Search

Abstract: PedvaxHIB is a pediatric vaccine that protects children from severe disease caused by the gram-negative bacterium Haemophilus influenzae type b (Hib). The vaccine is made by chemically conjugating Hib capsular polysaccharide to the outer membrane protein complex of Neisseria meningitidis. The protein-conjugated vaccine has proven to be extremely effective in preventing invasive Hib disease in infants and young children. This paper presents the nuclear magnetic resonance (NMR) methodology for the quantitative characterization of derivatized polysaccharide and its validation closely following ICH guidelines. The assay has been shown to be precise and accurate (relative standard deviation [RSD] ⩽1%), specific (no observable matrix interference), rugged (RSD <3% for day-to-day and operator-to-operator variations), and robust (to changes in temperature and polysaccharide concentration). Therefore, the quantitative NMR assay can be used as a reliable product release or process monitoring method and can replace several labor-intensive chromatographic and colorimetric methods. [Copyright &y& Elsevier]

Abstract: Purified capsular polysaccharide preparations from Streptococcus pneumoniae that are used for vaccine production typically contain residual levels of C-polysaccharide (C-Ps). Residual C-Ps is typically found in one of two forms, either chemically linked to the capsular polysaccharide (bound) or present by itself (free). Two analytical methods have been developed and applied to determine the relative percentages of the two C-Ps forms present in various capsular polysaccharide preparations. Both methods differentiate the two forms of C-Ps according to the difference of their hydrodynamic sizes. One method is based on labeling C-Ps with a fluorescent tag and separating the two forms of C-Ps by high-performance size exclusion chromatography with on-line refractive index and fluorescence detection, and the other method is based on measuring self-diffusion rates of the two forms of C-Ps by nuclear magnetic resonance (NMR) and quantifying each form with deconvolution. Both methods were evaluated for relative accuracy, precision, and ease of application, and they were found to provide comparable results for a large number of pneumococcal polysaccharide preparations. These analyses, combined with other quantitative NMR measurement of total C-Ps in the polysaccharide powder, provide a more refined means of evaluating the amount of each form of C-Ps in polysaccharide preparations targeted for vaccine production. [Copyright &y& Elsevier]

Indoleamine-2,3-dioxygenase 1 (IDO1) and tryptophan-2,3-dioxygenase 2 (TDO2) degrade tryptophan (Trp) to kynurenine (Kyn), and these enzymes have promise as therapeutic targets. A comprehensive characterization of potential safety liabilities of IDO1 and TDO2 inhibitors using knockout (KO) mice has not been assessed, nor has the dual Ido1/Tdo2 KO been reported. Here we characterized male and female mice with KOs for Ido1 , Tdo2 , and Ido1/Tdo2 and compared findings to the wild type (WT) mouse strain, evaluated for 14 days, using metabolomics, transcriptional profiling, behavioral analysis, spleen immunophenotyping, comprehensive histopathological analysis, and serum clinical chemistry. Multiple metabolomic changes were seen in KO mice. For catabolism of Trp to Kyn and anthranilic acid, both substrates were decreased in liver of Tdo2 and dual KO mice. Metabolism of Trp to serotonin and its metabolites resulted in an increase in 5-Hydroxyindole-3-acetic acid in the Tdo2 and dual KO mice. Ido1 and dual KO mice displayed a Kyn reduction in plasma but not in liver. Nicotinamide synthesis and conversion of glucose to lactic acid were not impacted. A slight decrease in serum alkaline phosphatase was seen in all KOs, and small changes in liver gene expression of genes unrelated to tryptophan metabolism were observed. Regarding other parameters, no genotype-specific changes were observed. In summary, this work shows metabolomic pathway changes for metabolites downstream of tryptophan in these KO mice, and suggests that inhibition of the IDO1 and TDO2 enzymes would be well tolerated whether inhibited individually or in combination since no safety liabilities were uncovered. • IDO1 & TDO2 degrade tryptophan to kynurenine, & these enzymes have promise as therapeutic targets. • To understand target liabilities, single & dual Ido1 & Tdo2 KO mice were thoroughly evaluated. • Tryptophan metabolite changes were seen, & other subtle findings had no toxicologic consequence. • This implies inhibition of these enzymes would be well tolerated. [ABSTRACT FROM AUTHOR]