Clinical Cancer Research CR Helping Patients Advances in Breast Cancer
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Cancer Research Clinical Cancer Research
Cancer Epidemiology Biomarkers & Prevention Molecular Cancer Therapeutics
Molecular Cancer Research Cancer Prevention Research
Cancer Prevention Journals Portal Cancer Reviews Online
Annual Meeting Education Book Meeting Abstracts Online
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Passebosc-Faure, K.
Right arrow Articles by Genin, C.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Passebosc-Faure, K.
Right arrow Articles by Genin, C.
Clinical Cancer Research Vol. 11, 6862-6867, October 1, 2005
© 2005 American Association for Cancer Research

Imaging, Diagnosis, Prognosis

Evaluation of a Panel of Molecular Markers for the Diagnosis of Malignant Serous Effusions

Karine Passebosc-Faure1, Guorong Li1, Claude Lambert1, Michèle Cottier3, Anne Gentil-Perret4, Pierre Fournel2, Maurice Pérol5 and Christian Genin1

Authors' Affiliations: Departments of 1 Immunology, 2 Pneumology, 3 Cytology, 4 Pathology, University Hospital Center, Saint Etienne, France and 5 Department of Pneumology, University Hospital Center, Lyon, France

Requests for reprints: Christian Genin, Department of Immunology, Bellevue University Hospital Center, University of Saint Etienne, Bd Pasteur, Pavillon 5 Bis, 42055 Saint Etienne, France. Phone: 33-4-77-82-83-79; Fax: 33-4-77-82-84-93; E-mail: Christian.Genin{at}univ-st-etienne.fr.


   Abstract
Top
 Abstract
Patients and Methods
 Results
 Discussion
 References
 
Purpose: Our main goal was to evaluate a panel of molecular markers for the detection of cancer cells in serous effusions and to determine their value as an adjunctive reverse transcription-PCR (RT-PCR) test to cytologic examination.

Experimental Design: One hundred fourteen serous effusions from 71 patients with tumors and 43 patients with benign diseases were subjected to RT-PCR for expression of carcinoembryonic antigen (CEA), epithelial cell adhesion molecule (Ep-CAM), E-cadherin, mammaglobin, mucin 1 (MUC1) isoforms MUC1/REP, MUC1/Y, and MUC1/Z, calretinin, and Wilms' tumor 1 susceptibility gene.

Results: CEA, Ep-CAM, E-cadherin, and mammaglobin were specifically expressed in malignant effusions. The sensitivity of RT-PCR in cytologically negative malignant effusions was 63.1% combining CEA and Ep-CAM (with 100% specificity) and reached 78.9% adding MUC1/Y or MUC1/Z (with 93% specificity). In the whole population of effusions, the combination of cytology with RT-PCR of CEA and Ep-CAM yielded a 90.1% sensitivity, a specificity and a positive predictive value of 100%, and a 86% negative predictive value for malignancy. Adding MUC1/Y or MUC1/Z to the panel, the sensitivity was 94.5% with 93% specificity, 95.7% PPV, and 90.9% negative predictive value. Moreover, CEA and mammaglobin were specifically expressed in epithelial malignancies, and mammaglobin was mainly expressed in effusions from breast carcinoma (97.3% of specificity).

Conclusions: A combination of cytology and RT-PCR analysis of CEA and Ep-CAM significantly improved the detection sensitivity of tumor cells in serous effusions. RT-PCR analysis of CEA, Ep-CAM, and mammaglobin in serous effusions could be a beneficial adjunct to cytology for the diagnosis of malignancy.

Serous effusions are commonly found in patients with cancer and they sometimes reveal the malignancy (1). As the diagnosis of the effusion acts directly on the prognosis and the treatment of the disease (the presence of tumor cells in effusions point out a poor prognosis and often implies an aggressive treatment, such as chemotherapy), it is clearly important to detect any cancer cells in the fluid with the greatest specificity and sensitivity. However, the search for cancer cells in serous effusions using classic cytologic technique is often difficult either because of their scarceness or because of their delicate distinction with reactive mesothelial or inflammatory cells (2). Then, when cancer cells are found in the fluid, the second question is to know if they are local or metastatic cancer cells, and if they are metastatic, from which organ? Cytology, which is the gold standard method for the diagnosis of serous effusion, has some difficulties to distinguish mesothelioma cells from carcinoma cells (2). Thus, many additional methods have been evaluated (3, 4) to improve the diagnostic accuracy to avoid invasive diagnostic techniques, such as thoracoscopy.

Recently, PCR techniques have been largely evaluated for the detection of cancer cells in blood, bone, and lymph nodes and they have proven to be more sensitive than conventional techniques (5, 6). However, PCR was less intensively used in serous effusions, although it provides an excellent material for molecular analysis. The few studies that have used PCR techniques for the detection of cancer cells in serous effusions have concluded it to be a beneficial adjunct to conventional techniques because they generally increase the detection sensitivity of tumor cells (7–11). However, the validated markers are very limited, which hinders clinical application.

The aims of our study were (a) to evaluate a panel of molecular markers for the detection of cancer cells in serous effusions, (b) to determine the value of the markers as an adjunctive reverse transcription-PCR (RT-PCR) test to cytologic examination, (c) and to assess their usefulness as differential diagnostic markers between epithelial and nonepithelial malignant effusions.

The markers have been chosen based on their published interest for tumor cell detection in serous effusions: mucin 1 (MUC1; designed as MUC1/REP; ref. 9), E-cadherin (CDH1; ref. 12), epithelial cell adhesion molecule (Ep-CAM or TACSTD1; refs. 10, 13), and carcinoembryonic antigen (CEA or CEACAM5; ref. 14). We also studied the expression of the two isoforms of MUC1, MUC1/Y and MUC1/Z, because they have been found to be preferentially expressed in epithelial tumors compared with MUC1/REP (15). Wilms' tumor 1 susceptibility gene (WT1; ref. 16) and calretinin (CALB2; refs. 12, 17), two markers of mesothelial cells, were tested to distinguish mesothelioma from carcinoma effusions. Mammaglobin (MGB1 or SCGB2A2) was used to specifically detect breast cancer cells because it was proven to be a mammary-specific marker (18). To our knowledge, CEA, CDH1, MUC1/Y, MUC1/Z, WT1, and CALB2 were never used to detect tumor cells in serous effusions by RT-PCR.


   Patients and Methods
Top
 Abstract
 Patients and Methods
Results
 Discussion
 References
 
Patients. Serous effusions from 114 patients (104 pleural and 10 ascitic fluids) were prospectively collected from the Departments of Pneumology, Cardiology, and Cancerology of the University Hospital Center of Saint Etienne and Department of Pneumology of the University Hospital Center of Lyon, France. All effusions had a reliable diagnosis documented by cytologic analysis, histologic analysis of pleural biopsy when it was done, and the review of clinical history and follow-up of the patients without histologic analysis or with conflicting results between cytology and histology. Effusions from 43 patients with nonmalignant disease caused by infection or cardiac failure served as controls (4 patients had an history of cancer but with a proven benign effusion). Among the 71 malignant effusions, 19 were cytologically negative. The cancer primary sites of the 71 patients with malignant serous effusions are detailed in Table 1. This study was approved by the institutional ethics committee.


View this table:
[in this window]
[in a new window]

  		Table 1. Expression of the different markers in serous effusions by RT-PCR

 
Preparation of cells from serous effusions. Each fluid sample was centrifuged in 50 mL tubes at 300 x g for 10 minutes at room temperature. Then, pellets were pooled together, washed once, and resuspended in RPMI 1640 with 10% FCS. Fractions of 900 µL were distributed in 1.5 mL cryotubes containing 100 µL DMSO and stored in liquid nitrogen until use. When needed, aliquots were thawed in a 37°C water bath and then washed once with Ca2+/Mg2+–free NaCl solution (0.01 mol/L), and a cell count was done.

RNA extraction. Total RNA was purified using the RNABle extraction solution (Eurobio, Les Ulis, France) following the manufacturer's instructions using 0.2 mL RNABle per 106 cells. Briefly, cell pellets were homogenized with the RNABle solution (containing guanidium isothiocyanate and phenol); then, total RNA was extracted by adding 0.1 volume of chloroform and precipitated using 1 volume of isopropanol. Then, it was washed twice in 75% ethanol and resuspended in RNase-free water. Total RNA was quantified spectrophotometrically at A260 nm.

Reverse transcription-PCR. Reverse transcription was done with the SuperScript First-Strand Synthesis System for RT-PCR (Invitrogen, Paisley, United Kingdom) using 0.5 µg total RNA and following the manufacturer's instructions.

PCR was done in 50 µL reaction mixture containing 1 µL cDNA, 5 µL of 10x PCR buffer (Invitrogen), 1 µL of 10 mmol/L deoxynucleotide triphosphate mix (Invitrogen), 1.5 µL of 50 mmol/L MgCl2, 100 ng of both sense and antisense primers (Table 2), and 1.25 units Taq polymerase (Invitrogen). Primers for ß-actin were also used to check RNA integrity and the efficiency of the reverse transcription step. The sequences of the primers were designed to span splice junctions or in two different exons so that genomic DNA contamination could be monitored. Positive (MCF7 cell line) and negative controls (PCR mix without cDNA) were included in each round of PCR. The primer sequences, their gene localization, and their annealing temperatures are listed in Table 2. After a denaturing step at 94°C for 3 minutes, PCR was done at 94°C for 1 minute, at the annealing temperature for 1 minute, and at 72°C for 1 minute for 30 cycles. PCR products were separated by electrophoresis on a 1.3% agarose gel. DNA fragments were visualized and photographed under UV light with ethidium bromide staining.


View this table:
[in this window]
[in a new window]

  		Table 2. Sequences and features of the primers used for RT-PCR

 
Statistical analysis. Marker expression between the different populations was compared using the {chi}2 test or the Fisher exact test when theoretical effective was insufficient. The level of statistical significance was set at P < 0.05. Diagnostic performance of each marker, alone or together, was assessed by computing sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV).


   Results
Top
 Abstract
 Patients and Methods
 Results
Discussion
 References
 
Evaluation of marker expressions in the whole population of serous effusions. WT1 and MUC1/REP were highly expressed in benign effusions (39.5% and 62.8%, respectively). Even if MUC1/REP was significantly more expressed in malignant than in benign serous effusions (P = 0.001), this two markers presented a too low specificity to supply a reliable analysis of the samples. Moreover, we did not see any correlation between tumor grades and marker expression in breast and pulmonary effusions (data not shown).

Results of marker expressions in the whole population of effusions are detailed in Table 1. MUC1/Y and MUC1/Z were always coexpressed and thus are considered as a unique marker, MUC1/YZ. CALB2 and MUC1/YZ were positive in 2.3% and 7% of benign effusions, respectively. Nevertheless, MUC1/YZ was significantly more expressed in malignant than in benign effusions (P < 0.0001). CEA, Ep-CAM, CDH1, and MGB1 were specifically expressed in malignant effusions.

The sensitivity, specificity, PPV, and NPV of cytology and of the molecular markers, alone or in combination, are presented in Table 3. Only the two marker combinations supplying the higher diagnostic information, in terms of sensitivity and specificity, were included in Table 3. Taken together, CEA, Ep-CAM, CDH1, and MGB1 yielded a positive result in 76% of the malignant effusions with 100% specificity. When they were combined to MUC1/YZ, we could reach a sensitivity of 81.7% for malignancy but with a slightly lower specificity (93%).


View this table:
[in this window]
[in a new window]

  		Table 3. Sensitivity, specificity, PPV, and NPV of cytology and of the molecular markers in the whole population (composed of 71 malignant and 43 benign effusions) of effusions

 
Finally, as expected, MGB1 was significantly more expressed in effusions from breast cancer patients than from other carcinoma (P < 0.0001): 71.4% of breast effusions were positive for MGB1, whereas only one effusion from an ovarian carcinoma was positive (Table 1). Thus, MGB1 presents a very good specificity (97.3%) for breast carcinoma effusions.

Marker expressions in the cytologically negative malignant effusions. Nineteen serous effusions, negative for tumor cell by cytologic examination, were diagnosed as malignant by histologic analysis of pleural biopsy and patient clinical data. Results of RT-PCR analysis are detailed in Table 4. Briefly, four samples were negative for all the markers (patients 6, 12, 15, and 16) and three were positive only with MUC1/YZ (patients 9, 17, and 18). Considering the malignant-specific markers, 12 samples were positive with at least one marker, and two molecular markers (CEA and Ep-CAM) were necessary and sufficient for the detection of tumor cells in these effusions. The sensitivity of RT-PCR in cytologically negative malignant effusions was 63.1% (12 of 19) combining CEA and Ep-CAM, with a specificity (43 of 43) and a PPV (12 of 12) of 100% and a NPV of 86% (43 of 50). Adding MUC1/YZ to this combination, the sensitivity for malignancy was 78.9% (15 of 19) with 93% specificity (40 of 43), 83.3% PPV (15 of 18), and 90.9% NPV (40 of 44).


View this table:
[in this window]
[in a new window]

  		Table 4. Marker expressions in cytologically negative malignant effusions

 
Moreover, among the three breast carcinoma effusions, two were positive for MGB1, which is of great interest to detect the tumor primary site without invasive diagnostic investigations.

Combination of cytology and reverse transcription-PCR. The combination of cytology and RT-PCR of CEA + Ep-CAM in the whole population of effusions allowed to confirm the tumoral diagnosis in 90.1% of malignant effusions (52 malignant effusions were positive with cytology and 12 more were positive with RT-PCR) with 100% specificity, 100% PPV, and 86% NPV (Table 3), whereas the combination of cytology and RT-PCR of CEA + Ep-CAM + MUC1/YZ yielded a sensitivity of 94.4% with a specificity of 93% for tumor cells detection in malignant effusions. The PPV and NPV were 95.7% and 90.9%, respectively (Table 3). The sensitivities of this two last combined test are significantly more elevated than that of cytology alone (P < 0.0001 each, Mac Nemar {chi}2 test).

Markers that could help to distinguish epithelial from nonepithelial malignant effusions. Then, we wanted to know which molecular markers would undoubtedly help to distinguish between epithelial and nonepithelial cancer cell in effusions. Only two markers, CEA and MGB1, were exclusively expressed in carcinoma effusions (Table 1). Surprisingly, Ep-CAM was positive in 45% of nonepithelial malignant effusions but was significantly more expressed in carcinoma effusions than in nonepithelial malignant ones (P = 0.018).

Considering the malignant effusions that were positive for tumor cells with the combination of cytology and CEA + Ep-CAM RT-PCR (64 effusion specimens), 48 had an epithelial origin and 16 a nonepithelial origin. In this population, the sensitivity, specificity, PPV, and NPV of CEA for carcinoma cells are 58.3% (28 of 48), 100% (16 of 16), 100% (28 of 28), and 44.4% (16 of 36), respectively. Combined with MGB1, the sensitivity reached 68.7% (33 of 48) and the NPV was 51.6% (16 of 31). Thus, CEA and MGB1 are very interesting molecular markers to determine the epithelial origin of tumor cells in serous effusions.

Moreover, CALB2 was significantly more expressed in mesothelioma than in carcinoma effusions (P = 0.018). Among the four carcinoma effusions that were positive for CALB2, three were also positive for CEA, thus excluding the possibility of a mesothelioma. Among the five CALB2-positive mesothelioma, three were suspected to be carcinoma by cytology and were diagnosed as mesothelioma by histology.


   Discussion
Top
 Abstract
 Patients and Methods
 Results
 Discussion
References
 
We aimed to evaluate an ancillary method that could improve the yield of cytologic analysis for cancer diagnosis in serous effusions. Thus, we focused our study on a rapid, easy, inexpensive, and sensitive technique that could be done in most analysis laboratories. Basic RT-PCR strategy seemed to be the best compromise between immunocytologic techniques that often lacks sensitivity and flexibility in the markers choice and more elaborated RT-PCR techniques, such as real-time quantitative RT-PCR that are more expensive and difficult to set.

As a first step, we evaluated the expression of the markers in the whole population of effusions to determine their interest for the detection of malignancy. Even if these markers have been studied at the mRNA level, one could expect them to be functional at the protein level according to the numerous publications that have studied them by immunocytochemistry or flow cytometry in serous effusions (12–14, 16, 19). We found two markers highly expressed (WT1 and MUC1/REP) and two other markers rarely expressed (CALB2 and MUC1/YZ) in benign effusions. WT1 and CALB2 are markers of mesothelial lineage, and as normal desquamated mesothelial cells can be present in any kind of effusion, they are of limited value, as molecular markers, for the distinction of mesothelioma effusions from other malignant or benign effusions. However, CALB2 was significantly more expressed in mesothelioma than in carcinoma effusions. Thus, CALB2 might help to orient a diagnosis in favor of a mesothelioma only when the tumoral nature of the effusion is ascertained and if it is used in combination with confirmed epithelial markers to exclude a maximum of false-positive results. In some immunocytochemical studies, CALB2 was positive in most of benign effusions (94-100%; refs. 12, 20). It is difficult to compare these results with ours, because the expression of CALB2 was detected at two different levels (protein or mRNA). The mRNA of CALB2 might be an unstable mRNA, but it is also a possibility that the polyclonal antibodies used in those studies produced nonspecific reactions. Moreover, in another immunocytochemical study using a different polyclonal anti-CALB2, only 27% of the benign effusions were stained (21). MUC1/REP, which is the full-length isoform of MUC1 (22), has been described to be expressed in nearly all normal and malignant epithelial cells (23) but also in normal and malignant hematopoietic cells (24, 25). This is probably why we found it expressed in benign serous effusions. Although MUC1/REP was significantly more expressed in malignant effusions, which is in agreement with Yu et al. (9), we think that it was too frequently expressed in benign samples to be a reliable molecular marker for malignancy. Thus far, the expression of alternatively spliced forms of MUC1 has been poorly studied. MUC1/Y and MUC1/Z have been detected in primary breast tumors by Western blot analysis (15, 26) and in ovarian cancer tissue by RT-PCR (27). Moreover, MUC1/Y has been detected by flow cytometry on the surface of ovary and breast malignant cells in effusions (19). We showed here that MUC1/Y and MUC1/Z were mostly expressed in malignant effusions of epithelial and nonepithelial origin and poorly expressed in benign effusions. They presented a very good specificity for malignancy (93%), and they were expressed alone in some malignant effusions, contributing to enhance the sensitivity of the RT-PCR combined test. Thus, they could be interesting molecular markers to add in some panels of markers to enhance the detection sensitivity of tumor cells, keeping in mind that they are not totally discriminant.

The other markers (CEA, Ep-CAM, CDH1, and MGB1) were specifically expressed in malignant effusions, therefore being very good candidates for molecular detection of malignancy in serous effusions. CEA was often used in immunohistochemical studies to distinguish carcinoma cells from reactive mesothelial or mesothelioma cells (14, 28). However, it was never used as a RT-PCR marker in serous effusions. In this study, we showed that CEA was not only specifically expressed in malignant effusions but was also restricted to the epithelial malignancies. This makes it an exceptional molecular marker to distinguish epithelial from nonepithelial malignant serous effusions. Ep-CAM is generally admitted as an epithelial marker expressed on the surface of most carcinomas and it was often used by immunocytochemistry to distinguish carcinoma from mesothelioma or reactive mesothelial cells in serous effusions (13, 29, 30). Using real-time RT-PCR, Nagel et al. found it weakly expressed in cultured mesothelial cells isolated from ascites of patients with benign disease (10). In our study, it was not expressed in benign serous effusions. This could be because conventional RT-PCR is less sensitive than real-time RT-PCR but also because their mesothelial cells were cultivated with FCS, which contained numerous cytokines susceptible to stimulate Ep-CAM expression as tumor necrosis factor-{alpha} (31). Moreover, Ep-CAM was expressed in most carcinomas (74.5%) and mesothelioma effusions (57.1%). Indeed, some reports described infrequent and weak expression of Ep-CAM in mesothelioma by immunocytochemistry (13, 29, 32). Thus, although it was significantly more expressed in carcinoma than in nonepithelial malignant effusions, we rather consider it as a tumor than a carcinoma marker in serous effusions. Therefore, if Ep-CAM seems to be of limited interest to distinguish carcinoma from mesothelioma effusions, it is a valuable molecular marker for the detection of tumor cells in serous effusions. CDH1, an epithelial-specific adhesion molecule, has been proven to reliably distinguish carcinoma cells from benign reactive mesothelial cells, but not from mesothelioma cells, in serous effusions by immunocytology (12, 20, 33). We also found that CDH1 was specifically expressed in malignant epithelial and nonepithelial serous effusions, but it was not very sensitive. It was expressed alone in none of the cytologically negative malignant effusions and in only one cytologically positive malignant effusion. That is why its usefulness in a panel of molecular markers is relative. MGB1 was reported previously as a specific marker of breast carcinoma cells (18). However, using nested RT-PCR, Grünewald et al. (34) recently found MGB1 expression in endometrial and ovarian carcinoma effusions as well as in stomach and lung effusions in another study (11). In this last study, they also found MGB1 expression in 12.9% of their control effusions. We only found MGB1 expression in breast and in one effusion from an ovarian carcinoma, and all our control effusions were negative. We think that the highest sensitivity of nested RT-PCR could explain the differences between our respective studies. With 97.3% of specificity for breast carcinoma effusions, MGB1 is therefore a marker of choice in the search of primary site in carcinoma effusions of unknown origin.

Then, after the determination of the most efficient markers for the detection of tumor cells in serous effusions, we evaluated their interest as an adjunctive RT-PCR test to cytology. Therefore, we studied their expression in a subgroup of cytologically negative malignant effusions. We found that there were two pertinent marker combinations: CEA + Ep-CAM + MUC1/YZ combination yielding the highest sensitivity (78.9%) with a very good specificity (93%) and CEA + Ep-CAM combination yielding the highest specificity (100%) with a good sensitivity (63.1%). The two marker combinations are interesting. However, our standpoint is that specificity is more important than sensitivity for a diagnosis of malignancy that has to be sure. Moreover, such molecular biology techniques have to be reliable to avoid invasive diagnostic techniques. Thus, the combination of CEA and Ep-CAM as an ancillary RT-PCR test in cytologically negative and suspicious serous effusions seems to us the best choice. Moreover, we would also complete this test with MGB1 because it can predict a breast or, less frequently, an ovarian origin of the effusion.

Then, we showed that the combination of cytology and CEA + Ep-CAM RT-PCR test improved the diagnosis accuracy, yielding 90.1% sensitivity, 100% specificity, 100% PPV, and 86% NPV for malignancy. These results are much better than the 81.1% sensitivity and 70.1% specificity obtained by Fiegl et al. (11) combining cytology, fluorescence in situ hybridization for aneuploidy, and nested RT-PCR of MGB1 and MGB1-B. Moreover, it seems that basic RT-PCR strategy allowed to abrogate most of markers' ectopic expression staying enough sensitive to be clinically useful to complement cytologic examination. Adding RT-PCR to clinical practice for the diagnosis of serous effusions would represent a nonexcessive extra workload in terms of time, cost, and manpower compared with the benefits that it would bring to the patients.

Finally, we also showed that CEA and MGB1 were specifically expressed in epithelial malignancies and that, among the 64 effusions detected as malignant with the combined test of cytology and CEA + Ep-CAM RT-PCR, they allowed to determine the epithelial origin of 68.7% of carcinoma effusions with 100% specificity.

In conclusion, this study showed that CEA and MGB1 are very reliable molecular markers for the detection of carcinoma cells in serous effusions and that a RT-PCR test of CEA, Ep-CAM, and MGB1 could be a helpful adjunct to cytologic examination for the diagnosis of malignancy and the orientation of the tumor origin.


   Acknowledgments
 
We thank C. Ruesch, M.D., P. Sagnol, M.D., and J.M. Vergnon, M.D., Ph.D., for providing pleural effusions; S. Laporte and E. Preysle from DRCC of St. Etienne for statistical analysis, and D. Bernard-Gallon, Ph.D., for critical reading of the article.


   Footnotes
 
Grant support: Ligue Contre le Cancer, Comité de la Loire, and French Ministry of Health.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Received 1/ 7/05; revised 7/ 7/05; accepted 7/13/05.


   References
Top
 Abstract
 Patients and Methods
 Results
 Discussion
 References
 

  1. Monte SA, Ehya H, Lang WR. Positive effusion cytology as the initial presentation of malignancy. Acta Cytol 1987;31:448–52.[Medline]
  2. Bedrossian CWM. Diagnostic problems in serous effusions. Diagn Cytopathol 1998;19:131–7.[CrossRef][Medline]
  3. Fetsch PA, Abati A. Immunocytochemistry in effusion cytology: a contemporary review. Cancer 2001;93:293–308.[CrossRef][Medline]
  4. Mohanty SK, Dey P. Serous effusions: diagnosis of malignancy beyond cytomorphology. An analytic review. Postgrad Med J 2003;79:569–74.[Abstract/Free Full Text]
  5. Gerhard M, Juhl H, Kalthoff H, Schreiber HW, Wagener C, Neumaier M. Specific detection of carcinoembryonic antigen-expressing tumor cells in bone marrow aspirates by polymerase chain reaction. J Clin Oncol 1994;12:725–9.[Abstract]
  6. Mori M, Mimori K, Ueo H, et al. Clinical significance of molecular detection of carcinoma cells in lymph nodes and peripheral blood by reverse transcription-polymerase chain reaction in patients with gastrointestinal or breast carcinomas. J Clin Oncol 1998;16:128–32.[Abstract/Free Full Text]
  7. Okamoto I, Morisaki T, Sasaki J, et al. Molecular detection of cancer cells by competitive reverse transcription-polymerase chain reaction analysis of specific CD44 variant RNAs. J Natl Cancer Inst 1998;90:307–15.[Abstract/Free Full Text]
  8. Yang CT, Lee MH, Lan RS, Chen JK. Telomerase activity in pleural effusions: diagnostic significance. J Clin Oncol 1998;16:567–73.[Abstract]
  9. Yu CJ, Shew JY, Liaw YS, et al. Application of mucin quantitative competitive reverse transcription polymerase chain reaction in assisting the diagnosis of malignant pleural effusion. Am J Respir Crit Care Med 2001;164:1312–8.[Abstract/Free Full Text]
  10. Nagel H, Werner C, Hemmerlein B. Real-time reverse transcription-polymerase chain reaction assay for GA733-2 m RNA in the detection of metastatic carcinoma cells in serous effusions. Am J Clin Pathol 2003;120:888–901.[CrossRef][Medline]
  11. Fiegl M, Haun M, Massoner A, et al. Combination of cytology, fluorescence in situ hybridization for aneuploidy, and reverse-transcriptase polymerase chain reaction for human mammaglobin/mammaglobin B expression improves diagnosis of malignant effusions. J Clin Oncol 2004;22:474–83.[Abstract/Free Full Text]
  12. Kitasume H, Kitamura K, Mukai K, et al. Cytologic differential diagnosis among reactive mesothelial cells, malignant mesothelioma, and adenocarcinoma. Cancer 2000;90:55–60.[CrossRef][Medline]
  13. Kuhlmann L, Berghauser KH, Schaffer R. Distinction of mesothelioma from carcinoma in pleural effusions. An immunocytochemical study on routinely processed cytoblock preparations. Pathol Res Pract 1991;187:467–71.[Medline]
  14. Ghosh AK, Gatter KC, Dunnill MS, Mason DY. Immunohistological staining of reactive mesothelium, mesothelioma, and lung carcinoma with a panel of monoclonal antibodies. J Clin Pathol 1987;40:19–25.[Abstract/Free Full Text]
  15. Baruch A, Hartmann M, Zrihan-Licht S, et al. Preferential expression of novel MUC1 tumor antigen isoforms in human epithelial tumors and their tumor-potentiating function. Int J Cancer 1997;71:741–9.[CrossRef][Medline]
  16. Hecht JL, Lee BH, Pinkus JL, Pinkus GS. The value of Wilms tumor susceptibility gene 1 in cytologic preparations as a marker for malignant mesothelioma. Cancer 2002;96:105–9.[CrossRef][Medline]
  17. Doglioni C, Tos AP, Laurino L, et al. Calretinin: a novel immunocytochemical marker for mesothelioma. Am J Surg Pathol 1996;20:1037–46.[CrossRef][Medline]
  18. Fleming TP, Watson MA. Mammaglobin, a breast-specific gene, and its utility as a marker for breast cancer. Ann N Y Acad Sci 2000;923:78–89.[Medline]
  19. Hartman M, Baruch A, Ron I, et al. MUC1 isoform specific monoclonal antibody 6E6/2 detects preferential expression of the novel MUC1/Y protein in breast and ovarian cancer. Int J Cancer 1999;82:256–67.[CrossRef][Medline]
  20. He DN, Zhu HS, Zhang KH, et al. E-cadherin and calretinin as immunocytochemical markers to differentiate malignant from benign serous effusions. World J Gastroenterol 2004;10:2406–8.[Medline]
  21. Simsir A, Fetsch P, Mehta D, Zakowski M, Abati A. E-cadherin, N-cadherin, and calretinin in pleural effusions: the good, the bad, the worthless. Diagn Cytopathol 1999;20:125–30.[CrossRef][Medline]
  22. Gendler SJ. MUC1, the renaissance molecule. J Mammary Gland Biol Neoplasia 2001;6:339–53.[CrossRef][Medline]
  23. Taylor-Papadimitriou J, Burchell J, Miles DW, Dalziel M. MUC1 and cancer. Biochim Biophys Acta 1999;1455:301–13.[Medline]
  24. Brugger W, Buhring HJ, Grunebach F, et al. Expression of MUC-1 epitopes on normal bone marrow: implications for the detection of micrometastatic tumor cells. J Clin Oncol 1999;17:1535–44.[Abstract/Free Full Text]
  25. Treon SP, Mollick JA, Urashima M, et al. Muc-1 core protein is expressed on multiple myeloma cells and is induced by dexamethasone. Blood 1999;93:1287–98.[Abstract/Free Full Text]
  26. Zrihan-Licht S, Vos HL, Baruch A, et al. Characterization and molecular cloning of a novel MUC1 protein, devoid of tandem repeats, expressed in human breast cancer tissue. Eur J Biochem 1994;224:787–95.[Medline]
  27. Obermair A, Schmid BC, Stimpfl M, et al. Expression of MUC1 splice variants in benign and malignant ovarian tumours. Int J Cancer 2002;100:166–71.[CrossRef][Medline]
  28. Lee JS, Nam JH, Lee MC, Park CS, Juhng SW. Immunohistochemical panel for distinguishing between carcinoma and reactive mesothelial cells in serous effusions. Acta Cytol 1996;40:631–6.[Medline]
  29. Sheibani K, Shin SS, Kezirian J, Weiss LM. Ber-EP4 antibody as a discriminant in the differential diagnosis of malignant mesothelioma versus adenocarcinoma. Am J Surg Pathol 1991;15:779–84.[Medline]
  30. Latza U, Niedobitek G, Schwarting R, Nekarda H, Stein H. Ber-EP4: new monoclonal antibody which distinguishes epithelia from mesothelia. J Clin Pathol 1990;43:213–9.[Abstract/Free Full Text]
  31. Grunberg E, Eckert K, Karsten U, Maurer HR. Effects of differentiation inducers on cell phenotypes of cultured nontransformed and immortalized mammary epithelial cells: a comparative immunocytochemical analysis. Tumour Biol 2000;21:211–23.[CrossRef][Medline]
  32. Ordonez NG. Value of the Ber-EP4 antibody in differentiating epithelial pleural mesothelioma from adenocarcinoma. The M.D. Anderson experience and a critical review of the literature. Am J Clin Pathol 1998;109:85–9.[Medline]
  33. Schofield K, D'Aquila T, Rimm DL. The cell adhesion molecule, E-cadherin, distinguishes mesothelial cells from carcinoma cells in fluids. Cancer 1997;81:293–8.[CrossRef][Medline]
  34. Grünewald K, Haun M, Fiegl M, et al. Mammaglobin expression in gynecologic malignancies and malignant effusions detected by nested reverse transcriptase-polymerase chain reaction. Lab Invest 2002;82:1147–53.[Medline]



This article has been cited by other articles:


Home page
 Cancer Res.Home page
H. C. Workman, C. Sweeney, and K. L. Carraway III
The Membrane Mucin Muc4 Inhibits Apoptosis Induced by Multiple Insults via ErbB2-Dependent and ErbB2-Independent Mechanisms
Cancer Res., April 1, 2009; 69(7): 2845 - 2852.
[Abstract] [Full Text] [PDF]

Home page
 Eur. J. Cardiothorac. Surg.Home page
S. Elia, R. Massoud, G. Guggino, B. Cristino, C. Cortese, A. R. De Massimi, and R. Zenobi
Tumor type M2-pyruvate-kinase levels in pleural fluid versus plasma in cancer patients: a further tool to define the need for invasive procedures
Eur. J. Cardiothorac. Surg., April 1, 2008; 33(4): 723 - 727.
[Abstract] [Full Text] [PDF]

Home page
 J. Clin. Pathol.Home page
F C Schmitt, A Longatto-Filho, A Valent, and P Vielh
Molecular techniques in cytopathology practice
J. Clin. Pathol., March 1, 2008; 61(3): 258 - 267.
[Abstract] [Full Text] [PDF]

Home page
 J. Mol. Diagn.Home page
F. Hube, Y. Myal, E. Leygue, J. Rollin, Y. Gruel, Y. Yatabe, and on behalf of the authors
Expression of Two Breast-Specific Molecules in the Lung
J. Mol. Diagn., July 1, 2006; 8(3): 390 - 393.
[Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Passebosc-Faure, K.
Right arrow Articles by Genin, C.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Passebosc-Faure, K.
Right arrow Articles by Genin, C.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Cancer Research Clinical Cancer Research
Cancer Epidemiology Biomarkers & Prevention Molecular Cancer Therapeutics
Molecular Cancer Research Cancer Prevention Research
Cancer Prevention Journals Portal Cancer Reviews Online
Annual Meeting Education Book Meeting Abstracts Online