City University of New York (CUNY) CUNY Academic Works All Dissertations, Theses, and Capstone Projects Dissertations, Theses, and Capstone Projects 9-2017 Synthesis and Evaluation of Tetrahydroprotoberberines as Dopamine Receptor Ligands Satishkumar V. Gadhiya The Graduate Center, City University of New York How does access to this work benefit ou?y Let us know! More information about this work at: https://academicworks.cuny.edu/gc_etds/2159 Discover additional works at: https://academicworks.cuny.edu This work is made publicly available by the City University of New York (CUNY). Contact: [email protected] SYNTHESIS AND EVALUATION OF TETRAHYDROPROTOBERBERINES AS DOPAMINE RECEPTOR LIGANDS BY SATISHKUMAR GADHIYA A dissertation submitted to the Graduate Faculty in Chemistry in partial fulfillment of the requirements for the degree of Doctor of Philosophy, The City University of New York. 2017 © 2017 All Rights Reserved SATISHKUMAR V GADHIYA ii Synthesis and Evaluation of Tetrahydroprotoberberines as Dopamine Receptor Ligands by Satishkumar Gadhiya This manuscript has been read and accepted for the Graduate Faculty in Chemistry in satisfaction of the dissertation requirement for the degree of Doctor of Philosophy Date Dr. Wayne Harding Chair of Examining Committee Date Dr. Brian Gibney Executive Officer Dr. Shengping Zheng Dr. Barbara Zajc Dr. Wayne Harding Supervisory Committee THE CITY UNIVERSITY OF NEW YORK iii ABSTRACT Synthesis and Evaluation of Tetrahydroprotoberberines as Dopamine Receptor Ligands By Satishkumar Gadhiya Advisor: Dr. Wayne Harding Dopamine (DA) receptors belong to the G-protein coupled receptors (GPCRs) family, divided in to two groups based on their high homology transmembrane domains; D1-like DA receptors (D1, D5) and D2-like DA receptors (D2-D4). DA receptor specific ligands have been exploited as a means for studying the prognosis and curing several CNS disorders. Though several efforts have been devoted to discover selective and potent DA receptor ligands, complete selectivity within the DA receptor subtypes remains a challenge. Tetrahydroprotoberberines (THPBs) are a group of naturally occurring tetracyclic alkaloids that belong to the tetrahydroisoquinoline family. A wide range of biological activities are associated with the THPB scaffold. Noted examples of THPB alkaloids, (-)-stepholidine 2.1 (SPD), (-)-tetrahydropalmatine 2.2 (THP), (-)-isocorypalmine 2.3 (ICP), and (-)-canadine 2.4, are known to possess high affinity for DA receptors and have been studied widely for their utility in various CNS disorders. (-)-Stepholidine is endowed with DA D1 agonist and D3 antagonist dual pharmacological profile, which makes it a potential candidate for the treatment of schizophrenia and psychostimulant drug abuse. Limitations on the unavailability of efficient synthetic methods, scarcity of structure-activity relationship (SAR) data at DA receptors and metabolic instability iv need to be addressed to make (-)-SPD 2.1 an ideal drug candidate for schizophrenia and psychostimulant drug addiction. In our work we have addressed prominent issues and missing gaps towards the synthesis and SAR studies of THPBs at dopamine D1, D2 and D3 receptors. A novel route to synthesize (±)-SPD (2.1) was developed over eight synthetic steps with 30% overall yield via the intermediacy of easily obtainable diester 3.7. In addition to the superior yield, this approach is advantageous in terms of conciseness and ease of synthetic manipulations. This route was deployed to produce C10 THPB analogues for SAR studies at DA receptors. Employing vital intermediate lactone 2.102, the enantioselective synthesis of (-)-SPD (2.1) was accomplished in 23% overall yield from commercially available starting materials in a fourteen step sequence. The crystal structure of (-)-SPD (2.1) was determined by single crystal X-ray diffraction. Synthesis of lactone 2.102 was straightforward, scalable and high yielding. This synthetic pathway was applied to generate C3 chiral THPB analogues of THPB for bioactivity studies. In order to understand the structural tolerance of the THPB core required for selective D1/D3 receptor binding, a thorough SAR study was designed. Consequently, a diverse library of C3 and C10 THPB analogues was synthesized and evaluated for affinity at DA D1, D2 and D3 receptors. Results from the SAR studies demonstrate that the C10 THPB analogues exhibit a general preference for the D1 receptor. Binding affinity evaluation at D3 receptor indicates that larger alkoxy groups are not tolerated at C10 position. In general, small alkoxy substituents are well tolerated at C10 position of THPB scaffold for D1 and D3 receptor affinity. Novel enantiopure C3 analogues of THPB were designed and synthesized to assess the optimum size of the alkoxy group at this position required for DA D1 and D3 receptor affinity. The results for binding affinity data suggest that C3 alkoxy substitution, decreases D1 and D3 receptor affinity while improving v selectivity at D3 versus D2 receptor when compared with parent compound (-)-SPD (2.1). Reduced affinity at D2 receptor for these analogues signifies the importance of the C3 alkoxy group to suppress affinity at the D2 receptor. These compounds will further expand our understanding of the tolerance of the THPB core required for DA D1 and D3 receptor activity. In order to rationalize the affinity of ligands, molecular docking studies were conducted at the D3 receptor. The ligand binding energies from the D3 receptor docking studies show some similarities and minor discrepancies compared to the experimentally derived affinities. These docking studies have revealed key receptor–ligand interactions, including the critical protonated tertiary N— Asp110 salt bridge motif, H-bonds to Ser192 and Cys181 and hydrophobic interactions to Phe106 and Phe345. Novel series of conformationally flexible analogues of (-)-SPD (2.1) were synthesized and evaluated in order to identify potent and selective DA D1 and D3 receptor ligands. Two groups of analogues were synthesized and assayed for binding affinity at DA receptors, one group having truncated THPB scaffold while the other featured a tetrahydroisoquinoline (THIQ)/arylamide hybridized pharmacophore core. Results of this study indicate that de-rigidification of the THPB scaffold nullifies the DA receptor activity and the intact THPB scaffold of (-)-SPD is necessary for the DA receptor activity. On the other hand, THIQ/arylamide hybrid analogues have demonstrated high potency and selectivity for D3 receptor. Representative compounds were studied in order to test the metabolic stability of synthesized THPB analogues. The results of this metabolic study indicate that having phenolic hydroxyl group at C2 and/or C10 on THPB scaffold engenders susceptibility to hepatic metabolism. vi Dedicated to Rev. Dadaji and Rev. Taiji, the source of my inspiration and blessings vii Acknowledgement This dissertation could not have been completed without the great support that I have received from so many people over the years. First and foremost, I offer my sincerest gratitude to my advisor, Professor Wayne Harding, for his patience, motivation, enthusiasm and immense knowledge. He patiently provided the vision, encouragement and advice necessary for me to proceed through the doctoral program and complete my dissertation. He has been generous with his knowledge, approachable to clarify doubts and willing to assist me overcome whatever hurdles I came across during my doctoral research. He showed genuine interest in my career development. He understood my career goals and provided me with ample opportunities to develop the skills required to achieve my goals. He always guided me to work towards substantial long-term objectives, structure my research, present my ideas effectively, and promote both myself and my research results. I could not have imagined having a better and more concerned mentor for my PhD study. I am also very grateful that Professor Barbara Zajc and Professor Shengping Zheng have been my committee members since 2012. They have provided valuable suggestions on my proposal and research, which guided me to interpret the research project from different perspectives and to improve my scientific skills. I am very much thankful to all the members of Harding lab who have been helpful and cheering at many different levels of graduate study. A special thanks to my senior colleagues Shashi, Sudharshan and Nirav for their continuous support and encouragement. I have learnt a lot from you guys, specially many tedious organic reactions and laboratory techniques. Anupam, Piarpaolo and Raj, thank you all for making the last phase of my PhD a joyous and cheerful period. viii I am thankful to all the undergrads of the Harding lab for their support and all the help they provided in my different research projects. I would like to thank all the members of Dr. Mootoo’s lab, Dr. Zheng’s Lab, Dr. Kawamura’s lab and Dr. Drain’s lab for their cooperation, direct and indirect favors during my graduate study. I am sincerely grateful to the Chemistry Department at Hunter College for providing the environment and equipment I have needed to produce and complete my thesis work. I would like to thank Prof. Thomas Kurtzman and his lab members at Lehman College for doing all the docking studies for my research project. I am very thankful to Dr. Bryan Roth and his colleagues at the University of North Carolina School of Medicine for screening compounds of my research projects. The research work I have done