Robert B. Jenkins, M.D., Ph.D. - US grants

glioma; neoplasm /cancer genetics; neoplasm /cancer classification /staging; molecular oncology; point mutation; oncogenes; genetic markers; cytogenetics; in situ hybridization; immunocytochemistry; pathology; flow cytometry; human tissue; histopathology;

neoplastic process; neoplasm /cancer genetics; cell differentiation; phenotype; oncogenic virus; histocompatibility antigens; neoplasm /cancer radiation therapy; neoplasm /cancer epidemiology; neoplasm /cancer pharmacology; prostate neoplasms; breast neoplasms; oncoproteins; hepatocellular carcinoma; multiple myeloma; lymphocytic leukemia; lymphoma; endocrine neoplasm; thyroid neoplasm; colon neoplasms; glioma; molecular pathology; neoplasm /cancer education; gene therapy; genetic regulation; gene mutation; pharmacogenetics; human population genetics; interleukin 6; molecular oncology; ovary neoplasms; small cell lung cancer; cancer registry /resource; neoplasm /cancer relapse /recurrence; human genetic material tag; dissection; human tissue; in situ hybridization;

neoplasm /cancer genetics; karyotype; biomedical facility; neoplastic process; tumor suppressor genes; oncogenes; cell transformation; cell line; animal tissue; human tissue; tissue /cell culture; in situ hybridization;

With the recognition that cancer is a genetic disease, many cancer researchers, including scientists in the Mayo Clinic Cancer Center (MCCC), require access to routine and advanced cytogenetics services. Such services can aid in the identification of genetic regions important for the development of specific malignancies. Such services can also be used to characterize the clonality of tumor specimens and determine the origin (e.g., mouse vs. human) of a specific tumor culture. Thus, the Cytogenetics Shared Resource will provide cytogenetic and molecular cytogenetic services to MCCC members. In addition, this Shared Resource will prepare EBV-transformed B-lymphocyte cell lines for MCCC members.

Alterations of chromosomes 1 and 19 are frequent events in human gliomas and have recently been suggested to be associated with chemotherapeutic response and prolonged survival in anaplastic oligodendrogliomas. However, the genes on these chromosome arms that are altered in gliomas and potentially associated with chemotherapeutic response and survival have not been identified. Furthermore, the exciting response and survival data have not been confirmed in an appropriately-treated, prospectively-collected cohort of patients nor have these clinical correlations been translated to lower grade oligodendrogliomas or other glial tumors. We have made considerable progress in mapping the genomic location of the 19q gene and have identified several genes in this region. In addition, we have assembled several prospectively-collected cohorts of patients to confirm and extend the clinical observations of Caimcross et al. In this application, we propose to 1) Complete the identification of the 19q gene (or genes) involved in the development of gliomas, 2) Begin an evaluation of the function of the 19q gene (or genes), 3) Finely map the minimal 1p deletion region in gliomas, to begin to identify the gene (or genes) on 1p involved in the development of gliomas, and 4) To perform clinical-translational studies designed to validate the postulated predictive value of 1p/19q alterations, to determine if alterations of the specific genes involved have similar associations, and to extend the associations to lower grade tumors and to tumors of different glial lineage. Thus, this project will identify and begin to study the function of the 19q gene, map the 1p deletion region, and further define the clinical significance of the 1p and 19q alterations. The overall result of this project will be an initial understanding of the genetics and biology of the 19q and 1p genes as well as the development of a combined histologic and genotypic assessment of gliomas that could potentially be used to stratify patients for therapy.

DESCRIPTION: (provided by Applicant) Gliomas are a significant cause of morbidity and mortality, and thus have been the focus of frequent basic biologic, basic genetic, and clinical investigations. Numerous genetic and biologic alterations associated with gliomas have been described. However, many of the specific genes involved in gliomas have yet to be identified. A great deal still needs to be learned about the mechanisms by which the known genes are involved in the pathogenesis of gliomas. Furthermore, while much has been learned about the biology and genetics of gliomas, little of this information has been translated into clinical practice. There are still problems with the morphologic classification of gliomas - especially oligodendrogliomas and mixed oligoastrocytomas. It is difficult to predict which patients with anaplastic astrocytomas will suffer early recurrence. And while some gliomas with specific genetic alterations seem to respond to chemotherapy (e.g., Cairncross et al., JNCI 90:1473,1998), these observations need to be confirmed and extended and additional therapeutic genetic targets identified and/or developed. Through four projects and two cores this program, which builds on the past experience of the Glioma Marker Network (GMN) consortium, will study the basic biology of several specific biochemical and genetic alterations associated with gliomas. It will also continue to evaluate the predictive, prognostic, and pathologic relevance of these alterations. Project 1 will study the biologic and clinical relevance of 7q gain in gliomas, with emphasis on the mechanism by which this alteration predicts a poorer prognosis. Project 2 will evaluate the function of mutated and amplified EGFR in gliomas and will test several potential therapeutic approaches targeting these mutant receptors. Project 3 will identify and study the function of the 19q gene associated with gliomas and associated with a prolonged response of some gliomas to chemotherapy. Project 4 will study the biologic and clinical relevance of glycolipid and glycosyltransferase alterations in gliomas. Thus, through these projects, this highly-interactive and experienced program will make significant progress toward understanding the pathogenesis of gliomas, and translating this knowledge into new diagnostic and therapeutic tools.

DESCRIPTION (provided by the applicant): This application addresses broad Challenge Area (08) Genomics, and specific Challenge Topic 08-CA-101* Augmenting Genome-Wide Association Studies. The development of glioblastoma (GBM) has been hypothesized to be associated with relatively common germline alterations with limited penetrance. Collaborating with the group at the University of California at San Francisco (UCSF) we recently reported that SNPs mapping to 9p and 20q are associated with the development of GBM. We propose to characterize the germline alterations within these 9p and 20q regions and to correlate these with glioma 9p deletion and 20q gain. Specific Aim 1: Perform detailed germline genetic analysis of the associated 9p and 20q regions using 1200 cases and controls to determine the prevalence of known polymorphisms and new alterations. Aim 1a: Perform custom genotyping of 600 previously genotyped cases and matched controls for all non-redundant SNPs to better define haplotypes. Impute haploytpes and evaluate their association with GBM development. Aim 1b: Perform high-throughput sequencing of the ~200kb and ~100kb regions within 9p and 20, respectively, in 50 GBM cases and 50 controls that carry the imputed at-risk haplotypes. To directly determine haplotype structure, perform high-through-put sequencing of 50 isolated at-risk and non-risk chromosomes. Aim 1c: Custom genotype candidate polymorphisms in 600 new cases and matched controls to validate new alterations from Aims 1a and 1b. Aim 1d: Perform custom aCGH analysis of the 9p and 20q regions to ascertain copy number variants. Specific Aim 2: Perform detailed genetic analysis and expression analysis of gliomas from the cases. Aim 2a: Using FISH and custom CGHa define glioma 9p deletion and 20q gain status. Aim 2b: Sequence all exons in the 9p and 20q regions. Assess methylation of target gene promoters. Aim 2c: Evaluate the tumor expression of all known exons and miRNAs within the targeted regions. Determine the underlying GBM genetic subtype. Specific Aim 3: Correlate polymorphism/ alteration/haplotype prevalence differences and with acquired glioma alterations to generate a list of candidate germline 9p and 20q mutations likely to be associated with the development of gliomas. Gliomas cause significant morbidity and mortality. Approximately 18,500 people in the U.S. are diagnosed with glioma each year. Because most gliomas are biologically aggressive, approximately 12,800 people in the U.S. succumb to these tumors every year. Understanding the predisposition to gliomas will have major implications for the prevention of gliomas as well as the management of these tumors.

The central focus of the Cytogenetics Shared Resource (CSR) is to provide cytogenetic and molecular cytogenetic analysis of human and animal model research samples. These samples may be in the form of cultured cells, fresh or frozen tissue, paraffin-embedded sections, or tissue microarrays (TMAs). Specific cell types include tumor cell lines, xenograft cells, hybrid cells from chimeric animal models, neural stem cells, and mouse embryonic fibroblasts, among others. The CSR provides specialized services including cell culture, routine cytogenetic analysis, chromosome breakage analysis, interphase and metaphase Fluorescence in situ Hybridization (FISH), Spectral Karyotyping (SKY) analysis of human and mouse metaphases, and homebrew FISH Probe Production. This last service provides novel FISH probes that are specific to investigators needs and are usually not commercially available. These probes are then used for FISH analysis studies. The CSR is expanding this service beyond homebrew enumeration FISH probes by creating custom translocation/fusion probes, break-apart probes, and multi-color FISH probes to ultimately assist in developing strategic probes that become clinical diagnostic and prognostic markers of disease. Other services provided by the CSR to meet the needs of investigators include utilization of the CSR laboratory for slide processing and fluorescent microscope and imaging system use. Briefly, applications of CSR services include: to characterize the clonality of tumor specimens;to determine the origin of a specific tumor culture (i.e., mouse vs. human), to aid in the identification of genetic regions important for the development of specific malignancies, and to map the location of DMA sequences, genes, or transgene insertions. With the recognition that cancer is a genetic disease, many cancer researchers, including scientists in the Mayo Clinic Cancer Center (MCCC), require access to CSR services. This Shared Resource is utilized by 9 of the 12 Cancer Center Programs. Approximately 66% of CSR users are CCSG members and represent 92% of the overall utilization.

Provocative Question 3: Summary/Abstract This proposal addresses Provocative Question #3, ?Do genetic interactions between germline variants and somatic mutations contribute to differences in tumor evolution?? The new 2016 WHO Classification of Tumors of the Central Nervous System utilizes two somatic alterations to molecularly classify adult diffuse glioma: IDH mutation and 1p/19q codeletion. We and others have shown that TERT promoter mutation further classifies gliomas into molecular subtypes with distinct clinical characteristics. In addition, during the last five years much has been learned about the germline predisposition to gliomas: genome-wide association studies (GWAS) have revealed that 25 regions in 24 genes are associated with glioma development. Our group identified many of these associations and fine-mapped two of them (MYC/CCDC26 and TP53) and demonstrated that the MYC/CCDC26 variant (rs55705857) is associated with IDH-mutant gliomas. Indeed we found rs55705857 to have an odds ratio>6 for development of IDH-mutated glioma and lowers the age of onset by ~10 years. It is clear that the risk allele of rs55705857 interacts with somatic IDH-mutation to accelerate low-grade glioma development. We hypothesize that germline variants interact with somatic alterations to accelerate the development of IDH-mutant and IDH wild-type gliomas. Our published and preliminary data provide strong evidence in support of this hypothesis; but it must be explored further. While each of the known 25 regions has been evaluated with respect to risk of the 2016 WHO molecular subtypes, an unbiased GWAS has yet to be performed. Thus, Aim 1 will cost- effectively utilize previously-collected GWAS data to identify novel germline variants that are associated with the WHO subtypes in order to provide better patient risk assessment. Aim 2 will translate these findings into the clinic by integrating the germline and somatic alterations to determine associations with patient survival. Lastly, Aim 3 will use functional genomics to begin to understand the mechanisms by with rs55705857 and other variants accelerate IDH-mutant glioma.