Pierantonio Bevilacqua

Tumor microvessel density, p53 expression, tumor size, and peritumoral lymphatic vessel invasion are relevant prognostic markers in node-negative breast carcinoma.

PURPOSE To determine the absolute and relative value of microvessel density (MVD), p53 and c-erbB-2 protein expression, peritumoral lymphatic vessel invasion (PLVI), and conventional prognosticators in predicting relapse-free (RFS) and overall survival (OS) rates in patients with node-negative breast carcinoma (NNBC). PATIENTS AND METHODS We monitored 254 consecutive patients with NNBC for a median of 62 months. Intratumoral MVD was measured after microvessels were immunostained using anti-CD31 antibody. p53 and c-erbB-2 protein and hormone receptors were also determined immunocytochemically. Results were analyzed by both univariate and multivariate statistical analysis. RESULTS Univariate analysis showed that MVD was significantly predictive of both RFS (odds ratio [OR], 8.30; P = .0001) and OS (OR, 4.50; P = .012) when tested as a continuous or dichotomous variable. Likewise, tumor size (OR, 3.16; P = .0012), PLVI (OR, 4.36; P = .0009), estrogen receptor (ER) status (OR, 2.35; P = .016), progesterone receptor (PR) status (OR, 2.00; P = .017), and expression of p53 protein (OR, 2.82; P = .004) were significantly associated with RFS. Tumor size (OR, 3.80; P = .0038) and expression of p53 protein (OR, 2.58; P = .024) were significantly associated with OS by univariate analysis. Multivariate analysis showed that MVD (P = .0004), p53 protein expression (P = .0063), tumor size (P = .0144), and PLVI (P = .0033) were all significant and independent prognostic factors for RFS. However, only tumor size (P = .004) and MVD (P = .047) were independent predictors for OS. c-erbB2 expression was not associated with outcome by either univariate or multivariate analysis. CONCLUSION MVD, p53 expression, PLVI, and tumor size are independent prognostic indicators of recurrence, which are useful in selection of high-risk NNBC patients who may be eligible to receive adjuvant therapies.

Prognostic value of intratumoral microvessel density, a measure of tumor angiogenesis, in node-negative breast carcinoma — results of a multiparametric study

SummaryIn the present study we update previous results on the prognostic value of intratumoral microvessel density (IMD), determined immunocytochemically using the monoclonal antibody CD-31 and a standard streptavidin-immunoperoxidase technique, published in theJ Clin Oncol 12:454–466, 1994. This study was undertaken in those 211 node-negative breast cancer (NNBC) cases of that series of which we had pathological material available to determine all the prognostic indicators. The median period of follow-up has been extended to 78 and 80 months for relapse-free survival (RFS) and overall survival (OS), respectively, and new biological indicators (i.e. Ki-67 labeling and 67 kDa laminin receptor expression) were included in the analysis.The main results obtained are:i) a confirmation that IMD is not associated with the other biological markers studied, i.e. expression of p53 protein, c-erbB-2 protein, 67 kDa laminin receptor, and cell kinetics; IMD was weakly associated only with histological grade (p=0.053);ii) IMD remains a highly significant prognostic factor for RFS and OS (p<0.0001 and p=0.018, respectively) in univariate analysis;iii) in multivariate analysis on RFS, IMD (likelihood ratio test (LRT)=30.16; p<0.0001), 67 kDa laminin receptor (LRT=9.80; p=0.0017), the IMD/67 kDa laminin receptor interaction (LRT=8.62; p=0.0033), tumor size (LRT=8.56; p=0.0034), and p53 protein (LRT=4.96; p=0.025) are significant and independent prognostic indicators. For OS, only tumor size (LRT=8.34; p=0.0038), menopausal status (LRT=5.16; p=0.023), p53 protein (LRT=4.37; p=0.036), and IMD (LRT=4.05; p=0.044) retain a significant and independent prognostic value.The results of this study confirm the prognostic importance on RFS of the variables previously tested, but not of peritumoral lymphatic vessel invasion. A novel finding is that 67 kDa laminin receptor and the IMD/67 kDa laminin receptor interaction are also significant and independent variables. For OS, the results confirm that both IMD and tumor size are significant and independent variables. With prolonged follow-up the novel finding that emerges is the prognostic importance of menopausal status and p53 protein.This new information could be useful for a more accurate selection of high-risk NNBC patients who require careful follow-up and may benefit from adjuvant therapy.

Correlation of growth fraction by Ki-67 immunohistochemistry with histologic factors and hormone receptors in operable breast carcinoma.

SummaryBreast cancer tissue samples obtained from 147 Stage I and II patients were tested with the monoclonal antibody Ki-67 and avidin-biotin-peroxidase complex in frozen sections. The percentage of cells with nuclear staining ranged from 5% to 65%. The frequency of Ki-67 positivity was classified in five groups: 0% (45/147 = 31%); 5–9% (38/147 = 26%); 10–19% (15/147 = 10%); 20–39% (24/147 = 16%) and ⩾ 40% (25/147 = 17%). The mean value was 20%, median 18% with standard deviation of 14.5%. A significant positive correlation was observed between the presence of high Ki-67 nuclear staining rate with pathological tumor size (p = 0.003), histologic grading (p = 0.04), and axillary lymph node metastases (p = 0.009). An inverse significant correlation was found between Ki-67 and estrogen receptor expression (p < 0.001). No correlation was observed with progesterone receptor expression or menopausal status. The overall picture is of an inverse relationship between high growth fraction determined with Ki-67 antibody and tumor differentiation parameters. These correlations confirm those already reported by thymidine labeling index and flow cytometry methods.The proliferative rate determined with Ki-67 antibody may provide information regarding cell kinetics of breast carcinoma, potentially useful in identifying patients with a different clinical course in order to improve the therapeutic approach, by a rapid, practical and easily performed immunohistochemical method.

A multiparametric study on the prognostic value of epidermal growth factor receptor in operable breast carcinoma

SummaryEpidermal growth factor receptor (EGFR) is a potentially useful new biological prognostic and predictive indicator in human breast cancer. Additional research on EGFR is warranted to enhance our information on: i) the method of choice for its detection and quality control issues; ii) its association with novel pathobiological markers of prognosis; iii) its prognostic value in multivariate analysis; and iv) its capability to predict response to hormone therapy and, in the future, to biological treatments using antibodies against the specific receptor or its ligands.In the present study we update previous data on EGFR status, determined immunocytochemically, by prolonging the period of observation up to 5 years and by including, in the multivariate analysis, several new biological indicators. The main results obtained are: i) EGFR is weakly associated with Ki-67 score (p=0.073) and with p53 expression (p=0.06); ii) EGFR is a significant indicator for recurrence (p<0.01 and odds ratio of 2.82) but not for death (p=0.27 and odds ratio of 1.49); iii) the prognostic power of EGFR is enhanced when combined with the knowledge of S-phase fraction; and iv) in multivariate analysis on relapse-free survival, EGFR and S-phase fraction (likelihood ratio test=26.40; p<0.01), c-erbB-2 protein and p53 mutant protein expression (likelihood ratio test= 5.94; p=0.05), cathepsin D (likelihood ratio test= 9.78; p<0.01), and nodal status (likelihood ratio test= 7.32; p<0.01) are significant and independent prognostic factors in early-stage breast carcinoma.This new information could be of help for a more rational approach in the use of EGFR as a marker in future clinical research.

Clinical management of a case of recurrent apocrine gland carcinoma of the scalp: Efficacy of a chemotherapy schedule with methotrexate and bleomycin

Apocrine carcinoma of the skin is a rare tumor. Wide surgical excision with complete removal of the neoplasm is the standard therapy and this appears to offer the best chance of cure. Radiotherapy may be used in case of local relapse or regional lymph node involvement. Systemic chemotherapy has not proved to be effective in the treatment of this tumor. We report on a 46-year-old woman with a recurrent apocrine carcinoma of the scalp that had previously been treated with surgery, radiotherapy and chemotherapy (Al-Saraff schedule). The patient was responsive to a second-line systemic chemotherapy regimen consisting of a weekly combination of methotrexate and bleomycin, and achieved long-term progression-free survival.

Immunocytochemical Determination of Epidermal Growth Factor Receptor with Monoclonal EGFR1 Antibody in Primary Breast Cancer Patients

Epidermal growth factor (EGF) has been shown to be important in regulating the growth of breast cancer cells in vivo because of its mitogenic action on some breast cancer cell lines in vitro. Immunocytochemical analysis of EGF receptor (EGFr) was carried out on frozen sections in 134 primary breast cancer patients. Overall 68 of 134 (51%) of the tumors were EGFr positive. There was no correlation between EGFr positivity and menopausal status. Regarding the histopathological features, no significant correlations were observed between EGFr expression and tumor size, grading and lymph nodes status. Estrogen (ER) and progesterone (PgR) receptors were detected by an immunocytochemical assay and an equal distribution of EGFr was found regarding steroid hormonal receptors expression. Finally, there was only a positive trend between the proliferative activity of the tumors, as measured by Ki-67 antibody, and the amount of EGFr. Our results suggest the presence of a subclass of breast tumors, characterized by the absence of ER and/or PgR and the presence of EGFr, whose growth appears to be mediated by autocrine growth factors rather than by steroid hormones. The overall picture is that of an independent relationship between EGFr expression and the known prognostic factors in breast cancer.

Lack of prognostic significance of the monoclonal antibody Ki-S1, a novel marker of proliferative activity, in node-negative breast carcinoma

In a series of 205 node-negative breast cancers (NNBC), we determined staining by the novel antibody Ki-S1, a marker of tumor cell proliferation, in order to test its association with other prognostic variables and its prognostic significance. Ki-S1 was determined in routinely formalin-fixed paraffin-embedded tumor samples. Ki-S1 gave a nuclear staining in the majority of the carcinomas (188 of 205), with percentages of reacting nuclei ranging from 2% to 90% (median value of 7%). In 107 tumors frozen sections were available to also assess the Ki-67 antibody. Among these, 94 had a nuclear staining of cancer cells ranging from 5% to 80% (median value of 7%). In 46 tumors we also determined the MIB-1 antibody. The percentage of MIB-1 nuclear staining ranged from 1% to 50% (median value of 20%). There was no significant relationship between Ki-S1 and the other two cell kinetic markers. Ki-S1 labeling was significantly associated only with tumor size (p = 0.03).With a median follow-up of 6 years, Ki-S1 had no significant prognostic value for either relapse-free survival (RFS) or overall survival (OS)(Ki-S1 as continuous logarithmic variable; p = 0.86 and p = 0.23, respectively). For RFS the following variables had a significant prognostic value: Ki-67 (≤ 10% vs > 10%; p = 0.037); progesterone receptor (PgR) expression (− vs +/++; p = 0.041); tumor size (pT1 vs pT2−3; p = 0.042) and grading (GI vs GII−III; p = 0.047). For OS, tumor size (p = 0.0044), age (continuous variable; p = 0.0060), and Ki-67 (p = 0.043) were significantly prognostic.In multivariate analysis (final model), only tumor size retained a significant and independent prognostic value for RFS (p = 0.0042). For OS, both tumor size (p = 0.0029) and age (≤ 55 years vs > 55 years; p = 0.041) retained significance in the multivariate model.In conclusion, Ki-S1 does not seem to have prognostic relevance in this series of NNBC. Possible hypotheses to explain this observation are discussed.

Peritumoral lymphatic vessel invasion compared with DNA ploidy, proliferative activity, and other pathologic features as prognostic indicators in operable breast cancer

SummaryIn 164 breast carcinomas the presence of peritumoral lymphatic vessel invasion (PLVI) was evaluated and correlated with other known indicators of prognosis and with the clinical outcome of the patients. Overall 22% of tumors were PLVI-positive. The presence of PLVI was significantly associated with axillary node involvement (p<0.0001) and tumor size (p=0.005), and tended toward an association with grading (p=0.065). No significant association was found between PLVI and steroid hormone receptors, DNA ploidy, or proliferative activity. Univariate analysis shows that peritumoral vessel invasion was significantly associated with a higher risk of recurrence (p=0.012) and with a trend toward shorter survival (p=0.074). Besides the presence of PLVI, prognosis was significantly worse also for patients with high proliferative aneuploid tumors and with axillary node metastases. Moreover, within the subsets of patients generally considered to have good prognosis, the presence of PLVI identified patients with a trend for higher risk such as those with PLVI-positive diploid tumors, PLVI-positive low-proliferative tumors, and PLVI-positive node-negative tumors. Adopting multivariate analysis, PLVI failed to retain prognostic importance when adjusted for node status, DNA ploidy, and proliferative activity.In conclusion, we found that the presence of PLVI has prognostic significance when singly evaluated. Multivariate analysis shows that PLVI is not an independent prognostic factor in stage I–II breast cancer.

Progesterone receptor determined by immunocytochemical and biochemical methods in human breast cancer

SummaryImmunocytochemical assay (ICA) of the progesterone receptor (PgR) was performed on 152 patients with stage I–II breast cancer. We employed the rat monoclonal antibody KD-68 and a peroxidase/antiperoxidase displaying system. The results obtained by ICA (PgRICA) were compared with those by the biochemical dextrancoated charcoal assay (PgRDCC). Comparing the two methods we found an overall agreement (accuracy) of 77.5%, a PgRICA sensitivity of 83.5% and a specificity of 73%. Both methods were significantly associated with oestrogen receptor expression, detected by DCC (P<0.001 for PgRDCC andP=0.0014 for PgRICA). No significant association was found between PgRICA or PgRDCC and the other clinicopathological features analysed. After a median follow-up of 36 months, the overall survival probability was 91% in PgRDCC-positive versus 81.5% in PgRDCC-negative patients (log-rank test, χ2=0.91) compared to 87.5% in PgRICA-positive versus 82% in PgRICA-negative ones (log-rank test, χ2=0.93). Disease-free survival probability was 74.5% in both PgRDCC-positive and PgRDCC-negative patients (log-rank test, χ2=0.02) compared to 78% in PgRICA-positive versus 71.5% in PgRICA-negative cases (log-rank test, χ2=0.37). The present study demonstrates that ICA is a reliable method to detect PgR, correlating well with the DCC assay. Moreover, the ICA assay seems to provide clinical information complementary to the biochemical method. The definition of its prognostic value in operable breast cancer needs additional studies, particularly in node-negative patients.

Immunocytochemical detection of progesterone receptor by monoclonal KD-68 antibody in operable breast cancer: Correlations with biochemical assay, pathological features and cell proliferative rate

A new immunocytochemical assay (ICA) for progesterone receptor (PgR), employing the rat monoclonal KD-68 antibody and a sensitive peroxidase-anti-peroxidase (PAP) technique as the displaying system, was performed in 129 human breast cancer specimens. PgR-ICA staining was almost all electively located in neoplastic cell nuclei with a substantial heterogeneity in distribution and intensity. To study the basic relationship of the results of the ICA method with the biochemical dextran-coated charcoal (DCC) assay we compared, in all the same specimens, the antibody nuclear staining with the PgR positivity by DCC (cut-off value of 10 fmol/mg of protein). We found an overall agreement of 77% between the two methods and a PgR-ICA sensitivity of 83% and a specificity of 72%, assuming that biochemical PgR is truth. PgR-ICA false-negative results were only nine out of 53 (17%); and false-positive were 21 out of 76 (28%). Using both methods no significant association was observed between PgR positivity with menopausal status, histological type, tumor size and lymph node status. The correlations between PgR expression and cell kinetics were assessed by an immunocytochemical method employing the monoclonal Ki-67 antibody. While a significant negative relationship was found between high Ki-67 score and PgR-ICA positivity (P less than 0.01) no correlation was found with DCC positivity. The present results demonstrate that ICA is a practical, reliable and inexpensive method with a good correlation to the conventional biochemical assay to determine the PgR status. Moreover, ICA recognizes PgR expression at the single cell level, thus providing additional information to the quantitative DCC assay that should improve the prognostic evaluation and the prediction of responsiveness to endocrine therapy in breast cancer.