Abstract 5210: Creation of endothelial-targeted adenoviral vectors for genetic engineering of the metastatic tumor microenvironment

Abstract

Abstract The endothelium is an attractive gene therapy target for metastatic cancer, providing access to systemic tumors. Previous work has focused on either viral entry/attachment (transductional targeting), or cell type specific enhancer promoter vector transgene expression regulation (transcriptional targeting). While previous studies demonstrated endothelial targeting, the tumor wide extent and transgene expression level quantification have been uncertain. Indeed, obtaining robust and stringent tumor expression has been challenging due to: insufficient transduction, focal rather than widespread intratumoral vascular expression, viral particle liver sequestration, and host preformed immunity. We developed a set of adenoviral vectors, containing 3 kb of the human ROBO4 enhancer/promoter, that are expressed at high levels throughout tumor vasculature. We now have three vectors representing four tiers of genetic modification. Transgenes encoding fluorescent proteins allowed us to quantify endothelial expression frequency by image analysis, and expression level using whole tissue Western blotting. Our first vector was Ad5.ROBO4. Compared with Ad5.CMV, Ad5.ROBO4 evidenced complete retargeting from hepatocytes to low-level expression in host organ and tumor endothelial cells. Warfarin-mediated hepatocyte detargeting produced a striking increase in tumor and bone marrow sinusoidal endothelial expression, without change in other host organs. Our second vector is Ad.RGD.H5/H3.ROBO4. Insertion of a cyclized RGD peptide in the fiber/knob HI loop engages enhanced tumor endothelial transduction via αv/β3 and αv/β5. Swapping hexon hypervariable regions responsible for Ad Type 5-Factor X binding with corresponding regions from serotype 3 enables marked diminution of hepatocyte sequestration. Use of the ROBO4 enhancer promoter produces high-level pan-intratumoral vascular expression particularly in cancers with markedly elevated VEGF production, such as orthotopic and metastatic renal cell carcinoma, orthotopic colonic liver metastases, and prostate cancer bone, brain, and liver metastases. Most striking, was enhancement of Ad.RGD.H5/H3.ROBO4 vector expression within interior tumor regions undergoing hypoxic necrosis; regions notoriously resistant to radiation or chemotherapies. Our third vector contains a polycistronic array, enabling triple transgene expression from a single vector within the tumor vasculature. Thus, we have panel of endothelial-targeted vectors with distinctive vascular tropism for use in cancers metastatic to individual host organs. These vectors enable expression of a palette of transgenes that can manipulate the tumor microenvironment to achieve metastatic growth inhibition alone, or “staggered” with chemo- or irradiation therapies. Note: This abstract was not presented at the meeting. Citation Format: Zhi Hong Lu, Sergey Kaliberov, Lyudmila Kaliberova, Rebecca E. Sohn, Yingqui Du, David T. Curiel, Jeffrey M. Arbeit. Creation of endothelial-targeted adenoviral vectors for genetic engineering of the metastatic tumor microenvironment. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5210. doi:10.1158/1538-7445.AM2015-5210