ABSTRACT
Mutations in KRAS and p53 signaling pathways contribute to loss of responsiveness to current therapies and a decreased survival in lung cancer. In this study, we have investigated the delivery and transfection of wild-type (wt-) p53 and microRNA-125b (miR-125b) expressing plasmid DNA, in SK-LU-1 human lung adenocarcinoma cells as well as in Kras(G12D)/p53(fl/fl) (KP) genetically engineered mouse model of lung cancer. Systemic plasmid DNA delivery with dual CD44/EGFR-targeted hyaluronic acid (HA)-based nanoparticles (NPs) resulted in a 2- to 20-fold increase in wt-p53 and miR-125b gene expression in SK-LU-1 cells. This resulted in enhanced apoptotic activity as seen with increased APAF-1 and caspase-3 gene expression. Similarly, in vivo evaluations in KP mouse model indicated successful CD44/EGFR-targeted delivery. Tumor growth inhibition and apoptotic induction were also observed with (wt-p53+miR125b) combination therapy in KP tumor model. Lastly, J774.A1 murine macrophages co-cultured with transfected SK-LU-1 cells showed a 14- to 35-fold increase in the iNOS-Arg-1 ratio, supportive of previous results demonstrating a role of miR-125b in macrophage repolarization. Overall, these results show tremendous promise of wt-p53 and miR-125b gene therapy using dual CD44/EGFR-targeting HA NP vector for effective treatment of lung cancer.
Subject(s)
Hyaluronic Acid/administration & dosage , Lung Neoplasms/therapy , MicroRNAs/genetics , Proto-Oncogene Proteins p21(ras)/genetics , Tumor Suppressor Protein p53/genetics , Animals , Apoptosis , Cell Line, Tumor , Cell Survival/drug effects , Combined Modality Therapy , ErbB Receptors/metabolism , Genetic Engineering , Genetic Therapy , Humans , Hyaluronan Receptors/metabolism , Hyaluronic Acid/chemistry , Lung Neoplasms/genetics , Mice , Nanoparticles/chemistry , Neoplasms, Experimental , Plasmids/genetics , TransfectionABSTRACT
Tumor multidrug resistance (MDR) is a serious clinical challenge that significantly limits the effectiveness of cytotoxic chemotherapy. As such, complementary therapeutic strategies are being explored to prevent relapse. The altered metabolic state of cancer cells, which perform aerobic glycolysis, represents an interesting target that can enable discrimination between healthy cells and cancer cells. We hypothesized that cosilencing of genes responsible for aerobic glycolysis and for MDR would have synergistic antitumor effect. In this study, siRNA duplexes against pyruvate kinase M2 and multidrug resistance gene-1 were encapsulated in hyaluronic acid-based self-assembling nanoparticles. The particles were characterized for morphology, size, charge, encapsulation efficiency, and transfection efficiency. In vivo studies included biodistribution assessment, gene knockdown confirmation, therapeutic efficacy, and safety analysis. The benefit of active targeting of cancer cells was confirmed by modifying the particles' surface with a peptide targeted to epidermal growth factor receptor, which is overexpressed on the membranes of the SKOV-3 cancer cells. To augment the studies involving transplantation of a paclitaxel-resistant cell line, an in vivo paclitaxel resistance model was developed by injecting repeated doses of paclitaxel following tumor inoculation. The nanoparticles accumulated significantly in the tumors, hindering tumor volume doubling time (P < 0.05) upon combination therapy in both the wild-type (2-fold) and resistant (8-fold) xenograft models. Although previous studies indicated that silencing of MDR-1 alone sensitized MDR ovarian cancer to paclitaxel only modestly, these data suggest that concurrent silencing of PKM-2 improves the efficacy of paclitaxel against MDR ovarian cancer.