Original Paper DiVerential Prognosis of Replication Error Phenotype and Loss of Heterozygosity in Sporadic Colorectal Cancer M.-J. Massa, 1 P. Iniesta, 1 R. Gonza Âlez-Quevedo, 1 C. de Juan, 1 T. Calde Âs, 2 A. Sa Ânchez-Pernaute, 3 J. Cerda Ân, 3 A.J. Torres, 3 J.L. Balibrea 3 and M. Benito 1 1 Department of Biochemistry and Molecular Biology, Faculty of Pharmacy, Complutense University, 28040 Madrid; and Services of 2 Immunology and 3 Surgery II, San Carlos Hospital, Madrid, Spain Several distinct genetic alterations have been associated with colorectal tumorigenesis. This study investigated the frequency of microsatellite instability, also known as replication error (RER), and loss of heterozygosity (LOH) at six chromosome regions in sporadic colorectal cancer (CRC). Eighty-six tumour and paired normal mucosa samples were included in the study. A polymerase chain reaction (PCR)-based technique was performed to analyse six (CA)n dinucleotide repeats located near or within regions containing important genes implicated in the complex process of colorectal tumor- igenesis (chromosomes 2p, 3p, 5q, 11p, 17p and 18q). Overall, LOH frequency was higher in RER  tumours (25/46, 54.3%) compared with RER + tumours (9/40, 22.5) (P = 0.04). To investigate prognostic implications, survival analysis was performed for 66 patients. Compared with RER  tumours, patients with RER + tumours at 2p, 3p, 5q, 11p or 18q were found to have an improved prognosis (overall survival, P = 0.02 and disease-free survival (DFS) P = 0.005) this variable being an independent prog- nostic factor by multivariate analysis (P = 0.001). Overall survival of patients whose tumours were LOH + was signi®cantly shorter compared with those without LOH (overall survival, P = 0.008 and DFS, P = 0.01). Thus, tumours displaying RER + and LOH + phenotype, as established by microsatellite analysis, show a diVerential prognosis. These data indicate that this may be a useful tool for the identi®cation of patients at diVerent risks aVected by CRC. # 1999 Published by Elsevier Science Ltd. All rights reserved. Key words: colorectal cancer, prognosis, replication error, loss of heterozygosity, microsatellite analysis Eur J Cancer, Vol. 35, No. 12, pp. 1676±1682, 1999 INTRODUCTION A series of genetic alterations involving both proto- oncogenes and tumour suppressor genes occur in colorectal tumorigenesis [1]. Thus, it is widely accepted that multiple genetic alterations aVecting proto-oncogenes, such as ras, and tumour suppressor genes, such as APC and TP53, are involved in the development and/or progression of colorectal cancer [1, 2]. Genome-wide searches for loss of heterozygosity (LOH) have been performed successfully to localise tumour sup- pressor gene loci [3, 4]. The observation that LOH may have prognostic implications has created a situation in which the study of this abnormality provides information signi®cant enough to suggest that, for clinical purposes, tumours of certain types should be typed routinely for LOH at speci®c loci [5]. In the past few years, a new potential mechanism that could increase the usually rare rate of spontaneous mutations has been described. This potential `mutator' defect has been referred to as either microsatellite instability (MSI) or the replication error (RER) phenotype [6]. This mechanism involves alterations on short tandem repeat DNA non-coding sequences (microsatellites). Although the function of these sequences is not yet clear, some researchers have speculated that such tandem repeats may be targets for certain proteins, which play a major role in the regulation of gene expression and DNA recombination [7, 8]. Moreover, four diVerent mutant genes, all homologous to bacterial DNA repair genes, have been associated with the RER phenotype [9±11]. European Journal of Cancer, Vol. 35, No. 12, pp. 1676±1682, 1999 # 1999 Published by Elsevier Science Ltd. All rights reserved. Pergamon Printed in Great Britain PII: S0959-8049(99)00158-6 0959-8049/99/$ - see front matter 1676 Correspondence to M. Benito, e-mail: benito@eucmax.sim.ucm.es Received 22 Jan. 1999; revised 17 May 1999; accepted 12 Jun. 1999.