Treatment of Clostridium difficile Infection with a Small-Molecule Inhibitor of Toxin UDP-Glucose Hydrolysis Activity.

Clostridium difficile infection (CDI) is the leading cause of hospital-acquired infectious diarrhea, with significant morbidity, mortality, and associated health care costs. The major risk factor for CDI is antimicrobial therapy, which disrupts the normal gut microbiota and allows C. difficile to flourish. Treatment of CDI with antimicrobials is generally effective in the short term, but recurrent infections are frequent and problematic, indicating that improved treatment options are necessary. Symptoms of disease are largely due to two homologous toxins, TcdA and TcdB, which are glucosyltransferases that inhibit host Rho GTPases. As the normal gut microbiota is an important component of resistance to CDI, our goal was to develop an effective nonantimicrobial therapy. Here, we report a highly potent small-molecule inhibitor (VB-82252) of TcdA and TcdB. This compound inhibits the UDP-glucose hydrolysis activity of TcdB and protects cells from intoxication after challenge with either toxin. Oral dosing of the inhibitor prevented inflammation in a murine intrarectal toxin challenge model. In a murine model of recurrent CDI, the inhibitor reduced weight loss and gut inflammation during acute disease and did not cause the recurrent disease that was observed with vancomycin treatment. Lastly, the inhibitor demonstrated efficacy similar to that of vancomycin in a hamster disease model. Overall, these results demonstrate that small-molecule inhibition of C. difficile toxin UDP-glucose hydrolysis activity is a promising nonantimicrobial approach to the treatment of CDI.

Solution stability of Captisol-stabilized melphalan (Evomela) versus Propylene glycol-based melphalan hydrochloride injection.

PURPOSE: The objective of this study was to compare the stability of recently approved Captisol-stabilized propylene glycol-free melphalan injection (Evomela™) against currently marketed propylene glycol-based melphalan injection. The products were compared as reconstituted solutions in vials as well as admixture solutions prepared from normal saline in infusion bags. METHODS: Evomela and propylene glycol-based melphalan injection were reconstituted in normal saline and organic custom diluent, respectively, according to their package insert instructions. The reconstituted solutions were diluted in normal saline to obtain drug admixture solutions at specific drug concentrations. Stability of the solutions was studied at room temperature by assay of melphalan and determination of melphalan-related impurities. RESULTS: Results show that based on the increase in total impurities in propylene glycol-based melphalan injection at 0.45 mg/mL, Evomela admixture solutions are about 5, 9, 15 and 29 times more stable at concentrations of 0.45, 1.0, 2.0 and 5.0 mg/mL, respectively. Results confirmed that reconstituted Evomela solution can be stored in the vial for up to 1 h at RT or for up to 24 h at refrigerated temperature (2-8 °C) with no significant degradation. After storage in the vial, it remains stable for an additional 3-29 h after preparation of admixture solution in infusion bags at concentrations of 0.25-5.0 mg/mL, respectively. In addition, Evomela solution in saline, at concentration of 5.0 mg/mL melphalan was bacteriostatic through 72 h storage at 2-8 °C. CONCLUSION: Formulation of melphalan with Captisol technology significantly improved stability compared to melphalan hydrochloride reconstituted with propylene-glycol based diluents.

Trichomonas vaginalis metronidazole resistance is associated with single nucleotide polymorphisms in the nitroreductase genes ntr4Tv and ntr6Tv.

Metronidazole resistance in the sexually transmitted parasite Trichomonas vaginalis is a problematic public health issue. We have identified single nucleotide polymorphisms (SNPs) in two nitroreductase genes (ntr4Tv and ntr6Tv) associated with resistance. These SNPs were associated with one of two distinct T. vaginalis populations identified by multilocus sequence typing, yet one SNP (ntr6Tv A238T), which results in a premature stop codon, was associated with resistance independent of population structure and may be of diagnostic value.

Age-appropriate health promotion education: roots firmly established in baccalaureate nursing pediatric rotation.

The Essentials of Baccalaureate Education for Professional Nursing Practice [American Association of Colleges of Nursing. (2008, October 8). Retrieved September 11, 2009, from http://www.aacn.nche.edu/Education/pdf/BaccEssentials08.pdf] addresses the importance of health promotion at the individual level across the lifespan to effect optimal population health. This qualitative study explores senior baccalaureate nursing students' perceptions (n = 85) of the teaching and learning experience related to health promotion during their pediatric clinical rotation. Four distinct learning factors of student, client, learning process, and subject matter emerged. Knowledge was enhanced, and students identified that the format of presentation influenced the quality of participant learning. Students' perception of the importance of patient and family teaching evolved over the course of the semester and highlights the importance of providing students with the tools necessary to be effective teachers and change agents to promote healthy behaviors across the lifespan. Fink's Taxonomy of Significant Learning can be applied to facilitate integrated course design for a pediatric baccalaureate nursing curriculum.

Evaluation of the first year of a pilot program in community pharmacy: HIV/AIDS medication therapy management for Medi-Cal beneficiaries.

BACKGROUND: The advent of combined antiretroviral therapy (ART) has increased treatment effectiveness but created new challenges for patients infected with human immunodeficiency virus (HIV) and for community pharmacists managing patients' drug therapy. The ability of pharmacist-provided medication therapy management (MTM) services to increase medication adherence, improve health outcomes, and reduce overall medical costs has been demonstrated in community pharmacies for chronic diseases such as diabetes and hypertension. However, the effectiveness of pharmacist-provided MTM services in HIV/acquired immune deficiency syndrome (AIDS) has not been well studied. In January 2005, a pilot program to evaluate MTM services for patients with HIV/AIDS began in California, allowing 10 HIV/AIDS specialty pharmacies to receive compensation for the MTM services that they provided to HIV/AIDS patients. OBJECTIVES: To examine the first year of the HIV/AIDS pharmacy MTM compensation pilot program, which described and compared pilot and nonpilot pharmacies with respect to (a) patient characteristics; (b) intermediate outcomes including type and number of ART medication regimens used, rates of adherence and excess medication fills for ART, use of contraindicated ART regimens, and occurrence of opportunistic infections; and (c) pharmacy and medical costs. METHODS: This was a cohort study examining 2005 Medi-Cal pharmacy and medical claims data for patients with HIV/AIDS who were served by pilot pharmacies versus other pharmacies. The HIV/AIDS patients were Medi-Cal beneficiaries aged 18 years or older as of January 1, 2005, who were continuously enrolled from January 1, 2004, through December 31, 2005, and diagnosed with HIV/AIDS, identified by receipt of at least 1 ART prescription and at least 1 medical claim with a diagnosis (primary or secondary) of HIV/AIDS (ICD-9-CM code 042.0) during both the index period (the year before pilot program implementation, 2004) and the intervention period (the study year, 2005). The only difference in the inclusion criteria for the 2 cohorts was that the pilot pharmacy patients were required to have filled 50% or more of their antiretroviral prescriptions in 2005 at 1 of the 10 pilot pharmacies. Adherence was defined as a medication possession ratio (MPR) of 80%-120% and excess medication fills as MPR greater than 120%. Comparisons were made between groups using bivariate statistics (Pearson chi-square for categorical variables and t-tests for continuous variables). For comparisons of costs, generalized linear models assuming a gamma distribution and log link function were used; predictor variables for the models included age, gender, race/ethnicity, and dual coverage under Medicare. RESULTS: A total of 7,018 HIV/AIDS patients in the Medi-Cal population were identified as meeting the study criteria. Of these, 19.3% (n=1,353) were pilot pharmacy patients. The demographic profile of pilot pharmacy patients was similar, but not identical, to that of patients receiving medications at other pharmacies. A larger percentage of pilot pharmacy patients were on protease inhibitor-based ART medication regimens (63.8% vs. 54.8%, P<0.001), remained on a single type of ART therapy throughout the study year (56.8% vs. 34.2%, P<0.001), and were classified as adherent (56.3% vs. 38.1%, P<0.001), compared with other pharmacy patients. Fewer pilot pharmacy patients used contraindicated regimens (11.6% vs. 16.6%, P<0.001) or had excess medication fills (19.7% vs. 44.8%, P<0.001). The rate of opportunistic infections did not differ significantly between groups (28.2% vs. 26.1%, P=0.121). The total mean (standard error) annual health care cost per patient was 10% higher in pilot pharmacies than in other pharmacies ($40,596 [$889] vs. $36,937 [$479], P=0.001); driven by use of (a) medications (primarily non-ART medications) and (b) mental health services. Payment from the California Department of Health Care Services for MTM services averaged $1,014 per pilot pharmacy study patient. CONCLUSION: Study findings for the first year of the MTM program suggest that the pilot pharmacy patients received more appropriate HIV treatment. The degree to which these differences are affected by selfselection of patients into the pilot pharmacies is unknown. Longer-term outcomes and costs of the pilot program will be examined when data for subsequent years are available.

The Fgl2/fibroleukin prothrombinase contributes to immunologically mediated thrombosis in experimental and human viral hepatitis.

Fibrin deposition and thrombosis within the microvasculature is now appreciated to play a pivotal role in the hepatocellular injury observed in experimental and human viral hepatitis. Importantly, the pathways by which fibrin generation is elicited in viral hepatitis may be mechanistically distinct from the classical pathways of coagulation induced by mechanical trauma or bacterial lipopolysaccharide (LPS). In the setting of murine hepatitis virus strain-3 (MHV-3) infection, a member of the Coronaviridae, activated endothelial cells and macrophages express distinct cell-surface procoagulants, including a novel prothrombinase, Fgl2/fibroleukin, which are important for both the initiation and localization of fibrin deposition. To assess the role of Fgl2/fibroleukin in murine viral hepatitis we generated a Fgl2/fibroleukin-deficient mouse. Peritoneal macrophages isolated from Fgl2/fibroleukin-/- mice did not generate a procoagulant response when infected with MHV-3. Fibrin deposition and liver necrosis were markedly reduced, and survival was increased in mice infected with MHV-3. To address the relevance of Fgl2/fibroleukin in human chronic viral hepatitis we studied patients with minimal and marked chronic hepatitis B. We detected robust expression of Fgl2/fibroleukin mRNA transcripts and protein in liver tissue isolated from patients with marked chronic hepatitis B. Fibrin deposition was strongly associated with Fgl2/fibroleukin expression. Collectively, these data indicate a critical role for Fgl2/fibroleukin in the pathophysiology of experimental and human viral hepatitis.

A novel predictive pharmacokinetic/pharmacodynamic model of repolarization prolongation derived from the effects of terfenadine, cisapride and E-4031 in the conscious chronic av node--ablated, His bundle-paced dog.

INTRODUCTION: Terfenadine, cisapride, and E-4031, three drugs that prolong ventricular repolarization, were selected to evaluate the sensitivity of the conscious chronic atrioventricular node--ablated, His bundle-paced Dog for defining drug induced cardiac repolarization prolongation. A novel predictive pharmacokinetic/pharmacodynamic model of repolarization prolongation was generated from these data. METHODS: Three male beagle dogs underwent radiofrequency AV nodal ablation, and placement of a His bundle-pacing lead and programmable pacemaker under anesthesia. Each dog was restrained in a sling for a series of increasing dose infusions of each drug while maintained at a constant heart rate of 80 beats/min. RT interval, a surrogate for QT interval in His bundle-paced dogs, was recorded throughout the experiment. RESULTS: E-4031 induced a statistically significant RT prolongation at the highest three doses. Cisapride resulted in a dose-dependent increase in RT interval, which was statistically significant at the two highest doses. Terfenadine induced a dose-dependent RT interval prolongation with a statistically significant change occurring only at the highest dose. The relationship between drug concentration and RT interval change was described by a sigmoid E(max) model with an effect site. Maximum RT change (E(max)), free drug concentration at half of the maximum effect (EC(50)), and free drug concentration associated with a 10 ms RT prolongation (EC(10 ms)) were estimated. A linear correlation between EC(10 ms) and HERG IC(50) values was identified. DISCUSSION: The conscious dog with His bundle-pacing detects delayed cardiac repolarization related to I(Kr) inhibition, and detects repolarization change induced by drugs with activity at multiple ion channels. A clinically relevant sensitivity and a linear correlation with in vitro HERG data make the conscious His bundle-paced dog a valuable tool for detecting repolarization effect of new chemical entities.

Utilization of molecular methods to identify prognostic markers for recurrent bacterial vaginosis.

BACKGROUND: Recurrent bacterial vaginosis (BV) after antimicrobial therapy is a major problem, affecting >50% of patients within 1 year. The objective of this study was to determine if prospective identification of patients at risk for recurrence using molecular methods is feasible. METHODS: Women were evaluated for BV by Amsel criteria and Nugent score. Vaginal specimens were analyzed using a panel of quantitative real-time polymerase chain reactions (qPCRs) at three times: pre-treatment, 7-10days post-treatment and 40-45days post-treatment. The PCRs quantified DNA of the following organisms: Gardnerella vaginalis; Atopobium vaginae; Bacterial Vaginosis-Associated Bacteria-1 (BVAB1), -2 (BVAB2) and -3 (BVAB3); Leptotrichia/Sneathia; Megasphaera Phylotypes 1 and 2; and Lactobacillus spp. (L. crispatus, L. gasseri, L. iners and L. jensenii). RESULTS: Out of 84 women diagnosed with BV (Amsel ≥3 and Nugent ≥4), 77 (91.7%) were successfully treated after 7-10days (asymptomatic and Amsel of either 0 or 1 with elevated vaginal pH and Nugent ≤6). Of these 77 women, 46 (59.7%) remained cured after 40-45days and 31 (40.3%) developed recurrent BV. In univariate analysis, we found that women who would have recurrent BV during the study had greater concentrations of Megasphaera Phylotype 2 (P=0.001) and BVAB2 (P=0.015) at initial diagnosis and greater vaginal pH (P=0.030), higher Nugent score (P=0.043) and a greater concentration of G. vaginalis (P=0.012) post-treatment, when compared to women who were cured during the study. These differences largely remained when cure was defined as Nugent ≤3 or when only women treated with intravaginal metronidazole were evaluated. CONCLUSION: Molecular analysis of BV is a useful adjunct to clinical and microscopic analysis to prospectively identify patients at high risk for recurrent BV.

Neuropsychiatric effects of guanfacine in children with mild tourette syndrome: a pilot study.

The objective of this study was to evaluate the neuropsychiatric effects of the alpha-2a adrenergic agonist guanfacine in children with Tourette syndrome (TS). Twenty-four children with TS participated in a 4-week, double-blind, placebo-controlled study of guanfacine. Tic severity, neuropsychologic functioning, and parent ratings of behavior were evaluated pre- and post-treatment. The sample had mild tic severity and subtle neuropsychologic dysfunction pretreatment. Post-treatment, patients receiving guanfacine were rated by parents as significantly improved (compared to placebo) on one measure of executive function (parent-rated metacognition). Improvement on tic severity, performance-based neuropsychologic measures, and all other parent ratings were not significantly better than placebo. At a moderate dose and short-term treatment duration, guanfacine did not provide significant neuropsychiatric benefits in this group of children with mild TS.

Professional development needs of registered nurses practicing at a military community hospital.

BACKGROUND: Focus groups were used to determine what factors need to be considered when designing a professional development program for RNs practicing at a military community hospital. METHOD: Six focus groups were formed with RNs from multiple directorates and settings within various specialties. FINDINGS: Five thematic groupings were identified: 1) specific professional development needs for leadership, clinical/specialty practice, competence development, and maintenance, 2) methods for providing continuing education, 3) methods for evaluating the effectiveness and efficiency of continuing education, 4) barriers to professional development, and 5) professional development issues impacting retention of military nurses. CONCLUSION: The professional development needs of RNs practicing in military hospitals are complex and multifaceted. The thematic groupings identified guided the design of a professional development program for RNs.

Clinical Escherichia coli isolates utilize alpha-hemolysin to inhibit in vitro epithelial cytokine production.

Uropathogenic Escherichia coli is the primary cause of urinary tract infections, which affects over 60% of women during their lifetime. UPEC exhibits a number of virulence traits that facilitate colonization of the bladder, including inhibition of cytokine production by bladder epithelial cells. The goal of this study was to identify the mechanism of this inhibition. We observed that cytokine suppression was associated with rapid cytotoxicity toward epithelial cells. We found that cytotoxicity, cytokine suppression and alpha-hemolysin production were all tightly linked in clinical isolates. We screened a UPEC fosmid library and identified clones that gained the cytotoxicity and cytokine-suppression phenotypes. Both clones contained fosmids encoding a PAI II(J96)-like domain and expressed the alpha-hemolysin (hlyA) encoded therein. Mutation of the fosmid-encoded hly operon abolished cytotoxicity and cytokine suppression. Similarly, mutation of the chromosomal hlyCABD operon of UPEC isolate F11 also abolished these phenotypes, and they could be restored by introducing the PAI II(J96)-like domain-encoding fosmid. We also examined the role of alpha-hemolysin in cytokine production both in the murine UTI model as well as patient specimens. We conclude that E. coli utilizes alpha-hemolysin to inhibit epithelial cytokine production in vitro. Its contribution to inflammation during infection requires further study.

Antiretroviral therapy adherence, medication use, and health care costs during 3 years of a community pharmacy medication therapy management program for Medi-Cal beneficiaries with HIV/AIDS.

BACKGROUND: The types of pharmacist-provided medication therapy management (MTM) services provided to patients with human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS) and the effects of MTM on medication adherence and patient outcomes have only recently begun to be studied. Although available studies suggest that patients receiving MTM services have better antiretroviral therapy (ART) adherence and outcomes, only 1 study has examined a large group of patients with HIV/AIDS, and none has examined adherence or outcomes for more than 1 year. A pilot program conducted by the California Department of Health Care Services (DHCS) and Medi-Cal (California's Medicaid program) provided an opportunity to examine ART adherence and outcomes in a large patient population receiving MTM services in community pharmacies over 3 years. OBJECTIVES: To examine an HIV/AIDS pharmacy MTM compensation pilot program over a 3-year period (2005- 2007) in a sample of Medi-Cal beneficiaries by describing the associations between use of pilot pharmacies and (a) adherence to ART regimens; (b) medication utilization, including number and type of ART medication regimens and use of contraindicated ART regimens; (c) occurrence of opportunistic infections; and (d) all-cause pharmacy and medical costs. METHODS: This was a cohort study examining Medi-Cal pharmacy and medical claims data (2005-2007) for patients with HIV/AIDS who were served by pilot pharmacies versus other (nonpilot) pharmacies. The study groups, pilot and nonpilot pharmacy patients with HIV/AIDS, consisted of Medi-Cal beneficiaries aged 18 years or older as of January 1, 2005, who were continuously enrolled from January 1, 2004, through December 31, 2007, and who received both a diagnosis of HIV/AIDS and at least 1 ART pharmacy claim during both the index period (2004) and the study period (January 1, 2005, through December 31, 2007). Pilot pharmacy patients were identified as having filled 50% or more of their ART prescriptions each year at 1 of the 10 pilot pharmacies. Patients for whom comprehensive medication data were not available, including those enrolled in managed care plans and/or Medicare, were excluded. Adherence was defined as a medication possession ratio (MPR) of 80%-120% and excess medication fills as MPR greater than 120%. Logistic regression was used to investigate the factors associated with adherence. Comparisons were made between groups using bivariate statistics (Pearson chi-square for categorical variables and t-tests for continuous variables). For comparisons of costs, generalized linear models were used including predictor variables for age, gender, and race/ethnicity. RESEARCH RESULTS: The study sample consisted of 2,234 patients meeting the study inclusion criteria. The proportion of study patients receiving the majority of their prescription medications (ART plus non-ART) at pilot pharmacies was 19.7% in 2005 and increased to 27.6% in 2006 and 28.1% in 2007. The demographic profile of pilot pharmacy patients was similar to that of patients receiving medications at nonpilot pharmacies, except that pilot pharmacies had a higher proportion of Latino patients (e.g., 19.7% vs. 14.9% in 2007, respectively, P = 0.006). A greater percentage of pilot than nonpilot pharmacy patients were adherent to their ART medication regimens (e.g., 2007: 69.4% vs. 47.3%, respectively, P < 0.001). After controlling for age, gender, and ethnicity/race in logistic regression analysis, use of a pilot pharmacy (odds ratio [OR] = 2.74, 95% CI = 2.44-3.10) was the most important factor associated with likelihood of adherence. Each year, pilot pharmacy patients were more likely than nonpilot pharmacy patients to remain on a single type of ART regimen (e.g., 2007: 71.7% vs. 49.1%, respectively, P < 0.001) and less likely to have excess fills (e.g., 2007: 12.9% vs. 35.5%, respectively, P < 0.001) and to use contraindicated regimens (e.g., 2007: 8.9% vs. 12.2%, respectively, P = 0.027). The percentages of patients experiencing opportunistic infections were similar between groups each year, approximately 35% (P = 0.809-0.945). In the generalized linear model analyses, the between-group differences in predicted mean (standard error [SE]) total health care costs per patient were not significantly different in any year (e.g., 2007: $38,983 [$1,023] vs. $38,856 [$633], respectively, P = 0.915). In each year, predicted non- ART medication costs were approximately 30%-40% greater in the pilot pharmacy than nonpilot pharmacy group (e.g., 2007: $10,815 [$538] vs. $8,190 [$252], respectively, P < 0.001); however, predicted expenditures for inpatient services were significantly lower (e.g., 2007: $3,083 [$293] vs. $5,186 [$300], respectively, P < 0.001). Payment from the DHCS Medi-Cal program for MTM services was approximately $1,000 per pilot pharmacy patient per year. CONCLUSIONS: Over a 3-year period, patients at pilot pharmacies consistently had higher medication adherence rates, were more likely to remain on a single type of ART regimen throughout the year, had fewer excess fills, and used fewer contraindicated regimens than nonpilot pharmacy patients. There were no significant differences in mean total cost per patient per group, and the additional MTM services payment added less than 3% to the total cost.

Medication therapy management services in community pharmacy: a pilot programme in HIV specialty pharmacies.

RATIONALE, AIMS AND OBJECTIVES: Pharmacist-provided medication therapy management services (MTMS) have been shown to increase patient's adherence to medications, improve health outcomes and reduce overall medical costs. The purpose of this study was to describe a pilot programme that provided pharmacy-based MTMS for patients with HIV/AIDS in the state of California, USA. METHOD: Pharmacists from the 10 pilot pharmacies were surveyed using an online data collection tool. Information was collected to describe the types of MTMS offered, proportion of patients actively using specific MTMS, pharmacist beliefs regarding effect on patient outcomes and barriers to providing MTMS, ability to offer MTMS without pilot programme funding and specialized pharmacist or staff training. RESULTS: Each responding pharmacy (7 of 10) varied in the number of HIV/AIDS patients served and prescription volume. All pharmacists had completed HIV/AIDS-related continuing education programmes, and some had other advanced training. The type of MTMS being offered varied at each pharmacy with 'individualized counselling by a pharmacist when overuse or underuse was detected' and 'refill reminders by telephone' being actively used by the largest proportion of patients. Most, but not all, pharmacists cited reimbursement as a barrier to MTMS provision. Pharmacists believed the MTMS they provide resulted in improved satisfaction (patient and provider), medication usage, therapeutics response and patient quality of life. CONCLUSION: The type of MTMS offered, and proportion of patients actively using, varied among participating pilot pharmacies.

Diels-Alder reactions of N-acyl-2-alkyl(aryl)-5-vinyl-2,3-dihydro-4-pyridones.

Readily available N-acyl-5-vinyl-2,3-dihydro-4-pyridones undergo Diels-Alder cyclization with various dienophiles to afford novel octahydroquinolines containing synthetically useful functionality. With dihydropyridone 5 and cis-disubstituted dienophiles, the resulting cycloadducts were obtained as single diastereomers in good to excellent yield. The corresponding reaction of 5 with methyl acrylate, acrylonitrile, and phenyl vinyl sulfone showed modest preference for the endo adducts. The effect of the dihydropyridone C-2 and C-4 substituents on the degree of diastereofacial control was examined. By using this methodology, the core decahydroquinoline skeleton of gephyrotoxin was prepared in a stereocontrolled fashion. Interesting reactivity was observed with certain dienophiles leading to ring-opening of the initially formed cycloadducts. This tandem reaction provides a route to uniquely substituted beta-aminoketones, alcohols, and unnatural amino acids.

Interactions of 1263W94 with other antiviral agents in inhibition of human cytomegalovirus replication.

Inhibition of human cytomegalovirus (HCMV) by 1263W94 was additive dosewise in combination with ganciclovir, acyclovir, and foscarnet. None of the commonly used anti-human immunodeficiency virus agents antagonized the inhibition of HCMV by 1263W94. The data were analyzed by a modified isobologram procedure that measures the strength and statistical significance of drug interactions.