Alveolar rhabdomyosarcoma is an aggressive soft tissue tumor which occurs in the pediatric population. Diagnosis is often complicated by the paucity of features of strained muscle differentiation and its similarity to a large group of small, round, blue cell tumors. Recent studies of this tumor have identified a characteristic translocation of chromosomes 2 and 13, t(2;13)(q35;q14). We hypothesize that genes at or near the t(2;13) breakpoint are involved in normal cell growth and development and that disruption of these genes by the t(2;13) translocation contributes to the pathogenesis of alveolar rhabdomyosarcoma. Using alveolar rhabdomyosarcoma cell lines developed by the core laboratories, we have performed conventional and pulsed-field gel electrophoresis with Southern blot analysis to rule out rearrangement of numerous candidate genes mapping to the relevant regions of 2q and 13q. To identify the gene(s) disrupted by the t(2;13) translocation, we plan an interdisciplinary approach combining molecular biology with somatic cell genetics and cytogenetics. Somatic cell hybrids in which the translocated chromosomes have been segregated from their normal counterparts will permit us to determine the position of the translocation breakpoint on published genetic maps of 2q and 13q. We will then apply those probes mapping closest to the 13q breakpoint to begin chromosomal walking towards the breakpoint with a yeast artificial chromosome (YAC) library constructed from a monochromosomal 13 somatic cell hybrid. Sequences cloned by this walking strategy and appropriately localized candidate genes will be tested for proximity to the breakpoint by Southern blot-gel electrophoresis analysis. Multiple tumors will be screened with hose probes which detect rearrangements to determine the distribution of the breakpoints and thereby focus on gene-encoding regions consistently disrupted by the t(2;13) translocation. Sequence-based assays will be developed for detection of the t(2;13) translocation in clinical material. Our unique ability to combine molecular biologic investigation with cytogenetic studies in a clinical setting will provide new information on the genetic etiology of this tumor, correlations of genetic changes with histopathologic and clinical features, and a molecular marker to facilitate diagnosis. Our long-term goals are to characterize the normal function of the genes at the t(2;13) breakpoint, analyze the consequences of rearrangement of these genes in rhabdomyosarcoma, and determine the utility of this molecular marker in clinical management.
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