Nanocarrier-enabled restoration of miR-34a by Ailanthus excelsa Roxb. phytochemicals directly target NOTCH1 signaling to suppress cancer stemness in triple-negative breast cancer

Abstract

Triple-negative breast cancer (TNBC) is a highly aggressive breast cancer subtype associated with poor prognosis, metastasis, and limited targeted therapies. This study developed a folate-functionalized chitosan nanoparticle system targeting the miR-34a/NOTCH1 signaling axis and evaluated its anticancer potential using in vitro, in vivo, and bioinformatics approaches. Nanoparticles were synthesized by ionic gelation and characterized for size, surface charge, polydispersity, and drug-release behavior. Tumor-targeting efficiency was enhanced through folic acid functionalization. Biological effects were assessed in MDA-MB-231 cells using viability, migration, invasion, and tumorsphere assays. Mechanistic studies included dual-luciferase reporter assays, rescue experiments, transcriptomic profiling, and TCGA-based bioinformatics analyses. Antitumor efficacy, pharmacokinetics, biodistribution, toxicity, and survival outcomes were evaluated in vivo. The nanoparticles exhibited nanoscale size, low polydispersity, positive zeta potential, and sustained drug release. Treatment significantly upregulated miR-34a and suppressed NOTCH1 signaling. Transcriptomic analysis revealed downregulation of epithelial–mesenchymal transition, stemness, and tumor progression pathways. Functionally, nanoparticle treatment reduced TNBC cell viability, migration, invasion, and tumorsphere formation. In vivo, the formulation inhibited tumor growth, reduced metastatic burden, prolonged survival, and showed minimal systemic toxicity relative to conventional chemotherapy. Pharmacokinetic studies demonstrated prolonged circulation and preferential tumor accumulation. Combination therapy with doxorubicin produced synergistic anticancer effects. TCGA analysis confirmed an inverse correlation between miR-34a and NOTCH1 and supported their prognostic relevance in TNBC. Folate-functionalized chitosan nanoparticles targeting the miR-34a/NOTCH1 axis represent a promising nanotherapeutic strategy for suppressing TNBC progression and metastasis while improving therapeutic efficacy and safety.