Evaluation of thermal regimes for transported ambassador ectotherms: One size does not fit all.

Providing appropriate environmental temperatures for captive ectotherms should be a husbandry priority. This can be especially challenging for ectotherms that are routinely transported, such as those used in education programs at zoos, because they are unable to thermoregulate while confined in non-temperature controlled, compact carriers. To assess if ectotherms used in the Fort Wayne Children's Zoo's outreach programs experienced appropriate transit temperatures during cold weather, we placed temperature loggers inside two sizes of transport carriers, half containing a heat source (bottle of hot water) and half not (control). While transport temperatures were appropriate for many ectotherms, this simple procedure failed to meet the thermal preferences of species with relatively low or high preferred temperatures such as the eastern tiger salamander (Ambystoma tigrinum) and the spiny-tailed lizard (Uromastyx maliensis), respectively. We found large heated carriers were warmer than small heated carriers, but the temperatures of control carriers did not differ. Despite considerable interspecific variation, large heated carriers provided higher thermal quality environments than both small heated and control carriers for all species except eastern tiger salamanders. We suggest further thermal monitoring of ectotherms during transit with the aim of identifying appropriate heat sources and developing efficient and effective transportation protocols. This could be achieved by modifying transport carriers so that animals are able to thermoregulate. Limiting or ceasing their use when appropriate temperatures cannot be provided may be necessary. Particular attention should be given to species with temperature preferences markedly different than the majority of others in a given collection. Zoo Biol. 35:339-345, 2016. © 2016 Wiley Periodicals, Inc.

Improving the Pharmacokinetics of GPR40/FFA1 Full Agonists.

We recently reported the discovery of a potent GPR40 full agonist AM-1638 (1). Herein, we describe our efforts in improving the drug-like properties of the full agonists through the systematic introduction of polar groups in the C-, D-, and A-rings. This led to the discovery of new GPR40 full agonists with significantly improved pharmacokinetic propeties. Compound 8 and 20 also showed potent in vivo efficacy in oral glucose tolerance tests in mice in addition to the improvement in properties.

Structure-assisted discovery of the first non-retinoid ligands for Retinol-Binding Protein 4.

Retinol-Binding Protein 4 (RBP4) is a plasma protein that transports retinol (vitamin A) from the liver to peripheral tissues. This Letter highlights our efforts in discovering the first, to our knowledge, non-retinoid small molecules that bind to RBP4 at the retinol site and reduce serum RBP4 levels in mice, by disrupting the interaction between RBP4 and transthyretin (TTR), a plasma protein that binds RBP4 and protects it from renal excretion. Potent compounds were discovered and optimized quickly from high-throughput screen (HTS) hits utilizing a structure-based approach. Inhibitor co-crystal X-ray structures revealed unique disruptions of RBP4-TTR interactions by our compounds through induced loop conformational changes instead of steric hindrance exemplified by fenretinide. When administered to mice, A1120, a representative compound in the series, showed concentration-dependent retinol and RBP4 lowering.

Discovery of AMG 925, a FLT3 and CDK4 dual kinase inhibitor with preferential affinity for the activated state of FLT3.

We describe the structural optimization of a lead compound 1 that exhibits dual inhibitory activities against FLT3 and CDK4. A series of pyrido[4',3':4,5]pyrrolo[2,3-d]pyrimidine derivatives was synthesized, and SAR analysis, using cell-based assays, led to the discovery of 28 (AMG 925), a potent and orally bioavailable dual inhibitor of CDK4 and FLT3, including many FLT3 mutants reported to date. Compound 28 inhibits the proliferation of a panel of human tumor cell lines including Colo205 (Rb(+)) and U937 (FLT3(WT)) and induced cell death in MOLM13 (FLT3(ITD)) and even in MOLM13 (FLT3(ITD, D835Y)), which exhibits resistance to a number of FLT3 inhibitors currently under clinical development. At well-tolerated doses, compound 28 leads to significant growth inhibition of MOLM13 xenografts in nude mice, and the activity correlates with inhibition of STAT5 and Rb phosphorylation.

Structure guided design of a series of sphingosine kinase (SphK) inhibitors.

Sphingosine-1-phosphate (S1P) signaling plays a vital role in mitogenesis, cell migration and angiogenesis. Sphingosine kinases (SphKs) catalyze a key step in sphingomyelin metabolism that leads to the production of S1P. There are two isoforms of SphK and observations made with SphK deficient mice show the two isoforms can compensate for each other's loss. Thus, inhibition of both isoforms is likely required to block SphK dependent angiogenesis. A structure based approach was used to design and synthesize a series of SphK inhibitors resulting in the identification of the first potent inhibitors of both isoforms of human SphK. Additionally, to our knowledge, this series of inhibitors contains the only sufficiently potent inhibitors of murine SphK1 with suitable physico-chemical properties to pharmacologically interrogate the role of SphK1 in rodent models and to reproduce the phenotype of SphK1 (-/-) mice.

Methodology for fitting and updating predictive accident models with trend.

Reliable predictive accident models (PAMs) (also referred to as Safety Performance Functions (SPFs)) have a variety of important uses in traffic safety research and practice. They are used to help identify sites in need of remedial treatment, in the design of transport schemes to assess safety implications, and to estimate the effectiveness of remedial treatments. The PAMs currently in use in the UK are now quite old; the data used in their development was gathered up to 30 years ago. Many changes have occurred over that period in road and vehicle design, in road safety campaigns and legislation, and the national accident rate has fallen substantially. It seems unlikely that these ageing models can be relied upon to provide accurate and reliable predictions of accident frequencies on the roads today. This paper addresses a number of methodological issues that arise in seeking practical and efficient ways to update PAMs, whether by re-calibration or by re-fitting. Models for accidents on rural single carriageway roads have been chosen to illustrate these issues, including the choice of distributional assumption for overdispersion, the choice of goodness of fit measures, questions of independence between observations in different years, and between links on the same scheme, the estimation of trends in the models, the uncertainty of predictions, as well as considerations about the most efficient and convenient ways to fit the required models.

Discovery of a new class of ghrelin receptor antagonists.

A series of benzodiazepine antagonists of the human ghrelin receptor GHSR1a were synthesized and their antagonism and metabolic stability were evaluated. The potency of these analogs was determined using a functional aequorin (Euroscreen) luminescent assay measuring the intracellular Ca(2+) concentration, and their metabolic stability was measured using an in vitro rat and human S9 hepatocyte assay. These efforts led to the discovery of a potent ghrelin antagonist with good rat pharmacokinetic properties.

Novel pyrrolidine melanin-concentrating hormone receptor 1 antagonists with reduced hERG inhibition.

We discovered novel pyrrolidine MCHR1 antagonist 1 possessing moderate potency. Profiling of pyrrolidine 1 demonstrated that it was an inhibitor of the hERG channel. Investigation of the structure-activity relationship of this class of pyrrolidines allowed us to optimize the MCHR1 potency and decrease the hERG inhibition. Increasing the acidity of the amide proton by converting the benzamide in lead 1 to an anilide provided single digit nanomolar MCHR1 antagonists while replacing the dimethoxyphenyl ring of 1 with alkyl groups possessing increased polarity dramatically reduced the hERG inhibition.

Identification of potent, noncovalent fatty acid amide hydrolase (FAAH) inhibitors.

Starting from a series of ureas that were determined to be mechanism-based inhibitors of FAAH, several spirocyclic ureas and lactams were designed and synthesized. These efforts identified a series of novel, noncovalent FAAH inhibitors with in vitro potency comparable to known covalent FAAH inhibitors. The mechanism of action for these compounds was determined through a combination of SAR and co-crystallography with rat FAAH.

Identification of piperazine-bisamide GHSR antagonists for the treatment of obesity.

Piperazine-bisamide analogs were discovered as partial agonists of human growth hormone secretagogue receptor (GHSR) in a high throughput screen. The partial agonists were optimized for potency and converted into antagonists through structure-activity relationship (SAR) studies. The efforts also led to the identification of potent antagonist with favorable PK profile suitable as a tool compound for in vivo studies.

Discovery and optimization of a novel Neuromedin B receptor antagonist.

The discovery and parallel synthesis of potent, small molecule antagonists of Neuromedin B receptor based on the ary-hexahydro-dibenzodiazepin-1-one core is described.

Identification of a PPARdelta agonist with partial agonistic activity on PPARgamma.

The discovery and optimization of a series of potent PPARdelta full agonists with partial agonistic activity against PPARgamma is described.

Identification and characterization of a non-retinoid ligand for retinol-binding protein 4 which lowers serum retinol-binding protein 4 levels in vivo.

Retinol-binding protein 4 (RBP4) transports retinol from the liver to extrahepatic tissues, and RBP4 lowering is reported to improve insulin sensitivity in mice. We have identified A1120, a high affinity (K(i) = 8.3 nm) non-retinoid ligand for RBP4, which disrupts the interaction between RBP4 and its binding partner transthyretin. Analysis of the RBP4-A1120 co-crystal structure reveals that A1120 induces critical conformational changes at the RBP4-transthyretin interface. Administration of A1120 to mice lowers serum RBP4 and retinol levels but, unexpectedly, does not improve insulin sensitivity. In addition, we show that Rpb4(-/-) mice display normal insulin sensitivity and are not protected from high fat diet-induced insulin resistance. We conclude that lowering RBP4 levels does not improve insulin sensitivity in mice. Therefore, RBP4 lowering may not be an effective strategy for treating diabetes.

Design and synthesis of protein superfamily-targeted chemical libraries for lead identification and optimization.

This review chronicles original literature dating back to 1992 outlining the applications of parallel synthesis and combinatorial chemistry to the synthesis of compound libraries focused towards specific superfamilies of molecular targets. Target families that have received significant literature coverage include kinases, proteases, nuclear hormone receptors and cell surface receptors, notably GPCRs.