ObjectiveProtein fermentation results in production of metabolites such as ammonia, amines and indolic, phenolic and sulfur-containing compounds. In vitro studies suggest that these metabolites might be toxic. However, human and animal studies do not consistently support these findings. We modified protein fermentation in healthy subjects to assess the effects on colonic metabolism and parameters of gut health, and to identify metabolites associated with toxicity.DesignAfter a 2-week run-in period with normal protein intake (NP), 20 healthy subjects followed an isocaloric high protein (HP) and low protein (LP) diet for 2 weeks in a cross-over design. Protein fermentation was estimated from urinary p-cresol excretion. Fecal metabolite profiles were analyzed using GC-MS and compared using cluster analysis. DGGE was used to analyze microbiota composition. Fecal water genotoxicity and cytotoxicity were determined using the Comet assay and the WST-1-assay, respectively, and were related to the metabolite profiles.ResultsDietary protein intake was significantly higher during the HP diet compared to the NP and LP diet. Urinary p-cresol excretion correlated positively with protein intake. Fecal water cytotoxicity correlated negatively with protein fermentation, while fecal water genotoxicity was not correlated with protein fermentation. Heptanal, 3-methyl-2-butanone, dimethyl disulfide and 2-propenyl ester of acetic acid are associated with genotoxicity and indole, 1-octanol, heptanal, 2,4-dithiapentane, allyl-isothiocyanate, 1-methyl-4-(1-methylethenyl)-benzene, propionic acid, octanoic acid, nonanoic acid and decanoic acid with cytotoxicity.ConclusionThis study does not support a role of protein fermentation in gut toxicity. The identified metabolites can provide new insight into colonic health.Trial RegistrationClinicalTrial.gov NCT01280513

B cell specific immunomodulatory drugs still remain an unmet medical need. Utilisation of validated simplified in vitro models would allow readily obtaining new insights in the complexity of B cell regulation. For this purpose we investigated which human B lymphocyte stimulation assays may be ideally suited to investigate new B lymphocyte immunosuppressants. Primary polyclonal human B cells underwent in vitro stimulation and their proliferation, production of immunoglobulins (Igs) and of cytokines, and expression of cell surface molecules were analysed using various stimuli. ODN2006, a toll-like receptor 9 (TLR9) agonist, was the most potent general B cell stimulus. Subsequently, we investigated on which human B cell lines ODN2006 evoked the broadest immunostimulatory effects. The Namalwa cell line proved to be the most responsive upon TLR9 stimulation and hence may serve as a relevant, homogeneous, and stable B cell model in an in vitro phenotypic assay for the discovery of new targets and inhibitors of the B cell activation processes. As for the read-out for such screening assay, it is proposed that the expression of activation and costimulatory surface markers reliably reflects B lymphocyte activation.

Our aim was to estimate the prevalence of mutations in the BRCA1 and BRCA2 genes among unselected incident cases of breast cancer in young women. We identified 158 incident breast cancer cases diagnosed before age 46 years in predefined geographic areas in Girona and Tarragona, Spain, during 1995-1997. Of these, 136 (86%) provided information on family history of cancer and were screened for BRCA1 and BRCA2 mutations. Nine of the 136 (6.6%) were found to carry BRCA deleterious mutations (MUT) (1 BRCA1 and 8 BRCA2), and 20 were detected with rare BRCA variants of unknown significance (UV). Both MUT and US BRCA alterations were more frequent in younger patients: 7 (11.6%) MUT and 12 (19.3%) UV carriers were found in the group of 62 patients younger than 40 years, whereas 2 (2.7%) MUT and 9 (12%) US carriers were identified in the group of 74 patients aged 40 -45. Family history of breast and ovarian cancers suggestive of hereditary condition (at least 2 first-or second-degree relatives affected with breast cancer or at least 1 relative affected with ovarian cancer or early-onset breast cancer) was absent for 5 of 9 MUT carriers. This suggests that BRCA screening policies based on family history of cancer would miss a considerable proportion of BRCA mutations. Mutations in the BRCA1 and BRCA2 genes explain at least 10% of breast cancer cases diagnosed before age 40 years. The contribution of these genes to early-onset breast cancer is likely to be even higher given that certain UV cases might be disease-associated. © 2003 Wiley-Liss, Inc. Key words: BRCA; breast cancer; SpainBreast cancer is the most frequent cancer type among women in the world, affecting up to 12% of all women in Europe and North America. 1 Germline mutations of the BRCA1 and BRCA2 genes substantially increase the lifetime risk of developing breast and ovarian cancers. 2,3 The estimated cumulative risk for breast cancer by age 70 in carriers of these mutations is about 80%. 2-5 Estimates of penetrance and contribution of BRCA1 and BRCA2 to cancer incidence have been mainly based on data from high-risk families and might not reflect the situation in all BRCA1/2 carriers. Several studies on founder mutations in populations like Ashkenazi Jews or Icelanders have provided penetrance estimates that were lower than those obtained in high-risk families. 4,5 The same trend was observed in Australian and UK studies on outbred populationbased series of breast cancers, estimating breast cancer penetrance to be 40 -50% by age 70. 6,7 Several reports estimating the prevalence of BRCA1 and BRCA2 mutations in large population-based samples of breast cancer cases have been published. 6 -11 However, all but the study of Loman et al. 11 confronted the problems of a relatively low proportion of incident cases included in the analysis and limited mutation screening protocols.Our aim was to evaluate in the Spanish population the prevalence of mutations in the BRCA1 and BRCA2 genes among unselected incident cases of breast cancer diagnosed before the age of 46 years....

Both clinical and experimental evidence illustrate that p190 and p210 BCR/ABL oncogenic tyrosine kinases induce resistance to DNA damage and confer an intrinsic genetic instability. Here, we investigated whether BCR/ABL expression could modulate nucleotide excision repair (NER). We found that ectopic expression of p210 BCR/ABL in murine lymphoid BaF3 cell line inhibited NER activity in vitro, promoting hypersensitivity of these cells to ultraviolet (UV) treatment and facilitating a mutator phenotype. However, expression of p210 BCR/ABL in human and murine myeloid cell lines and primary bone marrow cells resulted in the increased NER activity and resistance to UV irradiation. The ABL tyrosine kinase inhibitor STI571 reversed these effects, showing that p210 BCR/ABL tyrosine kinase activity is responsible for deregulation of NER. Hypoactivity of NER in p210 BCR/ABL-positive lymphoid cells was accompanied by the decreased interaction between proliferating cell nuclear antigen (PCNA) and xeroderma pigmentosum group B (XPB); conversely, this interaction was enhanced in p210 BCR/ABLpositive myeloid cells. p190 BCR/ABL did not affect NER in lymphoid and myeloid cells. In summary, our study suggests that p210 BCR/ABL reduced NER activity in lymphoid cells, leading to hypersensitivity to UV and mutagenesis. In contrast IntroductionBCR/ABL oncogenic tyrosine kinase is derived from translocation of the c-abl gene on chromosome 9 to the bcr locus on chromosome 22 (t(9;22); Philadelphia chromosome [Ph]1) and is present in essentially all cases of chronic myelogenous leukemia (CML) and a cohort of patients with acute lymphocytic leukemia (ALL). 1,2 The translocation may result in p190, p210, and p230 BCR/ABL kinases, depending on the position of the bcr breakpoint. 3 BCR/ABL oncogenic tyrosine kinase exhibits 2 complementary roles in cancer. The first and best known is stimulation of signaling pathways that render cells independent of their environment. BCR/ABL allows cells to proliferate in the absence of growth factors, protects them from apoptosis in the absence of external survival factors, and promotes invasion and metastasis. 4 The second role of BCR/ABL in hematologic malignancies, which is only just beginning to be fully recognized, is that it can render cells resistant to genotoxic therapies. [5][6][7][8][9][10][11][12] Treatment of CML with chemotherapy leads to a transient reduction of the number of leukemic cells. 13 However, the leukemic cells usually recover more quickly than normal counterparts. Therefore, these therapies do not mediate durable remissions in patients with CML. In addition, high-dose chemotherapy treatment combined with stem cell transplantation does not eradicate the disease in most patients as indicated by the poor efficacy of syngeneic twin transplantation in CML. 14 Although inhibition of caspase 3-dependent apoptotic pathways and prolongation of G 2 /M cell cycle phase seem to play important roles in the survival of BCR/ABL leukemia cells after DNA damage, 5,8,11 our recent findings indica...