Streamlined DNA Sequencing to Identify a Tumor Gene

Little, Randall

Abstract

Ependymomas account for approximately 10% of all brain tumors in children and young adolescents. Cytogenetic and loss of heterozygosity studies of ependymomas have shown frequent alterations of chromosome 22q. These data strongly support a tumor suppressor gene involved in this neoplasia on 22q. However, neither the chromosome 22q nor other genes involved in ependymoma have been identified. We have identified a ependymoma patient who exhibits a constitutional translocation [t(1;22)(p22;q11.2)]. The translocation in 22q11.2 lies between the DiGeorge syndrome and the breakpoint cluster (BCR) regions-an interval estimated to be approximately 4 Mb. We hypothesize that this translocation interrupts an ependymoma tumor suppressor gene in 22q11.2 and that it is a landmark for cloning the gene.
Our aim i n this proposal is to clone this ependymoma tumor suppressor gene by delimiting the 22q1 1.2 breakpoint to 150 kilobases (Phase I) and performing streamlined high-coverage (more than 90%), low- redundancy (2-2.5X) multiplex DNA sequencing and computer exon identification of this region (Phase II). The isolation of an ependymoma tumor suppressor gene would permit a mechanistic understanding of this tumor, as well as other cancers in which this gene may be involved. The cloning of this gene will have important applications to monitoring, diagnostics, and therapeutics for ependymoma. This proposal also will be an important evaluation of the streamlined large-scale DNA sequencing and exon identification approach to positionally clone genes. POTENTIAL COMMERCIAL APPLICATION: The identification of an ependymoma tumor suppressor gene is expected to have applications in monitoring, diagnostics, and therapeutics, as well as a basic understanding of this cancer. This gene may have an association with prognosis; this would be of great importance since there is no good prognostic marker. This gene may be involved in other cancers which show chromosome 22q alterations. Our streamlined sequencing approach should have applications to positionally cloning other disease genes.