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Peptide mapping with mass spectrometry (MS) is an important tool for protein characterization in the biopharmaceutical industry. Historically, peptide mapping monitors post-translational modifications (PTMs) of protein products and process intermediates during development. Multi-attribute monitoring (MAM) methods have been used previously in commercial release and stability testing panels to ensure control of selected critical quality attributes (CQAs). Our goal is to use MAM methods as part of an overall analytical testing strategy specifically focused on CQAs, while removing or replacing historical separation methods that do not effectively distinguish CQAs from non-CQAs due to co-elution. For example, in this study, we developed a strategy to replace a profile-based ion-exchange chromatography (IEC) method using a MAM method in combination with traditional purity methods to ensure control of charge variant CQAs for a commercial antibody (mAb) drug product (DP). To support this change in commercial testing strategy, the charge variant CQAs were identified and characterized during development by high-resolution LC-MS and LC-MS/MS. The charge variant CQAs included PTMs, high molecular weight species, and low molecular weight species. Thus, removal of the IEC method from the DP specification was achieved using a validated LC-MS MAM method on a QDa system to directly measure the charge variant PTM CQAs in combination with size exclusion chromatography (SE-HPLC) and capillary electrophoresis (CE-SDS) to measure the non-PTM charge variant CQAs. Bridging data between the MAM, IEC, and SE-HPLC methods were included in the commercial marketing application to justify removing IEC from the DP specification. We have also used this MAM method as a test for identity to reduce the number of QC assays. This strategy has received approvals from several health authorities.

Objective: To evaluate the long-term performance of the patch materials we have used to augment the pulmonary arterial tree across a wide spectrum of diagnoses and anatomical locations. Methods: Retrospective, single-center review of 217 consecutive pediatric patients at a tertiary referral center from 1993 to 2020 who underwent patch arterioplasty of the pulmonary arterial tree from the pulmonary bifurcation to the distal pulmonary arterial branches. Reintervention data were collected and analyzed. Lesion-specific anatomy and other variables were analyzed as risk factors for reintervention. Results: There were 280 total operations performed (217 initial operations and 63 reoperations) and 313 patches used. The patches used were autologous pericardium (166, 53.0%), pulmonary homograft (126, 40.3%), and a heterogeneous group of other materials (21, 6.7%). Overall patient survival was 86.2%, freedom from reoperation was 81.0% and freedom from reintervention (FFR) was 70.6%, with a median follow-up of 13.8 years (interquartile range, 6.3-17.9 years). For all patches, 10-, 20-, and 27-year FFR was 76.6%, 70.6%, and 70.6%, respectively. FFR was similar among all 3 patch type groups (P = .29). Multivariable Cox regression analysis showed that diagnoses of pulmonary atresia with ventricular septal defect and major aortopulmonary collateral arteries and hypoplastic left heart syndrome, patches placed at initial cardiac operation, and increasing number of cardiac operations were risk factors for reintervention. Conclusions: Autologous pericardium and pulmonary homograft patches performed similarly. Although patch type conferred no difference in need for reintervention, other risk factors did exist. Namely, diagnoses of pulmonary atresia with ventricular septal defect and major aortopulmonary collateral arteries and hypoplastic left heart syndrome, patch placement at a patient's first cardiac operation, and increasing number of cardiac operations were risk factors for reintervention.

Chronic alcohol consumption is associated with fatty liver disease in mammals. The object of this study was to gain an understanding of dysregulated lipid metabolism in alcohol-fed C57BL/6 mice using a targeted lipidomic approach. Liquid chromatography tandem mass spectrometry was used to analyze several lipid classes, including free fatty acids, fatty acyl-CoAs, fatty acid ethyl esters, sphingolipids, ceramides, and endocannabinoids, in plasma and liver samples from control and alcohol-fed mice. The interpretation of lipidomic data was augmented by gene expression analyses for important metabolic enzymes in the lipid pathways studied. Alcohol feeding was associated with i) increased hepatic free fatty acid levels and decreased fatty acyl-CoA levels associated with decreased mitochondrial fatty acid oxidation and decreased fatty acyl-CoA synthesis, respectively; ii) increased hepatic ceramide levels associated with higher levels of the precursor molecules sphingosine and sphinganine; and iii) increased hepatic levels of the endocannabinoid anandamide associated with decreased expression of its catabolic enzyme fatty acid amide hydrolase. The unique combination of lipidomic and gene expression analyses allows for a better mechanistic understanding of dysregulated lipid metabolism in the development of alcoholic fatty liver disease.