Efficacy of a Pseudomonas aeruginosa serogroup O9 vaccine.

There are currently no approved vaccines against the opportunistic pathogen Pseudomonas aeruginosa. Among vaccine targets, the lipopolysaccharide (LPS) O antigen of P. aeruginosa is the most immunodominant protective candidate. There are 20 different O antigens composed of different repeat sugar structures conferring serogroup specificity, and 10 are found most frequently in infection. Thus, one approach to combat infection by P. aeruginosa could be to generate immunity with a vaccine cocktail that includes all these serogroups. Serogroup O9 is 1 of the 10 serogroups commonly found in infection, but it has never been developed into a vaccine, due in part to the acid-labile nature of the O9 polysaccharide. Our laboratory has previously shown that intranasal administration of an attenuated Salmonella strain expressing the P. aeruginosa serogroup O11 LPS O antigen was effective in clearing bacteria and preventing mortality in mice following intranasal challenge with serogroup O11 P. aeruginosa. Consequently, we set out to develop a P. aeruginosa serogroup O9 vaccine using a similar approach. Here, we show that Salmonella expressing serogroup O9 triggered an antibody-mediated immune response following intranasal administration to mice and that it conferred protection from P. aeruginosa serogroup O9 in a murine model of acute pneumonia.

In Vitro and In Vivo Properties of CUO246, a Novel Bacterial DNA Gyrase/Topoisomerase IV Inhibitor.

CUO246, a novel DNA gyrase/topoisomerase IV inhibitor, is active in vitro against a broad range of Gram-positive, fastidious Gram-negative, and atypical bacterial pathogens and retains activity against quinolone-resistant strains in circulation. The frequency of selection for single step mutants of wild-type S. aureus with reduced susceptibility to CUO246 was <4.64 × 10-9 at 4× and 8× MIC and remained low when using an isogenic QRDR mutant (<5.24 × 10-9 at 4× and 8× MIC). Biochemical assays indicated that CUO246 had potent inhibitory activity against both DNA gyrase (GyrAB) and topoisomerase IV (ParCE). Furthermore, CUO246 showed rapid bactericidal activity in time-kill assays and potent in vivo efficacy against S. aureus in a neutropenic murine thigh infection model. These results suggest that CUO246 may be useful in treating infections by various causative agents of acute skin and skin structure infections, respiratory tract infections, and sexually transmitted infections.

Degree of Dietary Specialization on Furanocoumarin-Containing Hostplants in a Newly Invasive Web Building Caterpillar.

The genus Depressaria (Lepidoptera: Depressariidae) mostly comprises specialist herbivores with varying capacity for detoxification of defensive phytochemistry. Depressaria depressana, a Eurasian moth recently introduced into North America, is a family-level specialist of the Apiaceae, whose hosts include more than a dozen species in multiple tribes; Depressaria radiella is a super-specialist of Eurasian origin that feeds exclusively on species in the genera Pastinaca and Heracleum throughout its native and introduced range. In eastern North America, it feeds upon Pastinaca sativa, an invasive European species, and Heracleum maximum, a native species. We determined whether differences in furanocoumarin metabolism exist between D. depressana and two isolated populations of D. radiella, feeding exclusively on either P. sativa or H. maximum. We also compared gravimetric estimates of feeding efficiency to assess D. depressana larval performance on different diets. Both populations of D. radiella metabolized furanocoumarins at a greater rate than D. depressana. Although there was no difference in rates of metabolism of linear furanocoumarins in the two populations of D. radiella, individuals collected from H. maximum metabolized angular furanocoumarins more rapidly. The gravimetric assessments of feeding efficiencies revealed that D. depressana exhibited highest efficiencies consuming Daucus carota; moreover, this species survived to pupation consuming fruits of Zizia aurea, an apiaceous species native to North America. Our preliminary phylogenetic analysis, building on an earlier morphological analysis, incorporates mitochondrial cytochrome oxidase subunit 1 data from the BOLD database and revealed that the presence or absence of furanocoumarins is not a strong predictor of species-level evolution in Depressaria.

Personality Disorders as a Basis for Discharge and Denial of Benefits in the Military: Logical or Abusive?

ABSTRACT: The US Department of Defense specifically states that intellectual disability and personality disorders are not diseases for compensation purposes, and disabilities from them may not be service connected absent a superimposed mental disorder. In addition, the diagnosis of a personality disorder led to the discharge of 31,000 troops during the years 2001 to 2010. I review the history of these developments, and how the Diagnostic and Statistical Manual of Mental Disorders enabled these actions. In contrast, the United Kingdom and Canada do not allow such actions. Whether our approach is logical seems highly questionable, especially given the significant problems with the DSM's definitions of personality disorders, definitions at odds with the literature.

Animal Models of Peripheral Pain: Biology Review and Application for Drug Discovery.

Pain is a complex constellation of cognitive, unpleasant sensory, and emotional experiences that primarily serves as a survival mechanism. Pain arises in the peripheral nervous system and pain signals synapse with nerve tracts extending into the central nervous system. Several different schemes are used to classify pain, including the underlying mechanism, tissues primarily affected, and time-course. Numerous animal models of pain, which should be employed with appropriate Institutional Animal Care and Use approvals, have been developed to elucidate pathophysiology mechanisms and aid in identification of novel therapeutic targets. The variety of available models underscores the observations that pain phenotypes are driven by several distinct mechanisms. Pain outcome measurement encompasses both reflexive (responses to heat, cold, mechanical and electrical stimuli) and nonreflexive (spontaneous pain responses to stimuli) behaviors. However, the question of translatability to human pain conditions and potential treatment outcomes remains a topic of continued scrutiny. In this review we discuss the different types of pain and their mechanisms and pathways, available rodent pain models with an emphasis on type of pain stimulations and pain outcome measures and discuss the role of pathologists in assessing and validating pain models.

Mutations Reducing In Vitro Susceptibility to Novel LpxC Inhibitors in Pseudomonas aeruginosa and Interplay of Efflux and Nonefflux Mechanisms.

Upregulated expression of efflux pumps, lpxC target mutations, LpxC protein overexpression, and mutations in fabG were previously shown to mediate single-step resistance to the LpxC inhibitor CHIR-090 in P. aeruginosa Single-step selection experiments using three recently described LpxC inhibitors (compounds 2, 3, and 4) and mutant characterization showed that these mechanisms affect susceptibility to additional novel LpxC inhibitors. Serial passaging of P. aeruginosa wild-type and efflux pump-defective strains using the LpxC inhibitor CHIR-090 or compound 1 generated substantial shifts in susceptibility and underscored the interplay of efflux and nonefflux mechanisms. Whole-genome sequencing of CHIR-090 passage mutants identified efflux pump overexpression, fabG mutations, and novel mutations in fabF1 and in PA4465 as determinants of reduced susceptibility. Two new lpxC mutations, encoding A214V and G208S, that reduce susceptibility to certain LpxC inhibitors were identified in these studies, and we show that these and other target mutations differentially affect different LpxC inhibitor scaffolds. Lastly, the combination of target alteration (LpxCA214V) and upregulated expression of LpxC was shown to be tolerated in P. aeruginosa and could mediate significant decreases in susceptibility.

Target (MexB)- and Efflux-Based Mechanisms Decreasing the Effectiveness of the Efflux Pump Inhibitor D13-9001 in Pseudomonas aeruginosa PAO1: Uncovering a New Role for MexMN-OprM in Efflux of ß-Lactams and a Novel Regulatory Circuit (MmnRS) Controlling MexMN Expression.

Efflux pumps contribute to antibiotic resistance in Gram-negative pathogens. Correspondingly, efflux pump inhibitors (EPIs) may reverse this resistance. D13-9001 specifically inhibits MexAB-OprM in Pseudomonas aeruginosa Mutants with decreased susceptibility to MexAB-OprM inhibition by D13-9001 were identified, and these fell into two categories: those with alterations in the target MexB (F628L and ΔV177) and those with an alteration in a putative sensor kinase of unknown function, PA1438 (L172P). The alterations in MexB were consistent with reported structural studies of the D13-9001 interaction with MexB. The PA1438L172P alteration mediated a >150-fold upregulation of MexMN pump gene expression and a >50-fold upregulation of PA1438 and the neighboring response regulator gene, PA1437. We propose that these be renamed mmnR and mmnS for MexMN regulator and MexMN sensor, respectively. MexMN was shown to partner with the outer membrane channel protein OprM and to pump several ß-lactams, monobactams, and tazobactam. Upregulated MexMN functionally replaced MexAB-OprM to efflux these compounds but was insusceptible to inhibition by D13-9001. MmnSL172P also mediated a decrease in susceptibility to imipenem and biapenem that was independent of MexMN-OprM. Expression of oprD, encoding the uptake channel for these compounds, was downregulated, suggesting that this channel is also part of the MmnSR regulon. Transcriptome sequencing (RNA-seq) of cells encoding MmnSL172P revealed, among other things, an interrelationship between the regulation of mexMN and genes involved in heavy metal resistance.

Neural circuitry and precision medicines for mental disorders: are they compatible?

Given the failure of psychiatry to develop clinically useful biomarkers for psychiatric disorders, and the concomitant failure to develop significant advances in diagnosis and treatment, the National Institute of Mental Health (NIMH) in 2010 launched the Research Domain Criteria (RDoC), a framework for research based on the assumption that mental disorders are disorders of identifiable brain neural circuits, with neural circuitry at the center of units of analysis ranging from genes, molecules, and cells to behavior, self-reports, and paradigms. These were to be integrated with five validated dimensional psychological constructs such as negative and positive valence systems. Four years later, the NIMH stated that the ultimate goal of RDoC is precision medicine for psychiatry, with the assumption that precision medications will normalize dysfunctional neural circuits. How this could be accomplished is not obvious, given that neural circuits are widely distributed, have unclear boundaries, and exhibit a significant degree of neuroplasticity, with multiple circuits present in any given disorder. Moreover, the early focus on neural circuitry has been criticized for its reductionism and neglect of the more recent RDoC emphasis on the integration and equivalence of biological and psychological phenomena. Yet this seems inconsistent with the priorities of the NIMH director, an advocate of the central role of neural circuitry and projects such as the Brain Initiative and the Human Connectome Project. Will such projects, at a cost of at least $10 billion, lead to precision medications for mental disorders, or further diminish funding for clinical care and research?

Whither Research Domain Criteria?

In 2010, the National Institute of Mental Health launched the Research Diagnostic Criteria (RDoC) as a research framework aimed at advancing research into the etiology of mental disorders, the development of clinically actionable biomarkers, and the eventual development of precision medications. The foundation of RDoC in that first phase rested in the assumption that mental disorders are brain disorders that originate in aberrant neural circuitry, and that therapeutic advances could flow from alterations in that circuitry. RDoC proposed a matrix of psychological constructs with seven levels of analysis ranging from the cell to self-report, but with neural circuitry at the center. In 2016, another model was proposed in which neural circuitry became equivalent to other units of analyses. With the advent of a new Director of the NIMH, the emphasis returned to neural circuitry as a priority, along with computational psychiatry. Have these shifts undermined the RDoC project?

Nonclinical safety evaluation of erenumab, a CGRP receptor inhibitor for the prevention of migraine.

Calcitonin gene-related peptide (CGRP) and its receptor have been implicated as a key mediator in the pathophysiology of migraine. Thus, erenumab, a monoclonal antibody antagonist of the CGRP receptor, administered as a once monthly dose of 70 or 140 mg has been approved for the preventive treatment of migraine in adults. Due to the species specificity of erenumab, the cynomolgus monkey was used in the pharmacology, pharmacokinetics, and toxicology studies to support the clinical program. There were no effects of erenumab on platelets in vitro (by binding, activation or phagocytosis assays). Specific staining of human tissues with erenumab did not indicated any off-target binding. There were no erenumab-related findings in a cardiovascular safety pharmacology study in cynomolgus monkeys or in vitro in human isolated coronary arteries. Repeat-dose toxicology studies conducted in cynomolgus monkeys at dose levels up to 225 mg/kg (1 month) or up to 150 mg/kg (up to 6 months) with twice weekly subcutaneous (SC) doses showed no evidence of erenumab-mediated adverse toxicity. There were no effects on pregnancy, embryo-fetal or postnatal growth and development in an enhanced pre-postnatal development study in the cynomolgus monkey. There was evidence of placental transfer of erenumab based on measurable serum concentrations in the infants up to 3 months post birth. The maternal and developmental no-observed-effect level (NOEL) was the highest dose tested (50 mg/kg SC Q2W). These nonclinical data in total indicate no safety signal of concern to date and provide adequate margins of exposure between the observed safe doses in animals and clinical dose levels.

Impact of Inducible blaDHA-1 on Susceptibility of Klebsiella pneumoniae Clinical Isolates to LYS228 and Identification of Chromosomal mpl and ampD Mutations Mediating Upregulation of Plasmid-Borne blaDHA-1 Expression.

Twenty-three Klebsiella pneumoniae (blaDHA-1) clinical isolates exhibited a range of susceptibilities to LYS228, with MICs of ≥8 µg/ml for 9 of these. Mutants with decreased susceptibility to LYS228 and upregulated expression of blaDHA-1 were selected from representative isolates. These had mutations in the chromosomal peptidoglycan recycling gene mpl or ampD Preexisting mpl mutations were also found in some of the clinical isolates examined, and these had strongly upregulated expression of blaDHA-1.

In Vitro Activity of LYS228, a Novel Monobactam Antibiotic, against Multidrug-Resistant Enterobacteriaceae.

LYS228 is a novel monobactam with potent activity against Enterobacteriaceae LYS228 is stable to metallo-ß-lactamases (MBLs) and serine carbapenemases, including Klebsiella pneumoniae carbapenemases (KPCs), resulting in potency against the majority of extended-spectrum ß-lactamase (ESBL)-producing and carbapenem-resistant Enterobacteriaceae strains tested. Overall, LYS228 demonstrated potent activity against 271 Enterobacteriaceae strains, including multidrug-resistant isolates. Based on MIC90 values, LYS228 (MIC90, 1 µg/ml) was ≥32-fold more active against those strains than were aztreonam, ceftazidime, ceftazidime-avibactam, cefepime, and meropenem. The tigecycline MIC90 was 4 µg/ml against the strains tested. Against Enterobacteriaceae isolates expressing ESBLs (n = 37) or displaying carbapenem resistance (n = 77), LYS228 had MIC90 values of 1 and 4 µg/ml, respectively. LYS228 exhibited potent bactericidal activity, as indicated by low minimal bactericidal concentration (MBC) to MIC ratios (MBC/MIC ratios of ≤4) against 97.4% of the Enterobacteriaceae strains tested (264/271 strains). In time-kill studies, LYS228 consistently achieved reductions in CFU per milliliter of 3 log10 units (≥99.9% killing) at concentrations ≥4× MIC for Escherichia coli and K. pneumoniae reference strains, as well as isolates encoding TEM-1, SHV-1, CTX-M-14, CTX-M-15, KPC-2, KPC-3, and NDM-1 ß-lactamases.

Mode of Action of the Monobactam LYS228 and Mechanisms Decreasing In Vitro Susceptibility in Escherichia coli and Klebsiella pneumoniae.

The monobactam scaffold is attractive for the development of new agents to treat infections caused by drug-resistant Gram-negative bacteria because it is stable to metallo-ß-lactamases (MBLs). However, the clinically used monobactam aztreonam lacks stability to serine ß-lactamases (SBLs) that are often coexpressed with MBLs. LYS228 is stable to MBLs and most SBLs. LYS228 bound purified Escherichia coli penicillin binding protein 3 (PBP3) similarly to aztreonam (derived acylation rate/equilibrium dissociation constant [k2/Kd ] of 367,504 s-1 M-1 and 409,229 s-1 M-1, respectively) according to stopped-flow fluorimetry. A gel-based assay showed that LYS228 bound mainly to E. coli PBP3, with weaker binding to PBP1a and PBP1b. Exposing E. coli cells to LYS228 caused filamentation consistent with impaired cell division. No single-step mutants were selected from 12 Enterobacteriaceae strains expressing different classes of ß-lactamases at 8× the MIC of LYS228 (frequency, <2.5 × 10-9). At 4× the MIC, mutants were selected from 2 of 12 strains at frequencies of 1.8 × 10-7 and 4.2 × 10-9 LYS228 MICs were ≤2 µg/ml against all mutants. These frequencies compared favorably to those for meropenem and tigecycline. Mutations decreasing LYS228 susceptibility occurred in ramR and cpxA (Klebsiella pneumoniae) and baeS (E. coli and K. pneumoniae). Susceptibility of E. coli ATCC 25922 to LYS228 decreased 256-fold (MIC, 0.125 to 32 µg/ml) after 20 serial passages. Mutants accumulated mutations in ftsI (encoding the target, PBP3), baeR, acrD, envZ, sucB, and rfaI These results support the continued development of LYS228, which is currently undergoing phase II clinical trials for complicated intraabdominal infection and complicated urinary tract infection (registered at ClinicalTrials.gov under identifiers NCT03377426 and NCT03354754).

Optimization of novel monobactams with activity against carbapenem-resistant Enterobacteriaceae - Identification of LYS228.

Metallo-ß-lactamases (MBLs), such as New Delhi metallo-ß-lactamase (NDM-1) have spread world-wide and present a serious threat. Expression of MBLs confers resistance in Gram-negative bacteria to all classes of ß-lactam antibiotics, with the exception of monobactams, which are intrinsically stable to MBLs. However, existing first generation monobactam drugs like aztreonam have limited clinical utility against MBL-expressing strains because they are impacted by serine ß-lactamases (SBLs), which are often co-expressed in clinical isolates. Here, we optimized novel monobactams for stability against SBLs, which led to the identification of LYS228 (compound 31). LYS228 is potent in the presence of all classes of ß-lactamases and shows potent activity against carbapenem-resistant isolates of Enterobacteriaceae (CRE).

Determinants of Antibacterial Spectrum and Resistance Potential of the Elongation Factor G Inhibitor Argyrin B in Key Gram-Negative Pathogens.

Argyrins are natural products with antibacterial activity against Gram-negative pathogens, such as Pseudomonas aeruginosa, Burkholderia multivorans, and Stenotrophomonas maltophilia We previously showed that argyrin B targets elongation factor G (FusA). Here, we show that argyrin B activity against P. aeruginosa PAO1 (MIC = 8 µg/ml) was not affected by deletion of the MexAB-OprM, MexXY-OprM, MexCD-OprJ, or MexEF-OprN efflux pump. However, argyrin B induced expression of MexXY, causing slight but reproducible antagonism with the MexXY substrate antibiotic ciprofloxacin. Argyrin B activity against Escherichia coli increased in a strain with nine tolC efflux pump partner genes deleted. Complementation experiments showed that argyrin was effluxed by AcrAB, AcrEF, and MdtFX. Argyrin B was inactive against Acinetobacter baumannii Differences between A. baumannii and P. aeruginosa FusA proteins at key residues for argyrin B interaction implied that natural target sequence variation impacted antibacterial activity. Consistent with this, expression of the sensitive P. aeruginosa FusA1 protein in A. baumannii conferred argyrin susceptibility, whereas resistant variants did not. Argyrin B was active against S. maltophilia (MIC = 4 µg/ml). Spontaneous resistance occurred at high frequency in the bacterium (circa 10-7), mediated by mutational inactivation of fusA1 rather than by amino acid substitutions in the target binding region. This strongly suggested that resistance occurred at high frequency through loss of the sensitive FusA1, leaving an alternate argyrin-insensitive elongation factor. Supporting this, an additional fusA-like gene (fusA2) is present in S. maltophilia that was strongly upregulated in response to mutational loss of fusA1.

Etelcalcetide, A Novel Calcimimetic, Prevents Vascular Calcification in A Rat Model of Renal Insufficiency with Secondary Hyperparathyroidism.

Etelcalcetide, a novel peptide agonist of the calcium-sensing receptor, prevents vascular calcification in a rat model of renal insufficiency with secondary hyperparathyroidism. Vascular calcification occurs frequently in patients with chronic kidney disease (CKD) and is a consequence of impaired mineral homeostasis and secondary hyperparathyroidism (SHPT). Etelcalcetide substantially lowers parathyroid hormone (PTH) and fibroblast growth factor-23 (FGF23) levels in SHPT patients on hemodialysis. This study compared the effects of etelcalcetide and paricalcitol on vascular calcification in rats with adenine-induced CKD and SHPT. Uremia and SHPT were induced in male Wistar rats fed a diet supplemented with 0.75% adenine for 4 weeks. Rats were injected with vehicle, etelcalcetide, or paricalcitol for 4 weeks from the beginning of adenine diet. Rats fed an adenine-free diet were included as nonuremic controls. Similar reductions in plasma PTH and parathyroid chief cell proliferation were observed in both etelcalcetide- and paricalcitol-treated rats. Serum calcium and phosphorus were significantly lower in etelcalcetide-treated uremic rats and was unchanged in paricalcitol-treated rats. Both serum FGF23 and aortic calcium content were significantly lower in etelcalcetide-treated uremic rats compared with either vehicle- or paricalcitol-treated uremic rats. The degree of aortic calcium content for etelcalcetide-treated rats was similar to that in nonuremic controls and corroborated findings of lack of histologic aortic mineralization in those groups. In conclusion, etelcalcetide and paricalcitol similarly attenuated progression of SHPT in an adenine rat model of CKD. However, etelcalcetide differentially prevented vascular calcification, at least in part, due to reductions in serum FGF23, calcium, and phosphorus levels.

Nonclinical Safety Profile of Etelcalcetide, a Novel Peptide Calcimimetic for the Treatment of Secondary Hyperparathyroidism.

Etelcalcetide is a novel d-amino acid peptide that functions as an allosteric activator of the calcium-sensing receptor and is being developed as an intravenous calcimimetic for the treatment of secondary hyperparathyroidism in patients with chronic kidney disease on hemodialysis. To support clinical development and marketing authorization, a comprehensive nonclinical safety package was generated. Primary adverse effects included hypocalcemia, tremoring, and convulsions. Other adverse effects were considered sequelae of stress associated with hypocalcemia. Cardiovascular safety evaluations in the dog revealed an anticipated prolongation of the corrected QT interval that was related to reductions in serum calcium. Etelcalcetide did not affect the human ether-a-go-go gene ion channel current. Etelcalcetide was mutagenic in some strains of Salmonella, however, based on the negative results in 2 in vitro and 2 in vivo mammalian genotoxicity assays, including a 28-day Muta mouse study, etelcalcetide is considered nongenotoxic. Further support for a lack of genotoxicity was provided due to the fact that etelcalcetide was not carcinogenic in a 6-month transgenic rasH2 mouse model or a 2-year study in rats. There were no effects on fertility, embryo-fetal development, and prenatal and postnatal development. All of the adverse effects observed in both rat and dog were considered directly or secondarily related to the pharmacologic activity of etelcalcetide and the expected sequelae associated with dose-related reductions in serum calcium due to suppression of parathyroid hormone secretion. These nonclinical data indicate no safety signal of concern for human risk beyond that associated with hypocalcemia and associated QT prolongation.

Characterization of an Acinetobacter baumannii lptD Deletion Strain: Permeability Defects and Response to Inhibition of Lipopolysaccharide and Fatty Acid Biosynthesis.

UNLABELLED: Lipid A on the Gram-negative outer membrane (OM) is synthesized in the cytoplasm by the Lpx pathway and translocated to the OM by the Lpt pathway. Some Acinetobacter baumannii strains can tolerate the complete loss of lipopolysaccharide (LPS) resulting from the inactivation of early LPS pathway genes such as lpxC. Here, we characterized a mutant deleted for lptD, which encodes an OM protein that mediates the final translocation of fully synthesized LPS to the OM. Cells lacking lptD had a growth defect comparable to that of an lpxC deletion mutant under the growth conditions tested but were more sensitive to hydrophobic antibiotics, revealing a more significant impact on cell permeability from impaired LPS translocation than from the loss of LPS synthesis. Consistent with this, ATP leakage and N-phenyl-1-naphthylamine (NPN) fluorescence assays indicated a more severe impact of lptD deletion than of lpxC deletion on inner and outer membrane permeability, respectively. Targeted liquid chromatography-mass spectrometry (LCMS) analysis of LPS intermediates from UDP-3-O-R-3-hydroxylauroyl-N-acetyl-α-d-glucosamine through lipid IV(A) showed that the loss of LptD caused an accumulation of lipid IV(A). This suggested that pathway intermediate accumulation or mislocalization caused by the blockage of later LPS pathway steps impacts envelope integrity. Supporting this notion, chemical inhibition of lipid A precursor enzymes, including LpxC and FabB/F, in the lptD deletion strain partially rescued growth and permeability defects. IMPORTANCE: New antibiotics to treat Gram-negative bacterial infections are urgently needed. Inhibition of LPS biosynthesis is attractive because this would impact viability and cell permeability. Therefore, a better understanding of this pathway is important, especially in strains such as A. baumannii ATCC 19606, where LPS biosynthesis is not essential in vitro. We show that ATCC 19606 also survives the loss of the final translocation of LPS into the OM (lptD deletion). Intriguingly, this impaired cell envelope integrity more than the loss of LPS biosynthesis (lpxC deletion), presumably due to the accumulation of toxic intermediates. Supporting this, chemical inhibition of LPS biosynthesis partially reversed this permeability defect. This extends our understanding of the LPS machinery and provides insights into potential interrelationships of the target steps along this important pathway.

A quantitative measure of handwriting dysfluency for assessing tardive dyskinesia.

Tardive dyskinesia (TD) is a movement disorder commonly associated with chronic exposure to antidopaminergic medications, which may be in some cases disfiguring and socially disabling. The consensus from a growing body of research on the incidence and prevalence of TD in the modern era of antipsychotics indicates that this disorder has not disappeared continues to challenge the effective management of psychotic symptoms in patients with schizophrenia. A fundamental component in an effective strategy for managing TD is its reliable and accurate assessment. In the present study, we examined the clinical utility of a brief handwriting dysfluency measure for quantifying TD. Digitized samples of handwritten circles and loops were obtained from 62 psychosis patients with or without TD and from 50 healthy subjects. Two measures of dysfluent pen movements were extracted from each vertical pen stroke, including normalized jerk and the number of acceleration peaks. Tardive dyskinesia patients exhibited significantly higher dysfluency scores than non-TD patients and controls. Severity of handwriting movement dysfluency was correlated with Abnormal Involuntary Movement Scale severity ratings for some tasks. The procedure yielded high degrees of test-retest reliability. These results suggest that measures of handwriting movement dysfluency may be particularly useful for objectively evaluating the efficacy of pharmacotherapeutic strategies for treating TD.

Discovery of 2-methylpyridine-based biaryl amides as γ-secretase modulators for the treatment of Alzheimer's disease.

γ-Secretase modulators (GSMs) are potentially disease-modifying treatments for Alzheimer's disease. They selectively lower pathogenic Aß42 levels by shifting the enzyme cleavage sites without inhibiting γ-secretase activity, possibly avoiding known adverse effects observed with complete inhibition of the enzyme complex. A cell-based HTS effort identified the sulfonamide 1 as a GSM lead. Lead optimization studies identified compound 25 with improved cell potency, PKDM properties, and it lowered Aß42 levels in the cerebrospinal fluid (CSF) of Sprague-Dawley rats following oral administration. Further optimization of 25 to improve cellular potency is described.