Bcl-2 expression in pancreas development and pancreatic cancer progression

Apoptosis is important for both tissue development and differentiation; its deregulation may contribute to tumourigenesis. In order to clarify the role of Bcl-2, an apoptosis-inhibiting protein, in pancreatic morphogenesis and tumour progression, its immunohistochemical expression was evaluated in 12 samples of fetal pancreas, in 10 samples of adult pancreas with ductal hyperplastic lesions, in 120 cases of primary pancreatic ductal adenocarcinoma, and in 43 synchronous metastatic lymph nodes. To evaluate the role of apoptosis in pancreatic cancer, p53 expression was also studied in tumour samples. Bcl-2 cytoplasmic acinar and ductal immunostaining was found in all fetal and adult tissue samples; ductal hyperplastic lesions were constantly negative. Thirty out of 120 (25%) tumours and 3 out of 43 (7%) lymph nodes expressed Bcl-2, whereas 67 out of 120 (56%) expressed nuclear p53. Well-differentiated tumours (G1) were more frequently Bcl-2-positive (p=0.002); furthermore, there was an inverse correlation between Bcl-2 and p53 expression in primary tumours (p=0.02). Neither Bcl-2 nor p53 influenced patients' prognosis, which was instead affected by N (p=0.02) and M (p<0.0001) status and stage of the disease (p=0.002). It is concluded that Bcl-2 regulates pancreatic morphogenesis and tissue homeostasis from early fetal to adult life and can be considered a phenotypic marker of normal exocrine pancreas. On the other hand, the lack of expression in preneoplastic lesions and the low positivity found in primary tumours and lymph node metastases suggest that Bcl-2 does not play a centralrole in pancreatic tumourigenesis and cancer progression.     

Apoptosis and expression of Bax, Bcl-x, and Bcl-2 apoptotic regulatory proteins in colorectal carcinomas, and association with p53 genotype/phenotype.

AIMS: Spontaneous apoptosis and expression of the apoptotic regulatory proteins Bax, Bcl-x, and Bcl-2 were investigated in 50 colorectal carcinomas. The p53 genotypes/phenotypes and BAX genotypes were also determined, and possible associations of these with apoptosis and/or with expression of the different apoptotic regulatory proteins were studied. METHODS: Terminal deoxynucleotidyl transferase (TdT) mediated dUTP labelling of DNA fragments was used to detect apoptotic tumour cells in sections and peroxidase immunohistochemistry was used to assess protein expression. p53 genotype/phenotype was determined using constant denaturant gel electrophoresis/immunoblotting and bax genotype was determined using polymerase chain reaction based methods. RESULTS: The distribution of tumour apoptotic indices was bimodal with a natural cut off at 1.0% (range, 0.0-5.4%); the median fraction of apoptotic tumour cells was 0.8%. Tumour apoptosis was not associated significantly with tumour DNA ploidy status. Normal mucosal tissue had less than 0.1% apoptotic cells. Staining intensities for Bax, Bcl-x, and Bcl-2 were strong; that is, equivalent to or greater than positive normal mucosal cells, in 11 of 50, 20 of 49, and 20 of 48 carcinomas. Frameshift mutations in the bax gene were detected in three of 42 tumours analysed, all of which were DNA diploid, and Bax protein expression in these tumours was absent or very low. Bax, Bcl-x, and Bcl-2 protein expression were not correlated with tumour apoptosis or tumour DNA ploidy status. p53 was expressed in 34 of 50 tumours and p53 gene mutations were detected in 22 of 29 p53 positive tumours analysed. Apoptosis was significantly lower in a greater number of p53 positive tumours than p53 negative tumours. In addition, Bcl-2 protein expression was significantly higher in a greater number of p53 positive tumours compared with p53 negative tumours. Bax and Bcl-x protein expression were not significantly associated with p53 phenotype/genotype. CONCLUSIONS: The results indicate that acquisition of a p53 phenotype is associated with lower spontaneous apoptosis and higher expression of Bcl-2. The results also suggest that p53 is not a major determinant for Bax expression in colorectal carcinomas in vivo.     

Apoptosis and expression of Bax, Bcl-x, and Bcl-2 apoptotic regulatory proteins in colorectal carcinomas, and association with p53 genotype/phenotype.

AIMS: Spontaneous apoptosis and expression of the apoptotic regulatory proteins Bax, Bcl-x, and Bcl-2 were investigated in 50 colorectal carcinomas. The p53 genotypes/phenotypes and BAX genotypes were also determined, and possible associations of these with apoptosis and/or with expression of the different apoptotic regulatory proteins were studied. METHODS: Terminal deoxynucleotidyl transferase (TdT) mediated dUTP labelling of DNA fragments was used to detect apoptotic tumour cells in sections and peroxidase immunohistochemistry was used to assess protein expression. p53 genotype/phenotype was determined using constant denaturant gel electrophoresis/immunoblotting and bax genotype was determined using polymerase chain reaction based methods. RESULTS: The distribution of tumour apoptotic indices was bimodal with a natural cut off at 1.0% (range, 0.0-5.4%); the median fraction of apoptotic tumour cells was 0.8%. Tumour apoptosis was not associated significantly with tumour DNA ploidy status. Normal mucosal tissue had less than 0.1% apoptotic cells. Staining intensities for Bax, Bcl-x, and Bcl-2 were strong; that is, equivalent to or greater than positive normal mucosal cells, in 11 of 50, 20 of 49, and 20 of 48 carcinomas. Frameshift mutations in the bax gene were detected in three of 42 tumours analysed, all of which were DNA diploid, and Bax protein expression in these tumours was absent or very low. Bax, Bcl-x, and Bcl-2 protein expression were not correlated with tumour apoptosis or tumour DNA ploidy status. p53 was expressed in 34 of 50 tumours and p53 gene mutations were detected in 22 of 29 p53 positive tumours analysed. Apoptosis was significantly lower in a greater number of p53 positive tumours than p53 negative tumours. In addition, Bcl-2 protein expression was significantly higher in a greater number of p53 positive tumours compared with p53 negative tumours. Bax and Bcl-x protein expression were not significantly associated with p53 phenotype/genotype. CONCLUSIONS: The results indicate that acquisition of a p53 phenotype is associated with lower spontaneous apoptosis and higher expression of Bcl-2. The results also suggest that p53 is not a major determinant for Bax expression in colorectal carcinomas in vivo.     

Coexpression of p53 and c-erbB-2 proteins is associated with histological type, tumour stage, and cell proliferation in malignant salivary gland tumours

The development and progression of cancer are known to be regulated by various oncogenes and tumour suppressor genes. We analysed 63 primary malignant salivary gland tumours for the expression of p53 and c-erbB-2 proteins. Immunohistochemically, 7 of 63 tumours (11%) showed diffuse nuclear staining for p53 protein, and all 7 were also positive for c-erbB-2 protein. The overexpression of p53 protein correlated closely with the overexpression of c-erbB-2 protein (P<0.001). Overexpression of both p53 and c-erbB-2 proteins (coexpression) was found in tumours of certain histological types, such as adenocarcinoma, carcinoma in pleomorphic adenoma, and salivary duct carcinoma. Furthermore, it is noteworthy that coexpression was associated with high-grade carcinoma, advanced tumour stage, and a high Ki-67 labelling index (%) which is a marker of cell proliferation. In adenocarcinoma, we attempted to clarify the relationship between coexpression and histological grade. Coexpression was associated with histological grades showing high mitotic indices and necrotic areas, which reflected high cell-proliferative activity. These results suggest that the accumulation of genetic alterations, such as those involving p53 and c-erbB-2, plays an important part in the progression of malignant salivary gland tumours.     

Analysis of key cell-cycle checkpoint proteins in colorectal tumours

Aberrations in the components of cell-cycle checkpoints are a common feature of many tumours and several have been shown to have prognostic significance in colorectal cancer. In this study, seven components of cell-cycle control [cyclin D1, retinoblastoma (pRb), p21, p27, p16, p53, and proliferating cell nuclear antigen (PCNA)] were examined in a large series of well-characterized colorectal adenocarcinomas using immunohistochemistry to ascertain co-regulation and influence on survival. The majority (92%) of the tumours had abnormal staining of > or =2 cell-cycle control factors. Expression of cyclin D1 protein was correlated with both p21 (p<0.001) and p27 (p=0.033), suggesting co-regulation of these proteins in colorectal tumours. Only cyclin D1 (p=0.048) and p53 (p=0.025) were directly associated with PCNA levels, suggesting a more important role in the proliferative capacity of tumour cells. Significant associations between cell cycle-related proteins and clinicopathological data were observed: cyclin D1 and p53 proteins were correlated with patient age (p=0.042 and p<0.001, respectively) and p53 (p=0.01) and p21 (p=0.024) proteins were associated with tumour site. Expression of cyclin D1 protein was the only protein examined that was related to improved outcome in these patients (p=0.0266), but it was not an independent predictor of survival. These results suggest that loss of control of cell-cycle checkpoints is a common occurrence in colorectal tumours and may be an important therapeutic target.     

Loss of DCC expression in astrocytomas: relation to p53 abnormalities, cell kinetics, and survival

AIMS: Although frequent reduction or loss of DCC (deleted in colorectal carcinomas) has been demonstrated in gliomas, the association with cell kinetics and survival is still unclear. METHODS: A total of 119 astrocytomas, comprising 39 grade IV, 36 grade III, and 44 low grade tumours, were immunohistochemically investigated, along with 26 normal adult brain samples and two fetal brains. The results were compared with p53 abnormalities, Ki-67 labelling index (LI), mitotic index (MI), apoptotic index (AI), and survival. RESULTS: In normal adult and fetal brain tissues, DCC expression was detected in mature and terminally differentiated neuronal cells but not glial elements. In astrocytomas, whereas DCC expression was still clearly shown with low grade malignancy, DCC scores were significantly decreased in high histological grade malignancy, along with an increase in cell kinetics determined by AI, MI, and Ki-67 LI values. In addition, p53 LI values were significantly increased, although a direct link between DCC scores and p53 LI values was not evident. Univariate analysis revealed that high DCC scores and low p53 LI values were closely related to a favourable outcome for astrocytoma, although only the AI was an independent prognostic factor. CONCLUSIONS: The loss of DCC expression may be closely related to changes in cell kinetics and tumour phenotype in astrocytomas, independent of p53 abnormalities.     

Correlation between cathepsin D expression and p53 protein nuclear accumulation in oesophageal squamous cell carcinoma

AIM: The lysosomal protease cathepsin D has been reported to be associated with tumour progression in malignant tumours. Expression of the gene encoding cathepsin D is known to be stimulated by oestrogen in mammary cancer cells. Recent experiments revealed that a p53 DNA binding site is located in the promoter region of the cathepsin D gene. This fact indicates that cathepsin D expression may correlate with p53 protein expression. The purpose of this study is to evaluate the expression patterns of the cathepsin D and p53 proteins in oesophageal squamous cell carcinoma (SCC). METHODS: In 154 patients with oesophageal SCC, expression of the cathepsin D and p53 proteins was measured in tumours by means of immunohistochemistry using monoclonal antibodies against cathepsin D (clone, 1C11) and p53 (clone, BP53-12). RESULTS: Cathepsin D was detected in tumour cells, although it was not found in normal oesophageal epithelium adjacent to carcinoma. High cathepsin D expression (positive tumour cells > 10%) was detected in 76 of 154 cases (49%) and high p53 nuclear expression (positive tumour cells > 50%) was detected in 70 cases (46%). High cathepsin D expression was significantly associated with invasive tumour growth (p = 0.002), poor prognosis (p = 0.049), and nuclear accumulation of p53 protein (p = 0.001). Overexpression of both p53 and cathepsin D was seen in 45 of the 154 cases (29.2%). In addition, there was a positive correlation between the cathepsin D index (percentage of cathepsin D positive tumour cells) and Ki-67 labelling index (percentage of Ki-67 positive tumour cells) in 154 oesophageal SCCs (rho = 0.257; p = 0.009). However, in multivariate survival analysis, cathepsin D expression by the tumours was not an independent prognostic factor in patients with oesophageal SCC (p = 0.236). CONCLUSIONS: The expression of cathepsin D by cancer cells may play an important role in the invasive growth of oesophageal SCC. Overexpression of both p53 and cathepsin D was seen frequently in tumours; p53 gene abnormalities may correlate with cathepsin D overexpression in oesophageal SCC.     

Abl expression in human fetal and adult tissues,tumours and tumour microvessels

Abl kinases encoded by the abl oncogenes inhibit apoptosis without affecting cell proliferation. The aim of this study was to examine a wide range of normal fetal and adult human tissues and a variety of tumour types for Abl immunoreactivity. Sections from 193 paraffin blocks of normal fetal and adult tissues and 72 blocks from representative tumours were stained immunohistochemically using a polyclonal antibody to c-Abl/Bcr–Abl oncoprotein. Weak Abl immunoreactivity was observed in many adult tissues. Moderately intense or strong staining (cytoplasmic, nuclear or membranous) was consistently seen in hyaline cartilage, adipocytes, and ciliated epithelium. In fetal tissues, there was a broadly similar staining pattern, but Abl expression was also seen in muscle (all types) and occasionally in endothelial cells. The most intense staining was seen in sites of endochondral ossification and in the umbilical cord stroma. Negatively staining tissues included epidermis and squamous mucosa, lymph nodes, tonsil, spleen, hepatocytes, and adrenals. Most tumours showed focal or weak Abl immunoreactivity. The most intense staining was seen in chondrosarcoma, liposarcoma, and diffuse gastric (signet ring) adenocarcinoma. In the latter two tumour types, Abl expression was also observed in tumour microvessels. These results suggest that Abl not only functions as an apoptosis inhibitor, but also may have a role in connective tissue maturation and differentiation and in tumour growth and angiogenesis. © 1997 John Wiley & Sons, Ltd.     

Cytogenetics and Molecular Genetics of Malignant Gliomas and Medulloblastoma

Malignant gliomas and medulloblastomas which are the most common primary malignant brain tumours of adult and children, respectively, resemble other neurogenic tumours as they frequently contain gene amplification and show non-random loss of specific chromosomal regions. In gliomas the gene which is most often amplified, is the epidermal growth factor receptor gene. In many cases the gene is rearranged as well, producing abnormally small epidermal growth factor receptor proteins. More than 80% of tumours have lost chromosome 10 and losses of 9p13, 17p and 22 occur in subgroups of cases. 17p loss is associated with point mutations of the p53 gene, but the relevant genes in the other chromosomal regions remain to be identified. For medulloblastoma the most frequent chromosomal abnormality is i(17q). Whether or not p53 gene mutations are the targets of 17p losses in these tumours remains to be determined. Approximately 5% of medulloblastoma biopsies contain gene amplification, although the incidence in medulloblastoma cell lines is more than 80%. c-myc is the gene which is most frequently amplified in this tumour type. The relationship of these various molecular genetic abnormalities to the biology of the tumours and the course of the patients remains largely unexplored.     

A hybrid carcinoma: adenoid cystic carcinoma and salivary duct carcinoma of the salivary gland

Hybrid tumours of the salivary glands are very rare entities composed of two different tumours, each of which conforms with an exactly defined category. We describe an unusual hybrid carcinoma of the palate; it was comprised of an adenoid cystic carcinoma and a salivary duct carcinoma with a transitional region. These two different compartments showed different characteristics as regards cellular differentiation, proliferative activity, and expression of oncogene and tumour suppressor oncogene proteins, as revealed by using markers for muscle actin, keratin, vimentin, S-100 protein, GFAP, Ki-67, p53, and c-erbB-2 proteins. This case is the first reported with overexpression of p53 and c-erbB-2 proteins in the tumour entities. Salivary gland tumours consist of heterogeneous histological groups, and each has morphological diversity. This case indicates that some of the oncogene and tumour suppressor oncogene proteins may help to produce the histological heterogeneity of the salivary gland tumour.     

Deregulated expression of p16INK4a and p53 pathway members in benign and malignant myoepithelial tumours of the salivary glands

Aims: Myoepithelial salivary gland tumours are uncommon and follow an unpredictable biological course. The aim was to examine their molecular background to acquire a better understanding of their clinical behaviour. Methods and results: Expression of protein (E2F1, p16^INK4a, p53, cyclin D1, Ki67 and Polycomb group proteins BMI‐1, MEL‐18 and EZH2) was investigated in 49 benign and 30 primary malignant myoepithelial tumours and five histologically benign recurrences by immunohistochemistry and the findings correlated with histopathological characteristics. Benign tumours showed a higher percentage of cells with expression of p16^INK4a pathway members [p16^INK4a and E2F1 (both P < 0.001), and cyclin D1, P = 0.002] compared with normal salivary gland. Furthermore, malignant tumours expressed p53 (P = 0.003) and EZH2 (P = 0.09) in a higher percentage. Recurrences displayed more p53 + tumour cells (P = 0.02) than benign primaries. Amongst the benign tumours, the clear cell type had the highest proliferation fraction (P = 0.05) and a higher percentage of EZH2 was detected in the plasmacytoid cell type (P = 0.002). Conclusions: This study is the first to demonstrate that deregulation of the p16^INK4a senescence pathway is involved in the development of myoepithelial tumours. We propose that additional inactivation of p53 in malignant primaries and benign recurrences contributes to myoepithelial neoplastic transformation and aggressive tumour growth.     

The product of the imprinted H19 gene is an oncofetal RNA.

AIMS/BACKGROUND: The H19 gene is an imprinted, maternally expressed gene in humans. It is tightly linked and coregulated with the imprinted, paternally expressed gene of insulin-like growth factor 2. The H19 gene product is not translated into protein and functions as an RNA molecule. Although its role has been investigated for more than a decade, its biological function is still not understood fully. H19 is abundantly expressed in many tissues from early stages of embryogenesis through fetal life, and is down regulated postnatally. It is also expressed in certain childhood and adult tumours. This study was designed to screen the expression of H19 in human cancer and its relation to the expression of H19 in the fetus. METHODS: Using in situ hybridisation with a [35S] labelled probe, H19 mRNA was detected in paraffin wax sections of fetal tissues from the first and second trimesters of pregnancy and of a large array of human adult and childhood tumours arising from these tissues. RESULTS: The H19 gene is expressed in tumours arising from tissues which express this gene in fetal life. Its expression in the fetus and in cancer is closely linked with tissue differentiation. CONCLUSIONS: Based on these and previous data, H19 is neither a tumour suppressor gene nor an oncogene. Its product is an oncofetal RNA. The potential use of this RNA as a tumour marker should be evaluated.     

Interrelationship between Epstein-Barr virus infection in gastric carcinomas and the expression of apoptosis-associated proteins

AIMS: Epstein-Barr virus (EBV) and its associated proteins may be protective against the occurrence of apoptosis that would normally inhibit cancer development and progression. Alternatively, the viral infection may cause altered or mutated expression of oncogenes or tumour suppressor genes that are necessary for tumour development, an action that may also involve apoptosis. In this study, a relationship was sought between occurrence of EBV infection, expression of apoptosis-associated proteins (tumour suppressor gene p53 and oncogenes c-myc and bcl-2) and levels of cell death (apoptosis or necrosis) in 119 cases of gastric carcinoma. METHODS AND RESULTS: The EBV status of the gastric carcinomas (using the EBV-encoded small RNA I (EBER-1) and in-situ hybridization), stage and grade of tumour and sex of patients were compared for bcl-2, p53 and c-myc expression patterns. EBER-1 was detected in approximately 20% of cases studied. There was no significant correlation between levels of cell death in the tumour tissue and EBV status. In the protein analyses, development and progression of gastric carcinoma, with or without EBV infection, was independent of bcl-2 expression. However, in gastric cancers with EBV infection, p53 overexpression was inhibited and c-myc expression was increased in early stage cancers, in comparison with decreased c-myc expression in late stage cancers. CONCLUSIONS: The p53 and c-myc expression patterns indicate that EBV-infected gastric carcinomas are less likely to have a natural regression via apoptosis at an early stage and explain, in part, the resistance to treatment of late stage of gastric cancers.     

Genetic alterations of the tumour suppressor gene regions 3p, 11p, 13q, 17p, and 17q in human breast carcinomas.

Fifty-nine primary breast carcinomas and 11 metastases were examined to identify genetic alterations in the tumour suppressor gene regions 3p, 11p, 13q, 17p, and 17q. Loss of heterozygosity (LOH) was frequently observed on chromosome arms 17p (p144D6 lost in 75%, pYNZ22.1 in 55%, and TP53 in 48% of the primary tumours), 13q (RBI lost in 40% of the primary tumours), and 17q (pRMU3 lost in 35%, pTHH59 in 29%, and NM23HI in 26% of the primary tumours). Loss of all the markers except p144D6 was observed even more frequently in the metastases. Pairwise comparisons for concordance of allele losses on 17p indicated that there might be two genes on 17p implicated in breast cancer development; the TP53 gene and a gene located close to the p144D6 and pYNZ22.1 markers. LOH of the RBI gene was associated with LOH of pYNZ22.1 and p144D6, but not with LOH of TP53. LOH of RBI and TP53 was associated with occurrence of ductal carcinomas, RBI and p144D6 losses with tumour size, and p144D6 losses with positive node status as well. LOH of TP53 and the three 17q markers NM23HI, pTHH59, and pRMU3 was most frequently observed in tumours from postmenopausal women. p144D6 losses occurred most frequently in progesterone receptor-negative tumours, whereas pTHH59 losses occurred most frequently in oestrogen receptor-negative tumours. LOH of the investigated loci was not associated with ERBB2 protooncogene amplification, with positive family history of breast cancer, or with survival.     

Immunohistochemical analysis of bcl-2 and p53 expression in breast carcinomas: their correlation with Ki-67 growth fraction

We examined 59 breast cancers for p53 and bcl-2 protein expression by immunohistochemistry. The results were correlated with Ki-67 immunostaining. p53-negativity was noted in 40 cases and the remaining 19 tumours were p53-positive. Thirty-six tumours showed strong expression of bcl-2 and in 23 no staining for this protein was observed. We found statistically significant reverse correlation between expression of p53 and bcl-2 in majority of carcinomas: 31 cases were bcl-2 positive and p53-negative, and 14 tumours were bcl-2-negative and p53-positive. Six carcinomas showed no nuclear staining for Ki-67 and in the remaining 53 the percent of cancer cells positive for Ki-67 ranged from 1 to 60 (mean: 14.6). In these 53 cases we found that bcl-2-positive tumours were characterized by lower proliferation than bcl-2-negative tumours, the mean value of Ki-67 immunostaining being 10.7% and 23.0%, respectively. p53-negative tumours showed lower proliferation than p53-positive tumours: mean Ki-67 index was 10.2% and 23.9%, respectively.We conclude that immunohistochemically detected p53 and bcl-2 proteins show a significant inverse relationship in majority of breast carcinomas and their expression correlates with tumour proliferation (Ki-67 immunostaining).     

Expression of apoptosis-related proteins in endometriomas and benign and malignant ovarian tumours

Apoptosis is a physiological process by which multicellular organisms eliminate superfluous cells. Alterations in apoptosis play a key role in tumour development. The objective was to evaluate the immunohistochemical expression of p53, p21, bax, bak, fas, bcl-2 and bcl-x proteins in 10 endometriomas, 20 benign ovarian tumours (10 mucinous, 10 serous) and 30 malignant ovarian tumours (9 mucinous, 19 serous; 2 endometrioids). p53 positive cells (mean+/-SD) in endometriomas, and benign and malignant tumours were 1.9+/-3.2, 0 and 16.2+/-33.0, respectively. The difference was significant between benign tumours and endometriomas (P=0.003) but not between endometriomas and malignant tumours. P21 expression in endometriomas and benign and malignant tumours was 19.5+/-27.8, 1.7+/-6.7 and 4.1+/-8.6, respectively. Increased p21 expression was found in endometriomas compared with benign (P=0.001) and malignant (P=0.01) tumours. Bax expression was higher in endometriomas than in benign tumours (P=0.01), but no difference was found between endometriomas and malignant tumours. No difference in bak, fas, bcl-2 or bcl-x expression was observed among the groups. In endometriomas, a negative correlation was found between p53 and fas expression (P=0.04, r=0.66). Although endometriomas have histological features of benign ovarian tumours, endometriomas share with malignant ovarian tumours certain alterations in apoptosis-related proteins.     

Studies on p53 immunolocalisation in breast cancer and its prognostic significance

Immunocytochemical localisation of mutant p53 in breast tumours serves as a potential prognostic molecular marker. In order to study the expression of p53 protein in breast cancer which constitutes the second most common malignancy in the South Indian female population, MAb CIBCVMC12 has been generated against human p53 protein isolated and purified from bacterial cell lysate of E.coli carrying the plasmid T 7-7 Hup53 grown in Luria broth to induce the expression of p53. The positive clones selected by ELISA were found to exhibit strong staining of nuclear p53 in both fresh and archival paraffin embedded breast tumour tissue sections. Commercial MAb D 07 against p53 was used as control. In immunoprecipitation, this MAb of IgG2b isotype was found to bind specifically to a protein of 53 kD. Immuno cyto chemical assay of normal, benign and malignant breast tissues of different histological types revealed that the majority of tumour cells were strong positive in the case of infiltrating ductal and lobular carcinomas, the staining being less intense for in situ carcinoma. The test for normal and benign tissues was negative. The staining patterns were comparable with those of control antibody. These results suggest that the MAb generated is specific to p53. The p53 protein expression was compared with the estrogen receptor (ER) status for 50 breast tumours which revealed that 38% of these were p53 positive and of these two were ER+. Among the p53 negative tumours, 48% were found to be ER+. A comparison of the p53 expression for 100 breast cancer patients indicated that 57% of the tumours were p53 negative and these patients had a longer overall 5 year survival rate and recurrent free interval which is statistically very significant. These results might suggest that p53 positive tumours are more aggressive biologically with poor prognosis.