Co-mutations of epidermal growth factor receptor and BRAF in Chinese non-small cell lung cancer patients.

BACKGROUND: Epidermal growth factor receptor (EGFR) and BRAF are 2 driver genes in non-small cell lung cancer (NSCLC) which are normally mutually exclusive. It has been previously reported that the existence of BRAF V600E in EGFR-mutated NSCLC patients could cause resistance to EGFR tyrosine kinase inhibitors (TKIs), but the influence of other BRAF actionable mutations on resistance to EGFR-TKIs has not yet been investigated. Understanding the coexistence of EGFR and BRAF actionable mutations in Chinese NSCLC patients may be essential for further treatment and prognostic prediction. METHODS: A total of 127 Chinese NSCLC patients harboring EGFR and BRAF co-mutations were enrolled in this study. We analyzed the mutation profiles of these patients through next-generation sequencing (NGS). We explored the associations between somatic mutations and patient characteristics, including tumor stage and age, among others. RESULTS: The frequency of EGFR and BRAF co-mutation was 0.91% in Chinese NSCLC patients, compared with 0.97% in Western NSCLC patients (cBioPortal). Among the 127 patients with both EGFR and BRAF mutations, 93 of them harbored clinically significant mutations. The remaining 34 patients were found to have mutations of uncertain significance of either EGFR or BRAF. TP53 was the most frequently mutated gene in BRAF and EGFR co-mutation patients, accounting for around 58% (N=54/93). MET active mutations (amplification and exon 14 skipping) accounted for 12% (N=11/93). Approximately 18% of patients (N=17/93) with significant EGFR mutations were detected to have fusions/rearrangements of the BRAF gene. BRAF fusion was more likely detected in EGFR exon19del patients compared with non-exon19del patients (P value =0.015). In addition, EGFR T790M, the most TKI-resistant mutation, was not found in any patient with BRAF fusion/rearrangement. CONCLUSIONS: This study is the first to show different subtypes of EGFR and BRAF co-mutations in Chinese NSCLC patients. The prognosis of EGFR-TKI treatment may vary according to different BRAF actionable mutations. Aside from BRAF V600E, class II/III and BRAF fusions were found, which provides clues for investigating the resistance mechanisms of EGFR-TKIs in the future.

Bufadienolides induce p53-mediated apoptosis in esophageal squamous cell carcinoma cells in vitro and in vivo.

Bufadienolides are a type of cardiotonic steroids isolated from the skin and parotid venom glands of the toad Bufo bufo gargarizans Cantor, and exhibit wide-spectrum anticancer activities. However, the effects and mechanisms of bufadienolides on esophageal squamous cell carcinoma (ESCC) cells remain unknown. In the present study, the anticancer activities of two bufadienolides, bufotalin and bufalin, were examined in vitro and in vivo. The results demonstrated that bufotalin and bufalin effectively inhibited the viability of ESCC cells, with half-maximal inhibitory concentration (IC50) values of 0.8-3.6 µM. However, bufotalin and bufalin exhibited lower toxicity towards Het-1A human esophageal squamous cells, indicating their high selectivity towards cancer cells. Mechanistic studies revealed that bufotalin effectively induced ESCC cell apoptosis, as characterized by DNA fragmentation and nuclear condensation, which was primarily mediated through activation of caspase family members. In addition, treatment of ESCC cells with bufotalin markedly activated tumor protein p53 (p53) phosphorylation. Transfection of cells with p53 small interfering RNA markedly inhibited bufotalin-induced p53 phosphorylation and significantly inhibited bufotalin-induced cell apoptosis. Furthermore, bufotalin demonstrated in vivo anticancer efficacy in a tumor-bearing nude mice model, where bufotalin effectively inhibited Eca-109 xenograft tumor growth in a time- and dose-dependent manner, through activation of the p53 signaling pathway. Collectively, the results from the present study suggested that bufadienolides exert anticancer effects against ESCC by regulating the p53 signaling pathway.

Arenobufagin activates p53 to trigger esophageal squamous cell carcinoma cell apoptosis in vitro and in vivo.

Esophageal squamous cell carcinoma (ESCC) is often diagnosed at late incurable stage and lacks effective treatment strategy. Bufadienolides are cardiotonic steroids isolated from the skin and parotid venom glands of the toad Bufo bufo gargarizans Cantor with novel anticancer activity. However, there is little information about the effects and action mechanisms of bufadienolides on ESCC cells. In this study, the in vitro and in vivo anti-ESCC activities of bufadienolides, including bufalin (Bu) and arenobufagin (ArBu), were examined and the underlying molecular mechanisms were elucidated. The results showed that ArBu exhibited higher anticancer efficacy than Bu against a panel of five ESCC cells, with IC50 values ranging from 0.8 µM to 3.6 µM. However, ArBu showed lower toxicity toward Het-1A human normal esophageal squamous cells, indicating its great selectivity between cancer and normal cells. Moreover, ArBu effectively induced ESCC cell apoptosis mainly by triggering caspase activation through intrinsic and extrinsic pathways. Treatment of ESCC cells also significantly activated p53 signaling by enhancing its phosphorylation. Interestingly, transfection of cells with p53 small interfering RNA significantly inhibited the ArBu-induced p53 phosphorylation and the overall apoptotic cell death. Furthermore, ArBu also demonstrated novel in vivo anticancer efficacy by inhibiting the tumor growth through activation of p53 pathway. Taken together, these results demonstrate the p53-targeting therapeutic potential of bufadienolides against ESCC.