Molecular Evidence for Independent Origin of Multifocal Neuroendocrine Tumors of the Enteropancreatic Axis

Abstract

Neuroendocrine tumors of the enteropancreatic axis are often multifocal. We have investigated whether multifocal intestinal carcinoid tumors and multifocal pancreatic endocrine tumors arise independently or whether they originate from a single clone with subsequent intramural or intrapancreatic spread. Twenty-four cases, including 16 multifocal intestinal carcinoid tumors and eight multifocal pancreatic endocrine tumors, were studied. Genomic DNA samples were prepared from 72 distinct tumor nodules using laser capture microdissection. Loss of heterozygosity (LOH) assays were done using markers for putative tumor suppressor genes located on chromosomes 9p21 (p16), 11q13 (MEN1), 11q23 (SDHD), 16q21, 18q21, and 18q22-23. In addition, X chromosome inactivation analysis was done on the tumors from eight female patients. Twentytwo of 24 (92%) cases showed allelic loss in at least one tumor focus, including 15 of 16 (94%) cases of multifocal carcinoid tumors and 7 of 8 (88%) cases of multifocal pancreatic endocrine tumors. Eleven of 24 (46%) cases exhibited a different LOH pattern for each tumor. Additionally, 9 of 24 (38%) cases showed different LOH patterns among some of the coexisting tumors, whereas other coexisting tumors displayed the same allelic loss pattern. Two of 24 (8%) cases showed the same LOH pattern in every individual tumor. X chromosome inactivation analysis showed a discordant pattern of nonrandom X chromosome inactivation in two of six informative cases and concordant pattern of nonrandom X chromosome inactivation in the four remaining informative cases. Our data suggest that some multifocal neuroendocrine tumors of the enteropancreatic axis arise independently, whereas others originate as a single clone with subsequent local and discontinuous metastasis. (Cancer Res 2006; 66(9): 4936-42)

Inverted papilloma of the urinary bladder: a molecular genetic appraisal

“…37 The most consistently informative marker of the clonal composition of neoplasm in female subjects is the nonrandom pattern of X-chromosome inactivation. 26,29 Polyclonal proliferative lesions in female patients are composed of cells with different origins and randomly inactivated X-chromosomes will be identified within them, while neoplasms arising from a single clone are composed of cells in which the same X-chromosome is inactivated. 26,29,38,39 In our study, one inverted papilloma showed nonrandom inactivation of X-chromosomes, suggesting that inverted papillomas arise from a single progenitor cell and supporting the clonal process of inverted papilloma.…”

“…26,28 The clonality of the samples was evaluated on the basis of a polymorphism of the X-linked human androgen receptor gene (HUMARA) locus. 26,[28][29][30] This technique is dependent on digestion of DNA with the methylation-sensitive restriction enzyme HhaI, PCR amplification of the HUMARA locus and the detection of methylation at this locus. With this method, only the methylated HUMARA allele is amplified by PCR.…”

“…Nonrandom X chromosome inactivation indicates a clonal process. 29 The cases were considered informative if the control sample displayed two alleles after PCR amplification without HhaI digestion. Nonrandom inactivation of the X chromosomes was defined as a complete or nearly complete absence of one or the other allele after HhaI digestion, indicating predominance of one androgen receptor allele.…”

Loss of heterozygosity identifies genetic changes in chronic myeloid disorders, including myeloproliferative disorders, myelodysplastic syndromes and chronic myelomonocytic leukemia

“…6 Briefly, the cells of interest are identified and thermoplastic transfer film is placed over the area of interest with the film on an uncoverslipped, uncharged glass slide. A laser is activated and passes through the transfer film.…”

“…Polymerase chain reaction (PCR) was used to amplify genomic DNA at seven specific loci on six different chromosomes: 7q31.1 (D7S2554); 8q24 (D8S263); 9p21 (D9S157, D9S161); 13q14 (D13S319); 17p13 (TP53); 20q12 (D20S108). 6 PCR was performed in a 25 ml volume containing 0.8 mCi of a-32 P-labeled dATP, 2 mM of mappair primers, 0.3 mM of dNTP, 0.65 U of iTaq DNA Polymerase (Bio-Rad Lab, Hercules, CA, USA). PCR amplification has an initial denaturation step of 951C for 5 min, followed by 35 cycles at 951C for 30 s, at 581C for 30 s, and at 721C for 30 s, and then followed by a single final extension step at 721C for 7 min.…”