36: Mdsc-derived TNF Is a Novel Regulator of T-cell Dysfunction in Pancreatic Cancer

Abundance of myeloid-derived suppressor cells (MDSC) and a dysfunctional T-cell compartment are defining hallmarks of therapeutic resistance in pancreatic ductal adenocarcinoma (PDAC). Using congenic in vivo murine models to phenocopy extremes of T-cell enrichment or exclusion, we sought to interrogate central MDSC-mediated mechanisms that govern immune tolerance in PDAC.

 


Methods:

Orthotopically implanted T-cell-excluded (Tcell^lo) vs T-cell-enriched (T-cell^hi) congenic KPC tumors, and intratumoral Ly6G⁺F4/80^- MDSCs from both clones, were subjected to RNA sequencing. Ex vivo co-cultures evaluated effects of intratumoral MDSC on splenic T-cells. Orthotopically injected KPC-T-cell^lo mice were treated with etanercept vs. vehicle, and immunophenotyping via flow cytometry was performed.


Results:

RNA-seq of KPC T-cell^lo vs. T-cell^hi tumors revealed enrichment of myeloid immunoregulatory pathways, and downregulation of leukocyte activation/cytotoxicity pathways. Flow cytometry revealed dramatic increase in MDSCs infiltrating KPC-Tcell^lo tumors (P< 0.001). To decipher MDSC-intrinsic mechanisms associated with T-cell exclusion, RNA-seq of MDSCs infiltrating T-cell^hi clones revealed relative downregulation of MAPK signaling, and cytokine profiling of MDSCs conditioned with MAPK inhibitor trametinib revealed marked reduction in TNF secretion. Confocal microscopy confirmed striking decrease in TNF in MDSCs isolated from KPC- T-cell^hi vs. Tcell^lo tumors. Ex vivo MDSC-T-cell co-cultures significantly attenuated T-cell proliferation and activation (via IFN-γ release) while favoring T-cell apoptosis, which could be rescued by pre-conditioning MDSCs with either etanercept (TNFR2 decoy receptor) or MAPK pathway inhibitors. Orthotopically injected KPC-T-cell^lo tumor-bearing mice treated with etanercept demonstrated a remodeled TME vs. vehicle-treated mice, with attenuation in MDSC trafficking, enrichment in CD4⁺/CD8⁺ T-cell infiltration, and reduction in T-cell exhaustion.


Conclusions: MDSC-derived TNF regulates T-cell dysfunction in PDAC via a MAPK-dependent mechanism. Compartment-specific inhibition of TNF may be a provocative strategy to overcome immune tolerance in PDAC.