Document Type


Date of Award



College of Pharmacy and Health Sciences (COPHS)

Degree Name

Ph.D. in Pharmaceutical Science

Committee Chairperson

Dong Liang

Committee Member 1

Huan Xie

Committee Member 2

Song Gao

Committee Member 3

Selvam Chelliah

Committee Member 4

Mario Hollomon


CYP, Drug Metabolism, LC-MS/MS, Pharmacokinetics, Preclinical Development, UGT


Current treatments for cutaneous leishmaniasis suffer from toxic side effects, high cost, parenteral administration, and drug resistance. Thus, there is a critical need to develop oral drugs for the treatment of leishmaniasis. OJT007 is a novel class of drug with potent antiproliferative effects against Leishmania Major. The purpose of this project is to conduct preclinical drug development studies for OJT007 including bioanalytical assay development, pre-formulation studies, in vitro hepatic drug metabolism and in vivo pharmacokinetics. A sensitive, specific, and reproducible ultra-high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated. The separation was achieved on a UPLC BEH C18 column (2.1 × 50 mm, 1.7 μm) using a mobile phase consisting of 0.1% formic acid in acetonitrile and 0.1% formic acid in water as gradient elution at a flow rate of 0.4 mL/min. The mass analysis was performed with a 4000 QTRAP® mass spectrometer using multiple-ion reaction monitoring (MRM) in the positive mode, with the transition of m/z 325 → m/z 205 for OJT007 and m/z 350 → m/z 101 for voriconazole (as internal standard). Rat plasma and urine were extracted for OJT007 by protein precipitation in acetonitrile for quantification. Plasma protein binding of OJT007 was determined using the ultrafiltration method. In vitro phase I and II hepatic metabolism of OJT007 was evaluated in rat liver microsomes using standard reaction protocols. OJT007 metabolites were identified by LC-MS/MS using Q1MI, product ion, precursor ion and neutral loss scan and by β-glucuronidase hydrolysis. The OJT007 glucuronidation rates were determined by quantifying OJT007 glucuronide using UPLC, and the kinetic parameters of OJT007 were determined by measuring the initial glucuronidation rates. . Oral bioavailability of OJT007 was evaluated using a crossover study design. Serial plasma samples were collected at predetermined time points. Total urine samples were also collected from each rat for 24 hours after each dose. The pharmacokinetic parameters were calculated using Phoenix WinNonlin® 8.3 software. The intra- and inter-day precision and accuracy were within the acceptable limit of ≤20% for LLOQ and ≤15% for high, medium and low QC. The extraction recovery in rat plasma and urine samples were 95.1% and 83%, respectively. OJT007 exhibited a matrix factor in plasma and urine of 7.96% and 12.4%, respectively. The fraction of OJT007 bound to plasma protein had a mean value of 99.1%, suggesting the drug is highly bound to plasma proteins. OJT007 is glucuronidated rapidly in rat liver microsomes to form a mono-glucuronide, which was confirmed by LC-MS/MS and β-glucuronidase hydrolysis. The kinetic parameters of glucuronidation Vmax and Km were 1.125 nmol/min/mg and 10.73 μM., respectively. After intravenous administration, OJT007 displayed a bi-exponential disposition with a rapid distribution followed by a slower elimination. The mean AUC, volume of distribution, and clearance were 2.06, 6.90 L/Kg and 2.30 L/hr/Kg, respectively. Following oral administration, OJT007 was rapidly absorbed with a tmax of 1.4 hours. After oral dosing the mean AUC, volume of distribution, and clearance were 0.45, 78.6 L/Kg and 23.19 L/hr/Kg, respectively. Mean oral bioavailability of OJT007 in the co-solvent formulation was 10.9%. The mean percentage of OJT007 dose excreted unchanged in urine after 24 hours following intravenous and oral administration was less than 1%, suggesting OJT007 was extensively metabolized in vivo. A sensitive, specific and reproducible LC-MS/MS method was developed and validated to quantify OJT007 in rat plasma and urine. The method was successfully used for pharmacokinetic studies. This is the first time that oral bioavailability of OJT007 after oral administration in rat is determined. These results may prove valuable for further preclinical and clinical evaluation of OJT007


To view the content in your browser, please download Adobe Reader or, alternately,
you may Download the file to your hard drive.

NOTE: The latest versions of Adobe Reader do not support viewing PDF files within Firefox on Mac OS and if you are using a modern (Intel) Mac, there is no official plugin for viewing PDF files within the browser window.