Inhibitors of HIV-1 attachment. Part 7: Indole-7-carboxamides as potent and orally bioavailable antiviral agents

A series of substituted carboxamides at the indole C7 position of the previously described 4-fluoro-substituted indole HIV-1 attachment inhibitor 1 was synthesized and the SAR delineated. Heteroaryl carboxamide inhibitors that exhibited pM potency in the primary cell-based assay against a pseudotype virus expressing a JRFL envelope were identified. The simple methyl amide analog 4 displayed a promising in vitro profile, with its favorable HLM stability and membrane permeability translating into favorable pharmacokinetic properties in preclinical species.     

Structure-activity relationships of a novel series of melanin-concentrating hormone (MCH) receptor antagonists

A new series of 2-aminoquinolines has been identified as antagonists of the melanin concentrating hormone receptor (MCH-1R). Syntheses and structure-activity relationships are described leading to a compound having low nanomolar activity against the receptor and demonstrating functional antagonism. Studies also showed that some of the compounds were selective against a range of other G protein-coupled receptors.     

Quinazoline and benzimidazole MCH-1R antagonists

We have modified the previously reported 2-aminoquinoline 1 to provide two novel series of MCH-1R antagonists. Representative compounds from the quinazoline and benzimidazole series have been shown to be potent and selective, with promising in vitro eADME profiles.     

Estrogen withdrawal,increased breast cancer risk and the KRAS-variant

The KRAS-variant is a biologically functional, microRNA binding site variant, which predicts increased cancer risk especially for women. Because external exposures, such as chemotherapy, differentially impact the effect of this mutation, we evaluated the association of estrogen exposures, breast cancer (BC) risk and tumor biology in women with the KRAS-variant. Women with BC (n = 1712), the subset with the KRAS-variant (n = 286) and KRAS-variant unaffected controls (n = 80) were evaluated, and hormonal exposures, KRAS-variant status, and pathology were compared. The impact of estrogen withdrawal on transformation of isogenic normal breast cell lines with or without the KRAS-variant was studied. Finally, the association and presentation characteristics of the KRAS-variant and multiple primary breast cancer (MPBC) were evaluated. KRAS-variant BC patients were more likely to have ovarian removal pre-BC diagnosis than non-variant BC patients (p = 0.033). In addition, KRAS-variant BC patients also appeared to have a lower estrogen state than KRAS-variant unaffected controls, with a lower BMI (P < 0.001). Finally, hormone replacement therapy (HRT) discontinuation in KRAS-variant patients was associated with a diagnosis of triple negative BC (P < 0.001). Biologically confirming our clinical findings, acute estrogen withdrawal led to oncogenic transformation in KRAS-variant positive isogenic cell lines. Finally, KRAS-variant BC patients had greater than an 11-fold increased risk of presenting with MPBC compared to non-variant patients (45.39% vs 6.78%, OR 11.44 [3.42–37.87], P < 0.001). Thus, estrogen withdrawal and a low estrogen state appear to increase BC risk and to predict aggressive tumor biology in women with the KRAS-variant, who are also significantly more likely to present with multiple primary breast cancer.     

One Dose of Staphylococcus aureus 4C-Staph Vaccine Formulated with a Novel TLR7-Dependent Adjuvant Rapidly Protects Mice through Antibodies,Effector CD4+ T Cells,and IL-17A

A rapidly acting, single dose vaccine against Staphylococcus aureus would be highly beneficial for patients scheduled for major surgeries or in intensive care units. Here we show that one immunization with a multicomponent S. aureus candidate vaccine, 4C-Staph, formulated with a novel TLR7-dependent adjuvant, T7-alum, readily protected mice from death and from bacterial dissemination, both in kidney abscess and peritonitis models, outperforming alum-formulated vaccine. This increased efficacy was paralleled by higher vaccine-specific and α-hemolysin-neutralizing antibody titers and Th1/Th17 cell responses. Antibodies played a crucial protective role, as shown by the lack of protection of 4C-Staph/T7-alum vaccine in B-cell-deficient mice and by serum transfer experiments. Depletion of effector CD4⁺ T cells not only reduced survival but also increased S. aureus load in kidneys of mice immunized with 4C-Staph/T7-alum. The role of IL-17A in the control of bacterial dissemination in 4C-Staph/T7-alum vaccinated mice was indicated by in vivo neutralization experiments. We conclude that single dose 4C-Staph/T7-alum vaccine promptly and efficiently protected mice against S. aureus through the combined actions of antibodies, CD4⁺ effector T cells, and IL-17A. These data suggest that inclusion of an adjuvant that induces not only fast antibody responses but also IL-17-producing cell-mediated effector responses could efficaciously protect patients scheduled for major surgeries or in intensive care units.     

Clinical expression of cystic fibrosis in a large cohort of Italian siblings

Background

A clinical heterogeneity was reported in patients with Cystic Fibrosis (CF) with the same CFTR genotype and between siblings with CF.

Methods

We investigated all clinical aspects in a cohort of 101 pairs of siblings with CF (including 6 triplets) followed since diagnosis.

Results

Severe lung disease had a 22.2% concordance in sib-pairs, occurred early and the FEV₁% at 12?years was predictive of the severity of lung disease in the adulthood. Similarly, CF liver disease occurred early (median: 15?years) and showed a concordance of 27.8% in sib-pairs suggesting a scarce contribution of genetic factors; in fact, only 2/15 patients with liver disease in discordant sib-pairs had a deficiency of alpha-1-antitrypsin (a known modifier gene of CF liver phenotype). CF related diabetes was found in 22 pairs (in 6 in both the siblings). It occurred later (median: 32.5?years) and is strongly associated with liver disease. Colonization by P. aeruginosa and nasal polyposis that required surgery had a concordance >?50% in sib-pairs and were poorly correlated to other clinical parameters. The pancreatic status was highly concordant in pairs of siblings (i.e., 95.1%) but a different pancreatic status was observed in patients with the same CFTR mutations. This suggests a close relationship of the pancreatic status with the “whole” CFTR genotype, including mutations in regulatory regions that may modulate the levels of CFTR expression. Finally, a severe course of CF was evident in a number of patients with pancreatic sufficiency.

Conclusions

Physicians involved in care of patients with CF and in genetic counseling must be aware of the clinical heterogeneity of CF even in sib-pairs that, at the state of the art, is difficult to explain.

     

Genetic,clinical and radiographic signs in knee osteoarthritis susceptibility

Introduction

Osteoarthritis (OA) is considered to be a multifactorial and polygenic disease and diagnosis is mainly clinical and radiological. Correlation between radiographic data and clinical status has been reported. However, very few studies, especially in Caucasian people, describe the association between the Kellgren and Lawrence OA grading scale (KL) and genetic alterations to better understand OA etiopathogenesis and susceptibility. In order to update the knee OA grading, in this study we assessed the associations between KL grade, clinical features such as American Knee Society Score (AKSS), age, and polymorphisms in the principal osteoarthritis susceptibility (OS) genes in Sicilian individuals.

Methods

In 66 Sicilian individuals affected by primary knee OA, the clinical and radiographic evaluation was performed using 2 sub-scores of AKSS (knee score (KS) and function score (FS)) and KL. The patients were also classified according to age. Online Mendelian Inheritance in Man (OMIM) and Database of Single Nucleotide Polymorphisms (dbSNP) Short Genetic Variations databases were used to select gene regions containing the following polymorphisms to analyze: FRZB rs288326 and rs7775, MATN3 rs77245812, ASPN D14 repeats, PTHR2 rs76758470, GDF5 rs143383 and DVWA rs11718863. Patient genotypes were obtained using Sanger DNA sequencing analysis.

Results

In our cohort of patients a statistical association between the variables analyzed was reported in all associations tested (KL versus KS, FS and age). We observed that a mild to severe OA radiographic grade is related to severe clinical conditions and loss of articular function and that the severity of symptoms increases with age. Concerning the genotyping analysis, our results revealed a significant statistical association between KL grading and GDF5 rs143383 and DVWA rs11718863 genetic alterations. The latter was also associated with a more severe radiographic grade, displaying its predictive role as OA marker progression. Statistically significant association between clinical, radiographic and genetic signs observed, suggests extending the actual grading of knee OA based mainly on X-ray features.

Conclusions

This work represents a multidisciplinary and translational medicine approach to study OA where clinical, radiological, and OS5 and OS6 SNPs evaluation could contribute to better define grading and progression of OA and to the development of new therapies.     

Estimating lead time and overdiagnosis associated with PSA screening from prostate cancer incidence trends

Summary .   The introduction of the prostate-specific antigen (PSA) test has led to dramatic changes in the incidence of prostate cancer in the United States. In this article, we use information on the increase and subsequent decline in prostate cancer incidence following the adoption of PSA to estimate the lead time associated with PSA screening. The lead time is a key determinant of the likelihood of overdiagnosis, one of the main costs associated with the PSA test. Our approach conceptualizes observed incidence as the sum of the secular trend in incidence, which reflects incidence in the absence of PSA, and the excess incidence over and above the secular trend, which is a function of population screening patterns and the unknown lead time. We develop a likelihood model for the excess incidence given the secular trend and use it to estimate the mean lead time under specified distributional assumptions. We also develop a likelihood model for observed incidence and use it to simultaneously estimate the mean lead time together with a smooth secular trend. Variances and confidence intervals are estimated via a parametric bootstrap. Our results indicate an average lead time of approximately 4.59 years (95% confidence interval [3.24, 5.93]) for whites and 6.78 years [5.42, 8.20] for blacks with a corresponding secular trend estimate that is fairly flat after the introduction of PSA screening. These estimates correspond to overdiagnosis frequencies of approximately 22.7% and 34.4% for screen-detected whites and blacks, respectively. Our results provide the first glimpse of a plausible secular trend in prostate cancer incidence and suggest that, in the absence of PSA screening, disease incidence would not have continued its historic increase, rather it would have leveled off in accordance with changes in prostate patterns of care unrelated to PSA.     

Differential Expression and Network Inferences through Functional Data Modeling

Summary .  Time course microarray data consist of mRNA expression from a common set of genes collected at different time points. Such data are thought to reflect underlying biological processes developing over time. In this article, we propose a model that allows us to examine differential expression and gene network relationships using time course microarray data. We model each gene-expression profile as a random functional transformation of the scale, amplitude, and phase of a common curve. Inferences about the gene-specific amplitude parameters allow us to examine differential gene expression. Inferences about measures of functional similarity based on estimated time-transformation functions allow us to examine gene networks while accounting for features of the gene-expression profiles. We discuss applications to simulated data as well as to microarray data on prostate cancer progression.     

Modulation of the immune response by probiotic strains in a mouse model of gluten sensitivity

Probiotic strains play an important role in modulating activities in the gut-associated lymphoid tissue. Elucidation of the mechanisms that mediate probiotic-driven immunomodulation may facilitate their therapeutic application for specific immune-mediated diseases or for prophylaxis. In this study, we explored the effect of different Lactobacillus spp. and Bifidobacterium lactis in transgenic mice expressing the human DQ8 heterodimer, a HLA molecule linked to Celiac Disease (CD). In vitro analysis on immature bone marrow-derived dendritic cells (iBMDCs) showed that all strains up-regulated surface B7-2 (CD86), indicative of DC maturation, however, with different intensity. No strain induced appreciable levels of IL-10 or IL-12 in iBMDCs, whereas TNF-α expression was essentially elicited by Lactobacillus paracasei and Lactobacillus fermentum. Interestingly, these strains were found also to increase the antigen-specific TNF-α secretion in vivo, following co-administration of probiotic bacteria in mice mucosally immunized with the gluten component gliadin. Together these findings highlighted the ability of probiotics to exert strain-specific inductive rather than suppressive effects both on the innate and adaptive immunity in a mouse model of food antigen sensitivity.