2019

Human monoclonal antibody stability in rat cell cultures using surrogate peptide LC-MS/MS

Published:  04 June 2019

Evaluating stability of human monoclonal antibody in rat cell cultures using a surrogate peptide LC-MS/MS approach

Nadya Galeva, Reed Murbach, Krystal Gilligan, Kevin Westland, Seema Muranjan
Sekisui XenoTech, LLC, Kansas City, KS, USA

In vitro evaluations of monoclonal antibody (mAb) in drug development can be both useful and challenging. Cytotoxicity and stability of protein therapeutic candidates at relevant concentrations in cell culture are of interest in pharmaceutical research.  Bioanalytical strategies for mAb LC-MS based analysis involve using either intact or digested mAb. Our approach for determining the stability of a human mAb in cultures of primary rat hepatocytes was to use enzymatic protein digestion followed by LC-MS/MS quantitation of five surrogate peptides...
 
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Assessment of in vitro inhibition of CYPs, UGTs and transporters by oligonucleotides

Published:  28 May 2019

An assessment of the in vitro inhibition of cytochrome P450 enzymes (CYP), UDP-glucuronsyltransferases (UGT) and transporters by phosphodiester- or phosphorothioate-linked oligonucleotides

Dr. Brian W. Ogilvie
Sekisui XenoTech, LLC, Kansas City, KS, USA
 
What is already known about this subject?
  • Antisense oligonucleotides (ASOs) are increasingly being developed for many indications and are often modified with phosphorothioate linkages in lieu of phosphodiester linkages.
  • Few in vitro DDI studies with ASOs have been performed, including the investigational imetelstat, volanesorsen, and ISIS 681257.
  • No clinical DDIs have been reported.
 
What this study adds:
  • This study evaluated the inhibitory effects of two non‑therapeutic oligonucleotides with either phosphodiester (PD-GP and PD-Ac) or phosphorothioate (PT-GP and PT-Ac) linkages on the major drug-metabolizing CYPs and UGTs in both human liver microsomes (HLM) and cryopreserved human hepatocytes (CHH), and toward select transporters in expression systems.
 
Abstract:
Regulatory guidance documents do not have specific recommendations for ASO DDI evaluation, but these molecules do fall under the scope of overall DDI testing.
 
Aims:
  1. Evaluate different ASOs as inhibitors of CYPs, UGTs and transporters in vitro.
  2. Determine if in vitro inhibition of CYPs and UGTs in HLM by phosphorothioate ASOs is an artefact not observed in CHH...
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CYP1A2 Enzyme Localization in Normal and Diseased Pediatric Livers

Published:  13 March 2019

B. W. Ogilvie1, M. Czerwinski1, A. Kats2, M. T. Pritchard3,4, and S. E. Tague5.
1Sekisui XenoTech LLC, Kansas City, KS; 2Department of Pathology and Laboratory Medicine, Children’s Mercy Hospital, Kansas City, MO; 3Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS; 4The Liver Center, University of Kansas Medical Center, Kansas City, KS; and 5Kansas Intellectual and Developmental Disabilities Research Center, University of Kansas Medical Center, Kansas City, KS

CYP1A2 is a Phase 1 drug metabolizing enzyme whose expression begins between birth and 4 weeks of age, and gradually increases to about half of the adult levels by 6 years of age. The enzyme is expressed predominantly in liver where it constitutes 4 - 16% of the total hepatic CYP pool, and is a major determinant of the biotransformation of ~9% of clinically used drugs. Interestingly, CYP1A2 activity is decreased in adults with non-alcoholic fatty liver disease (NAFLD). Considering the increase in the number of medications given to children, as well as the rise in childhood obesity and NAFLD, this study utilized a pediatric liver tissue microarray (TMA) to determine if CYP1A2 abundance and localization was influenced by donor age and disease...

As this was selected to be a 20-min. Xenobiotic Disposition in Disease and Toxicities Topic platform presentation at the Society of Toxicology (SOT) Meeting & ToxExpo, no poster was presented. However, you may request information from the presentation or a webinar on the topic.

Patterns of Hyaluronan Deposition in Normal and Diseased Pediatric Livers

Published:  13 March 2019

Presented at the March 2019 Annual Society of Toxicology (SOT) Meeting in Baltimore, MD.
Maciej Czerwiński1, Alexander Kats2, Jordan Surgnier3, Sarah E. Tague4, and Michele T. Pritchard3,5
1
Sekisui XenoTech, Kansas City, KS., 2Department of Pathology and Laboratory Medicine, Children’s Mercy Hospital, Kansas City, MO., 3Department of Pharmacology, Toxicology and Therapeutics, 4Kansas Intellectual and Developmental Disabilities Research Center, 5The Liver Center, University of Kansas Medical Center, Kansas City, KS

The incidence of obesity and non-alcoholic fatty liver disease (NAFLD) is growing in children. Hyaluronan (HA), an extracellular matrix (ECM) glycosaminoglycan, is increased in liver and blood from adults with advanced liver disease. HA accumulation is also associated with inflammation and fibrosis in extrahepatic tissues. HA levels have not been examined in healthy or diseased livers from pediatric patients. Here, we sought to determine if hepatic HA deposition was associated with donor age and/or disease by using a pediatric liver tissue microarray (TMA)...