FAK inhibition reduces tumor infiltration of Tregs via restricting CCL5 release and boosts the present therapeutic regimen for BRAF(V600E)-mutated colorectal carcinoma

Background: BRAFV600E-mutated colorectal carcinoma (CRC) is a refractory disease with poor prognosis and limited therapeutic approaches. Recently, researches have indicated that the recruitment of Tregs in tumor tissue promoted by BRAFV600E is responsible for the weakened antitumor immunity. Whereas, as the main targeting therapy for this intractable carcinoma, BRAFV600E inhibitor has little response. In addition, FAK, being the novel therapeutic target in CRC, is considered to have an impact on the release of chemokines. In this study, we sought to explore the capacity of FAK inhibitor restricting Treg infiltration in tumor and its mechanism, as well as the synergistic antitumor effect with the present therapeutic strategy for BRAFV600E-mutated CRC. Methods: Cell proliferation was determined by CCK-8 assay and colony formation assay. Migration ability was detected by transwell assay. Cell cycle and cell group analysis were assessed by flow cytometry. Western blot, Elisa and RT-qPCR analysis were conducted for mechanism exploration. Immunohistochemistry and immunofluorescence analysis were performed to confirm the link between BRAFV600E, FAK and Tregs. CRC xenograft models were established to detect the Treg recruitment and evaluate the synergistic antitumor effect in vivo. Results: BRAFV600E-mutated CRC was of higher expression of FAK, and hence more sensitive to FAK inhibitor APG-2449. Besides, BRAFV600E-mutated in CRC was linked with more Treg infiltration in tumor site, therefore contributed to the attenuated antitumor immunity. What’s more, despite directly inhibiting tumor proliferation and migration, one of the important mechanisms of FAK blockade restricting the growth of BRAFV600E-mutated CRC, was by downregulating the release of Treg chemokine CCL5 from tumor cells through FAK-IL33 axis. With the recruitment of Tregs decreasing induced by FAK inhibitor APG-2449, the number of CD8+T cells relatively elevated in tumor tissue. In BRAFV600E-mutated CRC xenotransplantation models, we revealed that the combination of FAK inhibitor APG-2449 and Vemurafenib was of no significant difference with the combination of Vemurafenib and cetuximab. While adding APG-2449 to the current combination strategy achieved greater tumor suppressing effect than not adding. Conclusions: BRAFV600E-mutated CRC is associated with a higher expression level of FAK, as well as the tumor infiltration of Tregs. FAK inhibitor APG-2449 can promote antitumor immunity by restricting Treg number in tumor tissue, and shows a synergistic effect with the present therapeutic strategy for BRAFV600E-mutated CRC treatment. Citation Format: Luping Yuan, Wentao Pan, Suna Zhou, Qiuyun Luo, Xianglei Yan, Yuxin Zhang, Mengxian Pan, Lin Zhang, Miaozhen Qiu, Dajun Yang. FAK inhibition reduces tumor infiltration of Tregs via restricting CCL5 release and boosts the present therapeutic regimen for BRAFV600E-mutated colorectal carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3958.