A drug's selectivity for target receptors is essential to its therapeutic utility, but achieving selectivity between similar receptors is challenging. The serendipitous discovery of ligands that ...stimulate target receptors more strongly than closely related receptors, despite binding with similar affinities, suggests a solution. The molecular mechanism of such 'efficacy-driven selectivity' has remained unclear, however, hindering design of such ligands. Here, using atomic-level simulations, we reveal the structural basis for the efficacy-driven selectivity of a long-studied clinical drug candidate, xanomeline, between closely related muscarinic acetylcholine receptors (mAChRs). Xanomeline's binding mode is similar across mAChRs in their inactive states but differs between mAChRs in their active states, with divergent effects on active-state stability. We validate this mechanism experimentally and use it to design ligands with altered efficacy-driven selectivity. Our results suggest strategies for the rational design of ligands that achieve efficacy-driven selectivity for many pharmaceutically important G-protein-coupled receptors.
The development of an innovative detector technology for photon-counting in X-ray imaging is reported. This new generation of detectors, based on pixellated cadmium telluride (CdTe) and cadmium zinc ...telluride (CZT) detector arrays electrically connected to application specific integrated circuits (ASICs) for readout, will produce fast and highly efficient photon-counting and energy-dispersive X-ray imaging. There are a number of applications that can greatly benefit from these novel imagers including mammography, planar radiography, and computed tomography (CT). Systems based on this new detector technology can provide compositional analysis of tissue through spectroscopic X-ray imaging, significantly improve overall image quality, and may significantly reduce X-ray dose to the patient. A very high X-ray flux is utilized in many of these applications. For example, CT scanners can produce ~ 100 Mphotons/mm 2 /s in the unattenuated beam. High flux is required in order to collect sufficient photon statistics in the measurement of the transmitted flux (attenuated beam) during the very short time frame of a CT scan. This high count rate combined with a need for high detection efficiency requires the development of detector structures that can provide a response signal much faster than the transit time of carriers over the whole detector thickness. We have developed CdTe and CZT detector array structures which are 3 mm thick with 16 times 16 pixels and a 1 mm pixel pitch. These structures, in the two different implementations presented here, utilize either a small pixel effect or a drift phenomenon. An energy resolution of 4.75% at 122 keV has been obtained with a 30 ns peaking time using discrete electronics and a 57 Co source. An output rate of 6 times 10 6 counts per second per individual pixel has been obtained with our ASIC readout electronics and a clinical CT X-ray tube. Additionally, the first clinical CT images, taken with several of our prototype photon-counting and energy-dispersive detector modules, are shown.
Twenty-four patients with cancer met predetermined criteria for a diagnosis of zygomycosis over a 10-year period at our institution. All had hematologic malignancy, and most had either neutropenia or ...steroid use as a risk factor. Pulmonary involvement mimicking invasive aspergillosis was the most common presentation, and dissemination was seen in 58% of patients on whom autopsies were performed. Three-fourths of the patients with pulmonary zygomycosis had pathogenic microorganisms other than zygomycetes isolated from respiratory specimens. The sensitivity of cultures in detecting zygomycetes from respiratory specimens was low. A culture positive for zygomycetes was typically a preterminal finding in the fatal, acute cases. Two-thirds of the patients died. Favorable outcome seemed to correlate with lack of pulmonary involvement, surgical debridement, neutrophil recovery, and a cumulative total amphotericin B dose of 2000 mg. Therapy with high-dose amphotericin B, combined with aggressive surgery and immune reconstitution, offers the best chance for survival of cancer patients with zygomycosis.
Background
Since the 1950s, preoperative medical preparation has been widely applied in patients with pheochromocytoma to improve intraoperative hemodynamic instability and postoperative ...complications. However, advancements in preoperative imaging, laparoscopic surgical techniques, and anesthesia have considerably improved management in patients with pheochromocytoma. In consequence, there is no validated consensus on current predictive factors for postoperative morbidity. The aim of this study was to determine perioperative factors which are predictive for postoperative morbidity in patients undergoing laparoscopic adrenalectomy for pheochromocytoma.
Study design
It is a retrospective analysis of prospectively maintained databases in five medical centers from 2002 to 2013. Inclusion criteria were consecutive patients who underwent non-converted laparoscopic unilateral total adrenalectomy for pheochromocytoma.
Results
Two-hundred and twenty-five patients were included. All-cause and cardiovascular postoperative morbidity rates were 16 % (
n
= 36) and 4.8 % (
n
= 11), respectively. Preinduction blood pressure normalization after preoperative medical preparation had no impact on postoperative morbidity. However, past medical history of coronary artery disease (OR CI95 % = 3.39; 1.317–8.727) and incidence of intraoperative hemodynamic instability episodes (both SBP ≥ 160 mmHg and MAP < 60 mmHg) (OR CI95 % = 3.092; 1.451–6.587) remained independent predictors for postoperative all-cause morbidity. Similarly, past medical history of coronary artery disease (OR CI95 % = 14.41; 3.119–66.57), female sex (OR CI95 % = 12.05; 1.807–80.31), and incidence of intraoperative hemodynamic instability episodes (both SBP ≥ 200 mmHg and MAP < 60 mmHg) (OR CI95 % = 4.13; 1.009–16.90) remained independent predictors for postoperative cardiovascular morbidity.
Conclusions
This study identifies risk factors for cardiovascular and all-cause postoperative morbidity after laparoscopic adrenalectomy in current clinical setting. These data can help physicians to guide intra-operative blood pressure management and have to be taken into account in further studies.
Abstract
The M
4
muscarinic acetylcholine receptor (M
4
mAChR) has emerged as a drug target of high therapeutic interest due to its expression in regions of the brain involved in the regulation of ...psychosis, cognition, and addiction. The mAChR agonist, xanomeline, has provided significant improvement in the Positive and Negative Symptom Scale (PANSS) scores in a Phase II clinical trial for the treatment of patients suffering from schizophrenia. Here we report the active state cryo-EM structure of xanomeline bound to the human M
4
mAChR in complex with the heterotrimeric G
i1
transducer protein. Unexpectedly, two molecules of xanomeline were found to concomitantly bind to the monomeric M
4
mAChR, with one molecule bound in the orthosteric (acetylcholine-binding) site and a second molecule in an extracellular vestibular allosteric site. Molecular dynamic simulations supports the structural findings, and pharmacological validation confirmed that xanomeline acts as a dual orthosteric and allosteric ligand at the human M
4
mAChR. These findings provide a basis for further understanding xanomeline’s complex pharmacology and highlight the myriad of ways through which clinically relevant ligands can bind to and regulate GPCRs.
Recent breakthroughs and developments in structural biology have led to a spate of crystal structures for G protein-coupled receptors (GPCRs). This is the case for the muscarinic acetylcholine ...receptors (mAChRs) where inactive-state structures for four of the five subtypes and two active-state structures for one subtype are available. These mAChR crystal structures have provided new insights into receptor mechanisms, dynamics, and allosteric modulation. This is highly relevant to the mAChRs given that these receptors are an exemplar model system for the study of GPCR allostery. Allosteric mechanisms of the mAChRs are predominantly consistent with a two-state model, albeit with some notable recent exceptions. Herein, we discuss the mechanisms for positive and negative allosteric modulation at the mAChRs and compare and contrast these to evidence offered by pharmacological, biochemical, and computational approaches. This analysis provides insight into the fundamental pharmacological properties exhibited by GPCR allosteric modulators, such as enhanced subtype selectivity, probe dependence, and biased modulation while highlighting the current challenges that remain. Though complex, enhanced molecular understanding of allosteric mechanisms will have considerable influence on our understanding of GPCR activation and signaling and development of therapeutic interventions.
Allosteric modulation of G protein-coupled receptors (GPCRs) is a major paradigm in drug discovery. Despite decades of research, a molecular-level understanding of the general principles that govern ...the myriad pharmacological effects exerted by GPCR allosteric modulators remains limited. The M
muscarinic acetylcholine receptor (M
mAChR) is a validated and clinically relevant allosteric drug target for several major psychiatric and cognitive disorders. In this study, we rigorously quantified the affinity, efficacy, and magnitude of modulation of two different positive allosteric modulators, LY2033298 (LY298) and VU0467154 (VU154), combined with the endogenous agonist acetylcholine (ACh) or the high-affinity agonist iperoxo (Ipx), at the human M
mAChR. By determining the cryo-electron microscopy structures of the M
mAChR, bound to a cognate G
protein and in complex with ACh, Ipx, LY298-Ipx, and VU154-Ipx, and applying molecular dynamics simulations, we determine key molecular mechanisms underlying allosteric pharmacology. In addition to delineating the contribution of spatially distinct binding sites on observed pharmacology, our findings also revealed a vital role for orthosteric and allosteric ligand-receptor-transducer complex stability, mediated by conformational dynamics between these sites, in the ultimate determination of affinity, efficacy, cooperativity, probe dependence, and species variability. There results provide a holistic framework for further GPCR mechanistic studies and can aid in the discovery and design of future allosteric drugs.
We expanded on a series of pyrido2,1-fpurine-2,4-dione derivatives as human adenosine A3 receptor (hA3R) antagonists to determine their kinetic profiles and affinities. Many compounds showed high ...affinities and a diverse range of kinetic profiles. We found hA3R antagonists with very short residence time (RT) at the receptor (2.2 min for 5) and much longer RTs (e.g., 376 min for 27 or 391 min for 31). Two representative antagonists (5 and 27) were tested in 35SGTPγS binding assays, and their RTs appeared correlated to their (in)surmountable antagonism. From a k on–k off–K D kinetic map, we divided the antagonists into three subgroups, providing a possible direction for the further development of hA3R antagonists. Additionally, we performed a computational modeling study that sheds light on the crucial receptor interactions, dictating the compounds’ binding kinetics. Knowledge of target binding kinetics appears useful for developing and triaging new hA3R antagonists in the early phase of drug discovery.
PDF
