A multicenter phase III prospective randomized trial of high-dose epirubicin in combination with cyclophosphamide (EC) versus docetaxel followed by EC in node-positive breast cancer. GOIM (Gruppo Oncologico Italia Meridionale) 9902 study.

BACKGROUND: The Gruppo Oncologico Italia Meridionale 9902 trial compared four cycles of high-dose epirubicin plus cyclophosphamide (EC) with four cycles of docetaxel (Taxotere, D) followed by four cycles of EC as adjuvant treatment of node-positive breast cancer. PATIENTS AND METHODS: Patients were randomly assigned to EC (E 120 mg/m(2), C 600 mg/m(2), arm A) for four cycles or four cycles of D (100 mg/m(2)) followed by four cycles of EC (arm B), both regimens every 21 days. Hormone receptor-positive patients were given hormonal therapy for 5 years. Primary end point was 5-year disease-free survival (DFS). Secondary objectives were overall survival (OS) and safety. RESULTS: There were 750 patients enrolled. With a median follow-up of 64 months, 5-year DFS was 73.4% in both arms, and 5-year OS was 89.5% versus 90.7% in arm A and B [hazard ratio was 0.99 (95% confidence interval for DFS 0.75-1.31; P = 0.95)], respectively. Grade 3-4 toxicity was more common in arm B. CONCLUSIONS: This study did not show advantages from the addition of docetaxel to high-dose EC as adjuvant chemotherapy in node-positive breast cancer. The small sample size and low number of DFS events may have limited the ability to observe statistically significant difference between the two arms.

Attitude towards genetic testing for breast cancer susceptibility: a comparison of affected and unaffected women.

The objective of this study is to evaluate women's awareness and interest in genetic testing for breast cancer risk, to identify socio-demographic factors, to analyse the reasons for wanting or not wanting to be tested and finally to determine whether breast cancer patients and healthy women have different attitudes towards genetic testing. Consecutive series of 879 women without and with breast cancer participated in a 20-item self-completing questionnaire. Among breast cancer patients, 57% answered that they would definitely or probably accept being tested, compared with 84% of women without breast cancer. At the multiple logistic regression analysis only to have a diagnosis of breast cancer conditioned significantly the interest to have genetic testing. Surprisingly, a family history of breast cancer was found to have no significant impact. The most frequently cited reason for being interested in genetic testing was 'to learn about your children's risk'. Although women's awareness about breast cancer genes is inadequate, the interest in genetic testing is substantial and higher both in healthy women and in women with breast cancer. These results provide important indications for the development of educational strategies.

MMP-2, MMP-9, VEGF and CA 15.3 in breast cancer.

BACKGROUND: Matrix metalloproteinases (MMPs) are a family of extracellular matrix degrading proteinases. Owing to their matrix-degrading abilities and high expression in advanced tumours, MMPs were originally implicated in cancer progression, invasion and metastasis. PATIENTS AND METHODS: In this study, the correlation was determined between the expression of gelatinases (MMP-2 and MMP-9) in the sera of breast cancer patients from zymographic analysis and serum concentrations of VEGF and CA 15.3, before surgery and after 1 and 6 months; the association of both markers with clinicopathological features including histological type, stage of disease and estrogen (ER) and progesterone (PgR) receptors status were also analysed. In all, 88 breast cancer patients and 20 healthy women were involved in this study. RESULTS: No statistically significant correlation between pro MMP-2, pro MMP-9, VEGF and CA 15.3 serum levels was found (p>0.05). In breast cancer patients, a significant decrease of the pro MMP-2 serum expression 1 month after surgery with respect to serum levels before surgery (p=0.0008) was evident, as well as of CA 15.3 serum levels at baseline and after 1 month (p=0.017). Moreover a strong decrease of pro MMP-9 serum levels was found in 88 breast cancer patients after 1 month (p=0.028) and after 6 months (p =0.009) from surgery. On the other hand, no significant differences in the serum levels of VEGF, CA 15.3, pro MMP-2 or pro MMP-9 between 88 breast cancer patients preoperatively and 20 healthy women as controls were found. Our findings did indicate a significant positive association between higher preoperative levels of CA 15.3 and progression of disease (p=0.03), as well as a longer disease-free survival in patients who exhibited a decrease of serum pro MMP-9 expression compared to other biomarkers. No relationship between these four markers and the main clinical and pathological parameters was found. CONCLUSION: The present study failed to demonstrate any association between serum levels of MMPs, VEGF and CA 15.3 and well-known clinicopathological characteristics of breast carcinoma, while demonstrating the prognostic value of CA 15.3 and pro MMP-9 in the follow-up of breast cancer patients.

Randomized clinical trial of adjuvant fluorouracil, epirubicin, and cyclophosphamide chemotherapy for patients with fast-proliferating, node-negative breast cancer.

PURPOSE: The prospective applicability of new biologic tumor information to personalize adjuvant treatment of women with operable breast cancer remains to be demonstrated. The aim of the present study was to investigate whether patients with fast-proliferating, node-negative breast cancer could benefit from adjuvant chemotherapy with fluorouracil, epirubicin, and cyclophosphamide (FEC). PATIENTS AND METHODS: Beginning in November 1989, we analyzed the proliferative activity of primary tumors in a consecutive series of women with node-negative breast cancer to identify subgroups of patients with a worse prognosis and who were therefore suitable candidates for adjuvant systemic therapy. Proliferative activity was determined by means of the [3H]-thymidine incorporation assay using an autoradiographic technique. Women with fast-proliferating breast cancer ([3H]-thymidine labeling index, > 2.3%) were randomized to receive either six cycles of adjuvant FEC or no adjuvant therapy until disease progression. RESULTS: One-hundred twenty-five and 123 patients treated with radical surgery for pT1 to T2, N0, M0 breast cancer were randomized to the FEC and control arms, respectively. After a median follow-up of 70 months, 27 events (21.6%) were observed in the FEC arm and 39 (32.2%) in the control arm, with a significantly lower number of locoregional relapses in the FEC group. Five-year disease-free survival (DFS) was 81% in the FEC group and 69% in the control group (P <.02 by log-rank test). Cox multivariate analysis described the impact of adjuvant therapy with FEC on DFS as independent of the patients' main clinical-pathologic characteristics. CONCLUSION: FEC adjuvant polychemotherapy seems able to significantly improve the clinical outcome of patients with fast-proliferating, node-negative breast cancer.

Biomarkers predictive for clinical efficacy of taxol-based chemotherapy in advanced breast cancer.

BACKGROUND: Several studies looked for tumor biomarkers predictive for paclitaxel sensitivity but till now no reliable biomarkers are available. The aim of this study was to verify the potential predictive value of beta-tubulin III and IV, vascular endothelial growth factor-receptor (VEGFR-1), HER2/neu and microvessel density (mVD), in a group of 70 patients with advanced breast cancer (ABC) treated with paclitaxel-based chemotherapy. PATIENTS AND METHODS: Immunohistochemical analysis (ICA) of HER2, VEGFR-1, mVd, beta-tubulin III and beta-tubulin IV expression were performed in a series of 72 advanced breast cancer. Furthermore apoptotic fraction with TUNEL analysis was evaluated. RESULTS: beta-tubulin III ICA expression was predictive of progression after chemotherapy. In fact only 2% of the patients with low beta-tubulin III expression progressed after paclitaxel chemotherapy vs 38% of those with high beta-tubulin III tumor expression (P=0.000; by chi(2)). This evidence was confirmed by a logistic regression analysis (OR 28.789; 95% CI 3.212-258,072; P=0.004). There was not a significant association between other biomarkers' characteristics and clinical response to chemotherapy. A Cox multivariate analysis, with overall survival as a dependent variable, showed that only HER2 expression was independently associated (OR 2.39; 95% CI 1.09-5.23; P=0.03) with overall survival. CONCLUSIONS: We suggest that class III beta-tubulin immunohistochemical expression analysis could be a potentially relevant tumor biomarker for paclitaxel resistance in advanced breast cancer patients.

BRCA1 mutations and polymorphisms in a hospital-based consecutive series of breast cancer patients from Apulia, Italy.

INTRODUCTION: Hereditary breast cancer has been partly attributed to germline mutations in the BRCA1 gene that are deleterious for BRCA1 protein activity. This paper analyzes the incidence and characteristics of detectable BRCA1 mutations and polymorphisms in a hospital-based consecutive series of breast cancer patients from southern Italy to investigate the incidence and the association of these molecular alterations with breast cancer biology and family history. METHODS: One hundred cases with familial characteristics were selected from a consecutive series of 511 patients with a first diagnosis of breast cancer. DNA from peripheral blood was screened for whole BRCA1 gene mutations utilizing dHPLC as a pre-screening analysis and automatic DNA sequencing for the identification of specific alterations. RESULTS: In the overall series of 511 patients, 100 had a family history of breast cancer and were investigated for BRCA1 mutations. Two types of BRCA1 mutations were identified, 5382insC in six cases and 4566delA in one case. The 5382insC mutation was present in two out of six cases with ovarian cancer while 4566delA in one case of male cancer. The most frequent missense polymorphisms were E1038G, P871L, K1183R in exon 11, S1613G, M1652I in exon 16 and D1778G in exon 22. Confirming what found in previous studies, patients in whom pathological BRCA1 mutations were detected had early-onset breast cancer (p=0.05), positive nodal status (p=0.05), lower ER (p=0.02) and PgR (p=0.01) content. Interestingly, the K1183R polymorphism and, less strongly, S1613G polymorphism were associated to mutational risk (K1183R: OR 0.1 p=0.03; S1613G: OR 2.7 p=0.08). CONCLUSION: Mutations in the BRCA1 gene are frequent also in our consecutive series of patients from southern Italy. An association between two detected single nucleotide polymorphisms (SNPs) and BRCA1 mutational risk was ascertained. Finally, we confirm the fact that peculiar clinical-pathological features seem to characterize patients with a family history of breast cancer and BRCA1 alterations.

Heterogeneity of intratumour proliferative activity in primary breast cancer: biological and clinical aspects.

The present retrospective study was undertaken to verify whether the extent of intratumour proliferative activity variation or the method of quantifying tumour proliferative activity is related to biological characteristics and clinical outcome in a series of operable node-negative breast cancer patients. For tumour proliferative activity evaluation, the 3H-thymidine autoradiographic assay was used. After incubation of 3-8 samples from different areas of the equatorial section of each tumour for 1 h at 37 degrees C with 3H-thymidine, the following methods were used for evaluation of tumour cell labelling: mean tumour labelling index (LI), the highest labelling value from a specific area (LI-max), and the extent of intratumour labelling variation from several samples (LI-CV). LI-max was related to ER and PgR status, and linearly correlated with LI (c.c. = 0.92, P < 10(-6)) whereas LI-CV was independent of tumour size, grade ER and PgR status, but dependent on the number of tumour samples analysed for each tumour. After 5 years of median follow-up, disease-free survival was only related to tumour size (T1 versus T2: 84 versus 64%, P < 0.04 by log rank analysis) and different LI values (low versus high 3H-Tdr-LI:86 versus 61%, P < 0.03 by log rank analysis). LI-max and LI-CV values were not significantly related to clinical outcome. Cox multivariate analysis confirmed the independent prognostic value of LI and tumour size on disease-free survival.

Randomised clinical trial of adjuvant chemotherapy in patients with node-negative, fast-proliferating breast cancer.

A prospective randomised clinical trial in patients with node-negative, fast-proliferating breast cancer was initiated in January 1990 to verify the feasibility and reliability of a 3H-thymidine (3H-Tdr) autoradiographic assay in a prospective and consecutive series of node-negative patients and the therapeutic effect of adjuvant chemotherapy in patients with node-negative breast cancer and high tumour proliferative activity. Node-negative patients with a high 3H-Tdr Labelling Index (3H-Tdr-LI) were randomised to receive either no further treatment or combination chemotherapy consisting of fluorouracil, epirubicin plus cyclophosphamide (FEC) for 6 cycles. The autoradiographic assay was performed in 307 of 317 patients (97%) and was evaluable in 291 of 317 patients (92%). A total of 176 patients with a high 3H-Tdr-LI entered the clinical randomised study: 91 in the FEC arm and 85 in the control arm. Patient groups were fairly well balanced regarding the most important clinical and pathological characteristics. In total, 530 FEC cycles have been administered with an actual dose intensity of 90%. Patients receiving FEC demonstrated leucopenia in 35% of cases, alopecia in 70%, and loss of menses in premenopausal patients in an age-dependent manner. Patients are still being entered into the study.

Characterization of a h-3 GnRH method for the measurement of GnRH binding-sites in human breast-cancer and breast-cancer cell-lines.

The peptide hormone, GnRH has been hypothesized to play a direct antitumoral role in certain kinds of breast cancer. This direct effect is considered to function via high affinity membrane receptors to GnRH that have been demonstrated in tumors. However, no routine assay for GnRH receptors exist limiting further research in elucidating the mechanisms of GnRH action during treatment and both the clinical usefulness of this measure and the development or refinement of GnRH treatments. Part of the problem has been that previously reported procedures require expensive, difficult (for routine clinical laboratories) iodination of specific GnRH analogs. The use of iodinated substrates present the additional problems of radioactive waste disposal and safety of laboratory personal. Further, the use of different analogs complicates the comparison of data between laboratories. We present an assay that utilizes tritiated natural GnRH in order to assay the native receptor binding and reduce both safety problems and cost of assay/waste disposal. The use of nitrocellulose filters treated overnight with 1% BSA resolved the problems of high blank values. Kinetic data demonstrate that this assay is sensitive, accurate and give results comparable with other methods. We also show that the membranes derived from the tumor and the cell cultures may be frozen at -80 degrees C for up to 90 days without affecting the results of the assay.

Biological and clinical relevance of proliferative activity in inflammatory breast-cancer.

The hormone receptor status and proliferative activity characteristics of inflammatory breast cancer (IBC) were studied in a series of 46 patients. ER and PgR were measured by the DCC method and proliferative activity was by the H-3-thymidine autoradiographic labeling index (H-3-Tdr-LI). Tumors were ER and PgR positive in 42% and 38% of cases, respectively, whereas median H-3-Tdr-LI was 3.8%. With regard to clinical aspects, overall survival (OS) was not affected by either ER status (36 cases) or H-3-Tdr-LI value (33 cases). On the contrary, PgR(+) status was able to individualize women with a significantly higher probability of OS (X(2) by long rank test, p=0.03) after 35 months of follow-up. In the subgroup of 14 patients subjected to double biopsy performed before and after administration of primary polychemotherapy, the tumor proliferative activity variations were not related to clinical outcome.

Mucin-like carcinoma-associated antigen (mca) serum levels in breast-cancer - a critical-appraisal.

MCA serum levels were determined in 196 patients with benign breast disease (BD) and in 371 patients with breast cancer. MCA values were abnormal in 30.6% of the patients with BD. In breast cancer patients, significant increases were only found in metastatic disease (89.1%). MCA levels were not correlated with tumor size; on the contrary, the tumor marker showed a strict relation to axillary lymph node involvement. Moreover, the location of metastases and the number of sites involved seem to increase MCA concentrations. The MCA antigen seems to be a promising tool only for monitoring patients with advanced breast cancer.

Her-2/neu and fibrinolytic factors in human breast-cancer.

HER-2/neu status and t-PA, u-PA, and PAI-1 cytosol content were evaluated in 88 primary breast cancer patients to determine the relationships between these parameters. HER-2/neu was amplified in 24% (20/84) of tumor samples and overexpressed in 28% (14/50). In the overall series, median t-PA, u-PA and PAI-1 contents, measured by the enzyme-linked immmunoassay (ELISA), resulted in 1.7, 1.1 and 1.0 ng/mg cytosol protein (cyt prot), respectively. HER-2/neu overexpressed cases showed higher u-PA levels than those normally expressed whereas t-PA and PAI-1 levels did not vary in HER-2/neu altered and non altered cases. The t-PA levels did not differ in cases with or without HER-2/neu alterations, when separately considering the node-negative and node-positive cases. A significant relationship between t-PA levels and HER-2/neu alterations was observed only in the ER(+) tumors: t-PA levels were lower in amplified and overexpressed cases (1.4 versus 2.5 ng/mg cyt prot in amplified and single copy gene, respectively; 1.6 versus 2.3 ng/mg cyt prot in overexpressed and normally expressed cases, respectively). Therefore, t-PA and HER-2/neu could provide additional prognostic information for ER-negative patients.

HER-2/neu, c-Myc and cyclin-A in human breast cancer.

To better understand the role of HER-2/neu, c-myc and cyclin-A which seem to be activated in different steps of tumor cell growth, we analyzed a series of breast cancers from patients subjected to radical mastectomy with regard to HER-2/neu amplification (N=171) and expression of HER-2/neu (N=114), c-myc (N=31) and cyclin-A (N=71). Molecular evaluation demonstrated that HER-2/neu was amplified in 20% of cases and overexpressed in 27%, and its alterations were associated with a higher proliferative activity (H-3-Tdr-labeling index), although not statistically significant in patients without lymph node metastases, c-myc was overexpressed in 16% of cases and was weakly correlated to proliferation activity. Cyclin-A was overexpressed in 15% of cases and was significantly correlated to the percentage of the H-3-Tdr labeled cell fraction (p=0.002). Cyclin-A alterations were also significantly associated with well-differentiated tumors, suggesting that this gene could be involved in the control of the normal cell cycle rather than to cell proliferation during tumor growth.

Cytokeratins and proliferation in breast cancer patients.

Tissue polypeptide antigen (TPA) was studied in 242 sera and 165 tumor cell cytosols (both evaluations in 67 cases) of breast cancer patients, for which proliferative activity, determined by the TLI technique, was also available. The TPA serum and tumor cell cytosol median values (utilized for measure analysis as cut-off) were 70 U/1 and 377 U/mg cytosol protein, respectively. High serum TPA levels were associated with unfavourable clinicopathological characteristics whereas a higher tumor cell cytosol TPA level was associated with better cytohistological tumor differentiation. Furthermore, no overall correlation was found between serum and tumor cell cytosol TPA levels or between their levels and TLI. When analyzing cases in which serum and tumor cell cytosol TPA values were higher than 100 U/l and 500 U/mg cytosol protein, respectively (n = 28), serum TPA was positively associated with TLI (slope = 12.3 r = 0.55, p < 0.01), while cytosolic TPA resulted negatively associated with TLI (slope = -87.4 r = 0.41, p < 0.01). Finally, a strong inverse relationship between cytosolic and serum TPA (p < 0.0005) became evident. We suggest that TPA could represent a serum marker for tumor cell proliferation in specific patient subgroups with original high serum and/or cytosol TPA expression.

HER-2/neu in bladder carcinoma.

A total of 66 bladder cancer patients were studied to verify possible relationships between HER-2/neu alterations and pathological characteristics, and to define a poor prognosis patient subgroup with respect to time to recurrence, time to progression and survival. Tumor and healthy tissue specimens were analyzed for HER-2/neu DNA amplification and protein overexpression by Southern and Western blot techniques and evaluated statistically. 13% of cases were amplified and 39% were overexpressed. HER-2/neu alterations were not significantly associated with pathological staging or tumor grading. Multifocal tumors had a higher percentage and overexpression with respect to monofocal tumors. Actuarial analyses did not show a significant statistical correlation between HER-2/neu amplification and overexpression and clinical outcome. Clinical evaluation of HER-2/neu status showed that this gene is not related to tumor relapse, progression or patient survival.

Fec (Fluorouracil, epirubicin, cyclophosphamide) plus granulocyte macrophage-colony-stimulating factor (gm-csf) in advanced or inflammatory breast-cancer - a dose-finding study.

Seven patients with inflammatory and 11 with metastatic breast cancer were treated with high dose FEC chemotherapy plus GM-CSF; 5-fluorouracil and cyclophosphamide were administered at 500 mg/m2/iv/day 1, epirubicin at three dose levels: 100 mg, 120 mg and 140 mg/m2/iv/day 1 every three weeks (six patients per level). GM-CSF was administered at a dosage of 5 mg/kg/sc from day 5 to 12 of each cycle. The overall response rate was 83% (95% CI: 66%-100%) with 22% complete response. The median response duration for patients with metastatic disease was 7 months (range: 4-10). The hematological toxicity was moderate but reversible due to GM-CSF rescue; mucositis represented the dose limiting toxicity. In conclusion, the increase of dose intensity resulted in a higher response rate but not longer response duration, which must be taken into account when administering high-dose chemotherapy with growth factor rescue.

Neoadjuvant chemotherapy with accelerated CNF plus G-CSF in patients with breast cancer tumors larger than three centimeters: a pilot study.

From February 1992 to November 1993, forty patients with operable breast cancer tumors larger than three centimeters were enrolled in this study of accelerated neo-adjuvant chemotherapy. Thirty-seven patients are evaluable: one patient was excluded from the protocol and two refused to continue treatment after the first cycle. Chemotherapy consisted of three presurgical cycles of CNF [cyclophosphamide at 600 mg/m2, mitoxantrone (Novantrone) at 10 mg/m2 and 5-fluorouracil at 600 mg/m2] administered every 2 weeks, plus G-CSF (5 microg/kg s.c./day on days 7-12). Twenty-six of 37 patients (70%) achieved objective tumor response and were submitted to quadrantectomy. Toxicity was easily manageable. After a median 55-month follow-up (range 48-70), no locoregional recurrences were observed. Distant metastases occurred in 12/37 (32%) patients. The five-year disease-free (DFS) and overall (OS) survival were 58% and 80%, respectively. Accelerated CNF plus G-CSF proved to be a safe and tolerable regimen yielding a good clinical response thereby increasing the possibility of breast conservation surgery for patients otherwise candidates for mastectomy.

The impact of body mass index and type 2 diabetes on breast cancer: current therapeutic measures of prevention.

Epidemiological data have suggested a possible relationship between obesity, diabetes mellitus and cancer risk, particularly breast cancer. We set out to investigate the effect of body mass index and diabetes mellitus on the presence of breast cancer in the Apulian population. We selected 1,663 women affected with primary breast cancer and 4,702 control patients. All patients with breast cancer underwent surgical excision of the tumor and their tumors were histologically confirmed. The prevalence of type 2 diabetes (8%) in the women affected by breast cancer was significantly higher than in the control group (5%) (p<0.05). The majority of the diabetic women affected by breast cancer had a BMI value >25, both in premenopause and in postmenopause. With respect to BMI, the non-diabetic patients with breast cancer in postmenopause showed the same pattern as the diabetic ones. Instead, among the women in premenopause a higher percentage (55%) of patients with a BMI <24.9 was found (p<0.01). In the Apulian population, the presence of both type 2 diabetes and elevated values of BMI (that is in a condition of hyperinsulinemia) were found to enhance the frequency of breast cancer.

Breast cancer: biological characteristics in postmenopausal type 2 diabetic women. Identification of therapeutic targets.

HYPOTHESIS: Epidemiological data have suggested a possible relationship between diabetes mellitus and cancer risk, particularly breast cancer. We set out to investigate the effect of diabetes mellitus on the expression of estrogen and progesteron receptors and on the proliferative activity of primary breast cancer. METHODS: We selected 77 diabetic women and 578 control patients all in post-menopause and diagnosed with primary breast cancer. All patients underwent surgical excision of the tumor and on the specimens were performed an assessment of estrogen receptor and progesteron receptor and proliferative activity assay by (3)H-Thymidine incorporation. RESULTS: Diabetic women showed a decreased proliferative activity, while having the same estrogen receptor and progesteron receptor status and mean cytoplasmic concentration of their receptors than control group. Insulin treated women had a lower proliferative activity than non-insulin treated ones. CONCLUSION: Hyperinsulinemia and hyperglicemia influence in negative way the proliferative activity of diabetic women, likely inducing the expression of transforming growth factor beta, despite the high serum levels of Insulin-like growth factor and estrogen.

MCA performance in preoperative breast cancer patients.

Mucin-like carcinoma-associated antigen (MCA) is a new tumor associated antigen expressed on breast cancer cells independent of histological type and recognized by a two-site sandwich EIA (Roche) using the Mab b-12 as capture and detection antibody and a MCA cut-off of 11 U/ml. MCA serum levels were determined in 409 breast cancer patients and in 100 pts with benign breast disease histologically confirmed and staged according to UICC. The sensitivity of MCA regarding the stage of disease was elevated only in metastatic patients (89%) and the MCA values of this group were significantly different compared to those of patients with early or locally advanced breast cancer. In the group of benign patients, the specificity was 81%. The correlation between MCA values and number of involved axillary lymph nodes showed significant differences in the O versus 4-10 node subgroup (p < 0.001), in O versus > 10 and in 1-3 versus 4-10 node subgroup (p < 0.05); elevated MCA sensitivity was found in the 4-10 and > 10 subgroups. In our experience, the determination of MCA is not useful for early diagnosis of breast cancer nor does it indicate the extent of lymph node involvement. With regard to the prediction of tumor recurrence in postoperative patients free of disease, MCA levels increased concomitantly with the clinical evidence of relapse. These findings suggest that the MCA assay is highly sensitive only in patients with metastatic breast cancer; therefore one of the most promising test application seems to be for monitoring the clinical course of advanced disease patients.