase pDNA in vitro on cell culture and in vivo in mice.

【Results and discussion】 Bubbles were in the size range 500‐800 nm and could be re‐constituted by simple addition of water to the dry sample. Changes in the lipid composition had a big impact on the bubbles properties. A mixture of three different lipids in the stabilizing layer resulted in the most stable bubbles. In vivo imaging of B16BL6 tumours in mice, using the most stable bubbles showed circulation times longer than for the commercial bubble Sonazoid. Also, the bubbles were well suited for visualization of tumour neovasculature. Bubbles together with pDNA and US exposure increased the luciferase activity by about 300 times in vitro and 2000 times in vivo, compared to only pDNA+US. We believe this new formulation shows great promise for both diagnostic and therapeutic applications thanks to its good stability, relatively small bubble size and the simplicity of handling. REFERENCES :

1) Tumor specific ultrasound enhanced gene transfer in vivo with novel liposomal bubbles. Suzuki R, Takizawa T, Negishi Y, Maruyama K. et al. J Control Release 125, 137–144, 2008.

2) Tumor growth suppression by the combination of nanobubbles and ultrasound. Suzuki Ryo, Oda Yusuke, Omata Daiki, Unga Johan, Negishi Yoichi, Hashida Mitsuru, Maruyama Kazuo. et al. Cancer Science. 107, 217‐223, 2016.

3) Development of anionic bubble lipopolyplexes for efficient and safe gene transfection with ultrasound exposure in mice, Tomoaki Kurosaki, Ryo Suzuki, Kazuo Maruyama, Fumiyoshi Yamashita, and Mitsuru Hashida, et al. J. Control. Release 176, 24‐34, 2014.

ACKNOWLEDGEMENTS : This work was supported by JSPS KAKENHI (Grant # 16H03196), the MEXT‐Supported Program for the Strategic Research Foundation at Private Universities 2013‐2017.


CONCEPTION OF CONTRAST AGENTS OR NANOTHERAPEUTICS WITH THE HELP OF IMAGING MODALITIES

V. BOUDY1, B‐T. DOAN1, J. SEGUIN1, M. BESSODES1, D. SCHERMAN1, O. COUTURE2, A. DELALANDE3, G. RENAULT3, P. MIDOUX3, C. PICHON3, Nathalie MIGNET1

AFFILIATION :1 Chemical and Biological Technology for Health, CNRS UMR8258, INSERM U1022, Chimie

ParisTech, Université Paris Descartes 2 Institut Langevin, Paris, 3 CBM, Orléans, France

KEY WORDS : NANOPARTICLES, MICROBUBBLES, THERMOGELS, GENE TRANSFER, COLON CANCER MODEL,

IN VIVO MAGING

ABSTRACT : Nanoparticles have potentials in imaging in particular as enhanced contrast agent for techniques with low sensitivity, such as MRI or Ultrasound imaging. They also have a role to play in bimodal imaging. Indeed, thanks to their possible functionalization, various chromophores or contrastophor can be linked to the surface or within the core to provide new properties or to allow obtaining a high number of valuable informations in preclinical studies, while highly reducing the number of animals. The conception of a dedicated nanoparticle for long‐term optical imaging studies (Maldiney et al.) will be presented followed by few examples of the interest of bimodalities in preclinical evaluation will be shown. First, the conception of a protein scaffold for the measurement of the functional hepatic reserve (Chaumet‐Riffaud et al. 2010). A radioactive label provide quantitative informations on the liver function while an optical label will provide evidence on the specificity of the targeting. A second example will concern gaz microbubbles designed for gene transfer. Thanks to both ultrasound and optical imaging, microbubbles surface, injection and sonoporation parameters have been optimized to achieve liver gene transfer (Manta et al. 2015). In our quest of local delivery for colorectal primary cancer and metastases, thermogels were conceived and optimized thanks to physico‐chemical characteristics and in vivo imaging. Absence of recurrence post surgery was shown thanks to local administration of anticancer drug filled hydrogel (N. Zeng, 2014, 2015). REFERENCES :

1) Manta, S. Delalande, A., Bessodes, M., Bureau, M‐F., Scherman, D., Pichon, C., Mignet, N. Characterization of positively‐charged lipid shell microbubbles with Tunable Resistive Pulse Sensing (TRPS) method. A technical note, Ultrasound Med Biol. 2016, 42, 624‐30.

2) T. Maldiney, B.‐T. Doan, D. Alloyeau, M. Bessodes, D. Scherman, C. Richard. Gadolinium‐Doped Persistent Nanophosphors as Versatile Tool for Multimodal In Vivo Imaging. Advanced Functional Materials 2015, 25, 331–338.

3) N. Zeng, N. Mignet, G. Dumortier, E. Olivier, J. Seguin, M. Maury, D. Scherman, P. Rat, V. Boudy. Poloxamer bioadhesive hydrogel for buccal drug delivery: Cytotoxicity and trans‐epithelial permeability evaluations using TR146 human buccal epithelial cell line. Int. J. Pharm (2015) 495, 1028‐1037

4) Maldiney T, Bessière A, Seguin J, Teston E, Sharma SK, Viana B, Bos AJ, Dorenbos P, Bessodes M, Gourier D, Scherman D, Richard C. The in vivo activation of persistent nanophosphors for optical imaging of vascularization, tumours and grafted cells. Nature Materials. 2014, 13(4), 418‐426


5) Zeng N., Dumortier G., Maury M., Mignet N., Boudy V. Influence of additives on a thermosensitive hydrogel for buccal delivery of salbutamol: relation between micellization, gelation, mechanic and release properties Int J. Pharm 2014 467, 70‐83.

6) Delalande A, Postema M, Mignet N, Midoux P, Pichon C. Ultrasound and microbubble‐assisted gene delivery: recent advances and ongoing challenges Ther Deliv. 2012, 3, 1199

7) P. Chaumet‐Riffaud, I. Martinez‐Duncker, A.L. Marty, C. Richard, A. Prigent, F. Moati, L. Sarda‐

Mantel, D. Scherman, M. Bessodes, N. Mignet, Synthesis and Application of Lactosylated, 99mTc Chelating Albumin for Measurement of Liver Function, Bioconj. Chem., 2010, 21, 589‐596.

ACKNOWLEDGEMENTS : Thanks to the National Funding Agency for financial support, the Ligue foundation for PhD funding, CNRS, INSERM, CHimie ParisTech and Paris Descartes University for their continuous support


TO PASS OR NOT TO PASS THE BLOOD BRAIN BARRIER?

Patrick COUVREUR

Institut Galien, UMR CNRS 8612, Université Paris‐Sud (France) KEY WORDS : Nanoparticles, Brain Diseases, Alzheimer Disease, Brain Ischemia, Spinal Cord Injury, Blood

Brain Barrier, Polyalkylcyanoacrylate, Squalene ABSTRACT : Many drugs with potential neuroprotective activity don’t display any therapeutic efficacy because they are rapidly metabolized after intravenous administration and/or unable to diffuse through the blood brain barrier. Therefore, many drug nanocarriers have been designed with the dream to allow some drugs to translocate and diffuse into the brain parenchyma. This has been done by the functionalization of nanoparticles with polyethyleneglycol and various ligands able to target specific receptors of the brain endothelial cells. As an illustration of this approach, we developed a nano‐carrier system that could transfer chitosan nanospheres loaded with specific caspase‐3 inhibitor peptide Z‐DEV‐FMK, across the blood brain barrier (1). Polyethyleneglycol‐coated chitosan nanospheres were conjugated to an anti‐mouse transferrin receptor monoclonal antibody (TfRMAb) that selectively recognizes the transferrin receptor‐type‐1 on the cerebral vasculature. It was demonstrated with intravital microscopy that this nanomedicine was rapidly transported across the blood brain barrier and dose‐dependently decreased the infarct volume in mice experimental brain ischemia (1). We have also reported the design of a versatile and multifunctional targeted biodegradable polyalkylcyanoacrylate nanoparticulate platform (2, 3) and its successful in vitro application in Alzheimer's disease, the most common elderly dementia, affecting 35 million people worldwide. These nanoparticles were functionalized with a novel specific antibody, via the biotin/streptavidin binding strategy, in order to bind β‐amyloid peptide 1‐42 (Aβ1‐42) monomer and Aβ1‐42 fibrillar aggregates, usually located in Alzheimer's disease brains. Beyond the report of a new methodology for multifunctional nanoparticle construction, this also represented the first example of targeted polymeric nanoparticles for therapeutic application in Alzheimer’s disease (4). 15 month old Tg2576 transgenic mice receiving intravenous infusion of anti‐Aβ‐nanoparticles (3 days by week during 3 weeks) had complete correction of the memory defect in Alzheimer's disease transgenic mice. It was hypothesized that significant reduction of the Aβ soluble peptide and oligomer levels in brain accounted for this unique observation (5). In literature, some other examples demonstrated that well engineered nanoparticles or liposomes may help to translocate into the brain. However, the amount of nanoparticles able to reach the brain parenchyma remained always very modest and never exceeded 0.5% of injected dose/gr of tissue. This leads to limited ability to deliver drugs into the brain in a sufficient amount, also because of the poor nanoparticles and liposomes drug loading which rarely exceeds few %. In this context, we have developed the “squalenoylation/terpenoylation” concept (6, 13), a technology that takes advantage of the squalene's dynamically folded conformation to link this natural and biocompatible lipid to anticancer (7, 8), antimicrobial (9, 12) or neuroprotective compounds (10) in order to achieve the spontaneous formation of nanoassemblies (100–300 nm) in water, without the aid of surfactants. This new drug delivery concept has been applied to adenosine, an endogenous molecule representing a class of potential therapeutic agent with significant beneficial activity in several severe neurological disorders, such as stroke, spinal cord injury or multiple sclerosis. Nevertheless, adenosine has never been used for the treatment of severe neurological disorders and neurotrauma, as it possesses serious limitations, such as very short plasma half‐life due to rapid metabolization, the advent of moderate side effects and inability to


cross the blood brain barrier and the blood spinal cord barrier. It was found that the nanoparticles resulting from the bioconjugation of adenosine to squalene allowed: (i) an efficient protection from rapid metabolization and (ii) the induction of a dramatic neuroprotective effect in both an ischemia‐reperfusion model in mice and a spinal cord injury model in rats (10). The use of dual radio‐labeling and radio‐HPLC as analytical methods, as well as FRET, provided important information that neither the squalene‐adenosine nanoparticles, nor the squalene‐adenosine bioconjugate or even adenosine were delivered into the brain parenchyma (14, 15). On the contrary, our results indicated that the nanoparticles acted as an adenosine reservoir in the systemic circulation for at least 1h, allowing adenosine to interact with the brain microcirculation and the neurovascular unit, mainly composed by endothelial cells, pericytes, astrocytes endfeet and neurons, thereby triggering an indirect central effect. This demonstrates that treating neurological disorders doesn’t need, as commonly considered, to translocate the blood brain barrier but that just peripheral pharmacological activity may be beneficial (10, 14). REFERENCES : 1) H. Karatas et al., Journal of Neuroscience. 29, 13761‐13769, (2009)

2) L. Barraud et al., Journal of Hepatology, 42, 736‐743 (2005)

3) B. Le Droumaguet et al., ACS Nano, 6, 5866‐5879 (2012)

4) D. Brambilla et al., ACS Nano, 6, 5897‐5908 (2012)

5) D. Carradori et al., unpublished results (2016)

6) P. Couvreur et al., Nano Letters, 6, 2544‐2548 (2006)

7) A. Maksimenko et al., Proceedings of the National Academy of Science, 111 (2) E217‐E226 (2014)

8) A. Maksimenko et al., ACS Nano, 8, 2018‐2032 (2014)

9) N. Semiramoth et al., ACS Nano, 6, 3820‐3831 (2012)

10) A. Gaudin et al., Nature Nanotechnology, 9, 1054‐1063 (2014)

11) S. Mura et al., Nature Materials, 12, 991‐1003 (2013)

12) N. Abed et al., Scientific Reports (Nature), doi: 10.1038/srep13500 (2015)

13) S. Harisson et al., Angewandte Chemie Int. Edition, 52, 1678‐1682 (2013)

14) A. Gaudin et al., J Control Release, 212, 50‐58 (2015)

15) A. Gaudin et al., Chemistry of Materials, 27, 3636−3647 (2015)


SELF‐ASSEMBLED SUPRAMOLECULAR NANOSYSTEMS FOR SMART TARGETED THERAPY OF INTRACTABLE DISEASES

Kazunori KATAOKA

Innovation Center of NanoMedicine, Institute of Industry Promotion‐KAWASAKI, Japan Policy Alternatives Research Institute, The University of Tokyo, Tokyo, Japan

KEY WORDS : Nanomedicine, Polymeric Micelle, pH‐activatable systems, Cancer treatment

ABSTRACT : Nanotechnology‐based medicine (Nanomedicine) has received progressive interest for the treatment of intractable diseases, such as cancer, as well as for the non‐invasive diagnosis through various imaging modalities. Engineered polymeric nanosystems with smart functions play a key role in nanomedicine as drug carriers, gene vectors, and imaging probes. This presentation focuses present status and future trends of self‐assembled nanosystems from block copolymers for the therapy and the non‐invasive diagnosis of intractable cancer.

Nanosystems with 10 to 100 nm in size can be prepared by programmed self‐assembly of block copolymers in aqueous entity. Most typical example is polymeric micelles with distinctive core‐shell architecture. Several micellar formulations of antitumor drugs have been intensively studied in preclinical and clinical trials, and their utility has been demonstrated1. Compared with conventional formulations, such as liposomes, polymeric micelles have several advantages, including controlled drug release, tissue penetrating ability and reduced toxicity2. The development of smart polymeric micelles that dynamically change their properties due to sensitivity to chemical or physical stimuli is the most promising trend toward nanomedicines, directing to the targeting therapy with high efficacy and ensured safety. Notable anti‐tumor efficacy against intractable and metastatic cancer, including pancreatic cancer, of antitumor drug‐incorporated polymeric micelles with pH‐responding property was demonstrated to emphasize a promising utility of nanomedicines for cancer treatment3,4.

Versatility in drug incorporation is another feasibility of polymeric micelles. Polymeric micelles loaded with siRNA and mRNA have been successfully formulated with relevant properties for nanotherapeutics5‐7. Furthermore, loading of imaging reagents makes polymeric micelles with theranostic functions8,9. These results demonstrate the promising features of polymeric micelles as platform nanosystems for molecular therapy of various intractable diseases through the selective delivery of a variety of drugs having issues on pharmacokinetics and pharmacodynamics. REFERENCES :

1) H. Cabral, K. Kataoka, J. Contrl. Rel. 190 (2014) 465‐476. 2) Y. Matsumoto, et al, Nature Nanotech. 11 (2016) 533‐538. 3) H. Cabral, et al, ACS Nano 9 (2015) 4957‐4967. 4) H. Kinoh, et al, ACS Nano 10 (2016) 5643‐5655. 5) H. Nishida, et al, J. Contrl. Rel. 231 (2016) 29‐37. 6) H. Uchida, et al, J. Amer. Chem. Soc. 136 (2014) 12396‐12405. 7) C.‐Y. Lin, et al, J. Contrl. Rel. 235 (2016) 268‐275. 8) P. Mi, et al, ACS Nano 9 (2015) 5913‐5921. 9) P. Mi, et al, Nature Nanotech. (2016) Published Online.


MODULATION OF THE TUMOR MICROENVIRONMENT BY NANOTHERAPY

Iris MARANGON, Amanda SILVA, Florence GAZEAU

Laboratoire Matière et Systèmes Complexes, CNRS / Université Paris Diderot 10 rue Alice Domon et Léonie Duquet, 75205 Paris

ABSTRACT : Tumor stiffening, which stems from aberrant production of extracellular matrix and in particular from

collagen fiber network, has been considered a predictive marker of tumor malignancy. Moreover

experimental evidences suggest that tumor rigidity and altered mechanics are, per se, key modulators of

tumor progression. Our group has recently shown that thermal therapy denatures tumor stroma, directly

impacting on tumor permeation properties and drug/nanoparticles penetration1. In a recent work we

aimed to evaluate the therapeutic efficacy of the photothermal therapy (PTT) mediated by carbon

nanotubes2 by correlating tumor growth, stroma integrity and mechanical properties.3 At the microscopic

level, PTT impact on tumor stroma was characterized by biphoton and second‐harmonic generation (SHG)

imaging microscopy. At the macroscopic level, ultrasound elastography was used to assess mechanical

stiffness of the tumor tissue during the PTT treatment and over 10 days after moderate hyperthermia (43°C

‐ 45°C for 20 min, repeated twice) and thermal ablation (50°C for 3 min, repeated twice). We aimed to

determine the best compromise between overheating limitation, thermal damage and outcome on tumor

regression. While non‐treated tumors show increasing stiffness over time, both photothermal treatments

reversed tumor stiffening, while diminishing tumor volume. Such macroscopic features correlated to

microscopic damages of the tumor stroma induced by local heating of CNTs. This study higlights the

potential of ultrasonic shear wave elastography for monitoring the effects of hyperthermia on tumor

tissues via the evolution of their mechanic properties, in correlation with stroma integrity. Moreover it

enlarges the use of ultrasonic elastography as a tool for non‐invasive personalized monitoring method for

tumor therapy. Reversion of tumor stiffening by photothermal treatment could be a promising adjuvant

therapy for controlling tumor progression.3

The long term fate in the body and the biodegradation mechanisms of nanoparticles used for thermal

therapy are still a matter of debate. We will review some of our recent works discussing the degradability

of carbon nanotubes and the one year fate of gold/iron oxide nanoparticles.4,5

REFERENCES :

1. J. Kolosnjaj‐Tabi et al. Heat‐Generating Iron Oxide Nanocubes: Subtle “Destructurators” of the

Tumoral Microenvironment. ACS Nano 2014 8 (5), 4268‐4283

2. I. Marangon et al. Synergic mechanisms of photothermal and photodynamic therapies mediated by

photosensitizer/carbon nanotube complexes. Carbon, 97 : 110‐123 (2016)

3. I. Marangon et al. Tumor stiffening, a key determinant of tumor progression, is reversed by

nanoparticle‐ induced photothermal therapy. Submitted

4. Kolosnjaj‐Tabi J et al. The One Year Fate of Iron Oxide Coated Gold Nanoparticles in Mice

ACS Nano. 9(8):7925‐39 (2015). 5. Elgrabli D et al. Carbon Nanotube Degradation in Macrophages: Live Nanoscale Monitoring and

Understanding of Biological Pathway


DEVELOPMENT OF NEW BIO‐NANOTRANSPORTERS FOR BIOLOGICS DDS

Kazunari AKIYOSHI

Department of Polymer Chemistry, Graduate School of Engineering, Kyoto University,

ERATO JST, Nishikyo‐ku, Kyoto, 615‐8510, Japan

KEY WORDS : Nanogel/ proteoliposome/ exosome/ protein DDS/ We develop a new strategy of preparation of bio‐inspired functional nanoparticles (bio‐nanotransporters) and nanoparticle‐based “bottom‐up” design of biomaterials (functional gels or biointerfaces) for advanced medical technology. In particular, we focus on nanogel tectonics, proteoliposome engineering, and exosome engineering for the new cancer therapies, vaccines and regenerative medicine.

Exosomes are extracellular vesicles secreated by various cells and are known to be related to cell–cell communication through transport of informative constituents, including proteins, lipids, and microRNAs (miRNAs) [1]. For example, we reported the function of exosomes as nanocarriers of pathogen‐associated proteins CagA, which is a major virulence factor of Helicobacter pylori and involves in the development of upper gastrointestinal diseases and also diseases outside the stomach [2]. Recently, exosomes have been attracted growing interest in new drug delivery system of biological molecules. However, it is not always easy to control the delivery of exosomes to various cells and tissues. For applications of exosomal carriers in drug delivery systems, it may become necessary to modify and tune the exosomal interface. We propose new strategy for modification of exosomes with functional polymers or nanogels or hydrogels and also by hybrid with functional liposomes.

We proposed a new method, termed nanogel tectonics, to construct hybrid gel materials with a hierarchical structure for advanced biomedical technology [3]. Nanogels are used as individual components for building nano‐integrated functional hydrogel systems. Nanogel‐crosslinked (NanoClik) microspheres [4] or hybrid hydrogels [5] have been developed via the crosslinking of nanogels. We report here functional polysaccharide nanogel‐exosome hybrids. The cationic nanogels are useful for effective intracellular delivery of exosomes. Various functional self‐assembled nanogels containing cell‐specific, thermo responsive, pH sensitive, and photo responsive nanogels were applicable to use for preparation of functional exosomes [6]. Functionalization of exosomes with nanogel engineering is promising for wide applications of exosomes.

Another new membrane‐engineering strategy to modify the surface of exosomes was reported by using direct membrane fusion between synthetic liposomes and exosomes [7]. This method allows us to optimize the properties of the exosome surface in order to decrease its immunogenicity and increase its colloidal stability, and improve the half‐life of exosomes in blood. Recently, it was reported that exosomes transport not only water‐soluble cargoes like proteins and RNA, but also lipophilic molecules. Therefore, exosomes can carry bioactive lipids such as lipid signals or mediators between proximal or distal cells, and their fate is at least partly regulated by these lipids. Furthermore, exogenous membrane proteins can be incorporated into exosomes using this engineering method. We already reported a new one‐step method for preparing proteoliposome by cell‐free membrane protein synthesis in the presence of liposomes [8]. For example, liposomes containing connexin were developed using this method, and were capable of delivering a hydrophilic oligo‐peptide to cells via connexin‐mediated communication through gap junctions between the proteoliposome and cells [9]. Fusion of the proteoliposome and exosome membranes facilitated the incorporation of the membrane protein into the exosomes. The membrane‐engineering method used here represents a new strategy to yield hybrid exosomes as novel biological nanotransporters (bio‐nanotransporter) for advanced drug delivery systems. The exosome–liposome fusion method should also be useful for loading therapeutic agents into exosomes, a field that is now being researched by our group.


REFERENCES : [1] F. Momose, N. Seo, Y. Akahori, S. Sawada, N. Harada, T. Ogura, K. Akiyoshi, H. Shiku, Guanine‐rich sequences are a dominant feature of exosomal microRNAs across the mammalian species and cell types, PLOS ONE, 4, e0154134 (2016) [2] A. Shimoda, K. Ueda, S. Nishiumi, N. Murata‐Kamiya, S. Mukai, S. Sawada, T. Azuma,M. Hatakeyama, K. Akiyoshi, Exosomes as nanocarriers for systemic delivery of the Helicobacter pylori virulence factor CagA, Scientific Reports, 6, 18346 (2016) [3] Y. Tahara, K. Akiyoshi, Current advances in self‐assembled nanogel delivery systems for immunotherapy, Adv. Drug Deliv. Rev. 95, 65‐76 (2015) [4] Y. Tahara, S. Mukai, S. Sawada, Y. Sasaki, K. Akiyoshi, Nanocarrier‐integrated microspheres: Nanogel tectonic engineering for advanced drug delivery systems, Advanced Materials, 27, 5080‐5088 (2015) [5] Y. Hashimoto, S. Mukai, S. Sawada, Y. Sasaki, K. Akiyoshi, Nanogel tectonic porous gel loading biologics, nanocarriers and sells for advanced scaffold, Biomaterials, 37, 107‐115 (2015) [6] Y.Sasaki, K. Akiyoshi, Self‐assembled nanogel engineering for advanced biomedical technology, Chem. Lett. Highlight review, 41, 202‐208 (2012) [7] Y. Sato, K. Umezaki, S. Sawada, S. Mukai, Y. Sasaki, N. Harada, H. Shiku, K. Akiyoshi, Engineering hybrid exosomes by membrane fusion with liposomes, Scientific Reports, 6, 21933 (2016) [8] M. Ando, M. Akiyama, D. Okuno, M. Hirano, T. Ide, S. Sawada, Y. Sasaki, K. Akiyoshi, Liposome chaperon in cell‐free membrane protein synthesis: One‐step preparation of KcsA‐integrated liposomes and electrophysiological analysis by the planar bilayer method, Biomaterials Science, 4, 258‐264 (2016) [9] M. Kaneda, S. M. Nomura, S. Ichinose, S. Kondo, K. Nakahama, K. Akiyoshi, and I. Morita, Direct formation of proteo‐liposomes by in vitro synthesis and cellular cytosolic delivery with connexin‐expressing liposomes, Biomaterials, 30, 3971‐3977 (2009)


CHARM OF DENDRIMER NANOTECHNOLOGY IN CANCER THERAPY

PENG Ling Centre Interdisciplinaire de Nanoscience de Marseille, Aix‐Marseille University, CNRS, Marseille, FRANCE

KEY WORDS : drug delivery, gene therapy, dendrimer, nanovectors, RNA therapeutics

ABSTRACT : The application of nanotechnology to engineer nanovectors for drug delivery is widely expected to bring breakthrough in nanomedicine and create entirely novel therapeutics.1 Dendrimers are ideal nanocarriers for drug delivery by virtue of their uniquely well‐defined and precisely controlled structure as well as the multivalent cooperativity confined within nanosized volume. We have recently established bio‐inspired structurally flexible dendrimers 2‐4 and self‐assembled supramolecular dendrimers 5‐8 as excellent nanocarriers for gene and drug delivery. In particular, these amphiphilic dendrimers are able to form adaptive supramolecular nanostructures,6‐8 which encapsulate therapeutics with high loading efficiency for effective drug delivery to combat drug resistance. Our studies offer new perspectives in molecular engineering of functional dendrimers in nanotechnology‐based biomedical applications.

REFERENCES :

1) Blanco I, Haifa Shen H, Ferrari M, Nat. Biotechnol. 2015, 941‐951 2) Liu X, Wu J, Yamine M, Zhou J, Posocco P, Viel S, Liu C, Ziarrelli F, Fermeglia M, Pricl S, Vicotrero G,

Nguyen C, Erbacher P, Behr J‐P, Peng L, Bioconjugate Chem. 2011, 22, 2461‐2473 3) Liu X, Liu C, Chen C, Bentobji M, Cheillan FA, Piana JT, Qu F, Rocchi P, Peng L, Nanomedicine:

Nanotechnology, Biotechnology and Medicine, 2014, 10, 1627‐1636 4) Cui Q, Yang S, Ye P, Tian E, Sun G, Zhou J, Sun G, Liu X, Chen C, Murai K, Zhao C, Azizian KT, Yang L,

Warden C, Wu X, D’Apuzzo M, Brown C, Badie B, Peng L, Riggs A, Rossi JJ, Shi Y, Nat. Commun. 2016, 7,10637

5) Yu T, Liu X, Bolcato‐Bellemin A‐L, Wang Y, Liu C, Erbacher P, Qu F, Rocchi P, Behr J‐P, Peng L, Angew. Chem. Int. Ed. 2012, 51, 8478‐8484.

6) Liu X, Zhou J, Yu T, Chen C, Cheng Q, Sengupta K, Huang Y, Li H, Liu C, Wang Y, Posocco P, Wang M, Cui Q, Giorgio S, Fermeglia M, Qu F, Pricl S, Shi Y, Liang Z, Rocchi P, Rossi JJ, Peng L, Angew. Chem. Int. Ed., 2014, 53, 11822.

7) Chen C, Posocco P, Liu X, Cheng Q, Laurini E, Zhou J, Liu C, Wang Y, Tang J, Dal Col V, Yu T, Giorgio S, Fermeglia M, Qu F, Liang Z, Rossi JJ, Liu M, Rocchi P, Pricl S, Peng L, Small. 2016, doi: 10.1002/smll.201503866

8) Wei T, Chen C, Liu J, Liu C, Posocco P, Liu X, Cheng Q, Huo S, Liang Z, Fermeglia M, Pricl S, Liang XJ, Rocchi P, Peng L, Proc. Natl. Acad. Sci. U.S.A. 2015, 112, 2978

ACKNOWLEDGEMENTS : This work has been supported by the European Research projects “DENANORNA” and “Target4Cancer”, Ligue Nationale Contre le Cancer, Fondation pour la Recherche Médicale, Association Française contre les Myopathies, Association pour la Recherche sur les Tumeurs de la Prostate, INCa, Canceropôle PACA, ANR, CNRS and Aix‐Marseille University.


USE OF INFORMATICS TECHNOLOGIES FOR THE PREDICTION OF PHARMACOKINETIC BEHAVIORS

Fumioshi YAMASHITA

Graduate School of Pharmaceutical Sciences, Kyoto University

KEY WORDS : in silico, pharmacokinetics, modeling & simulation, QSAR, PBPK

ABSTRACT : Pharmacokinetic profiles of drugs are crucial in determining their safety and efficacy. In drug discovery and development, therefore, a number of efforts are being made to predict or characterize drug disposition behaviors. “In silico ADME” have often been used as a term representing prediction of drug disposition behavior from chemical structure. It aims to evaluate the risks of efficacy and safety of drug candidates a priori to their chemical synthesis, and is therefore expected to remove the bottleneck from early drug discovery. Quantitative structure‐activity relationship (QSAR) analysis derives a stochastic model that can describe a target physical or biological property from molecular descriptors (steric, electronic, or hydrophobic). Use of informatic approaches such as neural networks genetic algorithm greatly improves a predictive performance as compared to classical QSAR approaches. While such QSAR models enable prediction against each corresponding elementary process in pharmacokinetics, risk assessment must be done through integrative interpretation of these results. A physiologically based pharmacokinetic (PBPK) modeling is a technique in which pharmacokinetics is described with physiological and biochemical parameters such as blood flow rate, protein binding, and drug metabolism. It allows in vitro‐in vivo and interspecies extrapolations of pharmacokinetics from a mechanistic point of view. Packaged PBPK softwares are now available, in which interindividual differences associated with anatomical differences, pathophysiological states, and enzymatic polymorphisms are taken into account. However, models implemented in packaged softwares are difficult to reuse or extend for specialized purposes. Physiological Hierarchy Markup Language (PHML) is a new XML‐based specification to describe a wide variety of models of biological and physiological functions with hierarchical structures. A model is described by a set of functional modules, each of which specifies mathematical expressions of the module functions. The specifications of PHML are compatible in describing PBPK models that are composed of hierarchical kinetic processes. A notable feature of PHML is that SBML models can be included as a PHML module. It allows to reuse SBML models representing subcellular biochemical events without modeling over again by PHML. We demonstrate a SBML‐PHML hybrid model for drug‐drug interaction (DDI) caused by an inducer of drug metabolizing enzyme. Such a modeling & simulation platform greatly helps us to understand pharmacokinetics/pharmacodynamics relationships from systems pharmacology point of view. REFERENCES : 1) Yamashita F, Sasa Y, Yoshida S, Hisaka A, Asai Y, Kitano H, Hashida M, Suzuki H. Modeling of rifampicin‐

induced CYP3A4 activation dynamics for the prediction of clinical drug‐drug interactions from in vitro data. PLoS One. 8:e70330, 2013.

2) Baba H, Takahara J, Yamashita F, Hashida M. Modeling and prediction of solvent effect on human skin permeability using support vector regression and random forest. Pharm Res. 32:3604‐17, 2015.

3) Ose A, Toshimoto K, Ikeda K, Maeda K, Yoshida S, Yamashita F, Hashida M, Ishida T, Akiyama Y, Sugiyama Y. Development of a support vector machine‐based system to predict whether a compound is a substrate of a given drug transporter using its chemical structure. J Pharm Sci. 105:2222‐30, 2016.


CATALYTIC PROPERTIES OF ESTERASE IN SEVERAL ORGANS AND DESIGN OF

PRODRUG AND SOFT DRUG

Teruko IMAI

Faculty of Pharmaceutical Sciences, KUMAMOTO University

KEY WORDS: Prodrug, Hydrolysis, Esterase, Substrate Specificity Species Difference, Intestinal Hydrolysis

ABSTRACT :

The approach of prodrug and “soft” drug/antedrug is useful for resolving formulation, delivery, and toxicity limitations on problematic drugs. A prodrug is a pharmacologically inactive derivative of an active parent drug. The “soft” drug/antedrug have been designed to exert their desired effect locally but they are inactivated in the systemic circulation to reduce unwanted effects. Among small molecular weight drugs clinically used, 5‐7% of these drugs are grouped in prodrug and “soft” drug/antedrug. From 2000 to 2008, 191 new small molecular weight drugs were approved worldwide. Of these, 19.4% of drugs were identified as prodrugs and“soft”drug/antedrug. In recent drug repositioning, these drugs might be becoming an integral part of the drug discovery paradigm.

The metabolic consideration is most important in a design of prodrug and “soft” drug/antedrug. Since esterases are abundantly present in several tissues, most of prodrugs and “soft” drugs/antedrugs are designed to be metabolized by esterases and then fully converted to their active and inactive metabolites. Prodrug, aimed at improving the bioavailability of the parent drug is activated by esterases in the liver and intestine before entering systemic circulation, consequently enough high plasma concentration of parent drug results in an appropriate pharmacological effect. The “soft” drug/antedrug should be stably retained in a target tissue with very rapid inactivation into predictable metabolite in the systemic circulation such as blood and liver.

For bioconversion of prodrugs, carboxylesterase (CES), a serine esterase (active center: Ser‐His‐Glu triard), is the most important esterase due to its abundant ubiquitous presence in most organs. Among CES family (CES1‐CES5), CES1 and CES2 family play the main role for hydrolysis of xenobiotics. However, the intestinal hydrolysis of prodrug often results to less bioavailability, because the parent drug formed in intestinal mucosa is transported into luminal side as well as blood vessel. Human CES1, hCE1, is present in most organs except in intestine, and CES2 isozyme, hCE2, is present in limited organ such as intestine and kidney. So, it is able to design a successful prodrug that is hydrolyzed by hCE1 but not by hCE2. Most ester compounds are hydrolyzed by hCE1, but hCE2 hardly hydrolyses a compound which consists of a relatively large acyl group than alcohol group, for example temocapril and oseltamivir. The such different catalytic property of hCE1 and hCE2 is useful for design of prodrug. Although these characteristics are only in human CES, CESs in other animals including monkey show markedly different characteristics of substrate specificity and tissue expression. Therefore, it is difficult to predict the pharmacokinetics of a prodrug in human from experimental animals.


In particular, the difference of substrate specificity of intestinal CES led to significantly different absorption of prodrug between human and animals. This difficulty of prediction of human intestinal absorption is a bottle neck of development of prodrugs. Thereby, we proposed a predicting system of intestinal hydrolysis during absorption from an intrinsic clearance in human intestinal S9 fraction. A quantitative hydrolysis on the process of intestinal absorption was evaluated by in situ rat intestine single‐pass purfusion experiment. A sigmoidal relation was observed between in situ hydrolysis ratio and in vitro intrinsic clearance. This relation is useful for a prediction of intestinal hydrolysis of prodrug on the process of its trasnport across intesinal mucosa. Recently, we developed the noble intestinal membrane transport system using Caco‐2 cells. Generally, Caco‐2 cells expresses hCE1, although hCE2 is a major CES isozyme in human intestine. In order to prepare the model cell line of human intestine, hCE1 was knocked down and hCE2 was transfected in Caco‐2 cells. The noble Caco‐2 cells expressed only hCE2 and showed the expression of several transporters, indicating that its monolayer could be important in the transport experiment of prodrugs.

The bioconversion of “soft” drug/antedrug should be rapid in systemic circulation to enssure minimum toxicity, hence esterases in liver and blood are important. CES is predominantly available for detoxification of antedrug/”soft” drug in liver, and butyrylcholinesterase (BChE) and paraoxonase (PON; arylesterase) are the main esterases in plasma. Butyrylcholinesterase is abundantly present in human plasma. The catalytic center of BChE is same as CES, but its substrate specificity is markedly different from CES. The hydrolase activity of BChE is increased by presence of Ca and Mg ions, while completely inhibited by Zn, although CES is not affected by divalent cations. On the other hand, although expression level of PON is less than that of BchE in human plasma, it locates on high density lipoprotein (HDL) that easily interacts with hydrophobic ester compounds. PON’s catalytic mechanism (catalytic center: His‐His diard) and substrate specificity is different from CES, and hydrolyzes some of soft drugs that are resistant to CES. Furthermore, Esterase D is present in cytosol fraction of red blood cell. Its catalytic center is Ser‐His‐Asp triard, and it shows different substrate specificity and inhibition properties from CES. Esmorol, a “soft” drug, could be hydrolyzed by Esterase D in red blood cell.

There are useful esterases involved in prodrug activation and antedrug/”soft” drug inactivation in several organs and blood. The accurate information about esterases and an accurate prediction method proceeds a development of new prodrugs and “soft” drugs.

REFERENCES :

1) K. Ohura, K. Tasaka, M. Hashimoto, T. Imai, Distinct patterns of aging effects on the expression and activity of carboxylesterases in rat liver and intestine., Drug Metab Dispos., 42(2):264‒273, 2014

2) Y. Uno, S. Uehara, M. Hosokawa, T. Imai, Systematic identification and characterization of carboxylesterases in cynomolgus macaques., Drug Metab Dispos., 42(12):2002‒2006, 2014

3) F. G. Bahar, T. Imai, Aspirin hydrolysis in human and experimental animal plasma and the effect of metal cations on hydrolase activities. Drug Metab Dispos., 41: 1450‒1456, 2013

4) F. Bahar, K. Ohura, T. Ogihara, T. Imai, Species differences of esterase expression and hydrolase activity in plasma., J. Pharm. Sci., 101, 3264‐3274,2012.

5) T. Imai, K. Ohura, The role of intestinal carboxylesterase in the oral absorption of prodrugs, Curr.Drug Metab. 11, 793‐805, 2010

6) T. Imai, M. Hosokawa Prodrug approach using carboxylesterase activity: Catalytic properties and gene regulation of carboxylesterase in mammalian tissue., J. Pestic. Sci. 35, 229‐239, 2010.


BIOACTIVE GLYCOPOLYPEPTIDE SELF‐ASSEMBLED BIOHYBRID NANOMATERIALS

Sébastien LECOMMANDOUX

UNIVERSITÉ DE BORDEAUX, LCPO, UMR CNRS 5629, BORDEAUX‐INP, 33600 PESSAC, FRANCE

KEY WORDS : Copolymers, Self‐Assembly, Glycopolypeptides, Polymersomes The field of synthetic polypeptides has seen many significant advances in recent years, including studies on block and hybrid copolypeptides that form vesicles, fibrils, and other structures with potential applications in medicine and materials chemistry. However, the development of glycosylated polypeptides has not kept pace, primarily due to the inability to readily synthesize glycopolypeptides in a controlled manner. Glycosylation of natural proteins provides diverse functionality such as mediation of recognition events, modification of protein conformations, ect, that may find interest and application in biomedical field. In this context, we developed over the last years synthetic strategies for the design of glycosylated polypeptides and polysaccharide‐polypeptide biohybrids with controlled placement of sugar functionality. We were especially interested in designing amphiphilic copolymers able to self‐assemble into well‐ defined micelles and vesicles that can advantageously be loaded with drugs and present a surface with multivalent presentation of bioactive saccharides or oligosaccharides. The ability of these nanoparticles for different biomedical applications, from drug‐delivery to inhibitor, will be presented. [1] V. Jeannot, S. Mazzaferro, J. Lavaud, M. Arboléas, V. Josserand, J.‐L. Coll, S. Lecommandoux, C. Schatz, A. Hurbin, Targeting CD44 receptor‐positive lung tumors using polysaccharide‐based nanocarriers: influence of nanoparticle size and administration route, Nanomedicine: Nanotechnology, Biology, and Medicine 4 (2016) 921‐932. [2] A. Peyret, J.F. Trant, C. V Bonduelle, K. Ferji, N. Jain, S. Lecommandoux, E. R Gillies, Synthetic glycopolypeptides: synthesis and self‐assembly of poly (γ‐benzyl‐l‐glutamate)‐glycosylated dendron hybrids, Polymer Chemistry 6 (2015) 7902‐7912. [3] W. Liau, C. Bonduelle, M. Brochet, S. Lecommandoux, A. Kasko, Synthesis, characterization and biological interaction of glyconanoparticles with controlled branching, Biomacromolecules 16 (2015) 284‐294. [4] C. Bonduelle, J. Huang, T. Mena‐Barragán, C. Ortiz‐Mellet, C. Decroocq, E. Etamé, A. Heise, P. Compain, S. Lecommandoux, Iminosugar‐based glycopolypeptides: glycosidase inhibition with bioinspired glycoprotein analogue micellar self‐assemblies, Chem. Commun. 50 (2014) 3350‐3352. [5] C. Bonduelle, S. Lecommandoux, Synthetic glycopolypeptides as biomimetic analogues of natural glycoproteins, Biomacromolecules 14 (2013) 2976‐2983. [6] Huang, J. ; Bonduelle, C. ; Thévenot, J.; Lecommandoux, S. ; Heise, A., Biologically Active Polymersomes from Amphiphilic Glycopeptides, J. Am. Chem. Soc. 134 (2012) 119‐122 [7] Upadhyay, KK.; Bhatt, A.N.; Mishra, A.N.; Dwarakanath, B. S.; Jain, S.; Schatz, C.; Le Meins, JF.; Farooque, A.; Chandraiah, G.; Jain, AK.; Misra, AK.; Lecommandoux, The intracellular drug delivery and anti tumor

activity of doxorubicin loaded poly(‐benzyl L‐glutamate)‐b‐hyaluronan polymersomes, S. Biomaterials 31 (2010) 2882‐2992.


PROMISING CAPABILITIES OF AN ADENOSINE ANALOGUE, COA‐CL

Ikuko TSUKAMOTO and Ryoji KONISHI

Department of Pharmaco‐Bio‐Informatics, Faculty of Medicine, Kagawa University Miki, Kagawa 761‐0793, JAPAN

[email protected]‐u.ac.jp / [email protected]‐u.ac.jp

KEY WORDS : newly synthesized small molecule; COA‐Cl, alternative growth factor

ABSTRACT : COA‐Cl is a newly synthesized adenosine analogue with the molecular weight of 284. It is water soluble and very stable. We have found that it is angiogenic as well as neurotrophic. In this symposium, we will present various physiological activities of COA‐Cl and discuss about the application to regenerative medicine as a xeno free growth factor including DDS. <Angiogenic potency> COA‐Cl promoted tube formation of HUVEC (human umbilical vein endothelial cells) co‐cultured with NHDF (normal human dermal fibroblasts). Tube formation of HUVEC is a model for angiogenesis. We confirmed that not VEGF receptor but S1P receptor 1 contributed to the effects of COA‐Cl following the activation of ERK1/2 in HUVEC. As for NHDF, COA‐Cl increased VEGF production and secretion via cAMP/ PKA/CREB/PGC1� pathway. We also confirmed its angiogenic potency in vivo using chick chorioallantoic membrane (CAM) assay and rabbit cornea assay. <Neurotrophic potency> COA‐Cl promoted neurite outgrowth in PC12 and iPS derived human neuronal cells. In the oxygen/glucose deprivation study with PC12, COA‐Cl suppressed the LDH release. Further, COA‐Cl also suppressed the LDH release caused by H2O2. WB analysis showed that COA‐Cl protected PC12 cells from apoptosis caused by H2O2. Neuroprotective effects of COA‐Cl were also confirmed using a rat stroke model. Moreover, COA‐Cl was revealed to activate tyrosine hydroxylase, a rate limiting enzyme for the biosynthesis of catecholamine. It increased the secretion of dopamine from PC12 in a dose dependent manner. < Application to Regenerative medicine > Nowadays, regenerative medicine is one of the main issues in Japanese medical research. In this field, there are 3 main steps. The 1st step is the collection of healthy human cells from patients or volunteers. The 2nd step is the preparation of proper cells or tissues via cell culture in clean rooms. In some cases, cells are resetted to iPS cells and then differentiated to the purpose cells before preparing tissue or cell sheets. And the 3rd step is the transplantation of the products to the patients. In the 2nd step, different from our usual cell culture experiments, xeno free trophic factors are eagerly required and the usage of animal sera should be refrained. Here, the application of COA‐Cl to this field is very promising since COA‐Cl itself has VEGF‐like and NGF‐like potencies. It can be used as an alternative xeno free growth factor. In addition, it can promote endogenous VEGF production from fibroblasts. Further, COA‐Cl is a very stable compound. We can store it at room temperature for more than several months either in powder or solution form.

Commercially available as "2Cl‐C.OXT‐A" from WAKO


RATIONALE FOR THE USE OF CATIONIC EMULSIONS IN OPHTHALMOLOGY : FROM BENCH TO BEDSIDE

Jennifer DOURLAT, Jean‐Sébastien GARRIGUE

Innovation Center, Santen SAS

KEY WORDS : Ocular drug delivery, eye, ophthalmology, dry eye, emulsion, cyclosporine ABSTRACT :

Topical delivery has always been an issue in term of drug penetration. In the early 2000, emulsions have gained interest in ophthalmology with the launch of the first emulsion dedicated to the treatment of an eye condition. In the continuing effort of improving ocular bioavailability of drugs, cationic emulsions were then developed with new features. This positively charged dosage form has the property to interact with the negatively charged ocular surface providing a longer retention time and an enhanced tissue penetration. This original and patented concept was applied to the delivery of several actives of which cyclosporine A (CsA). CsA is a very lipophilic molecule which is very difficult to formulate as an eye drop. Cationic emulsions are adequate formulations to administer this molecule to the eye. However, cationic agents were often considered as deleterious to ocular surface and emulsions are unstable systems. A wide formulation development led to the choice of a safer cationic agent, cetalkonium chloride and very stable nanoemulsions designated as Novasorb® technology. Safety and ocular pharmacokinetics were evaluated in rabbits with a two‐fold increased penetration in cornea and conjunctiva compared to anionic emulsions and an improved healing of ocular surface lesions. Clinical trials revealed good tolerability and safety in patients and significant clinical efficacy in the treatment of severe dry eye with a once‐a‐day dosing regimen. This cationic cyclosporine A emulsion is now in the late stage of development.


FORMULATION DESIGN OF COMPOSITE NANO‐PARTICLES FOR INHALATION BY USING A TWO‐SOLUTION MIXING TYPE SPRAY NOZZLE

Ryo Maeda1, Tatsuaki Tagami1 , Takemasa Takii1, Jennifer Wong2, Philip Chi Lip Kwok3, Hak‐kim Chan2, and Tetsuya OZEKI1

1 Nagoya City University, 2 University of Sydney, 3 University of Hong Kong

KEY WORDS : Nano‐particles, Two‐solution mixing nozzle, Anti‐solvent, Inhalation

ABSTRACT : The marketing of inhaled formulation is expanding within the last decade and the inhaled formulations including inhalation devices have been actively developing. While the target diseases are mostly focused on lung related diseases (asthma, COPD), the inhaled formulation for the systemic diseases (inhaled insulin) is on market and other lung‐related diseases have focused attention. We previously developed “two‐solution mixing type nozzle” to prepare nanocomposite particles (Figure). Nanocomposite particles which means microparticles containing drug nanoparticles are useful for the preservation and handling of nanoscale particles. Nanoscale drug particles has advantage in the point of drug dissolution and bioavailability. The two solution mixing type nozzle can prepare nanocomposite particles in only one step, being useful for the simplicity and scalability. This spray nozzle is composed of two passages and the mixing part. After water solution (ex. sugar alcohol solution) and organic solution (ex. poorly water soluble drug solution) were flown in different passages, respectively, they are mixed in mixing part. The drug crystal starts grows there, as solvent and water are mixing, and they were immediately spray dried before the drug crystal have been grown.

We applied the technique to prepare nanocomposite particles for 1) improved drug dissolution and bioavailability, 2) taste masking and 3) inhaled formulation. Recently, 4) the nanocomposite particles for tuberculosis have been prepared by using two solution mixing type nozzle. After the nanocomposite particles are inhaled, they are expected to reach alveolar region where tuberculosis bacterium is hidden in alveolar macrophages. After reaching alveolar region, the nanocomposite particles are dissolved and drug nanoparticles exposed are expected to be taken up by alveolar macrophages. In our study, a high‐water soluble compound, sugar derivative that has effect against drug‐resistant tuberculosis bacterium was coated with lipophilic emulsifier by using dry emulsion method to deliver the drug into alveolar


macrophages at high concentration. The nanocomposite particles could remarkably accumulated into macrophage‐like cell line and alveolar macrophages in vivo. In the latter part, the other type of spray‐dried particles were prepared. The “nano‐matrix particles” which means nano‐porous microparticles could be prepared by two‐solution mixing type nozzle. The nano‐matrix particle is consist of only one compound (ex. water soluble drug) and the pore size on the particles can be controllable. The high purity and light nano‐matrix particles will be useful for inhaled formulation. REFERENCES :

1) Moeko Taki, Tatsuaki Tagami, Kaori Fukushige and Tetsuya Ozeki, Fabrication od nanocomposite particles using a two‐solution mixing‐type spray nozzle for use in an inhaled curcumin formulation, Int. J. Pharm., in press

2) Yukiko Nishino, Takanori Kanazawa, Yuuki Takashima, Tetsuya Ozeki, and Hiroaki Okada, Improved

intestinal absorption of a poorly water‐soluble oral drug using mannitol microparticles containing a nano‐solid drug dispersion, J.Pharm. Sci., 101(11), 4191‐4200( 2012)

3) Tetsuya Ozeki, Yusuke Akiyama, Norimitsu Takahashi, Tatsuaki Tagami, Toshiyuki Tanaka, Masashi

Fujii, and Hiroaki Okada, Development of Novel and Customizable Two‐Solution Mixing Type Nozzle for One‐step Preparation of Nanoparticle‐containing Microparticles, Biol. Pharm. Bull., 35(11), 1926‐1931(2012).

ACKNOWLEDGEMENTS : Ohkawara Kakohki Co., Ltd.


DEVELOPMENT OF VASCULARIZED 3D TISSUE MODELS USING LAYER‐BY‐LAYER TECHNIQUE AND APPLICATION FOR DDS RESEARCH

Mitsuru AKASHI1, Takami AKAGI1, Simona MURA2, Patrick COUVREUR2

1Graduate School of Frontier Bioscience, Osaka University, Japan. 2Institut Galien Paris‐Sud, Faculté de Pharmacie, Université Paris‐Sud.

KEY WORDS : Cell manipulation, three‐dimensinal tissue, extracellular matrix, cancer model

ABSTRACT : The appropriate construction technique for a three‐dimensional (3D) tissue is required to enhance the potential of cells to form engineered tissues. Recently, various techniques have been developed to construct 3D multilayered tissues, for example cell sheet engineering and multilayer scaffolds. We recently reported a simple and unique bottom‐up approach, termed “cell‐accumulation technique”, to develop 3D cellular multilayers with the desired layer number and location by the fabrication of layer‐by‐layer (LbL) fibronectin (FN)‐gelatin (G) (FN‐G) films onto the cell surfaces1). Less than 10 nm thickness of ECM films composed of FN‐G allowed all cells to adhere to each other through interactions between the FN‐G nanofilms and the cell membrane proteins to create various types of tissues, such as blood vessel wall, liver and heart2). Using this technique and a sandwich culture, highly dense and homogeneous endothelial tubular networks were formed in the 3D tissues. These hierarchical cell manipulations will be promising to achieve in vitro creation of artificial 3D‐tissue models and their application in drug effect, toxicology (especially for alternative systems to animal testing) and tissue engineering. In a preclinical study for drug developments, 2D cell culture models and animal models have been

employed. However, these data are not always a good predictor of human researches due to the differences of drug response between 2D cultured models and living tissues with 3D structures, and species difference. Especially, the oversimplified 2D cancer models do not reflect the 3D cell organization of in vivo solid tumors and thus disrupt crucial cell‐matrix interactions responsible for cancer cell invasion. As for in vivo models, they limit the monitoring of tumor growth or regression over a course of treatment. In this study, we applied the vascularized 3D tissue models for evaluation of proliferation and invation behavior of cancer cells, and effect of anti‐cancer drugs to 3D cancer models. These models can bridge this gap (2D vs 3D, vitro vs vivo), functioning as reliable models not only for cancer biology studies but also to serve as an efficient anti‐cancer drug screening platform.

Fig. 1 Evaluation of therapeutic efficacy of cancer drugs

using 3D cancer model.


REFERENCES : 1) A. Nishiguchi, H. Yoshida, M. Matsusaki, M. Akashi, Rapid construction of three‐dimensional

multilayered tissues with endothelial tube networks by the cell‐accumulation technique, Adv. Mater., 23, 3506‐3510 (2011).

2) Y. Amano, A. Nishiguchi, M. Matsusaki, H. Iseoka, S. Miyagawa, Y. Sawa, M. Seo, T. Yamaguchi, M. Akashi, Development of vascularized iPSC derived 3D‐cardiac myoblast tissues by filtration layer‐by‐layer technique and their application for pharmaceutical assays, Acta Biomater., 33, 110‐121 (2016).


SIMULATION OF CONTROLLED DRUG DELIVERY SYSTEMS IN VITRO AND IN VIVO

Juergen SIEPMANN and Florence SIEPMANN

Univ. Lille, Inserm, CHU Lille, U1008 ‐ Controlled Drug Delivery Systems and Biomaterials, F‐59000 Lille, France

KEY WORDS: Controlled drug delivery; mathematical modeling; in‐silico simulations

ABSTRACT:

Since the famous Professor Takeru Higuchi published his seminal paper in 1961 on the quantitative description of drug release from an ointment film (introducing the “Higuchi Equation”), mathematical modeling has become more and more popular in pharmaceutics. This can be explained by the crucial benefits in‐silico simulations of drug delivery systems can offer, including: (i) a better understanding of how a particular drug product releases the incorporated active agent, and (ii) the possibility to quantitatively predict the impact of the device design (e.g., composition, geometry and dimensions of the system) on the resulting drug release kinetics (and, ideally, also on the resulting pharmacokinetics and pharmacodynamics). Thus, based on rapid and inexpensive mathematical calculations, time‐consuming and cost‐intensive series of trial‐and‐error experiments can be avoided. This is particularly helpful, if long release periods (e.g. several years in the case of implants for long term treatments) are targeted.

Various mathematical theories have been proposed in the literature for a broad range of pharmaceutical dosage forms. This includes purely empirical (only descriptive) mathematical models (which do not offer any insight into the underlying mass transport mechanisms), mechanistically realistic theories (allowing to better understand how a specific system works), and semi‐empirical models (hybrid forms of these of two categories).

It has to be pointed out that certain drug delivery systems are relatively easy to describe, since eventually only one particular mass transport phenomenon is dominant (while all other involved processes can be neglected without introducing considerable errors). Examples for physico‐chemical phenomena, which might play a role in the control of drug release from a pharmaceutical dosage form include: water penetration into the system, polymer swelling, drug dissolution, drug diffusion through a polymeric network and/or water‐filled pores, polymer dissolution and/or degradation, osmotically driven water influx, creation of hydrostatic pressure acting against film coatings, potential crack formation, convective mass transport through cracks, local drops in micro‐pH, autocatalytic effects [e.g. in the case of poly(lactic‐co‐glycolic acid)‐based delivery systems], limited drug solubility effects, drug‐polymer interactions and drug re‐precipitation, to mention just a few.

To be able to identify, which particular phenomena are of importance in a specific type of drug delivery system, the latter should be thoroughly characterized before and after exposure to the release medium (or aqueous body fluids). Ideally, a broad range of experimental characterization techniques should be applied, such as in vitro drug release measurements under various conditions, X‐ray diffraction, differential scanning calorimetry, optical microscopy, scanning electron microscopy, dry mass loss and water uptake studies, gel permeation chromatography, synchrotron measurements, texture analysis, and long term stability studies.


Based on these experimental results, a mathematical theory can be developed and fitted to the experimental results. But caution must be paid at this stage: Even if good agreement between a fitted mathematical theory and experimental results is obtained, this is not a proof for the validity of the hypotheses the model is based on. This is particularly true, if several model parameters are fitted simultaneously. To evaluate the validity of a mathematical theory, it should be used to quantitatively predict the effects of certain device design parameters (e.g., of the drug loading or the dimensions of the system) on the resulting system performance. These theoretical predictions must then be compared with independent experimental results.

In this talk, several practical examples are given, including mathematical theories quantifying drug release from oral controlled release matrix tablets (based on different types of polymers) and lipid implants for controlled protein release. It has to be pointed out that very often in‐silico simulations of the impact of the device design of a controlled drug delivery system are limited to the in vitro performance of the systems (essentially to the in vitro drug release kinetics). Yet, there is a crucial lack of more comprehensive mathematical theories, allowing to predict the impact of the systems’ composition, geometry and dimensions also on the device performance in vivo: the resulting pharmacokinetics and pharmacodynamics (PK and PD). On the other hand, various (partially highly sophisticated) PK and PD models are available, but they generally only consider very much simplified drug release kinetics from a pharmaceutical dosage form.

This talk will also give an example for a combination of a mechanistically realistic mathematical theory describing drug release out of controlled release hot melt extrudates and the subsequent drug fate in the living organism, upon oral administration. The theoretical calculations are compared with experimental results obtained in vitro and in vivo. This type of in‐silico simulations has the potential to replace in vivo trials by theoretical calculations. In addition, the consequences of worst case scenarios can be predicted and, hence, the safety of pharmacotherapies be improved. REFERENCES:

Higuchi, T, 1961. Rate of release of medicaments from ointment bases containing drugs in suspensions. Journal of Pharmaceutical Sciences 50, 874‐875.

Siepmann, J; Siepmann, F. Mathematical modeling of drug delivery. International Journal of Pharmaceutics 364, 328‐343, 2008.

Siepmann, F; Herrmann, S; Winter, G; Siepmann, J. A novel mathematical model quantifying drug release from lipid implants. Journal of Controlled Release 128, 233‐240, 2008.

Siepmann, J; Siepmann, F. Modeling of diffusion controlled drug delivery. Journal of Controlled Release 161, 351‐362, 2012.

Siepmann, J; Siepmann, F. Mathematical modeling of drug dissolution. International Journal of Pharmaceutics 453, 12‐24, 2013.

Siepmann, J; Karrout, Y; Gehrke, M; Penz, FK; Siepmann, F. Predicting drug release from HPMC/lactose tablets. International Journal of Pharmaceutics 441, 826‐834, 2013.

Siepmann, J. In‐silico simulations of advanced drug delivery systems: What will the future offer? International Journal of Pharmaceutics 454, 512‐516, 2013.

ACKNOWLEDGEMENTS:

The authors are grateful for the support of this work by the “INTERREG V 2 Seas Mers Zeeën Cross‐border Cooperation Programme (2S01‐059‐IMODE)”.


DESIGN OF POLYMERIC NANOPARTICLE NAD MICELLE FOR TREATMENT OF BIOFILM INFECTION DISEASE

Hiromitsu YAMAMOTO, Chisato Takahashi, Noriko Ogawa

Aichi Gakuin University

KEY WORDS : Biofilm infection desease, PLGA nanoparticle, Polymeric micelle, Chitosan

ABSTRACT : Introduction Biofilm is formed by microorganisms for growing on a solid substrate. Biofilms are characterized by structural heterogeneity, genetic diversity, complex community interactions, and an extracellular matrix of polymeric substances. Bacteria living in a biofilm can have significantly different properties from free‐floating bacteria, as the dense and protected environment of the film allows them to cooperate and interact in various ways. This environment provides the resistance to detergents and antibiotics, as the dense extracellular matrix and the outer layer of cells protect the interior of the community. These biofilms cause the intractable infections. Therefore, the development of antibacterial drug delivery systems against the biofilm infection disease is strongly desired. We successfully established the DDS platform with poly (lactic‐co‐glycolic acid) nanoparticle (PLGA NPs) and are evaluating their functions for drug delivery device via orally, pulmonary and so on. In this paper, we intended to design the surface modified PLGA NPs and polymeric micelles for antibacterial drug delivery to the microorganisms in the biofilm. Method Preparation of PLGA NPs and polymeric micelles: PLGA NPs and polymeric micelles with Soluplus® were prepared by the emulsion solvent diffusion method. Chitosan (CS) modification on the surface of drug carrier was carried out by adding CS solution into outer aqueous solution during preparation process. Particle size and zeta potential were measured with Zetasizer (ZS90, Malvern inc). Formation of biofilm: Biofilm was prepared by incubation for 24hr after seeding Staphylococcus epidermidis in 12‐well plates. Evaluation of adsorption of drug carrier to biofilm: Fluorescence labeled drug carrier suspension was added into culture medium after biofilm formation. After incubation for 2 hours, biofilm was rinsed with distilled water. Obtained biofilm was observed with confocal laser scanning microscope. Fluorescence die was extracted from biofilm and quantitatively determined with fluorophotometer. Observation of biofilm treated with PLGA NPs using FE‐SEM: After growth of the biofilm in a 24‐well plate, it was washed with purified water and then treated with NP suspensions (N‐PLGA, CS‐PLGA and CAM‐PLGA NPs) at concentrations of 0.5, 1.0 and 3.0 mg/ml in TSB medium. After treatment with NPs, the plates were incubated for 2 h at 37°C in 0.5% CO2. The surface morphology of the biofilms was observed using FE‐SEM (JXA‐8530FA; JEOL Co., Japan). The incubated biofilms treated with NPs were added to phosphate‐buffered saline (2 ml) solution for 2 h. The biofilm was washed with purified water and treated with the IL solution. Following this, it was scraped using a cell scraper. The biofilm for observation using FE‐SEM was skimmed off using a copper mesh with carbon‐coated plastic microholes for organic samples. Results Particle size of CS coated PLGA NPs and Soluplus® micelles were ca. 390 nm and 80 nm, respectively. Zeta potentials were 31.6 mV and 15.3mV. Adsorbed amount of drug carriers on the biofilm were increased by CS‐modification. Furthermore, Soluplus micelles indicated the higher adsorption amount compared with PLGA NPs. Particle size of Soluplus® micelles was smaller than that of PLGA NPs. Biofilm has sponge like structure. Therefore, Soluplus® micelles with smaller size could access to the deeply part of biofilm. Clarithromycin(CAM)‐loaded drug carrier showed the antibacterial activity against biofilm formed by microorganisms. Especially, CS‐modified carrier indicated the more effective antibacterial activity and decreased the biofilm formation (Fig. 1).


Fig.2 shows the biofilm structure without any treatment (a) and after treatment with CS modified PLGA NPs (Ref.1). We can see the agglomerated bacterial flora covered by the thick film comprised EPS. Bacterial cells c.a.1 μm in size could not be observed because of existence of the thick film in Fig. 2‐a. The rough surface can be seen at high magnification. It was clear that the biofilm had holes <500 nm in size on its surface. These holes are well known to be water channels, which allow the diffusion of nutrients and oxygen through the biofilm agglomerate. FE‐SEM images of the biofilm treated with CS‐PLGA NPs are shown in Fig. 2‐b. The retraction of biofilm and the shape of bacteria could be observed in Fig.2‐b and the biofilm became thinner. Furthermore, we can see the adherence and penetration activity of CS‐PLGA NPs to the biofilm. It should be noted that the size of the bacterial cell and CS‐PLGA NPs are 1 and 294.6 nm, respectively. A void derived from erosion of the thick film of the biofilm was observed. The configuration of bacterial cell was drastically changed due to the antibacterial effect of CS‐PLGA NPs penetrating into the biofilm, eliminating the thick film and bacterial cells. Compared with N‐PLGA NPs, CS‐PLGA NPs showed a high antibacterial effect; PLGA NPs with a highly positive zeta potential can adhere to the biofilm. It seemed that the biofilm with negative charge attract positively charged CS‐PLGA NPs. CS‐PLGA NPs caused such retraction of biofilm and adhered cell surface, the antibacterial drug could be directly deliver to the bacteria. According to these behavior, CS modified drug carrier indicated strongly antibacterial activity of clarithromycin.

a) w/o treatment b) treated with CS PLGA NPs

PLGA NPs

Bacteria

Fig. 2 SEM images of biofilm after treated with CS PLGA NPs

REFERENCES :

1) C. Takahashi et al., Microscopy (Oxf) 169‐180 (2015)

Fig. 1 Antibacterial effect of clarithromycin loaded Solu- micelle and PLGA nanoparticle

CAM soln. CAM loaded PLGA NP

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NEW THERAPEUTIC NANO‐VECTORS ELABORATION STRATEGIES BASED ON SOFT CHEMISTRY

Cédric BOISSIERE, Clément SANCHEZ

LCMCP, Université Paris 6, CNRS, Collège de France

KEY WORDS : Vector, mesoporous, silica, Sol‐Gel, hybrid ABSTRACT :

Nano‐vectors are fairly recent nanomaterials aiming at helping diagnosis and treatment of heavy

diseases such as cancer. The huge research effort produced in the last fifteen years achieved an

impressive number of different organic or hybrid organic/inorganic nanomaterials integrating one

or several functionalities ranging from contrasting agent (helping at imaging diagnosis) or therapy

(drug release, hyperthermia, radio‐sensitising agent, etc.). Yet, so far, commercialized nanovectors

are fairly simple and organic‐based nanomaterials. The reason of this discrepancy comes from the

fact that multifunctional platforms (either organic or hybrid) require expensive and delicate

synthesis pathways, and/or use complex chemical compositions that strongly hinder their

pharmaceutical development and safety. In this presentation will be shown recent developments

of synthesis strategies of silica‐based nanovectors. The adequate coupling of soft‐chemistry and

processing allows developing in very simple ways multifunctional platforms with a constrained

number of constituents that offer a very good functionality/synthesis_complexity compromise and

are realistic from pharmaceutical development point of view.


X RAY MICRO CT IMAGING OF PENETRATED MICRONEEDLES

Yuji MAKINO1, C.Kato1, T.Kunimoto2, K.Kobayashi3, M.Ishibashi3, H.Hamamoto3

1Musashino University, Tokyo ; 2Tokushima Bunri University, Kagawa ; 3Medrx Co. Ltd., Kagawa

Microneedle, X ray micro CT, In vivo 3D imaging, Penetration depth ABSTRACT Microneedle‐mediated transdermal and intradermal drug delivery has attracted considerabke attentions as a breakthrough in the fields and clinical studies of influenza vaccinations are now going on in U.S.A.. However, many key factors that may affect therapeutic efficacies remain unsolved. Accurate measurement of penetration depths of all microneedles in an array, which is crucial to design specifications of microneedle arrays and application devices, is such an issue. Though optical coherence tomography (OCT) was shown to have potentials to assess the depth 1), 2), its application was limited to arrays made of photo‐permeable materials due to laser source. Therefore, X ray microCT was studied in vitro and in vivo to measure penetration depths by microneedles made of photo‐impermeable materials such as non‐transparent polymers or metals. An array was composed of 10 x12 microneedles (conical, two stages, height: 600μm, base diameter: 180μm) and a basement (square, thickness: 750μm), those composed of photo‐impermeable polymers. The array was attached to a spring type applicator and manually applied to animal skin at specific forces. In in vitro studies, after the arrays were applied to abdomens of minipigs (Goettingen, female, 6months), they were sacrificed and the areas were excised, followed by the CT observations. In in vivo imaging study, the array was manually applied to abdomen of a hairless mouse (male, 5weeks, anesthetized) by the applicator at 20N force. The mouse was then covered with lead film except the applied area and mounted on the stage of the CT. The CT images were recorded using TDM1000H‐II (2K) (Yamato Scientific Co., Ltd.) and analyzed using software myVGL 2.2 (Volume Graphics GmbH). Clear 3D images of penetrated microneedles were obtained in both in vitro and in vivo imaging studies. From the images, penetration depths of all microneedles in an array were read. As the images showed the depths and their variations in an array‐plane directly, they were useful in designing specifications of microneedles and applicators. In in vitro studies, the average penetrating depths by manual applications at 20, 40, 60N were 252±62, 294±64 and 287±66 μm, respectively. All of the needles were inserted to more than 100μm depth, which expects feasibility of the arrays. In vivo imaging study gave the similar images with high resolution, though the hairless mouse was anesthetized. The method was also successfully applied to a stainless needle. 1) Donnelly RF, et al. J Control Release. 2010;147:333‐341 2) Coulman SA, et al. Pharm Res. 2011;28:66‐81 This work was supported by Kagawa Industry Support Foundation and Ministry of Economy, Trade and Industry, Japan.


Career of speakers

and observers


NAME : Mitsuru AKASHI AFFILIATION AND ADDRESS: Graduate School of Frontier Bioscience, Osaka University Biosystems Builiding E704, 1‐3 Yamadaoka, Suita 565‐0871, Japan TELEPHONE N° : +81‐6‐6105‐5247

FAX N° : +81‐6‐6105‐9712

E‐MAIL : [email protected]‐u.ac.jp

CAREER HISTORY : M. Akashi is the Specially‐appointed Professor of Graduate School of Frontier Bioscience, Osaka University. He received his PhD degree in Engineering from Osaka University in 1978. He was a post‐doc at NIH, Gerontology Research Center (USA) and the University of Waterloo (Canada) in 1978‐1980. He joined the Department of Applied Chemistry and Chemical Engineering, Faculty of Engineering, Kagoshima University as an assistant professor in 1981. He was promoted to associate professor in 1984 and a full professor in 1989. From 2003 to 2015, he was a full professor for the Department of Applied Chemistry, Graduate School of Engineering, Osaka University. He received the Award of the Society of Polymer Science, Japan (1999), the Award of Japanese Society for Biomaterials (2004) and the Award of the Chemical Society of Japan (2014). SUMMARY OF PRESENT WORK : Functional biomaterials have been developed by employing weak interactions such as van der Waals interactions. By applying this technology to cell engineering, various types of three‐dimensional (3D) tissue models were constructed by layer‐by‐layer (LbL) assembly technique of extracellular matrix (ECM) on cell surface. The 3D tissues with vascular networks were constructed by a sandwich culture of endothelial cells between multi‐layers of human fibroblasts. These hierarchical cell manipulations will be promising to achieve in vitro creation of artificial 3D‐tissue models and their application in drug effect and toxicology. MAJOR PUBLICATIONS : 1) D. Hikimoto, A. Nishiguchi, M. Matsusaki, M. Akashi, High‐throughput blood‐ and lymph‐capillaries with

open‐ended pores which allow the transport of drugs and cells, Adv. Healthcare Mater., in press. 2) P. Chetprayoon, M. Matsusaki, U. Yokoyama, T. Tejima, Y. Ishikawa, M. Akashi, Use of three‐

dimensional arterial models to predict the in vivo behavior of nanoparticles for drug delivery, Angew. Chem. Int. Ed., 128, 4537‐4542 (2016).

3) Michiya Matsusaki, C. P. Case, Mitsuru Akashi, Three‐dimensional cell culture technique and pathophysiology, Adv. Drug Deliv. Rev., 74, 95‐103 (2014).

4) T. Akagi, T. Fujiwara, M. Akashi, Rapid fabrication of polylactide stereocomplex using layer‐by‐layer deposition by inkjet printing. Angew. Chem. Int. Ed., 51, 5493‐5496 (2012).

5) M. Matsusaki, H. Ajiro, T. Kida, T. Serizawa, M. Akashi, LbL assembly through weak interactions and their biomedical applications. Adv. Mater., 24, 454‐474 (2012).


NAME: Kazunari AKIYOSHI

AFFILIATION AND ADDRESS :

Department of Polymer Chemistry, Graduate School of Engineering

Kyoto University

Kyotodaigaku Katsura, Nishikyo‐ku, Kyoto, 615‐8510, Japan

TELEPHONE No: +81‐75‐383‐2589

FAX No: +81‐29‐853‐2590

E‐MAIL: [email protected]‐u.ac.jp

CAREER HISTORY : 2011 Research Director, ERATO Akiyoshi Bio‐nanotransporter project, JST, Japan 2010 Professor, Graduate School of Engineering, Kyoto University, Japan 2002 Professor, Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Japan 1999 ‐ 2002: Research member, PRESTO, JST, Japan 1997 Visiting Professor, Department of Chemistry, Luis Pasteur University, France 1993 Associate Professor, Graduate School of Engineering, Kyoto University, Japan 1989 Assistant Professor, Faculty of Engineering, Kyoto University, Japan 1987 Lecturer, Faculty of Engineering, Nagasaki University, Japan 1985 Postodoctral fellow (Professor Ei‐ichi Negishi), Purdue University, USA 1985 Ph.D, Faculty of Engineering, Kyushu University, Japan SUMMARY OF PRESENT WORK : Bio‐inspired nano‐organized system, Nanogel engineering for DDS and tissue engineering, Chaperon‐inspired materials, Protein delivery system for immunotherapy, Cancer vaccine, Mucosal vaccine, Nanogel‐integrated hydrogel for bone regeneration, Organic–inorganic hybrid materials, Artificial cell by liposome engineering, Membrane protein engineering. MAJOR PUBLICATIONS: 1) Y. Sasaki, K. Akiyoshi, Nanogel engineering for new nanobiomaterials: From chaperoning engineering to biomedical applications, Chemical Record, 10, 366‐376(2010)

2) T. Nochi, Y. Yuki, H. Takahashi, S. Sawada, M. Mejima, T. Kohda, N. Harada, G. Kong, A. Sato, N. Kataoka, D. Tokuhara, S. Kurokawa, Y. Takahashi, H. Tsukada, S. Kozaki, K. Akiyoshi, H. Kiyono, Nanogel antigenic protein delivery system for adjuvant‐free intranasal vaccines, Nature Materials, 9, 572‐578 (2010).

3) D. Muraoka, N. Harada, T. Hayashi, Y. Tahara, F. Momose, S. Sawada, S. Mukai, K. Akiyoshi, H. Shiku, A Nanogel‐based Immunologically Stealth Vaccine Targets Macrophages in the Medulla of Lymph Node and Induces Potent Anti‐Tumor Immunity, ACS Nano, 8, 9209‐9218 (2014)

4) Y. Tahara, S. Mukai, S. Sawada, Y. Sasaki, K. Akiyoshi, Nanocarrier‐integrated microspheres: Nanogel tectonic engineering for advanced drug delivery systems, Advanced Materials, 27, 5080‐5088 (2015)

5) T. Niwa, Y. Sasaki, E. Uemura, S. Nakamura, M. Akiyama, M. Ando, S. Sawada, S. Mukai, T. Ueda, H. Taguchi, K. Akiyoshi, Comprehensive study of liposome‐assisted synthesis of membrane proteins using a reconstituted cell‐free translation system, Scientific Reports, 5, 18025 (2015)

6) R. Kawasaki, Y. Sasaki, K. Katagiri, S. Mukai, S. Sawada, K. Akiyoshi, Magnetically guided protein transduction by hybrid of nanogel chaperone with iron oxide nanoparticles, Angew. Chem. Int. Ed. In press DOI: 10.1002/anie.201602577


NAME : Philippe BARTHELEMY

AFFILIATION AND ADDRESS: University of Bordeaux, ARNA laboratory, F‐33000 Bordeaux, France. 2INSERM, U1212, ARNA laboratory 3UMR CNRS 5320

TELEPHONE No: 33 (0)5 57 57 48 53,

FAX No: 33 (0)5 57 57 10 15

E‐MAIL: [email protected]

CAREER HISTORY : Barthélémy received his doctorate in chemistry from the University of Montpellier II, France in

1993. He was then a postdoctoral fellow at Emory University in the group of Pr Fredric Menger

(Lavoisier Grant and Emory Fellowship). In 1995 he was appointed as a temporary lecturer at the

University of Avignon and as Associate Professor at the same University in 1996. P. Barthélémy

worked also as a Visiting Associate Professor at Duke University in 2001. In 2005 he was appointed

as full Professor at the University of Bordeaux Segalen. He is leading the “ChemBioMed” team of

the INSERM U1212. Philippe Barthélémy was Vice President of the University of Bordeaux Segalen

(2011‐2013).

SUMMARY OF PRESENT WORK : One of the major objectives of his research is to develop novel bio‐inspired amphiphiles for

biomedical applications.

MAJOR PUBLICATIONS :

1) K. Oumzil, M. A. Ramin, C. Lorenzato, A. Hemadou, J. Laroche, M. J. Jacobin‐Valat, S. Mornet, C.‐E. Roy, T. Kauss, K. Gaudin, G. Clofent‐Sanchez, and P. Barthélémy (2016) Bioconjugate Chemistry 27 (3), pp. 569‐575.

2) Latxague, L., Ramin, M.A., Appavoo, A., Berto, P., Maisani, M., Ehret, C., Chassande, O., and Barthélémy, P. (2015) Angewandte Chemie, 54 (15), pp. 4517‐4521.

3) Patwa A. ; Labille, J. ; Bottero JY. ; Thiéry, A.; and Barthélémy, P. (2015) Chem. Commun. 51 (13), pp. 2547‐2550.

4) Oumzil, K. Benizri, S. Tonelli, G. Staedel, C. Appavoo, A. Chaffanet, M. Navailles L. and Barthélémy P. (2015) ChemMedChem 10 (11), pp. 1797‐1801.

5) Luvino, D. ; Khiati, S. ; Oumzil, K. ; Rocchi, P. ; Camplo, M. ; and Barthélémy P. (2013) J. Control. Release, 172, 954–961.

6) Khiati, S., Luvino, D., Oumzil, K., Chauffert, B., Camplo, M. and Barthélémy P. (2011) ACS Nano, 5 (11), pp 8649–8655. 7) Godeau, G., Bernard, J., Staedel, C., Barthélémy, P. (2009) Chem.Commun., 5127‐5129.


Name: Hassan BENAMEUR

AFFILIATION AND ADDRESS: CAPSUGEL R&D Strasbourg Parc d’Innovation, 180 Rue Tobias Stimmer, BP 30442 67400 Illkirch‐Graffenstaden, France

DIRECT LINE No: +33 (0)3 89 20 58 71 ASSISTANT No: +33 (0)3 89 20 49 86 E‐MAIL: [email protected]

CAREER HISTORY : Hassan Benameur, Ph.D., President of Capsugel France: he joined Capsugel R&D in 2002 and currently leads the Pharmaceutical Sciences department. Dr. Benameur is a Chemical Engineer and holds a PhD in Pharmaceutical Sciences from the Free University of Belgium. He is also a lecturer at several academic and industrial symposia (US, Europe, China, Japan) and was presented the Academy of Pharmaceutical Science and Technology, Japan Award in 2005. Dr. Benameur is a member of several major scientific associations (AAPS, APGI, BCRG, PSTJ and CRS) and scientific academies, and is also the author of 40 scientific publications and the inventor of 20 patents. Prior to joining Capsugel, Dr. Benameur held various positions in Pharmaceutical Research and Development within Therapeutica, SMB Galephar, and Gattefossé. SUMMARY OF PRESENT WORK : Dr. Hassan Benameur is Senior Director Pharmaceutical Sciences at Capsugel with over 20 years of expertise in advanced drug delivery development. Dr. Benameur provides expertise in novel polymer applications for drug delivery and optimized lipid‐based formulations for enhanced absorption to maximize success and reduce attrition in pharmaceutical product development.

RECENT PUBLICATIONS : (1) “Lipophilic Salts – Opportunities and Applications in Oral Drug Delivery”. Hywel Williams, Annabel Igonin, David

Vodak and Hassan Benameur. Drug Development & Delivery, June 1st 2016, Online edition.

(2) “Enteric Capsule Drug Delivery Technology – Achieving protection without coating”. Hassan Benameur. Drug Development & Delivery, June 2015, Vol. 15, N°5.

(3) “Toward the establishment of standardized in vitro tests for lipid‐based formulations. Part 5. Lipolysis of

representative formulations by gastric lipase.” Bakala‐N'Goma JC, Williams HD, Sassene PJ, Kleberg K, Calderone M, Jannin V, Igonin A, Partheil A, Marchaud D, Jule E, Vertommen J, Maio M, Blundell R, Benameur H, Müllertz A, Pouton CW, Porter CJ, Carrière F. Pharm Res. 2015 Apr;32(4):1279‐87. doi: 10.1007/s11095‐014‐1532‐y. Epub 2014 Oct 7.

(4) “Toward the establishment of standardized in vitro tests for lipid‐based formulations. Part 6. Effects of varying

pancreatin and calcium levels.” Sassene P, Kleberg K, Williams HD, Bakala‐N'Goma JC, Carrière F, Calderone M, Jannin V, Igonin A, Partheil A, Marchaud D, Jule E, Vertommen J, Maio M, Blundell R, Benameur H, Porter CJ, Pouton CW, Müllertz A. AAPS J. 2014 Nov;16(6):1344‐57. doi: 10.1208/s12248‐014‐9672‐x. Epub 2014 Oct 2.

(5) “Toward the establishment of standardized in vitro tests for lipid‐based formulations, part 4: proposing a new

lipid formulation performance classification system”. Williams HD, Sassene P, Kleberg K, Calderone M, Igonin A, Jule E, Vertommen J, Blundell R, Benameur H, Müllertz A, Porter CJ, Pouton CW; Communicated on Behalf of the LFCS Consortium. J Pharm Sci. 2014 Aug;103(8):2441‐55. doi: 10.1002/jps.24067. Epub 2014 Jul 1.

(6) “Digestion of phospholipids after secretion of bile into the duodenum changes the phase behavior of bile

components”. Birru WA, Warren DB, Ibrahim A, Williams HD, Benameur H, Porter CJ, Chalmers DK, Pouton CW. Mol Pharm. 2014 Aug 4;11(8):2825‐34. doi: 10.1021/mp500193g. Epub 2014 Jul 16.


NAME : Cédric BOISSIERE AFFILIATION AND ADDRESS : Laboratoire Chimie de la Matière Condensée de Paris University Paris 6 UMR CNRS 7574 4 place Jussieu, Tour 44 E4 75005 PARIS France TELEPHONE N° : 33144276290

E‐MAIL : [email protected]

CAREER HISTORY :

Dr C. Boissiere was born in France 1974. He was appointed as Fellow Researcher CNRS in 2002 and is now Research Director, head of the processing and hybrid materials group. He works on the synthesis of functional hierarchical nano‐materials by coupling of evaporation processing and bottom‐up soft‐chemistry. Most of his achievements concern nanostructured thin films, nanoparticles and aerosol materials for optics, heterogeneous catalysis and nano‐medicine. He is co‐authors of around 160 articles and 28 patents. His work was awarded by the European Membrane Society (EMS) in 2006, the Jean RIST medal of the French Society of Materials and Metallurgy (SF2M)) in 2007, and French Chemical Society in Solid Chemistry (2014). MAJOR PUBLICATIONS :

Advanced drug delivery vectors with tailored surface properties made of mesoporous binary oxides

submicronic spheres. M. Colilla, M. Manzano, M. Vallet‐Regi, C. Boissière, C. Sanchez, Chemistry of

Materials 2010, 22(5), 1821‐1830

Chemical Modification As a Versatile Tool for Tuning Stability of Silica Based Mesoporous Carriers in

Biologically Relevant Conditions, T. Fontecave, C. Sanchez, T. Azaïs, and C. Boissiere Chemistry of Materials

(2012) 24 4226‐4236

Using EISA for the direct drug templating of therapeutic vectors with high loading fractions, tunable drug

release, and controlled degradation, T. Fontecave, C. Boissiere, N. Baccile, FJ Plou, C. Sanchez Chemistry of

Materials, (2013) 25 (23) 4671‐4678

Gold‐silica quantum rattles for multimodal imaging and therapy, M. Hembury, C. Chiappini, et al., PNAS

(2015) 112 (7) 1959‐1964


NAME: Patrick COUVREUR

AFFILIATION AND ADDRESS: University Paris‐Sud, Faculté de Pharmacie,

UMR CNRS 8612, Institut Galien, 5 rue JB Clément

F‐92296 Chatenay‐Malabry (France)

TELEPHONE No: + 33 1 46 83 53 96

FAX No: +33 1 46 83 59 46

E‐MAIL: patrick.couvreur@u‐psud.fr

CAREER HISTORY:

Pr Patrick COUVREUR is Full Professor of Pharmacy at the Paris‐Sud University and holder of the

chair of “Innovation Technologique” (2009‐2010) at the prestigious « Collège de France ». He is

appointed as a Senior Member of the “Institut Universitaire de France”. He is also the recipient of

an “ERC Advanced Grant” (2010‐2015). Patrick COUVREUR’s research has led to the funding of two

start‐up companies (Bioalliance and Medsqual). The major scientific contribution of Patrick

COUVREUR to the Pharmaceutical Sciences is also recognized by numerous international and

national awards. His appointment as a member of eight academies (Académie des Sciences,

Académie des Technologies, Académie de Médecine and Académie de Pharmacie in France, as

well as the Académie Royale de Médecine in Belgium, the Royal Academy of Pharmacy in Spain,

the US National Academy, Institute of Medicine and the US National Academy of Engineering in

USA) is another recognition of major scientific and scholarly contributions of Patrick COUVREUR.

SUMMARY OF PRESENT WORK:

The research performed aims at developing new nanomedicines for the treatment of severe

diseases in oncology, infectiology and neuroscience.

MAJOR PUBLICATIONS:

1) Gaudin et al., Nature Nanotechnology, 9, 1054‐1063; 2014 2) Maksimenko et al, Proceed. Natl. Acad. Sci. USA, doi/10.1073/pnas.1313459110; 2014 3) Maksimenko et al, ACS Nano, 8, 2018–2032; 2014 4) Mura et al, Nature Materials, 12, 991‐1003; 2013 5) Harrisson S et al, Angewandte Chemie Int. Edition, 52, 1678‐82 ; 2013 6) Hillaireau et al, Biomaterials, 34, 4831‐4838 ; 2013 7) Semiramoth N et al, ACS Nano, 6, 3820‐3831 ; 2012 8) Reddy LH et al, J Hepatol, 55, 1461‐1466 ; 2011 9) Arias et al, ACS Nano, 22, 1513‐1521 ; 2011 10) McKinlay et al, Angewandte Chemie Int. Edition, 23, 6260‐6266 ; 2010 11) Horcajada P et al, Nature Materials, 9, 172‐178 ; 2010.


NAME: Jennifer DOURLAT AFFILIATION AND ADDRESS : SANTEN SAS 1 rue Pierre Fontaine Bâtiment Genavenir IV F‐91058‐Evry cedex France

TELEPHONE N° : 01 69 87 40 20

FAX N° : 01 69 87 40 33


CAREER HISTORY: Jennifer is an Engineer in Chemistry by training, and holds a Doctorate in Pharmacochemistry from University Paris Descartes. She has been doing several years of Research in the field of chemistry and pharmacology, prior joining biotech and pharma companies, where she has gathered significant experience and expertise in chemistry, analytical characterization, formulation and drug delivery over the past 8 years. Jennifer has occupied different positions in Onxeo (former BioAlliance Pharma) as a Project Coordinator and an Analytical Laboratory Head, before moving to Ipsen Innovation where she has established an Early Formulation platform at the R&D Center of Les Ulis site. Jennifer has joint recently Santen SAS Group, as a Senior Manager of Advanced Technology, where she will manage the ocular drug delivery project portfolio of the group. Her presentation will focus on existing Santen Drug Delivery Platform and introduce innovative aspects in the field of ocular delivery.


NAME : Elias FATTAL AFFILIATION AND ADDRESS : Institut Galien Paris‐Sud University of Paris‐Sud UMR CNRS 8612 5 Rue Jean‐Baptiste Clément 92290 Châtenay‐Malabry France FAX N° : 33146835511

E‐MAIL : elias.fattal@u‐psud.fr

CAREER HISTORY : Elias Fattal is a full professor in Drug Delivery Science at the University of Paris‐Sud in Châtenay‐Malabry, France and has been President of APGI from 2003 to 2010. He received his Pharmacy Degree (1983) and Ph.D. (1990) from the University of Paris‐Sud and followed an internship in Pharmacy at the University of Lille (1984‐1986). After visiting the Department of Pharmaceutical Chemistry at the University of California, San Francisco for a post‐doctoral position (1990‐1991), he became associate Professor (1992) and full Professor at the University of Paris‐Sud (2000). He is heading the Institut Galien Paris‐Sud. He has received the PSWC Research Achievement Award and the CRS College of Fellows Award. SUMMARY OF PRESENT WORK : Elias Fattal is heading a multidisciplinary group working on the targeted delivery of nucleic acids as well as lung delivery of small molecules and biomacromolecules for the treatment of inflammatory diseases. His research deals also with nanotoxicology towards lungs MAJOR PUBLICATIONS :

1) Dosio F., Arpicco S., Stella B., Fattal E. Hyaluronic acid for anticancer drug and nucleic acid delivery Advanced Drug Delivery Reviews, 97,204–236, 2016

2) Santiago, L, Hillaireau, H., Grabowski, N., Mura, S., Nascimento, T., Dufort, S., Coll, J‐L, Tsapis, N., Fattal, E., Compared in vivo toxicity in mice of lung delivered biodegradable and non‐biodegradable nanoparticles Nanotoxicology, 17:1‐11, 2015.

3) Alshaer, W., Hillaireau, H., Vergnaud, J.,Ismail S., Fattal, E., Functionalizing liposomes with anti‐CD44 aptamer for selective targeting of cancer cells. Bioconjugate Chemistry, 26 (7), 1307–1313, 2015.

4) Giacalone, G., Hillaireau, H., Capiau, P., Chacun, H., Reynaud, F., Fattal, E., Stabilization and cellular delivery of chitosan‐polyphosphate nanoparticles by incorporation of iron. Journal of Controlled Release, 194, 211‐219, 2014.

5) Ababneh, N., Alshaer, W., Allozi, O., Mahafzah, A., El‐Khateeb, M., Hillaireau, H., Noiray, M., Fattal, E., Ismail, S., In vitro selection of modified RNA aptamers against CD44 cancer stem cell marker. Nucleic Acid Therapeutics, 23(6), 401‐407, 2013.

6) Diou, O., Tsapis, N., Giraudeau, C., Valette, J., Gueutin, C., Bourasset, F., Zanna, S., Vauthier, C., Fattal, E., Long‐circulating perfluorooctyl bromide nanocapsules for tumor imaging by 19FMRI. Biomaterials, 33(22), 5593‐5602, 2012.


NAME : Florence GAZEAU AFFILIATION AND ADDRESS : Université Paris‐Diderot Laboratoire Matière et systems Complexes (MSC) Bât. Condorcet 10, rue Alice Domon et Léonie Duquet 75205 PARIS CEDEX 13

CAREER HISTORY :

Florence Gazeau obtained her Ph.D. from the University Paris 7‐ Diderot in 1997, focusing on the magnetic and hydrodynamic properties of ferrofluids. She joined the National Research Center for Scientific Research (“CNRS”) as a staff scientist in 1998, when she broadened her research on biomedical applications of magnetic nanoparticles. Her current research interests focus on the physics of nanomagnetism applied to nanomedicine, cell‐nanoparticles interactions, cellular MRI, nanoparticles‐mediated hyperthermia, magnetic targeting, nanoparticles behavior, biodegradation and long term in vivo fate and nanotoxicology. She also developed alternatives to cell therapy by extracellular vesicles as a novel class of drug delivery system. She is a senior CNRS scientist since 2009, and she works in the laboratory Matière et Systèmes Complexes at the University Paris Diderot. She is animator of the group Nanomedicine at the French observatoire of micro and nanotechnology. She is authors of more than 115 publications.


NAME: Hideyoshi HARASHIMA AFFILIATION AND ADDRESS: Laboratory for Molecular Design of Pharmaceutics, Faculty of Pharmaceutical Sciences Hokkaido University Kita 12, Nishi 6, Sapporo City, Hokkaido 060‐0812, Japan

TELEPHONE No: +81‐11‐706‐3919

FAX No: +81‐11‐706‐4879

E‐MAIL: [email protected] CAREER HISTORY:

1985‐1987 Assistant Professor of The University of Tokyo. 1987‐1989 Postdoctoral fellowship at Department of Anesthesiology, Stanford University School of Medicine 1989‐1999 Associate Professor of The University of Tokushima 1999‐ Professor of Laboratory for Molecular Design of Pharmaceutics, Graduate School of Pharm. Sci, Hokkaido University 2007‐ Professor of Laboratory for Molecular Design of Pharmaceutics,

Faculty of Pharmaceutical Sciences, Hokkaido University. 2009‐ Professor of Laboratory of Innovative Nanomedicine, Faculty of Pharmaceutical

Sciences, Hokkaido University

SUMMARY OF PRESENT WORK: Development of Multifunctional Envelope‐type Nano Device (MEND) to control intracellular trafficking of nucleic acids such as siRNA/pDNA as well as to control biodistribution by passive/active targeting. Associate Editor of Journal of Controlled Release and Cancer Science and Executive Editor of Advanced Drug Delivery Reviews. The president of Academy of Pharmaceutical Science and Technology of Japan (APSTJ:2012~2014) and a director of Japanese Society of Drug Delivery System since 2006 and a director of Pharmaceutical Society of Japan (2014‐‐‐2016). The Nagai Award from Japanese Society of Drug Delivery System in 2007, Distinguished Science Award from FIP in 2010 and Fellow from Controlled Release Society in 2013. The Presidential award from Hokkaido University in 2015 and 2016. The APSTJ Award from APSTJ in 2016. MAJOR PUBLICATIONS: 1) Hossen MN, Kajimoto K, Akita H, Hyodo M, Ishitsuka T, Harashima H. Therapeutic Assessment of

Cytochrome C for the Prevention of Obesity Through Endothelial Cell‐targeted Nanoparticulate System. Molecular Therapy 21(3): 533‐41(2013).

2) Kusumoto K, Akita H, Ishitsuka T, Matsumoto Y, Nomoto T, Furukawa R, El‐Sayed A, Hatakeyama H, Kajimoto K, Yamada Y, Kataoka K, Harashima H. A Lipid Envelope‐Type Nano Particle Incorporating a Multifunctional Peptide for the Systemic siRNA Delivery to the Pulmonary Endothelium. ACS Nano. 7: 7534‐41 (2013).

3) Nakamura T, Fukiage M, Suzuki Y, Yano I, Miyazaki J, Nishiyama H, Akaza H, Harashima H. Mechanism responsible for the antitumor effect of BCG‐CWS using the LEEL method in a mouse bladder cancer model. J Control Release. 196:161‐7 (2014).

4) Furukawa R, Yamada Y, Kawamura E, Harashima H. Mitochondrial delivery of antisense RNA by MITO‐Porter results in mitochondrial RNA knockdown, and has a functional impact on mitochondria. Biomaterials. 57:107‐15 (2015)

5) Sato Y, Hatakeyama H, Hyodo M, Harashima H. Relationship between the Physicochemical Properties of Lipid Nanoparticles and the Quality of siRNA Delivery to Liver Cells. Mol Ther. 24(4): 788‐795 (2016).


NAME : Teruko IMAI AFFILIATION AND ADDRESS : Faculty of Pharmaceutical Sciences, Kumamoto University 5‐1 Oe‐honmachi, Chuo‐ku, Kumam oto 862‐0973, Japan TELEPHONE N° : +81‐96‐371‐4626

FAX N° : +81‐96‐371‐4626


CAREER HISTORY : 1999 Professor in Kumamoto University 1985 Assistant professor in Kumamoto University SUMMARY OF PRESENT WORK : Objective of my research is rational design of prodrugs based on the functional analysis of metabolizing enzyme and the physiological pharmacokinetic analysis. I focused on hydrolases, especially the following esterases as an enzyme for activation of prodrug in the body, carboxylesterase, carboxymethylenebutenolidase and esterase D as a tissue esterase, and butyrylcholinesterase and paraoxonase as plasma esterase. My research is performing as follows: 1. Cloning and characterization of above esterases ; 2. Structural analysis of the substrate binding site of hydrolases; 3. Regulation of tissue expression of hydrolase isozymes in human and animals; 4. Development of predicting method of in vivo disposition of prodrugs from the in vitro experimental data. MAJOR PUBLICATIONS :

1) Isolation and characterization of arylacetamide deacetylase in cynomolgus macaques. Yasuhiro Uno, Masakiyo Hosokawa, Teruko Imai., J Vet Med Sci 2015

2) 3D‐Fibroblast Tissues Constructed by a Cell‐Coat Technology Enhance Tight‐Junction Formation of Human Colon Epithelial Cells. Michiya Matsusaki, Daichi Hikimoto, Akihiro Nishiguchi, Koji Kadowaki, Kayoko Ohura, Teruko Imai, Mitsuru Akashi, Biochem Biophys Res Commun., 457(3): 363‐9, 2015

3) Distinct patterns of aging effects on the expression and activity of carboxylesterases in rat liver and

intestine, Ohura K., Tasaka K., Hashimoto M., Imai T., Drug Metab Dispos., 371‐384, 2014.


NAME : Tatsuhiro ISHIDA AFFILIATION AND ADDRESS :

Tokushima University, Institute of Biomedical Sciences, Department of Pharmacokinetics and Biopharmaceutics, 1‐78‐1 Sho‐machi, Tokushima, 770‐8505, Japan TELEPHONE N° : +81 88 633 7260 FAX N° : +81 88 633 7259 E‐MAIL : ishida@tokushima‐u.ac.jp CAREER HISTORY : 2014 Professor at Tokushima University 2003 Associate Professor at the University of Tokushima 2000 Lecturer at the University of Tokushima 1998 Postdoctoral Fellow at the University of Alberta, Canada (group T.M. Allen) 1998 Ph.D. at the University of Tokushima (Prof. H. Kiwada) SUMMARY OF PRESENT WORK : TI has paid an attention to development of safe and efficient drug delivery system with liposome from beginning of his research carrier. He tries to deliver anticancer drugs, proteins and nucleic acids to diseased tissue in order to give patients cures. In addition, he tries to elucidate what happens in the body following systemic treatment with liposomal formulation or combined treatment with free drugs and liposomal formulation. He always pays an attention to not only efficacy but also side effect upon such treatment. Most of his projects are currently oriented on cancer chemotherapy and cancer gene therapy. MAJOR PUBLICATIONS :

1) Abu Lila, A.S., Kato, C., Fukushima, M., Huang, C., Wada, H., Ishida, T., Downregulation of thymidylate synthase by RNAi molecules enhances the antitumor effect of pemetrexed in an orthotopic malignant mesothelioma xenograft mouse model. Int. J. Oncol., 48, 1399‐1407 (2016)

2) Essam Eldin, N., Abu Lila, A.S., Kawazoe, K., Elnahas, H.M., Mahdy, M.A., Ishida, T., Encapsulation in

a rapid‐release liposomal formulation enhances the anti‐tumor efficacy of pemetrexed in a murine solid mesothelioma‐xenograft model. Eur. J. Pharm. Sci., 81, 60‐66 (2016)

3) Nakamura, H., Abu Lila A.S., Nishio, M., Tanaka, M., Ando, H., Kiwada, H., Ishida, T., Intra‐tumor

distribution of PEGylated liposome upon repeated injection: No possession by prior dose. J. Control. Release, 220, 406‐413 (2015)

4) Ando, H., Kobayashi, S., Abu Lila A.S., Essam Eldin N., Kato, C., Shimizu, T., Ukawa, M., Kawazoe, K.,

Ishida, T., Advanced therapeutic approach for the treatment of malignant pleural mesothelioma via the intrapleural administration of liposomal pemetrexed. J. Control. Release, 220, 29‐36 (2015)

5) Mima, Y., Hashimoto, Y., Shimizu, T., Kiwada, H., Ishida, T., Anti‐PEG IgM is a major contributor to

the accelerated blood clearance of polyethylene glycol‐conjugated protein. Mol. Pharmaceutics, 12, 2429–2435 (2015)


6) Hashimoto, Y., Abu Lila A., Shimizu,T., Ishida, T., Kiwada, H., B cell‐intrinsic toll‐like receptor 7 is

responsible for the enhanced anti‐PEG IgM production following injection of siRNA‐containing PEGylated lipoplex in mice. J. Control. Release, 184, 1‐8 (2014)

7) Abu Lila, A.S., Uehara, Y., Ishida, T., Kiwada, H., Application of polyglycerol‐coating to pDNA lipoplex

for the evasion of the accelerated blood clearance (ABC) phenomenon in nucleic acid delivery. J. Pharm. Sci., 103, 557‐566 (2014)

8) Abu Lila, A.S., Nawata, K., Shimizu, T., Ishida, T., Kiwada, H., Use of polyglycerol (PG), instead of

polyethylene glycol (PEG), prevents induction of the accelerated blood clearance phenomenon against long‐circulating liposomes upon repeated administration. Int. J. Pharm., 456, 235‐242 (2013)


NAME: Vincent JANNIN AFFILIATION AND ADDRESS: GATTEFOSSE S.A.S. 36 chemin de Genas 69804 Saint‐Priest cedex TELEPHONE N°: +33 4 72 22 98 38

FAX N°: +33 4 78 90 45 67


CAREER HISTORY: Jan. 2015 – present Research Director – Pharmaceuticals, Gattefossé S.A.S. Sept. 2001 – Dec. 2014 Pharm. R&D Project Director, Gattefossé S.A.S. Sept. 1999 – Aug. 2001 Pharmaceutical Laboratory Manager, Gattefossé S.A.S. SUMMARY OF PRESENT WORK: Dr Jannin’s current work is focused on the development and characterization of new lipid‐based excipients, particularly in the field of solubility and oral bioavailability enhancement for poorly water‐soluble drugs and peptides. His work comprises physical (polymorphism), physical‐chemical (colloidal sciences), biochemical (lipolysis) and biological (cell coculture) characterization of lipid‐based systems, notably of acylglycerols and polyoxylglycerides. MAJOR PUBLICATIONS: 1. Fernandez, S., Jannin, V., Rodier, J.D., Ritter, N., Mahler, B., Carrière, F. 2007. Comparative study of digestive

lipases performance on the self emulsifying excipient Labrasol®, medium chain glycerides and PEG esters. Biochimica et Biophysica Acta ‐ Molecular and Cell Biology of Lipids, 1771 (5), 633‐640.

2. Jannin, V., Musakhanian, J., Marchaud, D. 2008. Approaches for the development of solid and semi solid lipid‐based formulations. Advanced Drug Delivery Reviews, 60 (6) 734‐746.

3. Fernandez, S., Chevrier, S., Ritter, N., Mahler, B., Demarne, F., Carrière, F., Jannin, V. 2009. In vitro gastrointestinal lipolysis of four formulations of piroxicam and cinnarizine with the self emulsifying excipients Labrasol® and Gelucire® 44/14. Pharmaceutical Research, 26 (8) 1901‐1910.

4. Williams, H., Sassene, P., Kleberg, K., Bakala N’Goma, J.C., Calderone, M., Jannin, V., Igonin, A., et al., C.W. 2012. Towards the establishment of standardized in vitro tests for lipid‐based formulations: 1) Method parameterisation and comparison of in vitro digestion profiles across a range of representative formulations. Journal of Pharmaceutical Sciences, 101(9) 3360‐3380.

5. Béduneau, A., Tempesta, C., Fimbel, S., Pellequer, Y., Jannin, V., Demarne, F., Lamprecht, A. 2014. A tunable Caco‐2/HT29‐MTX co‐culture model mimicking variable permeabilities of the human intestine obtained by a new seeding procedure. European Journal of Pharmaceutics and Biopharmaceutics, 87 (2) 290‐298.

6. Jannin, V., Rosiaux, Y., Doucet, J. 2015. Exploring the possible relationship between the drug release of Compritol®‐containing tablets and its polymorph forms using micro X‐ray diffraction. Journal of Controlled Release. 197 158‐164.


NAME: Kazunori KATAOKA

AFFILIATION AND ADDRESS: Innovation Center of NanoMedicine (iCONM), Institute of Industrial Promotion – KAWASAKI, 3‐25‐14, Tonomachi, Kawasaki‐ku, Kawasaki 210‐0821, Japan. Policy Alternatives Research Institute, The University of Tokyo, 7‐3‐1, Hongo, Bunkyo‐ku, Tokyo 113‐1709, Japan

TELEPHONE No: +81‐44‐589‐5700

FAX No: +81‐44‐589‐5706

E‐MAIL: [email protected]‐tokyo.ac.jp, k‐kataoka@kawasaki‐net.ne.jp

CAREER HISTORY: 2016: Director General, Innovation Center of NanoMedicine, Institute of Industrial Promotion‐

KAWASAKI

2016: Professor, Policy Alternatives Research Institute, The University of Tokyo

2016: Adjunct Professor, Eshelman School of Pharmacy, The University of North Carolina at Chapel

Hill

2004: Professor (Joint Position), Graduate School of Medicine, The University of Tokyo

1998: Professor, Graduate School of Engineering, The University of Tokyo

1994: Professor, Faculty of Engineering Science, Tokyo University of Science

1989: Associate Professor, Faculty of Engineering Science, Tokyo University of Science

1979: Assistant Professor, Institute of Biomedical Engineering, Tokyo Women’s Medical College

1979: PhD at the University of Tokyo

SUMMARY OF PRESENT WORK: Polymeric Biomaterials for Drug Delivery, Block Copolymer Self‐Assembly, Stimuli‐Sensitive Polymers MAJOR PUBLICATIONS:

1) Harada, K. Kataoka, Chain length recognition: core‐shell supramolecular assembly from oppositely

charged block copolymers. Science 283(5398) 65‐67 (1999)

2) K. Kataoka, A. Harada, Y. Nagasaki, Block copolymer micelles for drug delivery: design,

characterization, and biological significance. Adv. Drug Deliv. Rev. 47(1) 113‐131 (2001)


NAME : Hiroshi KIKUCHI

AFFILIATION AND ADDRESS : Executive Director, Tsu kuba Research Laboratories, Eisai Co., Ltd. Tokodai 5‐1‐3, Tsukuba‐shi, Ibaraki 300‐2635, Japan

TELEPHONE No : +81‐29‐847‐5469

FAX No : +81‐29‐847‐2037

E‐MAIL : h4‐[email protected]

CAREER HISTORY : ‐ B.S. (1977) & Ph.D. (1991) degrees in Pharm. Sciences, The University of Tokyo ‐ Visiting Research Fellow (1993‐1994), School of Pharmacy, The Univ. of Michigan ‐ Adjunct Professor (1992‐present), Faculty of Pharm. Sci., The University of Tokyo ‐ Affiliate Professor (2007‐present), Graduate School of Pharm. Sci., Kyushu Univ. ‐ Adjunct Professor (2014‐present), Graduate School of Engineering, Kyoto Univ. ‐ Scientist, Principal Investigator, Daiichi Pharmaceutical Co., Ltd. (1977‐2007) ‐ Director (2007‐), Officer (2010‐) & Executive Director (2015‐present), Eisai Co., Ltd.

SUMMARY OF PRESENT WORK : Main research subject is drug delivery systems using particulate carriers such as liposomes, nanoparticles and emulsions. More than 410 presentations (95 international conferences) have been performed including 102 invited presentations (30 international conferences) and 91 invited lectures as of today. In addition, 116 publication papers (90 original scientific papers) have been published and more than 70 basic patents have been applied.

MAJOR PUBLICATIONS : 1) T. Suzuki, M. Ichihara, K. Hyodo, E. Yamamoto, T. Ishida, H. Kiwada, H. Kikuchi and H. Ishihara, Influence

of dose and animal species on accelerated blood clearance of PEGylated liposomal doxorubicin, Int. J.

Pharm., 476, 205‐212 (2014).

2) F. Okazaki, N. Matsunaga, H. Okazaki, H. Azuma, K. Hamamura, Y. Hara, A. Tsuruta, Y. Tsurudome, T.

Ogino, T. Suzuki, K. Hyodo, H. Ishihara, H. Kikuchi, H. To, H. Aramaki, S. Koyanagi and S. Ohdo, Circadian

clock in a mouse colon tumor regulates intracellular iron levels to promote tumor progression, J. Biol.

Chem., 291, 7017‐7028 (2016).

HONORS AND AWARDS:

‐ The 1st Nagai Award, The Japan Society of Drug Delivery System (2001)

‐ Takeru Aya Higuchi Memorial Prize, The Academy of Pharmaceutical Science & Technology, Japan (2007)

‐ The Award, The Academy of Pharmaceutical Science and Technology, Japan (2010)

‐ Profiled in Marquis Who’s Who in the World (2012, 2013, 2014, 2015 and 2016)

PROFESSIONAL SOCIETIES:

‐ Councilor (2001‐present), Trustee (2008‐2012), Auditor (2014‐2016) & Trustee (2016‐present), The Academy

of Pharmaceutical Science and Technology, Japan

‐ Councilor (2001‐present) & Auditor (2009‐present), The Japan Society of Drug Delivery System; Member of

Editorial Board (2010‐present) for “Drug Delivery System”

‐ Councilor (2002‐present), The Society of Cyclodextrins, Japan

‐ Trustee (2014‐present), The Pharmaceutical Society of Japan

‐ Auditor (2014‐present), The Pharmaceutical Society of Japan Kanto Branch


NAME : Kentaro KOGURE AFFILIATION AND ADDRESS : UNIVERSITY OF TOKUSHIMA, INSTITUTE OF HEALTH BIOSCIENCES Shomachi 1, Tokushima, 770‐8505, Japan TELEPHONE N° : +81‐88‐633‐7248

FAX N° : +81‐88‐633‐9572

E‐MAIL : kogure@tokushima‐u.ac.jp

CAREER HISTORY : 1994 Ph.D. at Tokushima University 1994 Assistant Professor, Toyama Medical and Pharmaceutical University 1998 Assistant Professor, Tokushima University 2003 Postdoctoral fellow, Japan Science and Technology Agency, CREST, Hokkaido University 2005 Lecturer, Hokkaido University 2007 Professor, Kyoto Pharmaceutical University 2016 Professor, Tokushima University SUMMARY OF PRESENT WORK : Transdermal delivery of macromolecules, such as functional nucleic acids and antigen peptides, by faint electricity Development of antioxidative nanoparticles encapsulating antioxidants, such as astaxantin, tocotrienol, for prevention of various oxidative stress MAJOR PUBLICATIONS : 1) Hasan M, Nishimoto A, Ohgita T, Hama S, Kashida H, Asanuma H, Kogure K. Faint electric treatment‐

induced rapid and efficient delivery of extraneous hydrophilic molecules into the cytoplasm. J. Control. Release 228, 20‐25 (2016).

2) Hama S, Kimura Y, Mikami A, Shiota K, Toyoda M, Tamura A, Nagasaki Y, Kanamura K, Kajimoto K, Kogure K. Electric stimulus opens intercellular spaces in skin. J. Biol. Chem. 289, 2450‐2456 (2014).


NAME: Hiromuo KONDO

AFFILIATION AND ADDRESS:

Drug Delivery, Pharmaceutical Research and Technology Labs,

Astellas Pharma Inc.

180, Ozumi, Yaizu‐shi, Shizuoka, 425‐0072, Japan

TELEPHONE No: +81‐54‐627‐9249

FAX No: +81‐54‐627‐9918


CAREER HISTORY:

Nagoya City University, Nagoya, Japan, M.S., March 1993

Astellas Pharma Inc. (former Yamanouch Pharmaceutical Co., Ltd. before April 2005) since April

1993

Ph.D. from Nihon University, Japan, March 2003

SUMMARY OF PRESENT WORK:

Senior Director of Drug Delivery

Research and development on new pharmaceutical technology

Proposal and development of new formulation for product life cycle extension MAJOR PUBLICATIONS:

1) Yoshida T., Nakanishi K., Yoshioka T., Tsutsui Y., Maeda A., Kondo H., Sako K., Euro J Pharm Biopharm.,

100, 58‐65 (2016).

2) Ishii T., Kobayashi N., Maeda A., Kondo H., Sako K., Yamada S., Kagawa Y., J Drug Del Sci Tech., 27, 1‐8

(2015).

3) Takemura S., Kondo H., Watanabe S., Sako K., Ogawara K., Higaki K., J. Pharm. Sci., 102(9), 3128‐35

(2013)

4) Takemura S., Kondo H., Suzumura K., Ogawara K., Watanabe S., Sako K., Higaki K., J. Pharm. Sci.,

101(6), 2134‐42 (2012)

5) Yasuji T., Kondo H., Sako K., Ther. Deliv., 3(1), 81‐90 (2012)

6) Kondo H., Shinoda T., Nakashima H., Watanabe T., Yokohama S., Biopharm. Drug Dispos., 24, 45‐51

(2003).

7) Kondo H., Takahashi Y., Watanabe T., Yokohama S., Watanabe J., Biopharm. Drug Dispos., 24, 131‐

140 (2003).


Name: Ryoji KONISHI

AFFILIATION AND ADDRESS: Dept. of Pharmaco‐Bio‐Informatics, Fac. of Med., Kagawa University Ikenobe, Miki, Kagawa, 761‐0793, Japan TEL +81‐(0)87‐ 898‐5111(Ext.2424) FAX +81‐(0)87‐ 891‐2383 E‐MAIL [email protected]‐u.ac.jp

CAREER HISTORY:

1991 Professor (visiting) Kagawa Medical University (Since 2003, Fac. of Med., Kagawa Univ.) 1983 Teikoku Seiyaku Co., Ltd., R&D 1969 Associate Professor, Nagasaki University, Faculty of Pharmacy 1967 PhD in Pharmaceutical Science, Kyoto University 1965 Assistant Professor, Kyoto University SUMMARY OF PRESENT WORK : Transdermal Delivery System MAJOR PUBLICATIONS: 1. Buccal/gingival drug delivery systems, T. Nagai & R. Konishi, J. Controlled Release 8, 353 (1987) 2. In vitro testing and transdermal delivery, T. Higuchi & R. Konishi, Therapeutic Research 6, 280 (1987) 3. Stage and region dependent expression of a radial glial marker in commissural fibers in kindled mice, S.

Tanaka et al. Epilepsy Research 67, 61 (2005) 4. A novel nucleic acid analogue shows strong angiogenic activity in vitro, I. Tsukamoto et al. Biochem.

Biophys. Res. Com. 399, 699 (2010) 5. Delayed administration of the nucleic acid analog 2Cl‐C.OXT‐A attenuates brain damage and enhances

functional recovery after ischemic stroke, N. Okabe et al. Brain Res. 1506, 115 (2013) 6. Involvement of S1P1 receptor pathway in angiogenic effects of a novel adenosine‐like nucleic acid

analog COA‐Cl in cultured human vascular endothelial cells, J. Igarashi et al. Pharma. Res. Per. 2, e00068 (2014)

7. Intestinal absorption, organ distribution, and urinary excretion of the rare sugar D‐psicose, I. Tsukamoto et al. Drug Design, Development and Therapy 8, 1955 (2014)

8. A Key Role of PGC‐1a Transcriptional Coactivator in Production of VEGF by a Novel Angiogenic Agent COA‐Cl in Cultured Human Fibroblasts, J. Igarashi et al. Physiol. Rep. 4, e12742 (2016)


NAME : Sebastien LECOMMANDOUX


Professor IPB‐ENSCBP, Université de Bordeaux

Laboratoire de Chimie des Polymères Organiques

UMR CNRS 5629, FRANCE

TELEPHONE No: 33 (0)5 40 00 22 41

FAX No: 33 (0)5 40 00 84 86


CAREER HISTORY : Sébastien Lecommandoux is Full Professor at the University of Bordeaux (Bordeaux‐INP). He is Director of the Laboratoire de Chimie des Polymères Organiques (LCPO‐CNRS) and is leading the group “Polymers Self‐Assembly and Life Sciences”. Sébastien Lecommandoux is recipient of the CNRS bronze medal (2004) and Institut Universitaire de France Junior Chair (IUF 2007). He is Associate Editor of Biomacromolecules (ACS) and in the Editorial Advisory Board of several international journals, including Bioconjugate Chemistry (ACS), Polymer Chemistry and Biomaterials Science (RSC).

SUMMARY OF PRESENT WORK : The aim of our current research is to develop block copolymers able to self‐assemble into nanostructured drug delivery systems, including spherical micelles or polymersomes. Specific attention is paid on these last ones, for which we try to make them biologically smart or responsive to specific triggers. We are developing a bio‐inspired approach by using polypeptide and/or polysaccharide building blocks. Most of our projects are currently oriented on cancer therapy and diagnosis.

MAJOR PUBLICATIONS : (1) Targeting CD44 receptor‐positive lung tumors using polysaccharide‐based nanocarriers: influence of nanoparticle size and administration route. V. Jeannot, S. Mazzaferro, J. Lavaud, M. Arboléas, V. Josserand, J.‐L. Coll, S. Lecommandoux, C. Schatz, A. Hurbin. Nanomedicine: Nanotechnology, Biology, and Medicine 4, 921‐932 (2016). (2) Quantitative side‐chain modifications of methionine‐containing elastin‐like polypeptides as a versatile tool to tune their properties. J.R. Kramer, R. Petitdemange, L. Bataille, A.‐L. Wirotius, K. Bathany, B. Garbay, T.J. Deming, E. Garanger, S. Lecommandoux. ACS Macro Letters 4, 1283‐1286 (2015). (3) Cascade Reactions in Multicompartmentalized Polymersomes. R. J. R. W. Peters, M. Marguet, S. Marais, M. W. Fraaije, J. C. M. Van Hest, S. Lecommandoux. Angew. Chem. Int. Ed. 53, 146‐150 (2014). (4) Polymersome Shape Transformation at the Nanoscale. R. Salva, J.‐F. Le Meins, O. Sandre, A. Brulet, M. Schmutz, P. Guenoun, S. Lecommandoux. ACS Nano 7, 9298‐9311 (2013). (5) Biologically Active Polymersomes from Amphiphilic Glycopeptides. J. Huang, C. Bonduelle, J. Thevenot, S. Lecommandoux, A. Heise. J. Am. Chem. Soc. 134 (1), 119‐122 (2012). (6) Polymersomes in polymersomes : multiple loading and permeability tuning. M. Marguet, L. Edembe, S. Lecommandoux. Angew. Chem. Int. Ed., 51, 1173‐1176 (2012).


NAME: Philippe LEFEVRE AFFILIATION AND ADDRESS: ROQUETTE, Customer Technical Service, Pharma Global Business Unit, 1 rue de la Haute Loge, 62136 Lestrem, France TELEPHONE N°: +33 321 633 670, MOBILE: +33 760 169 328

E‐MAIL: [email protected] CAREER HISTORY : After a chemical engineer degree (ENSCL, Lille, France, 1983), I was lucky enough to do my compulsory military service in an army scientific laboratory watching over nuclear contamination around civil or military nuclear sites… In 1985 I joined Roquette as manager of a quality control laboratory. In 1989 I moved to a Food Application Laboratory which included a tableting lab. I discovered a passion for the few questions arising from the pharma field and I was convinced that this field had to be developed. In 1991 I created the first Pharma Application Laboratory in Roquette, focusing on Pharmaceutical Technology, mainly granulation and tableting. I was highly active in the development of new excipients from the starch industry. In 2012, I was promoted Scientific Coordinator for Pharmaceutical Applications, coordinating the development of new excipients and coordinating the three regional Pharmaceutical Applications Laboratories (Asia, Europe, and Americas). In 2015, I was appointed Pharmaceutical Application Fellow, Senior‐Expert for the applications of ROQUETTE excipients and raw materials in Pharmaceutics & in Powder Technology. SUMMARY OF PRESENT WORK : Comprehension of mannitol behavior in high speed rotary press tableting using simulator The behavior of mannitol in tableting has always been difficult to apprehend and literature is conflicting on the nature of mannitol deformation mechanisms. We anticipated that compression kinetic was the key point when tableting mannitol. A Stylcam compression simulator was used to characterize the deformation behavior of textured mannitol in conditions of industrial tableting. The studied textured mannitol behaves more like a fragmentary material contrary to the plastic deformation mainly described in the literature. Comparison with a single punch press demonstrates the necessity to move to a high speed compression simulator when working on mannitol. Additionally it was used to determine what tableting parameters and powder characteristics are detrimental to the performance of mannitol in direct compression and to solve issues encountered in real pharmaceutical tablet production. MAJOR PUBLICATIONS: 33 articles and posters, focusing on direct compression excipients, orodispersible tablets, film‐coating and molecular inclusion with cyclodextrines Last article: TARLIER N., SOULAIROL I., BATAILLE B., RAVEL P., NOFRERIAS I., LEFEVRE P., SHARKAWI T., Compaction behavior and deformation mechanism of directly compressible textured mannitol in a rotary tablet press simulator, INTERNATIONAL JOURNAL OF PHARMACEUTICS 495.1 (Nov 10, 2015): 410‐419 29 patents, about half leading to commercial outlet


NAME : Yuji MAKINO

AFFILIATION AND ADDRESS : Facility for Industry‐Government‐Academia Collaboration, Musashino University, 1‐1‐20, Shinmachi, Nishi‐Tokyo, Tokyo 202‐8585, Japan

TELEPHONE N° :+81 3 3394 1374


CAREER HISTORY : 2013 Visiting Professor, Musashino University 2006 Professor, Tokushima Bunri University 2006 Ph.D. from University of Tokyo 1999 Director, Pharm. Product Res. Lab. Teijin Limited 1975 Research Chemist, Bio‐medical Res. Lab., Teijin Limited 1975 M.Sc. in Pharmaceutical Sciences from University of Tokyo 1971 Pharmacist (No.128333) 1970 Bachelor in Pharmaceutical Sciences from University of Tokyo SUMMARY OF PRESENT WORK : In 2013, after retirement from his life of meetings and lectures in the countryside, he soon returned to his home town Tokyo for laboratory works. His current interests include microneedle (presented here) and oral absorption enhancements (2nd generation product of highly absorbable oral curcumin is expected to be on European supplement market in the next year). He studies these projects with venture companies and focuses on overcoming barriers that block their way to commercializations. MAJOR PUBLICATIONS : (1) Kurita T, Makino Y: Novel Curcumin Oral Delivery Systems. Anticancer Research 33, 2807‐22, 2013 (2) Oshima H, Miyagishima A, Kurita T, Makino Y, Iwao Y, Sonobe T and Itai S: Preparation and stabilization of nifedipine

lipid nanoparticles. Int. J. Pharm., 377, 180‐184, 2009 (3) Makino Y, Nishibe Y, Matsugi H, Nishimura Y and Katsuyama I: A new in vivo animal model for nasal pharmacokinetic

studies. Drug Delivery System 20, 543‐550, 2005 (4) Makino Y, Nishibe Y, Matsugi H, Nishimura Y and Katsuyama I: Pharmacokinetics of nasal powder formulations. Drug

Delivery System 20, 656‐665, 2005 (5) Sakagami M, Sakon K, Kinoshita W, Makino Y: Enhanced pulmonary absorption following aerosol administration of

mucoadhesive powder microspheres. J Controlled Release, 77, 117‐129, 2001 (6) Sakagami M, Kinoshita W, Sakon K, Sato J and Makino Y: Mucoadhesive beclomethasone microspheres for powder

inhalation: their pharmacokinetics and pharmacodynamics evaluation. J Controlled Release, 80, 207‐218, 2002 (7) Sakagami M, Kinoshita W, Sakon K and Makino Y: Fractional contribution of lung, nasal and gastrointestinal absorption

to the systemic level following nose‐only aereosol exposure in rats: a case study of 3‐7 μm fluorescein aerosols. Arch Toxicol,, 77, 321‐329, 2003

(8) Suzuki Y and Makino Y: Mucosal drug delivery using cellulose derivatives as a functional polymer. J Controlled Release, 62, 101‐107, 1999

(9) Nagai T, Nishibe Y, Makino Y, Tomimori T and Yamada H: Enhancement of in vivo anti‐influenza virus activity of 5,7,4´‐trihydroxy‐8‐methoxyflavone by drug delivery system using hydroxypropyl cellulose. Biol Pharm Bull. 20

(10) 1082‐5, 1997 (10) Nishimura M, Makino Y and Matugi H: Tacalcitol Ointment for Psoriasis. Acta Derm Venereol (Stockh), Suppl. 186, 166‐168, 1994


NAME : Kazuo MARUYAMA AFFILIATION AND ADDRESS : Teikyo University ‐ Faculty of Pharma‐Sciences Laboratory of Drug Delivery System Research 2‐11‐1 Kaga, Itabashi‐ku, Tokyo JAPAN, 173‐8605 TELEPHONE N° : +81‐3‐3964‐8239

FAX N° : +81‐3‐3964‐8243


CAREER HISTORY : He has graduated Graduate School of Pharmaceutical Sciences, Kumamoto University in 1979. He has been full professor of Teikyo University since 2002. He received the 1st MIZUSHIMA AWARD in The Japan Society of Drug Delivery System on Jun 28, 2009. He is the Associate Editor of Journal of Liposome Research since 2002. He had organized the 11th Liposome Research Days International Conference in Yokohama on July 19th, 2008. SUMMARY OF PRESENT WORK : Recently, he has succeeded in developing a lipid bubble, which is a lipid micelle entrapping perfluorocarbon gas. Lipid bubble can use for both as a contrast agent in ultrasonography and a non‐invasive carrier for drug and gene delivery. The combination of lipid bubble and ultrasound is a good tool for “Theranostics” in cancer therapy. MAJOR PUBLICATIONS :

1) Tumor growth suppression by the combination of nanobubbles and ultrasound. Suzuki Ryo, Oda Yusuke, Omata Daiki, Nishiie Norihito, Koshima Risa, Shiono Yasuyuki, Sawaguchi Yoshikazu, Unga Johan, Naoi, Tomoyuki, Negishi Yoichi, Kawakami Shigeru, Hashida Mitsuru, Maruyama Kazuo. Cancer Science. 107, 217‐223, 2016

2) Enhancement of Blood‐Brain Barrier Permeability and Delivery of Antisense Oligonucleotides or plasmid DNA to Brain by the Combination of Bubble Liposomes and High‐intensity Focused Ultrasound. Yoichi Negishi, Masaya Yamane, Naho Kurihara, Yoko Endo‐Takahashi, Sanae Sashida, Norio Takagi, Ryo Suzuki, Kazuo Maruyama. Pharmaceutics 7,344‐362, 2015

3) Evaluation of the potential of doxorubicin loaded microbubbles as a theranostic modality using a murine tumor model. Shigeru Kawakami, Rodi Abdalkader, Johan Unga, Ryo Suzuki, Kazuo Maruyama, Fumiyoshi Yamashita, Mitsuru Hashida. Acta Biomater. 19, 112‐118. 2015

4) Development of fluorous lipid‐based nanobubbles for efficiently containing perfluoropropane, Yusuke Oda, Ryo Suzuki, Tatsuya Mori, Hideyo Takahashi, Hideaki Natsugari, Daiki Omata, Johan Unga, Hitoshi Uruga, Mutsumi Sugii, Shigeru Kawakami, Yuriko Higuchi, Fumiyoshi Yamashita, Mitsuru Hashida, Kazuo Maruyama, Int. J. Pharm. 487, 64‐71, 2015


NAME : Nathalie MIGNET AFFILIATION AND ADDRESS : CNRS UMR8258, INSERM U1022, Paris Descartes university, Chimie ParisTech Center of Pharmaceutical Research 4 avenue de l’observatoire 75006 PARIS

TELEPHONE N° :0153739581


CAREER HISTORY : Her scientific bases are issued from studies in physic sciences in Toulouse, France. After a PhD in Montpellier on oligonucleotide chemical synthesis in 1996, she was hired by the company Lynx Therapeutics in San Francisco, USA, working with Dr S. Gryaznov. She then joined the University of Sheffield, UK, to work on the method SELEX with Dr D. Williams. In 1999 she was hired by the French biotech company Capsulis (D. Roux, Bordeaux, France). She joined the CNRS as a research Scientist in 2000 to develop formulations for gene delivery in the mixed unit group CNRS/Aventis directed by Dr. D. Scherman. SUMMARY OF PRESENT WORK : Dr Nathalie MIGNET is performing her research at the Faculty of Pharmacy of the University Paris Descartes as CNRS research director. She in charge of the team “Nanovectors for targeted therapy and molecular imaging”. Her research dedicated to nanoparticles is interdisciplinary, at the interface between chemistry, physico‐chemistry, cell biology and in vivo studies. She is mostly interested in the development of formulations dedicated to therapy and targeted nanoparticles for imaging, development of vectors using imaging as a gain into in vivo understanding of nanovectors. The amin applications of her research are found in colon cancer and hepatic metastases, where her team developed very well described models. From 2012 to 2013, she was president of the GTRV (Vectorisation group). In 2014, she founded the French Society for Nanomedecine, SFNano. She is also actively involved in the working groups imaging and therapeutic of the European Technological Platform for nanomedicine.


NAME : Tsuneji NAGAI AFFILIATION AND ADDRESS :

The Nagai Foundation Tokyo Yayoi 2‐4‐16, Bunkyo‐ku, Tokyo 113‐0021, Japan

TELEPHONE N°: +81‐3‐3946‐8260

FAX N : +81‐3‐3946‐8260

E‐MAIL : nagai‐[email protected]

CAREER HISTORY: Ph.D., University of Tokyo, 1961. Research Associate, University of Toyo, 1961‐1971. Professor, Hoshi University, Tokyo, 1971‐1999. President, Hoshi University, Tokyo, 2001‐2014 SUMMARY OF PRESENT WORK: President, The Nagai Foundation Tokyo, 1986‐present. The Nagai Foundation Tokyo was

founded in 1986 in commemoration of Prof. Tsuneji Nagai receiving the 1986 Hoest‐Madsen Medal from FIP as the first for Japanese. Then the Foundation was officially approved as government‐licensed in 1994 for the purpose of promoting international exchange in pharmacy and pharmaceutical sciences. MAJOR PUBLICATIONS: Prof. Nagai’s research activity has been mainly on bioavailability studies and drug delivery

system (DDS) with 576 original refereed research papers, over 550 review and other scientific articles, 64 patents expired already (56 Japanese and 8 international), and 3 marketed ethical drug products ("Aftach®"; "Rhinocort®"; "Salcort®"). Around the time of retirement from his research position (1999), the citation of papers by Nagai and his group in the field of drug delivery in the world was No. 4 according to the data for the years of 1975‐1997 published by ISI: with 1636 citations for 287 papers of Nagai and his group; among about 6,000,000 chemists, among whom the first 10,800 researchers have more than 500 citations of all their papers. Two volumes of The Special Issue in Horner of Prof. Tsuneji Nagai were published in Journal of

Controlled Release (edited by Prof. Yuichi Sugiyama, Vol. 62, 1999) and in International Journal of Pharmaceutics (edited by Prof. Takashi Sonobe, Vol. 354, 2008), respectively.


NAME : Hiroaki OKADA AFFILIATION AND ADDRESS : Okada DDS Research Institute, Inc. ‐ 2‐83‐2‐1108 Yarimizu, Hachioji, Tokyo 192‐0375 Japan TELEPHONE & FAX N° : +81‐42‐670‐6489 E‐MAIL : okada@okada‐dds.com, [email protected]

CAREER HISTORY : BS, 1968, MS, 1970, PhD, 1982 School of Pharmacy, Kyushu University (Fukuoka, Japan) 1973‐1974 Graduate School, University of Kansas (USA, under Prof. Takeru Higuchi) 1970‐1993 Research Scientist, Pharmaceutical Research Labs., Takeda Pharm. Co. Ltd. 1993‐1997 Senior Research Head, DDS Research Labs., Takeda Pharm. Co. Ltd. 1997‐2001 Director, Pharmaceutical Business Development, Takeda Pharm. Co. Ltd. 2001‐2011 Professor, School of Pharmacy, Tokyo University of Pharmacy & Life Sciences 2011‐ Professor Emeritus, Tokyo University of Pharmacy & Life Sciences 2011‐ Director (CEO), Okada DDS Research Institute, Inc. SUMMARY OF PRESENT WORK : Consulting on the new drug discovery, formulation, production and reformulation of small molecular drugs and biologics such as peptides and oligonucleotides. MAJOR PUBLICATIONS : 1) H. Okada, Y. Sakura, H. Kawaji, T. Yashiki, and H. Mima, Regression of rat mammary tumors by a potent

luteinizing hormone‐regression of rat mammary tumors by a potent luteinizing hormone‐releasing hormone analogue (leuprolide) administered vaginally. Cancer Res., 43, 1869‐1874 (1983)

2) H. Okada, T. Heya, Y. Ogawa, and T. Shimamoto, One‐month release injectable microspheres of a luteinizing hormone releasing hormone agonist (leuprolide acetate) for treating experimental endometriosis in rats. J. Pharmacol. Exp. Ther., 244(2), 744‐750 (1988)

3) H. Okada, Y. Doken, Y. Ogawa, and H. Toguchi, Preparation of three‐month depot injectable microspheres of leuprorelin acetate using biodegradable polymers. Pharm. Res., 11(8), 1143‐1147 (1994)

4) M. Fukuta, H. Okada, S. Iinuma, S. Yanai, and H. Toguchi, Insulin fragments as a carrier for peptide delivery across the blood‐brain barrier. Pharm. Res., 11(12), 1681‐1688 (1994)

5) H. Okada, One‐ and three‐month release injectable microspheres of the LH‐RH superagonist leuprorelin acetate. Advanced Drug Delivery Review, 28, 43‐70 (1997)

6) N. Murata, Y. Takashima, K. Toyoshima, M. Yamamoto, and H. Okada, Anti‐tumor effects of anti‐VEGF siRNA encapsulated with PLGA microspheres in mice. J. Contr. Rel., 126(3), 246‐254 (2008)

7) T. Kanazawa Y. Takashima, S. Hirayama, and H. Okada, Effect of menstrual cycle on gene transfection through mouse vagina for DNA mucosal vaccine. Int. J. Pharm., 360(1‐2), 164‐170 (2008)

8) K. Tanaka, T. Kanazawa, Y. Shibata, Y. Suda, T. Fukuda, Y. Takashima, and H. Okada, Development of cell‐penetrating peptide‐modified MPEG‐PCL diblock copolymeric nanoparticles for systemic gene delivery. Int. J. Pharm., 396(1‐2), 229‐238 (2010)

9) T. Uchida, T. Kanazawa, M. Kawai, Y. Takashima, and H. Okada, Therapeutic effects on atopic dermatitis by anti‐RelA siRNA combined with functional peptides, Tat and AT1002, J. Pharmacol. Exp. Ther., 338, 443‐450 (2011)

10) H. Okada, Targeted siRNA therapy using cytoplasm‐responsive nanocarriers and cell‐penetrating peptides. J Pharm Inv 44, 505‐516 (2014)


NAME : Tetsuya OZEKI

AFFILIATION AND ADDRESS : Nagoya City University (NCU), Drug Delivery and Nano Pharmaceutics, Graduate School of Pharmaceutical Sciences, 3‐1 Tanabe‐dori, Mizuho‐ku, Nagoya, Aichi 467‐8603, Japan

TELEPHONE N° : +81 52 836 3463 FAX N° : +81 52 836 3463

E‐MAIL: [email protected]‐cu.ac.jp

CAREER HISTORY : 1995 Ph.D. at the Tokyo University of Pharmacy and Life Sciences 1995 Research Associate, Tokyo University of Pharmacy and Life Sciences 1999 Assistant Professor, Tokyo University of Pharmacy and Life Sciences 2003 Visiting Researcher, University of Kansas (Prof. Val. J. Stella) 2006 Associate Professor, Tokyo University of Pharmacy and Life Sciences 2009 Full Professor, Nagoya City University (NCU) 2013 Concurrent professor of graduate school between NCU and

Nagoya Institute of Technology (NIT)

SUMMARIES OF PRESENT WORKS ◆ Nanoparticles DDS for Inhalation Therapy of Tuberculosis ◆ Nanoparticles for Water‐insoluble drugs ◆ A novel HDL‐like gold nanoparticles formulation ◆ Stimuli‐responsive liposome formulation ◆ Nanoparticles DDS for Malaria Therapy ◆ Preparation of Drug Cocrystal Using Spray Dryer

RECENT MAJOR PUBLICATIONS : 1) Moeko Taki, Tatsuaki Tagami, Kaori Fukushige and Tetsuya Ozeki, Fabrication od nanocomposite

particles using a two‐solution mixing‐type spray nozzle for use in an inhaled curcumin formulation, Int. J. Pharm., in press

2) Hideki Mizusako et al., Active Drug Targeting of a Folate‐Based Cyclodextrin‐Doxorubicin Conjugate and the Cytotoxic Effect on Drug‐Resistant Mammary Tumor Cells In Vitro. J Pharm Sci. (2015) 104(9):2934‐40.

3) Tatsuaki Tagami et al., Effective Remote Loading of Doxorubicin into DPPC/Poloxamer 188 Hybrid Liposome to Retain Thermosensitive Property and the Assessment of Carrier‐Based Acute Cytotoxicity for Pulmonary Administration. J Pharm Sci. (2015) 104(11):3824‐32.

4) Tatsuaki Tagami et al., Simple and effective preparation of nano‐pulverized curcumin by femtosecond laser ablation and the cytotoxic effect on C6 rat glioma cells in vitro. Int J Pharm. (2014) 468(1‐2):91‐6.

5) Tetsuya Ozeki et al., Development of Novel and Customizable Two‐Solution Mixing Type Nozzle for One‐step Preparation of Nanoparticle‐containing Microparticles, Biol. Pharm. Bull., 35(11), 1926‐1931(2012)


NAME : Ling PENG AFFILIATION AND ADDRESS : Centre Interdisciplinaire de Nanoscience de Marseille (CINaM), Aix‐Marseille University, CNRS, 13288 Marseille FRANCE

TELEPHONE N° : 00 33 6 1724 8164 FAX N° : 00 33 4 9182 9301 E‐MAIL : ling.peng@univ‐amu.fr CAREER HISTORY :

11/1987 – 01/1993 PhD, Swiss Federal Institute of Technology, Zurich, Switzerland 02/1993 – 09/1997 Post‐doctoral researcher, Louis Pasteur University, Strasbourg, France 10/1997 – 03/1999 Research scientist of CNRS, Strasbourg, France since 03/1999 Tenured research scientist in CNRS, Strasbourg, France since 12/1999 Principal Investigator in CNRS, Marseille, France since 10/2008 CNRS Research Director in CINaM, Marseilles, France

SUMMARY OF PRESENT WORK :

‐ Functional Dendrimers for Gene and Drug Delivery ‐ Theranostics ‐ Personalized medicine and precision medicine


1) Laurent S, Chen H, Bedu S, Ziarelli F, Peng L, Zhang C‐C. Proc. Natl. Acad. Sci. U.S.A. 2005, 102, 9907‐9912.

2) Zhou J, Wu J, Hafdi N, Behr J‐P, Erbacher P, Peng L, "PAMAM dendrimers for efficient siRNA delivery and potent gene silencing", Chem. Commun. 2006, 2362‐2364

3) Yu T, Liu X, Bolcato‐Bellemin A‐L, Wang Y, Liu C, Erbacher P, Qu F, Rocchi P, Behr J‐P, Peng L, Angew. Chem. Int. Ed. 2012, 51, 8478‐8484.

4) Liu X, Zhou J, Yu T, Chen C, Cheng Q, Sengupta K, Huang Y, Li H, Liu C, Wang Y, Posocco P, Wang M, Cui Q, Giorgio S, Fermeglia M, Qu F, Pricl S, Shi Y, Liang Z, Rocchi P, Rossi JJ, Peng L, Angew. Chem. Int. Ed., 2014, 53, 11822.

5) Wei T, Chen C, Liu J, Liu C, Posocco P, Liu X, Cheng Q, Huo S, Liang Z, Fermeglia M, Pricl S, Liang XJ, Rocchi P, Peng L, Proc. Natl. Acad. Sci. U.S.A. 2015, 112, 2978

6) Cui Q, Yang S, Ye P, Tian E, Sun G, Zhou J, Sun G, Liu X, Chen C, Murai K, Zhao C, Azizian KT, Yang L, Warden C, Wu X, D’Apuzzo M, Brown C, Badie B, Peng L, Riggs A, Rossi JJ, Shi Y, Nat. Commun. 2016, 7,10637

7) Chen C, Posocco P, Liu X, Cheng Q, Laurini E, Zhou J, Liu C, Wang Y, Tang J, Dal Col V, Yu T, Giorgio S, Fermeglia M, Qu F, Liang Z, Rossi JJ, Liu M, Rocchi P, Pricl S, Peng L, Small. 2016, doi: 10.1002/smll.201503866.


NAME : Joël RICHARD

AFFILIATION AND ADDRESS: IPSEN 20, Rue Ethé VIRTON 28100 DREUX CEDEX France

TELEPHONE N° : +33 237 654 610

FAX No: +33 237 654 635


CAREER HISTORY : Dr Joël Richard graduated from Ecole Normale Supérieure (1984, Cachan, France). He then got his PhD in Materials Science/Colloids & Interface Science from University of Paris VI (1987) and Habilitation à Diriger les Recherches from University of Bordeaux I (1993). Dr Richard has more than 25 years of experience in chemistry and biopharmaceutical R&D, including several global senior leadership positions in various Biotech and Pharma companies, such as Ipsen, Merck Serono, Serono and Ethypharm. He also had an entrepreneurial experience, co‐funding Mainelab (France), a drug delivery company specialized in developing solvent‐free processes for protein delivery systems (1999‐2004). Dr Richard is presently leading all the CMC Development activities of Peptides & Small Molecules for the Ipsen Company, from Pre‐Clinical Development to Phase III, Filing, Registration and Life Cycle Management.

SUMMARY OF PRESENT WORK : Dr Richard has focused his research activity on new formulations and drug delivery systems (such

as microspheres, nanoparticles, chemically‐modified proteins, supercritical fluid technology ...),

especially for injectable protein and peptide formulations. Dr Richard has published 67 peer‐

reviewed scientific papers, 8 book chapters and 2 editorials in various fields (colloids and

interfaces, drug delivery, supercritical fluids, protein formulations, sustained‐release

formulations…). He is the author of more than 120 international communications and 53 patent

families.

MAJOR PUBLICATIONS: 1) Oral peptide delivery: technology status landscape and current status. L.N. Hassani, A.L. Lewis, J.

Richard, ONdrugDELIVERY (2015) 59, 12‐17. 2) Challenges in the delivery of peptide drugs: An industry perspective. A.L. Lewis and

J. Richard, Therapeutic Delivery, (2015) 6(2), 149–163. 3) Parenteral biologics delivery: recent progresses, key challenges and perspectives. J. Richard,

European Journal of Parenteral &Pharmaceutical Sciences 17 (3), 2012, 94‐109. 4) Challenges and opportunities in the delivery of cancer therapeutics. J. Richard, Therapeutic Delivery

2 (1), 2011, 107‐121. 5) The formulation and immunogenicity of therapeutic proteins: product quality is a key factor. J.

Richard, N. Prang, IDrugs 13 (8), 2010, 550‐558. 6) Formulation strategies based on supercritical fluid technology for the delivery of

biopharmaceuticals. J. Richard, Chemistry Today 26 (4), 2008, 48 –51.


NAME: Shigeki SASAKI AFFILIATION AND ADDRESS: Graduate School of Pharmaceutical Sciences, Kyushu University 3‐1‐1 Maidashi, Higashi‐ku, Fukuoka 812‐8582, Japan TELEPHONE No: +81‐92‐642‐6615

FAX N°: +81‐92‐642‐6615


CAREER HISTORY : 1977, Bachelor, Faculty of Pharmaceutical Sciences, The University of Tokyo. 1982, Doctor Degree, Faculty of Pharmaceutical Sciences, The University of Tokyo. 1982‐1984, Postdoctoral fellow at Department of Chemistry, Indiana University, USA. 1984‐1990, Assistant Professor, The University of Tokyo. 1990‐2002, Associate Professor, Kyushu University. 2002‐ Professor at Graduate School of Pharmaceutical Sciences, Kyushu University. SUMMARY OF PRESENT WORK: Design and synthesis of functional nucleosides and nucleotides for sequence specific modulation of gene expression.

MAJOR PUBLICATIONS : 1) Taniguchi Y., Kikukawa Y and Sasaki S. Discrimination Between 8‐Oxo‐2'‐Deoxy‐ guanosine and 2'‐

Deoxyguanosine in DNA by the Single Nucleotide Primer Extension Reaction with Adap TriphosphateAngew. Chem. Int. Ed. 54, 5147‐5151 (2015).

2) Jitsuzaki D., Onizuka K., Nishimoto A., Taniguchi Y. and Sasaki S.* Remarkable acceleration of the DNA/RNA inter‐strand functionality‐transfer reaction to modify a cytosine residue: the proximity effect via complexation with a metal cation, Nucleic Acids Res., 42, 8808‐8815 (2014).

3) Nishimoto A., Jitsuzaki D., Onizuka K., Taniguchi Y., Nagatsugi F. and Sasaki S.* 4‐Vinyl‐Substituted Pyrimidine Nucleosides Exhibit the Efficient and Selective Formation of Interstrand Cross‐Links with RNA and duplex DNA, Nucleic Acids Res.,41, 6774‐6781 (2013).

4) Taniguchi Y.* and Sasaki S.* An efficient antigene activity and antiproliferative effect by targeting the Bcl‐2 or survivin gene with triplex forming oligonucleotides containing a W‐shaped nucleoside analogue (WNA‐�T),Org. Biomol. Chem., 10, 8336‐8341. (2012).

5) Y. Taniguchi, R. Kawaguchi, S. Sasaki, Adenosine‐1,3‐diazaphenoxazine Derivative (Adap) for Selective Base Pair Formation with 8‐Oxo‐2’‐deoxyguanosine in DNA, J. Am. Chem. Soc. 133, 7272‐7275 (2011).

6) Onizuka K, Taniguchi Y., Sasaki S., A New Usage of Functionalized Oligodeoxy‐ nucleotide Probe for Site‐SpecificModification of a Guanine Base within RNA, Nucleic Acids Res. 38, 1760‐1766 (2010).


NAME : Juergen SIEPMANN

AFFILIATION AND ADDRESS :

University of Lille, College of Pharmacy INSERM U1008: Controlled Drug Delivery Systems and Biomaterials 3, rue du Professeur Laguesse, 59000 Lille, France

TELEPHONE N°: +33‐3 20 96 47 08 E‐MAIL: juergen.siepmann@univ‐lille2.fr CAREER HISTORY :

Juergen Siepmann studied Pharmacy and obtained his Ph.D. at the Freie Universitaet Berlin, Germany. Since 2004 he is Professor of Pharmaceutical Technology at the University of Lille, France. His awards include the “2012 APV Research Award for Outstanding Achievements in the Pharmaceutical Sciences”. He was visiting scientist/postdoctoral fellow at Purdue University, the University of Paris‐Sud, the University of Iowa and the University of Angers. So far, he published his work in more than 150 articles in international scientific journals, 250 poster and 100 oral presentations at national and international scientific meetings. He is editor of 3 books and author of 16 book chapters. Since 2010 Prof. Siepmann is heading the INSERM research group “Controlled Drug Delivery Systems and Biomaterials” (about 7 professors, 5 lecturers, 5 technical/administrative staff, 15 post‐docs & PhD students and 10 master students). In 2015 Prof. Siepmann served as Editor‐in‐Chief of the "Journal of Drug Delivery Sciences and Technology", since 2016 he is Editor‐in‐Chief of the “International Journal of Pharmaceutics”. Since 2010 he serves as president of the APGI (“International Society of Drug Delivery Sciences and Technology”). SUMMARY OF PRESENT WORK :

Prof. Siepmann’s research focuses on controlled drug delivery systems, in particular on the elucidation of the underlying mass transport phenomena and the optimization of the devices. Different types of systems (e.g. miniaturized implants for the inner ear, in‐situ forming implants for periodontitis treatment, coated pellets for colon targeting) are prepared and physico‐chemically characterized. Based on the experimental results, mechanistically realistic mathematical models are developed, able to predict the impact of the device design on the system performance. MAJOR PUBLICATIONS :

Siepmann, J; Peppas, NA. Hydrophilic matrices for controlled drug delivery: An improved mathematical model to predict the resulting drug release kinetics (the “sequential layer” model). Pharm. Res. 17, 1290‐1298, 2000.

Klose, D; Siepmann, F; Elkharraz, K; Krenzlin, S; Siepmann, J. How porosity and size affect the drug release mechanisms from PLGA‐based microparticles. Int. J. Pharm. 314, 198‐206, 2006.

Rosiaux, Y; Muschert, S; Chokshi, R; Leclercq, B; Siepmann, F; Siepmann, J. Ethanol‐resistant polymeric film coatings for controlled drug delivery. J. Control. Release 169, 1‐9, 2013.

Gasmi, H; Danede, F; Siepmann, J; Siepmann, F. Does PLGA microparticle swelling control drug release? New insight based on single particle swelling studies. J. Control. Release 213, 120‐127, 2015.

Fahier, J; Muschert, S; Fayard, B; Velghe, C; Byrne, G; Doucet, J; Siepmann, F; Siepmann, J. Importance of air bubbles in the core of coated pellets: Synchrotron X‐ray microtomography allows for new insights. J. Control. Release 237, 125‐137, 2016.


NAME : Ikuko TSUKAMOTO AFFILIATION AND ADDRESS : Dept. of Pharmaco‐Bio‐Informatics, Fac. of Med., Kagawa University Ikenobe, Miki, Kagawa, 761‐0793, Japan TEL/FAX +81‐(0)87‐ 898‐2383

E‐MAIL [email protected]‐u.ac.jp

CAREER HISTORY :

2016 Professor(Visiting), Kagawa University 2001 Associate Professor (Visiting), Kagawa Medical University (Since 2003, Fac. of Med., Kagawa Univ.) 1994 PhD in Medicine, Kagawa Medical University 1987 Assistant Professor, Dept. of Anesthesiology and Emergency Medicine 1986 Assistant Professor, Dept. of Radiology 1985 Assistant Professor, Dept. of Biochemistry, Kagawa Medical University 1985 PhD in Pharmaceutical Science, Kyoto University

SUMMARY OF PRESENT WORK: Cell physiology, Analytical Chemistry, Transdermal Delivery System

MAJOR PUBLICATIONS:

1) Effects of a precursor of endocrine disrupters on the interactions between acetylcholinesterase and halothane ‐Roles of biomembranes ‐ I. Tsukamoto et al. Progress in Anesthetic Mechanism 6, 552 (2000)

2) Fourier Transform Infrared (FTIR) analysis of volatile compounds in expired gas for the monitoring of poisonings ‐1.Ethanol ‐ T. Nishiyama et al. Pharm. Res. 18, 125 (2001)

3) A novel nucleic acid analogue shows strong angiogenic activity ‐ I. Tsukamoto et al. Biochem. Biophys. Res. Com. 399, 699 (2010)

4) Delayed administration of the nucleic acid analog 2Cl‐C.OXT‐A attenuates brain damage and enhances functional recovery after ischemic stroke ‐ N. Okabe et al. Brain Res. 1506, 115 (2013)

5) Involvement of S1P1 receptor pathway in angiogenic effects of a novel adenosine‐like nucleic acid analog COA‐Cl in cultured human vascular endothelial cells ‐ J. Igarashi et al. Pharma. Res. Per. 2, e00068 (2014)

6) Intestinal absorption, organ distribution, and urinary excretion of the rare sugar D‐psicose ‐ I. Tsukamoto et al. Drug Design, Development and Therapy 8, 1955 (2014)

7) A Key Role of PGC‐1a Transcriptional Coactivator in Production of VEGF by a Novel Angiogenic Agent COA‐Cl in Cultured Human Fibroblasts


NAME : Hiromitsu YAMAMOTO AFFILIATION AND ADDRESS : Aichi Gakuin University, School of Pharmacy, Lab. of Pharmaceutical Engineering 1‐100 Kusumoto

TELEPHONE N° : +81‐52‐757‐6770

FAX N° : +81‐58‐757‐6799


CAREER HISTORY : 1995/04 Assistant Professor of Laboratory of Pharmaceutical Engineering, Faculty of

Pharmacy, Gifu Pharmaceutical University, Japan 1999/10 Ph D thesis, Laboratory of Pharmaceutical Engineering, Gifu Pharmaceutical

University, Gifu, Japan (supervised by Prof. Y. Kawashima) 2004/02‐2005/01 Research Fellow in the institute of Pharmaceutical Technology at the Johann

Wolfgang Goethe University (German) 2006/04‐2012/03 Associate Professor at the Aichi Gakuin University in the Laboratory of

Pharmaceutical Engineering 2012/04‐ Professor at the Aichi Gakuin University in the Laboratory of Pharmaceutical

Engineering SUMMARY OF PRESENT WORK : Our researches are based on the powder technology. Especially, we focused on Drug Delivery System with surface modified polymeric nanoparticles and solubility enhancement of poorly water soluble drug by solid dispersion technique and inclusion complex with cyclodextrin. MAJOR PUBLICATIONS :

1) Nanomedical system for nucleic acid drugs created with the biodegradable nanoparticle platform., J Microencapsul.29, pp.54‐62 (2012)

2) Oral nuclear factor‐kB decoy oligonucleotides delivery system with chitosan modified poly (D,L‐lactide‐co‐glycolide) nanospheres for inflammatory bowel disease., Biomaterials32,pp.870‐878 (2011)

3) Observation of antibacterial effect of biodegradable polymeric nanoparticles on Staphylococcus epidermidis biofilm using FE‐SEM with an ionic liquid., Microscopy (Oxf) (2015)

4) Application of spherical silicate to prepare solid dispersion dosage forms with aqueous polymers, Int J Pharm. 493, pp.55‐62 (2015)

5) Physicochemical characterization of cyclodextrin‐drug interactions in the solid state and the effect of water on these interactions., J Pharm Sci., 104(3), pp.942‐954 (2015)


NAME: Fumiyoshi YAMASHITA


Graduate School of Pharmaceutical Sciences Kyoto University 46‐29 Yoshidashimoadachi‐cho, Sakyo‐ku, Kyoto 606‐8501 Japan TELEPHONE No: +81‐75‐75‐4535

FAX No: +81‐75‐753‐4575

E‐MAIL: [email protected]‐u.ac.jp

CAREER HISTORY: Dr. Fumiyoshi Yamashita graduated from School of Pharmaceutical Sciences, Kyoto University in 1988 and obtained his Ph.D. degree in Pharmaceutical Sciences from Kyoto University in 1995. He was appointed Research Assistant Professor at Kyoto University in 1991. Since 1997, he has been Associate Professor of Graduate School of Pharmaceutical Sciences, Kyoto University. He received Young Investigator Award of Academy of Pharmaceutical Science and Technology Japan (APSTJ) in 2001 and Young Investigator Award of Japanese Society for the Study of Xenobiotics (JSSX) in 2003. SUMMARY OF PRESENT WORK : His current research interests include ADME‐related chemoinformatics, system pharmacology

modeling, and development of receptor‐mediated drug delivery systems.


Yamashita, F and Hashida, M: Pharmacokinetic considerations for targeted drug delivery. Adv Drug Deliv

Rev, 65, 139‐147 (2013).

france-japan drug delivery systems symposium “recent ...€¦ · symposium “recent achievements...

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