Frederick R. Maxfield, PhD, is Professor and Chair of the Department of Biochemistry at Weill Cornell Medical College. He has used digital imaging microscopy to characterize pH changes in endocytic organelles, to measure the kinetics of transport of molecules among organelles, and to identify new endocytic organelles such as the endocytic recycling compartment. Dr. Maxfield has published extensively on trafficking of lipids and cholesterol.James M. Willard, PhD, has been a member of the Phospholipidosis Working Group at the Center for Drug Evaluation and Research (CDER) of the Food and Drug Administration since 2005 and Co-Chair of the group since 2011. Shuyan Lu, MSc, has been an Investigative Toxicologist of Drug Research and Development at Pfizer for over 10 years. She studies the role of lysosomal pathways and physical chemical properties of compounds in drug-induced toxicity.

• PREFACE xiiiLIST OF CONTRIBUTORS xvii1 Lysosomes: An Introduction 1Frederick R. Maxfield1.1 Historical Background, 2References, 42 Lysosome Biogenesis and Autophagy 7Fulvio Reggiori and Judith Klumperman2.1 Introduction, 72.2 Pathways to the Lysosomes, 102.2.1 Biosynthetic Transport Routes to the Lysosome, 102.2.2 Endocytic Pathways to the Lysosome, 102.2.3 Autophagy Pathways to the Lysosome, 122.2.4 The ATG Proteins: The Key Regulators of Autophagy, 142.3 Fusion and Fission between the Endolysosomal and Autophagy Pathways, 162.3.1 Recycling Endosomes and Autophagosome Biogenesis, 162.3.2 Autophagosome Fusion with Late Endosomes and Lysosomes, 172.3.3 Autophagic Lysosomal Reformation, 182.4 Diseases, 192.4.1 Lysosome-Related Disorders (LSDs), 192.4.2 Lysosomes in Neurodegeneration and Its Links to Autophagy, 202.4.3 Autophagy-Related Diseases, 202.5 Concluding Remarks, 22Acknowledgments, 23References, 233 Multivesicular Bodies: Roles in Intracellular and Intercellular Signaling 33Emily R. Eden, Thomas Burgoyne, and Clare E. Futter3.1 Introduction, 333.2 Downregulation of Signaling by Sorting onto ILVs, 353.3 Upregulation of Signaling by Sorting onto ILVs, 383.4 Intercellular Signaling Dependent on Sorting onto ILVs, 393.5 Conclusion, 44References, 454 Lysosomes and Mitophagy 51Dominik Haddad and Patrik Verstreken4.1 Summary, 514.2 Mitochondrial Significance, 514.3 History of Mitophagy, 524.4 Mechanisms of Mitophagy, 534.4.1 Mitophagy in Yeast, 544.4.2 Mitophagy in Mammals, 554.5 Conclusion, 57Acknowledgments, 57References, 585 Lysosome Exocytosis and Membrane Repair 63Rajesh K. Singh and Abigail S. Haka5.1 Introduction, 635.2 Functions of Lysosome Exocytosis, 635.2.1 Specialized Lysosome-Related Organelles, 645.2.2 Lysosome Exocytosis for Membrane Repair, 655.2.3 Lysosome Exocytosis as a Source of Membrane, 665.2.4 Lysosome Exocytosis for Extracellular Degradation, 665.2.5 Lysosome Exocytosis and Delivery of Proteins to the Cell Surface, 685.3 Mechanisms of Lysosome Exocytosis, 685.3.1 Maturation of Lysosomes and Lysosome-Related Organelles, 695.3.2 Transport of Lysosomes to the Plasma Membrane, 705.3.3 Tethering of Lysosomes to the Plasma Membrane, 725.3.4 Lysosome Fusion with the Plasma Membrane, 755.3.5 Calcium-Dependent Exocytosis, 765.4 Conclusion, 76Acknowledgments, 77References, 776 Role of Lysosomes in Lipid Metabolism 87Frederick R. Maxfield6.1 Introduction, 876.2 Endocytic Uptake of Lipoproteins, 886.3 Lipid Metabolism in Late Endosomes and Lysosomes, 916.4 Autophagy and Lysosomal Lipid Turnover, 946.5 Lysosomal Lipid Hydrolysis and Metabolic Regulation, 956.6 Summary, 96References, 967 TFEB, Master Regulator of Cellular Clearance 101Graciana Diez-Roux and Andrea Ballabio7.1 Lysosome, 1017.2 The Transcriptional Regulation of Lysosomal Function, 1027.3 TFEB Subcellular Regulation is Regulated by Its Phosphorylation, 1047.4 A Lysosome-to-Nucleus Signaling Mechanism, 1057.5 TFEB and Cellular Clearance in Human Disease, 1067.5.1 Lysosomal Storage Disorders, 1077.5.2 Neurodegenerative Disorders, 1097.5.3 Metabolic Syndrome, 1107.5.4 Cancer, Inborn Errors of Metabolism, Immunity, and Longevity, 110References, 1118 Lysosomal Membrane Permeabilization in Cell Death 115Urška Repnik and Boris Turk8.1 Introduction, 1158.2 Cell Death Modalities, 1168.3 Lysosomal Membrane Permeabilization (LMP) and Cell Death, 1178.3.1 Mechanisms of LMP, 1188.3.2 Upstream of LMP: Direct Insult Versus Molecular Signaling, 1218.3.3 Signaling Downstream of LMP, 1248.4 Conclusion, 127Acknowledgments, 127References, 1289 The Lysosome in Aging-Related Neurodegenerative Diseases 137Ralph A. Nixon9.1 Introduction, 1379.2 Lysosome Function in Aging Organisms, 1399.3 Lysosomes and Diseases of Late Age Onset, 1429.3.1 Cardiovascular Disease, 1429.4 Lysosomes in Aging-Related Neurodegenerative Diseases, 1449.4.1 Alzheimer’s Disease (AD), 1459.4.2 Parkinson’s Disease and Related Disorders, 1509.4.3 Diffuse Lewy Body Disease (DLB), 1559.4.4 Frontotemporal Lobar Degeneration (FTLD), 1559.5 Conclusion, 158Acknowledgments, 158References, 15910 Lysosome and Cancer 181Marja Jäättelä and Tuula Kallunki10.1 Introduction, 18110.2 Lysosomal Function and Its Importance for Cancer Development and Progression, 18110.3 Cancer-Induced Changes in Lysosomal Function, 18210.3.1 Increased Activity of Lysosomal Enzymes, 18210.3.2 Altered Lysosome Membrane Permeability, 18410.3.3 Increased Lysosome Size, 18410.3.4 Altered Lysosome Trafficking – Increased Lysosomal Exocytosis, 18510.4 Cancer-Induced Changes in Lysosome Composition, 18510.4.1 Changes in Lysosomal Hydrolases, 18510.4.2 Changes in the Lysosomal Membrane Proteins, 19210.5 Molecular Changes Involving Lysosomal Integrity, 19310.5.1 Cancer-Associated Changes in Lysosomal Sphingolipid Metabolism, 19310.5.2 Targeting Lysosomal Membrane Integrity, 19510.6 Conclusion, 196References, 19711 The Genetics of Sphingolipid Hydrolases and Sphingolipid Storage Diseases 209Edward H. Schuchman and Calogera M. Simonaro11.1 Introduction and Overview, 20911.2 Acid Ceramidase Deficiency: Farber Disease, 21011.3 Acid Sphingomyelinase Deficiency: Types A and B Niemann–Pick Disease, 21311.4 Beta-Glucocerebrosidase Deficiency: Gaucher Disease, 21511.5 Galactocerebrosidase Deficiency: Krabbe Disease/Globoid Cell Leukodystrophy, 21811.6 Arylsulfatase a Deficiency: Metachromatic Leukodystrophy, 21911.7 Alpha-Galactosidase a Deficiency: Fabry Disease, 22111.8 Beta-Galactosidase Deficiency: GM1 Gangliosidosis, 22411.9 Hexosaminidase A and B Deficiency: GM2 Gangliosidoses, 22611.10 Sphingolipid Activator Proteins, 229References, 23112 Lysosome-Related Organelles: Modifications of the Lysosome Paradigm 239Adriana R. Mantegazza and Michael S. Marks12.1 Differences Between LROs and Secretory Granules, 24012.2 Physiological Functions of LROs, 24012.3 LRO Biogenesis, 24412.3.1 Chediak–Higashi Syndrome and Gray Platelet Syndrome, 24412.3.2 Hermansky–Pudlak Syndrome, 24612.3.3 Melanosome Biogenesis, 24712.3.4 HPS and Melanosome Maturation, 24812.3.5 HPS and the Biogenesis of Other LROs, 25012.3.6 HPS and Neurosecretory Granule Biogenesis, 25012.3.7 Weibel–Palade Body Biogenesis, 25112.4 LRO Motility, Docking, and Secretion, 25212.5 LROs and Immunity to Pathogens, 25312.5.1 Cytolytic Granules, 25312.5.2 Familial Hemophagocytic Lymphohistiocytosis and Cytolytic Granule Secretion, 25412.5.3 Azurophilic Granules, 25512.5.4 NADPH Oxidase-Containing LROs, 25512.5.5 IRF7-Signaling LROs and Type I Interferon Induction, 25612.5.6 MIICs: LROs or Conventional Late Endosome/Lysosomes?, 25612.5.7 Phagosomes and Autophagosomes as New Candidate LROs, 25812.6 Perspectives, 260Acknowledgments, 260References, 26013 Autophagy Inhibition as a Strategy for Cancer Therapy 279Xiaohong Ma, Shengfu Piao, Quentin Mcafee, and Ravi K. Amaravadi13.1 Stages and Steps of Autophagy, 28213.2 Induction of Autophagy, 28313.3 Studies in Mouse Models Unravel the Dual Roles of Autophagy in Tumor Biology, 28513.4 Clinical Studies on Autophagy’s Dual Role in Tumorigenesis, 28613.5 Mouse Models Provide the Rationale for Autophagy Modulation in the Context of Cancer Therapy, 28813.6 Multiple Druggable Targets in the Autophagy Pathway, 29113.7 Overview of Preclinical Autophagy Inhibitors and Evidence Supporting Combination with Existing and New Anticancer Agents, 29213.8 Proximal Autophagy Inhibitors, 29313.9 Quinolines: From Antimalarials to Prototypical Distal Autophagy Inhibitors, 29313.10 Summary for the Clinical Trials for CQ/HCQ, 29513.11 Developing More Potent Anticancer Autophagy Inhibitors, 29813.12 Summary, Conclusion, and Future Directions, 30013.13 In Summary, 302References, 30214 Autophagy Enhancers, are we there Yet? 315Shuyan Lu and Ralph A. Nixon14.1 Introduction, 31514.2 Autophagy Impairment and Diseases, 31614.3 Autophagy Enhancer Screening, 31714.3.1 Methods for Monitoring Autophagy, 31714.3.2 Autophagy Enhancers Identified from Early Literature, 32614.3.3 mTOR Inhibitors, 33114.4 Other Agents that Boost Autophagy and Lysosomal Functions, 33514.4.1 HDAC Inhibition, 33614.4.2 pH Restoration, 33714.4.3 TRP Activator, 33714.4.4 TFEB Overexpression/Activation, 33814.4.5 Lysosomal Efficiency, 33814.4.6 MicroRNA, 33914.5 Concluding Remarks, 340References, 34115 Pharmacological Chaperones as Potential Therapeutics for Lysosomal Storage Disorders: Preclinical Research to Clinical Studies 357Robert E. Boyd, Elfrida R. Benjamin, Su Xu, Richie Khanna, and Kenneth J. Valenzano15.1 Introduction, 35715.2 Fabry Disease, 35915.3 Gaucher Disease, 36315.4 GM2 Gangliosidoses (Tay–Sachs/Sandhoff Diseases), 36715.5 Pompe Disease, 36815.6 PC-ERT Combination Therapy, 370References, 37216 Endosomal Escape Pathways for Delivery of Biologics 383Philip L. Leopold16.1 Introduction, 38316.2 Endosome Characteristics, 38416.3 Delivery of Nature’s Biologics: Lessons on Endosomal Escape from Pathogens, 38916.3.1 Viruses, 39016.3.2 Bacteria, Protozoa, and Fungi, 39216.3.3 Toxins, 39416.4 Endosomal Escape Using Engineered Systems, 39516.4.1 Peptides and Polymers, 39616.4.2 Lipids, 39816.4.3 Other Chemical and Physical Strategies, 39916.5 Conclusion, 399References, 40017 Lysosomes and Antibody–Drug Conjugates 409Michelle Mack, Jennifer Kahler, Boris Shor, Michael Ritchie, Maureen Dougher, Matthew Sung, and Puja Sapra17.1 Introduction, 40917.2 Receptor Internalization, 41017.3 Antibody–Drug Conjugates, 41317.4 Mechanisms of Resistance to ADCs, 41617.5 Summary, 417References, 41718 The Mechanisms and Therapeutic Consequences of Amine-Containing Drug Sequestration in Lysosomes 423Nadia Hamid and Jeffrey P. Krise18.1 Introduction, 42318.2 Lysosomal Trapping Overview, 42418.3 Techniques to Assess Lysosomal Trapping, 42718.4 Influence of Lysosomotropism on Drug Activity, 42918.5 Influence of Lysosomal Trapping on Pharmacokinetics, 43518.6 Pharmacokinetic Drug–Drug Interactions Involving Lysosomes, 438References, 44019 Lysosome Dysfunction: an Emerging Mechanism of Xenobiotic-Induced Toxicity 445Shuyan Lu, Bart Jessen, Yvonne Will, and Greg Stevens19.1 Introduction, 44519.2 Compounds that Impact Lysosomal Function, 44619.2.1 Lysosomotropic Compounds, 44619.2.2 Nonlysosomotropic Compounds, 45119.3 Cellular Consequences, 45219.3.1 Effect of Drugs on pH and Lysosomal Volume, 45219.3.2 Effects on Lysosomal Enzymes, 45319.3.3 Lysosomal Substrate Accumulation, 45419.3.4 Lysosomal Membrane Permeabilization (LMP) and Cell Death, 45419.3.5 Membrane Trafficking Changes, 45519.3.6 Other Cellular Impacts, 45819.4 Impaired Lysosomal Function as a Mechanism for Organ Toxicity, 46119.4.1 Liver Toxicity, 46219.4.2 Kidney Toxicity, 46419.4.3 Retinal, 46619.4.4 Peripheral Neuropathy, 46619.4.5 Muscle Toxicity, 46719.4.6 Tumorigenesis, 46819.4.7 General Considerations for Organ Toxicity, 46919.5 Concluding Remarks, 471References, 47220 Lysosomes and Phospholipidosis in Drug Development and Regulation 487James M. Willard and Albert De Felice20.1 Introduction, 48720.2 FDA Involvement, 48820.3 Autophagy and DIPL, 48920.4 Early Experience with Lethal DIPL, 48920.5 Clinical and Nonclinical Expressions of DIPL, 49020.5.1 Clinical, 49020.5.2 Nonclinical, 49120.6 Physical Chemistry, 49120.7 Quantitative Structure–Activity Relationship (QSAR), 49220.8 Toxicogenomics, 49320.9 Fluorescence, Dye, and Immunohistochemical Methods for Screening, 49420.10 FDA Database and QSAR Modeling, 49420.11 Linking Phospholipidosis and Overt Toxicity, 49420.12 Phospholipidosis and QT Interval Prolongation, 49620.13 DIPL Mechanisms, 50020.14 Treatment, 50120.15 Discussion, 50120.16 Future Directions and Recommendations, 505References, 506INDEX 513

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Frederick R. Maxfield, PhD, is Professor and Chair of the Department of Biochemistry at Weill Cornell Medical College. He has used digital imaging microscopy to characterize pH changes in endocytic organelles, to measure the kinetics of transport of molecules among organelles, and to identify new endocytic organelles such as the endocytic recycling compartment. Dr. Maxfield has published extensively on trafficking of lipids and cholesterol.James M. Willard, PhD, has been a member of the Phospholipidosis Working Group at the Center for Drug Evaluation and Research (CDER) of the Food and Drug Administration since 2005 and Co-Chair of the group since 2011. Shuyan Lu, MSc, has been an Investigative Toxicologist of Drug Research and Development at Pfizer for over 10 years. She studies the role of lysosomal pathways and physical chemical properties of compounds in drug-induced toxicity.

• PREFACE xiiiLIST OF CONTRIBUTORS xvii1 Lysosomes: An Introduction 1Frederick R. Maxfield1.1 Historical Background, 2References, 42 Lysosome Biogenesis and Autophagy 7Fulvio Reggiori and Judith Klumperman2.1 Introduction, 72.2 Pathways to the Lysosomes, 102.2.1 Biosynthetic Transport Routes to the Lysosome, 102.2.2 Endocytic Pathways to the Lysosome, 102.2.3 Autophagy Pathways to the Lysosome, 122.2.4 The ATG Proteins: The Key Regulators of Autophagy, 142.3 Fusion and Fission between the Endolysosomal and Autophagy Pathways, 162.3.1 Recycling Endosomes and Autophagosome Biogenesis, 162.3.2 Autophagosome Fusion with Late Endosomes and Lysosomes, 172.3.3 Autophagic Lysosomal Reformation, 182.4 Diseases, 192.4.1 Lysosome-Related Disorders (LSDs), 192.4.2 Lysosomes in Neurodegeneration and Its Links to Autophagy, 202.4.3 Autophagy-Related Diseases, 202.5 Concluding Remarks, 22Acknowledgments, 23References, 233 Multivesicular Bodies: Roles in Intracellular and Intercellular Signaling 33Emily R. Eden, Thomas Burgoyne, and Clare E. Futter3.1 Introduction, 333.2 Downregulation of Signaling by Sorting onto ILVs, 353.3 Upregulation of Signaling by Sorting onto ILVs, 383.4 Intercellular Signaling Dependent on Sorting onto ILVs, 393.5 Conclusion, 44References, 454 Lysosomes and Mitophagy 51Dominik Haddad and Patrik Verstreken4.1 Summary, 514.2 Mitochondrial Significance, 514.3 History of Mitophagy, 524.4 Mechanisms of Mitophagy, 534.4.1 Mitophagy in Yeast, 544.4.2 Mitophagy in Mammals, 554.5 Conclusion, 57Acknowledgments, 57References, 585 Lysosome Exocytosis and Membrane Repair 63Rajesh K. Singh and Abigail S. Haka5.1 Introduction, 635.2 Functions of Lysosome Exocytosis, 635.2.1 Specialized Lysosome-Related Organelles, 645.2.2 Lysosome Exocytosis for Membrane Repair, 655.2.3 Lysosome Exocytosis as a Source of Membrane, 665.2.4 Lysosome Exocytosis for Extracellular Degradation, 665.2.5 Lysosome Exocytosis and Delivery of Proteins to the Cell Surface, 685.3 Mechanisms of Lysosome Exocytosis, 685.3.1 Maturation of Lysosomes and Lysosome-Related Organelles, 695.3.2 Transport of Lysosomes to the Plasma Membrane, 705.3.3 Tethering of Lysosomes to the Plasma Membrane, 725.3.4 Lysosome Fusion with the Plasma Membrane, 755.3.5 Calcium-Dependent Exocytosis, 765.4 Conclusion, 76Acknowledgments, 77References, 776 Role of Lysosomes in Lipid Metabolism 87Frederick R. Maxfield6.1 Introduction, 876.2 Endocytic Uptake of Lipoproteins, 886.3 Lipid Metabolism in Late Endosomes and Lysosomes, 916.4 Autophagy and Lysosomal Lipid Turnover, 946.5 Lysosomal Lipid Hydrolysis and Metabolic Regulation, 956.6 Summary, 96References, 967 TFEB, Master Regulator of Cellular Clearance 101Graciana Diez-Roux and Andrea Ballabio7.1 Lysosome, 1017.2 The Transcriptional Regulation of Lysosomal Function, 1027.3 TFEB Subcellular Regulation is Regulated by Its Phosphorylation, 1047.4 A Lysosome-to-Nucleus Signaling Mechanism, 1057.5 TFEB and Cellular Clearance in Human Disease, 1067.5.1 Lysosomal Storage Disorders, 1077.5.2 Neurodegenerative Disorders, 1097.5.3 Metabolic Syndrome, 1107.5.4 Cancer, Inborn Errors of Metabolism, Immunity, and Longevity, 110References, 1118 Lysosomal Membrane Permeabilization in Cell Death 115Urška Repnik and Boris Turk8.1 Introduction, 1158.2 Cell Death Modalities, 1168.3 Lysosomal Membrane Permeabilization (LMP) and Cell Death, 1178.3.1 Mechanisms of LMP, 1188.3.2 Upstream of LMP: Direct Insult Versus Molecular Signaling, 1218.3.3 Signaling Downstream of LMP, 1248.4 Conclusion, 127Acknowledgments, 127References, 1289 The Lysosome in Aging-Related Neurodegenerative Diseases 137Ralph A. Nixon9.1 Introduction, 1379.2 Lysosome Function in Aging Organisms, 1399.3 Lysosomes and Diseases of Late Age Onset, 1429.3.1 Cardiovascular Disease, 1429.4 Lysosomes in Aging-Related Neurodegenerative Diseases, 1449.4.1 Alzheimer’s Disease (AD), 1459.4.2 Parkinson’s Disease and Related Disorders, 1509.4.3 Diffuse Lewy Body Disease (DLB), 1559.4.4 Frontotemporal Lobar Degeneration (FTLD), 1559.5 Conclusion, 158Acknowledgments, 158References, 15910 Lysosome and Cancer 181Marja Jäättelä and Tuula Kallunki10.1 Introduction, 18110.2 Lysosomal Function and Its Importance for Cancer Development and Progression, 18110.3 Cancer-Induced Changes in Lysosomal Function, 18210.3.1 Increased Activity of Lysosomal Enzymes, 18210.3.2 Altered Lysosome Membrane Permeability, 18410.3.3 Increased Lysosome Size, 18410.3.4 Altered Lysosome Trafficking – Increased Lysosomal Exocytosis, 18510.4 Cancer-Induced Changes in Lysosome Composition, 18510.4.1 Changes in Lysosomal Hydrolases, 18510.4.2 Changes in the Lysosomal Membrane Proteins, 19210.5 Molecular Changes Involving Lysosomal Integrity, 19310.5.1 Cancer-Associated Changes in Lysosomal Sphingolipid Metabolism, 19310.5.2 Targeting Lysosomal Membrane Integrity, 19510.6 Conclusion, 196References, 19711 The Genetics of Sphingolipid Hydrolases and Sphingolipid Storage Diseases 209Edward H. Schuchman and Calogera M. Simonaro11.1 Introduction and Overview, 20911.2 Acid Ceramidase Deficiency: Farber Disease, 21011.3 Acid Sphingomyelinase Deficiency: Types A and B Niemann–Pick Disease, 21311.4 Beta-Glucocerebrosidase Deficiency: Gaucher Disease, 21511.5 Galactocerebrosidase Deficiency: Krabbe Disease/Globoid Cell Leukodystrophy, 21811.6 Arylsulfatase a Deficiency: Metachromatic Leukodystrophy, 21911.7 Alpha-Galactosidase a Deficiency: Fabry Disease, 22111.8 Beta-Galactosidase Deficiency: GM1 Gangliosidosis, 22411.9 Hexosaminidase A and B Deficiency: GM2 Gangliosidoses, 22611.10 Sphingolipid Activator Proteins, 229References, 23112 Lysosome-Related Organelles: Modifications of the Lysosome Paradigm 239Adriana R. Mantegazza and Michael S. Marks12.1 Differences Between LROs and Secretory Granules, 24012.2 Physiological Functions of LROs, 24012.3 LRO Biogenesis, 24412.3.1 Chediak–Higashi Syndrome and Gray Platelet Syndrome, 24412.3.2 Hermansky–Pudlak Syndrome, 24612.3.3 Melanosome Biogenesis, 24712.3.4 HPS and Melanosome Maturation, 24812.3.5 HPS and the Biogenesis of Other LROs, 25012.3.6 HPS and Neurosecretory Granule Biogenesis, 25012.3.7 Weibel–Palade Body Biogenesis, 25112.4 LRO Motility, Docking, and Secretion, 25212.5 LROs and Immunity to Pathogens, 25312.5.1 Cytolytic Granules, 25312.5.2 Familial Hemophagocytic Lymphohistiocytosis and Cytolytic Granule Secretion, 25412.5.3 Azurophilic Granules, 25512.5.4 NADPH Oxidase-Containing LROs, 25512.5.5 IRF7-Signaling LROs and Type I Interferon Induction, 25612.5.6 MIICs: LROs or Conventional Late Endosome/Lysosomes?, 25612.5.7 Phagosomes and Autophagosomes as New Candidate LROs, 25812.6 Perspectives, 260Acknowledgments, 260References, 26013 Autophagy Inhibition as a Strategy for Cancer Therapy 279Xiaohong Ma, Shengfu Piao, Quentin Mcafee, and Ravi K. Amaravadi13.1 Stages and Steps of Autophagy, 28213.2 Induction of Autophagy, 28313.3 Studies in Mouse Models Unravel the Dual Roles of Autophagy in Tumor Biology, 28513.4 Clinical Studies on Autophagy’s Dual Role in Tumorigenesis, 28613.5 Mouse Models Provide the Rationale for Autophagy Modulation in the Context of Cancer Therapy, 28813.6 Multiple Druggable Targets in the Autophagy Pathway, 29113.7 Overview of Preclinical Autophagy Inhibitors and Evidence Supporting Combination with Existing and New Anticancer Agents, 29213.8 Proximal Autophagy Inhibitors, 29313.9 Quinolines: From Antimalarials to Prototypical Distal Autophagy Inhibitors, 29313.10 Summary for the Clinical Trials for CQ/HCQ, 29513.11 Developing More Potent Anticancer Autophagy Inhibitors, 29813.12 Summary, Conclusion, and Future Directions, 30013.13 In Summary, 302References, 30214 Autophagy Enhancers, are we there Yet? 315Shuyan Lu and Ralph A. Nixon14.1 Introduction, 31514.2 Autophagy Impairment and Diseases, 31614.3 Autophagy Enhancer Screening, 31714.3.1 Methods for Monitoring Autophagy, 31714.3.2 Autophagy Enhancers Identified from Early Literature, 32614.3.3 mTOR Inhibitors, 33114.4 Other Agents that Boost Autophagy and Lysosomal Functions, 33514.4.1 HDAC Inhibition, 33614.4.2 pH Restoration, 33714.4.3 TRP Activator, 33714.4.4 TFEB Overexpression/Activation, 33814.4.5 Lysosomal Efficiency, 33814.4.6 MicroRNA, 33914.5 Concluding Remarks, 340References, 34115 Pharmacological Chaperones as Potential Therapeutics for Lysosomal Storage Disorders: Preclinical Research to Clinical Studies 357Robert E. Boyd, Elfrida R. Benjamin, Su Xu, Richie Khanna, and Kenneth J. Valenzano15.1 Introduction, 35715.2 Fabry Disease, 35915.3 Gaucher Disease, 36315.4 GM2 Gangliosidoses (Tay–Sachs/Sandhoff Diseases), 36715.5 Pompe Disease, 36815.6 PC-ERT Combination Therapy, 370References, 37216 Endosomal Escape Pathways for Delivery of Biologics 383Philip L. Leopold16.1 Introduction, 38316.2 Endosome Characteristics, 38416.3 Delivery of Nature’s Biologics: Lessons on Endosomal Escape from Pathogens, 38916.3.1 Viruses, 39016.3.2 Bacteria, Protozoa, and Fungi, 39216.3.3 Toxins, 39416.4 Endosomal Escape Using Engineered Systems, 39516.4.1 Peptides and Polymers, 39616.4.2 Lipids, 39816.4.3 Other Chemical and Physical Strategies, 39916.5 Conclusion, 399References, 40017 Lysosomes and Antibody–Drug Conjugates 409Michelle Mack, Jennifer Kahler, Boris Shor, Michael Ritchie, Maureen Dougher, Matthew Sung, and Puja Sapra17.1 Introduction, 40917.2 Receptor Internalization, 41017.3 Antibody–Drug Conjugates, 41317.4 Mechanisms of Resistance to ADCs, 41617.5 Summary, 417References, 41718 The Mechanisms and Therapeutic Consequences of Amine-Containing Drug Sequestration in Lysosomes 423Nadia Hamid and Jeffrey P. Krise18.1 Introduction, 42318.2 Lysosomal Trapping Overview, 42418.3 Techniques to Assess Lysosomal Trapping, 42718.4 Influence of Lysosomotropism on Drug Activity, 42918.5 Influence of Lysosomal Trapping on Pharmacokinetics, 43518.6 Pharmacokinetic Drug–Drug Interactions Involving Lysosomes, 438References, 44019 Lysosome Dysfunction: an Emerging Mechanism of Xenobiotic-Induced Toxicity 445Shuyan Lu, Bart Jessen, Yvonne Will, and Greg Stevens19.1 Introduction, 44519.2 Compounds that Impact Lysosomal Function, 44619.2.1 Lysosomotropic Compounds, 44619.2.2 Nonlysosomotropic Compounds, 45119.3 Cellular Consequences, 45219.3.1 Effect of Drugs on pH and Lysosomal Volume, 45219.3.2 Effects on Lysosomal Enzymes, 45319.3.3 Lysosomal Substrate Accumulation, 45419.3.4 Lysosomal Membrane Permeabilization (LMP) and Cell Death, 45419.3.5 Membrane Trafficking Changes, 45519.3.6 Other Cellular Impacts, 45819.4 Impaired Lysosomal Function as a Mechanism for Organ Toxicity, 46119.4.1 Liver Toxicity, 46219.4.2 Kidney Toxicity, 46419.4.3 Retinal, 46619.4.4 Peripheral Neuropathy, 46619.4.5 Muscle Toxicity, 46719.4.6 Tumorigenesis, 46819.4.7 General Considerations for Organ Toxicity, 46919.5 Concluding Remarks, 471References, 47220 Lysosomes and Phospholipidosis in Drug Development and Regulation 487James M. Willard and Albert De Felice20.1 Introduction, 48720.2 FDA Involvement, 48820.3 Autophagy and DIPL, 48920.4 Early Experience with Lethal DIPL, 48920.5 Clinical and Nonclinical Expressions of DIPL, 49020.5.1 Clinical, 49020.5.2 Nonclinical, 49120.6 Physical Chemistry, 49120.7 Quantitative Structure–Activity Relationship (QSAR), 49220.8 Toxicogenomics, 49320.9 Fluorescence, Dye, and Immunohistochemical Methods for Screening, 49420.10 FDA Database and QSAR Modeling, 49420.11 Linking Phospholipidosis and Overt Toxicity, 49420.12 Phospholipidosis and QT Interval Prolongation, 49620.13 DIPL Mechanisms, 50020.14 Treatment, 50120.15 Discussion, 50120.16 Future Directions and Recommendations, 505References, 506INDEX 513

Select Guide Rating

Frederick R. Maxfield, PhD, is Professor and Chair of the Department of Biochemistry at Weill Cornell Medical College. He has used digital imaging microscopy to characterize pH changes in endocytic organelles, to measure the kinetics of transport of molecules among organelles, and to identify new endocytic organelles such as the endocytic recycling compartment. Dr. Maxfield has published extensively on trafficking of lipids and cholesterol.James M. Willard, PhD, has been a member of the Phospholipidosis Working Group at the Center for Drug Evaluation and Research (CDER) of the Food and Drug Administration since 2005 and Co-Chair of the group since 2011. Shuyan Lu, MSc, has been an Investigative Toxicologist of Drug Research and Development at Pfizer for over 10 years. She studies the role of lysosomal pathways and physical chemical properties of compounds in drug-induced toxicity.

• PREFACE xiiiLIST OF CONTRIBUTORS xvii1 Lysosomes: An Introduction 1Frederick R. Maxfield1.1 Historical Background, 2References, 42 Lysosome Biogenesis and Autophagy 7Fulvio Reggiori and Judith Klumperman2.1 Introduction, 72.2 Pathways to the Lysosomes, 102.2.1 Biosynthetic Transport Routes to the Lysosome, 102.2.2 Endocytic Pathways to the Lysosome, 102.2.3 Autophagy Pathways to the Lysosome, 122.2.4 The ATG Proteins: The Key Regulators of Autophagy, 142.3 Fusion and Fission between the Endolysosomal and Autophagy Pathways, 162.3.1 Recycling Endosomes and Autophagosome Biogenesis, 162.3.2 Autophagosome Fusion with Late Endosomes and Lysosomes, 172.3.3 Autophagic Lysosomal Reformation, 182.4 Diseases, 192.4.1 Lysosome-Related Disorders (LSDs), 192.4.2 Lysosomes in Neurodegeneration and Its Links to Autophagy, 202.4.3 Autophagy-Related Diseases, 202.5 Concluding Remarks, 22Acknowledgments, 23References, 233 Multivesicular Bodies: Roles in Intracellular and Intercellular Signaling 33Emily R. Eden, Thomas Burgoyne, and Clare E. Futter3.1 Introduction, 333.2 Downregulation of Signaling by Sorting onto ILVs, 353.3 Upregulation of Signaling by Sorting onto ILVs, 383.4 Intercellular Signaling Dependent on Sorting onto ILVs, 393.5 Conclusion, 44References, 454 Lysosomes and Mitophagy 51Dominik Haddad and Patrik Verstreken4.1 Summary, 514.2 Mitochondrial Significance, 514.3 History of Mitophagy, 524.4 Mechanisms of Mitophagy, 534.4.1 Mitophagy in Yeast, 544.4.2 Mitophagy in Mammals, 554.5 Conclusion, 57Acknowledgments, 57References, 585 Lysosome Exocytosis and Membrane Repair 63Rajesh K. Singh and Abigail S. Haka5.1 Introduction, 635.2 Functions of Lysosome Exocytosis, 635.2.1 Specialized Lysosome-Related Organelles, 645.2.2 Lysosome Exocytosis for Membrane Repair, 655.2.3 Lysosome Exocytosis as a Source of Membrane, 665.2.4 Lysosome Exocytosis for Extracellular Degradation, 665.2.5 Lysosome Exocytosis and Delivery of Proteins to the Cell Surface, 685.3 Mechanisms of Lysosome Exocytosis, 685.3.1 Maturation of Lysosomes and Lysosome-Related Organelles, 695.3.2 Transport of Lysosomes to the Plasma Membrane, 705.3.3 Tethering of Lysosomes to the Plasma Membrane, 725.3.4 Lysosome Fusion with the Plasma Membrane, 755.3.5 Calcium-Dependent Exocytosis, 765.4 Conclusion, 76Acknowledgments, 77References, 776 Role of Lysosomes in Lipid Metabolism 87Frederick R. Maxfield6.1 Introduction, 876.2 Endocytic Uptake of Lipoproteins, 886.3 Lipid Metabolism in Late Endosomes and Lysosomes, 916.4 Autophagy and Lysosomal Lipid Turnover, 946.5 Lysosomal Lipid Hydrolysis and Metabolic Regulation, 956.6 Summary, 96References, 967 TFEB, Master Regulator of Cellular Clearance 101Graciana Diez-Roux and Andrea Ballabio7.1 Lysosome, 1017.2 The Transcriptional Regulation of Lysosomal Function, 1027.3 TFEB Subcellular Regulation is Regulated by Its Phosphorylation, 1047.4 A Lysosome-to-Nucleus Signaling Mechanism, 1057.5 TFEB and Cellular Clearance in Human Disease, 1067.5.1 Lysosomal Storage Disorders, 1077.5.2 Neurodegenerative Disorders, 1097.5.3 Metabolic Syndrome, 1107.5.4 Cancer, Inborn Errors of Metabolism, Immunity, and Longevity, 110References, 1118 Lysosomal Membrane Permeabilization in Cell Death 115Urška Repnik and Boris Turk8.1 Introduction, 1158.2 Cell Death Modalities, 1168.3 Lysosomal Membrane Permeabilization (LMP) and Cell Death, 1178.3.1 Mechanisms of LMP, 1188.3.2 Upstream of LMP: Direct Insult Versus Molecular Signaling, 1218.3.3 Signaling Downstream of LMP, 1248.4 Conclusion, 127Acknowledgments, 127References, 1289 The Lysosome in Aging-Related Neurodegenerative Diseases 137Ralph A. Nixon9.1 Introduction, 1379.2 Lysosome Function in Aging Organisms, 1399.3 Lysosomes and Diseases of Late Age Onset, 1429.3.1 Cardiovascular Disease, 1429.4 Lysosomes in Aging-Related Neurodegenerative Diseases, 1449.4.1 Alzheimer’s Disease (AD), 1459.4.2 Parkinson’s Disease and Related Disorders, 1509.4.3 Diffuse Lewy Body Disease (DLB), 1559.4.4 Frontotemporal Lobar Degeneration (FTLD), 1559.5 Conclusion, 158Acknowledgments, 158References, 15910 Lysosome and Cancer 181Marja Jäättelä and Tuula Kallunki10.1 Introduction, 18110.2 Lysosomal Function and Its Importance for Cancer Development and Progression, 18110.3 Cancer-Induced Changes in Lysosomal Function, 18210.3.1 Increased Activity of Lysosomal Enzymes, 18210.3.2 Altered Lysosome Membrane Permeability, 18410.3.3 Increased Lysosome Size, 18410.3.4 Altered Lysosome Trafficking – Increased Lysosomal Exocytosis, 18510.4 Cancer-Induced Changes in Lysosome Composition, 18510.4.1 Changes in Lysosomal Hydrolases, 18510.4.2 Changes in the Lysosomal Membrane Proteins, 19210.5 Molecular Changes Involving Lysosomal Integrity, 19310.5.1 Cancer-Associated Changes in Lysosomal Sphingolipid Metabolism, 19310.5.2 Targeting Lysosomal Membrane Integrity, 19510.6 Conclusion, 196References, 19711 The Genetics of Sphingolipid Hydrolases and Sphingolipid Storage Diseases 209Edward H. Schuchman and Calogera M. Simonaro11.1 Introduction and Overview, 20911.2 Acid Ceramidase Deficiency: Farber Disease, 21011.3 Acid Sphingomyelinase Deficiency: Types A and B Niemann–Pick Disease, 21311.4 Beta-Glucocerebrosidase Deficiency: Gaucher Disease, 21511.5 Galactocerebrosidase Deficiency: Krabbe Disease/Globoid Cell Leukodystrophy, 21811.6 Arylsulfatase a Deficiency: Metachromatic Leukodystrophy, 21911.7 Alpha-Galactosidase a Deficiency: Fabry Disease, 22111.8 Beta-Galactosidase Deficiency: GM1 Gangliosidosis, 22411.9 Hexosaminidase A and B Deficiency: GM2 Gangliosidoses, 22611.10 Sphingolipid Activator Proteins, 229References, 23112 Lysosome-Related Organelles: Modifications of the Lysosome Paradigm 239Adriana R. Mantegazza and Michael S. Marks12.1 Differences Between LROs and Secretory Granules, 24012.2 Physiological Functions of LROs, 24012.3 LRO Biogenesis, 24412.3.1 Chediak–Higashi Syndrome and Gray Platelet Syndrome, 24412.3.2 Hermansky–Pudlak Syndrome, 24612.3.3 Melanosome Biogenesis, 24712.3.4 HPS and Melanosome Maturation, 24812.3.5 HPS and the Biogenesis of Other LROs, 25012.3.6 HPS and Neurosecretory Granule Biogenesis, 25012.3.7 Weibel–Palade Body Biogenesis, 25112.4 LRO Motility, Docking, and Secretion, 25212.5 LROs and Immunity to Pathogens, 25312.5.1 Cytolytic Granules, 25312.5.2 Familial Hemophagocytic Lymphohistiocytosis and Cytolytic Granule Secretion, 25412.5.3 Azurophilic Granules, 25512.5.4 NADPH Oxidase-Containing LROs, 25512.5.5 IRF7-Signaling LROs and Type I Interferon Induction, 25612.5.6 MIICs: LROs or Conventional Late Endosome/Lysosomes?, 25612.5.7 Phagosomes and Autophagosomes as New Candidate LROs, 25812.6 Perspectives, 260Acknowledgments, 260References, 26013 Autophagy Inhibition as a Strategy for Cancer Therapy 279Xiaohong Ma, Shengfu Piao, Quentin Mcafee, and Ravi K. Amaravadi13.1 Stages and Steps of Autophagy, 28213.2 Induction of Autophagy, 28313.3 Studies in Mouse Models Unravel the Dual Roles of Autophagy in Tumor Biology, 28513.4 Clinical Studies on Autophagy’s Dual Role in Tumorigenesis, 28613.5 Mouse Models Provide the Rationale for Autophagy Modulation in the Context of Cancer Therapy, 28813.6 Multiple Druggable Targets in the Autophagy Pathway, 29113.7 Overview of Preclinical Autophagy Inhibitors and Evidence Supporting Combination with Existing and New Anticancer Agents, 29213.8 Proximal Autophagy Inhibitors, 29313.9 Quinolines: From Antimalarials to Prototypical Distal Autophagy Inhibitors, 29313.10 Summary for the Clinical Trials for CQ/HCQ, 29513.11 Developing More Potent Anticancer Autophagy Inhibitors, 29813.12 Summary, Conclusion, and Future Directions, 30013.13 In Summary, 302References, 30214 Autophagy Enhancers, are we there Yet? 315Shuyan Lu and Ralph A. Nixon14.1 Introduction, 31514.2 Autophagy Impairment and Diseases, 31614.3 Autophagy Enhancer Screening, 31714.3.1 Methods for Monitoring Autophagy, 31714.3.2 Autophagy Enhancers Identified from Early Literature, 32614.3.3 mTOR Inhibitors, 33114.4 Other Agents that Boost Autophagy and Lysosomal Functions, 33514.4.1 HDAC Inhibition, 33614.4.2 pH Restoration, 33714.4.3 TRP Activator, 33714.4.4 TFEB Overexpression/Activation, 33814.4.5 Lysosomal Efficiency, 33814.4.6 MicroRNA, 33914.5 Concluding Remarks, 340References, 34115 Pharmacological Chaperones as Potential Therapeutics for Lysosomal Storage Disorders: Preclinical Research to Clinical Studies 357Robert E. Boyd, Elfrida R. Benjamin, Su Xu, Richie Khanna, and Kenneth J. Valenzano15.1 Introduction, 35715.2 Fabry Disease, 35915.3 Gaucher Disease, 36315.4 GM2 Gangliosidoses (Tay–Sachs/Sandhoff Diseases), 36715.5 Pompe Disease, 36815.6 PC-ERT Combination Therapy, 370References, 37216 Endosomal Escape Pathways for Delivery of Biologics 383Philip L. Leopold16.1 Introduction, 38316.2 Endosome Characteristics, 38416.3 Delivery of Nature’s Biologics: Lessons on Endosomal Escape from Pathogens, 38916.3.1 Viruses, 39016.3.2 Bacteria, Protozoa, and Fungi, 39216.3.3 Toxins, 39416.4 Endosomal Escape Using Engineered Systems, 39516.4.1 Peptides and Polymers, 39616.4.2 Lipids, 39816.4.3 Other Chemical and Physical Strategies, 39916.5 Conclusion, 399References, 40017 Lysosomes and Antibody–Drug Conjugates 409Michelle Mack, Jennifer Kahler, Boris Shor, Michael Ritchie, Maureen Dougher, Matthew Sung, and Puja Sapra17.1 Introduction, 40917.2 Receptor Internalization, 41017.3 Antibody–Drug Conjugates, 41317.4 Mechanisms of Resistance to ADCs, 41617.5 Summary, 417References, 41718 The Mechanisms and Therapeutic Consequences of Amine-Containing Drug Sequestration in Lysosomes 423Nadia Hamid and Jeffrey P. Krise18.1 Introduction, 42318.2 Lysosomal Trapping Overview, 42418.3 Techniques to Assess Lysosomal Trapping, 42718.4 Influence of Lysosomotropism on Drug Activity, 42918.5 Influence of Lysosomal Trapping on Pharmacokinetics, 43518.6 Pharmacokinetic Drug–Drug Interactions Involving Lysosomes, 438References, 44019 Lysosome Dysfunction: an Emerging Mechanism of Xenobiotic-Induced Toxicity 445Shuyan Lu, Bart Jessen, Yvonne Will, and Greg Stevens19.1 Introduction, 44519.2 Compounds that Impact Lysosomal Function, 44619.2.1 Lysosomotropic Compounds, 44619.2.2 Nonlysosomotropic Compounds, 45119.3 Cellular Consequences, 45219.3.1 Effect of Drugs on pH and Lysosomal Volume, 45219.3.2 Effects on Lysosomal Enzymes, 45319.3.3 Lysosomal Substrate Accumulation, 45419.3.4 Lysosomal Membrane Permeabilization (LMP) and Cell Death, 45419.3.5 Membrane Trafficking Changes, 45519.3.6 Other Cellular Impacts, 45819.4 Impaired Lysosomal Function as a Mechanism for Organ Toxicity, 46119.4.1 Liver Toxicity, 46219.4.2 Kidney Toxicity, 46419.4.3 Retinal, 46619.4.4 Peripheral Neuropathy, 46619.4.5 Muscle Toxicity, 46719.4.6 Tumorigenesis, 46819.4.7 General Considerations for Organ Toxicity, 46919.5 Concluding Remarks, 471References, 47220 Lysosomes and Phospholipidosis in Drug Development and Regulation 487James M. Willard and Albert De Felice20.1 Introduction, 48720.2 FDA Involvement, 48820.3 Autophagy and DIPL, 48920.4 Early Experience with Lethal DIPL, 48920.5 Clinical and Nonclinical Expressions of DIPL, 49020.5.1 Clinical, 49020.5.2 Nonclinical, 49120.6 Physical Chemistry, 49120.7 Quantitative Structure–Activity Relationship (QSAR), 49220.8 Toxicogenomics, 49320.9 Fluorescence, Dye, and Immunohistochemical Methods for Screening, 49420.10 FDA Database and QSAR Modeling, 49420.11 Linking Phospholipidosis and Overt Toxicity, 49420.12 Phospholipidosis and QT Interval Prolongation, 49620.13 DIPL Mechanisms, 50020.14 Treatment, 50120.15 Discussion, 50120.16 Future Directions and Recommendations, 505References, 506INDEX 513

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