TGF-β-responsive CAR-T cells promote anti-tumor immune function.

A chimeric antigen receptor (CAR) that responds to transforming growth factor beta (TGF-β) enables the engineering of T cells that convert this immunosuppressive cytokine into a potent T-cell stimulant. However, clinical translation of TGF-β CAR-T cells for cancer therapy requires the ability to productively combine TGF-β responsiveness with tumor-targeting specificity. Furthermore, the potential concern that contaminating, TGF-β?producing regulatory T (Treg) cells may preferentially expand during TGF-β CAR-T cell manufacturing and suppress effector T (Teff) cells demands careful evaluation. Here, we demonstrate that TGF-β CAR-T cells significantly improve the anti-tumor efficacy of neighboring cytotoxic T cells. Furthermore, the introduction of TGF-β CARs into mixed T-cell populations does not result in the preferential expansion of Treg cells, nor do TGF-β CAR-Treg cells cause CAR-mediated suppression of Teff cells. These results support the utility of incorporating TGF-β CARs in the development of adoptive T-cell therapy for cancer

Engineering bispecific chimeric antigen receptors to improve the efficacy of adoptive T-cell therapy

The recent FDA approval of CD19 chimeric antigen receptor (CAR) adoptive T-cell therapy for B-cell leukemias serves to highlight CAR-T cell therapy as a promising treatment approach for refractory cancers. More recently, adoptive transfer of T cells expressing CARs targeting B-cell maturation antigen (BCMA) has had numerous successes in clinical trial with 80-100% of multiple myeloma patients responding to treatment. However, CAR-T-cell therapy still faces several limitations including tumor antigen escape, a circumstance where tumor cells downregulate their surface antigen to avoid detection by CAR-T cells, and T-cell inhibition by cytokines such as transforming growth factor (TGF)-β in the solid tumor environment. These factors can significantly limit the efficacy of CAR-T-cell therapy. To overcome antigen escape, we designed single-chain bispecific CARs (OR-gate CARs) capable of signaling in the presence of two antigens instead of one. Using rational design principles, we constructed and evaluated CD19-OR-CD20 CARs that are able to prevent tumor antigen escape by CD19– leukemia. We further demonstrate that unlike single-input CD19 CARs, CD19-OR-CD20 CARs also prevent the emergence of spontaneous CD19-downregulated tumors in vivo. In a second study, we describe the rapid design and characterization of BCMA-OR-CS1 CARs and demonstrate that BCMA-OR-CS1 CARs can be rationally engineered to prevent antigen escape by BCMA– as well as CS1– myeloma cells. Finally, we explore the utility of the TGF-β CAR, a receptor capable of rewiring inhibitory TGF-β signaling to an activating response, in improving CD20 CAR function in TGF-β-rich environments. We evaluated three different bispecific targeting strategies, OR-gate CAR, DualCAR (co-expressing two receptors in one cells), and CARpool (pooling two different CAR-T cells), and demonstrate that TGF-β CAR-T cells are able to shield neighboring CD20 CAR-T cells from the inhibitory effects of TGF-β

Directed Evolution of Microcin J25 Towards Potency Against Resistant Escherichia coli

Microcin J25 (MccJ25) is a 21 amino acid antibacterial peptide whose lasso structure endows it with increased stability and resistance to many proteases. Capable of killing Gram-negative enterobacteria such as Escherichia coli and Salmonella newport, MccJ25 is a strong candidate for protein antibiotics. However, for MccJ25 to remain a promising antibiotic candidate, it must be shown that improved functionalities can be incorporated into the peptide even against a resistant strain. Previously, a strain of E. coli carrying the Thr931-Ile mutation in the G-G' section of the rpoC gene that encodes for the β' subunit of RNA polymerase (RNAP) was shown to be resistant to wild-type MccJ25. In addition, previous work demonstrated that incorporation of mutations at up to three positions in the peptide can produce active MccJ25 variants, some even more active than wild-type. In this work, a MccJ25-resistant strain of E. coli was generated by incorporating the mutation for resistance into the bacterial genome using genetic recombination via the Wanner method. Then, libraries of MccJ25 variants were screened against the resistant strain to select for mutants capable of overcoming the bacterial resistance. Showing that MccJ25 can be evolved to overcome a resistant strain would prove that natural evolution that bacteria have evolved for survival can be countered with laboratory directed evolution

Engineering bispecific chimeric antigen receptors to improve the efficacy of adoptive T-cell therapy

The recent FDA approval of CD19 chimeric antigen receptor (CAR) adoptive T-cell therapy for B-cell leukemias serves to highlight CAR-T cell therapy as a promising treatment approach for refractory cancers. More recently, adoptive transfer of T cells expressing CARs targeting B-cell maturation antigen (BCMA) has had numerous successes in clinical trial with 80-100% of multiple myeloma patients responding to treatment. However, CAR-T-cell therapy still faces several limitations including tumor antigen escape, a circumstance where tumor cells downregulate their surface antigen to avoid detection by CAR-T cells, and T-cell inhibition by cytokines such as transforming growth factor (TGF)-β in the solid tumor environment. These factors can significantly limit the efficacy of CAR-T-cell therapy. To overcome antigen escape, we designed single-chain bispecific CARs (OR-gate CARs) capable of signaling in the presence of two antigens instead of one. Using rational design principles, we constructed and evaluated CD19-OR-CD20 CARs that are able to prevent tumor antigen escape by CD19– leukemia. We further demonstrate that unlike single-input CD19 CARs, CD19-OR-CD20 CARs also prevent the emergence of spontaneous CD19-downregulated tumors in vivo. In a second study, we describe the rapid design and characterization of BCMA-OR-CS1 CARs and demonstrate that BCMA-OR-CS1 CARs can be rationally engineered to prevent antigen escape by BCMA– as well as CS1– myeloma cells. Finally, we explore the utility of the TGF-β CAR, a receptor capable of rewiring inhibitory TGF-β signaling to an activating response, in improving CD20 CAR function in TGF-β-rich environments. We evaluated three different bispecific targeting strategies, OR-gate CAR, DualCAR (co-expressing two receptors in one cells), and CARpool (pooling two different CAR-T cells), and demonstrate that TGF-β CAR-T cells are able to shield neighboring CD20 CAR-T cells from the inhibitory effects of TGF-β

La transition de la fécondité en Côte d'Ivoire (comment la politique démographique l'a influencée ?)

La Côte d Ivoire est le pays d Afrique occidentale francophone le plus avancé dans la transition de la fécondité. C est de l explication de cette transition que se charge cette thèse qui décrit les niveaux et tendances de la fécondité au cours de ces 45 dernières années. De 1960 à 1981 la fécondité est restée constamment élevée avant de baisser rapidement dans les années 1990. Les pionnières sont nées après les indépendances. Elles ont commencé à réduire leur fécondité quand elles avaient déjà fait leur cinquième enfant. Actuellement certains groupes de femmes contrôlent leurs naissances dans les unions alors que d autres continuent de se comporter naturellement. L évolution de la fécondité a été influencée par les actions du gouvernement en matière de santé, de scolarisation, de migration, de nuptialité et de planification familiale, actions ayant progressivement engendré de nouvelles attitudes face à la procréation. L augmentation de l âge au mariage et la diffusion de la contraception qui a touché certaines couches sociales qui ne contrôlaient pas auparavant leurs naissances en sont les principales manifestations. Les jeunes filles ne veulent plus reproduire les comportements de leurs mères et sont les plus avancées dans la transition de la fécondité.Ivory Coast is the Western Africa country which most advanced in the fertility transition. It is explanation of this transition that this rechearch takes care which describes the levels and tendencies of fertility during 45 last years. From 1960 to 1981 fertility constantly remained high but dropping quickly in the years 1990. The pioneers were born after independences. They started to reduce their fertility when they had already made their fifth child. Currently some groups of women control their births in the unions when others continue to behave naturally. The evolution of fertility was influenced by the government actions of the health, schooling, migration, nuptiality and family planning, actions having gradually generated new attitudes about procreation. Increase in the age to the marriage and diffusion of the contraception which touched some social layers which did not control their births before are the principal demonstrations. The girls do not want to reproduce any more the behaviors of their mothers and are advanced in the fertility transition.NANTERRE-BU PARIS10 (920502102) / SudocSudocFranceF

T Cells Expressing CD19/CD20 Bispecific Chimeric Antigen Receptors Prevent Antigen Escape by Malignant B Cells.

The adoptive transfer of T cells expressing anti-CD19 chimeric antigen receptors (CARs) has shown remarkable curative potential against advanced B-cell malignancies, but multiple trials have also reported patient relapses due to the emergence of CD19-negative leukemic cells. Here, we report the design and optimization of single-chain, bispecific CARs that trigger robust cytotoxicity against target cells expressing either CD19 or CD20, two clinically validated targets for B-cell malignancies. We determined the structural parameters required for efficient dual-antigen recognition, and we demonstrate that optimized bispecific CARs can control both wild-type B-cell lymphoma and CD19(-) mutants with equal efficiency in vivo To our knowledge, this is the first bispecific CAR capable of preventing antigen escape by performing true OR-gate signal computation on a clinically relevant pair of tumor-associated antigens. The CD19-OR-CD20 CAR is fully compatible with existing T-cell manufacturing procedures and implementable by current clinical protocols. These results present an effective solution to the challenge of antigen escape in CD19 CAR T-cell therapy, and they highlight the utility of structure-based rational design in the development of receptors with higher-level complexity. Cancer Immunol Res; 4(6); 498-508. ©2016 AAC

T Cells Expressing CD19/CD20 Bispecific Chimeric Antigen Receptors Prevent Antigen Escape by Malignant B Cells

The adoptive transfer of T cells expressing anti-CD19 chimeric antigen receptors (CARs) has shown remarkable curative potential against advanced B-cell malignancies, but multiple trials have also reported patient relapses due to the emergence of CD19-negative leukemic cells. Here, we report the design and optimization of single-chain, bi-specific CARs that trigger robust cytotoxicity against target cells expressing either CD19 or CD20, two clinically validated targets for B-cell malignancies. We determined the structural parameters required for efficient dual-antigen recognition, and we demonstrate that optimized bi-specific CARs can control both wild-type B-cell lymphoma and CD19(−) mutants with equal efficiency in vivo. To our knowledge, this is the first bi-specific CAR capable of preventing antigen escape by performing true OR-gate signal computation on a clinically relevant pair of tumor-associated antigens. The CD19-OR-CD20 CAR is fully compatible with existing T-cell manufacturing procedures and implementable by current clinical protocols. These results present an effective solution to the challenge of antigen escape in CD19 CAR T-cell therapy, and they highlight the utility of structure-based rational design in the development of receptors with higher-level complexity

Systematically optimized BCMA/CS1 bispecific CAR-T cells robustly control heterogeneous multiple myeloma.

Chimeric antigen receptor (CAR)-T cell therapy has shown remarkable clinical efficacy against B-cell malignancies, yet marked vulnerability to antigen escape and tumor relapse exists. Here we report the rational design and optimization of bispecific CAR-T cells with robust activity against heterogeneous multiple myeloma (MM) that is resistant to conventional CAR-T cell therapy targeting B-cell maturation antigen (BCMA). We demonstrate that BCMA/CS1 bispecific CAR-T cells exhibit superior CAR expression and function compared to T cells that co-express individual BCMA and CS1 CARs. Combination therapy with anti-PD-1 antibody further accelerates the rate of initial tumor clearance in vivo, while CAR-T cell treatment alone achieves durable tumor-free survival even upon tumor re-challenge. Taken together, the BCMA/CS1 bispecific CAR presents a promising treatment approach to prevent antigen escape in CAR-T cell therapy against MM, and the vertically integrated optimization process can be used to develop robust cell-based therapy against novel disease targets