Mechanisms Conferring Resistance to Pro-Apoptotic Cancer Gene Therapy

Recently, we have described a novel approach to the treatment of cancer that employs a series of vectors that encode surface expressed chimeric proteins in which the cytoplasmic death domain of Fas is fused in-frame to the extracellular domain of one of a number of cell surface receptors that recognize and bind various ligands that are differentially expressed within the tumor microenvironment. Although the majority of tumor cells transduced with such vectors are killed in the presence of the corresponding cognate ligand, a small percentage survive and in vivo may go on to repopulate a treated tumor. In order to understand the mechanisms employed by tumors to escape the cytotoxic effects of pro-apoptotic signals triggered via Fas, we isolated a large number of 293 tumor cell clones that survive following transfection with a plasmid vector encoding Flk-1/Fas, a chimeric receptor that induces tumor cell death in the presence of the pro-angiogenic cytokine VEGF. Characterization of Flk-1/Fas-positive clones revealed that while survival can most often be attributed simply to the down-regulation of VEGF ligand expression, in cells that express both receptor and ligand, other proteins involved in the regulation of apoptosis may be targeted. Specifically, a Flk-1/Fas-positive, VEGF-positive clone was identified in which expression of APAF-1 was almost completely abrogated.