Background: Cancer stem cells (CSCs) are able to survive after cancer therapies, resulting in tumor progression and recurrence, as is seen in colorectal cancer. Therapies targeting CSCs are regarded as novel and promising strategies for efficiently eradicating tumors. Berberine, an isoquinoline alkaloid extracted from the Chinese herbal medicine Coptis chinensis, was found to have antitumor activities against colorectal cancer, without knowing whether it exerts inhibitory effects on colorectal CSCs and the potential mechanisms.Methods: In this study, we examined the inhibitory roles of Berberine on CSCs derived from HCT116 and HT29 by culturing in serum-free medium. We also examined the effects of Berberine on m⁶A methylation via regulating fat mass and obesity-associated protein (FTO), by downregulating β-catenin.Results: We examined the effects of Berberine on the tumorigenicity, growth, and stemness of colorectal cancer stem-like cells. The regulatory effect of Berberine on N6-methyladenosine (m⁶A), an abundant mRNA modification, was also examined. Berberine treatment decreased cell proliferation by decreasing cyclin D1 and increasing p27 and p21 and subsequently induced cell cycle arrest at the G₁/G₀ phase. Berberine treatment also decreased colony formation and induced apoptosis. Berberine treatment transcriptionally increased FTO and thus decreased m⁶A methylation, which was reversed by both FTO knockdown and the addition of the FTO inhibitor FB23-2. Berberine induced FTO-related decreases in stemness in HCT116 and HT29 CSCs. Berberine treatment also increased chemosensitivity in CSCs and promoted chemotherapy agent-induced apoptosis. Moreover, we also found that Berberine treatment increased FTO by decreasing β-catenin, which is a negative regulator of FTO.Conclusions: Our observation that Berberine effectively decreased m⁶A methylation by decreasing β-catenin and subsequently increased FTO suggests a role of Berberine in modulating stemness and malignant behaviors in colorectal CSCs.

Background: Glioma is the most common and lethal form of brain cancer. It is highly malignant and is often characterized by chemoresistance and radioresistance, which are thought to mainly result from hypoxic microenvironments. Various tumour-promoting and tumour-suppressing microRNAs (miRNAs) have been identified in gliomas; however, it is still largely unknown how miRNAs are modified by hypoxia and subsequently affect glioma. In this study, we examined the expression of miR-210-3p, a well-characterized miRNA that responds to hypoxia in glioma cell lines. Methods: The expressions of miR-9 and miR-210-3p were analysed by using qPCR. Cell viability was measured by performing CCK-8 after eechinomycin treatment or introduction of miR-210 for 24 or 48 h. The correlation of HIF-1α expression with TGF-β were analysed using the REMBRANDT database. The biomarkers of EMT, including E-cadherin, N-cadherin and Vimentin, were detected by western blot. Apoptotic cell death was measured by performing Annexin V-FITC/PI double staining followed by flow cytometry. Results: We found that miR-210-3p was induced by a mechanism dependent on the hypoxia-induced transcriptional activity of HIF-1α. Then we established a positive association between the HIF-1α and TGF-β expression levels, and miR-210-3p upregulation induced TGF-β expression, indicating that hypoxia-induced HIF-1α activity upregulated TGF-β via miR-210-3p upregulation. Hypoxia-induced miR-210-3p activity was found to promote EMT by upregulating TGF-β, which subsequently enhanced the invasive ability in U87-MG cells. We further confirmed that miR-210-3p induced chemoresistance to TMZ in U87-MG cells via TGF-β upregulation under hypoxic conditions. Conclusion: These results help to reveal the potential regulatory mechanisms of hypoxia-induced miR-210-3p expression that affect malignant behaviors and chemoresistance via TGF-β upregulation in glioma cells.

Background: The mitochondrial-associated protein leucine-rich pentatricopeptide repeat-containing (LRPPRC) exerts multiple functions involved in physiological processes, including mitochondrial gene translation, cell cycle progression, and tumorigenesis. Previously, LRPPRC was reported to regulate mitophagy by interacting with Bcl-2 and Beclin-1 and thus modifying the activation of PI3KCIII and autophagy. Considering that LRPPRC was found to be negatively associated with survival rate, we hypothesize that LRPPRC may be involved in pancreatic cancer progression via its regulation of autophagy.Methods: Real-time quantitative polymerase chain reaction was performed to detect the expression of LRPPRC in 90 paired pancreatic cancer and adjacent tissues and five pancreatic cancer cell lines. Mitochondrial reactive oxidative species level and function were measured. Mitophagy was measured by performing to detect LC3 levels.Results: By performing a real-time quantitative polymerase chain reaction, the association of LRPPRC with the prognosis of pancreatic cancer was established, and pancreatic cancer tissues had significantly higher LRPPRC expression than adjacent tissues. LRPPRC was negatively associated with the overall survival rate. LRPPRC was also upregulated in pancreatic cancer cell lines. Knockdown of LRPPRC promoted reactive oxidative species accumulation, decreased mitochondrial membrane potential, promoted autophagy/mitophagy, and induced mitochondrial dysfunction. Subsequently, knockdown of LRPPRC inhibited malignant behaviors in PANC-1 cells, including proliferation, migration, invasion, tumor formation, and chemoresistance to gemcitabine. Finally, by inhibiting autophagy/mitophagy using 3-MA, the inhibitory effect of LRPPRC knockdown on proliferation was reversed.Conclusion: Taken together, our results indicate that LRPPRC may act as an oncogene via maintaining mitochondrial homeostasis and could be used as a predictive marker for patient prognosis in pancreatic cancer.

Cancer stem cells (CSCs) are the core factors leading to recurrence, insensitivity to radiotherapy and chemotherapy, and immunotherapy resistance in patients with colorectal cancer. AT7867, a potent oral AKT inhibitor, was found to have antitumor activity in colorectal cancer; however, the effect on colorectal cancer stem cells is still unclear. This study was conducted to clarify the molecular mechanism underlying the CSC growth inhibitory effects of AT7867. We cultured colorectal cancer cells (CRCs) in a serum-free medium and enriched colorectal cancer stem cells. Subsequently, the effects of AT7867 on CSCs were analyzed by CCK-8, colony formation, flow cytometry, and immunofluorescence assays. The results indicated that AT7867 induces G2/M phase arrest and cell apoptosis in cancer stem cells. Subsequently, we identified Ascl2 as the main gene affecting the stemness of colorectal cancer in AT7867 by RNA sequencing. The current study showed that Ascl2 is involved in the metastasis, invasion, and proliferation of CRCs. The next experiments demonstrated that overexpression of Ascl2 did affect the therapeutic effect of AT7867 on CRC stemness. Furthermore, compared with other Akt inhibitors, AT7867 could promote the differentiation of colorectal cancer stem cells. Thus, AT7867 might be a potential antitumor drug candidate to treat CRC by targeting CSCs.