The tumor microenvironment reprograms FAM227A expression: Implications for CRC metastasis and diagnostic biomarker development.

[BACKGROUND] Colorectal cancer (CRC) is one of the most common malignant diseases which imposing a substantial burden on global health and economies. We previously identified a novel molecule, FAM227A, through functional screening using an shRNA retroviral library.

[METHODS] We used mouse immunization, hybridoma preparation, antibody screening and purification to get several mouse antibodies targeting human FAM227A. We use Western blot and RT-qPCR to detect the expression of FAM227A in colorectal cancer cells and tissues. Confocal imaging, nuclear-cytoplasmic fractionation and mitochondrial isolation were used to demonstrate the subcellular localization and expression of FAM227A. We performed tumorigenesis assays to investigate the tumor-promoting ability of FAM227A. Finally, we employed ELSIA to detect the expression of FAM227A in the serum of pan-cancer patients and get a ROC curve.

[RESULTS] In this study, we developed specific monoclonal antibodies (mAbs) against human FAM227A, identifying clone #8 with high affinity and specificity. Analysis of FAM227A expression in colorectal cancer (CRC) and different in vitro TME stress simulations. Subcellular localization studies indicated FAM227A primarily resides cytoplasmically but undergoes nuclear translocation and mitochondrial enrichment under specific conditions. Notably, FAM227A undergoes C-terminal degradation, producing a ∼30 kDa fragment (ΔFAM227A), potentially mediated by FBXW7 via a predicted recognition motif. Functionally, FAM227A overexpression modulated mitophagy markers and p53 phosphorylation. In p53-mutant CRC cells, FAM227A overexpression suppressed tumor growth in vivo, contrasting with variable effects in p53 wild-type cells, indicating p53 status-dependent tumor suppression. Finally, serum FAM227A levels were significantly elevated across multiple cancer types compared to healthy controls, suggesting its potential as a pan-cancer diagnostic biomarker.

[CONCLUSIONS] Collectively, our findings reveal FAM227A as a dynamically regulated protein influenced by the TME, involved in p53 pathway modulation, and a promising circulating biomarker for cancer detection.