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Thymidine Analogues As Potent Tk-2 Inhibitors

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Thymidine Analogues As Potent Tk-2 Inhibitors SYNTHESIS AND BIOLOGICAL EVALUATION OF PYRIMIDINE NUCLEOSIDES AND NUCLEOSIDE PHOSPHONATES Apr. Sara Van Poecke Promoter Prof. Dr. Apr. Serge Van Calenbergh Academic year 2011-2012 SYNTHESIS AND BIOLOGICAL EVALUATION OF PYRIMIDINE NUCLEOSIDES AND NUCLEOSIDE PHOSPHONATES Apr. Sara Van Poecke Thesis submitted to the Faculty of Pharmaceutical Sciences to obtain the degree of Doctor in Pharmaceutical Sciences Promoter Prof. Dr. Apr. Serge Van Calenbergh Academic year 2011-2012 TABLE OF CONTENTS PART I: 3’-MODIFIED THYMIDINE ANALOGUES AS POTENT TK-2 INHIBITORS ........................... 1 Chapter 1: INTRODUCTION ................................................................................................................... 3 1.1 DEOXYRIBONUCLEOTIDES ................................................................................................... 3 1.1.1 The de novo Pathway ........................................................................................................ 4 1.1.2 Salvage Pathway ............................................................................................................... 4 1.2 DEOXYNUCLEOSIDE KINASES .............................................................................................. 4 1.3 THYMIDINE KINASES (TK-2 VS. TK-1)..................................................................................... 5 1.3.1 Localization and Expression .............................................................................................. 5 1.3.2 Substrate Specificities ....................................................................................................... 6 1.3.3 Structure ............................................................................................................................ 8 1.3.4 Mechanism of Action ....................................................................................................... 10 1.4 THE ROLE OF TK-2 ................................................................................................................ 11 1.4.1 mtDNA Synthesis ............................................................................................................. 11 1.4.2 TK-2 Deficiency ................................................................................................................ 12 1.5 TK-2 INHIBITORS ................................................................................................................... 15 1.5.1 Role of TK-2 Inhibitors ..................................................................................................... 15 1.5.2 Structure of TK-2 Inhibitors .............................................................................................. 15 Chapter 2: SYNTHESIS AND BIOLOGICAL EVALUATION OF 3’-TRIAZOLE-SUBSTITUTED THYMIDINE AND BVDU DERIVATIVES .............................................................................................. 21 2.1 OBJECTIVES .......................................................................................................................... 21 2.2 CHEMISTRY ........................................................................................................................... 23 2.2.1 Synthesis of 3’-Triazole-Substituted Thymidine Analogues ............................................ 23 2.2.2 Synthesis of 5’-O-Protected Isothiocyanate Analogue 1.59 ............................................ 25 2.2.3 Synthesis of 3’-Triazole-Substituted BVDU Analogues ................................................... 25 2.3 BIOLOGICAL EVALUATION ................................................................................................... 27 2.4 MOLECULAR MODELING AND STRUCTURE-ACTIVITY RELATIONSHIP ......................... 30 2.5 PHOTOPHYSICAL EVALUATION .......................................................................................... 34 2.6 CONCLUSIONS ...................................................................................................................... 35 2.7 EXPERIMENTAL PART .......................................................................................................... 36 2.7.1 Synthesis ......................................................................................................................... 36 2.7.2 Experimental Assays ....................................................................................................... 52 vii Chapter 3: SYNTHESIS AND BIOLOGICAL EVALUATION OF 3’-GUANIDINE-SUBSTITUTED THYMIDINE AND BVDU DERIVATIVES .............................................................................................. 55 3.1 OBJECTIVES .......................................................................................................................... 55 3.2 CHEMISTRY ........................................................................................................................... 56 3.2.1 Synthesis of 3’-Benzylamino Analogues of Thymidine .................................................... 56 3.2.2 Synthesis of 3’-Guanidine and 3’-Aminotetrazole Analogues of Thymidine .................... 56 3.2.3 Synthesis of 3’-Aminotetrazole Analogues of BVDU ....................................................... 60 3.3 BIOLOGICAL EVALUATION ................................................................................................... 61 3.4 MOLECULAR MODELING ...................................................................................................... 63 3.5 CONCLUSIONS ...................................................................................................................... 64 3.6 EXPERIMENTAL PART .......................................................................................................... 65 3.6.1 Synthesis ......................................................................................................................... 65 Part II: FURTHER EXPLORATION OF THE S.A.R. OF SUBSTRATE-BASED M. tuberculosis TMPK INHIBITORS ............................................................................................................................... 85 Chapter 1: INTRODUCTION ................................................................................................................. 87 1.1 TUBERCULOSIS .................................................................................................................... 89 1.1.1 Causative Organisms ...................................................................................................... 89 1.1.2 Transmission .................................................................................................................... 90 1.1.3 Pathophysiology............................................................................................................... 90 1.1.4 Symptoms ........................................................................................................................ 91 1.1.5 Diagnosis ......................................................................................................................... 92 1.1.6 Prevention and Control .................................................................................................... 93 1.1.7 Treatment ......................................................................................................................... 94 1.1.8 Resistance ....................................................................................................................... 97 1.2 THYMIDINE MONOPHOSPHATE KINASE (TMPK) ............................................................. 100 1.2.1 Structure of TMPKmt ..................................................................................................... 100 1.2.2 Known TMPKmt Inhibitors ............................................................................................. 102 Chapter 2: SYNTHESIS AND INHIBITORY ACTIVITY OF THYMIDINE ANALOGUES TARGETING TMPKmt ............................................................................................................................................... 107 2.1 OBJECTIVES ........................................................................................................................ 107 2.2 CHEMISTRY ......................................................................................................................... 110 2.2.1 Synthesis of 3’-Modified 4-Thio-β-Thymidine Analogues .............................................. 110 2.2.2 Synthesis of 5’-Modified β-Thymidine Analogues.......................................................... 111 2.2.3 Synthesis of 5’-Modified (4-Thio-)α-Thymidine Analogues ............................................ 113 2.2.4 Synthesis of 3’-Modified α-Thymidine Analogues.......................................................... 115 2.3 BIOLOGICAL EVALUATION ................................................................................................. 116 2.4 MOLECULAR DOCKING ...................................................................................................... 118 2.5 CONCLUSIONS .................................................................................................................... 119 viii 2.6 EXPERIMENTAL PART ........................................................................................................ 120 2.6.1 Synthesis ....................................................................................................................... 120 2.6.2 Biological Assays ..........................................................................................................
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