Stromal heterogeneity in pancreatic cancer

Abstract

Fibroblasts are a mesenchymal cell type present in most mammalian tissues, that make a major contribution to ECM deposition and modification in health and disease. Pancreatic ductal adenocarcinoma (PDA) tumours are typically infiltrated with large numbers of fibroblasts but their functional contributions to tumour progression are poorly understood. Research into dermal fibroblasts has revealed at least two distinct fibroblast lineages in mammalian skin, that diverge during embryonic development and have different, non-overlapping functions in adult tissue homeostasis and wound-healing. Whether such distinct lineages of fibroblasts exist in other tissues or tumours is not known. A major barrier to understanding fibroblast heterogeneity in tumours is a lack of robust fibroblast surface markers, which has precluded the isolation of live subpopulations for functional characterisation. I sought to address this by developing a single cell mass cytometry analysis workflow to annotate the PDA tumour microenvironment (TME). The analysis reveals extensive fibroblast heterogeneity in PDA tumours and highlights potential associations between fibroblast abundance and proliferation rates of certain immune cell subsets. A notable feature of murine and human normal pancreatic and PDA fibroblasts is the presence of two major fibroblast populations, demarked by differential expression of CD105. Isolation of these populations reveals that differential CD105 expression is stable under extended passaging and under various activating conditions, suggesting CD105+ and CD105- pancreatic fibroblasts to be stable, distinct lineages. CD105+ pancreatic fibroblasts are more sensitive to TGFβ1 and Il1α stimulation in vitro but both populations have equal sensitivity to IFNγ stimulation. In a syngeneic subcutaneous co-transplant model, CD105+ pancreatic fibroblasts are tumour permissive, and do not alter PDA tumour growth, and CD105- pancreatic fibroblasts are found to be highly tumour suppressive. The mechanism for tumour suppression is entirely dependent on functional adaptive immunity, with a major contribution from cDC1-mediated CD8+ T cell cross-priming. Despite CD105- PDA fibroblasts consistently having a fraction of cells with evidence of expression of MHCII antigen presentation in vivo, fibroblast MHCII antigen presentation does not contribute to the enhancement of anti-tumour immunity in this model. In addition, CD105+ fibroblasts cannot be made to become tumour suppressive by deletion of CD105, suggesting that whilst CD105 is a useful marker for separating functionally distinct pancreatic fibroblast lineages, lack of CD105 does not functionally contribute to the dominant in vivo phenotype of CD105- pancreatic fibroblasts. CD105+ and CD105- fibroblasts are found to be present in at least 12 other mammalian organs and 2 other spontaneous tumour types. This data highlights CD105 as marker of distinct fibroblast lineages in a large number of mammalian tissues and also indicates that fibroblast and immune cell interactions can drive dominant phenotypes in vivo.

Date of Award	31 Dec 2020
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Georges Lacaud (Supervisor) & Claus Jorgensen (Supervisor)