METTL14-mediated Bim mRNA m6A modification augments osimertinib sensitivity in EGFR-mutant NSCLC cells.

Resistance to osimertinib represents a significant challenge for the successful treatment of non-small cell lung cancer (NSCLC) harboring activating mutations in epidermal growth factor receptor (EGFR). N6-methyladenosine (m6A) on mRNAs is critical for various biological processes, yet whether m6A regulates osimertinib resistance of NSCLC remains unknown. In this study, we demonstrated that developing osimertinib-resistant phenotypes depends on m6A reduction resulting from downexpression of m6A methyltransferase METTL14 in EGFR-mutant NSCLCs. Both in vitro and in vivo assay showed that specific knockdown of METTL14 was sufficient to confer osimertinib resistance and elevated expression of METTL14 rescued the efficacy of osimertinib in the resistant NSCLC cells. Mechanistically, METTL14 promoted m6A methylation of pro-apoptotic Bim mRNA and increased Bim mRNA stability and expression, resulting in activating the Bim-dependent pro-apoptotic signaling and thereby promoting osimertinib-induced cell apoptosis. Analysis of clinical samples revealed that decreased expression of METTL14 was observed in osimertinib-resistant NSCLC tissues and significantly associated with a poor prognosis. In conclusion, our study reveals a novel regulatory mechanism by which METTL14-mediated m6A methylation of Bim mRNA inhibited osimertinib resistance of NSCLC cells. It offers more evidences for the involvement of m6A modification in regulation of osimertinib resistance, and provides potential therapeutic targets for novel approaches to overcome the tolerance of osimertinib and other EGFR-TKIs. Implications: This study offers more evidences for the involvement of METTL14-mediated m6A modification in regulation of osimertinib resistance, and provides potential therapeutic targets for novel approaches to overcome the tolerance of osimertinib and other EGFR-TKIs.

Resveratrol prevents Drp1-mediated mitochondrial fission in the diabetic kidney through the PDE4D/PKA pathway.

To explore the role of PDE4D in diabetic nephropathy (DN) and investigate whether resveratrol protects against DN via inhibiting PDE4D. Diabetic db/db mouse and glomerular mesangial cell line (GMCs) were used to investigate the role of PDE4D and the protective effect of resveratrol on renal fibrosis under high glucose (HG) environment. Resveratrol alleviated the progress of DN via inhibiting mitochondrial fragmentation and restoring the expression of PDE4D, PKA, phosphorylated Drp1-Ser637 and Drp1 in kidney of db/db mice. In HG-exposed GMCs, resveratrol treatment decreased the expression of PDE4D, increased PKA level, and inhibited Drp1-mediated mitochondrial fission. In contrast, PDE4D over-expression blunted the inhibitory effects of resveratrol on Drp1 expression and mitochondrial fission. Moreover, PKA inhibitor H89 blunted the effects of resveratrol on phosphorylated Drp1-Ser637 expression and mitochondrial fission in HG-treated GMCs. Inhibition of mitochondrial fission with Drp1 inhibitor Mdivi-1 alleviated mitochondrial dysfunction in GMCs under HG. These findings indicate PDE4D plays an important role in the process of DN. Resveratrol attenuates the development of DN by preventing mitochondrial fission through inhibiting PDE4D, which regulates the expression of phosphorylated Drp1-Ser637 directly.

Guanxinning injection ameliorates cardiac remodeling in HF mouse and 3D heart spheroid models via p38/FOS/MMP1-mediated inhibition of myocardial hypertrophy and fibrosis.

BACKGROUND: Heart failure (HF) is a cardiovascular disease with high morbidity and mortality. Guanxinning injection (GXNI) is clinically used for the treatment of coronary heart disease, but its therapeutic efficacy and potential mechanism for HF are poorly understood. This study aimed to evaluate the therapeutic potential of GXNI on HF, with a special focus on its role in myocardial remodeling. METHODS: 3D cardiac organoids and transverse aortic constriction (TAC) mouse models were established and utilized. Heart function and pathology were evaluated by echocardiography, hemodynamic examination, tail-cuff blood pressure and histopathology. Key targets and pathways regulated by GXNI in HF mouse heart were revealed via RNA-seq and network pharmacology analysis, and were verified by RT-PCR, Western blot, immunohistochemistry and immunofluorescence. RESULTS: GXNI significantly inhibited cardiac hypertrophy and cells death. It protected mitochondrial function in cardiac hypertrophic organoids and markedly improved cardiac function in HF mice. Analysis of GXNI-regulated genes in HF mouse hearts revealed that IL-17A signaling in fibroblasts and the corresponding p38/c-Fos/Mmp1 pathway prominently mediated cardiac. Altered expressions of c-Fos, p38 and Mmp1 by GXNI in heart tissues and in cardiac organoids were validated by RT-PCR, WB, IHC, and IF. H&E and Masson staining confirmed that GXNI substantially ameliorated myocardial hypertrophy and fibrosis in HF mice and in 3D organoids. CONCLUSION: GXNI inhibited cardiac fibrosis and hypertrophy mainly via down-regulating p38/c-Fos/Mmp1 pathway, thereby ameliorating cardiac remodeling in HF mice. Findings in this study provide a new strategy for the clinical application of GXNI in the treatment of heart failure.

Cip2a induces arginine biosynthesis and promotes tumor progression in non-small cell lung cancer.

Cancerous inhibitor of protein phosphatase 2A (Cip2a) is an oncoprotein, playing important roles in tumor progression. However, the underlying mechanisms by which Cip2a promotes tumor aggressiveness in NSCLC remain to be further investigated. In this study, we found that Cip2a expression is elevated in NSCLC and correlates with poor prognosis. Knockdown of Cip2a significantly reduced the ability of cell proliferation, invasion, and metastasis of NSCLC both in vitro and in vivo. Furthermore, we found that Cip2a promotes tumor progression partly by inducing arginine biosynthesis, and knockdown of Cip2a exhibited a significantly increased sensitivity to arginine deprivation and mTOR inhibition. In addition, we found that p53 mutants in NSCLC cells increased Cip2a expression by inhibiting the activity of wild-type p53. Our findings provide new insights into the mechanisms of Cip2a in promoting tumor progression and suggest that Cip2a represents a potential therapeutic target for treating NSCLC.

P21-activated kinase 2-mediated ß-catenin signaling promotes cancer stemness and osimertinib resistance in EGFR-mutant non-small-cell lung cancer.

Osimertinib (AZD9291) is a third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), used for treating patients with advanced non-small-cell lung cancer (NSCLC) harboring EGFR-activating mutations or the resistant T790M mutation. However, acquired resistance to osimertinib is inevitable in EGFR-mutant NSCLC. By employing a global mass spectrometry-based phosphoproteomics approach, we identified that the activated p21-activated kinase 2 (PAK2)/ß-catenin axis acts as a driver of osimertinib resistance. We found that PAK2 directly phosphorylates ß-catenin and increases the nuclear localization of ß-catenin, leading to the increased expression and transcriptional activity of ß-catenin, which in turn enhances cancer stem-like properties and osimertinib resistance. Moreover, we revealed that HER3 as an upstream regulator of PAK2, drives the activation of PAK2/ß-catenin pathways in osimertinib-resistant cells. The clinical relevance of these findings was further confirmed by examining tissue specimens from patients with EGFR-mutant NSCLC. The results demonstrated that the levels of HER3, phospho-PAK2 (p-PAK2) and ß-catenin in the tissues from patients with EGFR-mutant NSCLC, that had relapsed after treatment with osimertinib, were elevated compared to those of the corresponding untreated tissues. Additionally, the high levels of HER3, p-PAK2 and ß-catenin correlated with shorter progression-free survival (PFS) in patients with EGFR-TKI-treated NSCLC. We additionally observed that the suppression of PAK2 via knockdown or pharmacological targeting with PAK inhibitors markedly restored the response of osimertinib-resistant NSCLC cells to osimertinib both in vitro and in vivo. In conclusion, these results indicated that the PAK2-mediated activation of ß-catenin is important for osimertinib resistance and targeting the HER3/PAK2/ß-catenin pathway has potential therapeutic value in NSCLCs with acquired resistance to osimertinib.