The role of oncogenic KRAS in the immune evasion of non-small cell lung cancer

Lung cancer is a highly mutated tumour type in which, if effective, cancer immunotherapies have proven to be superior to other treatment modalities. However, tumours develop mechanisms by which they are able to evade the actions of the immune system, some of which may render them unresponsive to drugs aimed to boost antitumor immunity. KRAS is the most frequently mutated oncogene in cancer. Activating mutations in KRAS are found in up to a third of all lung cancer patients and have been previously shown to trigger immunosuppressive actions by tumour cells. Approximately a third of KRAS-mutant lung cancer patients harbour G12C mutations, which are amenable to therapeutic targeting thanks to the development of novel KRASG12C inhibitors, which are currently being tested in clinical trials. In this study, we aim to unveil new immunosuppressive mechanisms triggered by oncogenic KRAS signalling in vitro and in clinically relevant mouse models. Additionally, we try to elucidate the effects of novel KRASG12C inhibitors on the tumour microenvironment, with the ultimate goal of developing rational combination therapies. Using human cell lines, we have generated a gene expression dataset of KRAS-dependent genes. We have also developed a modified version of the murine lung cancer cell line 3LL to be able to perform experiments in an in vivo setting. Making use of these tools, we have established a link between KRAS signalling and the recruitment of immunosuppressive myeloid cells to the tumour microenvironment. Furthermore, the use of a therapeutic compound against KRASG12C has shown profound changes in the tumour microenvironment, further underscoring the fact that KRAS signalling can be immunosuppressive. This study highlights the possibility of therapeutically targeting components of the tumour microenvironment in KRAS mutant lung cancer, in combination with standard of care treatments or novel therapies such as KRASG12C inhibitors

Characterisation of tumour microenvironment remodelling following oncogene inhibition in preclinical studies with imaging mass cytometry

Mouse models are critical in pre-clinical studies of cancer therapy, allowing dissection of mechanisms through chemical and genetic manipulations that are not feasible in the clinical setting. In studies of the tumour microenvironment (TME), multiplexed imaging methods can provide a rich source of information. However, the application of such technologies in mouse tissues is still in its infancy. Here we present a workflow for studying the TME using imaging mass cytometry with a panel of 27 antibodies on frozen mouse tissues. We optimise and validate image segmentation strategies and automate the process in a Nextflow-based pipeline (imcyto) that is scalable and portable, allowing for parallelised segmentation of large multi-image datasets. With these methods we interrogate the remodelling of the TME induced by a KRAS G12C inhibitor in an immune competent mouse orthotopic lung cancer model, highlighting the infiltration and activation of antigen presenting cells and effector cells

CAR-T cell therapy targeting surface expression of TYRP1 to treat cutaneous and rare melanoma subtypes

A major limitation to developing chimeric antigen receptor (CAR)-T cell therapies for solid tumors is identifying surface proteins highly expressed in tumors but not in normal tissues. Here, we identify Tyrosinase Related Protein 1 (TYRP1) as a CAR-T cell therapy target to treat patients with cutaneous and rare melanoma subtypes unresponsive to immune checkpoint blockade. TYRP1 is primarily located intracellularly in the melanosomes, with a small fraction being trafficked to the cell surface via vesicular transport. We develop a highly sensitive CAR-T cell therapy that detects surface TYRP1 in tumor cells with high TYRP1 overexpression and presents antitumor activity in vitro and in vivo in murine and patient-derived cutaneous, acral and uveal melanoma models. Furthermore, no systemic or off-tumor severe toxicities are observed in an immunocompetent murine model. The efficacy and safety profile of the TYRP1 CAR-T cell therapy supports the ongoing preparation of a phase I clinical trial

CAR-T cell therapy targeting surface expression of TYRP1 to treat cutaneous and rare melanoma subtypes

Abstract A major limitation to developing chimeric antigen receptor (CAR)-T cell therapies for solid tumors is identifying surface proteins highly expressed in tumors but not in normal tissues. Here, we identify Tyrosinase Related Protein 1 (TYRP1) as a CAR-T cell therapy target to treat patients with cutaneous and rare melanoma subtypes unresponsive to immune checkpoint blockade. TYRP1 is primarily located intracellularly in the melanosomes, with a small fraction being trafficked to the cell surface via vesicular transport. We develop a highly sensitive CAR-T cell therapy that detects surface TYRP1 in tumor cells with high TYRP1 overexpression and presents antitumor activity in vitro and in vivo in murine and patient-derived cutaneous, acral and uveal melanoma models. Furthermore, no systemic or off-tumor severe toxicities are observed in an immunocompetent murine model. The efficacy and safety profile of the TYRP1 CAR-T cell therapy supports the ongoing preparation of a phase I clinical trial

An immunogenic model of KRAS-mutant lung cancer enables evaluation of targeted therapy and immunotherapy combinations

Mutations in oncogenes such as KRAS and EGFR cause a high proportion of lung cancers. Drugs targeting these proteins cause tumor regression but ultimately fail to elicit cures. As a result, there is an intense interest in how to best combine targeted therapies with other treatments, such as immunotherapies. However, preclinical systems for studying the interaction of lung tumors with the host immune system are inadequate, in part due to the low tumor mutational burden in genetically engineered mouse models. Here we set out to develop mouse models of mutant KRAS-driven lung cancer with an elevated tumor mutational burden by expressing the human DNA cytosine deaminase, APOBEC3B, to mimic the mutational signature seen in human lung cancer. This failed to substantially increase clonal tumor mutational burden and autochthonous tumors remained refractory to immunotherapy. However, establishing clonal cell lines from these tumors enabled the generation of an immunogenic syngeneic transplantation model of KRAS-mutant lung adenocarcinoma that was sensitive to immunotherapy. Unexpectedly, anti-tumor immune responses were not directed against neoantigens but instead targeted derepressed endogenous retroviral antigens. The ability of KRASG12C inhibitors to cause regression of KRASG12C-expressing tumors was markedly potentiated by the adaptive immune system, highlighting the importance of using immunocompetent models for evaluating targeted therapies. Overall, this model provides a unique opportunity for the study of combinations of targeted and immunotherapies in immune-hot lung cancer