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Construction and function of next generation carbonic anhydrase IX retargeted T cells for cancer immunotherapy

Grant number: 14/15739-4
Support type:Scholarships abroad - Research
Effective date (Start): December 01, 2014
Effective date (End): July 31, 2015
Field of knowledge:Health Sciences - Medicine
Principal Investigator:Eloah Rabello Suarez
Grantee:Eloah Rabello Suarez
Host: Wayne Anthony Marasco
Home Institution: Faculdade de Medicina do ABC (FMABC). Organização Social de Saúde. Fundação do ABC. Santo André , SP, Brazil
Local de pesquisa : Harvard University, Boston, United States  

Abstract

Carbonic anhydrase IX (CAIX) is an enzyme responsible for carbon dioxide hydration that is overexpressed in many hypoxic solid tumors, but not in normal tissues. Besides CAIX expression, many tumors overexpress programmed death-ligand 1 (PD-L1). PD-L1 binds to PD-1 receptor in T cells, B cells, and natural killer, resulting in an anergic phenotype of these cells that cannot restrain tumor progression. Many cancer cells can also downregulate MHC expression, inhibiting their recognition by the immune system. Therapeutic strategies targeting the recognition of tumor cells by T-cells and its activation without the necessity of MHC recognition by T-cell receptor (TCR) are very promising. A modified TCR called chimeric antigen receptor (CAR) containing single chain fragment variable (scFv) previously selected by high affinity against a tumor antigen is a current option to counteract this problem. Dr. Marasco's group, from Harvard Medical School, has recently developed a 2nd generation Anti-CAIX CAR that utilizes the signaling motifs of CD28 co-stimulatory molecule and CD3 zeta chain; which was transduced to T cells. The anti CAIX CAR T cells had strong anti-tumor effects in renal cell cancer (RCC) treatment both in vitro and in vivo. These results prompted us to propose adding anti-PD-L1 checkpoint control blockade to further optimize the effector functions of Anti-CAIX CAR T cells. Specifically for this project we will construct an anti-PD-L1 minibody (scFv-Fc) by cloning of anti-PD-L1 scFv, in a plasmid vector containing the Fc sequence and this minibody will be subcloned in a lentivirus containing the anti-CAIX CAR and transduced in T-cells. The effects of CART-cells in antigen and complement dependent cytotoxicity, viability, induction of cytokines release and clonal expansion will be evaluated in RCC cells. An orthotopic RCC model will be used for in vivo evaluation. Tumor specific targeted scFv-CART-cells that release antibodies in the tumor microenvironment are a next generation of CAR constructs with an incredible potential to markedly improve cancer treatment. (AU)