Abstract: Background:Glomeruloid microvascular proliferation (GMP), a novel histology-based angiogenesis marker, has been associated with decreased survival in several human cancers.Methods:In this study, we evaluated the ability of GMP to predict clinical response to neoadjuvant chemotherapy in a series of locally advanced breast cancers (n=112).Results:Presence of GMP (21% of the cases) was significantly associated with high-grade tumours and TP53 mutations in addition to the basal-like and HER2 subtypes of breast cancer as defined by gene expression data. GMP was correlated to a gene expression signature for tumour hypoxia response. The GMP pattern was also significantly associated with lack of treatment response and progressive disease (P=0.004).Interpretation:The findings suggest that GMP might be able to predict the lack of response to neoadjuvant chemotherapy in locally advanced breast cancer. Whether GMP may be an independent predictor compared with other factors including TP53 mutation status and tumour grade needs confirmation in larger studies.
Abstract: TP53 mutations have been associated with resistance to anthracyclines but not to taxanes in breast cancer patients. The MDM2 promoter single nucleotide polymorphism (SNP) T309G increases MDM2 activity and may reduce wild-type p53 protein activity. Here, we explored the predictive and prognostic value of TP53 and CHEK2 mutation status together with MDM2 SNP309 genotype in stage III breast cancer patients receiving paclitaxel or epirubicin monotherapy.
Abstract: Breast cancer tissue estrogen levels on an average exceed plasma as well as benign breast tissue levels. To evaluate the contribution of intra-tumor aromatization to individual tumor estrogen levels (estradiol, E2; estrone, E1; estrone sulfate, E1S), breast cancer tissue sections obtained during mastectomy in 28 postmenopausal breast cancer patients were stained for aromatase protein expression using the aromatase antibody 677. The findings were correlated to intra-tumor estrogen levels determined with a highly sensitive HPLC-RIA. Staining with 677 alone (irrespective of the hormone receptor status) revealed no difference in tumor E2 levels comparing 677+ versus 677- tumors, although a non-significant trend towards higher tumor E1 and E1S levels was observed in 677+ breast cancers. In contrast, tumor levels of E(2) were significantly higher in ER+ tumors compared to ER- tumors (P<0.001) and to benign breast tissue from the same breast (P<0.001). Analysing the additional effect of positive staining with the aromatase antibody 677 on tumor estrogen levels in the subgroup of ER+ tumors, revealed significantly higher tumor levels of E2 (mean level of 544.7 versus 197.1 fmol/g tissue) as well as a non-significant trend concerning tumor E1 (mean level of 296.9 versus 102.1 fmol/g tissue). The mean tumor tissue E1S level was observed somewhat lower in ER+677+ (103.5 fmol/g) versus ER+677- tumors (190.1 fmol/g). In the subgroup of ER+PgR+ tumors, tissue levels of E2 were also found to be significantly higher among 677+ compared to 677- tumors: 873.2 fmol/g (95% CI 395.9-1925.6) versus 217.9 fmol/g (95% CI 88.8-534.9) (P=0.015). In conclusion, our results indicate a moderate effect of aromatase enzyme expression evaluated by IHC using the antibody 677 on intra-tumor estrogen levels among ER+ breast cancers. A substantial interindividual variation in the ratios between the individual estrogen fractions suggests additional effects, like alterations in other enzymes to be involved in the intra-tumor estrogen homeostasis.
Abstract: Breast cancer is the most frequent cancer in women and consists of a heterogeneous collection of diseases with distinct histopathological, genetic and epigenetic characteristics. In this study, we aimed to identify DNA methylation based biomarkers to distinguish patients with locally advanced breast cancer who may benefit from neoadjuvant doxorubicin treatment.
Abstract: Previous studies have suggested elevated estrogen production in tumour-bearing breast quadrants as well as in breast cancers versus benign tissue. Using highly sensitive assays, we determined breast cancer tissue estrogen concentrations together with plasma and benign tissue estrogen concentrations in each quadrant obtained from mastectomy specimens (34 postmenopausal and 13 premenopausal women). We detected similar concentrations of each of the three major estrogens estradiol (E(2)), estrone (E(1)) and E(1)S in tumour-bearing versus non-tumour-bearing quadrants. Considering malignant tumours, intratumour E(1) levels were reduced in cancer tissue obtained from pre- as well as postmenopausal women independent of tumour ER status (average ratio E(1) cancer: benign tissue of 0.2 and 0.3, respectively; p<0.001 for both groups), suggesting intratumour aromatization to be of minor importance. The most striking finding was a significant (4.1-8.6-fold) increased E(2) concentration in ER positive tumours versus normal tissue (p<0.05 and <0.001 for pre- and postmenopausal patients, respectively), contrasting low E(2) concentrations in ER- tumours (p<0.01 and <0.001 comparing E(2) levels between ER+ and ER- tumours in pre- and postmenopausals, respectively). A possible explanation to our finding is increased ligand receptor binding capacity for E(2) in receptor positive tumours but alternative factors influencing intratumour estrogen disposition cannot be excluded.
Abstract: To evaluate the influence of the third-generation aromatase inhibitor letrozole (Femara) on breast cancer tissue levels of estrone (E(1)), estradiol (E(2)), and estrone sulfate (E(1)S) in postmenopausal women undergoing primary treatment for locally advanced estrogen receptor/progesterone receptor-positive breast cancers.
Abstract: The polycomb group protein enhancer of zeste homologue 2 (EZH2) has been linked to invasive properties of aggressive breast cancer. In this report, tissue microarray analysis of 190 breast carcinomas from a nested case-control study shows that EZH2 is significantly associated with interval breast cancers. Further, a strong relationship was found with tumor cell proliferation (by Ki-67 expression), locally advanced disease, metastasis at presentation, markers of the basal epithelial phenotype (positivity for cytokeratin 5/6 or P-cadherin), and p53 status. EZH2 expression was also significantly associated with glomeruloid microvascular proliferation, an aggressive angiogenic phenotype. For prediction of aggressive disease (any event of locally advanced disease, lymph node spread, or distant spread), EZH2 was the only variable of significance in multivariate analysis, whereas no additional information was given by Ki-67. Although EZH2 expression was significant in univariate survival analysis, only tumor cell proliferation and lymph node status were significant in the final multivariate model. In conclusion, our findings indicate an important relationship not only between EZH2 and markers of tumor cell proliferation but also with aggressive disease. These findings might be practically important and relevant because the polycomb group proteins have recently been suggested as candidates for targeted therapy.
Abstract: Neoadjuvant treatment offers an opportunity to correlate molecular variables to treatment response and to explore mechanisms of drug resistance in vivo. Here, we present a statistical analysis of large-scale gene expression patterns and their relationship to response following neoadjuvant chemotherapy in locally advanced breast cancers. We analyzed cDNA expression data from 81 tumors from two patient series, one treated with doxorubicin alone (51) and the other treated with 5-fluorouracil and mitomycin (30), and both were previously studied for correlations between TP53 status and response to therapy. We observed a low frequency of progressive disease within the luminal A subtype from both series (2 of 36 versus 13 of 45 patients; P = 0.0089) and a high frequency of progressive disease among patients with luminal B type tumors treated with doxorubicin (5 of 8 patients; P = 0.0078); however, aside from these two observations, no other consistent associations between response to chemotherapy and tumor subtype were observed. These specific associations could possibly be explained by covariance with TP53 mutation status, which also correlated with tumor subtype. Using supervised analysis, we could not uncover a gene profile that could reliably (>70% accuracy and specificity) predict response to either treatment regimen.
Abstract: We previously reported that high tumour cell proliferation evaluated by Ki-67 expression, high mitotic frequency and high histological grade were associated with resistance to primary doxorubicin monotherapy in locally advanced breast cancer harbouring wild-type (wt) TP53. The aim of our present study was to evaluate the predictive and prognostic impact of proliferation parameters assessed in tumour tissue obtained after chemotherapy, and alterations induced in tumour cell proliferation. While we found a significant reduction in Ki-67 expression and mitotic frequency in tumours with wtTP53 (p=0.001 and p=0.008, respectively), no significant change was recorded in tumours expressing mutant TP53. For histological grade there was no significant change in either group. There was a direct correlation between pre- and post-treatment values for Ki-67 and mitotic frequency in tumours harbouring wtTP53 (p=0.0001 for both), but no correlation in tumours harbouring mutated TP53. High post-treatment Ki-67 expression and mitotic frequency were found to predict doxorubicin resistance only in patients with wtTP53 (p=0.04 and p=0.03, respectively). The prognostic importance of proliferation markers and histological grade was found to be similar whether they were determined in the pre- or post-treatment samples (Ki-67; pre: p=0.02; post: p=0.03; mitotic frequency; p=0.002 and p=0.01, respectively; histological grade; p=0.0001 and p=0.002, respectively). While the reduction in mitotic frequency was associated with improved survival (p=0.03), no significant associations between changes in other parameters and outcome were recorded.
Abstract: We previously reported that defects in apoptotic pathways (mutations in the TP53 gene) predicted resistance to doxorubicin monotherapy. The aim of this study was to evaluate whether cell proliferation, as assessed by mitotic frequency and Ki-67 levels, may provide additional predictive information in the same tumours and to assess any potential correlations between these markers and mutations in the TP53 gene and erbB-2 overexpression. Surgical specimens were obtained from ninety locally advanced breast cancers before commencing primary chemotherapy consisting of weekly doxorubicin (14 mg/m2) for 16 weeks. 38% of the patients had a partial response (PR) to therapy, 52% had stable disease (SD) while 10% had progressive disease (PD). Univariate analysis showed a significant association between a high cell proliferation rate (expressed as a high mitotic frequency) and resistance to doxorubicin (P = 0.001). Further analyses revealed this association to be limited to the subgroup of tumour expressing wild-type TP53 (P = 0.016), and TP53 mutation status was the only factor predicting drug resistance in the multivariate analyses. The finding that a high mitotic frequency, as well as a high Ki-67 staining, correlated to TP53 mutations (P = 0.001 for both), suggests TP53 mutations are the key predictor of drug resistance, although cell proliferation may play an additional role in tumours harbouring wild-type TP53. Regarding overall (OS) and relapse-free survival (RFS), multivariate analyses (Cox' proportional hazards regression) revealed a high histological grade and negative oestrogen receptor (ER) status to be the variables that were most strongly related to breast cancer death (P = 0.001 and P = 0.001, respectively). A key reason for this difference with respect to the factors predicting chemotherapy resistance could be due to the adjuvant use of tamoxifen in all patients harbouring ER-positive tumours.
Abstract: Recent studies have found an association between certain TP53 mutations and resistance to anthracycline-based primary medical therapy in breast cancer. The purpose of this study was to investigate whether TP53 mutational status also might influence the response to a non-anthracycline-containing regimen in primary breast cancer.
Abstract: TP53 status [mutations, immunostaining, and loss of heterozygosity (LOH)], expression of c-erbB-2, bcl-2, and histological grading were correlated to the response to doxorubicin monotherapy (14 mg/m2) administered weekly to 90 patients with locally advanced breast cancer. Mutations in the TP53 gene, in particular those affecting or disrupting the loop domains L2 or L3 of the p53 protein, were associated with lack of response to chemotherapy (P = 0.063 for all mutations and P = 0.008 for mutations affecting L2/L3, respectively). Similarly, expression of c-erbB-2 (P = 0.041), a high histological grade (P = 0.023), and lack of expression of bcl-2 (P = 0.018) all predicted chemoresistance. No statistically significant association between either p53 immunostaining or TP53 LOH and response to therapy was recorded, despite the finding that both were associated with TP53 mutation status (p53 immunostaining, P < 0.001; LOH, P = 0.021). Lack of immunostaining for p53 despite mutation of the TP53 gene was particularly seen in tumors harboring nonsense mutations or deletions/splices (7 of 10 negative for staining compared with 4 of 16 with missense mutations). TP53 mutations (total/affecting L2/L3 domains) were associated with expression of c-erbB-2 (P < 0.001 for both), high histological grade (P = 0.001 and P = 0.025), and bcl-2 negativity (P = 0.003 and P = 0.002). TP53 mutations, histological grade, and expression of bcl-2 (but not LOH or c-erbB-2 expression) all predicted for relapse-free as well as breast cancer-specific survival in univariate analysis (Ps between <0.0001 and 0.0155), but only tumor grade was found to be predictive in multivariate analysis (P = 0.01 and P = 0.0007, respectively). Our data are consistent with the hypothesis that certain TP53 mutations predict for resistance to doxorubicin in breast cancer patients. However, the observation that the majority of patients with TP53 mutations affecting or disrupting the L2/L3 domains with LOH in addition (n = 12) obtained a partial response (n = 4) or stabilization of disease (n = 5) during chemotherapy suggests redundant mechanisms to compensate for loss of p53 function. Our findings are consistent with the hypothesis that other defects may act in concert with loss of p53 function, causing resistance to doxorubicin in breast cancers.
Abstract: Fasting blood samples were obtained before definitive surgery or biopsy in 128 patients referred to the department of surgery with suspected or manifest breast cancer. Insulin-like growth factor (IGF)-I, IGF-II and free IGF-I were measured by radioimmunoassay/immunoradiometric assay, while IGFBP-3 proteolysis was evaluated by Western immunoblot. 12 patients had ductal carcinoma in situ benign conditions, while staging revealed metastatic disease in 15 of 16 patients with invasive cancers. IGFBP-3 proteolysis above the normal range was recorded in 19 patients with invasive cancers, but in none of the patients suffering from DCIS/benign conditions. Increased IGFBP-3 proteolysis was most frequently recorded in patients harbouring large tumours and metastatic disease (Stage I: 0/19, 0%; Stage II: 3/45, 7%, Stage III: 9/37, 24%, and Stage IV: 7/15, 47%). IGFBP-3 proteolysis was significantly higher in Stage III (P =0.01) and IV (P< 0.001) patients compared to the other stage groups (P = 0.001). IGF-I and IGF-II correlated negatively to IGFBP-3 proteolysis and age. Plasma levels of IGF-I and -II were significantly lower in patients with elevated IGFBP-3 proteolysis compared to those within the normal range. Our findings reveal alterations in the IGF-system among a substantial number of patients with large primary breast cancers.
Abstract: The purpose of this study was to classify breast carcinomas based on variations in gene expression patterns derived from cDNA microarrays and to correlate tumor characteristics to clinical outcome. A total of 85 cDNA microarray experiments representing 78 cancers, three fibroadenomas, and four normal breast tissues were analyzed by hierarchical clustering. As reported previously, the cancers could be classified into a basal epithelial-like group, an ERBB2-overexpressing group and a normal breast-like group based on variations in gene expression. A novel finding was that the previously characterized luminal epithelial/estrogen receptor-positive group could be divided into at least two subgroups, each with a distinctive expression profile. These subtypes proved to be reasonably robust by clustering using two different gene sets: first, a set of 456 cDNA clones previously selected to reflect intrinsic properties of the tumors and, second, a gene set that highly correlated with patient outcome. Survival analyses on a subcohort of patients with locally advanced breast cancer uniformly treated in a prospective study showed significantly different outcomes for the patients belonging to the various groups, including a poor prognosis for the basal-like subtype and a significant difference in outcome for the two estrogen receptor-positive groups.
Abstract: The p53 tumour-suppressor gene is important in the regulation of cell growth and apoptosis, and loss of functional wild-type activity may be associated with tumour formation and resistance to therapy. Differentiation of functionally normal wild-type protein from mutant or abnormal protein remains difficult using either immunohistochemical assays or mutational DNA sequencing. p21(WAF1/CIP1) (p21) is induced by wild type p53 and plays an important role in promoting cell cycle arrest. To test the hypothesis that p21 protein expression may act as a downstream marker of tumours from patients with locally advanced breast cancer before treatment with doxorubicin, pretreatment p53 status had been characterized in 63 tumours by p53 protein immunostaining and DNA mutational analysis. There was a significant association between immunostaining for p53 and the presence of p53 mutations (P = 0.01). Of 56 patients available for determination of p21, 31 (55%) expressed p21 protein. Twenty-eight out of 31 patients (90%) positive for p21 had low negative p53 protein expression, whereas only 3 of 13 patients (23%) with high p53 expressed p21 (P = 0.009). No association was seen between p21 protein expression and p53 mutations (P = 0.24). The combination of p53 and p21 immunostaining results improved the specificity of the immunostaining but at a cost of significant reduction in sensitivity. Immunohistochemical assessment of p21 protein expression is inversely associated with abnormal p53 protein in human breast cancer. The detection of p21 protein expression in combination with p53 protein expression did not improve the ability of immunohistochemistry (IHC) to differentiate between normal and mutant p53 protein.
Abstract: The mechanisms causing resistance to chemotherapeutic drugs in cancer patients are poorly understood. Recent evidence suggests that different forms of chemotherapy may exert their cytotoxic effects by inducing apoptosis. The tumor suppressor gene P53 has a pivotal role inducing apoptosis in response to cellular damage. In vitro investigations have shown intact p53 to play a critical role executing cell death in response to treatment with cytotoxic drugs like 5-fluorouracil, etoposide and doxorubicin. Recently, mutations in the P53 gene were found to confer resistance to anthracyclines in a mouse sarcoma tumor model, and overexpression of the p53 protein (which, in most cases, is due to a mutated gene) was found to be associated with lack of response to cisplatin-based chemotherapy in non-small cell lung cancer. Previous studies have shown mutations in the P53 gene or overexpression of the p53 protein to predict a poor prognosis, but also a beneficial effect of adjuvant radiotherapy or chemotherapy in breast cancer. In this study we present data linking specific mutations in the P53 gene to primary resistance to doxorubicin therapy and early relapse in breast cancer patients.
Abstract: Sixty-three patients (median age 64 years) with locally advanced breast cancer (T3, T4 and/or N2) were treated with primary 'neoadjuvant' chemotherapy given as weekly doxorubicin monotherapy (14 mg/m2 per dose). Seven patients had solitary distant metastasis at the time of diagnosis. Twenty-eight patients (45%) achieved 'partial response' to primary chemotherapy. Twenty-nine patients (46%) had 'stable disease', and 6 patients (9%) had 'progressive disease' during treatment. Following chemotherapy, 52 patients were subjected to surgery and another 4 patients had surgery performed after radiotherapy. Surgery was considered impossible in only three patients. After a median observation time of 23 months, local recurrences were observed in 2 patients, one with progressive disease and one with stable disease during chemotherapy. Univariate analyses revealed that large tumour size, high histological grade and high mitotic frequency were associated with poor primary response to chemotherapy. Recent studies have demonstrated a correlation between p53-mutations and chemotherapy response.
Abstract: The aim of this project was to study the diagnostic value of DNA content and p53 protein expression in normal, hyperplastic and neoplastic parathyroid lesions. Tissue samples of 74 parathyroid glands from 34 patients with primary hyperparathyroidism were studied by DNA flow cytometry and p53 immunostaining. In 9 of 23 patients (39%) with parathyroid adenoma, a nondiploid cell population was present. Some normal looking glands removed from the same patients also had a nondiploid DNA index. Multiglandular hyperplasia was found in 11 patients, and in 5 of these (45%) the histograms showed nondiploid cells. The proliferative activity was generally low and S-phase fraction did not differ in glands with hyperplasia or adenoma, when compared with normal looking glands. One single case of hyperplasia showed a weak p53 positivity in scattered nuclei, probably representing wild type p53 protein. Thus, our present results suggest that DNA content and p53 protein staining are of no value in the routine work up of parathyroid glands removed from patients with primary hyperparathyroidism.
Abstract: Since February 1991, in Health Region 3, patients with recently diagnosed Stage III breast cancer have been treated with neoadjuvant chemotherapy. This treatment includes 16 weeks of chemotherapy prior to local treatment, usually consisting of mastectomy and postoperative radiotherapy. 48 patients have so far undergone this treatment. The majority (43 patients) received chemotherapy consisting of weekly doses of Adriamycin. Even with a high average age the therapy has been well tolerated. 60% of the patients partially responded to the therapy, an additional 15% experienced minimal response. At this point it would be premature to evaluate the long term effect of the therapy. It is very interesting, however, that at this stage we have not observed any local recidive in patients who showed primary response to chemotherapy. Following primary treatment of this group of patients, local recidive is usually a frequent and early observation.
Abstract: Progesterone binding cyst protein (PBCP) was measured in breast cancer cytosols from 128 pre- and post-menopausal women with operable node positive (pN+) breast cancer Stage II. All patients were included in a national multicenter study on the effect of adjuvant tamoxifen treatment in hormone sensitive breast cancer, i.e. estrogen receptor content of at least 10 pmol/g cytosol protein. Patients were randomised to receive adjuvant tamoxifen 20 mg once daily for two years or no endocrine treatment. At a median follow-up of 60 months, we found PBCP content in the primary tumor to be an important factor with regard to the effect of adjuvant tamoxifen treatment. The benefit of adjuvant tamoxifen treatment on relapse-free survival and overall survival was confined to the subpopulation of patients with PBCP negative tumors. PBCP should be further evaluated as a predictive factor for the effect of tamoxifen treatment.
Abstract: Production of proteolytic enzymes is important for the invasive properties of malignant tumours. In this series of 43 cases of various non-neoplastic and neoplastic thyroid lesions, the activity of dipeptidyl peptidase I and dipeptidyl peptidase IV was increased in papillary carcinomas. In addition, cathepsin B and cathepsin L were markedly elevated in 2 of 3 follicular carcinomas, and tryptase, which is a marker of mast cells, was also significantly elevated in follicular carcinomas. Our results indicate that proteolytic proteins are important in thyroid carcinomas, and in addition there seem to be some differences between papillary and follicular tumours. Larger studies are needed to confirm these findings.
Abstract: The results following operation for morbid obesity in 41 patients (8 men and 33 women) during the period 1983-1989 are reported. The median preoperative body mass index (BMI) was 43.6 (median excess weight 81%). All patients were preoperatively evaluated at the Department of Internal Medicine, and they had all tried several conservative regimens in order to lose weight. They were all operated on with gastric banding, creating a gastric stoma of 15 mm. Pneumonia developed in 10 patients, one patient got a wound infection, and one patient died postoperatively from a massive coronary embolism. The immediate postoperative weight loss was satisfactory, median BMI being reduced from 43.6 preoperative to 30.5 after 9 months. Median BMI after 3 and 5 years was 34 and 32 (median excess weight 42 and 41%, respectively). Six patients were reoperated, four having their band removed, two being converted to vertical gastric banding. In conclusion, gastric banding gives satisfactory results for most of the patients. The reoperation rate is, however, high and the long-term result is not favorable for all patients
Abstract: From 1983-88, 97 breast cancers in 94 women were treated with breast-conserving surgery at Haukeland University Hospital. 71 per cent of the tumours were less than 2 cm in diameter and 94 per cent less than 3 cm. 65 had negative axillary nodal status. 90 patients had ductal carcinoma, three of whom also had extensive intraductal carcinoma. Pathological findings in the resection margins (invasive carcinoma, intraductal carcinoma or atypical epithelial hyperplasia) were reasons for reoperation in 14 patients, eight of them by mastectomy. Thus, after completion of initial treatment the breast was preserved in 89 patients. Postoperative irradiation to the breast was given as a routine. Three elderly patients were excepted. Two patients developed ipsilateral breast recurrences after six and 24 months respectively. The first was treated by re-resection and the second by mastectomy. Neither of these showed evidence of distant metastases. Distant metastases were discovered in five patients, two of these have since died.
Abstract: From 1983-88, 97 breast cancers in 94 women were treated with breast conserving surgery at Haukeland University Hospital. 71% of the tumours were less than 2 cm in diameter and 94% less than 3 cm. 65% had negative axillary nodal status. 90 patients had ductal carcinoma, three of whom also had extensive intraductal carcinoma. Pathological findings in the resection margins (invasive carcinoma, intraductal carcinoma or atypical epithelial hyperplasia) were reasons for reoperation in 14 patients, eight of them by mastectomy. Thus, after completion of initial treatment the breast was preserved in 89 patients. Postoperative irradiation to the breast was given as a routine. Three elderly patients were excepted. Two patients developed ipsilateral breast recurrences after six and 24 months respectively. The first was treated by re-resection and the second by mastectomy. Neither of these showed evidence of distant metastases. Distant metastases were discovered in five patients, two of these have since died.
Abstract: From 1981 to 1988 six patients, five females and one male, with breast sarcoma were treated in the Department of Surgery, University Hospital, Bergen. Four of them had phyllodes tumour, one a malignant myoepithelioma and one a liposarcoma. Metastases to axillary lymph nodes or distant metastases were not observed. Sarcomas account for about 1% of malignant breast tumours in our department. The cases demonstrate diagnostic and therapeutical problems in dealing with breast sarcomas.
