Enhancing Anti-Tumour Cytotoxic Lymphocyte Responses by Targeting FES in Antigen Presenting Cells

Abstract

The non-receptor tyrosine kinase FES has been implicated in the suppression of inflammatory signaling pathways in antigen-presenting cells (APCs) proposing that it might be dampening APC production of inflammatory cytokines in the tumour niche. These cytokines constitute the third and final signal of cytotoxic lymphocyte (CTL) activation – one of the main anti-tumour immune cells. This thesis explores the effect of FES disruption in APCs, specifically, its effect on inflammatory signaling, Signal 3 cytokine production and CTL priming. Immunoblotting analysis of the inflammatory signaling cascade showed that lipopolysaccharide (LPS)-stimulated fes-/- bone marrow-derived macrophages (BMDMs) display enhanced induction of inflammatory signaling cascades, compared to fes+/+ BMDMs. Additionally, RNA sequencing analysis revealed that in response to LPS-stimulation, certain key Signal 3 cytokine mRNA transcripts show a trend of higher production by fes-/- BMDMs. While a multiplexed cytokine detection assay did not reveal differences between fes+/+ and fes-/- BMDM secretion of these cytokines into culture media, flow cytometry analysis revealed a significantly higher frequency of fes-/- BMDMs with detectable IL-12 levels at the cell surface. This provides a possible rationale for fes-/- BMDMs being more effective at presenting antigen to and activating CTLs; and this was experimentally observed using co-cultures of OT-I CTLs with BMDMs presenting chicken ovalbumin (OVA). Collectively, these observations suggest that the apparent immunosuppressive function of FES makes it an actionable target for improving the efficacy of immunotherapies aimed at enhancing anti-cancer adaptive immune responses, including immune checkpoint inhibitors.