Molecular Characterization of Gastrointestinal Tumors Using Tumor Suppresser Gene Associated Microsatellite Markers and their Relationship to Pathological and Clinical Features [original]

Menghong Sun

Dr. med.

Molecular Characterization of Gastrointestinal Tumors

Using Tumor Suppresser Gene Associated Microsatellite

Markers and their Relationship to Pathological and Clinical

Features

Geboren am 15. März, 1960

Studiengang der Fachrichtung Medizin vom SS/WS 1978 bis SS/WS 1983 (Bachelor

of Medicine), vom SS/WS 1985 bis SS/WS 1988 (Master of Medizin)

Klinisches Studium in Anhui Medizinische Universität

Staatsexamen am 05, 07, 1988 an der Anhui Medizinische Universität

Promotionsfach: Chirugie

Doktorvater: Prof. Dr.Med. M.v. Knebel Doeberitz

Tumors are the result of clonal stepwise alterations in multiple genes. Most of

our knowledge on the genetic nature of carcinogenesis has been derived from

the studies of gastrointestinal tumors. Current models provide strong

evidence for the existence of genetically heterogeneous preneoplastic cell

populations which furtheron lead to genetic heterogeneity in the tumors.

These findings outline the events responsible for determining the natural

history of gastrointestinal tumors. The prognostic impact of reading molecular

profiles of individual cancers is unclear at present. We addressed the

question whether a molecular profiling approach by analyzing allelic

imbalances of specific tumor suppressor gene loci as an evidence for allelic

loss (loss of heterozygosity) might correlate with the prognosis of the

respective cancers.

Matched tumor and normal DNA from CRC (colorectal cancer) of 79 patients

(1986 until 1988); 10 adenomas obtained from 10 FAP (familial adenomatous

polyposis loci) patients and two rare esophageal carcinosarcomas were

obtained after microdissection. 11 tumor associated loci were amplified by

PCR using fluorescein labeled microsatellite primers. PCR products were

analyzed on a DNA sequencer (A.L.F.) and allelic loss (LOH) was determined

using the Fragment Manager software.

In 86% (68/79) of the tumors frequent LOH was observed at the tumor

suppressor gene loci 5q12 (38%), DCC (72%), p53 (74%), NF2 (30%) and RB

(25%), as well as in loci not directly associated with colorectal carcinogenesis

such as 17q (BRCA1, 31%), 15q13 (ß2-microglobulin, 34%) and 6p21 (TAP1,

27%). Patients showing with LOH in RB or 5q12 had a better prognosis than

patients without LOH at these loci. In 14% (11/79) of colorectal tumors

genome wide instability was observed in more than two loci, reflecting a high

level of genetic instability. In an early adenoma LOH at the 5q12 (D5S107)

locus was detectable in tenascin positive crypts. Adenomas of a second

patient displayed variable LOH at the 18q12 (D18S34) locus, i.e. LOH in one

tenascin positive adenoma but wildtype in a different tenascin positive

adenomas. Such adenomas display genetic heterogeneity suggesting

temporally and spatially different mutagenic events.

In two esophageal carcinosarcomas LOH was also detected at the p53 locus,

hMLH1 and other loci. A rare splice donor site mutation at the boundary of

exon-intron 6 was detected in both carcinomatous and sarcomatous tumor

components which is a strong hint for the clonal origin of both tumor

elements.

We have established a complex LOH profile for sporadic colorectal tumors.

Molecular profiling of tumors represents a novel approach in defining different

types of genetic alterations arising during tumorigenesis.