The objective of this proposal is to examine the relationship between extrachromosomal (EC) DNA and tumor phenotype. Prior studies have revealed that most primary tumors contain EC DNA, often in the form of acentric structures called double minutes (DMs). DMs can harbor amplified oncogenes or drug resistance genes. Critical to this proposal is the finding that low concentrations of hydroxyurea can eliminate DMs and tumor phenotype. The elimination of c-myc-containing DMs in HL60 cells induces differentiation and reduces tumorigenicity. These and other data support the hypothesis that the genes associated with DMs are responsible for maintaining the malignant phenotype in vivo. This proposal will further investigate the mechanism of HU-induced differentiation in HL60 cells and identify novel DM- associated genes that are involved in regulating tumor phenotype.
The Specific Aims are: l. To conduct detailed in vitro and in vivo studies in HL60 cells to further characterize the mechanism of induction of differentiation through the elimination of EC DNA. This will be accomplished by utilizing hydroxyurea (HU) to effect EC DNA loss and comparing HU's effects to those of a known differentiating agent, retinoic acid (RA). Once these studies have been completed, the combination of HU and RA will be studied to determine if there is an additive or synergistic effect. 2. To identify genes associated with DMs in tumor cells and determine their role in tumor cell growth and differentiation. Tumor cell lines containing DMs will be obtained and the genes will be identified and characterized using microdissection and PCR amplification. The significance of EC DNA has been clearly established and further study is needed to determine its role in the growth and differentiation of cancer cells. The data generated from these studies will be used to design novel therapeutic strategies for patients with cancer.