Abstract 2374: Squamous cell carcinoma antigen regulates macrophage polarization, contributing to inhibition of T cell activity in cervical cancer chemo-radiotherapy
Authors: Chen Liyun , Shi Victoria , Inkman Matthew , Zhang Jin , Cosper Pippa , Schwarz Julie K. , Markovina Stephanie 
Published: April 2023

Abstract
Chemoradiation therapy (CRT) is the standard treatment for locally advanced cervical cancer. Anti-tumor T cell response is critical for immunogenic cell death and tumor regression enabled by CRT. Squamous cell carcinoma antigen (SCCA) is highly expressed in cervical tumors and passively released from tumor cells. Patients with elevated serum SCCA often showed radioresistance and recurrence. Despite its prognostic value, little is known about the function of extracellular SCCA. We analyzed SCCA-associated immune landscape using RNAseq from 20 cervical matched tumor biopsy pairs collected prior to (pre-) and 3 weeks into (mid-) CRT. Patients with low pretreatment SCCA (&lt;6 ng/ml, SCCA/L) and high SCCA (≥6ng/ml, SCCA/H) showed decreased CD4/CD8 T cells at mid-CRT, with a more significant decrease in SCCA/H patients. This corresponded to significantly reduced expression of T-cell trafficking chemokine CXCL9/10 during CRT, particularly in SCCA/H patients. Interestingly, granzyme B (GZMB) and interferon-γ (IFNγ), associated with T-cell cytotoxicity, were increased in 6 of the 10 SCCA/L patients, compared to 1 and 2 of the 10 SCCA/H patients with increased GZMB and IFNγ respectively. Another RNAseq cohort of 66 pre-CRT tumor biopsies also showed significantly higher LAG3 and CTLA4 expression in SCCA/H than SCCA/L patients. In vitro stimulation of human PBMC with CD3/28 antibody in the presence of SCCA1 protein resulted in inhibition of T cell proliferation and decreased cytotoxicity. However, when SCCA1 was incubated with purified CD3 T cells, no inhibition in T cell activity was observed, suggesting the involvement of myeloid cells in SCCA1 mediated T cell activity. To investigate the effect of SCCA1 on myeloid cells, we isolated human CD14+ monocytes, incubated with SCCA1, and found improved survival of CD14+ HLADRlo/neg monocytes. THP1 monocytes, possessed polarizing ability, were differentiated into M1/M2 macrophage with the supplement of SCCA1 during the polarization. The result showed that SCCA1 promoted M2 macrophage phenotypes, including increased M2 marker (CD209), PD-L1 expression, M2-associated cytokines (IL10, CCL18), and M2 polarizing signaling (STAT3 phosphorylation and Myc expression). Macrophages isolated from murine tumors with high SCCA1 expression demonstrated increased suppression on T cell activity in co-culture functional assays. Furthermore, tissue microarray from cervical cancer patients showed that high SCCA was associated with increased infiltration of CD11b+ myeloid cells and CD68+ macrophages in tumors. Overall, we concluded that SCCA promoted M2 polarization and macrophage infiltration, which may contribute to T cell exhaustion, leading to poor survival. Thus, therapeutic targeting of extracellular SCCA should be considered to improve treatment outcomes.
Citation Format: Liyun Chen, Victoria Shi, Matthew Inkman, Jin Zhang, Pippa Cosper, Julie K. Schwarz, Stephanie Markovina. Squamous cell carcinoma antigen regulates macrophage polarization, contributing to inhibition of T cell activity in cervical cancer chemo-radiotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2374.