Quinazolinones as γ-secretase modulators

Synthesis, SAR and evaluation of styrenyl quinazolinones as novel gamma secretase modulators are presented in this communication. Starting from literature and in-house leads we evaluated a range of quinazolinones which showed good modulation of γ-secretase activity.     

Cortactin releases the brakes in actin- based motility by enhancing WASP-VCA detachment from Arp2/3 branches

Cortactin is involved in invadopodia and podosome formation [1], pathogens and endosome motility [2], and persistent lamellipodia protrusion [ [3] and [4] ]; its overexpression enhances cellular motility and metastatic activity [ [5] , [6] , [7] and [8] ]. Several mechanisms have been proposed to explain cortactin's role in Arp2/3-driven actin polymerization [ [9] and [10] ], yet its direct role in cell movement remains unclear. We use a biomimetic system to study the mechanism of cortactin-mediated regulation of actin-driven motility [11]. We tested the role of different cortactin variants that interact with Arp2/3 complex and actin filaments distinctively. We show that wild-type cortactin significantly enhances the bead velocity at low concentrations. Single filament experiments show that cortactin has no significant effect on actin polymerization and branch stability, whereas it strongly affects the branching rate driven by Wiskott-Aldrich syndrome protein (WASP)-VCA fragment and Arp2/3 complex. These results lead us to propose that cortactin plays a critical role in translating actin polymerization at a bead surface into motion, by releasing WASP-VCA from the new branching site. This enhanced release has two major effects: it increases the turnover rate of branching per WASP molecule, and it decreases the friction-like force caused by the binding of the moving surface with respect to the growing actin network.     

New concepts of IL-10-induced lung fibrosis: fibrocyte recruitment and M2 activation in a CCL2/CCR2 axis

IL-10 is most commonly recognized as an anti-inflammatory cytokine possessing immunosuppressive effects necessary for regulated resolution of proinflammation. However, its role in the development of fibrosis during inflammatory resolution has not been clear. Few prior studies have linked IL-10 with the inhibition of fibrosis principally on the basis of regulating inflammation thought to be driving fibroproliferation. In contrast, in a model of long-term overexpression of IL-10, we observed marked induction of lung fibrosis in mice. The total cell number retrieved by bronchoalveolar lavage (BAL) increased 10-fold in the IL-10 overexpression (IL-10 OE) mice, with significant infiltration of T and B lymphocytes and collagen-producing cells. The presence of increased fibrocytes, isolated from collagenase-digested lungs, was identified by flow cytometry using dual staining of CD45 and collagen 1. Quantitative PCR analysis on an array of chemokine/chemokine receptor genes showed that receptor CCR2 and its ligand, CCL2, were highly upregulated in IL-10 OE mice, suggesting that IL-10-induced fibrocyte recruitment was CCL2/CCR2 specific. Given the prior association of alternatively activated (M(2)) macrophages with development of fibrosis in other disease states, we also examined the effect of IL-10 OE on the M(2) macrophage axis. We observed significantly increased numbers of M(2) macrophages in both BAL and whole lung tissue from the IL-10 OE mice. Administration of rabbit anti-CCL2 antiserum to IL-10 OE mice for three consecutive weeks significantly decreased fibrosis as evidenced by lung hydroxyproline content, compared with mice that received preimmune rabbit serum. These results indicate that overexpression of IL-10 induces fibrosis, in part, by fibrocyte recruitment and M(2) macrophage activation, and likely in a CCL2/CCR2 axis.     

IFITM3 inhibits influenza A virus infection by preventing cytosolic entry

To replicate, viruses must gain access to the host cell's resources. Interferon (IFN) regulates the actions of a large complement of interferon effector genes (IEGs) that prevent viral replication. The interferon inducible transmembrane protein family members, IFITM1, 2 and 3, are IEGs required for inhibition of influenza A virus, dengue virus, and West Nile virus replication in vitro. Here we report that IFN prevents emergence of viral genomes from the endosomal pathway, and that IFITM3 is both necessary and sufficient for this function. Notably, viral pseudoparticles were inhibited from transferring their contents into the host cell cytosol by IFN, and IFITM3 was required and sufficient for this action. We further demonstrate that IFN expands Rab7 and LAMP1-containing structures, and that IFITM3 overexpression is sufficient for this phenotype. Moreover, IFITM3 partially resides in late endosomal and lysosomal structures, placing it in the path of invading viruses. Collectively our data are consistent with the prediction that viruses that fuse in the late endosomes or lysosomes are vulnerable to IFITM3's actions, while viruses that enter at the cell surface or in the early endosomes may avoid inhibition. Multiple viruses enter host cells through the late endocytic pathway, and many of these invaders are attenuated by IFN. Therefore these findings are likely to have significance for the intrinsic immune system's neutralization of a diverse array of threats.     

Zoonotic Emerging Infectious Disease in Selected Countries in Southeast Asia: Insights from Ecohealth

Most emerging diseases of humans originate in animals, and zoonotic emerging infectious diseases (EIDs) threaten human, animal, and environment health. We report on a scoping study to assess actors, linkages, priorities, and needs related to management of these diseases from the perspective of key stakeholders in three countries in Southeast Asia. A comprehensive interview guide was developed and in-depth interviews completed with 21 key stakeholders in Vietnam, Lao People’s Democratic Republic, and Cambodia. We found numerous relevant actors with a predominance of public sector and medical disciplines. More capacity weaknesses than strengths were reported, with risk analysis and research skills most lacking. Social network analysis of information flows showed policy-makers were regarded as mainly information recipients, research institutes as more information providers, and universities as both. Veterinary and livestock disciplines emerged as an important “boundary-spanning” organization with linkages to both human health and rural development. Avian influenza was regarded as the most important zoonotic EID, perhaps reflecting the priority-setting influence of actors outside the region. Stakeholders reported a high awareness of the ecological and socioeconomic drivers of disease emergence and a demand for disease prioritization, epidemiological skills, and economic and qualitative studies. Evaluated from an ecohealth perspective, human health is weakly integrated with socioeconomics, linkages to policy are stronger than to communities, participation occurs mainly at lower levels, and equity considerations are not fully considered. However, stakeholders have awareness of ecological and social determinants of health, and a basis exists on which transdisciplinarity, equity, and participation can be strengthened.     

A de novo protein binding pair by computational design and directed evolution

The de novo design of protein-protein interfaces is a stringent test of our understanding of the principles underlying protein-protein interactions and would enable unique approaches to biological and medical challenges. Here we describe a motif-based method to computationally design protein-protein complexes with native-like interface composition and interaction density. Using this method we designed a pair of proteins, Prb and Pdar, that heterodimerize with a Kd of 130 nM, 1000-fold tighter than any previously designed de novo protein-protein complex. Directed evolution identified two point mutations that improve affinity to 180 pM. Crystal structures of an affinity-matured complex reveal binding is entirely through the designed interface residues. Surprisingly, in the in vitro evolved complex one of the partners is rotated 180° relative to the original design model, yet still maintains the central computationally designed hotspot interaction and preserves the character of many peripheral interactions. This work demonstrates that high-affinity protein interfaces can be created by designing complementary interaction surfaces on two noninteracting partners and underscores remaining challenges.     

The effectiveness of traditional and sling exercise strength training in women

Strength training often combines closed-kinetic-chain exercises (CKCEs) and open kinetic-chain exercises (OKCEs). The CKCE may be more effective for improving performance in lower-body training. Recently, we reported upper-body CKCE (using a commercially available system of ropes and slings, Redcord AS, Staubo, Norway) was as effective as OKCE training for strength gains and that CKCE was more effective than OKCE for improving throwing performance. To our knowledge the effectiveness of a strength training program that uses exclusively CKCE is unknown. In this study, we examined the effectiveness of CKCE vs. OKCE strength training programs in women enrolled in an introductory strength training program. Twenty-six participants were randomized to OKCE (traditional exercises) or CKCE (sling-based exercises). Participants completed 6 sets per week for 13 weeks. Pre and posttraining evaluations included the following: 1 repetition maximum (1RM) leg and bench press; sling exercise push-ups; isokinetic dynamometry; lateral step-down test; and the Star Excursion Balance Test. Both groups significantly improved bench press (by an average of 4-6 kg) and leg press (by an average of 23-35 kg) (p < 0.001). There was a significant group × time interaction (p < 0.001) for sling exercise push-ups (OKCE pre = 5.5 ± 8.6, OKCE post = 6.1 ± 8.2, CKCE pre = 6.8 ± 6.0, CKCE post = 16.9 ± 6.6). Isokinetic measures of knee extension, knee flexion, shoulder internal rotation, and shoulder external rotation increased (improvements ranged from 2.7 to 27.7%), with no group differences. Both OKCE and CKCE strength training elicited similar changes in balance. We conclude that CKCE training is equally as effective as OKCE training during the initial phases of a strength training program in women. The fact that only CKCE improved sling exercise push-ups supports previous findings suggesting functional superiority of CKCE.     

Male mating competitiveness of a Wolbachia-introgressed Aedes polynesiensis strain under semi-field conditions

Background

Lymphatic filariasis (LF), a global public health problem affecting approximately 120 million people worldwide, is a leading cause of disability in the developing world including the South Pacific. Despite decades of ongoing mass drug administration (MDA) in the region, some island nations have not yet achieved the threshold levels of microfilaremia established by the World Health Organization for eliminating transmission. Previously, the generation of a novel Aedes polynesiensis strain (CP) infected with an exogenous type of Wolbachia has been described. The CP mosquito is cytoplasmically incompatible (i.e., effectively sterile) when mated with wildtype mosquitoes, and a strategy was proposed for the control of A. polynesiensis populations by repeated, inundative releases of CP males to disrupt fertility of wild females. Such a strategy could lead to suppression of the vector population and subsequently lead to a reduction in the transmission of filarial worms.

Methodology/Principal Findings

CP males and F1 male offspring from wild-caught A. polynesiensis females exhibit near equal mating competitiveness with F1 females under semi-field conditions.

Conclusions/Significance

While laboratory experiments are important, prior projects have demonstrated the need for additional testing under semi-field conditions in order to recognize problems before field implementation. The results reported here from semi-field experiments encourage forward progression toward small-scale field releases.     

Inexplicable Inefficiency of Avian Molt? Insights from an Opportunistically Breeding Arid-Zone Species,Lichenostomus penicillatus

The majority of bird species studied to date have molt schedules that are not concurrent with other energy demanding life history stages, an outcome assumed to arise from energetic trade-offs. Empirical studies reveal that molt is one of the most energetically demanding and perplexingly inefficient growth processes measured. Furthermore, small birds, which have the highest mass-specific basal metabolic rates (BMR_m), have the highest costs of molt per gram of feathers produced. However, many small passerines, including white-plumed honeyeaters (WPHE; Lichenostomus penicillatus), breed in response to resource availability at any time of year, and do so without interrupting their annual molt. We examined the energetic cost of molt in WPHE by quantifying weekly changes in minimum resting metabolic rate (RMR_min) during a natural-molt period in 7 wild-caught birds. We also measured the energetic cost of feather replacement in a second group of WPHEs that we forced to replace an additional 25% of their plumage at the start of their natural molt period. Energy expenditure during natural molt revealed an energy conversion efficiency of just 6.9% (±0.57) close to values reported for similar-sized birds from more predictable north-temperate environments. Maximum increases in RMR_min during the molt of WPHE, at 82% (±5.59) above individual pre-molt levels, were some of the highest yet reported. Yet RMR_min maxima during molt were not coincident with the peak period of feather replacement in naturally molting or plucked birds. Given the tight relationship between molt efficiency and mass-specific metabolic rate in all species studied to date, regardless of life-history pattern (Efficiency (%)  = 35.720•10^−0.494BMRm; r² = 0.944; p = <0.0001), there appears to be concomitant physiological costs entrained in the molt period that is not directly due to feather replacement. Despite these high total expenditures, the protracted molt period of WPHE significantly reduces these added costs on a daily basis.