Abstract 1747: Hyaluronan (HA) accumulation restricts CD8+ T cell numbers and skews tumor-associated macrophage (TAM) phenotype in mouse syngeneic pancreatic tumors
Authors: Thompson Benjamin , Kimbler Trevor , Lee Jisook , Zhao Chunmei , Chen Kelly , Clift Renee , Thompson Curtis B. , Rosengren Sanna , Maneval Daniel C. 
Published: August 2018

The glycosaminoglycan hyaluronan (HA) is abundant in the tumor microenvironment (TME) of diverse solid tumor types, and tumor accumulation of HA is associated with poor patient outcome. We previously demonstrated, in multiple HA-rich mouse syngeneic tumor models, that enzymatic degradation of HA using PEGylated recombinant human hyaluronidase PH20 (pegvorhyaluronidase alfa; PEGPH20) enhanced tumor growth inhibition and increased tumor accumulation of T and NK cells when combined with a checkpoint inhibitor. However, the ability of HA accumulation in the TME to alter the composition of tumor-infiltrating immune cells has not been well characterized. Here, we addressed this question by performing a detailed immune phenotypic analysis of HA-poor and HA-rich tumors using a mouse syngeneic pancreatic tumor model. To achieve this, we transduced Pan02 cells with a lentivirus encoding hyaluronan synthase 3 (HAS3, Pan02-HAS3) or empty vector (EV, Pan02-EV). After peritibial implantation, Pan02-HAS3 tumors contained 13.4-fold higher levels of HA than Pan02-EV tumors, as determined by HA ELISA. Both variants grew with similar kinetics to the Pan02 parental line. Flow cytometric analysis of tumor-infiltrating immune cells in four independent and identical studies revealed an average 6.2-fold reduction in the number of CD8+ T cells in Pan02-HAS3 tumors compared with Pan02-EV. Likewise, numbers of CD4+ T cells and NK cells were reduced by an average of 4.8- and 2.9-fold, respectively, in Pan02-HAS3 tumors compared with Pan02-EV. In contrast, we did not observe consistent differences in the numbers of tumor-associated macrophages (TAMs), neutrophils/granulocytic myeloid-derived suppressor cells (G-MDSC), monocytes/monocytic (M)-MDSC, or dendritic cells (DCs) between the two tumor variants. However, within the TAM population, cells displaying an “M2-like” phenotype (CD206high MHCIIlow) were present at an average 10.5-fold higher frequency in Pan02-HAS3 tumors compared with Pan02-EV. Similarly, the percentage of DCs that were CD103+ CD11b- was reduced by 1.4-fold in Pan02-HAS3 tumors compared with Pan02-EV. Together, our data indicate that HAS3-driven HA accumulation in Pan02 mouse syngeneic tumors induces several aspects of immunosuppression: it restricts the numbers of tumor-infiltrating lymphocytes, and skews TAMs and DCs toward immunosuppressive phenotypes. These findings are consistent with our previous work, which demonstrated that combining PEGPH20 with anti-PD-L1 increased tumor-infiltrating lymphocyte numbers and reduced the frequency of CD206high MHCIIlow TAMs in Pan02-HAS3 tumors. Our results support the ongoing clinical evaluation of PEGPH20 in combination with immunotherapy in HA-rich tumor indications (NCT02563548 and NCT03193190).

Citation Format: Benjamin Thompson, Trevor Kimbler, Jisook Lee, Chunmei Zhao, Kelly Chen, Renee Clift, Curtis B. Thompson, Sanna Rosengren, Daniel C. Maneval. Hyaluronan (HA) accumulation restricts CD8+ T cell numbers and skews tumor-associated macrophage (TAM) phenotype in mouse syngeneic pancreatic tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1747.