Sexual Behavior of Older Adults Living with HIV in Uganda.

Sexual behavior among older adults with HIV in Sub-Saharan Africa has been understudied despite the burgeoning of this population. We examined sexual behavior among older adults living with HIV in Uganda. Participants were eligible for the study if they were 50 years of age or older and living with HIV. Quantitative data were collected through face-to-face interviews, including demographic characteristics, health, sexual behavior and function, and mental health. Of respondents, 42 were men and 59 women. More than one-quarter of these HIV-positive older adults were sexually active. A greater proportion of older HIV-positive men reported being sexually active compared to women (54 vs. 15%). Among those who are sexually active, a majority never use condoms. Sixty-one percent of men regarded sex as at least somewhat important (42%), while few women shared this opinion (20%). Multivariate logistic regression analyses revealed that odds of sexual activity in the past year were significantly increased by the availability of a partner (married/cohabitating), better physical functioning, and male gender. As more adults live longer with HIV, it is critical to understand their sexual behavior and related psychosocial variables in order to improve prevention efforts

Allosteric ligands for the pharmacologically dark receptors GPR68 and GPR65

At least 120 non-olfactory G protein-coupled receptors in the human genome are ”orphans” for which endogenous ligands are unknown, and many have no selective ligands, hindering elucidation of their biological functions and clinical relevance. Among these is GPR68, a proton receptor that lacks small molecule modulators for probing its biology. Yeast-based screens against GPR68 identified the benzodiazepine drug lorazepam as a non-selective GPR68 positive allosteric modulator. Over 3000 GPR68 homology models were refined to recognize lorazepam in a putative allosteric site. Docking 3.1 million molecules predicted new GPR68 modulators many of which were confirmed in functional assays. One potent GPR68 modulator—ogerin– suppressed recall in fear conditioning in wild-type, but not in GPR68 knockout mice. The same approach led to the discovery of allosteric agonists and negative allosteric modulators for GPR65. Combining physical and structure-based screening may be broadly useful for ligand discovery for understudied and orphan GPCRs

In silico design of novel probes for the atypical opioid receptor MRGPRX2

The primate-exclusive MRGPRX2 G protein-coupled receptor (GPCR) has been suggested to modulate pain and itch. Despite putative peptide and small molecule MRGPRX2 agonists, selective nanomolar potency probes have not yet been reported. To identify a MRGPRX2 probe, we first screened 5,695 small molecules and found many opioid compounds activated MRGPRX2, including (−)- and (+)-morphine, hydrocodone, sinomenine, dextromethorphan and the prodynorphin-derived peptides, dynorphin A, dynorphin B, and α- and β-neoendorphin. We used these to select for mutagenesis-validated homology models and docked almost 4 million small molecules. From this docking, we predicted ZINC-3573, which represents a potent MRGPRX2-selective agonist, showing little activity against 315 other GPCRs and 97 representative kinases, and an essentially inactive enantiomer. ZINC-3573 activates endogenous MRGPRX2 in a human mast cell line inducing degranulation and calcium release. MRGPRX2 is a unique atypical opioid-like receptor important for modulating mast cell degranulation, which can now be specifically modulated with ZINC-3573

Shooting in the dark: Strategies for discovering magic bullets and magic shotguns for orphan GPCRs

It is now widely accepted that drugs achieve efficacy via interactions with multiple targets simultaneously, a serendipitous biological symphony that is typically only deconvoluted retrospectively. While pan-profiling of these molecules has discovered varying activity against different proteins, efforts to recapitulate therapeutic effects through focused, specific interactions with singular targets have largely failed. This is especially true for G protein-coupled receptors (GPCRs), whose conserved topologies have introduced widespread degeneracy among their ligands. Yet, 38% of GPCRs are orphans, with no elucidated physiological function or no illuminating modulatory chemical matter. First, we use a robust yeast-screening platform to show that even in a small library of drugs, the NIH Clinical Collection, orphan GPCRs are potent targets for a variety of drugs. We focus on the benzodiazepine lorazepam and show that is also a strong positive allosteric modulator (PAM) of the pH-sensing orphan, GPR68. We generated over 3,000 receptor homology models for virtual screening campaigns that, among other hits, identified ogerin, a PAM that selectively and potently potentiated GPR68-Gs signaling (αβ= 27.5). Ogerin was prioritized to interrogate the in vivo function of GPR68, revealing that receptor activation suppressed recall in fear conditioning in wild-type, but not in GPR68 knockout, mice. Application of the same approach to a second orphan receptor, GPR65, led to the discovery of specific allosteric agonists and negative allosteric modulators. Second, we pick subsets of GPCRs to demonstrate that structure-based virtual screening can be used to directly identify molecules with a desired receptor binding profile. Although only a small number of hits were selective over the anti-target, a result observed in two parallel studies between three aminergic receptors and 2 opioid receptor subtypes, this performance is still better than that of ligand-based methods. Inexhaustive and incorrect treatment of receptor flexibility was subsequently deemed to be the culprit, a potentially receptor-specific nuisance that can be leveraged. At its core, this thesis describes test cases and methods that would ultimately encompass the machinery for a pipeline to generate molecules with a complete, prescribed GPCR polypharmacology

Teach-Discover-Treat 2014, Part 2; Molecular Docking with DOCK 3.7

<p>Tutorial for TDT2014, part 2. Finding potential anti-malarial compounds with molecular docking. For more info see https://github.com/ryancoleman/tdt2014-part2</p

Quantification of the transferability of a designed protein specificity switch reveals extensive epistasis in molecular recognition.

Reengineering protein-protein recognition is an important route to dissecting and controlling complex interaction networks. Experimental approaches have used the strategy of second-site suppressors, where a functional interaction is inferred between two proteins if a mutation in one protein can be compensated by a mutation in the second. Mimicking this strategy, computational design has been applied successfully to change protein recognition specificity by predicting such sets of compensatory mutations in protein-protein interfaces. To extend this approach, it would be advantageous to be able to transplant existing engineered and experimentally validated specificity changes to other homologous protein-protein complexes. Here, we test this strategy by designing a pair of mutations that modulates peptide recognition specificity in the Syntrophin PDZ domain, confirming the designed interaction biochemically and structurally, and then transplanting the mutations into the context of five related PDZ domain-peptide complexes. We find a wide range of energetic effects of identical mutations in structurally similar positions, revealing a dramatic context dependence (epistasis) of designed mutations in homologous protein-protein interactions. To better understand the structural basis of this context dependence, we apply a structure-based computational model that recapitulates these energetic effects and we use this model to make and validate forward predictions. Although the context dependence of these mutations is captured by computational predictions, our results both highlight the considerable difficulties in designing protein-protein interactions and provide challenging benchmark cases for the development of improved protein modeling and design methods that accurately account for the context

Selectivity Challenges in Docking Screens for GPCR Targets and Antitargets.

To investigate large library docking's ability to find molecules with joint activity against on-targets and selectivity versus antitargets, the dopamine D2 and serotonin 5-HT2A receptors were targeted, seeking selectivity against the histamine H1 receptor. In a second campaign, κ-opioid receptor ligands were sought with selectivity versus the μ-opioid receptor. While hit rates ranged from 40% to 63% against the on-targets, they were just as good against the antitargets, even though the molecules were selected for their putative lack of binding to the off-targets. Affinities, too, were often as good or better for the off-targets. Even though it was occasionally possible to find selective molecules, such as a mid-nanomolar D2/5-HT2A ligand with 21-fold selectivity versus the H1 receptor, this was the exception. Whereas false-negatives are tolerable in docking screens against on-targets, they are intolerable against antitargets; addressing this problem may demand new strategies in the field

Ask those who are ahead about a buffalo : Well-being of African grandparents with HIV

Older adults with HIV in sub-Saharan Africa often care for grandchildren, but the impact of caring on well-being has not been studied extensively. We examined 208 older adults with HIV in Uganda (n = 100) and South Africa (n = 108); 90% were grandparents and 47% cared for grandchildren (carers). Carers were vulnerable compared to noncarers and nongrandparents with a significantly higher average number of comorbidities (4.1 vs. 2.7 vs. 1.9, respectively) and depressive symptoms (11.3 vs. 9.1 vs. 7.3, respectively). However, carers had significantly lower average HIV stigma scores (32.2 vs. 35.5 vs. 42.5, respectively) and significantly higher perceived emotional support (6.1 vs. 5.7 vs. 5.4, respectively). Multiple regression analysis indicated that comorbidities and financial need mediated the positive association between grandparent carers and depression. The negative association between being a carer and stigma was no longer significant when controlling for other factors such as project site. We observed a significant Site x Comorbidities interaction indicating that the positive association between comorbidities and stigma was stronger for Ugandan participants. Findings highlight the need for improved management of comorbidities and the need for better financial security among older people with HIV in sub-Saharan Africa regardless of caregiving involvement