BackgroundCKLF-like MARVEL transmembrane domain-containing 6 (CMTM6), a programmed death-ligand 1 (PD-L1) regulator, is widely expressed in various tumors and regulates the immune microenvironment. However, its prognostic value remains controversial, and the roles of CMTM6 in colorectal cancer (CRC) are still unknown. In this study, we aimed to elaborate the expression patterns of CMTM6 and PD-L1 in CRC and investigate their relationship with the infiltration of T cells and the prognosis of patients with CRC.MethodsAnalysis of CMTM6 mRNA levels, gene ontology enrichment analysis and single-sample gene set enrichment analysis were performed in a The Cancer Genome Atlas colon cancer cohort. The expression of CMTM6 and PD-L1 and the infiltration of T cells in tumor tissues from our cohort containing 156 patients with CRC receiving adjuvant chemotherapy and 77 patients with CRC without chemotherapy were examined by immunohistochemistry assay.ResultsCMTM6 expression was upregulated in CRC compared with normal colon tissues, and CMTM6 levels were lower in advanced tumors than in early-stage tumors. High expression of CMTM6 correlated with lower pT stage and more CD4+/CD8+ tumor-infiltrating lymphocytes (TILs) and predicted a favorable prognosis in CRC. PD-L1 was expressed in CRC tissues at a low level, and PD-L1 positivity in tumor stroma (PD-L1(TS)), but not PD-L1 positivity in cancer cells (PD-L1(CC)), was associated with an increased density of CD4+ TILs and a favorable prognosis. The coexpression status of CMTM6 and PD-L1(TS) divided patients with CRC into three groups with low, moderate and high risks of progression and death, and patients with CMTM6High/PD-L1(TS)+ status had the longest survival. Moreover, the prognostic value of CMTM6/PD-L1 expression was more significant in patients with CRC treated with adjuvant chemotherapy than in those not treated with chemotherapy.ConclusionCMTM6 has a critical impact on the immune microenvironment and can be used as an independent prognostic factor for CRC. The coexpression status of CMTM6 and PD-L1 can be used as a new classification to stratify the risk of progression and death for patients with CRC, especially for patients receiving adjuvant chemotherapy. These findings may provide insights into improving responses to immunotherapy-included comprehensive treatment for CRC in the future.

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<p>Supplementary Tables S5 & S6 show the candidate genes picked out in HCT116 and HT29 cells; Supplementary Tables S7 & S8 show the proteins might bind with RFC4, detected by IP-MS in HCT116 and HT29 cells.</p>

<p>Supplementary Table S1 shows the sequences of siRNAs; Supplementary Table S2 shows the informations of shRNAs; Supplementary Table S3 shows the sequences of quantitative PCR Primers.</p>

<p>Supplementary Figure S1 shows the procedure to generate radioresistant HCT1166GyR cells; Supplementary Figure S2 shows reporter constructs of NHEJ and HR pathway; Supplementary Figure S3 shows representative images of IHC assays; Supplementary Figure S4 show the detailed data of screening candidate radioresistant genes; Supplementary Figure S5 shows RFC4 expression in CRC cells and tissues after radiation; Supplementary Figure S6 shows the representative clonogenic pictures for Figure 1C; Supplementary Figure S7 shows representative charts of Annexin V/PI double staining assays for Figure 1F; Supplementary Figure S8 shows the effects of RFC4 on γ-H2AX and pATM levels after radiation; Supplementary Figure S9 shows the additional evidences of RFC4 to promote NHEJ- but not HR-mediate DNA repair; Supplementary Figure S10 shows the effects of RFC4 on the chemotherapeutic drug resistance in HCT116 cells.</p>