Analytical method developments of antibody drug conjugates and disease biomarkers in microdialysis samples

Abstract

This dissertation focuses on developing analytical methods to study biomarkers in different pharmaceutical samples. Three different analytical methods were developed for microdialysis samples and antibody drug conjugates as anti-tumor drug. The first part of this dissertation is to develop a capillary electrophoresis with laser induced fluorescence (CE-LIF) method to monitor the change of amino acids in rat brain microdialysate as biomarkers of oxidative stress in epileptic seizures. Ornithine and citrulline was successfully separated and quantified. 3-Mercaptopropionic acid (3-MPA) was administrated to rat brain hippocampus region as a convulsant to induce epileptic seizures to free-moving rats. An increase of citrulline and ornithine level was observed after the seizure, and this confirmed nitric oxide were produced in epileptic seizures. In the second project, a high-performance liquid chromatography with mass spectrometry (HPLC-MS) method is developed to simultaneously monitor the change of 13 eicosanoids as biomarkers in rat colon microdialysate to study the enzymatic pathways of inflammatory bowel disease. Moreover, cyclodextrin were added into microdialysis perfusate to improve the microdialysis recovery and solubility of the hydrophobic eicosanoids. The last part is to utilize enzyme deconjugation and HPLC-UV-Vis to study the stability and compatibility of antibody drug conjugates (ADCs). Bromelain was utilized as an enzyme to fully cleave off the small molecule cytotoxic drug from the ADCs. This allows a direct analysis of the small molecule cytotoxic drug on the ADCs. The stability of antibody drug conjugates was studied with LC-UV-Vis after mixing with other conjugation reagents and formulation excipients.