Targeting proteasomal deubiquitinases USP14 and UCHL5 with b-AP15 reduces 5-fluorouracil resistance in colorectal cancer cells.

5-Fluorouracil (5-FU) is the first-line treatment for colorectal cancer (CRC) patients, but the development of acquired resistance to 5-FU remains a big challenge. Deubiquitinases play a key role in the protein degradation pathway, which is involved in cancer development and chemotherapy resistance. In this study, we investigated the effects of targeted inhibition of the proteasomal deubiquitinases USP14 and UCHL5 on the development of CRC and resistance to 5-FU. By analyzing GEO datasets, we found that the mRNA expression levels of USP14 and UCHL5 in CRC tissues were significantly increased, and negatively correlated with the survival of CRC patients. Knockdown of both USP14 and UCHL5 led to increased 5-FU sensitivity in 5-FU-resistant CRC cell lines (RKO-R and HCT-15R), whereas overexpression of USP14 and UCHL5 in 5-FU-sensitive CRC cells decreased 5-FU sensitivity. B-AP15, a specific inhibitor of USP14 and UCHL5, (1-5 µM) dose-dependently inhibited the viability of RKO, RKO-R, HCT-15, and HCT-15R cells. Furthermore, treatment with b-AP15 reduced the malignant phenotype of CRC cells including cell proliferation and migration, and induced cell death in both 5-FU-sensitive and 5-FU-resistant CRC cells by impairing proteasome function and increasing reactive oxygen species (ROS) production. In addition, b-AP15 inhibited the activation of NF-κB pathway, suppressing cell proliferation. In 5-FU-sensitive and 5-FU-resistant CRC xenografts nude mice, administration of b-AP15 (8 mg·kg-1·d-1, intraperitoneal injection) effectively suppressed the growth of both types of tumors. These results demonstrate that USP14 and UCHL5 play an important role in the development of CRC and resistance to 5-FU. Targeting USP14 and UCHL5 with b-AP15 may represent a promising therapeutic strategy for the treatment of CRC.

The role of bile acid in intestinal metaplasia.

A precancerous lesion of gastric cancer (GC), intestinal metaplasia (IM) is a pathological transformation of non-intestinal epithelium into an intestinal-like mucosa. It greatly raises the risk of developing the intestinal type of GC, which is frequently observed in the stomach and esophagus. It is understood that esophageal adenocarcinoma's precursor lesion, chronic gastroesophageal reflux disease (GERD), is what causes Barrett's esophagus (BE), an acquired condition. Recently, Bile acids (BAs), which are one of the compositions of gastric and duodenal contents, have been confirmed that it led to the occurrence and development of BE and gastric intestinal metaplasia (GIM). The objective of the current review is to discuss the mechanism of IM induced by bile acids. This review serves as a foundation for further research aimed at improving the way BE and GIM are currently managed.

[Anti-gastric cancer effects of Z Ajoene and its molecular mechanisms].

Objective: To investigate the effects of Z Ajoene on gastric cancer cell MGC-803 and its molecular mechanisms. Methods: The gastric cancer cells MGC-803 were treated with 0, 1, 5, 25 and 125 µmol/L Z Ajoene for 24 h, 48 h and 72 h, each with 3 replicate wells. The proliferation activity of MGC-803 cells was analyzed by MTS method, mitochondrial membrane potential was analyzed after JC-1 staining, nuclear type was observed after Hoechst 33342 staining, cytotoxicity was detected by LDH release method, and the apoptosis level and cell cycle were analyzed with flow cytometry. RT-qPCR and Western blot methods were used to evaluate the expression levels of P53, Caspase-3, RAS, ERK, BCL-2, AKT, mTOR and PI3K genes. At the same time, 4-week-old male BALB/C mice were randomly divided into 5 groups, 20 per group, and were subcutaneously inoculated with gastric cancer cell MGC-803 in the groin. Two days later, each group was injected with Z Ajoene at the doses of 0, 1, 5, 25 and 125 µmol/L, 0.1 ml/time, and was injected every other day. On the 20th day of the first injection of tumor cells, 10 mice in each group were killed, the tumor tissues were taken out and weighed. The survival period of the remaining mice was recorded and the effects of Z Ajoene on the growth and survival period of gastric cancer in tumor-bearing mice were observed. Results: After Z Ajoene treatment, the proliferation activity of MGC-803 cells was significantly inhibited and the apoptosis rate was significantly increased(P＜0.01). The transcription and expression levels of p53, Caspase-3 and BAX genes were significantly increased, while the transcription and expression levels of RAS, ERK1, BCL-2, AKT, mTOR and PI3K genes were decreased markedly(P＜0.01). The tumor inhibition experiments showed that the growth of the tumor could be inhibited and the survival time of the tumor-bearing animals could be greatly prolonged after Z Ajoene treatment(P＜0.01). Conclusion: Z Ajoene has therapeutic effects on gastric cancer, can inhibit the proliferation of gastric cancer cells and induce them apoptosis by regulating the expression of PI3K-AKT-mTOR and RAS-RAF-MEK-ERK signal pathways.

[The effects of tectochrysin on prostate cancer cells apoptosis and its mechanism].

OBJECTIVE: To investigate the effects of tectochrysin on prostate cancer cell line 22Rv.1 and reveal its molecular mechanism. METHODS: Tectochrysin at the concentrations of 0～20 µg/ml was applied to 22Rv.1 cells and normal prostate cell RWPE-1. The proliferation activity of the cells was detected by MTS assay. Flow cytometry and hoechst 33342 staining were used to analyze the effects of drugs on cell apoptosis, death, cell cycle and nuclear type changes. LDH release test was used to analyze the cytotoxicity of the drug to 22Rv.1 cells. QPCR and Western blot were used to analyze the effects of the drug on the expressions of genes in 22Rv.1 cells. Finally, the tumor inhibited effect of the drug on the bearing tumor BALB/c mice were confirmed though anti-tumor experiment. RESULTS: Tectochrysin could significantly inhibit the proliferation activity of 22Rv.1 cells and induced their apoptosis, and promoted the expressions of genes dr4, dr5, trail, p53, caspase-3, caspase-8, caspase-9, bid, bax and foxo3, inhibited the expressions of anti-apoptotic genes akt, pi3k and bcl-2. CONCLUSION: Tectochrysin can induce prostate cancer cells apoptosis through affecting TRAIL and PI3K/AKT signaling pathways, and has anti-prostate cancer effect.

[Anti-hepatoma effects of Smac analogue Birinapant and its related molecular mechanism].

OBJECTIVE: To investigate the effects of Birinapant on hepatocellular carcinoma cells and its related molecular mechanisms. METHODS: Human hepatocellular carcinoma cells QGY-7701 were treated with 0, 1, 5, 25 and 125 nmol/L Birinapant for 24, 48 and 72 hours respectively, each experiment 3 wells.The proliferation activity of cells, the apoptosis levels, the cells nuclear type, the mitochondrial membrane potential, the transcription and expression levels of genes and the cytotoxicity of Birinapant were analyzed.At the same time, 4-week-old male BALB/C mice were randomly divided into 5 groups, with 20 mice in each group.The mice were inguinal injected with QGY-7701 cells, and then subcutaneous injected with Birinapant (concentrations ranging from 0, 1, 5, 25, 125 µg/kg) in each group after two days, once every other day.On 18th day since first Birinapant injection, 10 mice were killed in each group to weigh tumor tissue and survival time was recorded from the remaining 10 mice.The effects of Birinapant on the growth of the tumor and the survival time of tumor-bearing mice were observed. RESULTS: Compared with the negative control (NC) group, the proliferation activity of QGY-7701 was inhibited significantly after Birinapant treatment and the apoptosis levels were increased significantly (P<0.01).The cell mitochondrial membrane potential was decreased and the karyotype was changed (P<0.01).At the same time, the transcription and expression levels of genes cellular inhibitor of apoptosis protein 1(cIAP-1), cellular inhibitor of apoptosis protein 2(cIAP-2), ras, raf, mek and erk were significantly decreased (P<0.01), while the expression levels of caspase-3 and caspase-9 genes were up-regulated (P<0.01).Compared with the model group (MG), the growth of the tumor was inhibited significantly and the survival time of the tumor-bearing mice was prolonged after Birinapant treatment (P<0.01). CONCLUSIONS: Birinapant can inhibit the expression of cIAP-1, cIAP-2 and the proteins of Ras-Raf-MEK-ERK signal pathways, so as to activate the mitochondria mediated endogenous apoptosis pathway.Birinapant shows a certain inhibitory effect on liver cancer.