Autologous Hematopoietic Stem Cell Transplantation for Autoimmune Diseases: From Mechanistic Insights to Biomarkers.

Phase I/II clinical trials of autologous hematopoietic stem cell transplantation (AHSCT) have led to increased safety and efficacy of this therapy for severe and refractory autoimmune diseases (AD). Recent phase III randomized studies have demonstrated that AHSCT induces long-term disease remission in most patients without any further immunosuppression, with superior efficacy when compared to conventional treatments. Immune monitoring studies have revealed the regeneration of a self-tolerant T and B cell repertoire, enhancement of immune regulatory mechanisms, and changes toward an anti-inflammatory milieu in patients that are responsive to AHSCT. However, some patients reactivate the disease after transplantation due to reasons not yet completely understood. This scenario emphasizes that additional specific immunological interventions are still required to improve or sustain therapeutic efficacy of AHSCT in patients with AD. Here, we critically review the current knowledge about the operating immune mechanisms or established mechanistic biomarkers of AHSCT for AD. In addition, we suggest recommendations for future immune monitoring studies and biobanking to allow discovery and development of biomarkers. In our view, AHSCT for AD has entered a new era and researchers of this field should work to identify robust predictive, prognostic, treatment-response biomarkers and to establish new guidelines for immune monitoring studies and combined therapeutic interventions to further improve the AHSCT protocols and their therapeutic efficacy.

In vitro evaluation of biomarkers of nephrotoxicity through gene expression using gentamicin.

Acute renal failure is one of the most frequent effects observed after taking medicine. Such situations have been tardily discovered, given that existing methods for assessing toxicity are not predictive. In this light, the present work evaluated the effects of gentamicin, a form of nephrotoxic drug, on HK-2 and HEK-293 cells. By using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and flow cytometry, both cells demonstrated that cytotoxicity occurs in a dose-dependent manner through the processes of apoptosis and cell necrosis. Gene expression analysis showed a relative increase of expression for genes related to cell processes and classic biomarkers, such as TP53, CASP3, CASP8, CASP9, ICAM-1, EXOC3, KIM-1, and CST3. A decrease in expression for genes BCL2L1 and EGF was observed. This study, therefore, indicates that, when the methods are used together, gene expression analysis is able to evaluate the nephrotoxic potential of a substance.

The role of PD-L1 expression as a predictive biomarker in advanced non-small-cell lung cancer: a network meta-analysis.

BACKGROUND: Tumor programmed death ligand one (PD-L1) expression has been studied in several trials in non-small-cell lung cancer. METHODS: We assessed the potential role of PD-L1 expression according to Cochrane Collaboration's Guidelines. RESULTS: 13 studies with 1979 patients were included. Among 915 PD-L1 negative patients this rate was 13% (RR 2.08; 95% CI: 1.49-2.91; p < 0.01). The response rate has increased concurrent to the PD-L1 expression (Pearson's correlation, r = 0.43). PD-L1 expression was also related to better 24-weeks progression-free rate (RR 0.79; 95% CI: 0.71-0.89) and a trend toward better 1-year overall survival rate (RR 0.96; 95% CI: 0.87-1.06). CONCLUSION: Taking this data in account, PD-L1 overexpression could not be currently considered a robust biomarker to tailor the immune checkpoint inhibitors treatment.

Urinary leukotriene and Bcl I polymorphism of glucocorticoid receptor gene in preschoolers with recurrent wheezing and high risk of asthma.

BACKGROUND: Urinary leukotriene (LTE4) is an important marker of airway inflammation presence. A relationship between single nucleotide polymorphism in the glucocorticoid receptor (GCR) gene promoter (Bcl I polymorphism), development of asthma and sensitivity to glucocorticoids has been hypothesised. OBJECTIVE: To explore the possible association between the Bcl I polymorphism and baseline levels of urinary LTE4 in preschoolers with recurrent wheezing episodes. We prospectively enrolled and classified 86 preschoolers based on the risk of developing asthma (by the Asthma Predictive Index [API]). METHODS: At admission standardised questionnaires for demographics and respiratory illness characteristics were completed. The Bcl I polymorphism of the GCR was determined by a PCR-RFLP assay from blood samples, and urinary leukotriene was assessed from urine samples by an enzyme immunoassay. RESULTS: We enrolled 86 preschoolers (46 with positive API and 40 with negative API). There were no statistical differences in demographic, respiratory illnesses and wheezing episodes characteristics between both groups. Also, the prevalence of Bcl I polymorphism was similar between positive vs. negative API groups (34.8% vs. 38.9% for homozygote GG, 56.5% vs. 52.8% for heterozygote GC, 8.7% vs. 8.3% for homozygote CC, respectively, p=0.94). However, urinary LTE4 (median [IQR]) was higher in preschoolers with positive than negative API (7.18 [5.57-8.96pg/ml] vs. 6.42 [3.96-8.07pg/ml], p=0.02, respectively). CONCLUSIONS: In our population, wheezing preschoolers with positive API exhibit higher levels of urinary LTE4 than those with negative API; but there were no differences in Bcl I polymorphism of the GCR.

Prospecção de bioativos farmacológicos em algas marinhas Rhodophyta e Heterokontophyta e avaliação de citotoxicidade / Prospection of pharmaceutical bioactive compounds in marine algae Rhodophyta and Heterokontophyta and cytotoxicity evaluation

Muitas drogas terapêuticas produzidas pela indústria farmacêutica são estruturas químicas isoladas de organismos encontrados na natureza ou moléculas baseadas nelas. Podem ser incluídas nesse grupo drogas isoladas de organismos marinhos, como corais, esponjas e algas marinhas, conhecidos como produtores de grandes quantidades de metabólitos secundários. Com base neste fato o presente estudo teve como objetivo realizar a prospecção de moléculas bioativas com propósito farmacológico, em extratos de algas marinhas vermelhas (Rhodophyta) e pardas (Heterokontophyta) coletadas no litoral brasileiro. A prospecção foi realizada por meio de avaliação de seus potenciais antioxidante, antibacteriano, antifúngico, anticancerígeno, e antiparasitário contra organismos causadores de leishmaniose e esquistossomose. Para as avaliações foram empregadas os extratos supercríticos de 5 espécies diferentes, sendo 2 pardas: Dictyota dichotoma e D. menstrualis e três vermelhas: Chondria littoralis, Spyridia hypnoides e Plocamium brasiliense. Os extratos foram avaliados quanto aos seus potenciais bioativos e os resultados mais promissores foram selecionados para as etapas seguintes do fracionamento. Em uma avaliação geral os extratos apresentaram bons resultados e representam uma potencial fonte de bioativos. Os extratos das espécies de D. dichotoma e D. menstrualis foram então submetidos a um procedimento de fracionamento bioguiado pela atividade esquistossomicida. Incorporou-se ainda um terceiro extrato de D. mertensii aos estudos e todas as etapas do fracionamento foram monitoradas por LC-MS. Comparando-se as massas detectadas em todas as frações que apresentaram atividade, para os 3 extratos, foi verificado que a substância de m/z 271,24 estava presente em todas elas, portanto os procedimentos de isolamento foram direcionados a esta molécula para a qual foi possível isolar 7 mg. Diferentemente do que era esperado a molécula quando avaliada isoladamente não apresentou atividade esquistossomicida, levando a hipótese de que a atividade seja decorrente de uma molécula diferente para cada espécie ou ainda que a mesma seja decorrente de uma interação com outras substâncias por um mecanismo de ação aditivo ou sinérgico. O trabalho avaliado de forma geral apresentou resultados promissores e representa um grande embasamento para servir como base para posteriores trabalho de fracionamento

Several therapeutic drugs manufactured by the pharmaceutical industry are chemical structures isolated from organisms that are found in nature or molecules based on that. May be included at this group drugs isolated from marine organisms, like corals, sponges and seaweeds, known as great secondary metabolites producers. Based on this facts the objective of the present study is to perform a prospection study to achieve bioactive molecules with pharmaceutical purposes, on extracts made from red (Rhodophyta) and brown (Heterokontophyta) seaweed collected in the Brazilian shore. The prospection studies was performed by means of evaluation of the antioxidant, antibacterial, antifungal, anticancer and antiparasitic (against Leishmania and Schistosoma) potential. In the evaluation were tested the supercritical extracts of 5 different species, including 2 brown species: Dictyota dichotoma and D. menstrualis and 3 red species: Chondria littoralis, Spyridia hypnoides and Plocamium brasiliense. The extracts were evaluated by their potential bioactive compounds and the most promising results were selected for the following fractionation steps. Overall the extracts have shown good results and may be represent a potential source of bioactive molecules. The extracts of both D. dichotoma and D. menstrualis were submitted to a bioguided fractionation process by their antischistosomal activities. It was still included a third extract from D. mertensii to the studies and every step was monitored by LC-MS techniques. Comparing the detected mass for each active fraction, it was observed the presence of a substance with m/z 271,24 in all of the extracts, so the isolating procedures were directed to obtain that specific molecule, which was obtained in a biomass of 7 mg. Differently than expected the molecule when evaluated isolated do not show the antischistosomal activity, leading to the hypothesis that the activity was related to different molecules for each species or even the observed effect is resulted by an interaction mechanism with another substances by an additive or synergist mechanism. The overall evaluation of the whole work show some promising results and it represent a great support for future fractionation works

Predictive role of RRM1 and BRCA1 mRNA expression on the clinical outcome of advanced non-small cell lung cancer.

This study aimed to evaluate the association between RRM1 and BRCA1 expressions and the therapeutic efficacy of platinum-based chemotherapy in non-small cell lung cancer patients in terms of their response and prognosis. In total, 377 patients agreed to participate in our study, and all of them received platinum-based combination chemotherapy between January 2008 and January 2009. The relative cDNA quantitation for RRM1 and BRCA1 was conducted using a fluorescence-based, real-time detection method, using ß-actin as a reference gene. The average age of the 377 patients was 64.6 years (range: 25.5-86.4 years), including 269 men and 108 women. Patients with high RRM1 expression benefited more from a platinum-containing regimen, and patients with high BRCA1 expression showed a high response rate to a platinum-containing regimen and reduced disease progression. Patients with high RRM1 expression were associated with a longer progression-free survival (PFS) and overall survival (OS) than those with low expression, and the hazard ratios (HRs) (95% confidence interval (CI)) were 0.67 (0.32-0.91) and 0.54 (0.30-0.95), respectively. Patients with high BRCA1 expression showed longer PFS and OS compared to those with low expression, and the HRs (95%CI) were 0.54 (0.30-0.95) and 0.62 (0.32-0.93), respectively. These results could be used in personalized chemotherapy decisions and to increase the response rate and prolonged survival, and could encourage exploration of the predictive value of other genes.

Influence of PCSK9 polymorphisms on plasma lipids and response to atorvastatin treatment in Brazilian subjects.

BACKGROUND: The proprotein convertase subtilisin/kexin type 9 (PCSK9) has a key role in the regulation of plasma low-density lipoprotein (LDL) cholesterol by enhancing the degradation of LDL receptor. Functional variants in PCSK9 have been associated with differences in plasma lipids and may contribute to the variability of the response to cholesterol-lowering drugs. OBJECTIVE: To investigate the influence of PCSK9 variants on plasma lipid profile and response to atorvastatin in Brazilian subjects. METHODS: PCSK9 E670G, I474V, and R46L single nucleotide polymorphisms (SNPs) and plasma lipids were evaluated in 163 hypercholesterolemics (HC) and 171 normolipidemics (NL). HC patients with indication for cholesterol-lowering drug therapy (n = 128) were treated with atorvastatin (10 mg/d/4 wk). PCSK9 SNPs were analyzed by real time polymerase chain reaction. RESULTS: Frequencies of the PCSK9 SNPs were similar between the HC and NL groups. Logistic regression analysis showed a trend of association between PCSK9 E670G and hypercholesterolemia after adjustment for covariates (P = .059). The 670G allele was associated with high basal levels of LDL cholesterol (P = .03) in HC patients using the extreme discordant phenotype method. No association tests were performed for R46L variant because of its very low frequency, whereas the I474V polymorphism and PCSK9 haplotypes were not related to hypercholesterolemia or variability on plasma lipids in both NL and HC groups (P > .05). LDL cholesterol reduction in response to atorvastatin was not influenced by PCSK9 genotypes or haplotypes. CONCLUSIONS: PCSK9 E670G polymorphism but not I474V contributes to the variability on plasma LDL cholesterol levels in hypercholesterolemic subjects. Both PCSK9 variants have no influence on cholesterol-lowering response to atorvastatin.

Dual role of IL-12 in the therapeutic efficacy or failure during combined PEG-Interferon-α2A and ribavirin therapy in patients with chronic hepatitis C.

Several efforts have been made to establish novel biomarkers with relevant predictive values to monitor HCV-infected patients under pegilated Interferon-α2A-(PEG-IFN-α2A)/ribavirin therapy. The aim of this study was to monitor the kinetics of HCV viral load, serum levels of pro-inflammatory/regulatory cytokines and leukocyte activation status before and after PEG-IFN-α2A/ribavirin therapy in 52 volunteers, including 12 chronic HCV patients and 40 controls. The HCV viral load, serum levels of cytokines (IL-8/IL-6/TNF-α/IL-12/IFN-γ/IL-4/IL-10) and the phenotype of peripheral blood leukocytes were evaluated before and after 4, 12 and 24 weeks following the PEG-IFN-α2A/ribavirin therapy. Our results demonstrated that sustained virological response-(SVR) is associated with early decrease in the viral load after 4 weeks of treatment. The presence of a modulated pro-inflammatory profile at baseline favors SVR, whereas a strong inflammatory response at baseline predisposes to therapeutic failure. Furthermore, a time-dependent increase on serum IL-12 levels in patients under treatment is critical to support the SVR, while the early predominance of IL-10 correlates to late virological relapse. On the other hand, a broad but unguided "cytokine storm" is observed in the non-responder HCV patients after 12 weeks of treatment. Corroborating these findings, monocyte/lymphocyte activation at baseline is associated with the non-responders to therapy whereas high CD8(+) T-cell numbers associate with SVR. All in all, these data suggest that the baseline pattern of serum pro-inflammatory/regulatory cytokines and the immunological activation status of chronic HCV patients undergoing PEG-IFN-α2A/ribavirin therapy are closely related with the therapeutic response.

Morphologic features do not influence response to trastuzumab-containing neoadjuvant chemotherapy in HER2-positive breast cancer.

Trastuzumab-containing neoadjuvant chemotherapy in patients with HER2-positive breast cancer is highly effective in reducing tumor volume and enables more patients to have breast-conserving surgery. The tumor hormone receptor level has been shown to significantly influence response to trastuzumab-containing neoadjuvant chemotherapy. In this study, we comprehensively evaluated various morphologic features and proliferative activity in 50 HER2-positive invasive breast carcinomas treated with trastuzumab-containing neoadjuvant chemotherapy to determine whether any of these features have the same predictive value as tumor hormone receptor content. The tumor proliferation activity as measured by Ki-67 and various morphologic parameters were compared between tumors that achieved >50% tumor volume reduction including pathologic complete response (pCR) and tumors that showed ≤50% tumor volume reduction. Thirty-seven cases (74%) showed >50% tumor volume reduction including 18 cases with pCR. Thirteen cases (26%) showed ≤50% tumor volume reduction. Neither morphologic variables nor Ki-67 labeling index were predictive of >50% tumor volume reduction. In contrast, hormone receptor status and semiquantitative H scores for hormone receptors were predictive of>50% tumor volume reduction. The mean estrogen and progesterone receptor H scores for tumors that showed >50% tumor volume reduction (including pCR) were 77 (median 10) and 31 (median 0), respectively, compared with 168 (median 200) and 79 (median 75) for cases that showed ≤50% tumor volume reduction. Semiquantitative scoring for hormone receptors provides useful information in terms of therapeutic response in HER2-positive tumors.

The antinociceptive and anti-inflammatory effects of the crude extract of Jatropha isabellei in a rat gout model.

ETHNOPHARMACOLOGICAL RELEVANCE: Jatropha isabellei Müll Arg. (Euphorbiaceae) is a medicinal plant that has been used in South American folk medicine for the treatment of arthritic diseases, particularly gout. AIM OF THE STUDY: This study was designed to verify the antinociceptive, anti-inflammatory and hypouricemic potential of Jatropha isabellei. MATERIALS AND METHODS: Rats were orally administered with the crude extract (100-300 mg/kg) or a fraction that is rich in alkaloids (0.15 mg/kg) of Jatropha isabellei. An intra-articular (i.a.) injection of 50 µl of monosodium urate (MSU) crystals (1.25mg/site) was used to generate the gout model to assess the effect of the treatment on nociception (thermal and mechanical hyperalgesia) and inflammation (oedema and neutrophil infiltration). The effect of Jatropha isabellei on the serum levels of uric acid was evaluated in a model of hyperuricaemia induced by the intraperitoneal injection of potassium oxonate (250 mg/kg). The side effects were analysed using an open-field test, gastric lesion assessment and by measuring the levels of the ALT and AST enzymes. RESULTS: Our study demonstrated that the crude extract of Jatropha isabellei and a fraction rich in alkaloids were able to prevent the thermal hyperalgesia, mechanical allodynia, oedema and neutrophil infiltration induced by intra-articular MSU injection in rats. On the other hand, treatment with Jatropha isabellei did not alter the uric acid levels increased by potassium oxonate in the hyperuricaemia model. In addition, Jatropha isabellei did not induce gastric lesions or liver damage and did not alter spontaneous locomotor activity. CONCLUSION: The crude extract of Jatropha isabellei and its fraction rich in alkaloid presents antinociceptive and anti-inflammatory effects in a rat gout model, similar to that observed after treatment with colchicine, supporting the traditional use of this plant in gouty patients.

Determinação de biomarcadores cardíacos em gatos Maine Coon geneticamente testados para a mutação da cardiomiopatia hipertrófica / Cardiac biomarkers in Maine Coon cats genetically tested for hypertrophic cardiomyopathy

A cardiomiopatia hipertrófica (CMH) é a cardiopatia mais diagnosticada em felinos e responsável por morbidade e mortalidade elevadas. A desorganização do sarcômero no miocárdio de gatos afetados com a CMH tem relação com a mutação do gene miosina ligado à proteína C (MYBPC3). A concentração plasmática de biomarcadores cardíacos como o aminoterminal peptídeo natriurético atrial (NTproANP) e o aminoterminal peptídeo natriurético tipo B (NT-proBNP) liberados, respectivamente, secundária ao estresse da parede miocárdica dos átrios e ventrículos; a troponina I cardíaca (cTnI), liberada secundariamente à lesão miocárdica; e a endotelina tipo 1 (ET-1), um peptídeo vasoconstrictor potente cuja concentração plasmática aumenta em pacientes com insuficiência cardíaca, tem incrementado o diagnóstico de cardiopatias em humanos. A CMH é diagnosticada pela ecocardiografia convencional pela evidenciação de hipertrofia cardíaca (HC) segmentar ou difusa. Este estudo utilizou 57 gatos da raça Maine Coon, testados geneticamente para a mutação (M), que foram separados em quatro grupos: GIA com M e com HC (n= 4); GIB com M e sem HC (n= 10); GIIA sem M e com HC (n= 5); GIIB sem M e sem HC (n= 38) e avaliados por meio de ecocardiografia convencional e determinação dos biomarcadores cardíacos NT-proANP, NTproBNP, cTnI e ET-1. A prevalência da mutação nos gatos estudados foi de 24,56%. Diferenças estatísticas significantes foram observadas nos valores de NT-proBNP entre os grupos GIA e GIIB e GIA e GIB; de cTnI entre GIA e GIIB. Quando considerado apenas a presença ou ausência da mutação ou da hipertrofia, foram encontrados valores maiores de NT-proBNP em animais com HC e de cTnI em animais com mutação. Com base na metodologia utilizada, estabeleceu-se um ponto de corte para o NT-proBNP, considerando a presença de hipertrofia de 189,9 pmol/L, com sensibilidade de 77,8%, especificidade de 81,3%, valor preditivo positivo de 43,8% e valor preditivo negativo de 95,1 para o NT-proBNP, e o outro ponto de corte de 0,015 ng/mL, com sensibilidade de 64,3%, especificidade de 81,4%, valor preditivo positivo de 52,9% e valor preditivo negativo de 87,5% para a cTnI. A perspectiva para novos estudos concerne à cTnI e sua relação com a presença da mutação MYBPC3

Hypertrophic cardiomyopathy (HCM) is the most common feline heart disease and is responsible for high morbidity and mortality rates. Sarcomere disorganization in the affected myocardium of cats with HCM is related to the myosin binding protein C (MYBPC3) gene mutation. The plasma concentration of cardiac biomarkers such as atrial aminoterminal natriuretic peptide (NT-proANP) and Type B aminoterminal natriuretic peptide (NT-proBNP) released, respectively, by atrial and ventricular myocardium secondary to wall stress; cardiac troponin I (cTnI), released secondary to myocardial injury; and type 1 endothelin (ET-1), a potent vasoconstrictor peptide have been increasingly used for evaluation of human heart disease which are increased in patients with heart failure (HF). HCM is diagnosed by the presence of segmental or diffuse cardiac hypertrophy (CH) through conventional echocardiography. This study enrolled 57 Maine Coon cats screened for mutation (M) that were assigned to four groups: GIA with M and with CH (n= 4); GIB with M and without CH (n= 10); GIIA without M and with CH (n= 5); GIIB without M and without CH (n= 38) and evaluated by conventional echocardiography and cardiac biomarkers NT-proANP, NT-proBNP, cTnI and ET-1 measurements. The prevalence of the mutation in this study was 24.56%. Statistically significantly differences were observed in NT-proBNP among GIA and GIIB groups and among GIA and GIB groups; and in cTnI between GIA and GIIB groups. When considering only mutation and CH presence or absence, higher values of NT-proBNP were found in CH cats, and higher values of cTnI in those with mutation. Based on proposed methodology, cut-off value to NT-proBNP considering CH presence of 189.9 pmol/L had a sensitivity of 77.8%, specificity of 81.3%, predictive positive value of 43,8% and predictive negative value of 95,1% and the cut-off value of 0.015 ng/mL for cTnI considering mutation presence had a sensitivity of 64.3%, specificity of 81.4%, predictive positive value of 52,9% and predictive negative value of 87,5%. This study opened new perspectives to studies related to cTnI and MYBPC3 mutation

Determinação de produtos de biotransformação de aflotoxina B1 e aplicabilidade na avaliação da eficiência de um adsorvente à base de aluminossilicato de cálcio e sódio hidratado em frangos de corte / Determination of aflatoxin B1 biotransformation products and applicability for the efficacy evaluation of a hydrated sodium calcium aluminosilicate-based adsorbent in broiler chicks

O objetivo do presente trabalho foi determinar resíduos de aflatoxinas M1 (AFM1), aflatoxicol (AFL), B1 (AFB1), B2 (AFB2), G1 (AFG1) e G2 (AFG2) não metabolizadas no fígado e rim de frangos de corte, com a finalidade de verificar sua aplicabilidade na avaliação da eficiência de um adsorvente comercial à base de aluminossilicato de cálcio e sódio (HSCAS) incorporado na dieta, bem como determinar o percentual de ligação do adsorvente com a AFB1em ensaios in vitro. Cem frangos de corte (Ross 708), machos, de 1 dia de idade, foram mantidos em baterias metálicas, com acesso ad libitum à ração e água. Utilizou-se um delineamento inteiramente casualizado com 4 tratamentos, sendo cada tratamento composto por 5 gaiolas contendo 5 frangos em cada uma. Os tratamentos foram os seguintes: A) dieta basal (DB), sem adição de HSCAS ou AFB1; B) DB com adição de 0,5% de HSCAS; C) DB com adição de 2,5 mg/kg de AFB1; e D) DB com adição de 2,5 mg/kg de AFB1 e 0,5% de HSCAS. As rações experimentais foram administradas de 1 a 21 dias de vida. No dia 21, 5 frangos de cada tratamento foram insensibilizados com dióxido de carbono, mortos por deslocamento cervical e amostras de fígado e rim foram coletadas para análise de resíduos de AFB1. O HSCAS foi efetivo em se ligar, in vitro, à AFB1, cujos percentuais de ligação variaram de 8,8 a 99,5%, para concentrações do adsorvente entre 0,05 e 100 mg/10 mL.

The objective of the study was to determine residues of aflatoxin M1 (AFM1), aflatoxicol (AFL), B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2) non-metabolized in liver and kidney of broiler chicks, aiming to verify its applicability for evaluation of a commercial adsorbent based-HSCAS efficacy incorporated into the diet, aswell as to determine the binding capacity of a HSCAS for AFB1 in an in vitro testing. One hundred day-old male broilers (Ross 708) were maintained in chick batteries and allowed libitum access to feed and water. A completely randomized design was used with 5 replicate pens of5 chicks assigned to each of 4 dietary treatments from hatch to 21 days. Dietary treatments included: A) basal diet (BD), with no HSCAS or AFB1; B) BD supplemented with 0.5% HSCAS only; C) BD supplemented with 2.5 mg AFB1/kg of feed; and D) BD supplemented with 2.5 mg AFB1/kg of feed and 0.5% HSCAS. On day 21, 5 chicks from each treatment wereinsensibilized with carbon dioxide, killed by cervical dislocation and samples of liverand kidney collected for analysis of AFB1 residues. The HSCAS was effective to binding AFB1 in vitro, with percentage of AFB1 bound varying from 8.8 to 99.5% for adsorbent concentrations between 0.05 and 100 mg/10mL.

Topoisomerase 2 alpha: a real predictor of anthracycline efficacy?

Anthracyclines are frequently used in the adjuvant setting for breast cancer treatment since it is considered that anthracycline-based chemotherapy treatment benefits breast cancer patients. Nonetheless, these drugs are associated with severe side effects and predictive factors, for sensitivity to anthracyclines, are warranted in clinical practice. Topoisomerase 2 alpha (TOP2A) is considered to be the molecular target of these drugs. The potential predictive value of TOP2A amplification and overexpression has been extensively studied in breast cancer patients treated with anthracyclines. However, results are not conclusive. In this paper, we review some of the published studies addressing the predictive value of TOP2A as well as the cellular functions of this enzyme and its status in breast cancer tissue.

Association of XRCC3 and XPD751 SNP with efficacy of platinum-based chemotherapy in advanced NSCLC patients.

INTRODUCTION: The aim of this study was to investigate whe- ther X-ray repair cross-complementing group 3 (XRCC3) and xeroderma pigmentosum group D (XPD) single nucleotide polymorphism (SNP) affects the outcome of platinum- based chemotherapy in advanced non-small-cell lung cancer (NSCLC) patients. MATERIAL AND METHODS: From September 2005 to June 2009, 355 advanced-stage NSCLC patients were accrued to compare the SNP variants with the outcome of platinum-based chemotherapy. TaqMan RT-PCR was used to evaluate the SNPs of the XRCC3 codon241 (Thr/Met) and XPD codon751 (Lys/Gln) DNA repair genes. RESULTS: There was a lack of association between the studied SNPs and the response rate, progression-free survival or overall survival (OS) in the whole population. However, subgroup analysis revealed that XRCC3 241 Thr/Met or Met/Met allele was associated with marginally significantly longer OS than XRCC3 Thr/Thr allele (19.0 m vs. 12.5 m, p=0.081) in the patients treated with gemcitabine/ cisplatin or carboplatin (GP/GC) while it was correlated with a significantly shorter OS in the patients treated with non GP/GC (9.3 m vs. 16 m, p=0.003). XPD751 Lys/Lys had an association with statistical significantly longer OS than XPD751 Lys/Gln or Gln/Gln in the subgroup of elderly patients (14.10 m vs. 10 m, p=0.019) and patients with non-adenocarcinoma (12.6 m vs. 9 m, p=0.042). CONCLUSIONS: XRCC3 Thr241Met played an opposite role in predicting prognosis of chemotherapy in NSCLC patients according to the first-line regimens. XPD 751 Lys/ Lys might be a better prognostic marker of elderly or noncarcinoma NSCLC subgroup treated with platinum-based chemotherapy.

Evaluación de marcadores moleculares asociados con resistencia a gota (Phytophthora infestans L) en papas diploides y tetraploides / Evaluation of molecular markers associated with resistance to late blight (Phytophthora infestans L) in diploid and tetraploid potatoes

La papa, cultivo de importancia a nivel mundial es gravemente afectado por gota, enfermedad ocasionada por el oomycete Phytophthora infestans. Actualmente la forma más efectiva para combatir la enfermedad es mediante el desarrollo de cultivares resistentes al patógeno. Para esto, una estrategia es identificar genes que confieran resistencia al patógeno, para lo cual se buscan marcadores asociados con el carácter de resistencia. En este estudio se evaluaron marcadores moleculares tipo SCAR (Sequence Characterized Amplified Region): CosA, GP179, BA47f2 y Prp1 asociados con resistencia a P. infestans y el gen de resistencia R1, en 22 cultivares tetraploides pertenecientes a la subespecie andigena y cinco especies silvestres. Se evaluó el polimorfismo y se determinó si los alelos polimórficos permitían diferenciar genotipos resistentes de susceptibles. Se comparó el tamaño de los fragmentos obtenidos con los fragmentos esperados asociados con resistencia de acuerdo a reportes. El análisis se realizó considerando presencia/ausencia de los fragmentos: CosA210, CosA250, R11400, R11800, BA47f2500, GP179570, Prp1300, Prp1600, y Prp1900. Los resultados indicaron que en los cultivares tetraploides y silvestres, se presentaron polimorfismos en todos los marcadores evaluados, con excepción del marcador GP179. No se encontró correlación entre el rasgo de resistencia y los alelos. Los resultados de este estudio muestran que hay repuesta diferencial a los marcadores entre las subsp. tuberosum y subsp. Andigena.

Potato is an important worldwide crop seriously affected by late blight disease caused by the oomycete Phytophthora infestans. Currently, the most effective way to control the disease is developing resistant cultivars to the pathogen by identifying genes that confer resistance to the pathogen. For this purpose it is important to find molecular markers associated with the trait. In this study, the SCAR (Sequence Characterized Amplified Region) markers: CosA, GP179, BA47f2 y Prp1, associated with late blight and the resistant gen R1 were evaluated in 22 tetraploid cultivars from subspecie andigena and five wild potato species. Polymorphism was evaluated and it was evaluated if polymorphic alleles allow differentiating resistant from susceptible genotypes. The fragment length for each marker was compared to the allele size reported associated to resistance. The analysis considered the presence/absence of the fragments: CosA210, CosA250, R11400, R11800, BA47f2500, GP179570, Prp1300, Prp1600 and Prp1900. The results indicated that both, tetraploid cultivars and wild potatoes, showed polymorphisms with all these markers, except with the GP179 marker. It was not found correlation between resistance and the presence of specific alleles. Evidence found in this study indicates that results obtained with molecular markers differed between subsp. tuberosum and subsp. andigena.

Serum and salivary IgE, IgA, and IgG4 antibodies to Dermatophagoides pteronyssinus and its major allergens, Der p1 and Der p2, in allergic and nonallergic children.

Allergic rhinitis (AR) is a public health problem with high prevalence worldwide. We evaluated levels of specific IgE, IgA, and IgG4 antibodies to the Dermatophagoides pteronyssinus (Dpt) house dust mite and to its major allergens (Der p1 and Der p2) in serum and saliva samples from allergic and nonallergic children. A total of 86 children were analyzed, from which 72 had AR and 14 were nonallergic healthy children. Serum IgE and serum/salivary IgG4 levels to Dpt, Der p1, and Der p2 were higher in allergic children whereas serum/salivary IgA levels to all allergens were higher in nonallergic children. IgE levels positively correlated with IgG4 and IgA to all allergens in allergic children, while IgA levels negatively correlated with IgG4 to Dpt and Der p1 in nonallergic children. In conclusion, mite-specific IgA antibodies predominate in the serum and saliva of nonallergic children whereas mite-specific IgE and IgG4 are prevalent in allergic children. The presence of specific IgA appears to have a key role for the healthy immune response to mucosal allergens. Also, specific IgA measurements in serum and/or saliva may be useful for monitoring activation of tolerance-inducing mechanisms during allergen specific immunotherapeutic procedures, especially sublingual immunotherapy.

Survivin as a predictive biomarker of complete pathologic response to neoadjuvant chemotherapy in patients with stage II and stage III breast cancer.

INTRODUCTION: Predictive biomarkers of response to neoadjuvant chemotherapy in patients with breast cancer are needed to better characterize tumors and enable more tailored therapies. METHODS: The expression levels of survivin, BCL-2, cyclin D1, ETS1, and PDEF in tumor samples obtained in the diagnostic biopsies of patients undergoing neoadjuvant chemotherapy for stage II and stage III disease were evaluated by immunohistochemistry (IHC). The mean expression score (range, 0-15) obtained by 3 different pathologists was used for analysis and correlated with complete pathologic response (pCR) and survival by standard univariate and multivariate methods. RESULTS: Forty-five female patients were included in this study and received preoperative standard anthracycline/taxane-based chemotherapy. The median age at diagnosis was 49 years (range, 25-70 years). Three patients (7.1%) achieved pCR. The mean expression score of survivin in the diagnostic biopsies was significantly higher (P = .01) in patients with pCR (9.3) than in those without (3.4). There was no significant association with pCR for the other biologic markers analyzed nor was there correlation with prognosis. Survivin levels were not associated with age, tumor grade, clinical stage, or receptor status. CONCLUSION: High expression levels of survivin in the primary tumor may be used as a potential predictive biomarker of pCR to neoadjuvant chemotherapy in patients with stage II and stage III breast cancer.

Expression of p53, Ki-67, and CD31 proteins in endometrial polyps of postmenopausal women treated with tamoxifen.

This study was undertaken to investigate the expression of p53, Ki-67, and CD31 proteins in endometrial polyps of postmenopausal women treated with tamoxifen (TAM). Postmenopausal women with endometrial polyps treated with TAM (n = 20), postmenopausal women with endometrial polyps without hormone use (n = 20), postmenopausal women with atrophic endometrium (n = 20), and postmenopausal women with endometrial adenocarcinoma (n = 20) were prospectively investigated. Tissue samples were immunohistochemically evaluated by monoclonal antibodies for p53, Ki-67, and CD31. The data were analyzed using the Student t test, analysis of variance, and χ2 to evaluate significant differences between the groups. The level of significance was set at P < 0.05. There was no difference in the expression of p53 between the groups (P = 0.067). The expression of Ki-67 was higher in the polyp samples from TAM-treated women compared with those from the women using no hormone (P = 0.0047) and those from the women with atrophic endometrium (P = 0.008). Samples from the women with endometrial cancer was associated with higher Ki-67 expression compared with the polyp samples from TAM-treated women (P = 0.004). The expression of CD31 was higher in the polyp samples of TAM-treated women compared with that of the samples from the women with atrophic endometrium (P < 0.001) and similar to the polyp samples from the women using no hormone (P = 0.319) and to the samples from the women with endometrial cancer (P = 0.418). The use of TAM in postmenopausal women might be associated with increased cellular proliferation in endometrial polyps without interfering angiogenesis or inactivation of tumor suppressor proteins.

QT interval prolongation: preclinical and clinical testing arrhythmogenesis in drugs and regulatory implications.

The assessment about the proarrhythmic risk associated with a particular drug is a major requirement for drugs under development, since many drugs have been withdrawn from market or got under strict pharmacological vigilance because of such a risk. Predicting the development of a life-threatening arrhythmia is a hard task but, in the case of TdP ("Torsades de Pointes"), there are some useful markers. Among them, the prolongation of the QT interval and its heart rate correction (QTc) are the most remarkable. Actually, QT prolongation is considered the surrogate marker of TdP from the clinical and regulatory standpoint. ICH E14 provides recommendations to sponsors concerning the design, conduct, analysis, and interpretation of clinical studies to assess the potential of a drug to delay cardiac repolarization. The regulatory information about preclinical safety evaluation is contained in ICH S7B. Both guidelines have been a matter of intense debate. False negative and false positive results have been found within the preclinical and clinical field. There still are grey areas in which further research would be necessary. Improvement of tools that may contribute to complement the data from the human ether-a-go-go-related gene HERG channel and QT/QTc studies, such as concentration-QT relationship (CQT) studies and other innovative techniques, will be more than welcome.