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Investigation on Nanoparticle Based Combination Therapy for Targeted Cancer Treatment

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Investigation on Nanoparticle Based Combination Therapy for Targeted Cancer Treatment Scholars' Mine Doctoral Dissertations Student Theses and Dissertations Fall 2020 Investigation on nanoparticle based combination therapy for targeted cancer treatment Muhammad Raisul Abedin Follow this and additional works at: https://scholarsmine.mst.edu/doctoral_dissertations Part of the Biomedical Engineering and Bioengineering Commons, Chemical Engineering Commons, and the Nanotechnology Commons Department: Chemical and Biochemical Engineering Recommended Citation Abedin, Muhammad Raisul, "Investigation on nanoparticle based combination therapy for targeted cancer treatment" (2020). Doctoral Dissertations. 2947. https://scholarsmine.mst.edu/doctoral_dissertations/2947 This thesis is brought to you by Scholars' Mine, a service of the Missouri S&T Library and Learning Resources. This work is protected by U. S. Copyright Law. Unauthorized use including reproduction for redistribution requires the permission of the copyright holder. For more information, please contact [email protected]. INVESTIGATION ON NANOPARTICLE BASED COMBINATION THERAPY FOR TARGETED CANCER TREATMENT by MUHAMMAD RAISUL ABEDIN A DISSERTATION Presented to the Faculty of the Graduate School of the MISSOURI UNIVERSITY OF SCIENCE AND TECHNOLOGY In Partial Fulfillment of the Requirements for the Degree DOCTOR OF PHILOSOPHY in CHEMICAL ENGINEERING 2020 Approved by: Sutapa Barua, Advisor Daniel Forciniti Yongjian Liu Jee Ching Wang Hu Yang © 2020 Muhammad Raisul Abedin All Rights Reserved iii PUBLICATION DISSERTATION OPTION This dissertation consists of the following five articles, formatted in the style used by the Missouri University of Science and Technology: Paper I, found on pages 25-67, is currently under review in Scientific Reports. Paper II, found on pages 68-123, is currently under review in Nano LIFE journal. Paper III, found on pages 124-162, is ready for submission to a peer reviewed journal. Paper IV, found on pages 163-194, was published in the Journal of Nanobiotechnology. Paper V, found on pages 195-230, is currently in press (Scientific Reports). iv ABSTRACT The current treatment methods in cancer are associated with toxicity in healthy tissues, partial therapeutic response, drug resistance and finally recurrence of the disease. The cancer drugs are challenged by non-specific binding, undesired toxicity in healthy cells, low therapeutic index and finally poor therapeutic outcome. In this work, a targeted nanoscale therapeutic system Antibody Drug Nanoparticle (ADN) was engineered to selectively inhibit the breast cancer cell growth with reduced toxicity in healthy cells. The ADNs were designed by synthesizing rod shaped nanoparticles using pure chemotherapeutic drug and covalently conjugating a therapeutic monoclonal antibody (mAb) on the surface of the drug nanorods. The rod shaped nanosized formulation of ADNs significantly enhanced the aqueous phase stability and therapeutic payload of the system while the conjugated mAb was utilized for specific targeting of breast cancer cells. The designed ADN was effective for active targeting and synergistic inhibition of breast cancer cells. The mechanisms of actions of ADN was investigated at the cellular, molecular and genetic levels in cancer cells. The engineered ADN synergistically inhibited the growth of >80% of the human epidermal growth factor receptor 2 (HER2) - positive breast cancer cells in vitro. The cell cycle and protein expression analysis showed that ADN arrested the cellular growth for a prolonged time and induced a programmed cell death mechanism in HER2-positive breast cancer cells in vitro. Finally, the gene regulatory analysis showed the genetic mechanisms of programmed cell death regulation induced by ADN in breast cancer cell lines. v ACKNOWLEDGMENTS I would like to express my gratitude to my advisor Dr. Sutapa Barua for her kind guidance throughout this uneven journey. Dr. Barua has been mentoring me from the very beginning till this date. All these years, Dr. Barua has been kind enough to keep her belief on me. The learnings from Dr. Barua will be the most valuable asset that I will take from this experience. I am also thankful to my other Ph.D. committee members, Dr. Daniel Forciniti, Dr. Yongjian Liu, Dr. Jee-Ching Wang and Dr. Hu Yang for agreeing to serve on my committee, investing their valuable time and providing valuable feedback. I would like to thank extremely talented fellow researchers in my lab, Dr. Sidharth Razdan, Kaitlyne Powers, Rachel Aiardo, Jawahar Khetan, Dr. Md Shahinuzzaman, Dr. Mohammad Aminul Islam. I appreciate their support during my hard times. My gratitude extends to all the faculty members of the Department of Chemical and Biochemical Engineering at Missouri S&T. Finally, I would like to thank my family: my parents, grandparents and my sister for their endless encouragement from nine thousand miles away in Bangladesh. My banker father and my loving homemaker mother sacrificed their whole lives for our better future. Today, this Ph.D. is their achievement more than mine. Vi TABLE OF CONTENTS Page PUBLICATION DISSERTATION OPTION...................................................................iii ABSTRACT.......................................................................................................................iv ACKNOWLEDGMENTS..................................................................................................v LIST OF ILLUSTRATIONS...........................................................................................xiv LIST OF TABLES..........................................................................................................xvii NOMENCLATURE......................................................................................................xviii SECTION 1. INTRODUCTION.................................................................................................... 1 1.1. NANOTECHNOLOGY IN CANCER............................................................... 1 1.2. CURRENT TREATMENT METHODS IN CANCER...................................... 3 1.2.1. Surgery......................................................................................................3 1.2.2. Radiation Therapy.....................................................................................4 1.2.3. Chemotherapy...........................................................................................5 1.2.4. Cancer Immunotherapy.............................................................................6 1.3. NANOPARTICLE BASED THERAPEUTICS IN CANCER TREATMENT . 7 1.4. COMBINATION THERAPY TO OVERCOME THE THERAPEUTIC RESISTANCE IN CANCER TREATMENT.................................................. 14 1.5. ANTIBODY DRUG CONJUGATES.............................................................. 15 1.5.1. Advantages and Challenges Associated with ADCs...............................17 1.5.2. Strategies for Developing Efficacious ADCs..........................................19 vii PAPER I. ANTIBODY-DRUG NANOPARTICLES INDUCES SYNERGISTIC TREATMENT EFFICACIES IN BREAST CANCER CELLS...............................25 ABSTRACT..............................................................................................................25 1. INTRODUCTION....................................................................................................26 2. MATERIALS AND METHODS.............................................................................29 2.1. SYNTHESIS OF PTXNRs...............................................................................29 2.2. CHARACTERIZATION OF PTXNR.............................................................. 30 2.3. ACTIVATION OF 2' OH NUCLEOPHILIC SITE OF PTXNR..................... 30 2.4. !H-NMR ANALYSIS OF ACTIVATED PTXNR........................................... 31 2.5. CONJUGATION OF TTZ WITH ACTIVATED PTXNR THROUGH THE LYSINE RESIDUE INTERACTION.............................................................. 31 2.6. FLUORESCENCE DATA ANALYSIS TO CONFIRM THE CONJUGATION OF TTZ WITH PTXNR...................................................... 32 2.7. OPTIMIZATION OF TTZ CONJUGATION USING RESPONSE SURFACE ANALYSIS...................................................................................32 2.8. IN VITRO THERAPEUTIC EFFICACY......................................................... 33 2.9. QUANTITATIVE ANALYSIS OF SYNERGISTIC EFFECTS OF PTXNR- TTZ...................................................................................................................34 2.10. CELL CYCLE ANALYSIS............................................................................ 34 2.11. WESTERN BLOT ANALYSIS..................................................................... 35 2.12. STATISTICAL ANALYSIS..........................................................................37 3. RESULTS ................................................................................................................ 37 3.1. SYNTHESIS OF PTXNRs...............................................................................37 3.2. CONFIRMATION OF LINKER CONJUGATION WITH PTXNRs USING 1H NMR ANALYSIS.......................................................................................38 viii 3.3. CONFIRMATION OF TTZ CONJUGATION WITH PTXNR-CDI..............40 3.4. OPTIMIZATION OF TTZ CONJUGATION EFFICIENCY..........................40 3.5. IN VITRO ANTICANCER EFFICACY........................................................... 42 3.6. THE EFFECTS OF PTXNR-TTZ IS SYNERGISTIC IN HER2 POSITIVE BREAST CANCER CELLS............................................................................45
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