Mitigating base-catalysed degradation of periodate-oxidized capsular polysaccharides: Conjugation by reductive amination in acidic media

Reductive amination coupling an aldehyde-containing polysaccharide, generated by periodate oxidation, with the amino groups in protein has been widely used in the synthesis of glycoconjugate vaccines. The conjugation is often achieved under slightly basic conditions via a Schiff’s base intermediate followed by its reduction with sodium cyanoborohydride. We observed that oxidized capsular polysaccharides such as Streptococcus pneumoniae type 6B (Pn-6B) and Haemophilus influenzae type a (HiA) underwent significant degradation during the conjugation in slightly basic media leading to sub-optimal glycoconjugates. Further study on oxidized Pn-3, Pn-6A, Pn-6C, Pn-2 polysaccharides and dextran provided evidence that the degradation is a result of base-catalysed β-elimination. In contrast to HiA, Pn-2, Pn-3, Pn-6B polysaccharides and dextran, oxidized Pn-6A and Pn-6C polysaccharides were stable under basic conditions due to lack of the leaving group at the β-position of the aldehyde. By performing conjugation of oxidized polysaccharides to bovine serum albumin (BSA) in phosphate buffer at pH 6.0, 6.8, 7.2 and 8.0, we concluded that the reductive amination proceeds best in slightly acidic media, particularly with those β-elimination susceptible polysaccharides.     

Half blind superiority tests for clinical trials of anti-infective drugs

This paper introduces a test of superiority of new anti-infective drug B over comparator drug A based on a randomized clinical trial. This test can be used to demonstrate assay (trial) sensitivity for noninferiority trials and rigorously tailor drug choice for individual patients. Our approach uses specialized baseline covariates X_A,X_B, which should predict the benefits of drug A and drug B, respectively. Using a response surface model for the treatment effect, we test for superiority at the (X_A,X_B) point that is most likely to show superiority. We identify this point based on estimates from a novel half-blind pseudo likelihood, where we augment a blinded likelihood (mixed over the treatment indicator) with likelihoods for the overall success rates for drug A and drug B (mixed over X_A,X_B). The augmentation results in much better estimates than those based on the mixed blinded likelihood alone but, interestingly, the estimates almost behave as if they were based on fully blinded data. We also develop an analogous univariate method using X_A for settings where X_B has little variation. Permutation methods are used for testing. If the “half-blind” test rejects, pointwise confidence interval can be used to identify patients who would benefit from drug B. We compare the new tests to other methods with an example and via simulations.     

Impact of bromocriptine-QR therapy on cardiovascular outcomes in type 2 diabetes mellitus subjects on metformin

Objectives: Type 2 diabetes mellitus (T2DM) is associated with a substantially increased risk of cardiovascular disease (CVD). Bromocriptine-QR (B-QR), a quick release sympatholytic dopamine D₂ receptor agonist, is a FDA-approved therapy for T2DM which may provide CVD risk reduction. Metformin is considered to be an agent with a potential cardioprotective benefit. This large placebo controlled clinical study assessed the impact of B-QR addition to existing metformin therapy on CVD outcomes in T2DM subjects.

Methods: 1791 subjects (1208 B-QR; 583 placebo) on metformin ± another anti-diabetes therapy at baseline derived from the Cycloset Safety Trial, a 12-month, randomized, multicenter, placebo-controlled, double-blind study in T2DM, were included in this study. The primary CVD endpoint evaluated was treatment impact on CVD event rate, prespecified as a composite of time to first myocardial infarction, stroke, coronary revascularization, or hospitalization for unstable angina/congestive heart failure. Impact on glycemic control was evaluated as a secondary analysis.

Results: The composite CVD end point occurred in 16/1208 B-QR treated (1.3%) and 18/583 placebo treated (3.1%) subjects resulting in a 55% CVD hazard risk reduction (intention-to-treat, Cox regression analysis; HR: 0.45 [0.23–0.88], p = 0.028). Kaplan-Meier curves demonstrated a significantly lower cumulative incidence rate of the CVD endpoint in the B-QR treatment group (Log-Rank p = 0.017). In subjects with poor glycemic control (HbA1c ≥ 7.5) at baseline, B-QR therapy relative to placebo resulted in a significant mean %HbA1c reduction of ?0.59 at week 12 and ?0.51 at week 52 respectively (p < 0.001 for both) and a 10 fold higher percent of subjects achieving HbA1c goal of ≤7% by week 52 (B-QR 30%, placebo 3%; p = 0.003).

Conclusion: These findings suggest that in T2DM subjects on metformin, BQR therapy may represent an effective strategy for reducing CVD risk.

Cycloset Safety Trial registration: ClinicalTrials.gov Identifier: NCT00377676.     

Shared Cognitive Features of Posttraumatic Cognitions and Obsessive–Compulsive Symptoms

Obsessive–compulsive disorder (OCD) and posttraumatic stress disorder (PTSD) frequently co-occur. However, the shared features of these conditions have been under-examined. Evaluation of the common aspects of posttraumatic and obsessive–compulsive (OC) symptoms could improve treatment responsivity for individuals with comorbid PTSD and OCD, for whom outcome is typically poorer than for those with either disorder alone. This study examined intolerance of uncertainty, inflated responsibility, and a global measure of posttraumatic cognitions as potential shared cognitive constructs that moderate distress associated with OC symptoms. A total of 211 undergraduate students reporting significant trauma histories participated. All participants completed measures of obsessive–compulsive symptoms and beliefs, as well as posttraumatic cognitions. Results indicated that posttraumatic cognitions moderated the relationship between inflated responsibility and intolerance of uncertainty, which in turn predicted all domains of obsessive–compulsive symptom distress (all βs > 0.41, all zs > 3.44). Further, posttraumatic cognitions alone significantly predicting OC symptoms related to doubting, obsessions, and neutralizing. These findings suggest that shared cognitive constructs play a role in co-occurring posttraumatic stress and OC symptoms, and thus may be a relevant treatment target when these disorders present simultaneously.     

Molecular epidemiology and prevalence of macrolide efflux genes mef(A) and mef(E) in Streptococcus pneumoniae obtained in Canada from 1997 to 2002

One hundred forty M phenotype Streptococcus pneumoniae isolates were evaluated by PCR-restriction fragment length polymorphism, serotyping, and pulsed-field gel electrophoresis. Molecular genotyping revealed that the predominant macrolide resistance mechanism in S. pneumoniae in Canada is mef(E) and resistance dissemination is due to both spread of the genetic element MEGA as well as clonal dissemination of penicillin- and/or macrolide-resistant strains.     

FURTHER STUDIES ON THE OPHTHALMO-TUBERCULIN REACTION IN CATTLE

Our conclusions differ in a few points from those of our first work on this subject. We are able to sum up our observations in this series of experiments as follows: 1. The ophthalmo-tuberculin test is of limited value in the diagnosis of tuberculosis in cattle. In some cases the reaction is very slight (hyperæmia). In others more pronounced congestion with profuse exudates are noted. Accuracy of observation is important. We are inclined to rely primarily on the results of the first instillation of tuberculin. Second instillations in a few instances elicit reaction in non-tubercular animals. 2. In the majority of animals tested the reaction increased in its intensity with each subsequent instillation of tuberculin. This fact indicates the development of a local hypersusceptibility or anaphylaxis associated with a partial immunity; von Pirquet calls this condition "allergie" (9). 3. It is possible in some cases to create a condition of "allergie" in healthy cattle, when spaced instillations of tuberculin are made. It is evident, therefore, that the result of the first instillation of tuberculin should be made the only basis of diagnosis. Rosenau and Anderson (II) have recently called attention to this point in regard to the human subject. 4. When repeated instillations of tuberculin are made on the conjunctiva at short intervals (twenty-four hours, etc.) a local immunity results (No. 3D et al.). If the instillations are separated two weeks or more anaphylaxis results. 5. We, therefore, hold that if tuberculin (0.1 cubic centimeter) is carefully instilled into the conjunctival sac and if careful comparison of the instilled eye with the opposite eye shows that a reaction of varying intensity results in from ten to twelve hours after the first instillation, a tubercular lesion is present. 6. In our first report (7) we were inclined to believe that subcutaneous tubercular injection given previous to the ocular test would slightly inhibit it. We have since become convinced that this is true only to a limited extent, and that in some cases the ophthalmo-reaction is exaggerated by a subcutaneous injection of tuberculin. 7. The primary ophthalmo-tuberculin reaction is in direct proportion to the extent of the tubercular processes in the body. The more extensive the tubercular processes, the more anaphylactic the animal is. This is in direct variance with the condition in the usual subcutaneous tuberculin test. (See Necropsy Report.) 8. We are inclined to believe that the ophthalmo-tuberculin test will reveal tuberculosis at as early a state as the usual subcutaneous test. 9. The ophthalmo-reaction is of no value in determining whether vaccinated cattle are actively tubercular or not, or in demonstrating any hypersusceptibility in the offspring of tubercular cattle. 10. The cutaneous test from our brief series of experiments does not seem to be as accurate as the ophthalmic test. This conclusion has been reached by several investigators.