Decreased expression of CD44 splicing variants in advanced colorectal carcinomas

Abstract

CD44v6 expression appears to be associated with adverse prognosis and propensity for metastasis in patients with colorectal cancer. However, expression of CD44 variants in different tumour stages has been poorly characterised. CD44 variant expression was investigated in normal colonic mucosa (n=36), colorectal adenomas (n=15), carcinomas (n=62) and metastases (n=6) by reverse transcriptase–polymerase chain reaction (RT-PCR) and Southern blotting with exon-specific probes. High frequencies of CD44standard (CD44s) and CD44 epithelial (CD44e) were observed in normal and neoplastic tissue. CD44v2 was seen predominantly in adenomas (27%) and UICCI carcinomas (29%). CD44v5 expression was low in normal mucosa (3%), higher in adenomas and carcinomas (29–33%), independent of tumour stage. CD44v6 expression was low in normal mucosa (6%) and higher in adenomas (47%) and carcinomas (42%). Surprisingly, a significant decrease of CD44v6 was observed in metastatic primary tumours (8%) and metastases (17%) (UICCIV) (P≤0.05). Therefore, the concept of CD44v6 conferring metastatic potential to malignant cells cannot be supported by our data.

Introduction

Colorectal carcinoma is the second most frequent cause of cancer mortality in Western countries. The prognosis of the disease is largely dependent on tumour stage at the time of surgery and is poor when development of metastasis has occurred. There is increasing evidence that the expression of variants of the glycoprotein CD44 is related to the invasiveness and metastatic potential of tumour cells. CD44 is a transmembranous cell adhesion glycoprotein that plays a role in cell–matrix interactions[1]. It is known to be a major ligand for hyaluronate and to bind collagen, laminin and fibronectin. Various isoforms of CD44 are generated by alternative splicing of exons 6 (v1–2) to exon 14 (v10)[2]. The significance of variant CD44 expression for the metastatic capacity of tumour cells was first reported by Günthert and colleagues[3]. In transfection studies, they demonstrated that expression of a CD44 protein variant encoded by alternatively spliced CD44 exon v6 conferred metastatic potential to a rat pancreas carcinoma cell line. Overexpression of CD44v6 and other variant exons in colorectal cancer and other gastrointestinal tumours was demonstrated in immunohistochemical studies4, 5Northern blots or reverse transcriptase–polymerase chain reaction (RT–PCR)6, 7, 8and in situ hybridisation9, 10. Data about CD44v6 expression in colorectal neoplasms depending on expansion of the tumour have been inconsistent. In most of these studies, however, stratification according to stage of the tumours was not carried out.

From a clinical point of view, an association of CD44v6 expression with poor prognosis in colorectal cancer patients has recently been observed. Mulder and coworkers[11]determined the CD44v6 status of primary tumours of 68 patients with colorectal cancer by immunohistochemistry and correlated CD44v6 status with survival. They found a significantly worse prognosis for patients with CD44v6 positive tumours compared with patients with negative tumours. A critical point in this study is that patients were not stratified according to stage of tumour. Thus, the question arose whether CD44v6 was an independent prognostic parameter in these patients. The aim of our study was to investigate the expression of CD44 variants in colorectal neoplasia in relation to tumour stage. CD44v5, another variant conferring metatatic potential to tumour cells in animal models and CD44v2, a variant first described in bladder cancer[12], were also investigated.