Mice received 200 mL of subcutaneous transplants(1x10^7 cells of PC-9-Gef). On day 14 post implantation, the mice were randomly divided into four groups (n = 5 per group): (1) control miRNA + control siRNA, (2) control miRNA + siNNMT, (3) control siRNA + miR-449a, and (4) miR-449a + siNNMT treated with multipoint intratumoral injection (10 mg per 100 mL per tumor for siRNAs two times per week and 20 mg per 100 mL per tumor for miRNAs three times per week) of these nucleic acids complexed with in vivo-jetPEI in 5% glucose for 3 weeks.

Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) are used clinically as target therapies for lung cancer patients, but the occurrence of acquired drug resistance limits their efficacy. Nicotinamide N-methyltransferase (NNMT), a cancer-associated metabolic enzyme, is commonly overexpressed in various human tumors. Emerging evidence also suggests a crucial loss of function of microRNAs (miRNAs) in modulating tumor progression in response to standard therapies. However, their precise roles in regulating the development of drug-resistant tumorigenesis are still poorly understood. Herein, we established EGFR-TKI-resistant non-small-cell lung cancer (NSCLC) models and observed a negative correlation between the expression levels of NNMT and miR-449a in tumor cells. Additionally, knockdown of NNMT suppressed p-Akt and tumorigenesis, while re-expression of miR-449a induced phosphatase and tensin homolog (PTEN), and inhibited tumor growth. Furthermore, yuanhuadine, an antitumor agent, significantly upregulated miR-449a levels while critically suppressing NNMT expression. These findings suggest a novel therapeutic approach for overcoming EGFR-TKI resistance to NSCLC treatment.