http://repub.eur.nl/res/aut/17591/ List of Publications en http://repub.eur.nl/static-eur/img/logo.png http://repub.eur.nl http://repub.eur.nl/res/pub/24207/ 2009-03-01T00:00:00Z Endocrine treatment of breast cancer is widely applied and effective. However, in advanced disease cases, the tumors will eventually progress into an estrogen-independent and therapy-resistant phenotype. To elucidate the molecular mechanisms underlying this endocrine therapy failure, we applied retroviral insertion mutagenesis to identify the main genes conferring estrogen independence to human breast cancer cells. Estrogen-dependent ZR-75-1 cells were infected with replication-defective retroviruses followed by selection with the anti-estrogen 4-hydroxy-tamoxifen. In the resulting panel of 79 tamoxifen-resistant cell lines, the viral integrations were mapped within the human genome. Genes located in the immediate proximity of the retroviral integration sites were characterized for altered expression and their capacity to confer anti-estrogen resistance when transfected into breast cancer cells. Out of 15 candidate BCAR (breast cancer anti-estrogen resistance) genes, seven (AKT1, AKT2, BCAR1, BCAR3, EGFR, GRB7, and TRERF1/BCAR2) were shown to directly underlie estrogen independence. Our results show that insertion mutagenesis is a powerful tool to identify BCAR loci, which may provide insights into the molecular and cellular mechanisms of breast tumor progression and therapy resistance thereby offering novel targets for the development of tailor-made therapeutical and prevention strategies. http://repub.eur.nl/res/pub/24889/ 2009-03-01T00:00:00Z Background: Crohn's disease and ulcerative colitis have a complex genetic background. We assessed the risk for both the development and severity of the disease by combining information from genetic variants associated with inflammatory bowel disease (IBD). Methods: We studied 2804 patients (1684 with Crohn's disease and 1120 with ulcerative colitis) and 1350 controls from seven university hospitals. Details of the phenotype were available for 1600 patients with Crohn's disease and for 800 with ulcerative colitis. Genetic association for disease susceptibility was tested for the nucleotide-binding and oligomerisation domain 2 gene (NOD2), the IBD5 locus, the Drosophila discs large homologue 5 and autophagy-related 16-like 1 genes (DLG5 and ATG16L1) and the interleukin 23 receptor gene (IL23R). Interaction analysis was performed for Crohn's disease using the most associated single nucleotide polymorphism (SNP) for each locus. Odds ratios were calculated in an ordinal regression analysis with the number of risk alleles as an independent variable to analyse disease development and severity. Results: Association with Crohn's disease was confirmed for NOD2, IBD5, DLG5, ATG16L1 and IL23R. Patients with Crohn's disease carry more risk alleles than controls (p = 3.85 × 10-22). Individuals carrying an increasing number of risk alleles have an increasing risk for Crohn's disease, consistent with an independent effects multiplicative model (trend analysis p = 4.25 × 10-23). Patients with Crohn's disease with a more severe disease course, operations or an age of onset below 40 years have more risk alleles compared to non-stricturing, non-penetrating behaviour (p = 0.0008), no operations (p = 0.02) or age of onset above 40 years (p = 0.028). Conclusion: Crohn's disease is a multigenic disorder. An increase in the number of risk alleles is associated with an increased risk for the development of Crohn's disease and with a more severe disease course. Combining information from the known common risk polymorphisms may enable clinicians to predict the course of Crohn's disease. http://repub.eur.nl/res/pub/14976/ 2009-02-01T00:00:00Z Objective: Prognosis in women with ovarian cancer mainly depends on International Federation of Gynecology and Obstetrics stage and the ability to perform optimal cytoreductive surgery. Since ovarian cancer has a heterogeneous presentation and clinical course, predicting progression-free survival (PFS) and overall survival (OS) in the individual patient is difficult. The objective of this study was to determine predictors of PFS and OS in women with advanced stage epithelial ovarian cancer (EOC) after primary cytoreductive surgery and first-line platinum-based chemotherapy. Design: Retrospective observational study. Setting: Two teaching hospitals and one university hospital in the south-western part of the Netherlands. Population: Women with advanced stage EOC. Methods: All women who underwent primary cytoreductive surgery for advanced stage EOC followed by first-line platinum-based chemotherapy between January 1998 and October 2004 were identified. To investigate independent predictors of PFS and OS, a Cox' proportional hazard model was used. Nomograms were generated with the identified predictive parameters. Main outcome measures: The primary outcome measure was OS and the secondary outcome measures were response and PFS. Results: A total of 118 women entered the study protocol. Median PFS and OS were 15 and 44 months, respectively. Preoperative platelet count (P = 0.007), and residual disease <1 cm (P = 0.004) predicted PFS with a optimism corrected c-statistic of 0.63. Predictive parameters for OS were preoperative haemoglobin serum concentration (P = 0.012), preoperative platelet counts (P = 0.031) and residual disease <1 cm (P = 0.028) with a optimism corrected c-statistic of 0.67. Conclusion: PFS could be predicted by postoperative residual disease and preoperative platelet counts, whereas residual disease, preoperative platelet counts and preoperative haemoglobin serum concentration were predictive for OS. The proposed nomograms need to be externally validated. http://repub.eur.nl/res/pub/16390/ 2009-01-01T00:00:00Z To unravel the mechanisms underlying failure of endocrine therapy of breast cancer, we have previously executed a functional genetic screen and identified the adaptor protein BCAR1 to be causative for tamoxifen resistance. As a consequence of the manifold of interactions with other proteins, we characterized the contribution of individual protein domains of BCAR1 to anti-estrogen-resistant proliferation of human breast cancer cells. We took advantage of the observation that the closely related family member HEF1 was unable to support long-term anti-estrogen-resistant cell proliferation. Chimerical proteins containing defined domains of BCAR1 and HEF1 were evaluated for anti-estrogen-resistant growth. Exchange of the SH3 and C-terminal domains did not modify the capacity to support cell proliferation. Full support of anti-estrogen resistant proliferation was observed for chimerical molecules containing the central part of BCAR1. The bi-partite SRC-binding site or the Serine-rich domain did not explain the differential capacity of BCAR1. These findings indicate that the differences between BCAR1 and HEF1 with respect to support of anti-estrogen resistance reside in the substrate domain which contains multiple sites for tyrosine phosphorylation. The crucial interactions required for anti-estrogen resistance occur within the substrate domain of BCAR1. Further deciphering of these interactions may resolve the growth regulatory mechanism and provide an explanation for the observation that primary tumors with high levels of BCAR1 are likely to fail on tamoxifen therapy. This information may also help to devise alternative personalized treatment strategies with improved outcome for breast cancer patients. http://repub.eur.nl/res/pub/25063/ 2009-01-01T00:00:00Z The introduction of prenatal screening requires rapid high-throughput diagnosis of common aneuploidies. Multiplex ligation-dependent probe amplification (MLPA) allows for quick, easily automated multiplex testing of these aneuploidies in one polymerase chain reaction. We performed a large prospective study using MLPA on 4000 amniotic fluid (AF) samples including all indications and compared its value to karyotyping and fluorescence in situ hybridization (FISH). MLPA can reliably determine common aneuploidies with 100% sensitivity and 100% specificity. Moreover, some mosaic cases and structural chromosome aberrations were detected as well. In cases of a male fetus, triploidies can be detected by an aberrant pattern of probe signals, which mimics maternal cell contamination (MCC). Macroscopic blood contamination was encountered in 3.2% of the AF samples. In 20% of these samples, an MLPA pattern was found consistent with MCC, although there were no false negatives of the most common aneuploidies. As the vast majority of inconclusive results (1.7%) is due to potential MCC, we designed a protocol in which we determine whether MLPA can be performed on blood-contaminated AF samples by testing if blood is of fetal origin. Then, the number of inconclusive results could be theoretically reduced to 0.05%. We propose an alternative interpretation of relative probe signals for rapid aneuploidy diagnosis (RAD). We discuss the value of MLPA for the detection of (submicroscopic) structural chromosome anomalies. MLPA is a reliable method that can replace FISH and could be used as a stand-alone test for RAD instead of karyotyping. http://repub.eur.nl/res/pub/13506/ 2004-09-15T00:00:00Z PURPOSE: BCAR1, the human homologue of the rat p130Cas protein, was identified in a functional screen for human breast cancer cell proliferation resistant to antiestrogen drugs. Here, we study the prognostic value of quantitative BCAR1 levels in a large series of breast cancer specimens. EXPERIMENTAL DESIGN: A specific ELISA was developed to measure BCAR1 protein levels in 2593 primary breast tumor cytosols. Tumor levels of BCAR1 were correlated with relapse-free survival (RFS) and overall survival (OS) and compared with collected data on urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor 1 (PAI-1). RESULTS: In tumor cytosols, BCAR1 protein levels varied between 0.02 and 23 ng/mg protein. BCAR1 levels exhibited a positive correlation with steroid hormone receptor levels, age and menopausal status, and uPA and PAI-1 levels. The level of BCAR1 (continuous or categorized as low, intermediate, or high) was inversely related with RFS and OS time. Multivariate analysis showed that BCAR1 levels contributed independently to a base model containing the traditional prognostic factors for both RFS and OS (both P < 0.0001). When added together with uPA and PAI-1 in the multivariate model, BCAR1 contributed independently of PAI-1 and was favored over uPA. Interaction tests allowed for additional analyses of BCAR1 protein levels in clinically relevant subgroups stratified by nodal and menopausal status. CONCLUSIONS: The quantitative BCAR1 protein level represents a prognostic factor for RFS and OS in primary breast cancer, independent of the traditional prognostic factors and the other novel marker PAI-1. http://repub.eur.nl/res/pub/13187/ 2003-09-01T00:00:00Z Since 1994, a population-based study of frontotemporal dementia (FTD) in The Netherlands has aimed to ascertain all patients with FTD, and first prevalence estimates based on 74 patients were reported in 1998. Here, we present new prevalence estimates after expansion of our FTD population to 245 patients, with emphasis on the prevalence in the province Zuid-Holland where the main study centre is located. All neurologists and physicians in nursing homes received a yearly postal enquiry about suspected FTD cases. FTD was diagnosed in 245 patients according to the Lund-Manchester criteria, supported by neuroimaging and neuropsychology. tau mutation analysis was performed in a subgroup of 154 patients (63%), and 40 out of 98 patients (41%) who died during follow-up were autopsied during the course of the study. The prevalence of FTD in the province Zuid-Holland was 3.6 per 100,000 at age 50-59 years, 9.4 per 100,000 at age 60-69 years and 3.8 per 100,000 at age 70-79 years. The median age at onset of the 245 patients (51% female) was 58.0 years (range 33-80 years). Dementia in one or more first-degree family members was found in 43% of patients and mutation analysis of the tau gene showed mutations in 34 patients (19 P301L, five L315R, four G272V, four R406W, one Delta K280 and one S320F), all with a positive family history for dementia (14% of the total population, 32% of patients with a positive family history). Pathological findings in the 40 autopsied patients consisted of dementia lacking distinctive histology in 22%, FTD with ubiquitin-positive inclusions in 33%, Pick's disease in 15% and tauopathy in the remaining 30% of patients, with tau mutations identified in more than half of the latter patients. We conclude that the prevalence of FTD in The Netherlands is higher than previously reported, confirming that FTD is more common than was previously thought. The finding of tau mutations in 32% of patients with a positive family history for dementia justifies mutation screening in FTD patients with a positive family history, while tau mutations in non-familiar cases are rare. http://repub.eur.nl/res/pub/9845/ 2002-01-01T00:00:00Z In a randomized multicenter study, the efficacies of two different GnRH agonists were compared with that of hCG for triggering final stages of oocyte maturation after ovarian hyperstimulation for in vitro fertilization. Ovarian stimulation was conducted by recombinant FSH (Puregon), and the GnRH antagonist ganirelix (Orgalutran) was coadministered for the prevention of a premature LH rise. Luteal support was provided by daily progestin administration. Frequent blood sampling was performed at midcycle in the first 47 eligible subjects included in the current study, who were randomized for a single dose of 0.2 mg triptorelin (n = 17), 0.5 mg leuprorelin (n = 15), or 10,000 IU hCG (n = 15). Serum concentrations of LH, FSH, E2, and progesterone (P) were assessed at variable intervals. LH peaked at 4 h after both triptorelin and leuprorelin administration, with median LH levels of 130 and 107 IU/liter (P < 0.001), respectively. LH levels returned to baseline after 24 h. Subjects receiving hCG showed peak levels of 240 IU/liter hCG 24 h after administration. A rise in FSH to 19 IU/liter (P < 0.001) was noted in both GnRH agonist groups 8 h after injection. Within 24 h the areas under the curve for LH and FSH were significantly higher (P < 0.001) in both GnRH agonist groups compared with that for hCG. E2 and P levels were similar for all groups up to the day of oocyte retrieval. Luteal phase areas under the curve for P and E2 were significantly elevated (P < 0.001) in the hCG group. The mean (+/-SD) numbers of oocytes retrieved were 9.8 +/- 5.4, 8.7 +/- 4.5, and 8.3 +/- 3.3; the percentages of metaphase II oocytes were 72%, 85%, and 86%; and fertilization rates were 61%, 62%, and 56% in the triptorelin, leuprorelin, and hCG group, respectively (P = NS for all three comparisons). These findings support the effective induction of final oocyte maturation in both GnRH agonist groups. In summary, after treatment with the GnRH antagonist ganirelix for the prevention of premature LH surges, triggering of final stages of oocyte maturation can be induced effectively by a single bolus injection of GnRH agonist, as demonstrated by the induced endogenous LH and FSH surge and the quality and fertilization rate of recovered oocytes. Moreover, corpus luteum formation is induced by GnRH agonists with luteal phase steroid levels closer to the physiological range compared with hCG. This more physiological approach for inducing oocyte maturation may represent a successful and safer alternative for in vitro fertilization patients undergoing ovarian hyperstimulation. http://repub.eur.nl/res/pub/8814/ 1998-01-01T00:00:00Z This case report describes the first attempt to treat imminent ovarian hyperstimulation syndrome (OHSS) by using a gonadotrophin-releasing hormone (GnRH) antagonist. A 33 year old, normo-ovulatory woman undergoing in-vitro fertilization received daily subcutaneous injections of 150 IU of recombinant follicle-stimulating hormone (recFSH) from cycle day 2, together with GnRH antagonist (ganirelix) 0.125 mg from cycle day 7 onwards. On cycle day 10 the patient was found to have a serum oestradiol concentration of 16 500 pmol/l and, on ultrasound examination, four preovulatory (>16 mm) and nine intermediate sized (10-16 mm) follicles. RecFSH injections were discontinued, human chorionic gonadotrophin (HCG) withheld, whereas the ganirelix dose was increased to 2 mg/d. This regimen led to a rapid decrease in serum oestradiol concentrations and the decrease in ovarian size on ultrasound. Since GnRH antagonists will become clinically available for in-vitro fertilization programmes in the near future this suggested regimen might have a role in preventing severe OHSS.