Squamous cell carcinoma antigen counteracts the radiation-induced antitumor response by driving an immune evasion phenotype through STAT signaling

Radiation therapy (RT)-induced immune response shapes the tumor microenvironment, and is recognized as an essential mechanism for anti-tumor effect. RT promotes the release of neoantigens, damage-associated molecular patterns and cytokines leading to cytotoxic T cell infiltration and enhancing immune surveillance. However, RT-induced anti-tumor response is sometimes hampered by inhibitory signals that suppress T cell function and facilitate tumor immune evasion. Squamous cell carcinoma antigen (SERPINB3/SCCA1) is highly expressed in epithelial cancers including cervix, head and neck, lung, and esophagus, and often associated with poor prognosis. We found that overexpressing mSerpinb3a in Lewis lung carcinoma cells (LL2/mB3a) promoted tumor growth compared to LL2 transduced with control vector (LL2/C). Prior to RT, increased MDSCs, TAMs, and Treg cells were observed in LL2/mB3a tumors compared to LL2/C, and is associated with a shorter RT-induced tumor growth delay in LL2/mB3a. The total number of infiltrating T cells was significantly reduced in LL2/mB3a and LL2/C at 2 days post-RT, with recovery by 5 days post-RT. However, CD8+ T remained low in the LL2/mB3a tumors and showed decreased proliferative capacity compared to LL2/C infiltrating T cells. Importantly, while RT improved the functionality in LL2/C-infiltrating T cells, as shown by ex vivo stimulation resulting in increased TNF and INFγ production compared to non-RT LL2/C-infiltrating T cells, the same improvement was not observed in LL2/mB3a-infiltrating T cells. In human T cell in vitro assay, SERPINB3 inhibited T cell activation by suppressing CD69, and CD25 expression, resulting in impaired proliferation capability. Further study on the association between SERPINB3 and immunosuppression showed that LL2/mB3a had increased suppressive chemokine CXCL1 and S100A8/A9 expression compared to LL2/C tumors and RT further enhanced expression of these chemokines in LL2/mB3a tumors. Likewise, human cancer cells overexpressing SERPINB3 also secreted high levels of CXCL1/8 and S100A8/A9. This chemokine induction was found regulated by elevated phosphorylation of STAT1/3 in SERPINB3-expressing cancer cells, while inhibiting STAT signaling by Ruxolitinib significantly suppressed the SERPINB3-dependent chemokine expression. We found that cervical cancer patients with high pre-treatment serum SCCA level (>16.1 ng/ml) and p-STAT3 level is associated with worse outcome with cancer specific survival at 2 years of about 10% compared to patients with serum SCCA <16.1 and low pSTAT3 of 95%. Our findings indicate that SERPINB3 high tumors produce high levels of suppressive chemokines through STAT signaling, leading to MDSCs, macrophages, and Treg infiltration, which dampen T cell functionality and mitigate the effect of radiation therapy. Citation Format: Liyun Chen, Victoria Shi, Songyan Wang, Rebecca Freeman, Jin Zhang, Pippa Cosper, Catherine Spina, Julie Schwarz, Stephanie Markovina. Squamous cell carcinoma antigen counteracts the radiation-induced antitumor response by driving an immune evasion phenotype through STAT signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 212.