Periodic Reporting for period 1 - NeoIDC (Neoantigen Identification with Dendritic Cell Reprogramming)
Periodo di rendicontazione: 2023-04-01 al 2024-09-30
In cancer, genetic mutations result in production of tumor neoantigens restrictively expressed in transformed cells and serve as ideal molecular targets for cancer vaccines and adoptive T cell therapies. However, available methods for the identification of clinically relevant neoantigens driving potent anti-tumor immunity are highly inefficient. Thus, there is an urgent need for innovative solutions to identify powerful immunogenic neoantigens to unlock the full potential of immunotherapy and treat currently untreatable cancers. The NeoIDC platform had the radical vision to use the type 1 conventional dendritic cell (cDC1) reprogramming technology, arising from the ERC-funded project TrojanDC, to directly identify highly immunogenic tumor neoantigens and neoantigen-specific T cell receptors (TCRs) for the development of powerful cancer vaccines and adoptive T cell therapies. To achieve this, we firstly aimed to identify tumor peptides presented on Major Histocompatibility Complex (MHC) by cDC1-reprogrammed cancer cells. Secondly, we aimed to expand neoantigen-specific T cells by co-culture with reprogrammed cancer cells and sequence expanded TCRs. We anticipated that this would facilitate the generation of potent cancer vaccines and tumor-reactive T cells for adoptive transfer. Experimental activities with collaborators and clinicians were coupled with exploitation plans including partnership with Asgard Therapeutics, ensuring commercialization and broad and targeted dissemination. NeoIDC combines cDC1’s antigen processing and presenting abilities with the unique mutational profile of cancer cells to enable the direct identification of immunogenic neoantigens and cognate TCRs. Ultimately, this project would set the stage for a new era of safe, personalized and effective cancer immunotherapies.
In this project, we developed two main activities that helped us get closer to the development of the next generation of neoantigen-based vaccines and adoptive T cell therapies.
Firstly, using cDC1 reprogramming on mouse cancer cell lines followed by immunopeptidomics, we profiled neoantigens present on MHC by liquid chromatography and tandem mass spectrometry (LC-MS/MS) analysis. For the data-dependent acquisition (DDA) of peptides, a library of MHC-I epitopes was generated, using bulk parental cells (100 million syngeneic B16 melanoma cells). In addition, we utilized the data independent acquisition (DIA) platform for peptide identification independently of a peptide library. Whole exome sequencing of B16 cells was also performed to filter somatic mutations out and focus on newly acquired mutations that could result in neoantigens.We identified 8 MHC class I neoantigens in the library that served as a reference. Interestingly, we identified 2 of the 8 neoantigens found in the library with reprogrammed cancer cells, while in B16 cells treated with IFN-γ (used as benchmark), we only detected 1 neoantigen at lower levels. Notably, one of the two identified neoantigens by reprogramming has never been characterized before and may represent a potent vaccine candidate. Quantification of the levels of the identified neoantigens against the number of peptides presented on the surface of the cancer cells showed an increase expression on the surface of reprogrammed cells, which indicates a preferential presentation of immunogenic antigens elicited by the upregulation of the professional antigen processing and presentation machinery. Overall, our data showed that reprogrammed mouse melanoma cells present tumor antigenic peptides at high levels leading to the discovery of new peptides not detected with standard approaches.
Our second activity and major achievement focused on T cells expansion. In the human, we co-cultured TILs from melanoma patients with reprogrammed cancer cells to expand for tumor-reactive T cells ex vivo. In collaboration with Inge Marie Svane’s lab and Ozcan Met (CCIT, Copenhagen, Denmark), we showed TIL reactivity and increased cytotoxicity upon monolayer co-culture with reprogrammed patient-derived melanoma cells. In vivo, we profiled T cells from tumors, tumor-draining lymph nodes and peripheral blood by single cell RNA-sequencing with T cell receptor enrichment using the syngeneic mouse melanoma model YUMM1.7.
In tumors containing reprogrammed melanoma cells, upon implantation of a mixture of reprogrammed melanoma cells and parental cells, we found increased frequencies of effector and effector memory CD8+ T cells, as well as a large cluster of type 1 T helper (Th1) and cytotoxic CD4+ T cells, with an interesting reduction in exhausted and regulatory T cell populations. Interestingly, we also found a reduction in exhausted and regulatory T cell populations.
These results were recently published in Science magazine (Ascic et al. Science 2024) and support the use of dendritic cell reprogramming for expansion of improved tumor-reactive T cell products for adoptive T cell treatments
Firstly, using cDC1 reprogramming on mouse cancer cell lines followed by immunopeptidomics, we profiled neoantigens present on MHC by liquid chromatography and tandem mass spectrometry (LC-MS/MS) analysis. For the data-dependent acquisition (DDA) of peptides, a library of MHC-I epitopes was generated, using bulk parental cells (100 million syngeneic B16 melanoma cells). In addition, we utilized the data independent acquisition (DIA) platform for peptide identification independently of a peptide library. Whole exome sequencing of B16 cells was also performed to filter somatic mutations out and focus on newly acquired mutations that could result in neoantigens.We identified 8 MHC class I neoantigens in the library that served as a reference. Interestingly, we identified 2 of the 8 neoantigens found in the library with reprogrammed cancer cells, while in B16 cells treated with IFN-γ (used as benchmark), we only detected 1 neoantigen at lower levels. Notably, one of the two identified neoantigens by reprogramming has never been characterized before and may represent a potent vaccine candidate. Quantification of the levels of the identified neoantigens against the number of peptides presented on the surface of the cancer cells showed an increase expression on the surface of reprogrammed cells, which indicates a preferential presentation of immunogenic antigens elicited by the upregulation of the professional antigen processing and presentation machinery. Overall, our data showed that reprogrammed mouse melanoma cells present tumor antigenic peptides at high levels leading to the discovery of new peptides not detected with standard approaches.
Our second activity and major achievement focused on T cells expansion. In the human, we co-cultured TILs from melanoma patients with reprogrammed cancer cells to expand for tumor-reactive T cells ex vivo. In collaboration with Inge Marie Svane’s lab and Ozcan Met (CCIT, Copenhagen, Denmark), we showed TIL reactivity and increased cytotoxicity upon monolayer co-culture with reprogrammed patient-derived melanoma cells. In vivo, we profiled T cells from tumors, tumor-draining lymph nodes and peripheral blood by single cell RNA-sequencing with T cell receptor enrichment using the syngeneic mouse melanoma model YUMM1.7.
In tumors containing reprogrammed melanoma cells, upon implantation of a mixture of reprogrammed melanoma cells and parental cells, we found increased frequencies of effector and effector memory CD8+ T cells, as well as a large cluster of type 1 T helper (Th1) and cytotoxic CD4+ T cells, with an interesting reduction in exhausted and regulatory T cell populations. Interestingly, we also found a reduction in exhausted and regulatory T cell populations.
These results were recently published in Science magazine (Ascic et al. Science 2024) and support the use of dendritic cell reprogramming for expansion of improved tumor-reactive T cell products for adoptive T cell treatments
The NeoIDC project pioneered the use of direct cell reprogramming to identify personalized neoantigen-derived peptides and TCRs for tailored immunotherapy. It allowed proof-of-concept and the identification of guarding neoantigens of reprogrammed cancer cells for potent vaccine design, as well as the expansion of T cells. NeoIDC went beyond current limitations and approaches to identify neoantigens, and provided proof-of-concept for expansion of tumor-reactive T cells for adoptive transfer. We will proceed with enhanced production of highly tumor-specific tumor-infiltrating lymphocytes (TILs), to offer a more effective immunotherapy for solid tumor patients. We will implement a simple step in step current state-of-the-art clinical protocols for expanding TILs leading to tumor reactive T cell expansion. We are currently seeking additional funding for implementation and expansion of patient-specific TILs in GMP conditions for personalized immunotherapy.