Drugs targeting the orthosteric, primary binding site of G-protein coupled receptors are the most common therapeutics. Allosteric binding sites, elsewhere on the receptors, are less well-defined, and so less exploited clinically. We report the crystal structure of the prototypic beta-2 adrenergic receptor in complex with an orthosteric agonist and Compound-6FA, a positive allosteric modulator of this receptor. It binds on the receptor’s inner surface in a pocket created by intracellular loop 2 and transmembrane segments 3 and 4, stabilizing the loop in an alpha helical conformation required to engage the G-protein. Structural comparison explains the selectivity of the compound for beta-2 over the beta-1 adrenergic receptor. Diversity in location, mechanism, and selectivity of allosteric ligands provides potential to expand the range of receptor drugs.
Conventional drug discovery efforts at the β2-adrenoceptor (β2AR) have led to the development of ligands that bind almost exclusively to the receptor’s hormone-binding orthosteric site. However, targeting the largely unexplored and evolutionarily unique allosteric sites has potential for developing more specific drugs with fewer side effects than orthosteric ligands. Using our recently developed approach for screening G protein–coupled receptors (GPCRs) with DNA-encoded small-molecule libraries, we have discovered and characterized the first β2AR small-molecule positive allosteric modulators (PAMs)—compound (Cmpd)-6 [(R)-N-(4-amino-1-(4-(tert-butyl)phenyl)-4-oxobutan-2-yl)-5-(N-isopropyl-N-methylsulfamoyl)-2-((4-methoxyphenyl)thio)benzamide] and its analogs. We used purified human β2ARs, occupied by a high-affinity agonist, for the affinity-based screening of over 500 million distinct library compounds, which yielded Cmpd-6. It exhibits a low micro-molar affinity for the agonist-occupied β2AR and displays positive cooperativity with orthosteric agonists, thereby enhancing their binding to the receptor and ability to stabilize its active state. Cmpd-6 is cooperative with G protein and β-arrestin1 (a.k.a. arrestin2) to stabilize high-affinity, agonist-bound active states of the β2AR and potentiates downstream cAMP production and receptor recruitment of β-arrestin2 (a.k.a. arrestin3). Cmpd-6 is specific for the β2AR compared with the closely related β1AR. Structure–activity studies of select Cmpd-6 analogs defined the chemical groups that are critical for its biologic activity. We thus introduce the first small-molecule PAMs for the β2AR, which may serve as a lead molecule for the development of novel therapeutics. The approach described in this work establishes a broadly applicable proof-of-concept strategy for affinity-based discovery of small-molecule allosteric compounds targeting unique conformational states of GPCRs.
Posterior cruciate ligament (PCL) injuries can be debilitating knee injuries, having involvement in up to 44% of traumatic knee injuries. However, isolated PCL injuries are relatively infrequent. Therefore, effective evidence-based rehabilitation protocols have proven to be elusive. This systematic review aims to summarize the latest evidence on postoperative rehabilitation protocols for patients undergoing PCL reconstruction. Studies included in this paper included those published from 1991 to 2019 with a grade 1 to 5 level of evidence discussing the postoperative PCL rehabilitation. A multidatabase search using largely PubMed and Ovid was conducted using relevant keywords such as "PCL," "postoperative," and "rehabilitation," initially leading to 955 papers, which were narrowed by relevance to 12 final published studies used in the analysis. Through careful review of the evidence, crucial principles of rehabilitation, such as an initial focus on protecting the graft during strengthening, as well as an optimized PCL reconstruction protocol are presented here. Rehabilitation following PCL reconstruction continues to be limited by a lack of high-quality evidencebased publications.
There remains debate surrounding partial (PN) versus radical nephrectomy (RN) for T1b-T2 renal cell carcinoma (RCC). PN offers nephron-sparing benefits but involves increased perioperative complications. RN putatively maximizes oncologic benefit with complex tumors. We analyzed newly available nephrectomy-specific NSQIP data to elucidate predictors of perioperative outcomes in localized T1b-T2 RCC. We identified 2094 patients undergoing nephrectomy between 2019–2020. Captured variables include surgical procedure and approach, staging, comorbidities, prophylaxis, peri-operative complications, reoperations, and readmissions. 816 patients received PN while 1278 received RN. Reoperation rates were comparable; however, PN patients more commonly experienced 30-day readmissions (7.0% vs. 4.7%, p = 0.026), bleeds (9.19% vs. 5.56%, p = 0.001), renal failure requiring dialysis (1.23% vs. 0.31%, p = 0.013) and urine leak or fistulae (1.10% vs. 0.31%, p = 0.025). Infectious, pulmonary, cardiac, and venothromboembolic event rates were comparable. Robotic surgery reduced occurrence of various complications, readmissions, and reoperations. PN remained predictive of all four complications upon multivariable adjustment. Several comorbidities were predictive of complications including bleeds and readmissions. This population-based cohort explicates perioperative outcomes following nephrectomy for pT1b-T2 RCC. Significant associations between PN, patient-specific factors, and complications were identified. Risk stratification may inform management to improve post-operative quality of life (QOL) and RCC outcomes.
IntroductionStereotactic Body Radiation Therapy (SBRT) has emerged as a definitive therapy for localized prostate cancer (PCa). However, more data is needed to predict patient prognosis to help guide which patients will benefit most from treatment. The FACIT-Fatigue (FACIT-F) is a well validated, widely used survey for assessing fatigue. However, the role of fatigue in predicting PCa survival has yet to be studied. Herein, we investigate the role of FACIT-F as a baseline predictor for overall survival (OS) in patients undergoing SBRT for localized PCa.MethodsA retrospective review was conducted of 1358 patients who received SBRT monotherapy between January 2008 to April 2021 at an academic, tertiary referral center. FACIT-F scores (range 0 to 52) were summed for patients who answered all 13-items on the survey. FACIT-F total scores of ≥35 represented severe fatigue. Patients receiving androgen deprivation therapy were excluded. Differences in fatigue groups were evaluated using chi-squared tests. OS rates were determined using the Kaplan-Meier method and predictors of OS were evaluated using Cox proportional hazard method.ResultsBaseline full FACIT-F scores and survival data was available for 891 patients. 5-year OS was 87.6% and 95.2%, respectively, for the severely fatigued and non-fatigued groups. Chi-squared analysis of fatigue groups showed no significant difference in the following categories: D’Amico risk group, age, ethnicity, grade group, T-stage, or PSA density. Severe fatigue was associated with a significant decrease in OS (hazard ratio 2.76; 95%CI 1.55 - 4.89). The Cox proportional hazard model revealed that age and FACIT-F were both statistically significant (p <0.05).ConclusionBaseline FACIT-F scores are significantly associated with OS. Higher FACIT-F scores, representing less fatigued patients, are associated with an overall survival benefit. These results indicate that the FACIT-F survey could serve as an additional metric for clinicians in determining prognostic factors for patients undergoing SBRT.
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