(615e) A Novel Application of Nanofiber Scaffolds for Gene Therapy

Electrospun nanofiber scaffolds have been used in cell growth and tissue engineering. Here we expand the application of nanofiber scaffolds to the genetic technique known as gene editing. Gene editing is a process by which single base mutations can be corrected, in the context of the chromosome, using a short piece of single stranded DNA in the form of an oligonucleotides (ssODN). The ssODNs are introduced into the cell to correct the mutant nucleotide using the inherent pathways of DNA replication and repair. This technique is being developed rapidly for use in the treatment of inherited diseases such as Sickle Cell Disease (SCD) and Phenylketonuria (PKU). The survival and proliferation of the cells bearing modified genes however appear to be impeded by a phenomenon known as reduced proliferation phenotype (RPP); this is the key barrier to practical implementation of gene editing in the clinical setting. To overcome RPP, we have utilized nanofiber scaffolds as templates on which modified cells were allowed to recover, grow and expand after gene editing. Here, we present evidence that HCT116-19, bearing an integrated, mutated eGFP gene and corrected by gene editing, proliferate on polylysine or fibronectin coated polycaprolactone (PCL) nanofiber scaffolds. In contrast, cells from the same reaction protocol plated on both regular plain surfaces and polylysine (or fibronectin) coated plate surfaces do not proliferate. Therefore, growing genetically modified (edited) cells on electrospun nanofiber scaffolds promotes the reversal of the Reduced Proliferation Phenotype and increases the potential of gene editing as an ex vivo gene therapy application.