Establishing Normal Testicular 18F-FDG PET/CT SUVs.

OBJECTIVE. Knowledge of normal testicular 18F-FDG PET/CT (FDG PET/CT) SUVs is crucial for accurate examination interpretation. The objective of this study was to establish normal testicular SUV ranges among adult men receiving health care in North America. MATERIALS AND METHODS. A retrospective review of an institutional electronic database identified adult men undergoing pretreatment clinical FDG PET/CT examinations from March 15, 2013, through March 15, 2018. An FDG PET/CT image review of 700 testicles in 350 male patients was performed. Data collected included testicular SUVmax, SUVmean, and visual PET pattern of uptake. RESULTS. Testicular SUVmean and SUVmax values (mean ± SD) by age group were as follows: 3.1 ± 0.7 and 3.8 ± 0.9 for the age group of 18-30 years; 3.2 ± 0.6 and 4.0 ± 0.8, 31-40 years; 3.1 ± 0.4 and 3.8 ± 0.5, 41-50 years; 3.0 ± 0.5 and 3.7 ± 0.7, 51-60 years; 2.9 ± 0.5 and 3.5 ± 0.7, 61-70 years; 2.8 ± 0.5 and 3.5 ± 0.7, 71-80 years; and 2.6 ± 0.5 and 3.3 ± 0.6, more than 80 years. A statistically significant difference exists between age groups for testicular SUVmean (p ≤ .001) and SUVmax (p < .001), with SUVs peaking in the 4th decade of life and subsequently declining with age. A small but significant negative correlation exists between blood glucose level and testicular SUVmean (r = -0.12). CONCLUSION. This study reports the largest currently known cohort of SUVs in normal testicles and may guide clinical interpretation of testicular FDG activity. Discrepancies in normal SUVs may exist because of differences in patient demographics and PET technology.

Binding of a potent small-molecule inhibitor of six-helix bundle formation requires interactions with both heptad-repeats of the RSV fusion protein.

Six-helix bundle (6HB) formation is an essential step for many viruses that rely on a class I fusion protein to enter a target cell and initiate replication. Because the binding modes of small molecule inhibitors of 6HB formation are largely unknown, precisely how they disrupt 6HB formation remains unclear, and structure-based design of improved inhibitors is thus seriously hampered. Here we present the high resolution crystal structure of TMC353121, a potent inhibitor of respiratory syncytial virus (RSV), bound at a hydrophobic pocket of the 6HB formed by amino acid residues from both HR1 and HR2 heptad-repeats. Binding of TMC353121 stabilizes the interaction of HR1 and HR2 in an alternate conformation of the 6HB, in which direct binding interactions are formed between TMC353121 and both HR1 and HR2. Rather than completely preventing 6HB formation, our data indicate that TMC353121 inhibits fusion by causing a local disturbance of the natural 6HB conformation.

An ensemble of structures of Burkholderia pseudomallei 2,3-bisphosphoglycerate-dependent phosphoglycerate mutase.

Burkholderia pseudomallei is a soil-dwelling bacterium endemic to Southeast Asia and Northern Australia. Burkholderia is responsible for melioidosis, a serious infection of the skin. The enzyme 2,3-bisphosphoglycerate-dependent phosphoglycerate mutase (PGAM) catalyzes the interconversion of 3-phosphoglycerate and 2-phosphoglycerate, a key step in the glycolytic pathway. As such it is an extensively studied enzyme and X-ray crystal structures of PGAM enzymes from multiple species have been elucidated. Vanadate is a phosphate mimic that is a powerful tool for studying enzymatic mechanisms in phosphoryl-transfer enzymes such as phosphoglycerate mutase. However, to date no X-ray crystal structures of phosphoglycerate mutase have been solved with vanadate acting as a substrate mimic. Here, two vanadate complexes together with an ensemble of substrate and fragment-bound structures that provide a comprehensive picture of the function of the Burkholderia enzyme are reported.

Switch control pocket inhibitors of p38-MAP kinase. Durable type II inhibitors that do not require binding into the canonical ATP hinge region.

Switch control pocket inhibitors of p38-alpha kinase are described. Durable type II inhibitors were designed which bind to arginines (Arg67 or Arg70) that function as key residues for mediating phospho-threonine 180 dependant conformational fluxing of p38-alpha from an inactive type II state to an active type I state. Binding to Arg70 in particular led to potent inhibitors, exemplified by DP-802, which also exhibited high kinase selectivity. Binding to Arg70 obviated the requirement for binding into the ATP Hinge region. X-ray crystallography revealed that DP-802 and analogs induce an enhanced type II conformation upon binding to either the unphosphorylated or the doubly phosphorylated form of p38-alpha kinase.

Identification of a peptide-peptide binding motif in the coating of nab-paclitaxel nanoparticles with clinical antibodies: bevacizumab, rituximab, and trastuzumab.

Antibody directed chemotherapy (ADC) takes advantage of the selectivity of the monoclonal antibody to increase the efficacy of the chemotherapeutic agent, while reducing toxicity. Previously we described three nab-paclitaxel (Abraxane) nanoparticles coated with commercial monoclonal antibodies. Identifying the binding sites responsible for these particles could allow reverse engineering of nab-paclitaxel binding antibodies, creating a modular platform for antibody directed chemotherapeutic nanoparticles. Herein, Biacore surface plasmon resonance is used to identify an antibody binding site, HSA Peptide 40, on human serum albumin with nanomolar affinity for all three monoclonal antibodies. This 18-mer peptide, which lies in Subdomain IIIA of human serum albumin, blocks binding of all three antibodies to nab-paclitaxel when added in excess. We furthermore show the complementary binding region on all three monoclonal antibodies to be the CDR H3 loop of the Fab region, and show that they all have nano to micromolar affinity for HSA Peptide 40 and nab-paclitaxel nanoparticles. The presented data identify the nature of the critical protein-protein interaction that enables antibody coating of nab-paclitaxel.

Frailty assessment in the geriatric outpatient clinic.

AIM: Frailty is a common phenomenon in geriatric patients. In the present translational research study, we assessed two frailty instruments (Fried 2001; Gill 2002), comparing the usefulness and scoring classifications for frailty screening in an academically affiliated geriatrics clinic. METHODS: Assessment was completed on 162 male veterans (mean age 83.7 years, 57% African American) enrolled in a geriatric clinic. The instruments' component criteria, which are well known to gerontological clinicians, were administered in a standard order and scoring was identical to original instruments. RESULTS: The five-item Fried frailty instrument required 15-20 min to complete; the two-item Gill frailty instrument required less than 2 min. Of the 162 participants assessed, 72 were determined to be frail by at least one of the instruments, but just 33 were frail by both instruments. Correlations between the instruments were Spearman = 0.55 (P < 0.001) and kappa = 0.25, (P < 0.001). There were no differences in frailty scores based on race, and there were equivocal results based on age, even though this was an older sample, with almost 17% ≥90 years. A total of 63% (103/162) of the sample met the criterion for weak grip strength, and decreasing grip strength correlated with increasing age (r = -0.238, P = 0.002). CONCLUSION: Expedient identification of the frailty syndrome remains an unmet necessity for clinical practice. The different results by the Fried and Gill frailty instruments are likely due to differences in component domains and testing methods. The present results support previous findings that showed that grip strength might be an important indicator of increasing frailty.

Treatment of Cholesterol Embolization Syndrome in the Setting of an Acute Indication for Anticoagulation Therapy.

Cholesterol embolization syndrome (CES) is a complication sometimes occurring after invasive endovascular procedures. CES is characterized by release of cholesterol crystals and particles from atheromatous plaques, which can occlude distal vessels and induce an inflammatory response, resulting in end-organ damage. We report the case of a 66-year-old man who presented with an acute ST-elevation myocardial infarction. An intra-aortic balloon pump was placed due to hemodynamic instability following percutaneous coronary intervention. Ten weeks after discharge, he presented with signs and symptoms of CES (e.g., livedo reticularis, acrocyanosis, acute renal failure), and a new left ventricular apical thrombus. Withdrawal of anticoagulation is often recommended in the setting of CES, on the presumption that anticoagulants favor plaque hemorrhage and subsequent cholesterol micro-embolization. Because of the potential disastrous consequences of an embolus, the patient was anticoagulated with warfarin concurrently with corticosteroids to suppress the inflammatory response to cholesterol crystals. His renal function continued to improve and was discharged without the need for dialysis. This case illustrates that anticoagulation therapy in CES is feasible and appears to be safe in patients with a coexisting urgent indication for anticoagulation.

Discovery of leukotriene A4 hydrolase inhibitors using metabolomics biased fragment crystallography.

We describe a novel fragment library termed fragments of life (FOL) for structure-based drug discovery. The FOL library includes natural small molecules of life, derivatives thereof, and biaryl protein architecture mimetics. The choice of fragments facilitates the interrogation of protein active sites, allosteric binding sites, and protein-protein interaction surfaces for fragment binding. We screened the FOL library against leukotriene A4 hydrolase (LTA4H) by X-ray crystallography. A diverse set of fragments including derivatives of resveratrol, nicotinamide, and indole were identified as efficient ligands for LTA4H. These fragments were elaborated in a small number of synthetic cycles into potent inhibitors of LTA4H representing multiple novel chemotypes for modulating leukotriene biosynthesis. Analysis of the fragment-bound structures also showed that the fragments comprehensively recapitulated key chemical features and binding modes of several reported LTA4H inhibitors.

Design of an engineered N-terminal HIV-1 gp41 trimer with enhanced stability and potency.

HIV fusion is mediated by a conformational transition in which the C-terminal region (HR2) of gp41 interacts with the N-terminal region (HR1) to form a six-helix bundle. Peptides derived from the HR1 form a well-characterized, trimeric coiled-coil bundle in the presence of HR2 peptides, but there is little structural information on the isolated HR1 trimer. Using protein design, we have designed synthetic HR1 peptides that form soluble, thermostable HR1 trimers. In vitro binding of HR2 peptides to the engineered trimer suggests that the design strategy has not significantly impacted the ability to form the six-helix bundle. The peptides have enhanced antiviral activity compared to wild type, with up to 30-fold greater potency against certain viral isolates. In vitro passaging was used to generate HR1-resistant virus and the observed resistance mutations map to the HR2 region of gp41, demonstrating that the peptides block the fusion process by binding to the viral HR2 domain. Interestingly, the activity of the HR2 fusion inhibitor, enfuvirtide (ENF), against these resistant viruses is maintained or improved up to fivefold. The 1.5 A crystal structure of one of these designs has been determined, and we show that the isolated HR1 is very similar to the conformation of the HR1 in the six-helix bundle. These results provide an initial model of the pre-fusogenic state, are attractive starting points for identifying novel fusion inhibitors, and offer new opportunities for developing HIV therapeutics based on HR1 peptides.