Abstract 5662: The development of SCLC specific nanoparticle-mediated p53 gene therapy

Abstract

Abstract Lung cancer is the most common cause of cancer-related deaths worldwide. Small cell lung cancer (SCLC) is a highly malignant and aggressive type of lung cancer with widespread metastases and poor prognosis. SCLC is characterized by chemoresistance and radioresistance with an average survival period of <1 year. Clinically, over 70% of SCLC cases harbor mutations in two key tumor suppressor genes, p53 and Rb, whose deletion or inactivation is associated with SCLC pathogenesis. Therefore the reinstatement of wild-type (WT) p53 expression provides an attractive gene therapy strategy for SCLC. We investigated restoration of WT p53 in p53-mutant SCLC cells, using non-viral, poly(β-amino ester) (PBAE) nanoparticles for gene delivery. PBAE nanoparticles possess numerous advantages over conventional viral carriers such as low risk of insertional mutagenesis, low immunogenicity, increased cargo capacity, and capabilities for cell targeting. PBAEs encapsulate DNA to form biodegradable cationic nanoparticles and have been studied in a variety of cancer cell types and in primary endothelial cells. This is the first study of PBAE-mediated gene therapy in SCLC. Two out of 112 PBAEs initially screened, 456 and 457, were validated for gene expression efficiency using a green fluorescent protein (GFP)-based reporter in the H446 SCLC cell line using microscopy and flow cytometry. Both 456 and 457 exhibited transfection efficiency of >40%; comparable to commercially available transfection reagents. We initiated efficacy assays by validating whether H446 cells possess mutant p53 and are a suitable target for p53 restoration; p53 status in H446 and H460 cells (WT control) was confirmed by direct sequencing. A plasmid encoding WT p53-GFP under the control of the CMV promoter (CMV-p53-GFP) was used to reconstitute exogenous expression of WT p53. An 8 h incubation with 456:CMV-p53-GFP complex induced WT p53 expression in H446 cells (p53-mutant) but not in H460 cells (p53 WT); leading to a moderate p21 induction, subsequent G1-arrest and anti-proliferative effects. A CMV-p53-GFP plasmid without p53 (CMV-GFP) served as a negative control for induction and activity of exogenous p53. Additionally, 456 and 457 exhibited in vivo tumoral transfection of CMV-LUC DNA after an intratumoral or an intravenous injection. These novel in vitro and in vivo effects by PBAEs in H446 cells suggested a possible therapeutic potential for targeted p53 gene therapy in other SCLC cells and mouse models with primary SCLC xenografts. Ongoing studies include genetic characterization of patient-derived primary SCLC xenografts and assessing systemic PBAE-p53 delivery in these xenografts. In parallel, we plan to employ PBAE-p53 delivery in combination with other target genes or with chemo-/radio-therapy in SCLC cell lines and mouse models. We strongly believe that PBAE-based gene therapy would provide higher efficacy and minimal adverse effects for improved SCLC treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5662. doi:1538-7445.AM2012-5662