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234 MUC1 in malignant mesothelioma
Sunday, October 22, 2006 / Workshop: Pathology: Current status of pathologic diagnosis of malignant mesothelioma phering of the step by step process leading from increased mesothelial cell proliferation to mesothelioma, from which we are expecting the development of new effective therapeutics.
234
MUC1 in malignant mesothelioma
J. Creaney 1 , A. Segal 2 , G. Sterrett 2 , E. Baker 3 , A.K. Nowak 1 , B.W.S. Robinson 1 , M. Millward 1 . 1 University of Western Australia, Nedlands, Australia; 2 PathWest Anatomincal Pathology, Australia 3 PathWest Cytogenics, Australia Background: Malignant mesothelioma (MM) is an aggressive tumour commonly caused by asbestos exposure. Distinction between benign and malignant mesothelial cells in effusions has been facilitated by the use of epithelial membrane antigen (EMA), a glycoprotein product of the MUC1 gene. EMA is equivalent to polymorphic epithelial mucin (PEM), sialomucin, episialin or CA15-3. MUC1 gene products have been extensively investigated in various common carcinomas and there are several ongoing clinical trials involving MUC1 targeted immunotherapy. The current study examined various characteristics of MUC1 in MM samples. Methods: Tumor, effusion and serum samples from patients with MM and from controls were identified and sourced from PathWest and the Australian Mesothelioma Tissue Bank. A tissue microarray was constructed from 20 MM and 16 benign-reactive effusion samples. Sections were stained with a panel of seven commercial antibodies using standard immunoperoxidase techniques. Levels of CA 15-3 were measured in sera from 34 MM patients and 22 controls using the IMMULITE 2000 BR-MA assay (Diagnostics Products Corporation). MUC1 mRNA expression was examined by conventional and quantitative PCR using a panel of primers. Fluorescence in situ hybridization (FISH) was performed on tissue microarray sections using three SpectrumGreen-labeled BAC clones which encompass the MUC1 gene, plus a SpectrumOrange-labeled control probe (Vysis). Results: Of the antibodies tested, the Dako anti-MUC1 antibody E29 was the most useful as a diagnostic discriminator between benign and malignant mesothelial cells, with a sensitivity of 84% and no false positive results. Serum CA15-3 was elevated in 70% of MM patients and 5% of healthy controls. Levels MUC1 mRNA were elevated in MM tumours relative to normal mesothelial cells. Different splicing profiles of MUC1 mRNA were observed between MM samples, from tumours and effusions, and non-malignant mesothelial cell samples. FISH studies suggested that gene amplification was not the major mechanism for MUC1 overexpression. Conclusion: MUC1 is an attractive target in MM. The use of MUC1 directed immunotherapy in MM should be investigated.
236
S57
Prolonged survival in malignant mesothelioma: A study of nineteen cases
F. Galateau-Sallé 1 , P. Vergani 2 , P. Astoul 3 , P. Rolland 4 , J.C. Pairon 5 , P. Brochard 6 , M. Matrat 5 , V. Abonnet 7 , E. Imbernon 4 , A. Gilg Soit Ilg 4 , M. Goldberg 4 , G. Launoy 8 . 1 French National Mesothelioma Registry CHU Caen, France; 2 ERI 3 Inserm CHU Caen, France; 3 Faculté de Médecine de la Timone CHU Marseille, France; 4 Institut de Veilel sanitaire, France; 5 Institut de Médecine du Travail de Paris Ile de France, France; 6 Laboratoire Santé Environnement Bordeaux, France; 7 CRB Mesotheliome ERI3 Inserm, France; 8 ERI Inserm CHU Caen, France Background: Prolonged survival in diffuse MM is rare. We have studied 19 cases with survival in excess of 3 years, in order to evaluate clinico-pathological features which enhance survival. Methods: Nineteen MM cases with a survival in excess of 3 years were evaluated for clinical and histopathological features. MM were classified according to the WHO classification [2004] and all cases presented calretinin (Zymed) and CK5/6 (DAKO) clear cut immunopositivity. The following immunohistochemical and molecular markers were studied: EMA, p53, desmin, mesothelin, p16. SV40 DNA sequences were studied by PCR analysis from paraffin embedded specimens and were sequenced. p16INK4and RASSF-1A genes was assessed by the method of methylation specific PCR [MSP]. Occupational histories were evaluated by a group of asbestos epidemiological experts. Results: The 19 long survivals range from 36 months, to 135 months with a mean survival time of 34 months The patients were 12 men and 7 women with an average age at diagnosis of 63 years. Asbestos exposure was identified in 14/19 cases. Clinical and anatomical location and immunohistochemical profiles (EMA, desmin, mesothelin) were similar to mesothelioma with poor prognosis. All were characterized by small nodules (with a size < 2 cm). Histological types comprised epithelioid 18/19 and one biphasic MM with osteocartilaginous differentiation. Except 2, all showed a prominent chronic inflammatory infiltrate. Molecular positive results were as follows; SV40(4/15), p16 INK4 methylation (2/18), and RASSF-1A(4/18). There was no correlation between asbestos exposure, SV40, p53 expression, p16 INK4 and RASSF-1A methylation. Conclusions: Favorable prognostic factors were small size (<2cm), epithelioid histological type and prominent chronic inflammatiory infiltrate. Immunohistochemical markers did not differ significantly from mesothelioma with poor prognosis. p16/CDKN2A and RASSF1A promoter methylation are significantly correlated in the process of tumorigenesis of mesothelioma (p<0.0006).
237
Accuracy of pathological diagnosis of mesothelioma (MM) in Japan
K. Inai, Y. Takeshima, K. Kushitani. Hiroshima University, Hiroshima, Japan
235
Diagnostic value of proliferative markers Ki-67 and Repp 86 in distinguishing between malignant and benign mesothelial proliferations
Z. Mohammadtaheri, M. Mehrafza, F. Mohammadi, M. Khodami, M. Masjedi, M. Bahadori. National Research Institude of Tuberculosis and Lung Disease, Teheran, Iran Background: The differentiation of benign pleural reactions from malignant mesothelioma may be difficult, especially in small biopsy specimens. Purpose: to evaluate 2 proliferative markers (Ki-67 and Repp 86) to distinguish between malignant mesothelioma (MM) and reactive mesothelial hyperplasia (MH). Methods: Thirty-six cases of malignant mesothelioma, including 26 men and 10 women, median age 62.9 (range 36-80 years) and 22 reactive mesothelial hyperplasia cases were investigated for proliferative status by immunohistochemistry method with two monoclonal antibodies; mAb Ki-S2 (repp 86) and Ki-S5 (Ki-67) and the labeling indices (LI) were calculated. Results: Both markers (Ki- 67 and Repp 86) show significant correlation with each other in differentiation of MM and MH (r= 77.5, p value < 0.0001). Average count reveals a significant increase in MM compared with reactive MH (p value < 0.0001). Assuming a threshold of 9% with both Ki-67 and Repp 86 a sensitivity of 88% and 92% and specificity of 94% respectively resulted. Conclusions: Both proliferative markers can be useful in distinction between malignant mesothelioma and reactive mesothelial hyperplasia (p value < 0.0001).
Background: The clinical diagnosis of MM is dependent on imaging including x-ray, CT or MRI, because the serum markers are not reliable at this time and so, definitive diagnosis by pathological examination has been required. However, there are many histological varieties in MM and the diagnostic criteria are not so clear and definite pathological diagnosis is not so easy. Therefore, we tried to make an antibody panel for immunohistochemistry in the differential diagnosis of MM and examined accuracy of pathological diagnosis of MM in Japan. Methods: Eighty-eight cases of epithelioid MM were compared to 51 cases of pulmonary adenocarcinoma, and 41 cases of sarcomatoid MM were compared to 47 cases of various types of sarcoma. In these cases, the representative tissue blocks were used for immunohistochemical stainings by S.A.B. method. In addition, 111 permitted cases among 878 cases with death certificate (2003) in Japan were examined in our laboratory to ascertain accuracy of the diagnosis of MM. Results: The antibody panel of calretinin, WT1 and CEA is useful for the differential diagnosis between epithelioid MM and pulmonary adenocarcinoma. On the other hand, the panel of CAM5.2, AE1/AE3 and other specified antibody positive for sarcomas is useful for the differential diagnosis between sarcomatoid MM and sarcoma. Using our criteria including immmunohistochemistry mentioned above, it was considered that 14 cases (13%) among 111 examined cases was diagnosed as unlikely or definitely not MM, and it is supposed that the diagnosis of MM in Japan is not correct in the proportion of 10-15%. Conclusions: For the accurate diagnosis of MM, immunohistochemistry using the special antibody panel according to histological type is indispensable. Knowledge of immunohistochemistry is essential for improvement of the accuracy of pathological diagnosis of MM.
234
MUC1 in malignant mesothelioma
J. Creaney 1 , A. Segal 2 , G. Sterrett 2 , E. Baker 3 , A.K. Nowak 1 , B.W.S. Robinson 1 , M. Millward 1 . 1 University of Western Australia, Nedlands, Australia; 2 PathWest Anatomincal Pathology, Australia 3 PathWest Cytogenics, Australia Background: Malignant mesothelioma (MM) is an aggressive tumour commonly caused by asbestos exposure. Distinction between benign and malignant mesothelial cells in effusions has been facilitated by the use of epithelial membrane antigen (EMA), a glycoprotein product of the MUC1 gene. EMA is equivalent to polymorphic epithelial mucin (PEM), sialomucin, episialin or CA15-3. MUC1 gene products have been extensively investigated in various common carcinomas and there are several ongoing clinical trials involving MUC1 targeted immunotherapy. The current study examined various characteristics of MUC1 in MM samples. Methods: Tumor, effusion and serum samples from patients with MM and from controls were identified and sourced from PathWest and the Australian Mesothelioma Tissue Bank. A tissue microarray was constructed from 20 MM and 16 benign-reactive effusion samples. Sections were stained with a panel of seven commercial antibodies using standard immunoperoxidase techniques. Levels of CA 15-3 were measured in sera from 34 MM patients and 22 controls using the IMMULITE 2000 BR-MA assay (Diagnostics Products Corporation). MUC1 mRNA expression was examined by conventional and quantitative PCR using a panel of primers. Fluorescence in situ hybridization (FISH) was performed on tissue microarray sections using three SpectrumGreen-labeled BAC clones which encompass the MUC1 gene, plus a SpectrumOrange-labeled control probe (Vysis). Results: Of the antibodies tested, the Dako anti-MUC1 antibody E29 was the most useful as a diagnostic discriminator between benign and malignant mesothelial cells, with a sensitivity of 84% and no false positive results. Serum CA15-3 was elevated in 70% of MM patients and 5% of healthy controls. Levels MUC1 mRNA were elevated in MM tumours relative to normal mesothelial cells. Different splicing profiles of MUC1 mRNA were observed between MM samples, from tumours and effusions, and non-malignant mesothelial cell samples. FISH studies suggested that gene amplification was not the major mechanism for MUC1 overexpression. Conclusion: MUC1 is an attractive target in MM. The use of MUC1 directed immunotherapy in MM should be investigated.
236
S57
Prolonged survival in malignant mesothelioma: A study of nineteen cases
F. Galateau-Sallé 1 , P. Vergani 2 , P. Astoul 3 , P. Rolland 4 , J.C. Pairon 5 , P. Brochard 6 , M. Matrat 5 , V. Abonnet 7 , E. Imbernon 4 , A. Gilg Soit Ilg 4 , M. Goldberg 4 , G. Launoy 8 . 1 French National Mesothelioma Registry CHU Caen, France; 2 ERI 3 Inserm CHU Caen, France; 3 Faculté de Médecine de la Timone CHU Marseille, France; 4 Institut de Veilel sanitaire, France; 5 Institut de Médecine du Travail de Paris Ile de France, France; 6 Laboratoire Santé Environnement Bordeaux, France; 7 CRB Mesotheliome ERI3 Inserm, France; 8 ERI Inserm CHU Caen, France Background: Prolonged survival in diffuse MM is rare. We have studied 19 cases with survival in excess of 3 years, in order to evaluate clinico-pathological features which enhance survival. Methods: Nineteen MM cases with a survival in excess of 3 years were evaluated for clinical and histopathological features. MM were classified according to the WHO classification [2004] and all cases presented calretinin (Zymed) and CK5/6 (DAKO) clear cut immunopositivity. The following immunohistochemical and molecular markers were studied: EMA, p53, desmin, mesothelin, p16. SV40 DNA sequences were studied by PCR analysis from paraffin embedded specimens and were sequenced. p16INK4and RASSF-1A genes was assessed by the method of methylation specific PCR [MSP]. Occupational histories were evaluated by a group of asbestos epidemiological experts. Results: The 19 long survivals range from 36 months, to 135 months with a mean survival time of 34 months The patients were 12 men and 7 women with an average age at diagnosis of 63 years. Asbestos exposure was identified in 14/19 cases. Clinical and anatomical location and immunohistochemical profiles (EMA, desmin, mesothelin) were similar to mesothelioma with poor prognosis. All were characterized by small nodules (with a size < 2 cm). Histological types comprised epithelioid 18/19 and one biphasic MM with osteocartilaginous differentiation. Except 2, all showed a prominent chronic inflammatory infiltrate. Molecular positive results were as follows; SV40(4/15), p16 INK4 methylation (2/18), and RASSF-1A(4/18). There was no correlation between asbestos exposure, SV40, p53 expression, p16 INK4 and RASSF-1A methylation. Conclusions: Favorable prognostic factors were small size (<2cm), epithelioid histological type and prominent chronic inflammatiory infiltrate. Immunohistochemical markers did not differ significantly from mesothelioma with poor prognosis. p16/CDKN2A and RASSF1A promoter methylation are significantly correlated in the process of tumorigenesis of mesothelioma (p<0.0006).
237
Accuracy of pathological diagnosis of mesothelioma (MM) in Japan
K. Inai, Y. Takeshima, K. Kushitani. Hiroshima University, Hiroshima, Japan
235
Diagnostic value of proliferative markers Ki-67 and Repp 86 in distinguishing between malignant and benign mesothelial proliferations
Z. Mohammadtaheri, M. Mehrafza, F. Mohammadi, M. Khodami, M. Masjedi, M. Bahadori. National Research Institude of Tuberculosis and Lung Disease, Teheran, Iran Background: The differentiation of benign pleural reactions from malignant mesothelioma may be difficult, especially in small biopsy specimens. Purpose: to evaluate 2 proliferative markers (Ki-67 and Repp 86) to distinguish between malignant mesothelioma (MM) and reactive mesothelial hyperplasia (MH). Methods: Thirty-six cases of malignant mesothelioma, including 26 men and 10 women, median age 62.9 (range 36-80 years) and 22 reactive mesothelial hyperplasia cases were investigated for proliferative status by immunohistochemistry method with two monoclonal antibodies; mAb Ki-S2 (repp 86) and Ki-S5 (Ki-67) and the labeling indices (LI) were calculated. Results: Both markers (Ki- 67 and Repp 86) show significant correlation with each other in differentiation of MM and MH (r= 77.5, p value < 0.0001). Average count reveals a significant increase in MM compared with reactive MH (p value < 0.0001). Assuming a threshold of 9% with both Ki-67 and Repp 86 a sensitivity of 88% and 92% and specificity of 94% respectively resulted. Conclusions: Both proliferative markers can be useful in distinction between malignant mesothelioma and reactive mesothelial hyperplasia (p value < 0.0001).
Background: The clinical diagnosis of MM is dependent on imaging including x-ray, CT or MRI, because the serum markers are not reliable at this time and so, definitive diagnosis by pathological examination has been required. However, there are many histological varieties in MM and the diagnostic criteria are not so clear and definite pathological diagnosis is not so easy. Therefore, we tried to make an antibody panel for immunohistochemistry in the differential diagnosis of MM and examined accuracy of pathological diagnosis of MM in Japan. Methods: Eighty-eight cases of epithelioid MM were compared to 51 cases of pulmonary adenocarcinoma, and 41 cases of sarcomatoid MM were compared to 47 cases of various types of sarcoma. In these cases, the representative tissue blocks were used for immunohistochemical stainings by S.A.B. method. In addition, 111 permitted cases among 878 cases with death certificate (2003) in Japan were examined in our laboratory to ascertain accuracy of the diagnosis of MM. Results: The antibody panel of calretinin, WT1 and CEA is useful for the differential diagnosis between epithelioid MM and pulmonary adenocarcinoma. On the other hand, the panel of CAM5.2, AE1/AE3 and other specified antibody positive for sarcomas is useful for the differential diagnosis between sarcomatoid MM and sarcoma. Using our criteria including immmunohistochemistry mentioned above, it was considered that 14 cases (13%) among 111 examined cases was diagnosed as unlikely or definitely not MM, and it is supposed that the diagnosis of MM in Japan is not correct in the proportion of 10-15%. Conclusions: For the accurate diagnosis of MM, immunohistochemistry using the special antibody panel according to histological type is indispensable. Knowledge of immunohistochemistry is essential for improvement of the accuracy of pathological diagnosis of MM.