Abstract:

Objective To investigate the prognostic role of transmembrane protein 205（TMEM205） in patients with colorectal cancer（CRC）. Methods The expression level of CRC tissue TMEM205 was analyzed in the Cancer Genome Atlas（TCGA） database in relation to the prognosis. Serum samples from 20 CRC patients at Yangpu Hospital of Tongji University School of Medicine from September 2022 to October 2023 were collected，and the research subjects were classified into recurrence group（10 cases） and non-recurrence group（10 cases） according to 1-year prognosis. Totally，5 CRC cell lines（HCT116，SW480，Lovo，RKO，HT29） and normal intestinal epithelial cell line NCM460 were selected. 5-Fluorouracil（5-FU）-resistant cell lines（HCT116/5-FU，SW480/5-FU） and oxaliplatin（OXA）-resistant cell lines（HCT116/OXA，SW480/OXA） were established. The drug-resistant cell lines were classified into knockdown group（transfected with TMEM205-targeting siRNA） and negative control group（transfected with siNC） according to different transfected siRNA. CCK-8 assay，cell colony formation assay，western blotting and real-time fluorescent quantitative polymerase chain reaction（qRT-PCR） were performed to determine the cell proliferation activity，chemotherapeutic drug sensitivity and the expression levels of epithelial-mesenchymal phenotype and cancer stemness markers in cell lines，knockdown group and negative control group. Exosomes were extracted from the culture media of parental HCT116 cells and drug-resistant cell lines，as well as the serum of CRC patients，and the expression level of TMEM205 was determined. Results Based on the analysis of the TCGA database，the overall survival of patients in low TMEM205 expression group was better than that in high TMEM205 expression group [hazard ratio（HR）=1.72，95% confidence interval（CI）1.107-2.659，P=0.016 1]. In CRC tissues，the expression of TMEM205，TNM pathological stage and serum carcinoembryonic antigen（CEA） level were related to poor prognosis in CRC patients with the score span of TMEM205 expression being superior to that of serum CEA. The TMEM205 relative mRNA and protein expression levels in 5 CRC cell lines were higher than those in NCM460 cell lines（P<0.01）. The TMEM205 relative mRNA and protein expression levels in drug-resistant isolates were higher than those in parental cells. Compared with negative control group，the knockdown group showed increased sensitivity to 5-FU and OXA（P<0.001），decreased half-maximal inhibitory concentration （IC₅₀）（P<0.001），and reduced cell colony formation ability（P<0.001）. The expression levels of epithelial phenotype-related markers [E-cadherin（E-Cad），beta-catenin and gamma-catenin] were enhanced（P<0.01），while the expression levels of mesenchymal phenotype-related markers [Vimentin，alpha-smooth muscle actin（α-SMA） and N-cadherin] were reduced（P<0.01）. The expression levels of tumor stemness markers [CD44，octamer-binding transcription factor 4（OCT4），Nanog homebox protein（Nanog） and sex-determining region Y-box protein 2（SOX2）] were decreased（P<0.01）. Conclusions In colorectal cancer cells，high expression of TMEM205 can maintain the drug-resistant phenotype to 5-FU and OXA through epithelial-mesenchymal transition and tumor stemness. TMEM205 may serve as a high-risk prognostic factor for CRC. Serum exosomal TMEM205 shows potential as a biomarker for predicting CRC recurrence.

Key words: Transmembrane protein 205, Exosome, 5-Fluorouracil, Oxaliplatin, Colorectal cancer, Recurrence, Prognosis