Gene Delivery of RNA Therapeutics for the Treatment of Brain Tumors

Abstract

The proliferation of glioblastoma multiforme (GBM) is often coupled to the dysregulation, amplification and overexpression of tyrosine kinase receptors (TKRs) including epidermal growth factor receptor (EGFR) and vascular endothelial growth factor receptor 2 (VEGFR2) among others. TKRs are cell‐surface receptors that bind to their cognate ligand, peptide hormones known as growth factors. Binding of the growth factor ligand stimulates TKRs, activating a signal‐transduction cascade that leads to cell growth, migration and the formation of tumor vasculature. A look at TKRs reveals a general architecture of the receptor. They are comprised of three distinct domains: an extracellular, transmembrane and an intracellular domain. As these receptors are often upregulated in GBM, an RNA Therapy to modulate the expression, inhibit the translation of critical TKR domains or reduce its activation would reduce the functionality of these oncogenic transcripts and proteins in GBM. Our research focuses on the development of AAV gene therapy vectors that encode novel RNA therapeutics to reduce the expression of oncogenic transcripts as well as inhibit their activation in GBM, the most common malignant primary brain tumor in adults. Individuals diagnosed with GBM have a short life expectancy of 12–15 months. The standard care and treatment as well as the developmental pipeline of new therapeutic strategies are often limited by the blood‐brain barrier (BBB), restricting systemically administered therapies from reaching the brain (Hicks et al., 2015). Thus, a novel strategy that circumvents this barrier is necessary to effectively reach the CNS tumor microenvironment. Direct AAV delivery of genes that encode RNA therapeutics at the time of tumor resection would bypass the BBB to deliver a persistent and constant dosage from the inside‐out. With the advancement of nanopore sequencing technology, we are able to read longer transcripts (>200kb). With this technology, we are able to examine the architecture of nascent pre‐mRNA transcripts, detect lower abundant alternatively spliced and polyadenylated isoforms, as well take advantage of RNA structurome analysis to reveal RNA elements within TKRs which are more susceptible to RNA anti‐sense directed therapies. To this end, we have developed a strategy to identify novel targets within oncogenic transcripts and incorporate these therapies directed against these targets into our AAV directed RNA gene therapy platforms. In addition, we use RNA mimics to take advantage of the RNA interference pathway to modulate and reduce expression of oncogenic transcripts. This research has the potential to identify novel RNA elements and alternatively spliced and polyadenylated isoforms of TKRs in health and disease. The ultimate goal of our research project is to build on current strategies to identify even better RNA elements of the EGFR and VEGFR2 transcripts which are structurally available for both microRNA degradation and anti‐sense directed alternative splicing therapy.