Project 1: Molecular T-cell Therapy in Allogeneic Hematopoietic Stem-cell Transplantation
Immunotherapy based on the adoptive transfer of ex vivo expanded and activated autologous tumor-reactive lymphocytes can mediate tumor regression in cancer patients. However, high-avidity T cells specific for tumor-associated antigens (TAAs) are usually absent in patients because of self-tolerance. Using HLA-A2.1 transgenic mice, we have demonstrated the feasibility of T-cell receptor (TCR) gene transfer into T cells to circumvent self-tolerance to the widely expressed human p53 and MDM2 TAAs. A safety concern of TCR gene transfer is the risk of pairing between introduced and the naturally expressed endogenous TCR chains, resulting in the generation of self-reactive T cells. To prevent mixed TCR chain dimer formation, we engineered and optimized TAA-specific single chain TCRs. We successfully applied this strategy to the human TAAs p53, gp100 and MDM2-specific TCRs as promising antigens-driven immunotherapy for both melanoma and hematologic malignancies. Here, we evaluated the safety issues raised by the risk of TAA-TCR gene transfer-associated on/off-target toxicities and the therapeutic potential in relevant transgenic mouse models of adoptive transfer.
Project 2: Chimeric antigen receptors (CAR)-engineered T cells have demonstrated remarkable efficacy in patients with B-cell malignancies.
Yet, serious adverse events (e.g. cytokine release syndrome or fatal outcome) were frequently observed. NK cells can enhance T-cell activation without production of cytokines linked to adverse reactions. This joint (Frankfurt/Mainz, Dresden, Essen, Heidelberg) translational immunotherapy project aims to combine NK cells as effector cells with universal epitope-specific CAR (UniCAR) with tunable activity. The UniCAR is only activated upon cross-linking through a soluble tumor-targeting module carrying the UniCAR binding epitope, thereby achieving highest tumor specificity. Previous work from the contributing DKTK partner sites has demonstrated the versatility of this approach in T cells, and showed that it is feasible to derive CAR-engineered, established human NK cells as an off-the-shelf cellular therapeutic. Focusing on advanced melanoma, we will investigate the antitumor activity of UniCAR NK cells in clinically relevant models. Recombinant targeting modules will be based on antibodies and soluble single-chain fragments of TCR specific for melanoma antigens. Promising combinations of UniCAR NK cells and distinct targeting molecules will be identified for subsequent translation into a phase clinical study.
[Funding 2017-2019: Deutsches Konsortium für Translationale Krebsforschung (DKTK)]
Project 3: Impact of mutational pathways on immune environment, neoantigens and tumor rejection for the development of comprehensive biomarker panels in cancer immunotherapy
Cancer immunotherapy has demonstrated remarkable efficacy by checkpoint modulation in a large variety of neoplasms. Therapeutic response correlates with the mutational load of the patient tumors, suggesting that neoantigens contribute to tumor rejection. However, a relevant proportion of patients with high mutational load do not benefit while some patients with low mutational burden do. This underscores the need for an improved understanding of underlying factors limiting the efficacy of immune checkpoint therapy. To address this need, we will link an ImmuNEO module to the DKTK MASTER program that will include exome and transcriptome-sequenced patients on a pan-cancer basis who receive treatment with checkpoint modulators. Tumors from these patients will be used for peptide ligandome analyses, predictive in-situ biomarker analysis by immunohistochemistry, MALDI-TOF imaging mass spectrometry and RNA sequencing (RNAseq). T cells and TCR will be obtained from blood and tumor tissue and tested for neoantigen specificity and tumor reactivity. This multi-center (Müchen, Heidelberg, Frankfurt/Mainz, Essen, Freiburg, Dresden, Berlin) project will provide a better understanding of the interplay between mutational signatures and the relevant components of the immune system aiming at biomarker discovery and the design of combinatorial treatment strategies based on the identified mechanisms of action.
[Funding 2017-2019: Deutsches Konsortium für Translationale Krebsforschung (DKTK)]
Project 4: Molecular T cell immunotherapy and inhibition of tumor immune escape mechanisms
T cells can be retrovirally equipped with T cell receptors (TCRs) that are able to recognise tumor-associated antigens (TAA) with high affinity and thereby redirected to kill tumor cells in vitro and in vivo. A heterogenous population of immature or activated myeloid cells (myeloid-derived suppressor cells, MDSC) has emerged as one of the key effector populations of tumor immune escape, which potentially inhibits antitumoral effectiveness of TAA reactive T cells. This joint project (together with Dr. Markus Munder) aims to improve conventional T cell-based tumor immunotherapy by analysis and suppression of MDSC-mediated tumor immune escape mechanisms. For this purpose, we will use adoptive transfer of T cells redirected with HLA.A*02:01-restricted TCRs of defined specificities (p53(264-272), MDM2(81-88), XBP-1(18-27)) in both a syngeneic mouse tumor model and a NSG-A2 xenograft model. MDSC frequency in the circulation and in the tumor micromilieu as well as MDSC-associated tumor immune escape mechanisms (arginase and downstream products of arginine metabolism, reactive oxygen and nitrogen species) and suppressive properties will be analyzed in vitro and in vivo. In parallel, T cell expansion, phenotype and effector functions will be studied and correlated with the MDSC data. MDSC-mediated immunosuppressive pathways will then be specifically targeted by (a) pharmacological inhibitors for arginase (ABH, BEC, nor-NOHA), COX-2 (coxibs) or nitric oxide synthase (L-NMMA), (b) cytokine receptor blocking antibodies, (c) inhibition of the generation or expansion of MDSC (sunitinib, PDE5 inhibitor sildenafil) or (d) maturation / differentiation of MDSC into effective antigen presenting cells (all trans retinoic acid, ATRA). T cell proliferation and effector functions will be further boosted by modulating arginine availability via supplementation of citrulline, a non-toxic precursor amino acid for cellular arginine biosynthesis. In summary, our novel combined approach has the potential to synergistically enhance the efficacy of anti-tumor immunotherapy as a prerequisite for its successful translational implementation in clinical practice as powerful anti-cancer treatment.
(Project TP06 within the Collaborative Research Center SFB 1292; https://sfb1292.uni-mainz.de/)
[Funding 2018-2021: German Science Foundation (DFG)]
Project 5: Exploring a soluble T-cell receptor (TCR)-interleukin-15 superagonist complex as a novel, targeted immunostimulatory adjuvant to improve T/NK cell-mediated antitumor immunity to melanoma
This is a joint project together with AG Hartwig. It aims to explore the therapeutic potential of a novel targeted IL-15 superagonist composed of a soluble TCR specific for a cell surface melanocytic differentiation antigen, fused to an IL15/huIgG1-Fc (ILR-Fc) complex.
Therapeutic efficacy of adoptive cellular therapy in solid cancer such as e.g. melanoma is often hampered by nondurable antitumor immunity associated with insufficient trafficking, homing and persistence of T cells in an immunosuppressive tumor microenvironment. Interleukin (IL)-15 plays a pivotal role not only in T-cell but also NK-cell mediated immunity and might therefore represent a very promising candidate for improving antitumor responses under these conditions. In the proposed study we will thus explore a novel targeted IL-15 superagonist composed of a soluble HLA-independent single-chain (sc) TCR specific for a cell surface melanocytic differentiation antigen, fused to an IL15/IL15Rα/huIgG1-Fc (ILR-Fc) complex to confer high and prolonged bioactivity. The therapeutic potential of recombinant scTCR-ILR-Fc fusion protein to improve anti- melanoma responses will be examined in vitro and in vivo using clinically relevant mouse models after adoptive transfer of non-directed or antigen-TCR modified murine and human T cells or NK cells and combinations thereof. We propose that upon binding of the scTCR to the melanoma-associated cell surface antigen, signaling through the IL15/IL15Rα will substantially promote T-cell and NK-cell activation and induce cytolytic activity as recently reported for a HLA-A2-restricted p53-specific TCR linked to IL-2. Finally, targeting IL-15 might contribute to reduced nonspecific off-target activity often observed with cytokines used to boost immunity.
[Funding 2017: Inneruniversitäre Forschungsförderung (IUFF)]