Ester Rozenblum

DPC4, A Candidate Tumor Suppressor Gene at Human Chromosome 18q21.1

About 90 percent of human pancreatic carcinomas show allelic loss at chromosome 18q. To identify candidate tumor suppressor genes on 18q, a panel of pancreatic carcinomas were analyzed for convergent sites of homozygous deletion. Twenty-five of 84 tumors had homozygous deletions at 18q21.1, a site that excludes DCC (a candidate suppressor gene for colorectal cancer) and includes DPC4, a gene similar in sequence to a Drosophila melanogaster gene (Mad) implicated in a transforming growth factor-β (TGF-β)-like signaling pathway. Potentially inactivating mutations in DPC4 were identified in six of 27 pancreatic carcinomas that did not have homozygous deletions at 18q21.1. These results identify DPC4 as a candidate tumor suppressor gene whose inactivation may play a role in pancreatic and possibly other human cancers.

Mad-related genes in the human

Resistance to the growth inhibitory effects of TGF-β is common in human cancers1,2. However, the mechanism(s) by which tumour cells become resistant to TGF-β are generally unknown. We have identified five novel human genes related to a Drosophila gene called Mad which is thought to transduce signals from TGF-β family members3–5. One of these genes was found to be somatically mutated in two of eighteen colorectal cancers, and three of the other genes were located at chromosomal positions previously suspected to harbor tumour suppressor genes. These data suggest that this gene family may prove to be important in the suppression of neoplasia, imparting the growth inhibitory effects of TGF-β-like ligands.

Multi-Layered Cancer Chromosomal Instability Phenotype

Whole-chromosomal instability (W-CIN) – unequal chromosome distribution during cell division – is a characteristic feature of a majority of cancer cells distinguishing them from their normal counterparts. The precise molecular mechanisms that may cause mis-segregation of chromosomes in tumor cells just recently became more evident. The consequences of W-CIN are numerous and play a critical role in carcinogenesis. W-CIN mediates evolution of cancer cell population under selective pressure and can facilitate the accumulation of genetic changes that promote malignancy. It has both tumor-promoting and tumor-suppressive effects, and their balance could be beneficial or detrimental for carcinogenesis. The characterization of W-CIN as a complex multi-layered adaptive phenotype highlights the intra- and extracellular adaptations to the consequences of genome reshuffling. It also provides a framework for targeting aggressive chromosomally unstable cancers.

Novel near-diploid ovarian cancer cell line derived from a highly aneuploid metastatic ovarian tumor

A new ovarian near-diploid cell line, OVDM1, was derived from a highly aneuploid serous ovarian metastatic adenocarcinoma. A metastatic tumor was obtained from a 47-year-old Ashkenazi Jewish patient three years after the first surgery removed the primary tumor, both ovaries, and the remaining reproductive organs. OVDM1 was characterized by cell morphology, genotyping, tumorigenic assay, mycoplasma testing, spectral karyotyping (SKY), and molecular profiling of the whole genome by aCGH and gene expression microarray. Targeted sequencing of a panel of cancer-related genes was also performed. Hierarchical clustering of gene expression data clearly confirmed the ovarian origin of the cell line. OVDM1 has a near-diploid karyotype with a low-level aneuploidy, but samples of the original metastatic tumor were grossly aneuploid. A number of single nucleotide variations (SNVs)/mutations were detected in OVDM1 and the metastatic tumor samples. Some of them were cancer-related according to COSMIC and HGMD databases (no founder mutations in BRCA1 and BRCA2 have been found). A large number of focal copy number alterations (FCNAs) were detected, including homozygous deletions (HDs) targeting WWOX and GATA4. Progression of OVDM1 from early to late passages was accompanied by preservation of the near-diploid status, acquisition of only few additional large chromosomal rearrangements and more than 100 new small FCNAs. Most of newly acquired FCNAs seem to be related to localized but massive DNA fragmentation (chromothripsis-like rearrangements). Newly developed near-diploid OVDM1 cell line offers an opportunity to evaluate tumorigenesis pathways/events in a minor clone of metastatic ovarian adenocarcinoma as well as mechanisms of chromothripsis.

Abstract 4208: Integrated oligo aCGH and SKY analysis of genomic alterations in malignant mesothelioma cell lines.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Malignant mesothelioma (MMt) is a rare and very aggressive tumor of the mesothelium. Its most prevalent subtype is malignant pleural mesothelioma, eighty percent of which are associated with asbestos exposure. MMt incidence is 0.9 new cases per 100,000 persons per year. The five year relative survival is 7% overall, which positions MMt immediately above pancreatic cancer (6%), at the bottom of the survival list by cancer site. We applied a comprehensive molecular cytogenetic analysis using spectral karyotyping (SKY) and oligo aCGH to a panel of MMt cell lines. We found the largest set of recurrent and non-recurrent homozygous deletions (HD) for MMt (88 HDs in 17 cell lines; 52 recurrent HDs that fall into ten genomic areas). Detailed analysis of the most frequently homozygously deleted area located at 9p21.3 revealed that the main targets are CDKN2A/p14ARF (in 100% of the MM cell lines), CDKN2A/p16INK4A and CDKN2B /p15 (94% of cell lines). p14ARF seems to be the main target of HDs in the 9p21.3 area (deleted in all 17 MMt cell lines studied), followed by p16INK4A (deleted in 16 cell lines) indicating the inactivation of two major tumor suppressing pathways, p53 and Rb. Accordingly, mutations or HDs of p53 and Rb are infrequent events in MMt tumors. Other recurrent HDs were located at 9p21.2 (targeting LINGO2) and at16p13.3 (RBFOX1) in 41% of cell lines, 22q11.23 (GSTT1) in 29%, 22q12.2 (NF2) and 3q26 (no known genes) in 23% each, 8p11.22 -23 (ADAM5P/ADAM3A) in 18%, 3p21.3-p21.2 (RPL29), 3p21 (DUSP7), 4q22.1 (FAM190A) and 13q11-q12 (LATS2) in 12% each of the 17 MM cell lines While p16INK4A, p14ARF, p15, LATS2 and NF2 have been shown to behave as tumor suppressors in MMts, other genes in the HD areas presented here could be new tumor suppressor candidates. Structural chromosomal rearrangements were mainly non-reciprocal translocations (74% of aberrations detected by SKY) and deletions (23%). No recurrent balanced or unbalanced translocations have been found. Structural chromosomal rearrangements resulted in recurrent losses of fragments or whole chromosomal arms of 1p, 3p, 8p, 9p, 13q, 14q, 15q, 18q, 22q and gains of 1q, 5p, 7p, 8q, 17q, 20q. Small HDs were frequently embedded in the areas of focal losses. Integration of SKY and aCGH data allowed reconstruction of chromosomal rearrangements leading to the formation of HDs. Our data imply that only with acquisition of structural or numerical karyotypic abnormalities can MMt cells attain a complete loss of the 9p21.3 genomic area and loss of tumor suppressor genes located there. Tetraploidization seems to be a late event in the karyotypic progression of MMt cells, after HD in the 9p21.3 area has already been acquired. Citation Format: Ester Rozenblum, Geula Klorin, Oleg Glebov, Robert L. Walker, Yoonsoo Park, Paul S. Meltzer, Ilan R. Kirsch, Frederic J. Kaye, Anna V. Roschke. Integrated oligo aCGH and SKY analysis of genomic alterations in malignant mesothelioma cell lines. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4208. doi:10.1158/1538-7445.AM2013-4208

Abstract 3265: Immortalization and comprehensive characterization of a human ovarian adenocarcinoma cell line derived from an Ashkenazi Jewish patient

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC We have established and characterized a new ovarian cancer cell line derived from stage IV serous ovarian adenocarcinoma metastatic tissue from a 47 year old Ashkenazi Jewish patient. The patient developed ovarian cancer at age 44 and died of the disease at age 48. The tissue was obtained after an extensive secondary cytoreductive intraperitoneal surgery. Prior to the surgery the patient underwent primary cytoreduction followed by several systemic therapies with recurrences. A culture of the metastatic tissue was transfected with Simian Virus 40 Large T-Antigen and telomerase. The cell line has been in culture on and off during 3 years. The identity of the cell line was confirmed by Short Tandem Repeat genotyping on genomic DNA extracted from early and late passages as well as from frozen metastatic tissue. The cell line was further characterized by array-Comparative Genomic Hybridization and gene expression microarray; and by Spectral Karyotyping and Fluorescent In Situ Hybridization. The results of these analyses were compared, when appropriate, with those of frozen metastatic tissue. A low level of genetic instability was detected in the cell line, but the karyotypes seem to be near diploid and relatively stable, after several years of cultivation. We believe that this novel ovarian cancer cell line could contribute to the understanding of the molecular genetic basis of the disease in this advanced stage, and be representative of the tumors of a great number of patients that become resistant to the majority if not all known cancer therapies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3265.