Abstract / Summary
Cisplatin (CDDP) resistance reduces the treatment effect in non-small cell lung cancer. Lung cancer stem-like cells are closely linked to drug tolerance. Mitochondrial dysfunction may affect their stemness and response to treatment. This study examined whether peptide nucleic acid conjugated with triphenylphosphonium (PNA-TPP) could alter the response of A549-derived stem-like spheres to cisplatin. We also explored whether this effect was associated with Drp1 (dynamin-related protein 1)-related mitochondrial changes. A549-derived stem-like spheres were established by single-cell cloning and serum-free suspension culture. PNA-TPP was designed to target the mitochondrial DNA D-loop region and was used at 10 nmol/mL for 24 h. Cell viability, colony formation, sphere formation, apoptosis, migration, and invasion were assessed using CCK-8, colony formation, sphere formation, Annexin V-FITC/PI flow cytometry, wound healing, and Transwell assays. Mitochondrial membrane potential, reactive oxygen species (ROS), and ATP levels were detected using JC-1 staining, DCFH-DA flow cytometry combined with DHE fluorescence staining, and ELISA, respectively. Drp1 mRNA expression was examined by qRT-PCR, and Drp1 protein levels in the mitochondrial fraction were assessed by Western blotting. Drp1 overexpression and shRNA-mediated knockdown were used to evaluate its involvement. PNA-TPP increased Drp1 mRNA expression and mitochondrial Drp1 levels. PNA-TPP reduced mitochondrial membrane potential and ATP levels, increased ROS levels, and decreased sphere-forming ability and octamer-binding transcription factor 4 (OCT4), SRY-box transcription factor 2 (SOX2), and Nanog homeobox (NANOG) expression. Compared with CDDP treatment alone, PNA-TPP pretreatment plus CDDP resulted in lower cell viability, colony formation, migration, and invasion, together with increased apoptosis and a higher cleaved caspase-9/total caspase-9 ratio. Drp1 overexpression partly mimicked these effects, whereas Drp1 knockdown partially restored mitochondrial membrane potential and stemness-related phenotypes after PNA-TPP treatment. PNA-TPP treatment was associated with Drp1-related mitochondrial dysfunction, weakened stem-like phenotypes, and an enhanced response to cisplatin treatment in A549-derived stem-like spheres. Not applicable.