Expression of c-myc protein is related to cell proliferation and expression of growth factor receptors in transitional cell bladder cancer

Archival biopsy specimens from transitional cell bladder tumours (n=185) were analysed immunohistochemically for expression of c-myc protein. The results were compared with compared with histopathological and clinical parameters and survival. Forty-three per cent of the tumours were negative for c-myc protein and weak, moderate, or strong cytoplasmic expression was found in 34, 14, and 9 per cent of cases, respectively. Nuclear positivity for c-myc protein was detected in 35 per cent of tumours and nuclear opositivity was related to overexpression of c-erb B-2 (P=0.01) and a high proportion of nuclei were also positive for p53 oncoprotein (p<0.05). Cytoplasmic expression of c-myc protein was related to histological grade (P=0.005), papillary status (P=0.007), the S-phase fraction (P=0.008), the mitotic index (P=0.021), overexpression of epidermal growth factor receptor (P=0.045), and c-erb B-2 (P=0.17). Expression of c-myc protein was not significantly related to the progression of tumours and it had no prognostic value in survival analysis. Independent predictors were the T-category (P<0.001), papillary status. (P=0.001), and S-phase fraction (P=0.061). The results show that while c-myc gene product participates in growth regulation of human bladder cancer cells, it has no independent prognostic significance.     

Prognostication of astrocytoma patient survival by Ki-67 (MIB-1), PCNA,and S-phase fraction using archival paraffin-embedded samples

The prognostic power of three proliferation estimation methods, Ki-67 (MIB-1) and PCNA immunohistochemistry, and flow cytometry (S-phase and S+G2/M fractions, respectively), were evaluated in 50 cases of astrocytoma. Each proliferation index showed a strong association with the grade of malignancy (grades I-IV). The MIB-1 labelling index (LI) provided additional information, as it could be used for the discrimination of grade II and grade III astrocytomas (P=0·0357). All three proliferation estimation methods also had strong prognostic potential (MIB-1 LI: P<0·0001; PCNA Li: P<0·0001; S-phase: P=0·0004; S+G2/M: P=0·0124). According to the receiver operating characteristics (ROC) curve, the MIB-1 LI showed generally the best sensitivity and specificity in placing the patients correctly into groups of survivors and non-survivors, which was further confirmed in the multivariate analysis. Only 4 per cent of the patients having high MIB-1 scores (>15·3 per cent) were alive after 2-years' follow-up. In contrast, 72 per cent of patients with tumours of low proliferation activity survived. It appears that Ki-67 (MIB-1) immunolabelling using archival paraffin-embedded samples is of value in predicting prognosis in astrocytic tumours.     

Expression of p53 protein has no independent prognostic value in breast cancer

A series of 392 female breast carcinomas was analysed immunohistochemically for expression of p53 protein with special emphasis on the role of p53 as an independent prognostic factor. Altogether, 54·8 per cent of the carcinomas expressed p53 protein, with the mean [standard error (SE)] fraction of positive nuclei being 17·1 per cent (1·2 per cent). Expression of p53 protein was independent of tumour metastasis at diagnosis, axillary lymph node status, tumour diameter, histological type, tubule formation, proportion of intraductal growth, margin formation, necrosis, DNA ploidy, and S-phase fraction. A high fraction of p53-positive nuclei was significantly related to patient age under 70 years, high grade, severe nuclear pleomorphism, dense infiltration of tumour by lymphocytes, high mitotic index, and high apoptotic index (for all, P<0·05). Impaired survival probability in the entire cohort (P=0·05) and in the axillary lymph node-positive (ANP) tumours (P=0·015) was associated with a fraction of p53-positive nuclei less than 25 per cent, while in the axillary lymph node-negative (ANN) tumours, expression of p53 had no prognostic value. In multivariate analysis, independent prognostic predictors included axillary lymph node status, tumour diameter, and mitotic index. In the ANN tumours, tumour diameter, fraction of p53-positive nuclei, and tumour grade were independent prognostic factors, whereas in the ANP tumours, diameter and mitotic index were the two independent prognostic factors. The results suggest that abnormal expression of p53 protein is only a weak independent prognostic factor in female breast cancer.     

p53 OVEREXPRESSION AND HUMAN PAPILLOMAVIRUS INFECTION IN TRANSITIONAL CELL CARCINOMA OF THE URINARY BLADDER: CORRELATION WITH HISTOLOGICAL PARAMETERS

Seventy-nine transitional cell carcinomas (TCCs) of the urinary bladder (25 grade 1, 22 grade 2, and 32 grade 3 tumours) were examined for p53 overexpression by immunohistochemistry with a monoclonal antibody and for human papillomavirus (HPV) infection by the polymerase chain reaction (PCR). Positive immunostaining for p53 was detected in 40·5 per cent of the cases; the percentage of positive cases was significantly lower in low-grade (G1 and G2) TCCs than in high-grade (G3) tumours (10·6 per cent vs. 84·4 per cent; P <0·0001). The overall rate of HPV infection was 32·9 per cent; 20·3 per cent of the cases were positive for HPV 16, 3·8 per cent for HPV 18, and 8·9 per cent for both. Consensus primers as well as type-specific primers for HPV types 6, 11, and 33 failed to detect any additional case with HPV infection. The prevalence of HPV 16 and/or HPV 18 infection was significantly higher in low-grade than in high-grade tumours (44·7 per cent vs. 15·6 per cent; P =0·0061). p53-positive cases were more common among papillary, deeply infiltrating tumours, and HPV-positive cases among papillary, non-infiltrating lesions. According to these data, p53 overexpression and HPV 16/18 infection are common findings in bladder TCC and there appears to be an inverse correlation of p53 overexpression and of HPV infection with tumour aggressiveness. The possibility of different molecular pathways in superficial low-grade and in invasive high-grade tumours is suggested.     

Apoptosis suppressing protein bcl-2 is expressed in well-differentiated breast carcinomas with favourable prognosis

The expression of bcl-2 protein was analysed immunohistochemically in 202 female breast carcinomas. The intensity of bcl-2 expression was inversely related to tumour grade (P<0·0001), tumor necrosis (P<0·0001), mitotic index (P<0·0001), oestrogen receptor content (P<0·0001), progesterone receptor content (P=0·0007), S-phase fraction (P=0·00047), and apoptotic index (P=0·087). A high fraction of bcl-2-positive cells was related to ductal or lobular type (P=0·03) and slight nuclear pleomorphism (P=0·03). Heterogeneous expression of bcl-2 protein was associated with high grade (P=0·02), severe nuclear pleomorphism (P=0·02), DNA aneuploidy (P=0·018), high S-phase fraction (P=0·05), and early metastasis (P=0·03). Intense expression of bcl-2 protein was significantly related to favourable outcome in the entire cohort (P=0·0013), as well as in axillary lymph node-negative (ANN) tumours (P=0·0124). Long recurrence-free periods in the entire cohort (P=0·037) and in ANN tumours (P=0·08) were confined to cases with intense expression of bcl-2 protein. In multivariate analysis, bcl-2 expression had no independent prognostic value in the entire cohort or in axillary lymph node-negative breast carcinomas, whereas it was a weak independent prognostic factor in axillary lymph node-positive breast carcinomas.     

Expression of pS2 protein in transitional cell bladder tumours

A series of 201 bladder cancer biopsy specimens was analysed immunohistochemically for the expression of pS2 protein. Altogether, 61 per cent of the tumours were pS2-negative; in 16 per cent less than 1 per cent and in 23 per cent of cases more than 1 per cent of cells were pS2-positive. Normal transitional epithelium was negative for pS2. The fraction of positive cells was higher in poorly differentiated non-papillary tumours and in invasive tumours with pelvic lymph-node (P=0.05) and distant metastasis (P=0.10). pS2 expression was not related to sex, while patients aged 60–70 years had low fractions of pS2-positive cells (P=0.03). DNA ploidy, S-phase fraction, mitotic index, morphometric nuclear features, and expression of c-erbB-2, p53, and epidermal growth factor receptor were independent of expression of pS2. Tumours expressing pS2 in over 10 per cent of cells had a lower survival probability (P=0.0486). The results show that pS2 is expressed in 40 per cent of transitional cell bladder tumours, but that this marker has no clinical significance over established prognostic factors.     

p53 Protein,PCNA staining,and DNA content in follicular neoplasms of the thyroid gland

Forty-nine follicular adenomas and 11 follicular carcinomas of the thyroid were investigated by immuno-histochemistry for the expression of p53 protein and proliferating cell nuclear antigen (PCNA). The DNA ploidy and the S-phase fraction (SPF) of the neoplasms were analysed by flow cytometry. Twelve adenomas (24 per cent) and six carcinomas (55 per cent) were DNA non-diploid (P=0·07). The carcinomas had a higher proliferation rate than the adenomas when assessed either by SPF size (median 9·9 per cent vs. 2·9 per cent, P=0·0003) or by PCNA staining intensity (P<0·0001). Some scattered nuclei in two (4 per cent) adenomas and in three (27 per cent) carcinomas stained positively for p53 (P=0·04). The two adenomas with positive staining for p53 were subserially sectioned, but no signs of invasion were found; both patients are alive and well 6 and 7 years after surgery. One of the two adenomas showing positive p53 nuclear staining was DNA aneuploid, and both were positive in PCNA staining, but their SPFs were low (2·1 and 3·3 per cent). We conclude that p53 protein expression is not confined to follicular carcinomas; scattered p53-positive cells may also be present in histologically and clinically benign follicular adenomas. Because both follicular adenomas and carcinomas may be DNA aneuploid and their SPF and PCNA staining distributions overlap, the distinction between follicular adenoma and carcinoma should still be based on histological criteria.     

High-affinity monomeric 67-kd laminin receptors and prognosis in pancreatic endocrine tumours

Cell-surface high-affinity monomeric 67-kD laminin receptors have been proposed to promote the invasion and metastasis of a variety of tumours, but there are, as yet, no data regarding the expression of these molecules in pancreatic endocrine tumours (PETs). The prognosis of these very rare tumours is problematic and the only irrefutable evidence of their malignancy still continues to be the occurrence of local invasion and metastases. In this retrospective investigation, 34 functioning and 48 non-functioning sporadic PETs were evaluated for the expression of the MLuC5 monoclonal antibody, which specifically recognizes the 67-kD laminin receptors. Laminin receptors were found in 42/82 cases (51 per cent) and their expression was associated with metastatic disease (P<0·001), high proliferative activity expressed by a Ki-67 index above 5·0 per cent (P<0·001), absence of progesterone receptors (P=0·013), immunoreactivity for hormones other than insulin (P<0·001), a tumour diameter more than 3·0 cm (P=0·001), and a fatal clinical outcome (P<0·001). Laminin receptors were also expressed by most metastatic foci and all intravascular emboli of tumour cells. Positivity for laminin receptors was associated with shorter survival in functioning (P=0·026) and non-functioning (P=0·042) tumours, as well as in the whole series of pancreatic endocrine tumours (P<0·001). On multivariate analysis, laminin receptor expression was not an independent prognostic factor, while a Ki-67 index above 5·0 per cent was the most powerful predictor of survival. However, the association of laminin receptor expression and Ki-67 index could identify a group of malignant PETs with low proliferative activity characterized by an intermediate prognosis. In conclusion, these data suggest that monomeric laminin receptors may play a role in the invasion and metastasis of PETs and that their expression may be an additional prognostic factor, along with proliferative activity. © 1997 by John Wiley & Sons, Ltd.     

NUCLEAR p53 OVEREXPRESSION IS AN INDEPENDENT PROGNOSTIC PARAMETER IN NODE-NEGATIVE NON-SMALL CELL LUNG CARCINOMA

The prognosis of operated patients with non-small cell lung cancer (NSCLC) is poor despite thorough pre-operative staging. An improved preselection is needed of patients likely to profit from surgery. This study was undertaken to evaluate the prognostic significance of nuclear p53 overexpression in a cohort of 247 surgically treated patients with NSCLC. It showed that the prevalence of immunohistochemically detectable p53 overexpression varied between different tumour types. p53 overexpression was equally frequent in large cell carcinoma (53 per cent) and in squamous cell carcinoma (54 per cent), but significantly less frequent in adenocarcinoma (34 per cent; P =0·009). p53 overexpression was particularly rare in bronchioloalveolar carcinoma (positivity in 1 of 17 cases). These variations may reflect aetiological differences between the histological subtypes. p53 overexpression was also associated with high tumour grade ( P =0·0157) and the presence of lymph node metastasis ( P =0·0259), but not with advanced tumour stage. Survival analysis showed no difference in clinical outcome between p53-positive and p53-negative tumours within 101 node-positive tumours. In contrast, survival time was significantly better in p53-negative tumours than in p53-positive tumours within the group of 113 node-negative tumours ( P =0·032). Stepwise regression analysis showed that p53 overexpression is an independent prognostic factor in node-negative NSCLC.     

IMMUNOHISTOCHEMICAL ASSAY FOR OESTROGEN RECEPTORS IN PARAFFIN WAX SECTIONS OF BREAST CARCINOMA USING A NEW MONOCLONAL ANTIBODY

The aim of this study was to evaluate the utility of a new monoclonal antibody (AER311) that targets the oestrogen receptor (ER) in an immunohistochemical assay (IHA) applied to breast cancers. Ninety-seven cases of invasive ductal carcinoma were studied by AER311-IHA using a pressure-cooking antigen retrieval technique applied to formaldehyde-fixed, paraffin-embedded tissue sections; immunostaining was assessed by semi-quantitative scoring ( H score). There was 80 per cent concordance between the ER status measured by dextran-coated charcoal (DCC) assay and AER311-IHA, with 63/97 (65 per cent) tumours positive and 15/97 (15 per cent) tumours negative by both assays. Of the 12 DCC-positive cases that were negative by AER311-IHA, 11 were borderline positive (3–8 fmol/mg). Similarly, six of seven DCC-negative cases that scored positive by AER311-IHA had only borderline positive H scores (<50). When AER311-IHA was compared with 1D5-IHA, there was good concordance in ER status (77 per cent) and a significant correlation ( r =0·7, P <0·001) between H scores. Nevertheless, the correlation between ER level determined by AER311-IHA and that measured by DCC ( r =0·53, P <0·001) was higher than that for 1D5-IHA ( r =0·32, P =0·002). AER311-IHA can therefore provide reliable information about the ER status of breast carcinoma on paraffin sections and is an acceptable alternative to other commercially available monoclonal antibodies.     

Increased cell proliferation activity and decreased cell death are associated with the emergence of hormone-refractory recurrent prostate cancer

The tumour growth kinetics (cell proliferation and apoptosis) of ten hormone-refractory locally recurrent prostate cancers were compared with their matched untreated primary tumour specimens. All recurrent tumours had a higher cell proliferation activity, as defined by Ki-67 immunohistochemistry, than corresponding primary tumours from the same patients. The mean cell proliferation activity in recurrences (13·5±3·8 per cent) was over two times higher (P<0·0001) than that in primary tumours (5·5±2·4 per cent), suggesting that cell clones which progress during androgen withdrawal are actively stimulated to proliferate. The mean percentage of apoptotic cells, as estimated by the in situ end-labelling technique, was 5·4±4·7 per cent in untreated primary tumours, whereas it was 2·3±1·5 per cent in locally recurrent tumours (P=0·05). In all but two cases, the apoptotic index was lower in recurrent than in corresponding primary tumours, suggesting that recurrent prostate carcinomas are able to avoid apoptosis in the androgen-deprived environment. In conclusion, the clinical progression of prostate cancer during androgen withdrawal is associated with increased cell proliferation and decreased apoptosis. © 1997 by John Wiley & Sons, Ltd.     

MICROSATELLITE INSTABILITY IN INTESTINAL- AND DIFFUSE-TYPE GASTRIC CARCINOMA

To investigate the role of genetic instability in the development of intestinal- and diffuse-type gastric cancers, six microsatellite loci were analysed in 98 carcinomas of the two main histotypes, at both early and advanced stages of progression, and in five preneoplastic lesions. RER+ phenotype frequency proved to be significantly higher ( P =0·013) in intestinal (23 per cent) than in diffuse cancers (5 per cent) and slightly higher in advanced (19 per cent) than in early (12 per cent) tumours. When comparing early and advanced tumours of the same histotype, a similar frequency was found for diffuse tumours (4 per cent vs. 6 per cent), and an increase from 19 to 30 per cent for intestinal cancers. Instability at more than one locus was limited to intestinal tumours and replication errors were also detected in an intestinal dysplasia. On the whole, these data suggest that genetic instability has an important and early role in gastric carcinogenesis of the intestinal type and a less important role in gastric carcinogenesis of the diffuse type. Most tumours of this panel had previously been characterized for p53 gene mutations. p53 screening was extended to all samples, to investigate the possible association between gene mutations and microsatellite instability. Analysis showed a trend ( P =0·07, Fisher's exact test) towards a negative association between these two genetic lesions in tumours of the intestinal type. © 1997 John Wiley & Sons, Ltd.     

Expression of E-cadherin (E-CD) as related to other prognostic factors and survival in breast cancer

A series of 208 breast cancer biopsies were analysed immunohistochemically for expression of E-cadherin (E-CD). Altogether, 72 per cent of the tumours showed E-CD positivity in over 50 per cent of cells, the staining being heterogeneous in nearly all tumours. In only 16 per cent of ductal carcinomas was positive staining seen in less than 1 per cent of cells. Expression of E-CD was not related to tumour diameter, nodal status, metastasis at diagnosis, histological grade, DNA ploidy, S-phase fraction, nuclear area, mitotic frequency, or PR content. There was a significant relationship between expression of E-CD, histological type (P=0.01), the proportion of intraductal growth (P=0.008), the density of tumour-infiltrating lymphocytes (P=0.0007), ER content (P=0.012), and morphometric nuclear factors (P=0.02). Expression of E-CD showed a weak association with a high survival probability (P=0.02), while the relation to recurrence-free survival was not significant (P=0.11). In axillary lymph node-negative tumours, E-CD expression was not related significantly to survival (P=0.11) or to recurrence-free survival (P=0.06). In multivariate analysis, E-CD expression had no independent prognostic value, while the axillary lymph node status, tumour diameter, patient age, and mitotic frequency were independent prognostic factors. The results indicate that E-CD expression is related to several histological features in breast cancer, but has no independent prognostic value over standard prognostic factors.     

CYCLIN D1 AND RETINOBLASTOMA GENE EXPRESSION IN HUMAN BREAST CARCINOMA: CORRELATION WITH TUMOUR PROLIFERATION AND OESTROGEN RECEPTOR STATUS

Cyclin D1 (CCND1) and retinoblastoma (Rb) genes are cell cycle regulators which are altered in some breast carcinomas. However, the possible cooperation between CCND1 and Rb, as well as the influence and coincidence of their abnormalities in the proliferative capacity of mammary carcinoma cells in vivo, is still unknown. In order to assess both the significance of the CCND1 gene and Rb alterations in breast carcinomas and their relationship with the proliferative capacity of the tumours and other clinico-pathological factors, CCND1 mRNA expression was studied in 46 cases of primary breast carcinomas and matched normal tissue, 45 of which were also studied immunohistochemically. Rb expression was analysed in the same cases by immunohistochemistry, whereas the proliferative activity of the carcinomas was evaluated by flow cytometry. CCND1 mRNA was overexpressed in 19 tumours (41 per cent). Sixteen cases showed diffuse immunohistochemical expression, ten carcinomas had few positive cells, and 19 were absolutely negative. CCND1 mRNA and protein overexpression was associated with oestrogen receptor (ER) expression by the tumour. Interestingly, lack of ER expression was associated with a decreased CCND1 mRNA signal in non-overexpressed tumours. No association was observed between CCND1 mRNA or protein overexpression and tumour proliferation or other clinico-pathological parameters. Loss of Rb expression was observed in 26 per cent of the tumours. This abnormality was significantly associated with increased mean S-phase (P=0·017) and decreased CCND1 mRNA expression in non-overexpressed tumours, supporting in vivo the postulated regulatory loop between Rb and CCND1 in vitro. We conclude that CCND1 up-regulation is not associated with increased proliferative activity in breast carcinomas, whereas its expression might be regulated in vivo by hormones and Rb. Loss of Rb expression is significantly associated with an increased proliferation of tumour cells, suggesting an important role in the progression of a subset of breast carcinomas, regardless of CCND1 abnormalities. © 1997 John Wiley & Sons, Ltd.     

Alterations of expression of Rb,p16INK4A and cyclin D1 in non-small cell lung carcinoma and their clinical significance

Inactivation of the Rb pathway in non-small cell lung carcinoma (NSCLC) occurs mostly through inactivation of the cyclin-dependent kinase inhibitor p16^INK4A and/or up-regulation of cyclin D1. In order to assess the frequency and the prognostic value of these abnormalities in NSCLC, immunohistochemical analysis of Rb, p16^INK4, and cyclin D1 has been performed on 168 cases of NSCLC including 77 squamous cell carcinomas, 43 adenocarcinomas, and 48 basaloid carcinomas. The reduced survival rate of basaloid carcinoma (stage I–II) compared with other histological types of NSCLC was confirmed (p = 0·008). Loss of protein expression of Rb and p16^INK4A was observed in 12 per cent and 58 per cent of NSCLC cases respectively and cyclin D1 overexpression in 43 per cent. There was an inverse correlation between Rb and p16 expression ( p < 0·0001) and a direct correlation between Rb and cyclin D1 expression ( p = 0·0007). In univariate analysis, Rb-negative adenocarcinomas at stages I–II had a significantly shorter survival than Rb-positive cases ( p = 0·04) and stages I–II p16-positive cases had a shorter survival than p16-negative cases ( p = 0·02), which was more significant in basaloid carcinoma ( p = 0·003). p16 status retained its influence on survival in multivariate analysis at stage I–II for all cases ( p = 0·01) and for basaloid carcinoma ( p = 0·005). Cyclin D1 overexpression did not influence survival. Combined Rb/p16/cyclin D1 phenotypes in univariate analysis showed a shorter survival for Rb-negative/p16-positive/cyclin D1-negative tumours ( p = 0·002). These results, linked to previous data, indicate that the Rb pathway of G1 arrest is initially disrupted in the vast majority of NSCLCs (83 per cent), but could not confirm an unfavourable role for each individual event (p16^INK4A loss or cyclin D1 up-regulation) in prognosis. Copyright © 1999 John Wiley & Sons, Ltd.     

Loss of membranous E-cadherin expression in pancreatic cancer: Correlation with lymph node metastasis,high grade,and advanced stage

Epithelial cadherin (E-cadherin) is a Ca²⁺-dependent cell-cell adhesion molecule that connects cells via homotypic interactions. Its function is critical in the induction and maintenance of cell polarity and differentiation, and its loss of downregulation is associated with an invasive and poorly differentiated phenotype in colon and other tumours. We have used an avidin-biotin immunoperoxidase technique to localize E-cadherin in microwave-treated, paraffin-embedded sections from 36 patients with pancreatic adenocarcinomas. E-cadherin was expressed by normal ductal and acinar cells with typical membranous staining at the intercellular junctions. Loss of normal surface E-cadherin expression was found in 19/36 (53 per cent) tumours compared to the adjacent normal ductal cells. Abnormal E-cadherin expression was found more frequently in poorly differentiated (grade III) (6/7, 86 per cent) than in well-differentiated tumours (grade I) (4/14, 28 per cent) (P=0·012). Membranous E-cadherin expression was also lost more frequently in primary tumours with lymph node (stage III) (14/23, 61 per cent) and distant metastasis (stage IV) (2/2, 100 per cent) compared with 3/11 (27 per cent) lymph node-negative tumours (stage I) (P=0·043). In conclusions, our data indicate that loss of membranous E-cadherin expression is associated with high grade and advanced stage in pancreatic cancer.     

The prognostic significance of immunohistochemically detected lymph node micrometastases in colorectal carcinoma

Micrometastases have been detected by immunocytochemical means in the lymph nodes of patients with otherwise node-negative cancer of the colon and rectum. This study examines the incidence and prognostic significance of nodal micrometastases in Dukes' B carcinoma. Five hundred and fifty-nine lymph nodes from 77 cases of Dukes' B carcinoma were examined for lymph node micrometastases by immunocytochemical staining for cytokeratin AE1:AE3. Micrometastases were detected in 19 cases (25 per cent). Cell clusters were present in ten cases, the remaining nine cases displaying only single cells. The presence of micrometastases was unrelated to age (P = 0·06), sex (P = 0·32), tumour site (P = 0·37), tumour size (P = 0·67), or tumour differentiation (P = 0·66). Ten-year survival estimates by the Kaplan–Meier lifetable method was 47 per cent in patients with and without micrometastases (χ² = 0·35 and 1 df, P = ns). The presence of nodal micrometastases detectable only by immunocytochemistry in patients with Dukes' B colorectal cancer does not justify reassignment to a more advanced disease stage.     

Expression of CD44 standard and variant-v6 proteins in transitional cell bladder tumours and their relation to prognosis during a long-term follow-up

The expression of the standard CD44 (CD44s) and its v6 isoform (CD44v6) was analysed immunohistochemically in 173 cases of transitional cell bladder cancer. The results of immunohistochemical analyses were related to established prognostic factors and clinical follow-up data. The expression intensity of CD44s in non-basal tumour cells was significantly related to TN classification, S-phase fraction (SPF), mitotic index, grade, density of tumour infiltrating lymphocytes. The expression intensity of CD44v6 in non-basal tumour cells was inversely related to DNA ploidy, SPF, and mitotic index. The expression intensity of CD44v6 in basal tumour cells was also inversely related to T-category, grade, papillary status, DNA ploidy, SPF, and mitotic index. Strong expression of CD44s in non-basal tumour cells was related to unfavourable outcome in univariate analysis (p=0·008), whereas the strong expression of CD44v6 in both non-basal cells (p=0·005) and basal cells (p=0·0008) was related to high survival probability. In multivariate survival analysis, the expression intensity of CD44v6 was independently related to favourable outcome in muscle invasive tumours, while in superficial tumours, CD44s was an independent prognostic factor. The results suggest that the expression of CD44s and CD44v6 is associated with malignant features and prognosis in bladder cancer. Copyright © 1998 John Wiley & Sons, Ltd.     

p21WAF1/Cip1 is associated with cyclin D1CCND1 expression and tubular differentiation but is independent of p53 overexpression in human breast carcinoma

p21^WAF1/Cip1 is an inhibitor of cdk/cyclin complexes, and thus regulates the cell cycle. p21 is also related to cell differentiation and is regulated by wild-type p53, although p53-independent regulatory pathways have been proposed. In order to analyse p21 expression as well as its relationship with p53 in human breast cancer, an immunohistochemical analysis was undertaken of 77 breast carcinomas, 16 of them with an in situ component; 30 adjacent normal tissue samples; and five non-neoplastic specimens. Forty-four infiltrating carcinomas (57 per cent) were p21-positive. Expression of p21 was also observed in pre-invasive lesions, whereas normal ducts were negative or focally and weakly positive. p21 expression was associated with high histological grade (II+III) (P-0·017) and poor tubule formation (P-0·002), and was significantly less frequent in lobular carcinomas (P-0·0001). p21 positivity also correlated with increased proliferation, but this seemed to be dependent on the histological grade. Twenty carcinomas (26 per cent) showed p53 overexpression, but this was not associated with p21 negativity, suggesting the existence of p53-independent mechanisms for p21 regulation in vivo. Cyclin D1^CCND1 expression was analysed in the same series and an association between p21 and cyclin D1 expression was found, since 23 of 26 cyclin D1-positive carcinomas were p21-positive (P<0·001 …). In conclusion, p21 is frequently overexpressed in breast carcinomas and this occurs in the early stages of neoplastic progression. This overexpression seems to be independent of p53 status and might be involved in cyclin D1 modulation. © 1998 John Wiley & Sons, Ltd.     

APC in the regulation of intestinal crypt fission

The functional effects of APC (adenomatous polyposis coli gene) germ-line mutations on crypt fission and cell proliferation were investigated in the normal intestine of human familial adenomatous polyposis (FAP) and multiple intestinal neoplasia (MIN) mice. Compared with controls, there was a 19-fold increase in the proportion of crypts in fission in FAP colon [95 per cent confidence interval (CI):11–32, P<0·0001], and a 75 and 61 per cent increase in MIN colon (95 per cent CI:1·08–2·82, P<0·02) and small bowel, respectively (95 per cent CI:1·31–1·99, P<0·001). In marked contrast, no significant differences in intra-cryptal epithelial cell proliferation or mitotic distribution were seen. Furthermore, 10·9 per cent of crypts in FAP were in asymmetrical fission as opposed to only 1 per cent in controls (P=0·001). The largest relative increases in MIN crypt fission were in the colon (proximal and distal colon:190 per cent, P=0·02 and 83 per cent, P=0·01), suggesting that Apc mutations exert their maximal influence site-specifically. However, sites with the highest relative increases were also those with the largest eventual tumour sizes, but not the highest polyp counts. Three-dimensional serial section reconstruction analysis corroborated that FAP adenomas enlarge by crypt fission, which was frequently both asymmetrical and atypical. It is proposed that the absence of an increase in intestinal cell division infers that APC regulates intestinal crypt differentiation, specifically through the crypt cycle. This role appears analogous to the control of axis re-duplication in embryonic development, when downstream targets of APC are over-expressed. It is concluded that in vivo, the major defect in pre-neoplastic intestine harbouring APC mutations is elevated rates of crypt fission, and that this is also the mode by which micro-adenomas enlarge. © 1998 John Wiley & Sons, Ltd.