Project 1 has had two major objectives. The first was to enhance effectiveness of candidate therapeutic viruses constructed by deletion of specific genes. These viruses are more effective in treatment that combines virus administration to the tumor and radiation. Nevertheless, the effectiveness of combined therapy is tumor genotype-dependent. In the past 5 years, we have collaborated with Project 2 to (a) identify the molecular basis for enhancement of attenuated viruses by ionizing radiation (IR) and (b) identify a cellular gene (MEK) whose product, when used properly in conjunction with IR, could overcome the restriction to attenuated virus replication imposed by tumor genotypes. The results of these studies are described in the Progress Report for Project 2. Our second objective was to construct viruses that can only infect cancer cells but not normal cells. In essence we ablated the ability of HSV-1 to attach to heparin sulfate proteoglycans and to enter cells by way of its natural protein receptors, HveA and nectin 1. Thus, the engineered virus R5141 infects cells solely via the IL13 a2 receptor while R5181 enters cells via the urokinase plasminogen activator receptor. Our findings have significantly contributed to understanding the biology and biochemistry of glycoprotein (g) D. Our new objectives are as follows:
AIM 1 will develop methods for production of clinical grade targeted viruses. Since targeted viruses must be produced in noncancerous cells that stably express the novel receptor(s), we propose several ways in which we can produce clinical grade viruses. The objective of AIM 2 is to develop more effective viruses for therapy of GBM with AY34.5 mutant viruses. Therapeutic viruses currently in clinical trials extend survival time in a small fraction of treated patients in a tumor genotype dependent manner. We have constructed a virus in which the constitutively active MEK gene is driven by an IR inducible promoter (R2660). In preliminary studies R2660 plus IR blocked growth of a tumor resistant to virus or IR alone. The safety features of this and other mutant viruses will be studied. The objective of AIM 3 is to render the viruses targeting specific receptors on cell surfaces more effective. We have established proof of principle but do not consider the current generation optimal. We have identified specific shortcomings and ways to improve these viruses The objective of AIM 4 is based on the novel observation that a component of the amino-terminal domain of the urokinase plasminogen activator receptor interacts with the carboxyl-terminus of the gD ectodomain. The amino terminus of urokinase plasminogen activator contains a Kringle domain with affinity for lysines. Thus, we will determine whether the Kringle domain of human plasminogen can be used to target gD and by extension, target HSV-1 to its ligand, the glucose regulatory protein 78.
Project 1 is a component of a Program Project Grant designed to cure or at least effectively prolong quality of life of patients with malignant gliomas. At this point in time the proof of principle has been established at both basic and clinical levels. The task confronting Project 1 is to design, construct and test the next generation of therapeutic viruses characterized by enhanced therapeutic profile and to develop methods for their production in GMP facilities. Validation ofthe novel viruses will be done in collaboration with Projects 2 and 3 and, ultimately, with Project 4.
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