Histopathologic classification of pediatric rhabdomyosarcoma (RMS) has established subtypes that are associated with distinct clinical characteristics. The distinction between the subtypes is strengthened by molecular studies that identified PAX3/PAX7-FKHR gene fusions in alveolar RMS and allelic loss of chromosomal region 11p15.5 in embryonal RMS. Despite the utility of this classification system, clinical heterogeneity exists within these subtypes in such parameters as site, stage, and outcome. As a potential correlate of this heterogeneity, alterations of other prot-oncogenes and tumor suppressor genes have been detected in smaller subsets of RMS tumors. In particular, mutations of p53 and RAS genes, amplification of MYCN and 12q13-15 loci, and deletions of CDKN2A have been reported in cases of RMS. Some of these changes occur preferentially in a single subtype whereas other alterations occur at a low frequency in all subtypes. These findings indicate that RMS pathogenesis can be characterized as a multistep process involving fundamental subtype-specific events that collaborate with other subtype-specific and non-specific events. The occurrence of alterations in only a subset of each RMS subtype provides the basis for genetic heterogeneity within these categories that may provide an explanation for clinical heterogeneity. In this application, the Biology Committee of the Intergroup Rhabdomyosarcoma Study (IRS) Group will examine the clinical significance of p53 and RAS family mutations, MYCN and 12q13-15 amplification, and CDKN2A alterations in a large panel of well-characterized RMS tumors collected from the closely monitored and uniformly treated patients of the IRS trials. In particular, we will assay for each of these genetic alterations as well as PAX3/PAX7- FKHR fusion status in a set of 98 frozen tumor samples from IRS-IV to establish the overall frequencies and associations with pathologic and clinical characteristics of these cases. Based on these findings, specific associations will be further examined by directed analysis of specific genetic alterations in selected tumor specimens from the collection of over 900 paraffin blocks of IRS-III. These studies will provide a definitive analysis of the occurrence and clinical significance of these genetic alterations in RMS, and identify novel markers that will be analyzed further in prospective IRS studies.
