Histopathologg 1992, 21, 331-334
p53 expression is common in malignant rnesothelioma G .KAFIRI, D.M.THOMAS, N. A.SHEPHERD*, T.KRAUSZ, D.P .LANE? & P. A .HALL* Department of Histopathology, Royal Postgraduate Medical School, London and *Department of Histopathology. Gloucester Royal Hospital, Gloucester and tGll Transformation Research Group, CRC Laboratories, Department of Biochemistry, Dundee University, Dundee and $Department of Histopathology, UMDS, St Thomas's Hospital Campus, London, UK Date of submission 5 November 1991 Accepted for publication 1 May 1992
KAPIRI G . . THOMAS D.M., SHEPHERD N . A . , KRAUSZ T . , LANE D.P. & HALL P . A .
(1992) Histopathology 21, 331-334
p53 expression is common in malignant mesothelioma The p53 tumour suppressor gene has been shown to be frequently mutated in a wide range of human neoplasms. This is accompanied by increased levels of p5 3 protein which become immunologically detectable in pathological material. We have investigated the possibility that the differential diagnosis between reactive and neoplastic mesothelium might be resolved using a polyclonal serum raised to human p53 protein, CM-1.None of 20 cases of reactive mesothelial proliferation showed p53 immunoreactivity while 70% (14 of 20) of cases of malignant mesothelioma showed p53 staining. We can thus infer that abnormalities of p53 appear to be a common event in malignant mesothelioma and that p53 immunostaining may be of value in the distinction of malignant mesothelioma from reactive hyperplasia. Keywords: tumour suppressor gene, p5 3, mesothelioma
Introduction p5 3 is encoded by a tumour suppressor gene discovered by Lane & Crawford' as a consequence of its protein product binding to the SV40 virus large T antigen. For some time this nuclear protein was believed to have a role in neoplasia as a dominantly transforming oncoprotein, but there is now considerable evidence pointing to p53 being a tumour suppressor genezs3.Mutation of p53 has been recently reported to be a frequent event in a range of human tumour types2e3,including b r e a s P , stomach7,lung8v9, pancreas12,l i ~ e r ' ~ .uter'~, us7, brain and bladder cancerlo, melanoma', ovary and testis cancer15J6,and bone and soft tissue sarcomas17. The majority of abnormalities are mis-sense mutations that inactivate the tumour suppressor activity of p53, although growth promoting activity of mutant p53 has Address for correspondence: Professor P.A.Hal1. Department of Histopathology. UMDS, St Thomas's Hospital Campus, Lambeth Palace Road, London SEl 7EH. UK.
also been des~ribed~.~. Wild type p53 has a very short half-life and is not detectable by immunological methods. It has been consistently found that mutant protein, which takes on an abnormal conformation, is more stable, than wild type protein and accumulates in neoplastic ~ e l l s ~ . to ~ .become ~ . ' ~ immunologicallydetectable. The identification of p53 may thus prove to be a useful marker of neoplasia in human pathological material19. The differential diagnosis between reactive mesothelial hyperplasia and malignant mesothelioma can frequently be difficult and is conventionally based upon a combination of clinical and morphological features such as papillary foci, necrosis, nuclear atypia and abnormal mitoses20*21. Immunohistochemical studies have in the past been advocated to assist with the differential diagnosis but have not been confirmed as being Consequently, we have investigated the possible diagnostic utility of employing the immunolocal i t i o n of p53 as an adjunct to morphology in the differential diagnosis of mesothelioma. 331
332 G.Kafiri et al.
Materials and methods Formalin-fixed and wax-embedded blocks from 20 cases of pleural mesothelioma were obtained from the departments of histopathology at the Royal Postgraduate Medical School and Gloucester Royal Hospital. The diagnosis of mesothelioma, in all cases, was based upon the clinical findings, morphological features and histochemical and immunohistochemical profile. Twenty cases of reactive mesothelial proliferation were also examined. STAINING TECHNIQUE
Sections (4 pm thick) were cut and dried at 37OC overnight. After blocking endogenous peroxidase with 30% hydrogen peroxide for 30 min, the sections were immunostained by the ABC method. The primary antibody was CM-1, a rabbit polyclonal antiserum raised against genetically engineered human p5 3 expressed in E. cdi7 (Midgley et al. In preparation). Primary antibody was supplied at a dilution of 1:1000, incubated overnight at 4°C and subsequently detected by biotinylated swine anti-rabbit serum at a dilution of 1:500 followed by ABC complex (DakopattsLtd, High Wycombe, Bucks, UK) using standard protocols. ABC complex was visua: lized with di-amino benzidine and hydrogen peroxide. Sections were then lightly counterstained with Harris's haematoxylin, dehydrated and mounted. In all experiments negative controls consisted of substitution of primary antiserum with phosphate buffered saline and positive controls involved the use of cases of human colon carcinoma previously shown to have mutant p5 3. Histochemical characterization of the tumours was performed using standard methodsz3.Antibodies used to characterize tumours included CAM 5.2 (Becton Dickinson, Mountain View, CA, USA),Leu M 1 (Becton Dickinson, Mountain View, CA, USA), epithelial membrane antigen (EMA) (Dakopatts Ltd, High Wycombe, Bucks, UK), AUAl (generous gift of the Imperial Cancer Research Fund), carcino-embryonic antigen (CEA) (DakopattsLtd, High Wycombe,Bucks, UK) and staining was performed using an ABC technique.
age 61.6 years: range 42-82), all but one of whom were male. The patients presented typically with pleural effusions or ascites and weight loss. The morphological features, both macroscopically and microscopically, were typical of mesothelioma. The histochemical profile was diastase-periodic acid-Schfl negative and alcian blue with hyaluronidase treatment negative. Immunophenotyping showed low molecular weight cytokeratin (CAM 5.2) immunoreactivity, but absence of CEA, Leu M1, AUAl and EMA. In 10 cases the diagnosis was confirmed at autopsy. In no case of reactive mesothelium was there any detectable p53 immunoreactivity. In contrast, in 1 4 of the 20 cases (70%) of mesothelioma nuclear p53 immunoreactivity could be identified (Figure 1).Three positive cases showed expression in more than 75% of neoplastic cells, whereas in the other 11 cases only occasional tumour cells showed staining, ranging from 1 to 30% of neoplastic cells. In all cases there was evidenceof heterogeneity of expression of p5 3 with areas of negativity and foci of staining.
Discussion There is considerable evidence that mutation of the p53 gene is a common event in human neopla~ia~.~*'~'~.
ASSESSMENT
Morphological diagnosis was made independently of assessment of p53 immunostaining by two observers (TK81NAS). p53 immunostaining was assessed independently by two other observers (GK & PAH).
Results The 20 mesotheliomas arose in elderly patients (mean
Flgure 1. Nuclear p53 immunoreactivity in the nuclei of neoplastic mesothelial cells. Immunoperoxidase with haematoxylin counterstain.
p53 in mesothelioma
333
Furthermore there is a strong association between al.) has now been published: Midgley CA, Fisher CJ, detectable p53 immunoreactivity and the presence of Bartek J, Vojtesek B, Lane DP, Barnes DM. Analysis of m ~ t a t i o n ~ * ~The * ’ *molecular . basis of this appears to p53 expression in human tumours: an antibody raised involve altered protein conformation and subsequent against human p53 expressed in E. coli. I. Cell Science prolongation of protein half-life with accumulation of 1992; 101; 183-189. p5 3 protein, although this might in a small proportion of cases occur as a consequence of other molecular abnormalities, including alterations of degradative pathways. References This allows the pathologist to employ the detection of 1. Lane DP. Crawford LV. T antigen is bound to host protein in SVp53 as a marker of p53 mutation. A number of studies 40-transformed cells. Nature 1979: 278; 261-263. have indicated the general validity of this approach in a 2. Lane DP. Benchimol S. p53: oncogene or anti-oncogene? Genes range of tumour types. In those cases where sequencing D~v.1990; 4; 1-8. 3. Levine A]. Momand J, Finlay CA. The p53 tumour suppressor of tumours with p53 immunoreactivity has been pergene. Nature 1991; 351; 453-456. formed, mutations have been identified2*3*6-8*12. Until 4. Cattoretti G, Rilke F. Andreola S, D’Amato L. Delia D. p53 recently antibodies that recognize p5 3 would only work expression in breast cancer. Int. 1. Cancer 1988: 41; 178-183. in cryostat sections, thus limiting the range of studies 5. Bartek J, Bartkova 1, Vojtesek B et al. Patterns of expressionof the that could be performed and the applicability to surgical p53 tumour-suppressor in human breast tissues in situ and in vitro. Int. 1. Cancer 1990: 46; 839-844. pathology. The availability of an anti-serum raised 6. Bartek J, Iggo R. Gannon J. Lane DP. Genetic and immunocytocheagainst genetically engineered human p5 3 that is active mica1 analysis of mutant p53 in human breast cancer cell lines. in conventionally fixed and wax-embedded material Oncogene 1 9 9 0 5; 893-899. greatly extends the range of material that can be 7. Bartek J, Bartkova J. Vojtesek B et al. Aberrant expression of the investigated, although it may under-estimate the true p53 oncoprotein is a common feature of a wide spectrum of level of expression, possibly as a consequence of partial human malignancies. Oncogene 1991: 6 1699-1703. 8. lggo R, Gatter KC. Bartek J. Lane DP, Harris AL. Increased alteration of the p53 protein by cross-linking fixatives12. expression of the mutant forms of p5 3 oncogene in primary lung In this study we have demonstrated p5 3 immunoreaccancer. Lancet 1 9 9 0 335; 675-679. tivity in 70% of mesotheliomas. A recent molecular 9. Chiba I. Takahashi T. Nau M et al. Mutations in the p53 gene are analysis has confirmed these data24. frequent in primary, resected non-smallcell lung cancer. Oncogene In the context of surgical pathology the high fre1 9 9 0 5; 1603-1610. 10. Nigro JM, Baker SJ,Preisinger AC et al. Mutations in the p53 gene quency of p53 immunoreactivity in malignant mesotheoccur in diverse human tumour types. Nature 1989: 342; 705lioma and its absence in reactive mesothelial prolifera708. tions suggests that this will be a useful adjunct to 11. Rodrigues N. Rowan A. Smith MEF et. al. p53 mutations in morphological diagnosis. However, p5 3 immunoreaccolorectal cancer. Proc. Natl. Acad. Sci. USA 1 9 9 0 87; 7555tivity is common in adenocarcinoma of the lung; thus 7559. 12. Barton CM.StaddonSL. Hughes CM et al. Abnormalitiesof the p5 3 p53 staining will not assist in this differentialdiagnosis. tumour suppressorgene in human pancreatic cancer. Br. 1.Cancer It is important to recognize, however, that the absence of 1991: 6 4 1076-1082. p53 immunoreactivity does not exclude a diagnosis of 13. Bressac €3, Keco M. Wands J, Ozturk M. Selective G to T mutations neoplasia. In addition, the presence of heterogeneity of p53 gene in hepatocellular carcinoma from Southern Africa. of p53 staining may lead to problems of sampling. It Nature 1991; 350; 429-431. 14. Hsu IC. Metcalf RA, Sun T. Welsh JA. Wang NJ, Harris CC. has recently been shown that p53 immunostaining may be a generally valuable technique in cytopathol~gy~~ Mutational hotspot in the p53 gene in human hepatocellular carcinomas. Nature 1991: 350; 427-428. and therefore p5 3 immunolocalizationmay assist in the 15. Bartkova J. Bartek J, Lukas J et al. p53 proteii alterations in differential diagnosis of mesothelioma and reactive human testicular cancer including preinvasive intratubular germ mesothelial cells in cytological preparations. cell neoplasia. Int. J. Cancer In press.
Addendum Since submission of this manuscript it has become clear that situations exist where p53 immunoreactivity is detectable without mutation of the p53 gene. This does not invalidate the observations reported here, but cautions against the uncritical equating of p5 3 staining and mutation of the p53 gene. The manuscript described in preparation (Midgley et
16. Marks JR. Davidoff AM, Kerns BJ et al. Overexpression and mutation of p5 3 in epithelial ovarian cancer. Cancer Res. 1991: 51; 2979-2984. 17. Stratton MR. Moss P.Warren W et al. Mutation of the p53 gene in human soft tissue sarcomas: association with abnormalitiesRB-1 gene. Oncogene 1990: 5; 1297-1301. 18. Gannon JV, Greaves R. Iggo R. Lane DP. Activating mutations in p53 produce a common conformational effect. A monoclonal antibody speciRc for the mutant form. EMBO 1. 1990: 9; 15951602. 19. Lane DP. Mutation of the p5 3 gene and accumulation of the p5 3 protein:common steps found in the majority of human cancers. In
334
GXafri et al.
Fortner JG, Rhodes JE eds. Accomplishments in Cancer Research. Philadelphia: JB Lippincott Co.. 1 9 9 0 252-266. 20. McCaughlhey WTE, Al-Jabi M. DUTerentiation of serosal hyperplasia and neoplasia in biopsies. Patho/. Annu. 1986: 21; 271293. 21. Bolen JW, Hammar SP. McNutt MA. Reactive and neoplastic serosal tissue. A light, ultrastructural and immunocytochemical study. Am. 1. Surg. Puthol. 1986; 1 0 34-47. 22. Soosay GN, GrifEths M.Papadaki L,Happerfireld L,Bobrow L. The differential diagnosis of epithelial type mesothelioma from adenocarcinoma and reactive mesothelial proliferation.1. Patho/. 1991: 163: 299-305.
23. Ghosh AK, Gatter KC, Dunnill MS,Mason DY. Immunohlstochemica1 staining of reactive mesotheiium, mesotheliorna and lung carcinoma with a panel of monoclonal antibodies. 1. Clin. Puthol. 1987: 40; 19-25. 24. Cote RJ, Jhanwar SC, Novick S,Pellicer A. Genetic alterations of the p53 gene are a featureofmalignant rnesothelioma.Cancer Res. 1991: 51; 5410-5416. 25. Hall PA, Ray A, Lemoine NR.Midgley CA. Krausz T. Lane DP. p53 immunostaining as a marker of malignant disease in diagnostic cytopathology. Lancet 1991: 338; 513.
p53 expression is common in malignant rnesothelioma G .KAFIRI, D.M.THOMAS, N. A.SHEPHERD*, T.KRAUSZ, D.P .LANE? & P. A .HALL* Department of Histopathology, Royal Postgraduate Medical School, London and *Department of Histopathology. Gloucester Royal Hospital, Gloucester and tGll Transformation Research Group, CRC Laboratories, Department of Biochemistry, Dundee University, Dundee and $Department of Histopathology, UMDS, St Thomas's Hospital Campus, London, UK Date of submission 5 November 1991 Accepted for publication 1 May 1992
KAPIRI G . . THOMAS D.M., SHEPHERD N . A . , KRAUSZ T . , LANE D.P. & HALL P . A .
(1992) Histopathology 21, 331-334
p53 expression is common in malignant mesothelioma The p53 tumour suppressor gene has been shown to be frequently mutated in a wide range of human neoplasms. This is accompanied by increased levels of p5 3 protein which become immunologically detectable in pathological material. We have investigated the possibility that the differential diagnosis between reactive and neoplastic mesothelium might be resolved using a polyclonal serum raised to human p53 protein, CM-1.None of 20 cases of reactive mesothelial proliferation showed p53 immunoreactivity while 70% (14 of 20) of cases of malignant mesothelioma showed p53 staining. We can thus infer that abnormalities of p53 appear to be a common event in malignant mesothelioma and that p53 immunostaining may be of value in the distinction of malignant mesothelioma from reactive hyperplasia. Keywords: tumour suppressor gene, p5 3, mesothelioma
Introduction p5 3 is encoded by a tumour suppressor gene discovered by Lane & Crawford' as a consequence of its protein product binding to the SV40 virus large T antigen. For some time this nuclear protein was believed to have a role in neoplasia as a dominantly transforming oncoprotein, but there is now considerable evidence pointing to p53 being a tumour suppressor genezs3.Mutation of p53 has been recently reported to be a frequent event in a range of human tumour types2e3,including b r e a s P , stomach7,lung8v9, pancreas12,l i ~ e r ' ~ .uter'~, us7, brain and bladder cancerlo, melanoma', ovary and testis cancer15J6,and bone and soft tissue sarcomas17. The majority of abnormalities are mis-sense mutations that inactivate the tumour suppressor activity of p53, although growth promoting activity of mutant p53 has Address for correspondence: Professor P.A.Hal1. Department of Histopathology. UMDS, St Thomas's Hospital Campus, Lambeth Palace Road, London SEl 7EH. UK.
also been des~ribed~.~. Wild type p53 has a very short half-life and is not detectable by immunological methods. It has been consistently found that mutant protein, which takes on an abnormal conformation, is more stable, than wild type protein and accumulates in neoplastic ~ e l l s ~ . to ~ .become ~ . ' ~ immunologicallydetectable. The identification of p53 may thus prove to be a useful marker of neoplasia in human pathological material19. The differential diagnosis between reactive mesothelial hyperplasia and malignant mesothelioma can frequently be difficult and is conventionally based upon a combination of clinical and morphological features such as papillary foci, necrosis, nuclear atypia and abnormal mitoses20*21. Immunohistochemical studies have in the past been advocated to assist with the differential diagnosis but have not been confirmed as being Consequently, we have investigated the possible diagnostic utility of employing the immunolocal i t i o n of p53 as an adjunct to morphology in the differential diagnosis of mesothelioma. 331
332 G.Kafiri et al.
Materials and methods Formalin-fixed and wax-embedded blocks from 20 cases of pleural mesothelioma were obtained from the departments of histopathology at the Royal Postgraduate Medical School and Gloucester Royal Hospital. The diagnosis of mesothelioma, in all cases, was based upon the clinical findings, morphological features and histochemical and immunohistochemical profile. Twenty cases of reactive mesothelial proliferation were also examined. STAINING TECHNIQUE
Sections (4 pm thick) were cut and dried at 37OC overnight. After blocking endogenous peroxidase with 30% hydrogen peroxide for 30 min, the sections were immunostained by the ABC method. The primary antibody was CM-1, a rabbit polyclonal antiserum raised against genetically engineered human p5 3 expressed in E. cdi7 (Midgley et al. In preparation). Primary antibody was supplied at a dilution of 1:1000, incubated overnight at 4°C and subsequently detected by biotinylated swine anti-rabbit serum at a dilution of 1:500 followed by ABC complex (DakopattsLtd, High Wycombe, Bucks, UK) using standard protocols. ABC complex was visua: lized with di-amino benzidine and hydrogen peroxide. Sections were then lightly counterstained with Harris's haematoxylin, dehydrated and mounted. In all experiments negative controls consisted of substitution of primary antiserum with phosphate buffered saline and positive controls involved the use of cases of human colon carcinoma previously shown to have mutant p5 3. Histochemical characterization of the tumours was performed using standard methodsz3.Antibodies used to characterize tumours included CAM 5.2 (Becton Dickinson, Mountain View, CA, USA),Leu M 1 (Becton Dickinson, Mountain View, CA, USA), epithelial membrane antigen (EMA) (Dakopatts Ltd, High Wycombe, Bucks, UK), AUAl (generous gift of the Imperial Cancer Research Fund), carcino-embryonic antigen (CEA) (DakopattsLtd, High Wycombe,Bucks, UK) and staining was performed using an ABC technique.
age 61.6 years: range 42-82), all but one of whom were male. The patients presented typically with pleural effusions or ascites and weight loss. The morphological features, both macroscopically and microscopically, were typical of mesothelioma. The histochemical profile was diastase-periodic acid-Schfl negative and alcian blue with hyaluronidase treatment negative. Immunophenotyping showed low molecular weight cytokeratin (CAM 5.2) immunoreactivity, but absence of CEA, Leu M1, AUAl and EMA. In 10 cases the diagnosis was confirmed at autopsy. In no case of reactive mesothelium was there any detectable p53 immunoreactivity. In contrast, in 1 4 of the 20 cases (70%) of mesothelioma nuclear p53 immunoreactivity could be identified (Figure 1).Three positive cases showed expression in more than 75% of neoplastic cells, whereas in the other 11 cases only occasional tumour cells showed staining, ranging from 1 to 30% of neoplastic cells. In all cases there was evidenceof heterogeneity of expression of p5 3 with areas of negativity and foci of staining.
Discussion There is considerable evidence that mutation of the p53 gene is a common event in human neopla~ia~.~*'~'~.
ASSESSMENT
Morphological diagnosis was made independently of assessment of p53 immunostaining by two observers (TK81NAS). p53 immunostaining was assessed independently by two other observers (GK & PAH).
Results The 20 mesotheliomas arose in elderly patients (mean
Flgure 1. Nuclear p53 immunoreactivity in the nuclei of neoplastic mesothelial cells. Immunoperoxidase with haematoxylin counterstain.
p53 in mesothelioma
333
Furthermore there is a strong association between al.) has now been published: Midgley CA, Fisher CJ, detectable p53 immunoreactivity and the presence of Bartek J, Vojtesek B, Lane DP, Barnes DM. Analysis of m ~ t a t i o n ~ * ~The * ’ *molecular . basis of this appears to p53 expression in human tumours: an antibody raised involve altered protein conformation and subsequent against human p53 expressed in E. coli. I. Cell Science prolongation of protein half-life with accumulation of 1992; 101; 183-189. p5 3 protein, although this might in a small proportion of cases occur as a consequence of other molecular abnormalities, including alterations of degradative pathways. References This allows the pathologist to employ the detection of 1. Lane DP. Crawford LV. T antigen is bound to host protein in SVp53 as a marker of p53 mutation. A number of studies 40-transformed cells. Nature 1979: 278; 261-263. have indicated the general validity of this approach in a 2. Lane DP. Benchimol S. p53: oncogene or anti-oncogene? Genes range of tumour types. In those cases where sequencing D~v.1990; 4; 1-8. 3. Levine A]. Momand J, Finlay CA. The p53 tumour suppressor of tumours with p53 immunoreactivity has been pergene. Nature 1991; 351; 453-456. formed, mutations have been identified2*3*6-8*12. Until 4. Cattoretti G, Rilke F. Andreola S, D’Amato L. Delia D. p53 recently antibodies that recognize p5 3 would only work expression in breast cancer. Int. 1. Cancer 1988: 41; 178-183. in cryostat sections, thus limiting the range of studies 5. Bartek J, Bartkova 1, Vojtesek B et al. Patterns of expressionof the that could be performed and the applicability to surgical p53 tumour-suppressor in human breast tissues in situ and in vitro. Int. 1. Cancer 1990: 46; 839-844. pathology. The availability of an anti-serum raised 6. Bartek J, Iggo R. Gannon J. Lane DP. Genetic and immunocytocheagainst genetically engineered human p5 3 that is active mica1 analysis of mutant p53 in human breast cancer cell lines. in conventionally fixed and wax-embedded material Oncogene 1 9 9 0 5; 893-899. greatly extends the range of material that can be 7. Bartek J, Bartkova J. Vojtesek B et al. Aberrant expression of the investigated, although it may under-estimate the true p53 oncoprotein is a common feature of a wide spectrum of level of expression, possibly as a consequence of partial human malignancies. Oncogene 1991: 6 1699-1703. 8. lggo R, Gatter KC. Bartek J. Lane DP, Harris AL. Increased alteration of the p53 protein by cross-linking fixatives12. expression of the mutant forms of p5 3 oncogene in primary lung In this study we have demonstrated p5 3 immunoreaccancer. Lancet 1 9 9 0 335; 675-679. tivity in 70% of mesotheliomas. A recent molecular 9. Chiba I. Takahashi T. Nau M et al. Mutations in the p53 gene are analysis has confirmed these data24. frequent in primary, resected non-smallcell lung cancer. Oncogene In the context of surgical pathology the high fre1 9 9 0 5; 1603-1610. 10. Nigro JM, Baker SJ,Preisinger AC et al. Mutations in the p53 gene quency of p53 immunoreactivity in malignant mesotheoccur in diverse human tumour types. Nature 1989: 342; 705lioma and its absence in reactive mesothelial prolifera708. tions suggests that this will be a useful adjunct to 11. Rodrigues N. Rowan A. Smith MEF et. al. p53 mutations in morphological diagnosis. However, p5 3 immunoreaccolorectal cancer. Proc. Natl. Acad. Sci. USA 1 9 9 0 87; 7555tivity is common in adenocarcinoma of the lung; thus 7559. 12. Barton CM.StaddonSL. Hughes CM et al. Abnormalitiesof the p5 3 p53 staining will not assist in this differentialdiagnosis. tumour suppressorgene in human pancreatic cancer. Br. 1.Cancer It is important to recognize, however, that the absence of 1991: 6 4 1076-1082. p53 immunoreactivity does not exclude a diagnosis of 13. Bressac €3, Keco M. Wands J, Ozturk M. Selective G to T mutations neoplasia. In addition, the presence of heterogeneity of p53 gene in hepatocellular carcinoma from Southern Africa. of p53 staining may lead to problems of sampling. It Nature 1991; 350; 429-431. 14. Hsu IC. Metcalf RA, Sun T. Welsh JA. Wang NJ, Harris CC. has recently been shown that p53 immunostaining may be a generally valuable technique in cytopathol~gy~~ Mutational hotspot in the p53 gene in human hepatocellular carcinomas. Nature 1991: 350; 427-428. and therefore p5 3 immunolocalizationmay assist in the 15. Bartkova J. Bartek J, Lukas J et al. p53 proteii alterations in differential diagnosis of mesothelioma and reactive human testicular cancer including preinvasive intratubular germ mesothelial cells in cytological preparations. cell neoplasia. Int. J. Cancer In press.
Addendum Since submission of this manuscript it has become clear that situations exist where p53 immunoreactivity is detectable without mutation of the p53 gene. This does not invalidate the observations reported here, but cautions against the uncritical equating of p5 3 staining and mutation of the p53 gene. The manuscript described in preparation (Midgley et
16. Marks JR. Davidoff AM, Kerns BJ et al. Overexpression and mutation of p5 3 in epithelial ovarian cancer. Cancer Res. 1991: 51; 2979-2984. 17. Stratton MR. Moss P.Warren W et al. Mutation of the p53 gene in human soft tissue sarcomas: association with abnormalitiesRB-1 gene. Oncogene 1990: 5; 1297-1301. 18. Gannon JV, Greaves R. Iggo R. Lane DP. Activating mutations in p53 produce a common conformational effect. A monoclonal antibody speciRc for the mutant form. EMBO 1. 1990: 9; 15951602. 19. Lane DP. Mutation of the p5 3 gene and accumulation of the p5 3 protein:common steps found in the majority of human cancers. In
334
GXafri et al.
Fortner JG, Rhodes JE eds. Accomplishments in Cancer Research. Philadelphia: JB Lippincott Co.. 1 9 9 0 252-266. 20. McCaughlhey WTE, Al-Jabi M. DUTerentiation of serosal hyperplasia and neoplasia in biopsies. Patho/. Annu. 1986: 21; 271293. 21. Bolen JW, Hammar SP. McNutt MA. Reactive and neoplastic serosal tissue. A light, ultrastructural and immunocytochemical study. Am. 1. Surg. Puthol. 1986; 1 0 34-47. 22. Soosay GN, GrifEths M.Papadaki L,Happerfireld L,Bobrow L. The differential diagnosis of epithelial type mesothelioma from adenocarcinoma and reactive mesothelial proliferation.1. Patho/. 1991: 163: 299-305.
23. Ghosh AK, Gatter KC, Dunnill MS,Mason DY. Immunohlstochemica1 staining of reactive mesotheiium, mesotheliorna and lung carcinoma with a panel of monoclonal antibodies. 1. Clin. Puthol. 1987: 40; 19-25. 24. Cote RJ, Jhanwar SC, Novick S,Pellicer A. Genetic alterations of the p53 gene are a featureofmalignant rnesothelioma.Cancer Res. 1991: 51; 5410-5416. 25. Hall PA, Ray A, Lemoine NR.Midgley CA. Krausz T. Lane DP. p53 immunostaining as a marker of malignant disease in diagnostic cytopathology. Lancet 1991: 338; 513.
