BCl-2 EXPRESSION IS ASSOCIATED WITH IMPROVED PROGNOSIS IN PATIENTS WITH DISTAL COLORECTAL ADENOCARCINOMAS Upender MANNE 1 , Heidi L. WEISS 2 and William E. GRIZZLE 1 * 1 Department of Pathology, University of Alabama at Birmingham, Birmingham, Alabama, USA 2 Medical Statistics Section, Comprehensive Cancer Center, University of Alabama at Birmingham, Birmingham, Alabama, USA Several recent studies have suggested that the anatomic location of the tumor and ethnicity of the patient should be considered in the evaluation of prognostic markers of colo- rectal neoplasia. The phenotypic expression of Bcl-2 has been reported to be a useful prognostic marker in colorectal ad- enocarcinoma (CRC). However, its prognostic importance in CRCs based on their anatomic location and on the ethnicity of the patients has not been reported. Therefore, we evalu- ated Bcl-2 expression by immunohistochemistry in CRCs collected from 107 African-American and 149 Caucasian pa- tients from a southern U.S. population. In univariate Kaplan- Meier survival analyses, Bcl-2 expression was associated with better overall survival of both African-American (log-rank, p  0.040) and Caucasian (log-rank, p  0.032) patients with distal but not with proximal CRCs. In multivariate Cox re- gression analyses, when the pathologic features of tumors were not included, the expression of Bcl-2 was associated with better survival in either ethnic patient populations with distal CRCs after adjusting for other confounding variables and p53 nac status; however, it was not significant in either race when tumor stage was included in multivariate analyses. Thus, these studies suggest that the expression of Bcl-2 in CRCs is a valuable indicator of good prognosis in either race when CRCs are located in the distal colorectum. Also, these studies suggest that the expression of Bcl-2 is useful in deter- mining prognosis before pathologic-clinical staging and it can aid in selection of treatment after evaluation of diagnostic biopsy specimens of patients with distal colorectal adenocar- cinomas. Int. J. Cancer (Pred. Oncol.) 89:423– 430, 2000. © 2000 Wiley-Liss, Inc. Currently, the pathologic stage of primary colorectal adenocar- cinomas (CRCs) is used to predict the clinical outcomes of pa- tients. Nevertheless, the pathologic stage of a CRC may not be a perfect indicator of clinical outcome, as even a group of patients with tumors of identical stage have different responses to treat- ment; thus, there may be specific identifiable subgroups of patients with poorer clinical outcomes. Also, it would be useful to predict clinical outcome before definitive surgery and without complete knowledge of the tumor stage. Thus, recent studies have focused on identifying subgroups of patients with more aggressive CRCs and in predicting the clinical outcome of CRC patients using molecular markers. The genetic lesions that occur during tumorigenesis of CRC (and other cancers) are influenced by several external and internal factors including genetic and ethnic background of patients (re- viewed in Neuhausen, 1999; Grizzle et al., 1999). One notable example is the prognostic importance of abnormalities in p53 in CRCs (reviewed in Manne et al., 1997a; Grizzle et al., 1999; McLeod and Murray, 1999). Recently, we reported that the prog- nostic significance of nuclear accumulation of p53 (p53 nac ) varied with the ethnic group of the patient and the anatomic location of the CRCs (Manne et al., 1998). The usefulness of p53 nac as an indicator of poor prognosis was limited to Caucasians, specifically to patients with adenocarcinomas located in the proximal colon; however, p53 nac was not useful as a prognostic marker in Cauca- sian patients with distal CRCs or in African-American patients with CRCs from either the proximal or distal colorectum. Thus, the usefulness of specific molecular markers in predicting clinical outcome might vary among specific subgroups of patients with CRCs based on the anatomic location of the tumor and the eth- nicity of patients. Furthermore, in a recent review, Neuhausen (1999) described the importance of ethnic differences in cancer risk and in the underlying genetic variations and suggested that the knowledge gained by studying the effect of a single, frequent genetic alteration in a well-defined population can be applied to larger populations and the findings can be useful in designing effective prevention and treatment strategies. Several oncogenes and tumor suppressor genes such as bcl-2 and p53 are involved in regulating programmed cell death and cellular proliferation. The dysregulation and/or alteration of bcl-2 and p53 genes have been identified during the development and progression of CRCs (Hague et al., 1994; Bosari et al., 1995; Sinicrope et al., 1995b; Baretton et al., 1996; Hao et al., 1997; Yang et al., 1999). In addition, lack of Bcl-2 expression was correlated with tumor local invasion, metastasis and recurrence in CRC (Ilyas et al., 1998; Ishijima et al., 1999; Giatromanolaki et al., 1999). Patients with CRCs who exhibit high levels of Bcl-2 expression have been reported to have a good clinical outcome (Sinicrope et al., 1995a, 1999; Ofner et al., 1995; Baretton et al., 1996; Manne et al., 1997a; Kaklamanis et al., 1998; Buglioni et al., 1999); however, some studies have not observed such an association (Bosari et al., 1995; Schneider et al., 1997; Tollenaar et al., 1998; Giatromanolaki et al., 1999). Conversely, in a small group of CRC patients (n = 48), Bhatavdekar et al. (1997) corre- lated Bcl-2 expression with poor prognosis. Although the reasons for this controversy are not known, we hypothesize that the prog- nostic importance of Bcl-2 expression in CRCs may be limited to specific subgroups of patients similar to the clinical usefulness of p53 nac in CRC (Manne et al., 1998). In order to identify high-risk subgroups of patients based on the anatomic location of tumor in the 2 major ethnic groups of the United States, we evaluated primary CRCs from 107 African- American and 149 Caucasian patients for the expression of Bcl-2. All these tumors were analyzed previously for p53 nac (Manne et al., 1998). As shown in a lymphoma cell line (Wang et al., 1993) and in a mammary epithelial cell line (Haldar et al., 1994), Bcl-2 and p53 oncoproteins are interrelated; thus, a better understanding of tumor behavior or aggressiveness may be gained by examining the co-expression of Bcl-2 and p53 nac . Therefore, we correlated the concomitant expression of Bcl-2 and p53 nac with patient survival. MATERIAL AND METHODS Tissues Formalin-fixed paraffin-embedded tissue blocks of CRCs from 204 African-American and 300 Caucasian (non-Hispanic) patients were collected randomly from files of the University of Alabama at Birmingham (UAB) Hospital and the affiliated Birmingham Veterans Administration (VA) Hospital. The selection of blocks was restricted to patients with “first primary” CRCs resected between 1981 and 1993 and to the patients for whom the clinical outcome data were available (Manne et al., 1998). For our study, due to limitations of resources, subgroups of 107 African-Ameri- *Correspondence to: William E. Grizzle, Department of Pathology, University of Alabama at Birmingham, University Station, Birmingham, AL 35294. Fax: (205) 975-7128. E-mail: grizzle@path.uab.edu Received 22 February 2000; Revised 20 April 2000 Int. J. Cancer (Pred. Oncol.): 89, 423– 430 (2000) © 2000 Wiley-Liss, Inc. Publication of the International Union Against Cancer