Cytokeratin-positive Cells In Bone Marrow Research Articles

Perioperative Activation of Disseminated Tumor Cells in Bone Marrow of Patients With Prostate Cancer

The outcome of prostate cancer is highly unpredictable. To assess the dynamics of systemic disease and to identify patients at high risk for early relapse we followed the fate of disseminated tumor cells in bone marrow for up to 10 years and genetically analyzed such cells isolated at various stages of disease. Nine hundred bone marrow aspirates from 384 patients were stained using the monoclonal antibody A45-B/B3 directed against cytokeratins 8, 18, and 19. Log-rank statistics and Cox regression analysis were applied to determine the prognostic impact of positive cells detected before surgery (244 patients) and postoperatively (214 patients). Samples from primary tumors (n = 55) and single disseminated tumor cells (n = 100) were analyzed by comparative genomic hybridization. Detection of cytokeratin-positive cells before surgery was the strongest independent risk factor for metastasis within 48 months (P < .001; relative risk [RR], 5.5; 95% CI, 2.4 to 12.9). In contrast, cytokeratin-positive cells detected 6 months to 10 years after radical prostatectomy were consistently present in bone marrow with a prevalence of approximately 20% but had no influence on disease outcome. Characteristic genotypes of cytokeratin-positive cells were selected at manifestation of metastasis. Cytokeratin-positive cells in the bone marrow of prostate cancer patients are only prognostically relevant when detected before surgery. Because we could not identify significant genetic differences between pre- and postoperatively isolated tumor cells before manifestation of metastasis, we postulate the existence of perioperative stimuli that activate disseminated tumor cells. Patients with cytokeratin-positive cells in bone marrow before surgery may therefore benefit from adjuvant therapies.

Expression of extracellular matrix metalloproteases inducer on micrometastatic and primary mammary carcinoma cells.

EMMPRIN (extracellular matrix metalloprotease inducer) is a glycosylated member of the immunoglobulin superfamily known to stimulate the production of matrix metalloproteases (MMPs) 1, 2, and 3 and MT1-MMP in peritumoral fibroblasts. We here evaluated whether EMMPRIN expression is related to tumor progression in human breast cancer. An immunohistochemical study using high-density tissue microarrays (n = 2222 breast cancer samples) and EMMPRIN-specific antibodies HIM6 and MEM-M6/1 was performed, and staining results were statistically correlated with various clinicopathological parameters. To analyze the putative association between EMMPRIN expression and bone marrow (BM) micrometastasis, an additional set of 55 breast tumors from patients with or without micrometastatic cells as determined with anti-cytokeratin antibody A45-B/B3 were included in our study. Cytokeratin-positive cells in BM were costained with EMMPRIN-specific antibody 1G6.2. Positive EMMPRIN staining correlated significantly with various histopathological risk factors (higher tumor grade, increased tumor size, negative estrogen receptor status and progesterone receptor status, and higher mitotic index) as well as decreased tumor-specific survival (log-rank, P = 0.0027). In particular, in patients > 50 years (i.e., postmenopausal women), EMMPRIN expression was an independent prognosticator as shown by Cox regression analysis (relative risk = 1.7, 95% confidence interval 1.4-4.3, P = 0.036). An involvement of EMMPRIN in tumor progression was also supported by the fact that it was expressed on approximately 90% of micrometastatic cells in BM. EMMPRIN expression in primary tumor predicts an unfavorable prognosis in breast cancer, suggesting a crucial role of EMMPRIN in progression of human mammary carcinomas.

The prognostic impact of cytokeratin-positive cells in bone marrow of patients with localized prostate cancer.

Our study evaluates the prognostic significance of the cytokeratin-positive mononuclear cells (CK+ cells) in the bone marrow (BM) and peripheral blood (PB) as detected by immunocytochemistry in patients with locoregionally confined prostate cancer. BM and PB samples were obtained from 66 newly diagnosed patients with T1-4pN0M0 prostate cancer. All samples were analyzed by standardized immunocytochemical methods (anticytokeratin mononuclear antibody; AE1/AE3) applying a negative immunomagnetic cell enrichment technique. A second sampling was obtained in 60 of the 66 patients >or=2 years after definitive radiotherapy. The median follow-up after high-dose radiotherapy of the patients was 65 months. For the analysis of the postradiotherapy clinical progression-free survival (PFS) treatment, failure was defined as pelvic tumor growth or development of distant metastases. At diagnosis CK+ cells were found in BM in 14 of 66 (21%) prostate cancer patients. This was not associated with an increased risk of progression. On the other hand, the presence of CK+ cells in 12 of 60 (20%) patients at the second BM aspiration was significantly related to a shorterPFS (p = 0.02). In the multivariate analysis, the presence of CK+ cells in the posttreatment BM did not remain as an independent variable of PFS assessment if posttreatment PSA was entered into the analysis. CK+ cells in PB were found in 12% of the patients. After therapy, none of the patients had detectable CK+ cells in PB. The presence of CK+ cells in the posttreatment but not in the pretreatment BM was associated with decreased PFS in patients irradiated for pelvis-confined nonmetastatic prostate cancer. Although this association was not retained in multivariate analysis, our observations indicate that the presence of CK+ cells after local therapy define a group of patients that have a high risk of developing distant metastases.

Cytokeratin-positive cells in preoperative peripheral blood and bone marrow aspirates of patients with colorectal cancer

Detection of cytokeratin-positive cells in bone marrow and peripheral blood may have prognostic significance in cancer patients. Furthermore, a correlation between uPAR expression on micrometastases and patient prognosis has been suggested. However, in patients with colorectal cancer, preoperative detection and characterization of tumour cells in bone marrow and peripheral blood, using an immunocytochemical approach, have not yet been substantiated as a prognostic tool. Methods: Forty-one bone marrow aspirates and 38 peripheral blood aspirates, obtained preoperatively from patients with colorectal cancer, were immunocytochemically screened for cytokeratin-positive cells. Where cytokeratinpositive cells were observed, an additional microslide was double immunostained for simultaneous detection of cytokeratin and uPAR/CD87. Results: Cytokeratinpositive cells were observed in 4 out of 41 bone marrow aspirates (10%). None of the isolated cytokeratin-positive cells expressed uPAR. In peripheral blood, no cytokeratin-positive cells were observed. Conclusion: The present study does not support the introduction of a routine immunocytochemical identification of cytokeratin-positive cells in preoperatively obtained bone marrow aspirates or peripheral blood from patients with colorectal cancer.

Detection of Cytokeratin-Positive Cells in Bone Marrow in Breast Cancer and Colorectal Carcinoma in Comparison with Other Factors of Prognosis

A prospective study is presented in which 293 patients suffering from breast cancer and colorectal carcinoma were analyzed for prognostic relevance of detected isolated disseminated tumor cells in bone marrow (IDTBM). The patients underwent surgery in the period from 1995 to 1997 and remained under observation until 1999. The monoclonal antibody A 45-B/B3 was used in the standard immuno-cytochemical method for detecting IDTBM, which represented an independent prognostic factor for survival time in patients with breast cancer or colorectal cancer. In breast cancer, when IDTBM were detected, the survival period was reduced by at least half. When disseminated tumor cells containing the A45-B/B3 antibody were detected in bone marrow, the risk of an earlier relapse of the tumor increased at least fourfold. In colorectal cancer, detection of IDTBM reduced survival time by a factor of 1.2-4.3. The risk of earlier relapse increased when disseminated tumor cells containing the A45-B/B3 antibody were detected in bone marrow by 2.8-8.1. Therefore, the use of IDTBM as an independent prognostic factor would provide an important method for determining the pathological stage of various cancers. Standardization of this technique into a generally accepted method would be especially desirable in treatment of patients with breast or colorectal cancer.

The presence of cytokeratin-positive cells in bone marrow increases the risk of relapse in patients with breast cancer

The presence of cytokeratin-positive cells in bone marrow increases the risk of relapse in patients with breast cancer

Cytokeratin-positive cells in bone marrow in comparison with other prognostic factors in colon carcinoma.

Despite the use of radical locoregional therapeutic methods and although conventional methods of diagnosis give no indication of metastases at the time of operation, distant metastases develop in approximately 50% of carcinoma patients within 5 years. While local relapses after the R0 resection of solid tumors are mainly a matter of concern for the surgeon, distant metastases can be traced back to the systemic dissemination of tumor cells at the time of operation. A prospective study is presented in which 145 patients suffering from colon carcinoma were analyzed for the prognostic relevance of isolated disseminated tumor cells detected in the bone marrow (IDT BM). The patients were operated on between 1993 and 1997 and subsequently observed until 1999. The monoclonal antibody A45-B/B3 was used with the immunocytochemical standard method for detecting IDT BM. For the purpose of cell cultivation, the cells were marked with the HEA-125 antibody and separated by means of magnetic cell sorting (MACS). In this investigation the presence of isolated disseminated tumor cells, as indicated by the A45-B/B3 antibody, proved to be an independent prognostic factor for survival time. The risk of an earlier death increased in node-negative and metastases-free patients with the detection of IDT BM by a factor of 12.60. The detection of IDT BM also represented an independent prognostic factor for the time until advancement of the tumor. The risk of an earlier relapse increased with the detection of disseminated tumor cells in the bone marrow containing the A45-B/B3 antibody by a factor of 18.02. A generally acknowledged standardization of the method is desirable. Due to the importance of the independent prognostic IDT BM factor, this method of ascertaining the pathological stage should be established at institutions of higher learning.

Cytokeratin-Positive Cells in the Bone Marrow and Survival of Patients with Stage I, II, or III Breast Cancer

Background Cytokeratins are specific markers of epithelial cancer cells in bone marrow. We assessed the influence of cytokeratin-positive micrometastases in the bone marrow on the prognosis of women with breast cancer. Methods We obtained bone marrow aspirates from both upper iliac crests of 552 patients with stage I, II, or III breast cancer who underwent complete resection of the tumor and 191 patients with nonmalignant disease. The specimens were stained with the monoclonal antibody A45-B/B3, which binds to an antigen on cytokeratins. The median follow-up was 38 months (range, 10 to 70). The primary end point was survival. Results Cytokeratin-positive cells were detected in the bone marrow specimens of 2 of the 191 control patients with nonmalignant conditions (1 percent) and 199 of the 552 patients with breast cancer (36 percent). The presence of occult metastatic cells in bone marrow was unrelated to the presence or absence of lymph-node metastasis (P=0.13). After four years of follow-up, the presence of micrometastases in bone marrow was associated with the occurrence of clinically overt distant metastasis and death from cancerrelated causes (P<0.001), but not with locoregional relapse (P=0.77). Of 199 patients with occult metastatic cells, 49 died of cancer, whereas of 353 patients without such cells, 22 died of cancer-related causes (P<0.001). Among the 301 women without lymphnode metastases, 14 of the 100 with bone marrow micrometastases died of cancer-related causes, as did 2 of the 201 without bone marrow micrometastases (P<0.001). The presence of occult metastatic cells in bone marrow, as compared with their absence, was an independent prognostic indicator of the risk of death from cancer (relative risk, 4.17; 95 percent confidence interval, 2.51 to 6.94; P<0.001), after adjustment for the use of systemic adjuvant chemotherapy. Conclusions The presence of occult cytokeratinpositive metastatic cells in bone marrow increases the risk of relapse in patients with stage I, II, or III breast cancer. (N Engl J Med 2000;342:525-33.)

Cytokeratin-positive cells in the bone marrow and survival of patients with stage I, II, or III breast cancer.

Cytokeratins are specific markers of epithelial cancer cells in bone marrow. We assessed the influence of cytokeratin-positive micrometastases in the bone marrow on the prognosis of women with breast cancer. We obtained bone marrow aspirates from both upper iliac crests of 552 patients with stage I, II, or III breast cancer who underwent complete resection of the tumor and 191 patients with nonmalignant disease. The specimens were stained with the monoclonal antibody A45-B/B3, which binds to an antigen on cytokeratins. The median follow-up was 38 months (range, 10 to 70). The primary end point was survival. Cytokeratin-positive cells were detected in the bone marrow specimens of 2 of the 191 control patients with nonmalignant conditions (1 percent) and 199 of the 552 patients with breast cancer (36 percent). The presence of occult metastatic cells in bone marrow was unrelated to the presence or absence of lymph-node metastasis (P=0.13). After four years of follow-up, the presence of micrometastases in bone marrow was associated with the occurrence of clinically overt distant metastasis and death from cancer-related causes (P<0.001), but not with locoregional relapse (P=0.77). Of 199 patients with occult metastatic cells, 49 died of cancer, whereas of 353 patients without such cells, 22 died of cancer-related causes (P<0.001). Among the 301 women without lymph-node metastases, 14 of the 100 with bone marrow micrometastases died of cancer-related causes, as did 2 of the 201 without bone marrow micrometastases (P<0.001). The presence of occult metastatic cells in bone marrow, as compared with their absence, was an independent prognostic indicator of the risk of death from cancer (relative risk, 4.17; 95 percent confidence interval, 2.51 to 6.94; P<0.001), after adjustment for the use of systemic adjuvant chemotherapy. The presence of occult cytokeratin-positive metastatic cells in bone marrow increases the risk of relapse in patients with stage I, II, or III breast cancer.

Does the immunocytochemical detection of epithelial cells in bone marrow (micrometastasis) influence the time to biochemical relapse after radical prostatectomy?

The detection of cytokeratin-positive bone marrow cells has been considered a prognostic factor in numerous malignant tumors. We investigated whether this was also valid for localized prostate cancer. Bone marrow aspirates were taken prior to radical prostatectomy from 169 consecutive patients with pT1/2 pNO G1-3 adenocarcinoma of the prostate. The immunocytochemical detection of cytokeratin no. 18 (CK 18)-positive cells using monoclonal antibody CK 2 was interpreted as micrometastasis. Repeat marrow aspirations were performed at 6 months postoperatively and once a year thereafter. The patients were re-examined over a period of at least 10 and a maximum of 72 months (median 32 months). An increase in prostate specific antigen > or = 0.5 ng/ml was considered a biochemical "relapse". One hundred and fifty-four patients had evaluable bone marrow aspirates, of which 74.7% were CK 18-negative and 25.3% positive. The latency period for biochemical relapse was 1481 days (median) in the CK 18-negative group and 1106 days (median) in the CK 18-positive group. This difference was not statistically significant. The CK 18-positive aspirates (n = 39) showed one positive cell in 20 cases, two positive cells in 8 and three or more positive cells in 11 cases. The preoperative number of cells had no statistically significant effect upon the onset of biochemical relapse. Only patients with three or more CK 18-positive cells tended to have a poorer prognosis. One hundred and thirteen patients had evaluable bone marrow aspirates pre- and postoperatively. Postoperative persistence or occurrence of CK 18-positive cells did not affect the outcome of the disease. The detection of CK 18-positive cells in bone marrow does not influence the prognosis of patients with localized prostate cancer within a period fo 32 months (median). Solely a subgroup of patients showing a large preoperative number of CK 18-positive cells seems to tend to an unfavorable course of the disease. Thus, further studies are necessary aiming at a more detailed characterization of these cells.

Clinical significance of micrometastasis in bone marrow of patients with gastric cancer and its relation to angiogenesis.

BACKGROUND: Detection of micrometastasis is an important problem of clinical significance for a better understanding and control of tumor progression, which will improve patients' survival time.METHODS: To identify micrometastases in bone marrow, an immunocytochemical assay for epithelial cytokeratin protein was performed in 106 patients with primary gastric cancer. Also, in 40 of the 106 patients, vascular endothelial growth factor (VEGF) expression and intratumoral vessel density were examined by an immunohistochemical staining method.RESULTS: Of the 106 patients, 22 (20.8%) presented with cytokeratin-positive cells in bone marrow at the time of primary surgery. The positive findings were related to depth of invasion, peritoneal dissemination, and liver metastasis. Patients with cytokeratin-positivity in bone marrow had a higher VEGF positive rate (73%; 8/11) than did cytokeratin-negative patients (48%; 14/29). Intratumoral vessel density in VEGF-positive patients was 26.9 +/- 10.3, which was significantly higher than that in VEGF-negative patients (13.2 +/- 8.7, P < 0.05). Thus, the presence of cytokeratin-positive cells in bone marrow was closely related to angiogenesis in the primary tumor.CONCLUSIONS: Cytokeratin staining can be useful for identifying patients at high risk for metastasis. Prophylactic lymph node dissection, adjuvant chemotherapy, and antiangiogenic treatment may be necessary for patients with micrometastasis.

Immunocytologic staining of cytokeratin in bone marrow aspirates to detect micrometastatic cells fails in patients with metastatic urologic carcinoma

The detection of cytokeratin-positive cells in bone marrow of patients with various carcinomas is considered an adverse marker. To establish a routine diagnostic procedure to detect these cells in patients with urologic carcinoma, we evaluated different immunocytological techniques in parallel. The study included 50 patients with urologic carcinoma (16 with metastatic disease) and 10 healthy volunteers. Bone marrow was aspirated from the iliac crest; after density centrifugation, cytocentrifuged specimens were stained according to optimized modifications of conventional immunocytologic techniques. Overall only 3% of patients with locally restricted carcinoma and only 19% with metastatic disease showed cytokeratin-positive bone marrow cells. Nonspecific labeling of bone marrow cells in the immunoglobulin isotype controls was a major problem, demonstrating the need for numerous detailed controls. If all nonspecific reactions were excluded, up to 20% of the healthy controls yielded positive results as well. The presence of cytokeratin-positive cells in bone marrow aspirates shows only slight correlation with systemic metastasis. The biologic and clinical relevance of this immunocytologic marker must be judged cautiously.