Yibo Fan

miR-103/107 modulates multidrug resistance in human gastric carcinoma by downregulating Cav-1.

MicroRNAs (miRNAs) are a class of non-protein-coding small RNAs with the capacity to regulate fundamental biological processes essential for cancer initiation and progression. In the present study, we analyzed miRNA expression levels between multidrug-resistant gastric carcinoma cell line SGC7901/ADR and its parent cell line SGC7901 using a miRNA microarray. MiR-103/107 was downregulated compared with parental SGC7901 cells. Overexpression of miR-103/107 sensitized SGC7901/ADR cells to doxorubicin (DOX), as demonstrated by in vitro and in vivo drug sensitivity assay. We further confirmed that miR-103/107 inhibited P-gp function in gastric cancer SGC7901/ADR cells. Finally, we verified that caveolin-1 (Cav-1), a critical component of lipid rafts, was a target of miR-103/107.

Trastuzumab and oxaliplatin exhibit a synergistic antitumor effect in HER2-postive gastric cancer cells

Trastuzumab has recently been recommended for the treatment of epidermal growth factor receptor-2 (HER2)-positive advanced gastric cancer in combination with the capecitabine/cisplatin (XP) versus continuous infusion of 5-fluorouracil/cisplatin (FP) regimen. However, it is unclear whether it is rational to combine trastuzumab with other chemotherapy regimens in clinical practice. Our study demonstrates that adding trastuzumab to oxaliplatin, a commonly used third-generation platinum derivative, increases the antitumor effect in vitro. In MTT assays, combination treatment with oxaliplatin and trastuzumab significantly decreased the concentration of oxaliplatin required to induce 50% growth inhibition in HER2-positive gastric cancer cells. Further investigation revealed that the trastuzumab–oxaliplatin combination induced cell cycle arrest and decreased expression of both p-AKT and p-ERK. Notably, this treatment combination induced downregulation of the excision repair cross-complementation group 1 (ERCC1) protein, which is involved in the key repair process of the oxaliplatin-DNA platinum adduct at the protein level. Similar changes were also observed in HER2-positive breast cancer cells. These findings suggest that trastuzumab synergizes the cytotoxic effect of oxaliplatin on HER2-positive gastric and breast cancer cells. Our study provides preclinical evidence for the optimization of this combination regimen in the treatment of HER2-positive gastric cancer patients.

Bufalin inhibits TGF-β-induced epithelial-to-mesenchymal transition and migration in human lung cancer A549 cells by downregulating TGF-β receptors.

The epithelial-to-mesenchymal transition (EMT) is a well-known prerequisite for cancer cells to acquire the migratory and invasive capacity, and to subsequently metastasize. Bufalin is one of the major active components of the traditional Chinese medicine Chan Su, and accumulating evidence has shown its anticancer effect in multipe types of cancer. However, the role of bufalin in transforming growth factor-β (TGF-β)-induced EMT and migration remains unclear. In the present study, the effect of bufalin on TGF-β-induced EMT and migration was investigated in human lung cancer A549 cells. TGF-β induced EMT in A549 cells and increased their migratory ability, which were markedly suppressed by bufalin. Additionally, TGF-β-induced upregulation of Twist2 and zinc finger E-box binding homeobox 2 (ZEB2), as well as the phosphorylation of Smad2 and Smad3 were also inhibited by bufalin. However, the Smad-independent signaling pathways were not affected. Further analysis showed that the TGF-β receptor I (TβRI) and TGF-β receptor II (TβRII) were downregulated in the presence of bufalin. Pretreatment with SB431542, a potent inhibitor of the phosphorylation of TβRI, significantly attenuated TGF-β-induced EMT, mimicking the effect of bufalin on A549 cells. Taken together, these results suggest that bufalin suppresses TGF-β-induced EMT and migration by downregulating TβRI and TβRII in A549 cells.

Gastric cancer‐derived exosomes promote peritoneal metastasis by destroying the mesothelial barrier

An intact mesothelium serves as a protective barrier to inhibit peritoneal carcinomatosis. Cancer‐derived exosomes can mediate directional tumor metastasis; however, little is known about whether gastric cancer‐derived exosomes will destroy the mesothelial barrier and promote peritoneal dissemination. Here, we demonstrate that gastric cancer‐derived exosomes facilitate peritoneal metastasis by causing mesothelial barrier disruption and peritoneal fibrosis. Injury of peritoneal mesothelial cells elicited by gastric cancer‐derived exosomes is through concurrent apoptosis and mesothelial‐to‐mesenchymal transition (MMT). Additionally, upregulation of p‐ERK in peritoneal mesothelial cells is primarily responsible for the MMT while contributing little to apoptosis. Together, these data support the concept that exosomes play a crucial role in remodeling the premetastatic microenvironment and identify a novel mechanism for peritoneal metastasis of gastric carcinoma.

Tamoxifen reverses epithelial–mesenchymal transition by demethylating miR-200c in triple-negative breast cancer cells

BackgroundAlthough the efficacy of tamoxifen (TAM) for breast cancer has been attributed to inducing cell cycle arrest and apoptosis by inhibiting estrogen receptor (ER) signaling, recent evidence indicates that TAM also possesses ER-independent antitumor activity through an unclear mechanism. The present study investigated the anti-tumor mechanism of TAM on mesenchymal triple-negative breast cancer (TNBC).MethodsThe inhibitory effect of TAM on tumor migration and metastasis was analyzed by transwell chamber in vitro and by murine xenograft model in vivo. The promoter sequence of miR-200c was predicted by an online CpG island predictor. Relative expression of miR-200c was measured by quantitative real-time PCR.ResultsAfter treatment with TAM, mesenchymal TNBC cells (MCF-7/ADR and MDA-MB-231) morphologically changed from mesenchymal to epithelial types. Meanwhile, cell migration ability was also significantly decreased in ER-positive breast cancer cells after exposure to TAM. Consistent with these in-vitro results, TAM significantly suppressed lung metastasis rate of mesenchymal TNBC cells in murine xenograft tumors. miRNA array analysis of two types of breast cancer cells showed that miR-200c expression was inhibited in mesenchymal TNBC cells, but increased after TAM treatment due to demethylation of miR-200c promoters.ConclusionsOur results indicate that TAM inhibits cell migration and enhances chemosensitivity of mesenchymal TNBC cells by reversing their EMT-like property; and that this EMT-reversal effect results from upregulation of miR-200c through demethylating its promoter. To our knowledge, this is the first explanation of a non-ER-related mechanism for the effect of TAM on mesenchymal TNBC cells.

MicroRNA-891b is an independent prognostic factor of pancreatic cancer by targeting Cbl-b to suppress the growth of pancreatic cancer cells

Growing evidence has revealed that microRNAs could regulate the proliferation of pancreatic ductal adenocarcinoma (PDAC) cells and predict the prognosis of PDAC. Here the comparative microRNA expression profiles of the good and poor prognosis groups were performed by microRNA microarray. MicroRNA-891b (miR-891b) was screened and validated to be a prognostic predictor of PDAC in the initial group and further evaluated to be an independent predictor for the overall survival of resectable PDACs in an independent cohort. By a series of cellular and animal experiments, as well as clinical specimen analyses, miR-891b was confirmed to target the Cbl-b gene, promot the expression of tumor suppressor p21 protein and inhibit the proliferation of PDAC cells. The results provide a theoretical basis for the study of miR-891b as an independent prognostic predictor of PDAC and the role of miR-891b/Cbl-b pathway in this prediction, as well as the identification of new targets for PDAC.

E3 Ubiquitin Ligase Cbl-b Prevents Tumor Metastasis by Maintaining the Epithelial Phenotype in Multiple Drug-Resistant Gastric and Breast Cancer Cells

Multiple drug resistance (MDR) and metastasis are two major factors that contribute to the failure of cancer treatment. However, the relationship between MDR and metastasis has not been characterized. Additionally, the role of the E3 ubiquitin ligase Cbl-b in metastasis of MDR gastric and breast cancer is not well known. In the present study, we found that MDR gastric and breast cancer cells possess a typical mesenchymal phenotype and enhanced cell migration capacity. Additionally, Cbl-b is poorly expressed in MDR gastric and breast cancer cells. In MDR gastric adenocarcinoma tissues, gastric cancer patients with low Cbl-b expression were more likely to have tumor invasion (P = .016) and lymph node metastasis (P = .007). Moreover, overexpression of Cbl-b reduced cell migration in MDR cell cultures both in vitro and in vivo. Cbl-b overexpression also prevented EMT by inducing ubiquitination and degradation of EGFR, leading to inhibition of the EGFR-ERK/Akt-miR-200c-ZEB1 axis. However, further overexpression of EGFR on a background of Cbl-b overexpression restored both the mesenchymal phenotype and cell migration capacity of MDR gastric and breast cancer cells. These results suggest that Cbl-b is an important factor for maintenance of the epithelial phenotype and inhibition of cell migration in MDR gastric and breast cancer cells.

DR5‐Cbl‐b/c‐Cbl‐TRAF2 complex inhibits TRAIL‐induced apoptosis by promoting TRAF2‐mediated polyubiquitination of caspase‐8 in gastric cancer cells

Ubiquitination of caspase‐8 regulates TNF‐related apoptosis‐inducing ligand (TRAIL) sensitivity in cancer cells, and the preligand assembly complex plays a role in caspase‐8 polyubiquitination. However, whether such a complex exists in gastric cancer cells and its role in TRAIL‐triggered apoptosis is unclear. In this study, DR5, casitas B‐lineage lymphoma‐b (Cbl‐b)/c‐Cbl, and TRAF2 formed a complex in TRAIL‐resistant gastric cancer cells, and Cbl‐b and c‐Cbl were the critical adaptors linking DR5 and TRAF2. Treatment with TRAIL induced caspase‐8 translocation into the DR5‐Cbl‐b/c‐Cbl‐TRAF2 complex to interact with TRAF2, which then mediated the K48‐linked polyubiquitination of caspase‐8. The proteasome inhibitor bortezomib markedly enriched the p43/41 products of caspase‐8 activated by TRAIL, indicating proteasomal degradation of caspase‐8. Moreover, TRAF2 knockdown prevented the polyubiquitination of caspase‐8 and thus increased TRAIL sensitivity. In addition, the inhibition of Cbl‐b or c‐Cbl expression and overexpression of miR‐141 targeting Cbl‐b and c‐Cbl partially reversed TRAIL resistance by inhibiting the interaction between TRAF2 and caspase‐8 and the subsequent polyubiquitination of caspase‐8. These results indicate that the DR5‐Cbl‐b/c‐Cbl‐TRAF2 complex inhibited TRAIL‐induced apoptosis by promoting TRAF2‐mediated polyubiquitination of caspase‐8 in gastric cancer cells.

E3 ubiquitin ligases Cbl‐b and c‐Cbl downregulate PD‐L1 in EGFR wild‐type non‐small cell lung cancer

Anti‐PD‐1/PD‐L1 therapies have demonstrated prominent clinical effects in the treatment of non‐small cell lung cancer (NSCLC). However, limited understanding of the regulatory mechanisms of PD‐L1 has become one of the biggest challenges for further improving efficacy. In this study, we observed that in wild‐type EFGR cell lines A549 and H460, the ubiquitin ligases Cbl‐b and c‐Cbl inhibit PD‐L1 by inactivating STAT, AKT, and ERK signaling. MiR‐181a and miR‐940 were screened and validated to target Cbl‐b and c‐Cbl, respectively. Furthermore, in NSCLC tissues, the expression of Cbl‐b/c‐Cbl is negatively correlated with PD‐L1 expression. Taken together, these findings indicated a new regulatory mechanism for PD‐L1 in wild‐type EGFR NSCLC cell lines by Cbl‐b and c‐Cbl.

The Chemokine Receptor CXCR4 and c-MET Cooperatively Promote Epithelial-Mesenchymal Transition in Gastric Cancer Cells

The C-X-C motif chemokine receptor 4 (CXCR4) pathway can promote tumor metastasis but is dependent on cross talk with other signaling pathways. The MET proto-oncogene (c-MET) participates in metastasis and is highly expressed in gastric cancer. However, the relationship between CXCR4 and c-MET signaling and their mechanisms of action in gastric cancer metastasis remain unclear. In this study, in vitro experiments demonstrated that C-X-C motif chemokine ligand 12 (CXCL12)/CXCR4 induces epithelial-mesenchymal transition (EMT) and promotes migration in gastric cancer cells, which is accompanied by c-MET activation. These phenomena were reversed by c-MET inhibition. Further investigation revealed that c-MET activation correlated with its interaction with caveolin 1 in lipid rafts, induced by CXCL12. In clinical samples, we observed a significant positive association between CXCR4 expression and c-MET phosphorylation (r = 0.259, P = .005). Moreover, samples expressing both receptors were found to indicate significantly poorer patient prognosis (P < .001). These results suggest that CXCL12 induces EMT at least partially through cross talk between CXCR4 and c-MET signaling. In addition, changes in these pathways could have clinical importance for the treatment of gastric cancer.