Screening of the NIH Clinical Collection for inhibitors of HIV-1 integrase activity

Drug repurposing offers a validated approach to reduce drug attrition within the drug discovery and development pipeline through the application of known drugs and drug candidates to treat new indications. Full exploitation of this strategy necessitates the screening of a vast number of molecules against an extensive number of diseases of high burden or unmet need and the subsequent dissemination of the findings. In order to contribute to endeavours within this field, we screened the 727 compounds comprising the US National Institutes of Health (NIH) Clinical Collection through an HIV-1 (human immunodeficiency virus type 1) integrase stand transfer inhibition assay on an automated scintillation proximity assay platform. Only two compounds were identified within the initial screen, with cefixime trihydrate and epigallocatechin gallate found to reduce integrase strand transfer activity at IC50 values of 6.03±1.29 ?M and 9.57±1.62 ?M, respectively. However, both cefixime trihydrate and epigallocatechin gallate retained their low micromolar inhibitory activity when tested against a raltegravir-resistant integrase double mutant (FCIC50 values of 0.83 and 0.06, respectively), were ineffective in an orthogonal strand transfer ELISA (<30% inhibition at 100 ?M) and produced negligible selectivity index values (<1) in vitro. While no useful inhibitors of HIV-1 integrase strand transfer activity were found within the NIH Clinical Collection, the identification of two assay-disrupting molecules demonstrates the importance of consideration of non-specific inhibitors in drug repurposing screens. Significance: • This study is the first to screen the US NIH Clinical Collection for potential HIV-1 integrase inhibitors. • The pervasive nature of promiscuous inhibitors is emphasised

HIV-1 Phenotypic Reverse Transcriptase Inhibitor Drug Resistance Test Interpretation Is Not Dependent on the Subtype of the Virus Backbone

To date, the majority of HIV-1 phenotypic resistance testing has been performed with subtype B virus backbones (e.g. HXB2). However, the relevance of using this backbone to determine resistance in non-subtype B HIV-1 viruses still needs to be assessed. From 114 HIV-1 subtype C clinical samples (36 ARV-naïve, 78 ARV-exposed), pol amplicons were produced and analyzed for phenotypic resistance using both a subtype B- and C-backbone in which the pol fragment was deleted. Phenotypic resistance was assessed in resulting recombinant virus stocks (RVS) for a series of antiretroviral drugs (ARV's) and expressed as fold change (FC), yielding 1660 FC comparisons. These Antivirogram® derived FC values were categorized as having resistant or sensitive susceptibility based on biological cut-off values (BCOs). The concordance between resistance calls obtained for the same clinical sample but derived from two different backbones (i.e. B and C) accounted for 86.1% (1429/1660) of the FC comparisons. However, when taking the assay variability into account, 95.8% (1590/1660) of the phenotypic data could be considered as being concordant with respect to their resistance call. No difference in the capacity to detect resistance associated with M184V, K103N and V106M mutations was noted between the two backbones. The following was concluded: (i) A high level of concordance was shown between the two backbone phenotypic resistance profiles; (ii) Assay variability is largely responsible for discordant results (i.e. for FC values close to BCO); (iii) Confidence intervals should be given around the BCO's, when assessing resistance in HIV-1 subtype C; (iv) No systematic resistance under- or overcalling of subtype C amplicons in the B-backbone was observed; (v) Virus backbone subtype sequence variability outside the pol region does not contribute to phenotypic FC values. In conclusion the HXB2 virus backbone remains an acceptable vector for phenotyping HIV-1 subtype C pol amplicons

Phylodynamics of HIV-1 from a Phase-III AIDS Vaccine Trial in North America

In 2003, a phase III placebo-controlled trial (VAX004) of a candidate HIV-1 vaccine (AIDSVAX B/B) was completed in 5,403 volunteers at high risk for HIV-1 infection from North America and the Netherlands. A total of 368 individuals became infected with HIV-1 during the trial. The envelope glycoprotein gene (gp120) from the HIV-1 subtype B viruses infecting 349 patients was sequenced from clinical samples taken as close as possible to the time of diagnosis, rendering a final data set of 1,047 sequences (1,032 from North America and 15 from the Netherlands). Here, we used these data in combination with other sequences available in public databases to assess HIV-1 variation as a function of vaccination treatment, geographic region, race, risk behavior, and viral load. Viral samples did not show any phylogenetic structure for any of these factors, but individuals with different viral loads showed significant differences (P = 0.009) in genetic diversity. The estimated time of emergence of HIV-1 subtype B was 1966–1970. Despite the fact that the number of AIDS cases has decreased in North America since the early 90s, HIV-1 genetic diversity seems to have remained almost constant over time. This study represents one of the largest molecular epidemiologic surveys of viruses responsible for new HIV-1 infections in North America and could help the selection of epidemiologically representative vaccine antigens to include in the next generation of candidate HIV-1 vaccines

Beneficial effect of the oxygen free radical scavenger amifostine (WR-2721) on spinal cord ischemia/reperfusion injury in rabbits

<p>Abstract</p> <p>Background</p> <p>Paraplegia is the most devastating complication of thoracic or thoraco-abdominal aortic surgery. During these operations, an ischemia-reperfusion process is inevitable and the produced radical oxygen species cause severe oxidative stress for the spinal cord. In this study we examined the influence of Amifostine, a triphosphate free oxygen scavenger, on oxidative stress of spinal cord ischemia-reperfusion in rabbits.</p> <p>Methods</p> <p>Eighteen male, New Zealand white rabbits were anesthetized and spinal cord ischemia was induced by temporary occlusion of the descending thoracic aorta by a coronary artery balloon catheter, advanced through the femoral artery. The animals were randomly divided in 3 groups. Group I functioned as control. In group II the descending aorta was occluded for 30 minutes and then reperfused for 75 min. In group III, 500 mg Amifostine was infused into the distal aorta during the second half-time of ischemia period. At the end of reperfusion all animals were sacrificed and spinal cord specimens were examined for superoxide radicals by an ultra sensitive fluorescent assay.</p> <p>Results</p> <p>Superoxide radical levels ranged, in group I between 1.52 and 1.76 (1.64 ± 0.10), in group II between 1.96 and 2.50 (2.10 ± 0.23), and in group III (amifostine) between 1.21 and 1.60 (1.40 ± 0.19) (p = 0.00), showing a decrease of 43% in the Group of Amifostine. A lipid peroxidation marker measurement ranged, in group I between 0.278 and 0.305 (0.296 ± 0.013), in group II between 0.427 and 0.497 (0.463 ± 0.025), and in group III (amifostine) between 0.343 and 0.357 (0.350 ± 0.007) (p < 0.00), showing a decrease of 38% after Amifostine administration.</p> <p>Conclusion</p> <p>By direct and indirect methods of measuring the oxidative stress of spinal cord after ischemia/reperfusion, it is suggested that intra-aortic Amifostine infusion during spinal cord ischemia phase, significantly attenuated the spinal cord oxidative injury in rabbits.</p

Trauma networks: present and future challenges

In England, trauma is the leading cause of death across all age groups, with over 16,000 deaths per year. Major trauma implies the presence of multiple, serious injuries that could result in death or serious disability. Successive reports have documented the fact that the current ad hoc unstructured management of this patient group is associated with considerable avoidable death and disability. The reform of trauma care in England, especially of the severely injured patient, has already begun. Strong clinical leadership is embraced as the way forward. The present article summarises the steps that have been made over the last decade that led to the recent decision to move towards a long anticipated restructure of the National Health Service (NHS) trauma services with the introduction of Regional Trauma Networks (RTNs). While, for the first time, a genuine political will and support exists, the changes required to maintain the momentum for the implementation of the RTNs needs to be marshalled against arguments, myths and perceptions from the past. Such an approach may reverse the disinterest attitude of many, and will gradually evolve into a cultural shift of the public, clinicians and policymakers in the fullness of time

Burden of Illness and Quality of Life in Tuberous Sclerosis Complex: Findings From the TOSCA Study

Research on tuberous sclerosis complex (TSC) to date has focused mainly on the physical manifestations of the disease. In contrast, the psychosocial impact of TSC has received far less attention. The aim of this study was therefore to examine the impact of TSC on health, quality of life (QoL), and psychosocial well-being of individuals with TSC and their families. Questionnaires with disease-specific questions on burden of illness (BOI) and validated QoL questionnaires were used. After completion of additional informed consent, we included 143 individuals who participated in the TOSCA (TuberOus SClerosis registry to increase disease Awareness) study. Our results highlighted the substantial burden of TSC on the personal lives of individuals with TSC and their families. Nearly half of the patients experienced negative progress in their education or career due to TSC (42.1%), as well as many of their caregivers (17.6% employed; 58.8% unemployed). Most caregivers (76.5%) indicated that TSC affected family life, and social and working relationships. Further, well-coordinated care was lacking: a smooth transition from pediatric to adult care was mentioned by only 36.8% of adult patients, and financial, social, and psychological support in 21.1, 0, and 7.9%, respectively. In addition, the moderate rates of pain/discomfort (35%) and anxiety/depression (43.4%) reported across all ages and levels of disease demonstrate the high BOI and low QoL in this vulnerable population

Phylodynamics of HIV-1 from a Phase III AIDS Vaccine Trial in Bangkok, Thailand

In 2003, a phase III placebo-controlled trial (VAX003) was completed in Bangkok, Thailand. Of the 2,546 individuals enrolled in the trial based on high risk for infection through injection drug use (IDU), we obtained clinical samples and HIV-1 sequence data (envelope glycoprotein gene gp120) from 215 individuals who became infected during the trial. Here, we used these data in combination with other publicly available gp120 sequences to perform a molecular surveillance and phylodynamic analysis of HIV-1 in Thailand.Phylogenetic and population genetic estimators were used to assess HIV-1 gp120 diversity as a function of vaccination treatment, viral load (VL) and CD4(+) counts, to identify transmission clusters and to investigate the timescale and demographics of HIV-1 in Thailand. Three HIV-1 subtypes were identified: CRF01_AE (85% of the infections), subtype B (13%) and CRF15_AE (2%). The Bangkok IDU cohort showed more gp120 diversity than other Asian IDU cohorts and similar diversity to that observed in sexually infected individuals. Moreover, significant differences (P<0.02) in genetic diversity were observed in CRF01_AE IDU with different VL and CD4(+) counts. No phylogenetic structure was detected regarding any of the epidemiological and clinical factors tested, although high proportions (35% to 50%) of early infections fell into clusters, which suggests that transmission chains associated with acute infection play a key role on HIV-1 spread among IDU. CRF01_AE was estimated to have emerged in Thailand in 1984.5 (1983-1986), 3-6 years before the first recognition of symptomatic patients (1989). The relative genetic diversity of the HIV-1 population has remained high despite decreasing prevalence rates since the mid 1990s.Our study and recent epidemiological reports indicate that HIV-1 is still a major threat in Thailand and suggest that HIV awareness and prevention needs to be strengthened to avoid AIDS resurgence

Natural clusters of tuberous sclerosis complex (TSC)-associated neuropsychiatric disorders (TAND): new findings from the TOSCA TAND research project.

BACKGROUND: Tuberous sclerosis complex (TSC)-associated neuropsychiatric disorders (TAND) have unique, individual patterns that pose significant challenges for diagnosis, psycho-education, and intervention planning. A recent study suggested that it may be feasible to use TAND Checklist data and data-driven methods to generate natural TAND clusters. However, the study had a small sample size and data from only two countries. Here, we investigated the replicability of identifying natural TAND clusters from a larger and more diverse sample from the TOSCA study. METHODS: As part of the TOSCA international TSC registry study, this embedded research project collected TAND Checklist data from individuals with TSC. Correlation coefficients were calculated for TAND variables to generate a correlation matrix. Hierarchical cluster and factor analysis methods were used for data reduction and identification of natural TAND clusters. RESULTS: A total of 85 individuals with TSC (female:male, 40:45) from 7 countries were enrolled. Cluster analysis grouped the TAND variables into 6 clusters: a scholastic cluster (reading, writing, spelling, mathematics, visuo-spatial difficulties, disorientation), a hyperactive/impulsive cluster (hyperactivity, impulsivity, self-injurious behavior), a mood/anxiety cluster (anxiety, depressed mood, sleep difficulties, shyness), a neuropsychological cluster (attention/concentration difficulties, memory, attention, dual/multi-tasking, executive skills deficits), a dysregulated behavior cluster (mood swings, aggressive outbursts, temper tantrums), and an autism spectrum disorder (ASD)-like cluster (delayed language, poor eye contact, repetitive behaviors, unusual use of language, inflexibility, difficulties associated with eating). The natural clusters mapped reasonably well onto the six-factor solution generated. Comparison between cluster and factor solutions from this study and the earlier feasibility study showed significant similarity, particularly in cluster solutions. CONCLUSIONS: Results from this TOSCA research project in an independent international data set showed that the combination of cluster analysis and factor analysis may be able to identify clinically meaningful natural TAND clusters. Findings were remarkably similar to those identified in the earlier feasibility study, supporting the potential robustness of these natural TAND clusters. Further steps should include examination of larger samples, investigation of internal consistency, and evaluation of the robustness of the proposed natural clusters