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Esophageal squamous cell carcinoma (ESCC) is a highly malignant gastrointestinal cancer with a high recurrence rate and poor prognosis. Although N6-methyladenosine (m6A), the most abundant epitranscriptomic modification of mRNAs, has been implicated in several cancers, little is known about its participation in ESCC progression. We found reduced expression of ALKBH5, an m6A demethylase, in ESCC tissue specimens with a more pronounced effect in T3-T4, N1-N3, clinical stages III-IV, and histological grade III tumors, suggesting its involvement in advanced stages of ESCC. Exogenous expression of ALKBH5 inhibited the in vitro proliferation of ESCC cells, whereas depletion of endogenous ALKBH5 markedly enhanced ESCC cell proliferation in vitro. This suggests ALKBH5 exerts anti-proliferative effects on ESCC growth. Furthermore, ALKBH5 overexpression suppressed tumor growth of Eca-109 cells in nude mice; conversely, depletion of endogenous ALKBH5 accelerated tumor growth of TE-13 cells in vivo. The growth-inhibitory effects of ALKBH5 overexpression are partly attributed to a G1-phase arrest. In addition, ALKBH5 overexpression reduced the in vitro migration and invasion of ESCC cells. Altogether, our findings demonstrate that the loss of ALKBH5 expression contributes to ESCC malignancy.

Background: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide and has a poor prognosis due to the high incidence of invasion and metastasis-related progression. However, the underlying mechanism remains elusive, and valuable biomarkers for predicting invasion, metastasis, and poor prognosis of HCC patients are still lacking. Methods: Immunohistochemistry (IHC) was performed on HCC tissues (n = 325), and the correlations between MST4 expression of the clinical HCC tissues, the clinicopathologic features, and survival were further evaluated. The effects of MST4 on HCC cell migratory and invasive properties in vitro were evaluated by Transwell and Boyden assays. The intrahepatic metastasis mouse model was established to evaluate the HCC metastasis in vivo. The PI3K inhibitor, LY294002, and a specific siRNA against Snail1 were used to investigate the roles of PI3K/AKT pathway and Snail1 in MST4-regulated EMT, migration, and invasion of HCC cells, respectively. Results: In this study, by comprehensively analyzing our clinical data, we discovered that low MST4 expression is highly associated with the advanced progression of HCC and serves as a prognostic biomarker for HCC patients of clinical-stage III-IV. Functional studies indicate that MST4 inactivation induces epithelial-to-mesenchymal transition (EMT) of HCC cells, promotes their migratory and invasive potential in vitro, and facilitates their intrahepatic metastasis in vivo, whereas MST4 overexpression exhibits the opposite phenotypes. Mechanistically, MST4 inactivation elevates the expression and nuclear translocation of Snail1, a key EMT transcription factor (EMT-TF), through the PI3K/AKT signaling pathway, thus inducing the EMT phenotype of HCC cells, and enhancing their invasive and metastatic potential. Moreover, a negative correlation between MST4 and p-AKT, Snail1, and Ki67 and a positive correlation between MST4 and E-cadherin were determined in clinical HCC samples. Conclusions: Our findings indicate that MST4 suppresses EMT, invasion, and metastasis of HCC cells by modulating the PI3K/AKT/Snail1 axis, suggesting that MST4 may be a potential prognostic biomarker for aggressive and metastatic HCC.

In recent years, the development of adjunctive therapeutic hyperthermia for cancer therapy has received considerable attention. However, the mechanisms underlying hyperthermia resistance are still poorly understood. In this study, we investigated the roles of cold-inducible RNA binding protein (Cirbp) in regulating hyperthermia resistance and underlying mechanisms in nasopharyngeal carcinoma (NPC). Our results firstly revealed that hyperthermia significantly attenuated the stemness property of NPC cells, while combination treatment of hyperthermia and oridonin dramatically increased the killing effect on NPC cells and cancer stem cell (CSC)-like population. Moreover, hyperthermia substantially improved the sensitivity of radiation-resistant NPC cells and CSC-like cells to radiotherapy. Hyperthermia noticeably suppressed Cirbp expression in NPC cells and xenograft tumor tissues. Furthermore, Cirbp inhibition remarkably boosted anti-tumor-killing activity of hyperthermia against NPC cells and CSC-like cells, whereas ectopic expression of Cirbp compromised tumor-killing effect of hyperthermia on these cells, indicating that Cirbp overexpression induces hyperthermia resistance. ThermomiR-377–3p improved the sensitivity of NPC cells and cancer stem-like cells to hyperthermia in vitro by directly suppressing Cirbp expression. More importantly, our results displayed the significantly boosted sensitization of tumor xenografts to hyperthermia by Cirbp silencing in vivo, but ectopic expression of Cirbp nearly completely counteracted hyperthermia-mediated tumor cell-killing effect against tumor xenografts in vivo. Mechanistically, Cirbp silencing-induced inhibition of DNA damage repair by inactivating ATM-Chk2 and ATR-Chk1 pathways, decrease in stemness and increase in cell death contributed to hyperthermic sensitization; conversely, Cirbp overexpression-induced promotion of DNA damage repair, increase in stemness and decrease in cell apoptosis contributed to hyperthermia resistance. Taken together, these findings reveal a previously unrecognized role for Cirbp in positively regulating hyperthermia resistance and suggest that thermomiR-377–3p and its target gene Cirbp represent promising targets for therapeutic hyperthermia.

Objective: To investigate the effects of Maytenus compound on the proliferation of hepatocellular carcinoma (HCC) cells in vitro and in vivo and to explore the underlying mechanism. Methods: The half maximal inhibitory concentration (IC50) values of Maytenus compound in HepG2 and BEL-7402 cells were determined by the MTS assay. HepG2 and BEL-7402 cells were treated with different concentrations of Maytenus compound. MTS assays, colony formation assays and cell cycle analyses were performed to clarify the inhibitory effect of Maytenus compound on the proliferation of HepG2 and BEL-7402 cells in vitro. After subcutaneous injection of HepG2 cells, nude mice were randomly divided into a vehicle control group and a drug intervention group, which were intragastrically administered ddH2O or Maytenus compound, respectively. The inhibitory effect of Maytenus compound on the proliferation of HepG2 cells in vivo was analyzed using subcutaneous tumor growth curves, tumor weight, the tumor growth inhibition rate and the immunohistochemical detection of BrdU-labeled cells in S phase. The organ toxicity of Maytenus compound was initially evaluated by comparing the weight difference and organ index of the two groups of nude mice. The main proteins in the EGFR-PI3K-AKT signaling pathway were detected by Western blot after Maytenus compound intervention in vivo and in vitro. Results: Maytenus compound showed favorable antiproliferation activity against HepG2 and BEL-7402 cells with IC50 values of 79.42±11.71 µg/mL and 78.48±8.87 µg/mL, respectively. MTS assays, colony formation assays and cell cycle analyses showed that Maytenus compound at different concentration gradients within the IC50 concentration range significantly suppressed the proliferation of HepG2 and BEL-7402 cells in vitro and inhibited cell cycle progression from G1 to S phase. Additionally, Maytenus compound, at an oral dose of 2.45 g/kg, dramatically inhibited, without obvious organ toxicity, the proliferation of subcutaneous tumors formed by HepG2 cells in nude mice. In addition, the tumor growth inhibition rate for Maytenus compound was 66.94%. Furthermore, Maytenus compound inhibited the proliferation of liver orthotopic transplantation tumors in nude mice. Western blot analysis showed that Maytenus compound significantly downregulated the expression of p-EGFR, p-PI3K, and p-AKT and upregulated the expression of p-FOXO3a, p27, and p21 in vivo and in vitro. Conclusion: Maytenus compound significantly inhibited the proliferation of HCC cells in vitro and in vivo. The downregulation of the EGFR-PI3K-AKT signaling pathway and subsequent inhibition of cell cycle progression from G1 to S phase is one of the possible mechanisms. Maytenus compound has a high tumor growth inhibition rate and has no obvious organ toxicity, which may make it a potential anti-HCC drug, but the results from this study need to be confirmed by further clinical trials in HCC patients.