Ferroptosis is a newly identified iron-dependent form of death that is becoming increasingly recognized as a promising avenue for cancer therapy. N6-methyladenosine (m⁶A) is the most abundant reversible methylation modification in mRNA contributing to tumorigenesis. However, the crucial role of m⁶A modification in regulating ferroptosis during colorectal cancer (CRC) tumorigenesis remains elusive. Herein, we find that m⁶A modification is increased during ferroptotic cell death and correlates with the decreased m⁶A demethylase fat mass and obesity-associated protein (FTO) expression. Functionally, we demonstrate that suppressing FTO significantly induces CRC ferroptotic cell death, as well as enhancing CRC cell sensitivity to ferroptosis inducer (Erastin and RSL3) treatment. Mechanistically, high FTO expression increased solute carrier family 7 member 11 (SLC7A11) or glutathione peroxidase 4 (GPX4) expressions in an m⁶A-YTHDF2 dependent manner, thereby counteracting ferroptotic cell death stress. In addition, we identify Mupirocin as a novel inhibitor of FTO, and Mupirocin induces CRC ferroptosis and inhibits tumor growth. Clinically, the levels of FTO, SLC7A11, and GPX4, are highly correlated expression in CRC tissues. Our findings reveal that FTO protects CRC from ferroptotic cell death in promoting CRC tumorigenesis through triggering SLC7A11/GPX4 expression.

Colorectal cancer exhibits a notable prevalence and propensity for metastasis, but the current therapeutic interventions for metastatic colorectal cancer have yielded suboptimal results. ICIs can decrease tumor development by preventing the tumor’s immune evasion, presenting cancer patients with a new treatment alternative. The increased use of immune checkpoint inhibitors (ICIs) in CRC has brought several issues. In particular, ICIs have demonstrated significant clinical effectiveness in patients with MSI-H CRC, whereas their efficacy is limited in MSS. Acquired resistance can still occur in patients with a positive response to ICIs. This paper describes the efficacy of ICIs currently in the clinical treatment of CRC, discusses the mechanisms by which acquired resistance occurs, primarily related to loss and impaired presentation of tumor antigens, reduced response of IFN-λ and cytokine or metabolic dysregulation, and summarizes the incidence of adverse effects. We posit that the future of ICIs hinges upon the advancement of precise prediction biomarkers and the implementation of combination therapies. This study aims to elucidate the constraints associated with ICIs in CRC and foster targeted problem-solving approaches, thereby enhancing the potential benefits for more patients.

Background: With the recent emergence of immune checkpoint inhibitors, microsatellite instability (MSI) status has become an important biomarker for immune checkpoint blockade therapy. There are growing technical demands for the integration of different genomic alterations profiling including MSI analysis in a single assay for full use of the limited tissues. Methods: Tumor and paired control samples from 64 patients with primary colorectal cancer were enrolled in this study, including 14 MSI-high (MSI-H) cases and 50 microsatellite stable (MSS) cases determined by MSI-PCR. All the samples were sequenced by a customized NGS panel covering 2.2 MB. A training dataset of 28 samples was used for selection of microsatellite loci and a novel NGS-based MSI status classifier, USCI-msi, was developed. NGS-based MSI status, single nucleotide variant (SNV) and tumor mutation burden (TMB) were detected for all patients. Most of the patients were also independently detected by immunohistochemistry (IHC) staining. Results: A 9-loci model for detecting microsatellite instability was able to correctly predict MSI status with 100% sensitivity and specificity compared with MSI-PCR, and 84.3% overall concordance with IHC staining. Mutations in cancer driver genes (APC, TP53, and KRAS) were dispersed in MSI-H and MSS cases, while BRAF p.V600E and frameshifts in TCF7L2 gene occurred only in MSI-H cases. Mismatch repair (MMR)-related genes are highly mutated in MSI-H samples. Conclusion: We established a new NGS-based MSI classifier, USCI-msi, with as few as 9 microsatellite loci for detecting MSI status in CRC cases. This approach possesses 100% sensitivity and specificity, and performed robustly in samples with low tumor purity.

Background: Aberrant alternative splicing (AS) is a pervasive event during colorectal cancer (CRC) development. SF3B3 is a splicing factor component of U2 small nuclear ribonucleoproteins which are crucial for early stages of spliceosome assembly. The role of SF3B3 in CRC remains unknown. Methods: SF3B3 expression in human CRCs was analyzed using publicly available CRC datasets, immunohistochemistry, qRT-PCR, and western blot. RNA-seq, RNA immunoprecipitation, and lipidomics were performed in SF3B3 knockdown or overexpressing CRC cell lines. CRC cell xenografts, patient-derived xenografts, patient-derived organoids, and orthotopic metastasis mouse models were utilized to determine the in vivo role of SF3B3 in CRC progression and metastasis. Results: SF3B3 was upregulated in CRC samples and associated with poor survival. Inhibition of SF3B3 by RNA silencing suppressed the proliferation and metastasis of CRC cells in vitro and in vivo, characterized by mitochondria injury, increased reactive oxygen species (ROS), and apoptosis. Mechanistically, silencing of SF3B3 increased mTOR exon-skipped splicing, leading to the suppression of lipogenesis via mTOR-SREBF1-FASN signaling. The combination of SF3B3 shRNAs and mTOR inhibitors showed synergistic antitumor activity in patient-derived CRC organoids and xenografts. Importantly, we identified SF3B3 as a critical regulator of mTOR splicing and autophagy in multiple cancers. Conclusions: Our findings revealed that SF3B3 promoted CRC progression and metastasis by regulating mTOR alternative splicing and SREBF1-FASN-mediated lipogenesis, providing strong evidence to support SF3B3 as a druggable target for CRC therapy.

Hepatocellular carcinoma (HCC) is not only one of the four highest malignancies, but also the principal reason of cancer-related death worldwide, yet no effective medication for anti-HCC is available. Stachydrine hydrochloride (SH), an alkaloid component in Panzeria alaschanica Kupr, exhibits potent antitumor activity in breast cancer. However, the anti-HCC effects of SH remain unknown. Our study assessed the therapeutic effect of SH on HCC and tried to clarify the mechanisms by which it ameliorates HCC. No studies involving using SH for anti-HCC activity and molecular mechanism have been reported yet. We examined the cell viability of SH on HCC cells by MTT assay. The effect of SH on cell autophagy in HCC cells was verified by Western blot and Immunofluorescence test. Flow cytometry was performed to assess cell-cycle arrest effects. Cell senescence was detected using β-Gal staining and Western blot, respectively. An inhibitor or siRNA of autophagy, i.e., CQ and si LC-3B, were applied to confirm the role of autophagy acted in the anti-cancer function of SH. Protein expression in signaling pathways was detected by Western blot. Besides, molecular docking combined with cellular thermal shift assay (CETSA) was used for analysis. Patient-derived xenograft (PDX) model were built to explore the inhibitory effect of SH in HCC in vivo. In vitro studies showed that SH possessed an anti-HCC effect by inducing autophagy, cell-cycle arrest and promoting cell senescence. Specifically, SH induced autophagy with p62 and LC-3B expression. Flow cytometry analysis revealed that SH caused an obvious cell-cycle arrest, accompanied by the decrease and increase in Cyclin D1 and p27 levels, respectively. Additionally, SH induced cell senescence with the induction of p21 in HCC cell lines. Mechanistically, SH treatment down-regulated the LIF and up-regulated p-AMPK. Moreover, PDX model in NSG mice was conducted to support the results in vitro. This study is the first to report the inhibitory function of SH in HCC, which may be due to the induction of autophagy and senescence. This study provides novel insights into the anti-HCC efficacy of SH and it might be a potential lead compound for further development of drug candidates for HCC.