Targeting HDAC2 with bruceine D epigenetically restores SMAD3 expression via H4K16ac enrichment to suppress malignant progression in EGFR-TKI-resistant lung adenocarcinoma.

ETHNOPHARMACOLOGICAL RELEVANCE: Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) are a cornerstone therapy for lung adenocarcinoma (LUAD), but their efficacy is often limited by primary or acquired resistance. Natural bioactive compounds such as bruceine D (BD) may offer alternative strategies for patients with EGFR-TKI resistance; however, the mechanism underlying BD's effects in resistant LUAD remains unclear.

AIM OF THE STUDY: This study aimed to determine whether BD suppresses malignant phenotypes of primary and acquired EGFR-TKI-resistant LUAD cells and to elucidate the shared molecular mechanisms.

MATERIALS AND

METHODS

Using H1975 (primary resistant) and PC-9/OR (acquired resistant) cells, cytotoxicity assays determined optimal BD concentration. Antitumor effects were evaluated in vitro via functional assays and in vivo using a xenograft mouse model. Integrative approaches including network pharmacology, Mendelian randomization, molecular docking, and dynamics simulations identified potential targets, with functional rescue experiments. CPTAC, Western blot, mRNA-seq, and CUT&Tag explored epigenetic mechanisms, validated by rescue.

RESULTS

BD suppressed proliferation, migration, invasion, and induced intrinsic apoptosis in both resistant cells in vitro and in vivo. HDAC2 was identified as a direct binding target. BD binding to HDAC2 increased H4K16ac enrichment at the SMAD3 promoter, upregulating SMAD3. Rescue experiments confirmed that the HDAC2/H4K16ac/SMAD3 axis mediates the antitumor effects of BD. Clinical samples and database analyses suggested diagnostic, prognostic, and therapeutic potential for H4K16ac, HDAC2, and SMAD3.

CONCLUSION

This study reveals for the first time a common mechanism by which the BD/HDAC2/H4K16ac/SMAD3 axis regulates malignant phenotypes in EGFR-TKI-resistant LUAD, offering a novel natural compound-based therapy.