Suicide Gene Approaches Using RA-Responsive Promoters to Drive DTA Gene, and Delivery to Neuroblastoma Cells with Integrin Receptor Targeted Nanoparticles

Abstract

Incidences of neuroblastoma are approximately 10.5% and are the cause of up 15% of mortality rates among children with cancer. The treatment of neuroblastoma is difficult with low survival rates of 40-50% due to the amplification of the MYCN oncogene leading to poor survival rates. It is in this backdrop that the current study aimed to examine the suicide gene approaches using RA- responsive promoters to drive the DTA gene, and delivery to neuroblastoma cells with Integrin receptor-targeted nanoparticles. Specifically, the study sought to develop a proof of principle that the DTA suicide gene can be made ATRA-responsive and used to selectively kill neuroblastoma tumor cells by cloning the DTA gene downstream of a RARE promoter. In this primarily study, the first aim was to demonstrate the viability of the DTA gene as a suicide gene that can be transcriptionally activated by ARTA in the neuroblastoma cells. The second aim was to assess the inducibility of the CYP26 promoter in response to ATRA into Neuroblastoma cells. The final objective was to assess the effectiveness of delivering these plasmid vectors to neuroblastoma cells using Integrin receptor-targeted nanoparticles. The results showed that CYP26 was successfully inducible in the neuroblastoma cells after incubating with 100 and 1000 nM ATRA. Moreover, the RARE-DTA plasmid was cloned successfully using Gibson assembly method and was inducible in Neuroblastoma cells by ATRA. However, the IRTN delivery system was not found to be successful in transducing plasmids into the cells compared to the lipofectamine 2000 reagent. Study findings were used to draw the conclusion that RA-responsive promoters such as CYP26 can successfully drive the DTA gene into Neuroblastoma cells.