The most prevalent problem in cancer therapy is the re-growth and metastasis of malignant cells after standard treatment with surgery, radiation, and chemotherapy. Gene therapy approaches have suffered from the inadequate transduction efficiencies of replication-defective vectors that have been used thus far. Replication-competent vectors represent an emerging technology that shows considerable promise as a novel treatment option, particularly for locally advanced or recurrent cancer. In contrast to conventional replication-defective retrovirus vectors, gene transfer using replication-competent murine leukemia virus (MLV)-based retrovirus vectors has proven to be highly efficient, resulting in >98% transduction throughout entire tumors over a period of several weeks in breast cancer, prostate cancer, and glioma models, even with an initial inoculum of vector supematant as low as 10e(4-5) total infectious units, corresponding to MOIs as low as 0.001. While various other replicating oncolytic viruses are now in development as cancer therapeutics, the use of replication-competent retrovirus (RCR) vectors has rarely been contemplated due to the potential risks that might be associated with uncontrolled spread of virus. In fact, MLV-based RCR vectors exhibit a significant degree of inherent tumor-selectivity, as extratumoral spread was undetectable by sensitive PCR assays in all normal tissues tested, presumably due to the intrinsic inability of MLV to infect quiescent cells, and MLV exhibits additional advantageous characteristics including a simple genome and well-characterized life cycle, fidelity of transcriptional regulation, and efficient non-lyric transmission of suicide genes which are stably expressed until maximal intratumoral vector spread and optimal timing for pro-drug administration is reached. Using RCR vectors expressing the suicide gene cytosine deaminase, we have now demonstrated highly efficient killing of cancer cells both in culture and in tumor models in vivo. Treatment of intracranial gliomas with RCR vector-mediated suicide gene therapy resulted in 100% survival for >120 days, compared to 0% survival of control groups in < 40 days, and viral persistence was observed in all metastatic ectopic foci. We now propose that the efficient and stable conferral of viral neo-antigens to the tumor cells present an opportunity to further improve the long-term efficacy of this therapy by combining multiple approaches, including (1) seroimmunotherapy by follow-up administration of radioisotope-conjugated antibodies directed against viral proteins, and (2) active immunotherapy by peripheral sensitization with tumor vaccines expressing viral antigens, combined with intratumoral gene transfer of immunostimulatory cytokines. In this application, we have combined the expertise of multiple collaborators to directly test these approaches in syngeneic intracranial glioma models in vivo. ? ?
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