Project overview
Background: UK incidence of head & neck cancer (HNSCC; comprising human papillomavirus (HPV)-ve and HPV+ve subgroups) is rising and has increased by 20% in the last decade (>12000 cases/year). HNSCC is difficult to treat. Surgery and radiotherapy are standard but associated with significant morbidity and a static 5-year survival rate (50-60%). Response to anti-PD1/PDL1 immunotherapy is ~15%, with most HNSCC tumours classed as 'immune-cold', containing few T-lymphocytes (TILlow) and a stroma rich in myofibroblastic cancer-associated fibroblasts (myoCAF). CAF have been a putative therapeutic target for many years. However, these cells are functionally heterogenous and remain poorly characterised, which has hampered the development of clinically effective targeting strategies. We have shown that myoCAF exclude CD8 T-cells from tumours and confer immunotherapy resistance. Strategies for overcoming myoCAF-mediated lymphocyte exclusion are required to improve immunotherapy response. Aims: To characterise CAF subgroups in HPV-ve HNSCC, HPV+ve HNSCC and normal mucosa, determine how these phenotypes differentially regulate the immune microenvironment and optimise manipulation of myoCAF regulatory pathways to improve immunotherapy response. Methods: We hypothesise that HNSCC contains functionally distinct CAF subgroups that differentially regulate the immune microenvironment and therefore effective CAF targeting requires an understanding of this functional heterogeneity. We will use single cell transcriptomics and multiplexed immunochemistry, integrated with bulk RNA sequencing to characterise CAF phenotypes, spatial distribution and immune cell interactions in HNSCC/healthy tissues and pre/post radiotherapy. Functional analyses will study how CAF subtypes shape the immune microenvironment, focusing on interactions with CD8 T-cells using established in vitro assays and in vivo models. Preliminary work identified myoCAF involvement in immunotherapy resistance. We propose three potential strategies for targeting myoCAF and expect these strategies to represent highly-effective immunotherapy adjuncts: (1) skewing tumour-promoting myoCAF to a tumour-suppressive phenotype, through inhibition of myoCAF-specific downstream mediators of TGF-β1 signalling; (2) depleting myoCAF; and (3) targeting the interaction between myoCAF and macrophages, to reshape the immune microenvironment. To test these strategies, we will compare the efficacy of NOX4/ATM/Wee1 inhibitors on myoCAF 'normalisation'; investigate the impact of myoCAF depletion by vaccination (targeting MageD4B, which we recently found expressed by myoCAF and tumour cells in HNSCC); and test whether anti-CD47 (promoting macrophage phagocytosis) can overcome myoCAF-mediated CD8+ T-cell exclusion. In each case, analysing immune cell phenotype/distribution and anti-tumour efficacy in vivo, and in combination with immunotherapy. Summary: If successful, this work will lead to clinical-testing myoCAF targeting as part of combination immunotherapy in HNSCC.
