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dc.contributor.advisorSchöneich, Christian
dc.contributor.authorBommana, Rupesh
dc.date.accessioned2019-05-10T16:12:24Z
dc.date.available2019-05-10T16:12:24Z
dc.date.issued2017-12-31
dc.date.submitted2017
dc.identifier.otherhttp://dissertations.umi.com/ku:15617
dc.identifier.urihttp://hdl.handle.net/1808/27834
dc.description.abstractProtein-based pharmaceuticals are a fast growing class of therapeutics, which are widely used in the treatment of various diseases such as cancers and autoimmune diseases. Proteins are susceptible to multiple degradation pathways including oxidation, deamidation and photodegradation. Over the past decade, the pharmaceutical industry has become increasingly cognizant of the sensitivity of proteins towards light. The exposure of proteins to light during their development and patient administration is inevitable, and may result in protein degradation. In the recent past there have been multiple reports indicating formation of aggregates, discoloration, oxidation, covalent crosslinking and fragmentation on exposure of protein pharmaceuticals to light. These degradation pathways could enhance immunogenicity or cause inactivation. Therefore, to increase the stability of these proteins, an understanding of their degradation is necessary. The research covered in this dissertation focused on two major degradation mechanisms for an IgG monoclonal antibody (mAb): chemical and physical degradation by photo irradiation. First, this dissertation explored the effects of certain pharmaceutical excipients on the extent and site-specificity of epimerization in UV irradiated mAb formulations. Amino acid analysis of UV irradiated mAb revealed formation of D-Ala, D-Glu, and D-Val in mAb formulations. Secondly, the underlying mechanism behind the increased aggregation propensity of a photo irradiated IgG1 mAb was investigated using hydrogen deuterium exchange mass spectrometry and biophysical characterization. Specific correlations were established between changes in dynamics of distinct segments in the CH2 domain of the IgG1 mAb and decreased thermal stability as well as increase in aggregation propensity of the IgG1 mAb on light exposure. This dissertation also describes development of a rapid screening methodology to identify protein oxidation by monitoring Tyr and Phe oxidation using fluorogenic derivatization. The Fluorogenic derivatization technique was adapted to a 96-well plate, in which several protein formulations can be screened in a short time. In addition, this dissertation also outlines the capability of an extreme ultra pressure liquid chromatography system in investigating complex chemical degradation problems.
dc.format.extent234 pages
dc.language.isoen
dc.publisherUniversity of Kansas
dc.rightsCopyright held by the author.
dc.subjectPharmaceutical sciences
dc.subjectChemistry
dc.subjectBiochemistry
dc.subjectamino acid analysis
dc.subjectfluorogenic derivatization
dc.subjecthydrogen deuterium exchange mass spectrometry
dc.subjectlight exposure
dc.subjectmonoclonal antibody
dc.subjectprotein degradation
dc.titleDegradation of therapeutic proteins: Screening methods and identification of epimerized amino acids and local conformational changes in light exposed proteins
dc.typeDissertation
dc.contributor.cmtememberVolkin, David B
dc.contributor.cmtememberWang, Zhou (Michael)
dc.contributor.cmtememberTolbert, Thomas J
dc.contributor.cmtememberDeeds, Eric
dc.thesis.degreeDisciplinePharmaceutical Chemistry
dc.thesis.degreeLevelPh.D.
dc.identifier.orcid
dc.rights.accessrightsembargoedAccess


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