Expression and purification of native and functional influenza A virus matrix 2 proton selective ion channel.

Influenza A virus displays one of the highest infection rates of all human viruses and therefore represents a severe human health threat associated with an important economical challenge. Influenza matrix protein 2 (M2) is a membrane protein of the viral envelope that forms a proton selective ion channel. Here we report the expression and native isolation of full length active M2 without mutations or fusions. The ability of the influenza virus to efficiently infect MDCK cells was used to express native M2 protein. Using a Calixarene detergents/surfactants based approach; we were able to solubilize most of M2 from the plasma membrane and purify it. The tetrameric form of native M2 was maintained during the protein preparation. Mass spectrometry shows that M2 was phosphorylated in its cytoplasmic tail (serine 64) and newly identifies an acetylation of the highly conserved Lysine 60. ELISA shows that solubilized and purified M2 was specifically recognized by M2 antibody MAB65 and was able to displace the antibody from M2 MDCK membranes. Using a bilayer voltage clamp measurement assay, we demonstrate a pH dependent proton selective ion channel activity. The addition of the M2 ion channel blocker amantadine allows a total inhibition of the channel activity, illustrating therefore the specificity of purified M2 activity. Taken together, this work shows the production and isolation of a tetrameric and functional native M2 ion channel that will pave the way to structural and functional characterization of native M2, conformational antibody development, small molecules compounds screening towards vaccine treatment.

Major depressive disorder, personality disorders, and coping strategies are independent risk factors for lower quality of life in non-metastatic breast cancer patients.

OBJECTIVE: Our aim was to identify risk factors for lower quality of life (QOL) in non-metastatic breast cancer patients. METHODS: Our study included 120 patients from the University Hospital Centers of Tours and Poitiers. This cross-sectional study was conducted 7 months after patients' breast cancer diagnosis and assessed QOL (Quality of Life Questionnaire Core 30 = QLQ-C30), socio-demographic characteristics, coping strategies (Brief-COPE), physiological and biological variables (e.g., initial tumor severity and types of treatment received), the existence of major depressive disorder (Mini International Neuropsychiatric Interview), and pain severity (Questionnaire de Douleur Saint Antoine). We assessed personality disorders 3 months after diagnosis (Vragenlijst voor Kenmerken van de Persoonlijkheid questionnaire). We used multiple linear regression models to determine which factors were associated with physical, emotional, and global QOL. RESULTS: Lower physical QOL was associated with major depressive disorder, younger age, a more severe initial tumor stage, and the use of the behavioral disengagement coping. Lower emotional QOL was associated with major depressive disorder, the existence of a personality disorder, a more severe pain level, higher use of self-blame, and lower use of acceptance coping strategies. Lower global QOL was associated with major depressive disorder, the existence of a personality disorder, a more severe pain level, higher use of self-blame, lower use of positive reframing coping strategies, and an absence of hormone therapy. CONCLUSIONS: Lower QOL scores were more strongly associated with variables related to the individual's premorbid psychological characteristics and the manner in which this individual copes with the cancer (e.g., depression, personality, and coping) than to cancer-related variables (e.g., treatment types and cancer severity). Copyright © 2015 John Wiley & Sons, Ltd.

Self-Monitoring of Blood Glucose and Hypoglycemia-Related Hospitalization in a Population-Based Cohort of Canadian Patients With Type 1 or Type 2 Diabetes.

OBJECTIVES: The aim of this work was to determine whether: 1) blood glucose test strip use in the population is associated with hypoglycemia hospitalization rates, and 2) blood glucose test strip use among individuals is associated with a reduced risk of hypoglycemia hospitalization. METHODS: Administrative databases from Saskatchewan, Canada, were used to ascertain population-level hypoglycemia hospitalizations and test strip utilization over the period from 1996 to 2014. For objective 1, a generalized linear model with generalized estimating equations was fit to provincial data stratified by age group, sex and year. For objective 2, a nested case-control study was conducted for a cohort of insulin users with diagnosed diabetes. Multivariable conditional logistic regression was used to test the association of test strip use with hospitalization, after adjusting for clinical and demographic factors and health services use. Odds ratios (ORs) and 95% confidence intervals (95% CIs) are reported. RESULTS: A total of 5,166 hospitalizations for hypoglycemia were identified in the observation period. Annual glucose test strip use increased by over 350%; however, no association was found with provincial hypoglycemia hospitalization rate during the same period, even after controlling for all-cause hospitalizations and population demographics. In the case-control analysis, test strip use was not associated with hospitalization for hypoglycemia among insulin users (n=10,617; adjusted OR, 1.08; 95% CI, 0.88 to 1.31). A sensitivity analysis in an independent cohort of noninsulin users produced a similar finding (n=47,501; adjusted OR, 1.04; 95% CI, 0.55 to 1.94). CONCLUSION: Our findings add to the body of evidence against a protective effect of blood glucose test strip use for serious hypoglycemia.

Novel calixarene-based surfactant enables low dose split inactivated vaccine protection against influenza infection.

Influenza A viruses cause major morbidity and represent a severe global health problem. Current influenza vaccines are mainly egg-based products requiring the split of whole viruses using classical detergents such as Triton X-100, which implies certain limitations. Here, we report the use of the novel calixarene-based surfactant CALX133ACE as an alternative to classical detergents for influenza inactivated split vaccine preparation. We confirmed that CALX133ACE-based split HA antigens are fully functional and quantifiable by the "gold standard" method SRID. Additionally, as in the case of the Triton X-100-based split, the CALX133ACE-based split antigens are stable for at least 6 months at 4 °C. Moreover, immunization of mice with CALX133ACE-based split NYMC X-179A (H1N1) antigens harboring 10 to 30-fold less antigen than the commercialized trivalent inactivated vaccines Vaxigrip® or Fluviral® induced comparable efficient protection and neutralizing antibody responses against A(H1N1)pdm09 infection. Taken together, our results demonstrate for the first time the use of a calixarene-based detergent as an efficient splitting agent for the production of optimized influenza split antigens, paving the way for significant improvement in the vaccine manufacturing process, notably with regard to the current regulation on the prohibition of endocrine disruptors, such as Triton X-100.

Trends in Blood Glucose Test Strip Utilization: A Population-Wide Analysis in Saskatchewan, Canada.

OBJECTIVES: To describe trends in blood glucose test strip (TS) utilization and cost in Saskatchewan. METHODS: A retrospective analysis of TS use between January 1, 1996, and December 31, 2013, was conducted using population-based health administrative databases in Saskatchewan. The prescription drug database was used to describe the annual number of TS dispensations, the number of strips dispensed, the number of unique beneficiaries and the total costs. A patient-level analysis was also carried out to describe the patterns of TS use (i.e. light, moderate or heavy) by the entire cohort and by diabetes treatments. Potential cost savings due to a newly implemented restriction policy were estimated based on the most recent data (2013). RESULTS: TS utilization increased dramatically between 1996 and 2013 in terms of the number of users and the average number of TSs received. The percentage of TS users receiving fewer than 4 TSs per week (i.e. light users) decreased by 20%, while the percentage of heavy users (i.e. those receiving more than 8 TSs per week) increased by 19%. During the same period, the use of high-risk oral hypoglycemic medications declined by 30% among all TS users. Heavy TS use was observed in at least one-third of all users, irrespective of treatment type. CONCLUSIONS: If Saskatchewan's newly imposed coverage limits had been applied in 2013, the costs of strips exceeding those limits would have totalled $2.5 million. Although TS use aligns with chronic disease care paradigms, the substantial costs and lack of evidence of patient outcomes demand better strategies to help reduce unnecessary use.

Personality disorders, but not cancer severity or treatment type, are risk factors for later generalised anxiety disorder and major depressive disorder in non metastatic breast cancer patients.

This study aimed to determine whether personality disorders were associated with later Major Depressive Disorder (MDD) or Generalised Anxiety Disorder (GAD) in breast cancer patients. This longitudinal and multicentric study included 120 French non-metastatic breast cancer patients. After cancer diagnosis (T1) and 7 months after diagnosis (T3), we assessed MDD and GAD (Mini International Neuropsychiatric Interview 5.0). We assessed personality disorders 3 months after diagnosis (VKP). We used multiple logistic regression analysis to determine what were the factors associated with GAD and MDD at T3. At T3, prevalence rate was 10.8% for MDD and 19.2% for GAD. GAD at T3 was significantly and independently associated with GAD at T1 and with existence of a personality disorder, no matter the cluster type. MDD at T3 was significantly and independently associated with MDD at T1 and with the existence of a cluster C personality disorder. Initial cancer severity and the type of treatment used were not associated with GAD or MDD at T3. Breast cancer patients with personality disorders are at higher risk for GAD and MDD at the end of treatment. Patients with GAD should be screened for personality disorders. Specific interventions for patients with personality disorders could prevent psychiatric disorders.

Community Pharmacists Assisting in Total Cardiovascular Health (CPATCH): A Cluster-Randomized, Controlled Trial Testing a Focused Adherence Strategy Involving Community Pharmacies.

STUDY OBJECTIVE: To test a brief intervention for preventing statin nonadherence among community pharmacy patrons. DESIGN: Prospective, cluster-randomized, controlled trial (the Community Pharmacists Assisting in Total Cardiovascular Health [CPATCH] trial). SETTING: Thirty community pharmacies in Saskatchewan, Canada. PATIENTS: Participating pharmacies were randomized to 15 intervention pharmacies where a brief statin adherence intervention was delivered by pharmacists (intervention group [907 patients]) or 15 usual care pharmacies where no statin adherence intervention was delivered (usual care group [999 patients]) to new users of statins (defined as less than 1 yr of statin therapy). INTERVENTION: Staff (pharmacy managers, staff pharmacists, and technicians) from intervention pharmacies attended a 2.5-hour workshop on the CPATCH program that prepared pharmacists to deal with the adherence barriers most likely associated with statin use (e.g., safety, cost, patient-provider relationship, and tolerability). Intervention pharmacists screened for new statin users and assessed these adherence barriers. Pharmacists were then instructed to tailor their follow-up plan based on the individual patient's situation. Investigators contacted the intervention pharmacies monthly to assess their compliance with the protocol and to offer additional support to motivate ongoing participation. MEASUREMENTS AND MAIN RESULTS: The primary outcome was mean difference in statin adherence between the intervention and usual care groups. Adherence was measured by the proportion of days covered (PDC) between 6 and 12 months following the original prescription fill date. General estimating equations were used to evaluate the difference in mean adherence between groups. Secondary outcomes included the percentage of new statin users exhibiting optimal adherence (defined as PDC of 80% or higher) and the percentage exhibiting nonpersistence (defined as the cessation of all statin dispensations within 3 mo of the first dispensation). Among 1906 eligible patients, no significant differences in mean adherence were observed between those receiving the intervention and those receiving usual care (71.6% vs 70.9%, p=0.64), the percentage of patients achieving optimal adherence (57.3% vs 55.9%, p=0.51), or the percentage exhibiting nonpersistence (9.4% vs 8.3%, p=0.41). However, compliance to the study protocol was extremely low in several intervention pharmacies. In a post hoc analysis, a higher level of protocol compliance among intervention pharmacies was significantly associated with higher adherence (p<0.01 for trend). Pharmacies falling in the highest tertile of compliance to the study protocol exhibited higher mean adherence among their patients compared with those in the usual care group (ß = 0.056, 95% confidence interval [CI] 0.010-0.101, p=0.01), and a significantly higher percentage of patients achieving optimal adherence (odds ratio 1.32, 95% CI 1.08-1.61; p<0.01); however, nonpersistence did not significantly differ between the two groups (5.5% vs 8.3%, p=0.27). CONCLUSION: The CPATCH intervention was ineffective for improving patient adherence to statin therapy in community pharmacies. However, poor effectiveness may have resulted from a failure to deliver the protocol consistently in several intervention pharmacies.

Trends in prevalence, incidence and pharmacologic management of diabetes mellitus among seniors newly admitted to long-term care facilities in Saskatchewan between 2003 and 2011.

OBJECTIVE: We aimed to describe trends in the prevalence and incidence of diabetes mellitus and also report the overall use of diabetes medications among patients newly admitted to a long-term care facility (LTCF). METHODS: A retrospective cohort study was done using health administrative databases in Saskatchewan. Eligible patients were newly admitted to LTCF in Saskatchewan between 2003 and 2011 and maintained LTCF residency for at least 6 months. Prevalence of diabetes was defined with physician or hospital claims in the 2 years preceding admission. Antihyperglycemic medication use was estimated from prescription claims data during the first 6 months after LTCF admission. All data were descriptively analyzed. RESULTS: The validated case definition for diabetes (≥2 diagnostic claims) in the 2 years before or 6 months after admission was met by 16.9% of patients (2471 of 14,624). An additional 965 patients (6.6%) had a single diabetes diagnostic claim or antihyperglycemic prescriptions only. Among patients receiving antihyperglycemic therapies, 64.9% (1518 of 2338) were exclusively managed with oral medications, and metformin was the most commonly used medication. Glyburide was commonly withdrawn after LTCF admission. Insulin use was observed in 23.9% of diabetes patients, with a mean daily average consumption of 54.7 units per day. CONCLUSIONS: Use of diabetes medications appear to generally align with Canadian practice recommendations as evidenced by declining use of glyburide and frequent use of metformin. Future studies should examine clinical benefits and safety of hypoglycemic agent use in LTCFs.

Do prior authorization policies discourage first-line antipsychotic use in patients newly discharged from a hospitalization for schizophrenia in Saskatchewan?

BACKGROUND: Drug benefit providers can decrease prescribing of specific medications through prior authorization policies. In Saskatchewan, certain second generation antipsychotics (SGAs) are recognized as first-line agents to manage schizophrenia; but, require prior authorization because their coverage is restricted in other conditions. We aimed to determine if the need for prior-authorization substantially diminishes prescribing of first-line SGAs in comparison to unrestricted agents. OBJECTIVES: To conduct an ecological comparison of SGA prescribing with changes in prior- authorization policies between 1997 and 2005 using health-administrative databases in Saskatchewan, Canada. METHOD: Eligible subjects were discharged from hospital with a first-time primary diagnosis of schizophrenia between 1997 and 2005. SGAs dispensed within 7 days of discharge were used to estimate prescribing preferences for olanzapine and quetiapine relative to risperidone. Percentages of SGA use were age and sex standardized to the 2000 cohort. RESULTS: Out of 1,277 eligible patients, 521 (41%) received 564 SGA dispensations within 7-days of hospital discharge. Between 1997 and 1998, risperidone was the only SGA covered for first-line use and made up 72.6% (82/113) of SGA use while olanzapine made up 27.4% (31/113) for a crude preference ratio of 0.38 (27.4/72.6). Risperidone use decreased to 65.8% in 1999-2002 and to 47.4% in 2003-2005 as a percentage of SGA dispensations. Correspondingly, the preference ratios for olanzapine and quetiapine increased from 0.40 to 0.57 and from 0.12 to 0.54 in these respective periods. CONCLUSIONS: The requirement for prior-authorization does not appear to substantially diminish prescribing of first-line SGAs for the treatment of schizophrenia in Saskatchewan, Canada.