Prognostic significance of free gastrointestinal tumor cells in peritoneal lavage detected by immunocytochemistry and polymerase chain reaction.

AIMS: The aim of our study was to identify tumor cells in peritoneal lavage comparatively with immunocytochemistry (ICC) and half-nested reverse transcriptase-polymerase chain reaction (RT-PCR) using carcinoembryonic antigen (CEA) as marker and to evaluate their prognostic significance. PATIENTS AND METHODS: In 75 patients who underwent surgery for a carcinoma of the colorectum (n=49), stomach (n=17) or pancreas (n=9) and 13 patients with an abdominal aortic aneurysm (control group) the abdomen was irrigated with saline solution immediately after laparotomy. Cells were separated by Ficoll-density centrifugation and divided into 2 equal volumes for ICC and RT-PCR. For ICC cells were spun onto slides by cytospin centrifugation and stained with a monoclonal antibody (mab) against CEA using the APAAP method. For RT-PCR total RNA was extracted from the cells, transcribed into cDNA and amplified with CEA-specific primers. Lavages of 13 patients with an abdominal aortic aneurysm and blood samples of 6 healthy donors served as controls. RESULTS: Immunostained tumor cells were found in peritoneal lavage in 23% (17/75) of all patients, whereas 63% (47/75) of patients gave a positive result by RT-PCR analysis. In the control group (n=13) no patient presented with tumor cells in ICC, however 5 of 13 (38%) showed amplified CEA-mRNA by RT-PCR, and so did one of six blood samples. Using ICC technique, we found significant correlations between detection rates and pT-, pN-, pM-categories as well as tumor stage. On the contrary, by RT-PCR significant correlations were observed only between pT- and pM-categories and detection rates. Detection of tumor cells in peritoneal lavage with both techniques was associated with poor prognosis. Moreover, these tumor cells are an independent prognostic factor and may have an influence on the development of peritoneal carcinomatosis. CONCLUSION: ICC is a useful method for detection of tumor cells in peritoneal lavage. In contrast, half-nested RT-PCR cannot be recommended, as the detection rates are unproportionally high, obviously as a result of CEA-mRNA expression in nontumor cells.

Apoptosis in rectal cancer: prognostic significance in comparison with clinical histopathologic, and immunohistochemical variables.

PURPOSE: The aim of this study was to evaluate the prognostic value of the apoptotic index for recurrence and disease-free survival after curative surgery for rectal cancer, particularly in relation to clinicopathologic variables, p53- and bcl-2 expression. METHODS: Formalin-fixed, paraffin-embedded tissue samples of rectal carcinomas resected curatively within a five-year period were used (N = 160). Apoptotic cells with fragmented DNA were detected by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphatase-biotin nick-end-labeling method. The ratio of apoptotic tumor cells (in percent) was classified into low apoptotic index (less than 10 percent) and high apoptotic index (10 percent or more). Immunohistochemical analysis was performed using monoclonal antibodies (DO-1 for p53 and clone 124 for bcl-2). Statistics included univariate and multivariate analysis, and survival was calculated using the Kaplan-Meier method. RESULTS: Seventy-five percent of tumors showed a low apoptotic index, and 25 percent had a high apoptotic index. No correlation was found between apoptotic index and International Union Against Cancer stage (P > 0.05). However, significant correlations were documented with histologic differentiation (mean apoptotic index, 5.74 percent in moderately vs. 3.98 percent in poorly differentiated carcinomas; P = 0.0173), lymph node involvement (mean apoptotic index, 6.11 percent in pN1 vs. 3.72 percent in pN2; P = 0.0074), p53 status (mean apoptotic index, 6.26 percent in p53- vs. 4.42 percent in p53+; P = 0.0085), and bcl-2 expression (mean apoptotic index, 5.13 percent in bcl-2- vs. 6.51 percent in bcl-2+; P = 0.0418). Tumors of the lower rectum had a lower apoptotic index than those of the upper rectum (P = 0.0277). Neither univariate nor multivariate analysis assessed apoptotic index as predictor of prognosis: Recurrence rates did not differ between tumors related to apoptotic index (22 percent with low apoptotic index vs. 15 percent with high apoptotic index; P > 0.05), and no significant differences were found regarding survival (P > 0.05). On multivariate analysis, International Union Against Cancer stage (P = 0.0002), p53 (P = 0.0002), gender (P = 0.0136), and bcl-2 (P = 0.0243) were independent predictors of recurrence. These variables, except for bcl-2, were also independently related to disease-free survival. CONCLUSIONS: Reflecting tumor biology, apoptotic index as single variable showed no prognostic significance, whereas p53 was an independent predictor for both recurrence and survival, and bcl-2 was independently related to recurrence, but not to survival. Clinically, International Union Against Cancer stage and gender were independent prognostic factors after curative surgery for rectal cancer.

p53 and Bcl-2 as significant predictors of recurrence and survival in rectal cancer.

The aim of this study was to evaluate the prognostic value of p53 nuclear accumulation and Bcl-2 expression after curative surgery for rectal cancer. Immunohistochemistry was performed using monoclonal antibodies (MAb) (DO-1 for p53; anti-human Bcl-2 MAb, clone 124, for Bcl-2) on formalin-fixed, paraffin-embedded tissues of 160 rectal carcinomas (UICC stages I-III), and results were compared with data from the prospective registry of rectal cancer by univariate and multivariate logistic regression model focusing specifically on recurrence. Survival was calculated by the Kaplan-Meier method and proportional hazards model. p53 nuclear accumulation was documented in 39% (n=63) of tumours and was associated with a higher incidence of tumour progression (local or distant recurrence) and poorer disease-free survival (P<0.0001). Bcl-2 expression was detected in 29% (n=47), and was associated with longer disease-free survival and lower incidence of recurrence (P<0.0086). Multivariate logistic regression analysis demonstrated that gender (P=0.0136), UICC stage (P=0.0002), p53 expression (P=0.0002) and Bcl-2 expression (P=0. 0243) were independent factors predictive of recurrence. The proportional hazards model identified p53 (P=0.0009), UICC stage (P=0.0480), gender (P=0.0049), but not Bcl-2 (P=0.1503), as independently related to disease-free survival. Looking at the p53/Bcl-2 subgroups, the poorest prognosis was observed in the p53+/Bcl-2- subgroup, whereas patients whose tumours were p53-/Bcl-2+ had the best prognosis (P<0.0001). Immunohistochemical assessment of both p53 and Bcl-2 status may be valuable in predicting recurrence and survival after curative surgery for rectal cancer. Therefore, they play a role as prognostic factors in rectal cancer. p53 is a stronger predictor of prognosis than Bcl-2.