Tumor-specific cytolytic CD4 T cells mediate immunity against human cancer

CD4 T cells have been implicated in cancer immunity for their helper functions. Moreover, their direct cytotoxic potential has been shown in some patients with cancer. Here, by mining single-cell RNA-seq datasets, we identified CD4 T cell clusters displaying cytotoxic phenotypes in different human cancers, resembling CD8 T cell profiles. Using the peptide-MHCII-multimer technology, we confirmed ex vivo the presence of cytolytic tumor-specific CD4 T cells. We performed an integrated phenotypic and functional characterization of these cells, down to the single-cell level, through a high-throughput nanobiochip consisting of massive arrays of picowells and machine learning. We demonstrated a direct, contact-, and granzyme-dependent cytotoxic activity against tumors, with delayed kinetics compared to classical cytotoxic lymphocytes. Last, we found that this cytotoxic activity was in part dependent on SLAMF7. Agonistic engagement of SLAMF7 enhanced cytotoxicity of tumor-specific CD4 T cells, suggesting that targeting these cells might prove synergistic with other cancer immunotherapies.

Anti-angiogenic immunotherapy for lung cancer

Ece Kadioglu

Lung cancer is the leading cause of cancer-associated deaths worldwide. Platinum-based chemotherapy is the most common therapeutic approach in advanced non-small cell lung cancer (NSCLC), particularly for tumors that carry non-druggable activating mutations. However, these tumors are generally not eradicated when diagnosed at an advanced stage and are frequently resistant to chemotherapy. Therefore, there is an urgent medical need for new treatments for this frequent cancer type. Immunotherapies are treatments based on increasing the strength of immune responses against cancer. In particular, monoclonal antibodies that block inhibitory "immune-checkpoints" expressed on T cells (e.g., programmed cell death protein 1, or PD1) show clinical efficacy in increasing tumor types. However, these therapies improve survival in only a minority of NSCLC patients, and ongoing efforts are being made for increasing their efficacy through combinatorial treatments. In this regard, growing evidence indicates that tumor angiogenesis is closely associated with immunosuppression. Pro-angiogenic factors, such as vascular endothelial growth factor A (VEGFA) and angiopoietin 2 (ANG2), cause abnormalities and dysfunctions in tumor blood vessels that impair T cell extravasation and anti-tumoral activity. Recent findings in preclinical cancer models show that reprogramming the features of tumor-associated blood vessels by anti-angiogenic drugs may help to increase intratumoral T cell abundance and sensitize refractory tumors to immunotherapy. Nevertheless, the potential of anti-angiogenics in the context of lung cancer immunotherapy is poorly understood. Therefore, the main objective of my project was to audit combinations of anti-angiogenic therapy and immune checkpoint blockade in a genetically engineered mouse model of NSCLC. I first studied the anti-tumoral activity of dual VEGFA and ANG2 inhibition using a bispecific antibody (A2V) in the KrasLSL-G12D/+;p53fl/fl (KP) NSCLC model. I found that A2V, but not single VEGFA or ANG2 blockade, had robust tumor-inhibitory activity in KP mice, which was at least comparable to standard-of-care chemotherapy. However, A2V only moderately increased cytotoxic T cells in the tumors. To potentially increase the immunogenicity of KP tumors, I also generated two KP mouse variants by either co-expressing a surrogate neoantigen or genetically deleting a DNA mismatch repair enzyme in the cancer cells. However, the therapeutic efficacy of A2V was not further improved in spite of a slightly more activated immune response. I next investigated combination therapies involving A2V and PD1 blockade. In contrast to previous findings in other cancer types, the addition of PD1 antibodies deteriorated the efficacy of A2V, likely through enhancing immunosuppressive mechanisms sustained, at least in part, by regulatory T cells and T cell-antagonizing cytokines. In summary, my work supports the potential clinical efficacy of combined VEGFA and ANG2 blockade for lung cancer therapy but provided little evidence for its synergy in association with anti-PD1 therapy.

EPFL2019

Mechanical Immunoengineering of T Cells for Enhanced Cancer Immunotherapy

Kewen Lei

Cancer is one of the leading causes of death globally. Immunotherapy represents one of the most promising cancer treatments inducing complete and durable responses in a fraction of cancer patients. Despite the unprecedented clinical success achieved by T cell-based cancer immunotherapies, there are tremendous outstanding challenges, including low response rate and severe toxicities, which substantially limit the clinical benefits in a majority of cancer patients. At present, enormous efforts have been spent on modulating biochemical interactions for improving the efficacy and safety of T cell-based cancer immunotherapy. Besides biochemical exchanges, mechanical interactions are universal in every step of T cell immunity and are crucial in regulating T cell functions. Thus, immunoengineering approaches that modulate mechanical interactions in T cell immunity hold great promise to improve current cancer immunotherapy. This thesis aims to develop novel mechanical immunoengineering approaches for improving cancer immunotherapy. Specifically, it includes the following parts:Developing a spiky artificial antigen-presenting cell (aAPC) to enhance ex vivo T cell expansion. Ex vivo expansion of T cells is critical in adoptive T cell therapy (ACT) to ensure an effective therapeutic dosage for cancer treatment. By mimicking the dendritic morphology of dendritic cells, I developed a spiky TiO2 microparticle presenting T cell stimulatory ligands as an aAPC to enhance ex vivo T cell expansion. The spiky aAPC outperformed the smooth counterpart, as well as Dynabead®, a standard aAPC used in clinical manufacturing of ACT therapy, for T cell activation and expansion depending on F-actin polymerization in T cells. Therefore, this finding identifies the morphology of aAPCs as a critical parameter to improve ex vivo T cell manufacturing for ACT therapy.Boosting T cell-mediated cytotoxicity by stiffening cancer cells. I discovered that cancer cell softening derived from cholesterol enrichment in the plasma membrane led to resistance to T cell-mediated cytotoxicity. Stiffening cancer cells via membrane cholesterol depletion substantially enhanced T cell-mediated cytotoxicity against cancer cells through augmenting T cell mechanical force but not any known biochemical pathways of T cell-mediated cytotoxicity. Cancer cell stiffening intervention in vivo enhanced the efficacy of ACT therapy in multiple preclinical solid tumor models. Cancer cell softness thus represents a new immune checkpoint of mechanical basis, and therapeutically targeting this mechanical immune checkpoint has the potential to improve the clinical response of T cell-based cancer immunotherapy.Utilizing T cell force as a trigger to achieve specific release of anti-cancer drugs. Specific delivery of T cell supporting drugs and/or anti-cancer drugs into tumors has the potential to increase the response rate of ACT therapy without causing systemic toxicity. To this end, I developed a T cell force-responsive drug release system based on a mesoporous silica microparticle with DNA force sensors as gatekeepers. T cell force as a highly specific signal upon recognizing cancer cells presenting cognate antigens was shown to trigger specific drug release when T cell receptor signaling was activated. Further, the T cell force-responsive drug release system could deliver anti-cancer drugs in vitro and in vivo in a T cell force-dependent manner and holds promise to enhance cancer immunotherapy.

EPFL2022

New roles of the lymphatic endothelium in modulating adaptive immunity: implications for immune tolerance and cancer immunotherapy

Lambert Charles François Potin

Lymphatic vessels have traditionally been considered as passive conduits for interstitial fluid drainage and for immune cell trafficking to lymph nodes. In the last two decades, many studies have challenged this classical dogma: lymphatic endothelial cells (LECs) were shown to actively interact with immune cells by secreting chemokines and cytokines, and presenting antigens to T cells. These newly described functions pinpoint a multi-faceted role of LECs in orchestrating the immune response through both passive and active mechanisms. Yet, precise mechanisms leading to antigen-specific fine-tuning of the T cell response remain unexplored. Moreover, although lymphatics have been shown to play important roles in cancer, the contributions of LECs in regulating antitumor immunity have been largely overlooked by the biomedical community. This doctoral thesis aimed at filling these gaps. First, in order to investigate the mechanisms and implications of LEC antigen presentation to CD4+ T cells, we engineered a mouse model where LECs lacked the ability to present antigens to CD4+ T cells. In these mice, antigen-specific responses to vaccination were increased when compared to wild-type controls. Moreover, we demonstrated that LECs were poor inducers of CD4+ T cell activation in vitro, and that they could dampen CD4+ T cell activation by dendritic cells. Taken together, these findings suggest that LECs participate to the peripheral immune tolerance of CD4+ T cells. Second, we investigated the immunological consequences of the development of lymphatic vessels, called lymphangiogenesis, in mouse melanoma. Upon induction of tumor lymphangiogenesis, we observed striking tumor enrichment with immune cells, which resulted in a higher sensitivity of those tumors to immunotherapy. We subsequently demonstrated that inducing lymphangiogenesis in a mouse model of cold tumors, which lack immune infiltrates and do not respond to immunotherapies, could restore their responsiveness to immunotherapy treatments. Third, to predict which cancer types were the most likely to replicate the findings we obtained in mouse melanoma, we mined The Cancer Genome Atlas database for gene expression. We identified a set of cancers, such as colon adenocarcinoma and cholangiocarcinoma, where high expression of the lymphangiogenic growth factor was associated with a strong T cell signature. This doctoral thesis unveiled new immunomodulatory functions of LECs by which antigen-specific immune tolerance is induced. Additionally, it opened new avenues to target lymphatics therapeutically in cancer to potentiate immunotherapies. We believe this work contributes to demonstrating the immunological importance of LECs, and hope that lymphatics will be considered as an important immunomodulatory player of the tumor microenvironment when developing novel immunotherapies.

EPFL2018