Research Strategy for Phyto Pharmaceuticals

Dr. Arun Balakrishnan Sr. Vice President

Head - Operations, Natural Product & External Liaison

Brief description of the use of Natural products in drug discovery and research.

Use of Natural Product plant extracts as Phyto Pharmaceuticals.

Regulatory challenges in Herbal product registrations.

Outline of the talk

3

• Natural Products have been applied to human healthcare for thousands of years.

• Drug Discovery in ancient times was by chance and was based on clinical practices.

• Understanding therapeutic benefits deepens Natural Product and increase previously serendipitous discoveries.

• Evolve into active researches for new medicines.

• Progress in Life Sciences has not only revealed many pathological processes of diseases and also the molecular mechanisms.

• Leads to the development of molecular and cellular assays in conjunction with HTS.

• The transition from traditional to empirical and molecular screening will increase the probability of discovering new leads and drug candidate from Natural products.

Natural Products and Healthcare

CONFIDENTIAL • Natural product and obtaining pure material in large quantities is generally difficult,

if the active molecule is present in small amounts.

• HTS and the new biological assays can handle microgram or less amounts to test invitro.

• This is advantageous since many Natural products are difficult to isolate, purify and many a times difficult to synthesize.

• However, innovations in combinatorial chemistry were large and structurally diverse chemical libraries can be generated including parallel synthesis have also helped in producing several analogs faster using NP scaffolds.

• Innovations in computer applications, automation technologies, microfluid management, software design has enhanced speed to screen large number of compounds in short time.

• Identify HITS, that can be developed into a potential drug lead with desired therapeutic actively.

Automation in Biology - Natural Product approach to

isolation faster

New approaches in Natural Products Chemistry

• Many pharma companies find Natural Products unattractive owing to the difficulties

concerning isolation and structure elucidation of bioactive natural products.

• However, more rapid methodologies by which purification can be enhanced, have made people to relook into NP resources.

• Using High Performance Liquid Chromatography (HPLC) and subsequent fractionation analysis by Liquid Chromatography / Man Spectrometry (LC / MS).

• Combining MS data with those from libraries of known compounds, net novel molecules can be distinguished – Dereplication.

• Kills projects without novel compounds in the beginning itself.

• Automated sample injection and fraction collection enables to study multiple samples at one time.

Screening for Bio-molecules from microbial diversity collected from different ecological niches

Screening for Bio-molecules from microbial diversity collected from different ecological niches

•The largest screening program done in the country. •245000 microbes and > 600000 extracts •Funded by DBT, New Delhi

DBT – PEL Public Private Partnership in Drug Discovery

High Throughput Screening 7

Effluent Treatment plants /contaminated sites Western Ghats,

Insect guts

Hot Springs of Himachal

Pradesh

Extremophiles from Orissa, Bihar and West Bengal

Wet land ecosystems of North West India

Eastern Ghats, and Mangroves area

North East area

River Sediments

7000 isolates /month

Marine isolates

NIO IGIB

ILS

Univ

Delhi

NCCS NEERI

GNDU

MSSRF

IBSD

High Throughput Screening 8

NCCS

GNDU

Univ

Delhi

ILS

IBSD

MSSRF

NEERI

IGIB

PLSL

NIO

Natural Products Phytotechnology

Year Trade Name Lead Compound Disease area Class

2000 Artemotil Artemisinin Antimalarial Sesquiterpene lactone; Semi-synthetic NP

2002 Orfadin Leptospermone antityrosinaemia NP-derived

2002 Reminyl Galantamine Alzheimer's disease Alkaloid; NP

2004 Apokyn Morphine Parkinson's disease Alkaloid; semi-synthetic NP

2004 Spiriva Atropine Chronic obstructive pulmonary

disease Alkaloid; semi-synthetic NP

2005 Sativex Dronabinol/ Cannabidol Pain Cannabinoids; NP

2007 Vyvanse Amphetamine Attention deficit hyperactivity

disorder Amine; NP-derived

2008 Relistor Naltrexone Opioid-induced constipation

and pain Alkaloid; NP-derived

2009 Qutenza Capsaicin Pain Vaniloid; NP

2013 Crofelemer Oligomeric proanthocyanidin Antidiarrheal Agents Polyphenolic; NP

Plant-derived drugs launched (2000- 2010)

Confidential

Lead compound/Name Compound class Disease area Development

status Developer

Epipodophyllotoxin Alkaloid Oncology Phase I Pierre Fabre Laboratories

Curcumin Polyphenol Inflammation; oncology Phase I/ II Various concerns worldwide

Betulinic acid/ Bevirimat Triterpenoid Anti-HIV Phase II Panacos Pharmaceuticals

Castanospermine/ Celgosivir Indolizine alkaloid Antiviral (HCV) Phase II MIGENIX

4-Methylumbelliferone/ Heparvit Coumarin Antiviral (HCV & HBV) Phase II MTmedical Inst. of Health and BioMonde

Huperzine-A Alkaloid Alzheimer's Disease Phase II Chinese scientists

Lobeline Piperidine alkaloid Neurological Phase II Yaupon Therapeutics and NIH

Resveratrol Triphenolic Stibeline Diabetes & obesity Phase II Sirtris Pharmaceuticals

Genistein Isoflavone Oncology Phase II Astellas, Bausch & Lomb

Phlorizin/ Dapagliflozin Polyphenolic glycoside Antidiabetic Phase III Bristol-Myers Squibb

Himbacine Alkaloid Cardiovascular Phase III Schering-Plough

Camptothecin/Karenitecin Quinoline alkaloid Oncology Phase III BioNumerk and ASKA Pharmaceutical

Combretastatin A-4/ Zybrestat Stilbenoid phenol Oncology Phase III OXiGENE

Paclitaxel/ Cabazitaxel Diterpene taxoid Oncology Phase III Sanofi-Aventis

Vinblastine/ Vinflunine Vinca alkaloids Oncology Phase III Pierre Fabre Laboratories

Homoharringtonine Alkaloid Oncology Phase III ChemGenex

Daidzein Isoflavone Oncology Phase III Marshall Edwards (Novogen)

Plant-derived compounds in clinical trials

Confidential

•The global market for botanical and plant-derived drugs was expected to increase from $19.5 billion in 2008 to $32.9 billion in 2013, a compound annual growth rate (CAGR) of 11.0%.

•Botanical drugs generated $127,000.0 in 2008. This segment was expected to reach $2.4 billion in 2013 for a CAGR of 651.7%.

•All other plant derived drugs generated $19.5 billion in 2008. This was expected to reach $30.5 billion in 2013, for a CAGR of 9.4%.

Global market for botanicals

Confidential

Moist deciduous forests, Maharashtra

Pine forest: Almora, Uttarakhand

Valley of Flowers, Garhwal Himalaya

Plant

Collection Sites

Distribution of Plant Collection Sites

Plant

Diversity in

India

Medicinal text: 25-volume Descriptive Catalogue of Palm-leaf Manuscripts (Institute of Asian Studies )

Plants in Indian Traditional

Medicines

ISM: Knowledge base

Confidential

In monasteries

Which stream?

Herbal NCE

Bioactive

Lead

Synthetic/ Semi-synthetic

(Lower tox. & high efficacy

Assure Patent protection

Standardized extract

Biomarkers

Novel Formulations

(single/polyherbal)

FOCUS ON-

Leads from ethnomedicines

Random Collection- unexplored areas

Unique / endemic species

Allied of clinically proven species

Creation of Ethnomedicinal extract library

Evidence based Phyto-Pharmaceutical

ANTIVIRAL - Viracure

Phase II

Coleus forskohlii - Forskolin

Traditional use: Digestive, Respiratory, Circulatory, Nervous disorders, Antipain

Modern use: Included in many formulations for obesity

(e.g. Lipitrex ), glaucoma, hypertension etc. Effect

of Coleus forskohlii on various types of cancer

being evaluated

Primary Hits 722

Primary Screening

No. of samples: 5200

Med lit. search

Selected:

226

Sec. Actives: 93

Prioritized species

52

ANTIINFLAMMATION

Screening Development Cultivation

Evidence-based Phyto-pharmaceuticals

NCE / NCA

Project workflow

• Phase 1: dereplication – Small-scale samples – Dereplication for known

compounds – Prioritisation

• Phase 2: large scale

fractionation – Large-scale samples – Automated fractionation – Analysis of fractions

• Phase 3: purification &

structure elucidation – Final purification – Analysis of purified

fractions – Structure elucidation

Analytical HPLC/MSMS of extracts

Retain aliquot Send fractions for testing

Analyse MSMS data HR-MSMS of active fractions

Phase 1 Report

Prepare large-scale extract

Automated prep HPLC fractionation

Send aliquots for testing

Collect MSMS data

Bioassay of fractions

HR-MSMS & NMR of active fractions

Further material required? yes

no

Structure elucidation

Select samples for isolation

Phase 1:

Phase 2:

Phase 2 Report

Further purification

HR-MSMS & NMR of active fractions

Phase 3 Report

Phase 3:

Bioassay of fractions

Send aliquots for testing

Bioassay of fractions

Quality Raw material

Insights from Traditional Chinese medicines: Quality Control

Regulatory Challenges in Herbal Product Registration

Regulatory Challenges

• Standardization of Products

• Collection strategy

• Controls on adulterants

• Reproducibility

• Safety & Efficacy

Standardization of Products

Herbs are natural products and chemical composition varies depending upon factors:

• Botanical species.

• Used chemotypes.

• Anatomical part used (seed, flower. Root, leaf etc).

• Storage: Sun, humidity, type of ground time of harvest.

• Concentration of active principle or Biomarker.

• Ascertain consistent chemical profile and biological activity.

Need validation of above as part of regulatory process as lack of information about composition can turn to be black box for the herbal based treatment.

Collection Strategy

It is observed that concentration of bio-marker and active principal varies due to:

• Weather conditions.

• Soil quality.

• Ground water and Moisture content.

• Season etc.

To ensure consistent quality product:

• Need selection and validation of Collection strategy from wild or Good validated harvesting practices.

• Need good manufacturing practices for extraction modes and related parameters.

Control on Adulterants

Regulatory approval require to ascertain control of adulterants which can be found in the herbal material through different sources:

• Pesticide residue (more likely if it is harvested in the agricultural fields)

• Aflatoxin content

• Bacterial/fungal growth

• Heavy metal contamination

The control need to be demonstrated from collection through different processing including extraction and final product manufacturing.

Reproducibility

One of the biggest challenge due to variation in chemical profile due to various factors which can lead to possible variation in biological activity:

• Must to identify and characterize the bio-marker or active principal (constituent).

• Repetitive testing different batches to control batch to batch variation.

• Establish the relation of active marker or constituent through pre-clinical and clinical studies.

• Standardize collection strategy and validate the process to reduce the variability.

• Develop appropriate analytical method to analyze the bio-marker or active principal.

29

Extraction methods, physico-chemical tests, microbial loads, heavy metal contaminants, chromatographic finger print profile with phytochemical reference markers, assay for active constituents or characteristic markers, if active constituents are not known.

Data on the active Phytopharmaceutical establishing the natural window of constituents (marker compounds) reflecting geographical and seasonal changes in a year, wherever applicable.

Information on any excipients (diluents or builders or stabilizers or preservatives used, if any) and their proportions.

Details of packaging of the extract or Phytopharmaceutical, storage conditions and labeling.

Processing of extract or Phytopharmaceutical (if used as an extract)

30

Composition, dosage, excipients, stabilizers, agents used, packaging materials, test(s) for the identification for the Phytopharmaceutical.

Quality specifications for actives and inactive Phytopharmaceutical, chromatographic finger print profile with phytochemical reference maker(s) and assay for active constituent(s) or characteristic maker(s), if active constituents are not known.

• Manufacturing process of formulation • Stability data : for 0, 1, 2, 3 and 6 months • Safety and pharmacological information

Provide comparative chemical and chromatographic profile, and spectroscopic information

• Formulation of Phytopharmaceutical drug applied for :

Safety and Efficacy

• There is very limited scientific evidence to establish the safety and efficacy of the herbal products.

• Out of top herbal products which has scientific evidence of efficacy has some concern over safety.

• More advance technologies need to be used to characterize the herbal products to control the variance and establish the safety and efficacy.

• Regulatory need more scientific / evidence based medicine which are designed through pre-clinical and clinical trial studies instead of tradition or supposed millenarian belief.

• Need more funding for research and understanding of herbal products to meet regulatory requirements.

• Current regulation of standard medical therapies can not be applied to herbal products, may need regulation which is suitable to herbal products.

32

28 to 90 days repeat dose oral toxicity on two species of animals of which one should be non rodent as per Schedule Y;

In-vitro Genotoxicity data (Ame’s test and Chromosomal aberration test as per Schedule Y);

Dermal toxicity tests for topical use products;

Teratogenicity study (only if Phytopharmaceutical (s) is intended for usage during pregnancy).

• Animal toxicity and safety data :

33

For all Phytopharmaceutical- Data from Phase I (Max. Tolerated Dose and associated toxicities) and the protocols to be submitted prior to performing the studies.

Data of results of dose finding studies performed and the protocols shall be submitted prior to performing the studies.

If already marketed for > 5 years or there is adequate published evidence regarding the safety, the studies may be abbreviated, modified or relaxed.

• Human Studies :

34

• Proof of concept or confirmatory clinical trials Protocols for approval for any specific or special safety and efficacy study proposed specific to the product.

Proposed protocol for approval for human clinical studies appropriate to generate or validate safety and efficacy data for the dosage form or product.

Submit information on how the quality of the formulation would be maintained during the above studies.

• Regulatory status Status in any country under any category like functional food or dietary supplement or as traditional medicine or as an approved drug.

• Marketing Information • Post Marketing Surveillance [PMS]

3rd February 2009 Slide 30 Asha Almeida