Regulation of epidermal growth factor receptor tyrosine kinase inhibitor resistance via Ambra1-mediated autophagy in non-small cell lung cancer.

To explore the molecular mechanisms related to epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKI) resistance, along with potential therapeutic targets and strategies. The autophagy and Beclin 1 regulator 1 (Ambra1) short hairpin ribonucleic acid (shRNA) lentivirus vector and Ambra1 overexpression plasmid, constructed with a plasmid cloning deoxyribonucleic acid (pcDNA) 3.1 vector, were used to down-regulate and up-regulate Ambra1 expression in the human lung adenocarcinoma erlotinib-resistant cell line (PC9/ER), respectively, as well as to screen stable transgenic cell lines. The IC50 of Erlotinib in these cell lines were measured to determine their resistance status. The real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to measure messenger ribonucleic acid (mRNA) expression of resistance-related genes like multidrug resistance protein 1 (MDR1), multidrug resistance-associated protein 1 (MRP1), and lung drug-resistant-related protein (LRP). Western blot was performed to analyze the protein expressions of the autophagy-related genes Beclin 1, LC3II/I, and p62. Each stable transgenic line formed a tumor under the skin in nude mice; the mice with subcutaneous tumorigenesis of PC9/ER cells and shAmbra1-PC9/ER cells were subsequently treated with rapamycin (RAPA) and chloroquine (CQ), respectively. The mRNA expressions of MDR1, MRP1, and LRP in each tumor tissue sample were detected by qRT-PCR. The protein expressions of adenosine monophosphate-activated protein kinase (AMPK), phosphorylated-AMPK (p-AMPK), forkhead box O3 (FoxO3a), and phosphorylated forkhead box O3 (p-FoxO3a) in the AMPK/FoxO3a signaling pathway were analyzed via Western blot. The qRT-PCR result revealed that the level of Ambra1 in EGFR-TKI-resistant cells had increased. This was further exacerbated by the overexpression of Ambra1 and was reduced after its inhibition. Additionally, Ambra1 upregulated the mRNA expression of drug-resistant genes and the expression of autophagy-related proteins. Subcutaneous tumorigenesis of RAPA-treated shAmbra1-PC9/ER cells resulted in increased expression of drug resistance-related genes and a concomitant decrease in p-AMPK and increase in p-FoxO3a. The results revealed that Beclin-1/ß-actin, p62/ß-actin, and LC3II/I in the model group were all significantly increased compared to the control group, with P<0.05. Compared to the model group, Beclin-1/ß-actin, p62/ß-actin, and LC3II/I were all significantly higher in the pcDNA-Ambra1 group, with P<0.05. Compared to the model group, Beclin-1/ß-actin, p62/ß-actin, and LC3II/I were all significantly decreased in the shAmbra1 group, with P<0.05. Thus, these data suggest that Ambra1 promotes cellular autophagy. In addition, subcutaneous tumorigenesis of CQ-treated shAmbra1-PC9/ER cells resulted in reduced expression of drug resistance-related genes, and a concomitant increase in p-AMPK and decrease in p-FoxO3a. The results of this study revealed that Ambra1-mediated autophagy regulated EGFR-TKI resistance in NSCLC, most probably through the AMPK/FoxO3a signaling pathway.

Efficacy and predictive factors of glucocorticoid therapy for patients with hepatitis B virus-related acute-on-chronic liver failure.

Background and study aims: Glucocorticoid (GC) treatment for liver failure is controversial. This study sought to evaluate the efficacy and predictive factors of glucocorticoid therapy for hepatitis B virus-related acute-on-chronic liver failure (HBV- ACLF). Patients and methods: A total of 302 patients with HBV- ACLF were enrolled and categorized by treatment modality (GC vs. Control). Baseline characteristics, liver function, disease complications, and mortality were recorded. Univariate and multivariate analysis were used to identify predictive factors for HBV-ACLF-related mortality. Results: GC therapy significantly improved the 30- and 60-day mortality of HBV-ACLF patients (4.64% vs. 11.92%, P=0.022 and 16.56% vs. 25.83%, P=0.049 for the Control and GC groups, respectively) and GC was an independent prognostic factor for 30-day mortality (OR = 0.177, 95% CI 0.051-0.616, P = 0.007). However, enhanced survival was not associated with improved liver function. There were no significant differences in the incidence of complications (i.e., ascites, bacterial infection, encephalopathy, hepatorenal syndrome, and gastrointestinal bleeding) between the GC and Control groups (P >0.05), except that fungal infection occurred with higher frequency in the GC group (P = 0.037). A significant improvement in the 30-day survival associated with GC therapy was observed among patients <40 years of age, a Model for End-stage Liver Disease (MELD) score of 25-35 or a CLIF- Consortium ACLF (CLIF-C ACLF) grade 0-1 (all P <0.05). Conclusions: GC therapy improved the short-term (30- and 60- day) mortality of patients with HBV-ACLF. This treatment may be of particular benefit to patients who are <40 years of age, have a MELD score of 25-35, or have a CLIF-C ACLF grade of 0-1. (Acta gastroenterol. belg., 2022, 85, 593-600).

Long non-coding RNA NEAT1 promotes viability and migration of gastric cancer cell lines through up-regulation of microRNA-17.

OBJECTIVE: Gastric carcinoma (GC) is a common cancer with heavy mortality and poor outcome at advanced stages and metastasis. Long non-coding RNA nuclear-enriched abundant transcript 1 (NEAT1) has been reported to be an oncogene in GC recently. However, the underlying mechanism is far from understood. We aimed to explore the role of NEAT1 in GC as well as the underlying mechanisms. PATIENTS AND METHODS: The expression of NEAT1 in clinical human GC tissues and GC cell lines were assessed by quantitative reverse transcription PCR. Then, NEAT1 was non-physiologically expressed in GC cells (SGC-7901 and MKN45 cells), followed by estimation of cell viability, migration, invasion, apoptosis, activation of the phosphatidylinositol-3-kinase (PI3K)/AKT and glycogen synthase kinase 3ß (GSK3ß) pathways, and microRNA (miR)-17 level. Moreover, the effects of miR-17 inhibition on cell viability, migration, and activation of the PI3K/AKT and GSK3ß pathways in GC cells overexpressing NEAT1 were also explored. RESULTS: NEAT1 was up-regulated in GC tissues and cell lines. Then, cell viability and migration of GC cells were markedly increased by NEAT1 overexpression, while the cell invasion and apoptosis were unchanged. The phosphorylated level of PI3K, AKT, and GSK3ß were increased by NEAT1 overexpression. Subsequently, we found miR-17 level was positively correlated with NEAT1 expression, and NEAT1 functions through up-regulating miR-17. CONCLUSIONS: NEAT1 was up-regulated in GC tissues and cell lines. Its overexpression enhanced cell viability and migration through up-regulating miR-17, along with activation of the PI3K/AKT and GSK3ß pathways.

miR-494 inhibits invasion and proliferation of gastric cancer by targeting IGF-1R.

OBJECTIVE: The aim of this study was to investigate the target gene of miR-494 and its roles in tumor growth of gastric cancer (GC). PATIENTS AND METHODS: Expression of miR-494 was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR) in gastric cancer tissues and cell lines. Then, luciferase reporter assay was used to elucidate whether insulin-like growth factor 1 receptor (IGF1R) is a target gene of miR-494. Finally, the roles and mechanism of miR-494 in the regulation of tumor invasion were further investigated. RESULTS: Relative miR-494 level was found to be significantly lower in patients with GC than healthy controls (p < 0.01). Over-expression of miR-494 could inhibit gastric cancer cell proliferation, migration, and invasion in vitro. Furthermore, we demonstrated that miR-494 binds to the 3'-untranslated region (UTR) of IGF1R and inhibits the expression of the IGF1R protein. CONCLUSIONS: Our data showed that miR-494 acted as a tumor suppressor in GC.

Role of IL-8 gene polymorphisms in glioma development in a Chinese population.

This case-control study aimed to investigate the role of -251 T>A (rs4073) and -781 C>T (rs2227306) polymorphisms in the interleukin-8 (IL-8) gene in the development of glioma in a Chinese population. One hundred and twenty-seven glioma patients and 284 healthy control subjects were recruited to this study between February 2013 and December 2014. The IL-8 -251 T>A (rs4073) and -781 C>T (rs2227306) polymorphisms were genotyped by polymerase chain reaction coupled with restriction fragment length polymorphism. The patients and control subjects were comparable by gender (X2 = 1.24, P = 0.27), tobacco smoking status (X2 = 0.80, P = 0.37), alcohol consumption status (X2 = 0.97, P = 0.32), and family history of cancer (X2 = 1.54, P = 0.22). The age of glioma patients was statistically lower than that of control subjects (t = 2.87, P = 0.002). The chi-square test revealed the lack of any statistically significant differences in the genotype distributions of IL-8 rs4073 (X2 = 0.89, P = 0.64) and rs2227306 (X2 = 2.58, P = 0.28) between the glioma patients and control subjects. Unconditional logistic regression analysis revealed that the IL-8 rs4073 and rs2227306 gene polymorphisms did not contribute to the development of glioma. In conclusion, we determined that there is a lack of evidence suggesting a significant association between the IL-8 rs4073 and rs2227306 gene polymorphisms and the development of glioma in a Chinese population.

Correlation between non-metastatic protein 23 expression and clinicopathological features of colorectal cancer in Asians.

The current meta-analysis was performed to investigate the association between non-metastatic protein 23 (NM23) expression, tumor pathology, and disease prognosis in colorectal cancer (CRC) among Asians. English and Chinese language-based electronic databases (e.g., PubMed, EBSCO, Ovid, Springerlink, Wiley, Web of Science, Wanfang databases, China National Knowledge Infrastructure, VIP databases) were searched using search terms to identify published studies relevant to NM23 and CRC with immunohistochemistry. In total, 289 studies were identified through database searches, and 16 cohort studies (4 studies in English, 12 in Chinese) were chosen for meta-analysis, which included 1592 CRC patients. The results revealed that NM23 protein expression in CRC tissue was higher in patients with Dukes stages A and B than in patients with Dukes stages C and D. The NM23 protein was expressed at higher levels in well- and moderately differentiated tumors than in poorly differentiated tumors. The 5-year survival rate was also higher in CRC patients with NM23-positive tumors than in CRC patients with NM23-negative tumors. Significantly, 5-year tumor relapse and metastasis were lower in patients with NM23-positive tumors than in CRC patients with NM23-negative tumors. The findings suggest that NM23 expression status is associated with tumor aggressiveness and survival in CRC among Asians. Importantly, CRC patients with NM23-positive tumors had a better prognosis, and thus NM23 expression maybe used as a key prognostic indicator for CRC.

Preparation, characterisation and evaluation of curcumin with piperine-loaded cubosome nanoparticles.

OBJECTIVE: In this study, curcumin was designed into the nanoformulation called cubosome with piperine in order to improve oral bioavailability and tissue distribution of curcumin. METHODS: The characteristic of the cubosome was studied by using scanning electron microscope (SEM), Infrared spectrum and small angle X-ray scattering (SAXS) techniques. Tissue distribution of cubosome was measured by liquid chromatography-mass spectrometry (LC-MS) method in mice. RESULTS: The characteristic of the cubosome was demonstrated that the curcumin and piperine were encapsulated in the interior of the cubosome and the crystal form was Pn3m space. The pharmacokinetic test revealed that the cubosome could improve the oral bioavailability significantly compared to the suspension of curcumin with piperine and be mainly absorbed by the spleen. CONCLUSION: These findings provide the reference to a preferable choice of the curcumin formulation and contribute to therapeutic application in clinical research.