PREX2 contributes to radiation resistance by inhibiting radiotherapy-induced tumor immunogenicity via cGAS/STING/IFNs pathway in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) lacks established biomarkers or molecular targets for predicting or enhancing radiation response. Phosphatidylinositol-3,4,5-triphosphate-dependent Rac exchange factor 2 (PREX2) exhibits intricate implications in tumorigenesis and progression. Nevertheless, the precise role and underlying mechanisms of PREX2 in CRC radioresistance remain unclear. METHODS: RNA-seq was employed to identify differentially expressed genes between radioresistant CRC cell lines and their parental counterparts. PREX2 expression was scrutinized using Western blotting, real-time PCR, and immunohistochemistry. The radioresistant role of PREX2 was assessed through in vitro colony formation assay, apoptosis assay, comet assay, and in vivo xenograft tumor models. The mechanism of PREX2 was elucidated using RNA-seq and Western blotting. Finally, a PREX2 small-molecule inhibitor, designated PREX-in1, was utilized to enhance the efficacy of ionizing radiation (IR) therapy in CRC mouse models. RESULTS: PREX2 emerged as the most significantly upregulated gene in radioresistant CRC cells. It augmented the radioresistant capacity of CRC cells and demonstrated potential as a marker for predicting radioresistance efficacy. Mechanistically, PREX2 facilitated DNA repair by upregulating DNA-PKcs, suppressing radiation-induced immunogenic cell death, and impeding CD8+ T cell infiltration through the cGAS/STING/IFNs pathway. In vivo, the blockade of PREX2 heightened the efficacy of IR therapy. CONCLUSIONS: PREX2 assumes a pivotal role in CRC radiation resistance by inhibiting the cGAS/STING/IFNs pathway, presenting itself as a potential radioresistant biomarker and therapeutic target for effectively overcoming radioresistance in CRC.

Comprehensive characterization of tumor microenvironment in colorectal cancer via molecular analysis.

Colorectal cancer (CRC) remains a challenging and deadly disease with high tumor microenvironment (TME) heterogeneity. Using an integrative multi-omics analysis and artificial intelligence-enabled spatial analysis of whole-slide images, we performed a comprehensive characterization of TME in colorectal cancer (CCCRC). CRC samples were classified into four CCCRC subtypes with distinct TME features, namely, C1 as the proliferative subtype with low immunogenicity; C2 as the immunosuppressed subtype with the terminally exhausted immune characteristics; C3 as the immune-excluded subtype with the distinct upregulation of stromal components and a lack of T cell infiltration in the tumor core; and C4 as the immunomodulatory subtype with the remarkable upregulation of anti-tumor immune components. The four CCCRC subtypes had distinct histopathologic and molecular characteristics, therapeutic efficacy, and prognosis. We found that the C1 subtype may be suitable for chemotherapy and cetuximab, the C2 subtype may benefit from a combination of chemotherapy and bevacizumab, the C3 subtype has increased sensitivity to the WNT pathway inhibitor WIKI4, and the C4 subtype is a potential candidate for immune checkpoint blockade treatment. Importantly, we established a simple gene classifier for accurate identification of each CCCRC subtype. Collectively our integrative analysis ultimately established a holistic framework to thoroughly dissect the TME of CRC, and the CCCRC classification system with high biological interpretability may contribute to biomarker discovery and future clinical trial design.

Recurrence risk assessment for stage III colorectal cancer based on five methylation biomarkers in plasma cell-free DNA.

For stage III colorectal cancer (CRC) patients with a high risk of recurrence, intensified adjuvant chemotherapy can improve overall survival. We aimed to develop a circulating tumor DNA (ctDNA) methylation marker model for predicting the relapse risk of stage III CRC patients. Differentially methylated markers identified between 53 normal mucosa samples and 165 CRC tissue samples, as well as between plasma samples from 75 stage I/II (early-stage) CRC patients and 55 stage IV (late-stage) CRC patients, were analyzed using Student's t-tests. The overlapping methylation markers shared by plasma and tissue samples were used to establish a methylation marker model to evaluate the tumor burden in the peripheral blood of CRC patients using the random forest method. This model was verified in the validation cohort (n = 44) and then applied to predict recurrence risk in 50 stage III CRC patients and monitor the clinical disease course in serial samples from four CRC patients. We built a five-marker-based ctDNA methylation model that had high sensitivity (84.21%) and specificity (84%) in identifying late-stage CRC in a validation cohort containing 24 stage I/II CRC patients and 20 stage IV CRC patients. The model achieved high sensitivity (87.5%) and specificity (94.12%) in predicting tumor relapse in an independent cohort of 50 stage III CRC patients and could be an independent recurrence risk factor for stage III patients [Hazard ratio (HR), 60.4; 95% confidence interval (CI): 7.68-397; p = 9.73e-5]. Analysis of serial blood samples of CRC showed that the model could monitor disease relapse earlier than imaging examination and serum carcinoembryonic antigen (CEA) and so may provide an opportunity for the early adjustment of therapeutic strategies. Moreover, the model could potentially monitor the clinical course and treatment response dynamically. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

HAO1-mediated oxalate metabolism promotes lung pre-metastatic niche formation by inducing neutrophil extracellular traps.

Metabolic reprogramming has been shown to be involved in cancer-induced pre-metastatic niche (PMN) formation, but the underlying mechanisms have been insufficiently explored. Here, we showed that hydroxyacid oxidase 1 (HAO1), a rate-limiting enzyme of oxalate synthesis, was upregulated in the alveolar epithelial cells of mice bearing metastatic breast cancer cells at the pre-metastatic stage, leading to oxalate accumulation in lung tissue. Lung oxalate accumulation induced neutrophil extracellular trap (NET) formation by activating NADPH oxidase, which facilitated the formation of pre-metastatic niche. In addition, lung oxalate accumulation promoted the proliferation of metastatic cancer cells by activating the MAPK signaling pathway. Pharmacologic inhibition of HAO1 could effectively suppress the lung oxalate accumulation induced by primary cancer, consequently dampening lung metastasis of breast cancer. Breast cancer cells induced HAO1 expression and oxalate accumulation in alveolar epithelial cells by activating TLR3-IRF3 signaling. Collectively, these findings underscore the role of HAO1-mediated oxalate metabolism in cancer-induced lung PMN formation and metastasis. HAO1 could be an appealing therapeutic target for preventing lung metastasis of cancer.

CMTM6 expression in M2 macrophages is a potential predictor of PD-1/PD-L1 inhibitor response in colorectal cancer.

BACKGROUND: CMTM6 is a novel key regulator of PD-L1. High expression of both CMTM6 and PD-L1 may predict the benefit of PD-1 axis blockade in lung cancer. We aimed to investigate the expression pattern of CMTM6 between mismatch repair-defective (dMMR) and mismatch repair-proficient (pMMR) colorectal cancer (CRC) tissues and assess its correlation with the response to PD-1/PD-L1 pathway blockade. METHODS: Immunohistochemistry (IHC) was used to analyze CMTM6 and PD-L1 expression and immune cell density in dMMR/pMMR CRC. Quantitative multiplex immunofluorescence (IF) was performed to detect CMTM6, PD-L1, CD4, CD8, CD68 and CD163 expression in CRC patients treated with PD-1/PD-L1 inhibitors. RESULT: IHC analysis showed that CMTM6 and PD-L1 were both expressed in tumor cells (TCs) and invasion front immune cells (ICs). CMTM6 and PD-L1 expression and CD4+, CD8+, CD68+ or CD163+ cell density were significantly higher in dMMR CRC patients than in pMMR CRC patients. CMTM6 expression was positively correlated with PD-L1 expression and CD163+ M2 macrophage density in dMMR CRC. IF analysis showed that the coexpression rate of CMTM6/PD-L1 and the expression rate of CMTM6 in CD8+ T cells and CD163+ M2 macrophages were significantly increased in the group that exhibited clinical benefit. CMTM6 expression in M2 macrophages was identified as the best biomarker for predicting the responsiveness to PD-1/PD-L1 inhibitors. CONCLUSIONS: CMTM6 expression in M2 macrophages may predict the PD-1/PD-L1 inhibitor response rate in CRC patients more accurately than dMMR/microsatellite instability-high (MSI-H) status. It can also identify pMMR CRC patients who could benefit from PD-1/PD-L1 inhibitors.