University of Angers MiNt: Micro et Nanomedicines Biomimetiques France

Instituto Oncologico Veneto Italy

Dr. Giovanna Lollo, MiNT, France

Dr. Ilaria Marigo, IOV, Italy

Nano Immuno Chemotherapy to treat cancers

Lymphonanocarriers for the treatment of Metastatic Cancer

LYMPHOTARG

The LYMPHOTARG project proposes to develop specifically targeted anticancer treatments, by associating anticancer drug to specific nanostructures composed of lipids and polymers, which have a specific affinity for the lymph nodes. In this way, we expect to prevent the process of metastatic spreading through lymphatic vessels.

50 nm

PEG-HS (Solutol)

Surfactant (Span 80)

Triglyceride (labrafac)

Cooling

Heating

60-85°C

Dilution

(Water 20°C)

Heating

W/O

PIZ

Temperature

O/W

Water

Oil

Design and development of Gemcitabine loaded LNC

Gem-C12

Gem-C12: Modified Gemcitabine

Biodistribution Studies

n=5

Biodistribution studies on healthy mice following i.v. and s.c. administration of fluorescent labelled LNC

LNC reach the lymph nodes and remain there for a while (4 days for i.v. and 2 weeks for s.c.). Accumulation in liver and spleen following i.v. and s.c. administration was also evidenced.

Biodistribution Studies

0 10 20 30 40

Day post tumor-graft

0.0

0.2

0.4

0.6

0.8

1.0

Cu

mu

lativ

e P

rop

ortio

n

Su

rviv

ing

salin iv

gemzar iv

gemC12

LNC blank iv

LNC gemC12 iv

LNC blank sc

LNC gemC12 sc

Antitumor efficacy C

um

ula

tive p

rop

ort

ion

su

rviv

ing

mic

e

SCID-CB17 mice grafted at day 0 with 2.104 Ma44-3 cells and treated at day 5, 7 and 9 for IV treatment and at day 5 and 9 for SC treatment (N = 10). A total maximal tolerated dose of 40mg/kg (equivalent molar of gemcitabine) was used to treat groups with Gemzar, GemC12 micelles (GemC12) or LNC GemC12.

P < 0.05

Evaluation of systemic side effects

• Hematologic toxicity

Complete blood count (CBC) complete granulocyte count (WBC)

platelet count (PLT)

WBC (G/L) RBC (T/L) HGB (g/dL) HCT (%) PLT (G/L)

Saline IV 0,2 ± 0,1 9,8 ± 0,3 15,1 ± 0,3 47 ± 1 469 ± 50

Gemzar IV 0,2 ± 0,1 9,2 ± 0,3 14,0 ± 0,4 45 ± 1 245 ± 63

GemC12 IV 0,3 ± 0,2 9,1 ± 0,6 14 ± 1 45 ± 2 275 ± 63

Lnc Blank IV 0,6 ± 0,3 9,8 ± 0,2 15,1 ± 0,3 47,7 ± 0,9 465 ± 44

Lnc GemC12 IV 0,14 ± 0,03 8,9 ± 0,2 13,7 ± 0,3 43,1 ± 0,9 284 ± 24

Lnc Blank SC 0,4 ± 0,1 9,4 ± 0,7 14 ± 1 46 ± 3 453 ± 20

Lnc GemC12 SC 0,26 ± 0,06 9,4 ± 0,5 14,5 ± 0,8 45 ± 2 281 ± 83

*

* P < 0,05 (Kruskall-

Wallis test)

Severe combined immunodefiency mice very low WBC level Deficient lymphocytes B and T Normal natural killer cells, macrophages, granulocytes Abnormal thymus and lymph nodes

Gemzar induce significant myelosuppression on SCID-CB17 mice as revealed by platelet count in comparaison to saline group. Neither blank LNC or LNC loaded by GemC12 delivered by i.v. or s.c. induced significant myelosuppresion

Next step....

The global objective of the project is to develop a novel, synergistic strategy between nanomedicine and immunotherapy to treat cancer. A double approach combining the relief of tumor-induced immunosuppression and the stimulation of a specific immune response against the tumor is proposed.

NICHE: Nano Immuno CHEmotherapy

Good nanocarrier (LNC) prototype from an immunological point of view was developed. Gem-loaded LNC suppress BM-MDSC differentiation in vitro and in vivo

Nano Immuno Carriers

Cross talk between neoplastic and remote sites, such as the bone marrow and spleen, determine the release of soluble factors that drive the accumulation of myeloid cells (MDSC); these myeloid cells subsequently promote neovascularization and metastasis.

Bronte and Gabrilovich, Nat. Rev. Immunol., 2010

LNC accumulate into spleen and could probably interact with the MDSC

Immunesuppressive cells impair cancer immunotherapy

Adoptive Cell Transfer therapy (ACT)

T T

Adoptively

transferred T cells

Endogenous suppressive

cells

Adapted from Restifo, Dudley and Rosenberg. Nature reviews in Immunology 2012

Primary tumor

Mechanism of action

Stimulate immune response

Spleen

Lymph node

Nanomedicine

Target

Revert immunosuppression

MDSC

Monocyte

Tumor microenvironment

TAM

Antigens

Cross Presentation Activation of CD8+ and CD4+

CRT HMGB-1 ATP

DC

CD8+ CD4+

Tumor cell

ICD

NICHE Project