Abstract 4701: Verteporfin inhibits surface PD-L1 expression in triple-negative breast cancer (TNBC) cells

Abstract

Abstract Background: Programmed death ligand 1 (PDL1) is commonly expressed on the surface of many tumor cells, including breast cancer. The activity of tumor-infiltrating lymphocytes (TIL) is inhibited by PDL1. High PD-L1 basal cells (particularly basal B) overexpress genes involved in invasion, motility, and chemoresistance (Soliman et al., 2014). Targeting and blocking PD-L1 may enhance eradication of aggressive breast cancer cells by the immune system. Verteporfin (VP), a photosensitizer used to treat macular degeneration, was found to inhibit PD-L1 expression in a high-throughput screen. Studies demonstrated VP inhibits YAP activation by disrupting YAP-TEAD interactions and preventing YAP induced oncogenic growth (Zhang et al., 2016). Here we demonstrate the inhibitory effect of VP on PD-L1 expression in TNBC cell lines. Methods: MDA-MB-231 cells co-cultured with human PBMCs were treated with VP (Sigma) in presence of concanavalin A (ConA) and analyzed using flow cytometry to study levels of CD8+IFNg+ cells. TNBC cell lines (MDA-MB-231, BT-20, HCC-1143, Hs-578T) were treated with VP at doses ranging from 1µM-10µM for 24 hrs. The cells were processed for flow cytometry, Western blot and RT-PCR to check PD-L1 in mechanistic studies looking at manipulation of YAP pathway genes. Results: When MDA-MB-231 cells were co-cultured with normal human PBMCs in the presence of ConA, the CD8+IFNg+ stained cells were reduced compared to PBMC + ConA alone. Interestingly, in the group treated with VP, rescue of CD8+IFNg+ cells was observed. Moreover, MDA-MB-231, BT-20, HCC-1143 and Hs-578T TNBC cells treated with VP showed a significant dose-dependent inhibition of PD-L1 expression by flow cytometry. Western blot analysis also showed complete clearance of PD-L1 protein band with the lowest dose (1µM) used. However, RT-PCR analysis did not show a significant fold change in mRNA levels of PD-L1 in MDA-MB-231 treated cells. Surprisingly, mechanistic studies performed by silencing YAP1, E2F1, and TBK1 in MDA-MB-231, BT-20 and HS-578 T showed a decline in PD-L1 in E2F1 silenced cells, highlighting a plausible role of E2F1-PDL1 signaling axis. However, no change in PD-L1 expression was seen in cells silenced with YAP1 and TBK1. Further, chromatin-immunoprecipitation assay demonstrated E2F1 binding to PD-L1 promoter. Conclusion: Our data so far demonstrate that verteporfin treatment leads to inhibition of PD-L1 in TNBC cell lines and improvement in CD8+IFNg+ cells, indicating that VP might have potential for treatment approaches. Our study warrants further attention towards understanding the mechanism of action of VP in inhibiting PD-L1 and the role of E2F1 in the process. Citation Format: Fatema Khambati, Neha Jaiswal, Srikumar Chellappan, Hatem Soliman. Verteporfin inhibits surface PD-L1 expression in triple-negative breast cancer (TNBC) cells [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 4701.