Validation of 70-gene prognosis signature in node-negative breast cancer.

PURPOSE: The 70-gene prognosis signature (van't Veer et al., Nature 415(6871):530-536, 2002) may improve the selection of lymph node-negative breast cancer patients for adjuvant systemic therapy. Optimal validation of prognostic classifiers is of great importance and we therefore wished to evaluate the prognostic value of the 70-gene prognosis signature in a series of relatively recently diagnosed lymph node negative breast cancer patients. METHODS: We evaluated the 70-gene prognosis signature in an independent representative series of patients with invasive breast cancer (N = 123; <55 years; pT1-2N0; diagnosed between 1996 and 1999; median follow-up 5.8 years) by classifying these patients as having a good or poor prognosis signature. In addition, we updated the follow-up of the node-negative patients of the previously published validation-series (Van de Vijver et al., N Engl J Med 347(25):1999-2009, 2002; N = 151; median follow-up 10.2 years). The prognostic value of the 70-gene prognosis signature was compared with that of four commonly used clinicopathological risk indexes. The endpoints were distant metastasis (as first event) free percentage (DMFP) and overall survival (OS). RESULTS: The 5-year OS was 82 +/- 5% in poor (48%) and 97 +/- 2% in good prognosis signature (52%) patients (HR 3.4; 95% CI 1.2-9.6; P = 0.021). The 5-years DMFP was 78 +/- 6% in poor and 98 +/- 2% in good prognosis signature patients (HR 5.7; 95% CI 1.6-20; P = 0.007). In the updated series (N = 151; 60% poor vs. 40% good), the 10-year OS was 51 +/- 5% and 94 +/- 3% (HR 10.7; 95% CI 3.9-30; P < 0.01), respectively. The DMFP was 50 +/- 6% in poor and 86 +/- 5% in good prognosis signature patients (HR 5.5; 95% CI 2.5-12; P < 0.01). In multivariate analysis, the prognosis signature was a strong independent prognostic factor in both series, outperforming the clinicopathological risk indexes. CONCLUSION: The 70-gene prognosis signature is also an independent prognostic factor in node-negative breast cancer patients for women diagnosed in recent years.

[A young woman with abnormal vaginal tissue loss]. / Een jonge vrouw met abnormaal vaginaal weefselverlies.

A 18-year-old woman experienced abnormal and painful vaginal tissue loss while using an oral contraceptive. A pregnancy test was negative. Histopathological examination of the tissue showed a decidualised endometrium. We made the diagnosis of progesterone-induced membranous dysmenorrhoea.

Analysis of local recurrence after breast conservative treatment for invasive breast cancer: a single institution cohort.

In a group of 900 patients treated for carcinoma of the breast, we evaluated patient-, tumour- and treatment-related parameters, predicting the success or failure of conservative treatment for invasive breast cancer. Thirty-one patients developed local recurrences which were detected within 0.1-12.1 years after treatment of the primary tumour (with a median of 6.2 years), providing a risk of 2% at 5 years and 9% at 10 years. The locally recurrent tumours and their original primary tumours of 28 patients could be retrieved from the pathology laboratory archives. These 28 tumours of the recurrence group (RG) were matched with tumours without local recurrence, the non-recurrence group, for age at time of diagnosis, duration of follow-up and T- and N-stage. The tumours were studied for type and grade of invasive tumour including the in situ component and involvement of surgical margins. In addition, the expression of cell-cycle proteins, p53, Ki-67 (MIB-1) and BCL-1, as well as HER-2/ neu oncoprotein, estrogen and progesterone receptor were investigated. We found a mean age at diagnosis for the RG of 50 years, and the mean age at time of diagnosis for the whole group of 900 patients was 56 years (p=0.003). Thirty-nine percent of the RG had a positive surgical margin, which was the case for only 18% in the control group (p=0.09). The presence of the in situ component was also correlated with increased local recurrence (p=0.022). Furthermore, local recurrence was also associated with a significantly increased occurrence of distant metastases (p=0.001). We conclude that breast conservative treatment is safe with a low local recurrence rate.

Atypical HNPCC owing to MSH6 germline mutations: analysis of a large Dutch pedigree.

Hereditary non-polyposis colorectal cancer (HNPCC) is the most common genetic susceptibility syndrome for colorectal cancer. HNPCC is most frequently caused by germline mutations in the DNA mismatch repair (MMR) genes MSH2 and MLH1. Recently, mutations in another MMR gene, MSH6 (also known as GTBP), have also been shown to result in HNPCC. Preliminary data indicate that the phenotype related to MSH6 mutations may differ from the classical HNPCC caused by defects in MSH2 and MLH1. Here, we describe an extended Dutch HNPCC family not fulfilling the Amsterdam criteria II and resulting from a MSH6 mutation. Overall, the penetrance of colorectal cancer appears to be significantly decreased (p<0.001) among the MSH6 mutation carriers in this family when compared with MSH2 and MLH1 carriers (32% by the age of 80 v >80%). Endometrial cancer is a frequent manifestation among female carriers (six out of 13 malignant tumours). Transitional cell carcinoma of the urinary tract is also relatively common in both male and female carriers (10% of the carriers). Moreover, the mean age of onset of both colorectal cancer (MSH6 v MSH2/MLH1 = 55 years v 44/41 years) and endometrial carcinomas (MSH6 v MSH2/MLH1 = 55 years v 49/48 years) is delayed. As previously reported, we confirm that the pattern of microsatellite instability, in combination with immunohistochemical analysis, can predict the presence of a MSH6 germline defect. The detailed characterisation of the clinical phenotype of this kindred contributes to the establishment of genotype-phenotype correlations in HNPCC owing to mutations in specific mismatch repair genes.

Radiotherapy for oesophagus carcinoma: the impact of p53 on treatment outcome.

BACKGROUND AND PURPOSE: Wildtype p53 protein plays an important role in the cellular response to ionizing radiation and other DNA damaging agents and is mutated in many human tumours. We evaluated the relationship of the immunohistochemically determined p53 protein status and the disease control with radiotherapy alone for carcinoma of the oesophagus. MATERIALS AND METHODS: Immunostaining for p53 protein was performed on paraffin-embedded specimens from 69 patients with adeno- and squamous cell carcinoma of the oesophagus. All patients were treated by radiotherapy exclusively, consisting of a combination of external irradiation and intraluminal brachytherapy, using two different dose levels. RESULTS: Fifty-four percent (37/69) of the tumours showed overexpression of the p53 protein. No difference in pre-treatment parameters for p53-positive and p53-negative cases was detected. In multivariate analysis p53 was significantly associated with overall survival (OS) next to weight loss, tumour stage and N-stage. For metastatic-free survival (MFS) p53 status proved to be the sole independent prognostic factor. The influence of p53 on local recurrence-free survival (LRFS), however, was not as strong as on OS and MFS. CONCLUSIONS: Immunohistochemically detected overexpression of mutated p53 protein in oesophagus carcinoma was an independent prognostic factor in a group of patients treated with radiotherapy alone.

E-cadherin expression in oesophageal carcinoma treated with high-dose radiotherapy; correlation with pretreatment parameters and treatment outcome.

BACKGROUND AND PURPOSE: E-cadherin plays an important role in the cell-cell contact of normal epithelium. Loss of E-cadherin expression may be related to tumour invasiveness and metastatic potential. In a group of patients treated for oesophageal carcinoma by radiotherapy only, we found that immunohistochemically detected p53 expression correlated with reduced survival, mainly because of the occurrence of distant metastases. We questioned whether, in this group of patients, E-cadherin expression was concomitantly altered and served as a predictive factor for the development of distant metastases. MATERIALS AND METHODS: Immunostaining for E-cadherin was performed on paraffin- embedded biopsy specimens from patients with adenocarcinoma and squamous cell carcinoma of the oesophagus. E-cadherin status and its correlation with regard to pretreatment parameters and treatment outcome were determined. RESULTS: An aberrant staining pattern of E-cadherin did not correlate with any of the pretreatment parameters. In a univariate analysis, a significantly reduced metastatic potential was found for tumours that had an aberrant cellular staining pattern for E-cadherin, which was strongest for squamous cell carcinomas. However, in a multivariate analysis only p53 status correlated significantly with the occurrence of distant metastases. CONCLUSION: Although, in univariate analysis, aberrant E-cadherin expression served as a better, rather than a worse prognostic factor, p53 status remained the only significant parameter in multivariate analysis, in this group of patients with oesophageal carcinoma. No relationship between p53 status and E-cadherin expression was found.

The prognostic influence of neuroendocrine differentiation in breast cancer: results of a long-term follow-up study.

Reports about neuroendocrine (NE) differentiation in breast carcinomas and its possible relation with prognosis are scarce. Furthermore the results of some studies have not been subjected to multivariate survival analysis and the follow-up periods were relatively short. Therefore, in the present long-term follow-up study, the prognostic influence of immunohistochemically defined NE cells, present in the tumours of 40 out of 317 (12.6%) curatively operated breast cancer patients, was studied. The mean follow-up period was 104 months. NE differentiation (NED) was determined by the immunohistochemical detection of chromogranin A and/or synaptophysin. This is concordant with other studies focussing on NED in breast cancer. In contrast to the literature in our series only in 9 out of 40 cases (23%) we were able to detect coexpression of chromogranin A and synaptophysin. This might be due to the characteristics of the antibodies we used. Although most tumours in our series were of the usual type, some tumours with NED were of a special type. Neither univariately, nor taking account of various known prognostic factors, does focal NED appear to carry a special prognostic significance. This finding is in line with results of previous studies.

P53 and radiotherapy for oesophageal carcinoma: A comparison between 4 different antibodies.

Wild-type p53 protein plays an important role in the cellular response to ionising radiation and other DNA damaging agents and is mutated in many human tumours. Immunohistochemistry is a rapid method to detect elevated protein levels. The p53 status in a group of patients treated by radiotherapy exclusively for oesophagus carcinoma was examined and correlated with pre-treatment parameters and treatment outcome with regard to overall survival, local recurrence-free survival and distant metastases-free survival. Four different antibodies were used to evaluate their predictive power to detect p53 expression. Immunostaining for p53 protein with 4 different antibodies was performed on paraffin-embedded specimens from 69 patients with adenocarcinoma and squamous cell carcinoma of the oesophagus. All patients were treated with radiotherapy exclusively, consisting of external irradiation combined with intraluminal brachytherapy. Detection of p53 using the antibody (DO7) was significantly correlated with overall survival and distant metastases-free survival. For local recurrence-free survival no statistical significance was reached. The use of the other 3 antibodies all showed the same trend with regard to distant metastases, however statistical significance was not reached. The use of multiple antibodies did not increase the predictive value of DO7. Both for survival and metastases-free survival the use of DO7 alone was sufficient as a significant prognostic factor. We conclude that in this series, only the use of DO7 was correlated with prognosis in oesophagus carcinoma treated by radiotherapy only and that addition of 3 antibodies did not improve the predictive power.

The influence of pre-operative radiotherapy on the expression of p53 and adhesion molecules: correlation with treatment results in patients with squamous cell carcinoma or adenocarcinoma.

E-cadherin and the catenins are responsible for inter-cellular adhesion in epithelial tissues. E-cadherin and/or catenin expression is often altered in malignancies, leading to increased invasiveness and metastatic activity of tumour cells. Intact adhesion molecules reduce the risk on distant metastases. This is confirmed by studies mostly performed on surgical series, correlating e-cadherin expression in tumours with prognosis and treatment outcome. It has become more apparent that anti-cancer treatment by itself can also affect the expression of adhesion molecules. We therefore suggest that the prognostic value of e-cadherin expression may depend on the treatment modality and the sequence of therapies administered for malignant tumours. We used paraffin embedded specimens from patients with rectal tumours and patients with laryngeal tumours treated by short course radiotherapy before definitive surgery. Expression of p53, e-cadherin, and beta-catenin was determined in pre-radiotherapy biopsies and in the surgical specimens. Material was available from 37 patients. We found no correlation between the expression of p53, e-cadherin or beta-catenin and pre-treatment parameters. Mutated p53 in pre-radiation biopsy correlated with increased occurrence of distant metastases and there was an unexpected trend for abnormal e-cadherin expression to correlate with reduced metastases. The prognostic value of p53 no longer existed after examination of the surgical specimens (post-radiotherapy). There was a trend for e-cadherin to reverse from abnormal to normal expression in laryngeal squamous cell carcinomas after radiotherapy, in 5/7 cases it was accompanied with p53 conversion from positive to negative expression. Based on this study it is suggested that the predictive value of the e-cadherin expression for the occurrence of distant metastases in tumours treated by radiotherapy before surgery may be different from that found in tumours treated by surgery only. This may be related to the influence of radiotherapy on e-cadherin expression, especially in squamous cell carcinomas. Alteration in p53 expression was of predictive value only in pre-treatment biopsies and the beta-catenin status did not correlate with treatment outcome in this series.

Death from chemotherapy in gestational trophoblastic disease.

Multiple cytotoxic drug administration is the generally accepted treatment of patients with a high-risk stage of choriocarcinoma. Based on this principle a 27-year old woman, classified as being in the high-risk group (Goldstein and Berkowitz score: 11), was treated with multiple cytotoxic drugs. The multiple drug schema consisted of: Etoposide 16.213, Methotrexate, Cyclophosphamide, Actomycin-D, and Cisplatin. On the first day of the schedule, moderate high doses of Methotrexate, Etoposide and Cyclophosphamide were administered. Within 8 hours after initiation of therapy the patient died with a clinical picture resembling massive pulmonary obstruction due to choriocarcinomic tissue plugs, probably originating from the uterus. Formation of these plugs was probably due to extensive tumor necrosis at the level of the walls of the major uterine veins, which resulted in an open exchange of tumor plugs to the vascular spaces; decrease in tumor tissue coherence secondary to chemotherapy may have further contributed to the formation of tumor emboli. In view of the close time association between the start of chemotherapy and the acute onset of massive embolism other explanations, such as spontaneous necrosis, must be considered less likely. Patients with large pelvic tumor loads are, according to existing classifications, at high risk to die and to develop drug resistance. Notwithstanding these facts our findings suggest that these patients might benefit from relatively mild initial treatment, especially true for patients not previously exposed to this drug. Close observation of the response status both clinically and with beta-hCG values may indicate whether and when more agressive combination chemotherapy should be started.(ABSTRACT TRUNCATED AT 250 WORDS)

Haemolytic disease of the newborn due to anti-K antibodies.

We describe a case of intrauterine foetal death at 32 weeks gestation with signs of hydrops foetalis without erythroblastosis. The hydrops foetalis appeared to be caused by blood group incompatibility. Irregular antibodies of anti-K specificity were found in the serum of the mother, while the father was positive for the K-antigen. The antibody dependent cell-mediated cytotoxicity (ADCC) test with serum of the mother was positive. This case shows the characteristics of haemolytic disease of the newborn by anti-K. Moreover, it underlines the necessity of both adequate follow-up of individual cases, and screening for irregular antibodies in all pregnancies as well as central registration of screening results.

[Less operations required due to perioperative frozen section examination of sentinel nodes in 275 breast cancer patients]. / Minder operaties vereist door peroperatief vriescoupeonderzoek van schildwachtklieren bij 275 patiënten met mammacarcinoom.

OBJECTIVE: To determine the reliability of a peroperative frozen section examinations of sentinel lymph nodes in mammary carcinoma. DESIGN: Retrospective. METHOD: In the Reinier de Graaf Hospital and Diagnostic Centre SSDZ Delft, the Netherlands, the results of frozen section from sentinel lymph node investigations of mammary carcinomas from 1997-2000 were compared with the final pathological results. If axillary dissection had been performed on these patients, the histopathological findings of the dissected lymph nodes were also studied. RESULTS: Frozen sections were made of 287 sentinel lymph nodes from 275 patients. A tumour was found in the sentinel lymph nodes of 64 patients and these patients immediately underwent a complete axillary lymph node dissection. For 31 of these patients a tumour was also found in the other lymph nodes. In 29 of these 31 patients, histological examination had shown extranodal extension. The frozen sections from the sentinel nodes of the remaining 211 patients were considered negative. However, in 13 of these patients, the paraffin sections of the sentinel node nevertheless showed a tumour and the remaining axillary lymph nodes were removed in a second operation. In the last 89 patients studied, the sentinel lymph nodes were cut at four levels and stained immunohistochemically at one level for cytokeratins. Accordingly micrometastases were found in the sentinel lymph nodes of 4 of the 13 patients with (false-)negative frozen sections. False-positive results did not occur. CONCLUSION: The major advantage of the sentinel node method in breast cancer is that for women without metastasis present in the sentinel node, axillary dissection is avoided. By means of a peroperative examination of frozen sections, 83% of the patients with a metastasis in the sentinel lymph node (or about one quarter of all patients) were spared from having a second operation for axillary dissection at a later stage.

[The detection of melanoma metastases in sentinel nodes by polymerase chain reaction]. / Opsporing van metastasen van melanomen in schildwachtklieren met de polymerasekettingreactie.

OBJECTIVE: To investigate the possibility of detecting melanoma metastases using molecular biological techniques in patients with a primary malignant melanoma. DESIGN: Descriptive. SETTING: Reinier de Graaf Gasthuis Hospital Delft and diagnostic centre SSDZ Delft, the Netherlands. METHODS: A melanoma specific tyrosinase mRNA was amplified using the reverse transcriptase polymerase chain reaction (RT-PCR) technique. RESULTS: A total of 23 sentinel nodes derived from 9 patients were examined. In 14 sentinel nodes metastases were found using RT-PCR. In microscopic slides stained with haematoxylin and eosin (HE) melanoma metastases were found in 2 sentinel nodes of 2 patients. With immunohistochemistry melanoma metastases were found in the same 2 sentinel nodes. CONCLUSION: It is possible to detect melanoma metastases in sentinel nodes using molecular biological techniques.

Impact of Ovabloc intratubal polymer on the morphology of the fallopian tube.

Treatment of the fallopian tubes by Ovabloc silicone intratubal polymer (ITP; Dow Corning silastic 382 Medical-Grade Elastomer) is used for permanent female contraception. Studies on animals show minor structural changes in the tubal epithelial lining caused by the ITP and suggest reversibility of tubal integrity after removal of the polymer. The aim of this study was to examine the impact of the ITP on the human tubal epithelial lining. From 13 patients treated with ITP, 23 fallopian tubes were studied by light microscopy and transmission electron microscopy. The examined 19 isthmical and 10 ampullary parts show marked structural changes. In the isthmical parts of the fallopian tubes, cellular and ciliary changes develop immediately after insertion of the silicone polymer. As for the ampullary parts, the cellular and ciliary changes are related to the ITP exposure time. The changes in the isthmus and ampulla are persistent for at least 15 months after removal of the ITP. It is not known whether the effects are reversible.

Angiotensin II receptor blockade after myocardial infarction in rats: effects on hemodynamics, myocardial DNA synthesis, and interstitial collagen content.

Angiotensin-converting enzyme (ACE) inhibitors are widely used for treatment of heart failure after myocardial infarction (MI). The beneficial effects consist of a combination of hemodynamic effects and interference with cardiac structural alterations. These effects are believed to depend on inhibition of angiotensin II (AII) formation and thus diminished angiotensin receptor stimulation. We administered the angiotensin II-1 (AT-1) receptor antagonist losartan during and after completion of the repair phase of an MI to investigate involvement of the AT-1 receptor in the above described effects of captopril. MI reduced cardiac output (CO) (sham 94 +/- 4 ml/min, MI 78 +/- 5 ml/min) and maximal CO (sham 154 +/- 4, MI 107 +/- 5 ml/min, respectively). Losartan (15 mg/kg/day) resulted in a rightward shift of the AII pressor dose-response curve by a factor of 32-40. Neither CO nor COVL,max was affected by losartan treatment in either phase (late treatment CO = 78 +/- 5, COVL,max = 118 +/- 9 ml/min). Although early treatment with losartan reduced cardiac hypertrophy measured as heart weight, DNA synthesis was reduced only slightly. In contrast, collagen deposition was inhibited completely. The results suggest that the effects of captopril in rats after MI are not dependent on AT-1 receptor-mediated mechanisms.

Cardiac remodeling in hypertension and following myocardial infarction: effects of arteriolar vasodilators.

Cardiac remodeling constitutes a risk factor in cardiovascular disease. It may occur in a variety of circumstances. In hypertension and following myocardial infarction, pharmacological intervention in the remodeling process has been the subject of several studies. But the mechanisms of action of drugs that do contribute to regression of the remodeling response are still a matter of debate. Much of the confusion around the subject comes from the fact that classic arteriolar vasodilators do not result in such regression in hypertensive cardiac hypertrophy. This paper reviews some of the literature to examine whether there is indeed an exceptional position for vasodilators in hypertensive heart disease. Although, conceptually, arteriolar dilatation, and thus afterload reduction, might also have favorable effects on the remodeling response following myocardial infarction, clinical studies suggest the opposite. In the present paper, possible mechanisms are discussed, and evidence is presented that shows that hydralazine has an unexpected effect on the remodeling response at the level of the extracellular matrix.

The prognostic value of pretreatment expression of androgen receptor and bcl-2 in hormonally treated prostate cancer patients.

PURPOSE: We determined the prognostic value of oncoprotein bcl-2 and androgen receptor expression in pretreatment transurethral resection specimens of hormonally treated prostate cancer patients. MATERIALS AND METHODS: A total of 68 pretreatment transurethral resection specimens, 30 radical prostatectomy specimens and 21 palliative transurethral resection specimens with androgen independent prostate cancer was stained with a monoclonal antibody against bcl-2. Androgen receptor immunohistochemistry was performed on pretreatment transurethral resection specimens only. Results were scored semiquantitatively and were correlated with tumor stage and grade and with the occurrence of clinical progression or tumor related death. RESULTS: Bcl-2 expression by adenocarcinoma cells was found in 32, 17 and 24% of pretreatment transurethral resection, radical prostatectomy and palliative transurethral resection specimens, respectively. The bcl-2 scores did not correlate with tumor stage or grade. Androgen receptor was expressed in 88% of pretreatment transurethral resection specimens. Androgen receptor scores were marginally related to tumor grade, but not to tumor stage. A prognostic value of bcl-2 or androgen receptor in pretreatment transurethral resection specimens was not found. When a combined bcl-2/androgen receptor score was used, this parameter was an independent prognostic marker to predict clinical progression with Gleason grade and stage classification. Gleason grade was the only independent prognostic marker to predict tumor related death. CONCLUSIONS: The expression of bcl-2 and androgen receptor in pretreatment prostate cancer specimens is not related to the prognosis of hormonally treated prostate cancer. Bcl-2 expression is not increased in endocrine therapy resistant prostate cancer. Surprisingly, a combined bcl-2/androgen receptor score acts as an independent prognosticator for clinical progression.