Machine Learning Models to Predict Inhibition of the Bile Salt Export Pump.

Cholestatic liver injury is frequently associated with drug inhibition of bile salt transporters, such as the bile salt export pump (BSEP). Reliable in silico models to predict BSEP inhibition directly from chemical structures would significantly reduce costs during drug discovery and could help avoid injury to patients. We report our development of classification and regression models for BSEP inhibition with substantially improved performance over previously published models. We assessed the performance effects of different methods of chemical featurization, data set partitioning, and class labeling and identified the methods producing models that generalized best to novel chemical entities.

Rethinking drug design in the artificial intelligence era.

Artificial intelligence (AI) tools are increasingly being applied in drug discovery. While some protagonists point to vast opportunities potentially offered by such tools, others remain sceptical, waiting for a clear impact to be shown in drug discovery projects. The reality is probably somewhere in-between these extremes, yet it is clear that AI is providing new challenges not only for the scientists involved but also for the biopharma industry and its established processes for discovering and developing new medicines. This article presents the views of a diverse group of international experts on the 'grand challenges' in small-molecule drug discovery with AI and the approaches to address them.

Impact of External Quality Assurance on the Quality of Xpert MTB/RIF Testing in Viet Nam.

Following the endorsement of the Xpert MTB/RIF assay (Cepheid, Sunnyvale, CA, USA) by the World Health Organization (WHO) in 2010, Viet Nam's National Tuberculosis Control Program (NTP) began using GeneXpert instruments in NTP laboratories. In 2013, Viet Nam's NTP implemented an Xpert MTB/RIF external quality assurance (EQA) program in collaboration with the U.S. Centers for Disease Control and Prevention (CDC) and the Foundation for Innovative New Diagnostics (FIND). Proficiency-testing (PT) panels comprising five dried tube specimens (DTS) were sent to participating sites approximately twice a year from October 2013 to July 2016. The number of enrolled laboratories increased from 22 to 39 during the study period. Testing accuracy was assessed by comparing reported and expected results; percentage scores were assigned; and feedback reports were provided to sites. On-site evaluation (OSE) was conducted for underperforming laboratories. The results from the first five rounds demonstrate the positive impact of PT and targeted OSE visits on testing quality. On average, for every additional round of feedback, the odds of achieving PT scores of ≥80% increased 2.04-fold (95% confidence interval, 1.39- to 3.00-fold). Future work will include scaling up PT to all sites and maintaining the performance of participating laboratories while developing local panel production capacity.

Development of MK-8353, an orally administered ERK1/2 inhibitor, in patients with advanced solid tumors.

BACKGROUND: Constitutive activation of ERK1/2 occurs in various cancers, and its reactivation is a well-described resistance mechanism to MAPK inhibitors. ERK inhibitors may overcome the limitations of MAPK inhibitor blockade. The dual mechanism inhibitor SCH772984 has shown promising preclinical activity across various BRAFV600/RAS-mutant cancer cell lines and human cancer xenografts. METHODS: We have developed an orally bioavailable ERK inhibitor, MK-8353; conducted preclinical studies to demonstrate activity, pharmacodynamic endpoints, dosing, and schedule; completed a study in healthy volunteers (P07652); and subsequently performed a phase I clinical trial in patients with advanced solid tumors (MK-8353-001). In the P07652 study, MK-8353 was administered as a single dose in 10- to 400-mg dose cohorts, whereas in the MK-8353-001 study, MK-8353 was administered in 100- to 800-mg dose cohorts orally twice daily. Safety, tolerability, pharmacokinetics, pharmacodynamics, and antitumor activity were analyzed. RESULTS: MK-8353 exhibited comparable potency with SCH772984 across various preclinical cancer models. Forty-eight patients were enrolled in the P07652 study, and twenty-six patients were enrolled in the MK-8353-001 study. Adverse events included diarrhea (44%), fatigue (40%), nausea (32%), and rash (28%). Dose-limiting toxicity was observed in the 400-mg and 800-mg dose cohorts. Sufficient exposure to MK-8353 was noted that correlated with biological activity in preclinical data. Three of fifteen patients evaluable for treatment response in the MK-8353-001 study had partial response, all with BRAFV600-mutant melanomas. CONCLUSION: MK-8353 was well tolerated up to 400 mg twice daily and exhibited antitumor activity in patients with BRAFV600-mutant melanoma. However, antitumor activity was not particularly correlated with pharmacodynamic parameters. TRIAL REGISTRATION: ClinicalTrials.gov NCT01358331. FUNDING: Merck Sharp & Dohme Corp., a subsidiary of Merck & Co. Inc., and NIH (P01 CA168585 and R35 CA197633).

A dual role for Integrin α6ß4 in modulating hereditary neuropathy with liability to pressure palsies.

Peripheral myelin protein 22 (PMP22) is a component of compact myelin in the peripheral nervous system. The amount of PMP22 in myelin is tightly regulated, and PMP22 over or under-expression cause Charcot-Marie-Tooth 1A (CMT1A) and Hereditary Neuropathy with Pressure Palsies (HNPP). Despite the importance of PMP22, its function remains largely unknown. It was reported that PMP22 interacts with the ß4 subunit of the laminin receptor α6ß4 integrin, suggesting that α6ß4 integrin and laminins may contribute to the pathogenesis of CMT1A or HNPP. Here we asked if the lack of α6ß4 integrin in Schwann cells influences myelin stability in the HNPP mouse model. Our data indicate that PMP22 and ß4 integrin may not interact directly in myelinating Schwann cells, however, ablating ß4 integrin delays the formation of tomacula, a characteristic feature of HNPP. In contrast, ablation of integrin ß4 worsens nerve conduction velocities and non-compact myelin organization in HNPP animals. This study demonstrates that indirect interactions between an extracellular matrix receptor and a myelin protein influence the stability and function of myelinated fibers.

Rapid Ascent From Zero Quality to International Organization for Standardization Accreditation: A Case Study of Hai Duong Preventive Medicine Center in Vietnam, 2012-2013.

OBJECTIVES: In 2012, the Vietnam Ministry of Health sought to improve the quality of health laboratories by introducing international quality standards. METHODS: Strengthening Laboratory Management Toward Accreditation (SLMTA), a year-long, structured, quality improvement curriculum (including projects and mentorship) was piloted in 12 laboratories. Progress was measured using a standardized audit tool (Stepwise Laboratory Quality Improvement Process Towards Accreditation). RESULTS: All 12 pilot laboratories (a mix of hospital and public health) demonstrated improvement; median scores rose from 44% to 78% compliance. The public health laboratory in Hai Duong Province entered the program with the lowest score of the group (28%) yet concluded with the highest score (86%). Five months after the completion of the program, without any additional external support, they were accredited. Laboratory management/staff describe factors key to their success: support from the facility senior management, how-to guidance provided by SLMTA, support from the site mentor, and strong commitment of laboratory staff. CONCLUSIONS: Hai Duong preventive medical center is one of only a handful of laboratories to reach accreditation after participation in SLMTA and the only laboratory to do so without additional support. Due to the success seen in Hai Duong and other pilot laboratories, Vietnam has expanded the use of SLMTA.

Investigation of an outbreak of bloody diarrhea complicated with hemolytic uremic syndrome.

In July-August 2009, eight patients with bloody diarrhea complicated by hemolytic uremic syndrome (HUS) were admitted to hospitals in Tbilisi, Georgia. We started active surveillance in two regions for bloody diarrhea and post-diarrheal HUS. Of 25 case-patients who developed HUS, including the initial 8 cases, half were ⩾15 years old, 67% were female and seven (28%) died. No common exposures were identified. Among 20 HUS case-patients tested, Shiga toxin was detected in the stools of 2 patients (one with elevated serum IgG titers to several Escherichia coli serogroups, including O111 and O104). Among 56 persons with only bloody diarrhea, we isolated Shiga toxin-producing E. coli (STEC) O104:H4 from 2 and Shigella from 10; 2 had serologic evidence of E. coli O26 infection. These cases may indicate a previously unrecognized burden of HUS in Georgia. We recommend national reporting of HUS and improving STEC detection capacity.

Thiophene carboxamide inhibitors of JAK2 as potential treatments for myleoproliferative neoplasms.

A series of carboxamide-substituted thiophenes demonstrating inhibition of JAK2 is described. Development of this chemical series began with the bioisosteric replacement of a urea substituent by a pyridyl ring. Issues of chemical and metabolic stability were solved using the results of both in vitro and in vivo studies, ultimately delivering compounds such as 24 and 25 that performed well in an acute PK/PD model measuring p-STAT5 inhibition.

Disease surveillance system evaluation as a model for improved integration and standardization of the laboratory component in the Field Epidemiology and Laboratory Training Program (FELTP) curriculum worldwide.

Integration of laboratory training into the Centers for Disease Control and Prevention's (CDC) Field Epidemiology Training Program (FETP) began in 2004 and has advanced the training of laboratory scientists worldwide on the basic principles of epidemiology, disease surveillance, and outbreak investigation. The laboratory component of the FE(L)TP training has traditionally been disease specific, revolving around classroom and bench training on laboratory methods, and field placement in areas where services are needed. There is however a need to improve the integration of epidemiology elements used in surveillance, outbreak investigation, and evaluation activities with specific measurable laboratory activities that could in turn impact the overall disease surveillance and response. A systematic and clear evaluation guideline for the laboratory components of disease surveillance systems alongside the corresponding epidemiological indicators can better identify, address, and mitigate weaknesses that may exist in the entire surveillance system, and also help to integrate and standardize the FE(L)TP curriculum content. The institution of laboratory Quality Management System principles linked to a comprehensive surveillance evaluation scheme will result in improved disease surveillance, response, and overall laboratory capacity over time.

The discovery of tricyclic pyridone JAK2 inhibitors. Part 1: hit to lead.

This paper describes the discovery and design of a novel class of JAK2 inhibitors. Furthermore, we detail the optimization of a screening hit using ligand binding efficiency and log D. These efforts led to the identification of compound 41, which demonstrates in vivo activity in our study.

Efficient identification of novel leads by dynamic focused screening: PDK1 case stud.

A dynamic, focused screening strategy that utilized a limited but diversified set of target-specific compounds was explored as an efficient means for the identification of inhibitors of the protein kinase PDK1. Approximately 21,500 compounds, including a 19,000 molecule kinase-focused compound collection (KFCC), were screened at two concentrations to identify initial leads. The KFCC included several empirically-derived, general kinase libraries and molecules chosen by PDK1-specific virtual screens. As was expected, this initial screen mostly identified potent leads with limited novelty. In order to overcome this limitation, the data from the screen were used to drive several rounds of a customized iterative focused screening (IFS) campaign. A machine-learning technique was used to build a predictive model to identify compounds to be screened in subsequent rounds. Molecules deemed not to be novel were removed from the training set for the next round, which allowed this campaign to progressively walk away from the chemical space covered by the KFCC. This resulted in the identification of PDK1 inhibitors which are uniquely different from publicly known chemotypes after just three rounds of screenings. A retrospective analysis of this IFS approach against an ultra-high throughput screen (uHTS) indicated that while uHTS is still the most prolific paradigm for lead identification, this dynamic, focused screening approach was successful in discovering novel scaffolds for a medicinal chemistry effort. Finally, a theoretical optimization suggested the dynamic, focused screening approaches could provide either a complementary or alternative approach to uHTS for the efficient and rapid lead identification.

Development of high-throughput TR-FRET and AlphaScreen assays for identification of potent inhibitors of PDK1.

The PI3K/Akt signaling pathway plays a key role in cancer cell growth, survival, and tumor angiogenesis. 3-Phosphoinositide-dependent protein kinase 1 (PDK1) is a Ser/Thr protein kinase, which catalyzes the phosphorylation of a conserved residue in the activation loop of a number of AGC kinases, including proto-oncogenes Akt, p70S6K, and RSK kinases. To find new small-molecule inhibitors of this important regulator kinase, the authors have developed PDK1-specific high-throughput enzymatic assays in time-resolved fluorescence resonance energy transfer (TR-FRET) and AlphaScreen formats, monitoring phosphorylation of a biotinylated peptide substrate derived from the activation loop of Akt. Development of homogeneous assays enabled screening of a focused kinase library of approximately 21,500 compounds in 1536-well TR-FRET format in duplicate. Upon validation of hits in an alternative 384-well AlphaScreen assay, several classes of structurally diverse PDK1 inhibitors, including tetracyclics, tricyclics, azaindoles, indazoles, and indenylpyrazoles, were identified, thus confirming the utility and sensitivity of the developed assays. Further testing in PC3 prostate cancer cells confirmed that representatives of the tetracyclic series showed intracellular modulation of the PDK1 activity, as evident from decreased phosphorylation levels of AKT, RSK, and S6-ribosomal protein.

3,4-Disubstituted benzofuran P1' MMP-13 inhibitors: optimization of selectivity and reduction of protein binding.

Potent 3,4-disubstituted benzofuran P1' MMP-13 inhibitors have been prepared. Selectivity over MMP-2 was achieved through a substituent at the C4 position of the benzofuran P1' moiety of the molecule. By replacing a backbone benzene with a pyridine and valine with threonine, compounds (e.g., 44) with greatly reduced plasma protein binding were also obtained.

Molecular mechanism of action of pharmacoperone rescue of misrouted GPCR mutants: the GnRH receptor.

The human GnRH receptor (hGnRHR), a G protein-coupled receptor, is a useful model for studying pharmacological chaperones (pharmacoperones), drugs that rescue misfolded and misrouted protein mutants and restore them to function. This technique forms the basis of a therapeutic approach of rescuing mutants associated with human disease and restoring them to function. The present study relies on computational modeling, followed by site-directed mutagenesis, assessment of ligand binding, effector activation, and confocal microscopy. Our results show that two different chemical classes of pharmacoperones act to stabilize hGnRHR mutants by bridging residues D(98) and K(121). This ligand-mediated bridge serves as a surrogate for a naturally occurring and highly conserved salt bridge (E(90)-K(121)) that stabilizes the relation between transmembranes 2 and 3, which is required for passage of the receptor through the cellular quality control system and to the plasma membrane. Our model was used to reveal important pharmacophoric features, and then identify a novel chemical ligand, which was able to rescue a D(98) mutant of the hGnRHR that could not be rescued as effectively by previously known pharmacoperones.

Discovery of novel inhibitors of the ZipA/FtsZ complex by NMR fragment screening coupled with structure-based design.

ZipA is a membrane anchored protein in Escherichia coli that interacts with FtsZ, a homolog of eukaryotic tubulins, forming a septal ring structure that mediates bacterial cell division. Thus, the ZipA/FtsZ protein-protein interaction is a potential target for an antibacterial agent. We report here an NMR-based fragment screening approach which identified several hits that bind to the C-terminal region of ZipA. The screen was performed by 1H-15N HSQC experiments on a library of 825 fragments that are small, lead-like, and highly soluble. Seven hits were identified, and the binding mode of the best one was revealed in the X-ray crystal structure. Similar to the ZipA/FtsZ contacts, the driving force in the binding of the small molecule ligands to ZipA is achieved through hydrophobic interactions. Analogs of this hit were also evaluated by NMR and X-ray crystal structures of these analogs with ZipA were obtained, providing structural information to help guide the medicinal chemistry efforts.

Bed occupancy, turnover intervals and MRSA rates in English hospitals.

This article (a follow on from an article concentrating on Northern Ireland) examines the relationship between percentage bed occupancy (PO), turnover interval (TI) and methicillin-resistant Staphylococcus aureus (MRSA) rates in the acute beds of specialist English hospital trusts and describes the TI and levels of bed occupancy. The data were collected from publicly available data: MRSA rates of blood-borne infection per 1000 bed days from the Department of Health; average length of stay from Hospital Episode Statistics; and percentage occupancy from the Department of Health Hospital Activity statistics were used. Pearson's Correlation coefficients were used as basis for inferential analysis. The mean TI for all trusts was as 0.94 days, median 0.95 days. Twenty percent of trusts had TIs, on average, of less than 0.58 days (13.9 hours) and 10% had a TI less than 0.32 days (7.6 hours). The mean PO was 84.98% and the median was 84.76%. Seventy percent of the trusts exceeded the recommended 82% bed occupancy. The inference from this study is that there is a relationship between TI and PO and rate of MRSA infection in specialist English hospitals and that PO rates are at a level which may interfere with good infection control procedures.

Functional, biophysical, and structural bases for antibacterial activity of tigecycline.

Tigecycline is a novel glycylcycline antibiotic that possesses broad-spectrum activity against many clinically relevant species of bacterial pathogens. The mechanism of action of tigecycline was delineated using functional, biophysical, and molecular modeling experiments in this study. Functional assays showed that tigecycline specifically inhibits bacterial protein synthesis with potency 3- and 20-fold greater than that of minocycline and tetracycline, respectively. Biophysical analyses demonstrated that isolated ribosomes bind tigecycline, minocycline, and tetracycline with dissociation constant values of 10(-8), 10(-7), and >10(-6) M, respectively. A molecular model of tigecycline bound to the ribosome was generated with the aid of a 3.40-angstrom resolution X-ray diffraction structure of the 30S ribosomal subunit from Thermus thermophilus. This model places tigecycline in the A site of the 30S subunit and involves substantial interactions with residues of H34 of the ribosomal subunit. These interactions were not observed in a model of tetracycline binding. Modeling data were consistent with the biochemical and biophysical data generated in this and other recent studies and suggested that tigecycline binds to bacterial ribosomes in a novel way that allows it to overcome tetracycline resistance due to ribosomal protection.

Bed occupancy, turnover interval and MRSA rates in Northern Ireland.

The data describe bed turnover intervals (TI), bed percentage occupancy (PO) and methicillin-resistant Staphylococcus aureus (MRSA) rates per 1000 bed days of patient episodes. It was collected from annual hospital statistics in Northern Ireland (NI) and from the Communicable Diseases Surveillance Centre (CDSC) NI. The descriptive data show 6 of the general 11 surgical Trusts, out of a total of 12 Trusts examined, had PO greater than 85%; and all 11 medical facilities in these Trusts had occupancy rates greater than 85%. A significant correlation was established between turnover interval and MRSA per 1000 bed days of patient episodes in acute services beds. The correlation of PO with MRSA rates was 0.49 (ns). The conclusions drawn from the study are that in many Trusts the rates of bed occupancy for general surgery and general medicine is in excess of national guidelines and rapid turnover of patients is related to rates of MRSA infection. The implications for nurses and managers are discussed.

Synthesis and biological evaluation of biphenylsulfonamide carboxylate aggrecanase-1 inhibitors.

Aggrecanases are recently discovered enzymes that cleave aggrecan, a key component of cartilage. Aggrecanase inhibitors may provide a unique means to halt the progression of cartilage destruction in osteoarthritis. The synthesis and evaluation of biphenylsulfonamidocarboxylic acid inhibitors of aggrecanase-1 are reported. Compound 24 demonstrated 89% inhibition of proteoglycan degradation at 10 microg/mL and has an oral bioavailability in rat of 35%.

Structure-based identification of the binding site for the hemiasterlin analogue HTI-286 on tubulin.

A binding mode of HTI-286, a synthetic analogue of the peptidic antimitotic agent hemiasterlin, to tubulin is proposed. The binding mode was derived from iterative docking experiments directed at regions of the tubulin interdimer interface that are believed to be consistent with all current experimental data regarding the HTI-286/tubulin interaction. These data include (1) competitive inhibition of the tubulin binding of the Vinca alkaloids and other antimitotic agents, (2) proximity to stretches of amino acid residues identified in two separate photoaffinity-labeling experiments, (3) structure-activity relationships for HTI-286 and its analogues, (4) saturation transfer difference nuclear magnetic resonance (NMR) experiments, and (5) NMR transfer nuclear Overhauser effect spectroscopy (NOESY) experiments that potentially identify the bioactive conformation. The predicted binding mode thus affords a means to understand the mode of action of hemiasterlin, HTI-286, and other closely related molecules.