We cloned the human reduced folate carrier gene a critical determinant of antifolate sensitivity of tumor cells and have begun to study its regulation. We have found that this gene is cell cycle regulated and has maximal expression during S-phase. We have isolated 5' genomic RFC sequences and identified the presence of multiple promoters regulating the RFC expression. We have constructed a series of RFC promoter reporter fusion genes and have identified sequences involved in the basal expression from each of the upstream promoter regions. We are currently studying the sequences involved in cell cycle regulation of this gene. """"""""We are also studying the use of recombinant adenoviruses for gene therapy of cancer. Recombinant viral vectors have been constructed that express the cyclin kinase inhibitors p16/INK4. We have shown that each of this vector (AdP16) is cytotoxic to breast cancer cells that express wild type RB, resulting in cell cycle inhibition and apoptosis."""""""" We have also constructed an adenoviral vector expressing von Hipple Lindau gene (AdVHL) and found that overexpression of VHL results in inhibition of growth and cell cycle arrest of renal cancer cell lines expressing mutant VHL. Furthermore adenoviral mediated of VHL results in increased p27kip gene expression through both increased protein stability and increased p27 gene transcription. """"""""We are studying the ability to reconstitute patients with metastatic breast cancer with multidrug resistance gene (MDR1) modified hematopoietic cells. CD34+ cells are transduced ex vivo with retroviral vectrors containing either the MDR1 gene or the bacterial NeoR phosphotransferase gene. 6/6 patients have evidence for MDR1 gene marking following reconstitution. While only 3/6 patients have evidence of NeoR gene marking. Following treatment with MDR1 chemotherapy, 3/6 patients show increased levels of granulocytes containing the MDR1 vector while only 1/6 patients have evidence of NeoR marking in granultocytes following chemotherapy. These results may suggest that tMDR1 overexpression may enhance engraftment potential and/or protect hematopietic cels from toxicity of chemotherapy."""""""" """"""""We also have performed two phase I clinical trials investigating the efficacy and toxicity of novel agents. The first involves study of a bispecific antibody MDX210 which binds both erbB2 receptors on tumor cells and FcgRI receptors on monocytes and granulocytes. Administration of this MDX210 to patients with erbB2 positive tumors results in rapid decrease in circulating monocytes and marked increase in release of several cytokines including gamma interferon, IL-6, and G-CSF."""""""" """"""""The other phase I trials involves study of a farnesyl transferase inhibitor R11577, one of the first of this new class of agents to enter clinical trials. Inhibition of protein farnseylation inhibits the posttranslational modification of several proteins including the of proteins including ras protein. Inhibition of ras farnesylation blocks insertion of the ras protein int op cell membranes and inhibits growth of tumor cell lines expressing mutant ras as well as wild type ras. In this trial R11577 is given orally bid for 5 days every two weeks. Results indicate that R15577 is orally bioavailable and that peak serum concentrations increase with doses up to 500 mg po. The drug is well tolerated with nausea, vomiting, and fatigue being the major toxicities. There was little hematopoietic toxicity when R11577 is given in this intermitant schedule. .""""""""
