Insertional Mutagenesis Strategies in Cancer Genetics (eBook)

Adam Dupuy, Ph.D., is an Assistant Professor at the Carver College of Medicine at the University of Iowa, and a rising star in the field of insertional mutagenesis; two of the last of his publications have been in Nature, and he was the lead author on one of these. Dr. Dupuy earned his B.A. at Central College in Pella, Iowar, went on to earn his Ph.D. at the University of Minnesota in the Program in Molecular, Cellular, Developmental Biology & Genetics, and did his Postdoctoral fellowship in the Mouse Cancer Genetics Program at the NCI-Frederick.

Contents 5
Contributors 6
1 Insertional Mutagenesis: A Powerful Tool in Cancer Research 8
1.1 Introduction 8
1.2 Historical Perspective 9
1.3 The Site of Insertion can Provide Mechanistic Insights 11
1.4 Methodologies 12
1.5 Pairing Insertional Mutagenesis with Other Genetic Tools 15
1.6 Mining Hidden Information in Mutagenesis Screens 17
1.7 Importance of Thorough Bioinformatics Analysis 19
1.8 The Added Value of Large Numbers 20
1.9 Crossvalidation with Other Genetic Datasets 20
1.10 Are New Cancer Causing Pathways to be Found 21
1.11 Conclusions and Recommendations 22
References 23
2 Retroviral Insertional Mutagenesis in Mouse Models of Leukemia and Lymphoma 26
2.1 Introduction to the Murine Leukemia Viruses 26
2.2 Viral and Host Determinants of Disease 28
2.3 Challenges in MLV Mutagenesis Studies 30
2.4 Comparative Oncogenomics 36
2.5 Validation of Candidates 38
2.6 Do Murine Retroviruses Cause Human Cancer 40
2.7 Summary and Future Perspectives 40
References 41
3 Gene Discovery by MMTV Mediated Insertional Mutagenesis 46
3.1 Introduction 46
3.1.1 Mouse Mammary Gland Biology 48
3.1.2 Mouse Mammary Tumor Virus 48
3.1.3 Viral Genes and Genomic Organization 50
3.1.4 Transcription and Translation of the Viral Genes 51
3.1.5 Virion Structure and Assembly 52
3.1.6 Infectious Life Cycle 53
3.1.7 Tissue Specificity and Hormonal Regulation of MMTV Replication 54
3.1.8 Exogenous MMTV 55
3.1.9 Endogenous MMTV 56
3.1.10 MMTV-Induced Tumors 56
3.1.11 MMTV Induced Hormone Dependent Premalignant Lesions 57
3.1.12 Hormone-Dependent Mammary Tumors and MMTV 58
3.1.13 MMTV-Induced Tumors in other Tissues 58
3.2 Gene Discovery by MMTV Mediated Insertional Mutagenesis 59
3.2.1 Common Integration Sites 59
3.2.2 Modes by Which Insertional Mutagenesis Affects Cancer Genes 60
3.3 Gene Families and Pathways Frequently Targeted by MMTV 63
3.3.1 Infrequently Targeted Genes and Pathways 65
3.4 Comparison of MMTV Targets in Mammary Tumors and MuLV Targets in Lymphomas 66
3.5 Core and Sporadic Activated Cancer Pathways 66
3.5.1 Wnt Signaling 67
3.5.2 Fgf Signaling 68
3.5.3 Notch Signaling 69
3.6 MMTV Targets and Mammary Stem Cells 70
3.7 Relevance of Insertional Mutagenesis for Human Breast Cancer 70
3.8 Concluding Remarks 72
References 73
4 Chicken Models of Retroviral Insertional Mutagenesis 83
4.1 The Beginning of the Story: the Case of Chicken Bursal Lymphomas 85
4.2 Efficient Activation of C_MYC Requires Defective Provirus: the Transcriptional Interference and Related Phenomena 87
4.3 Readthrough Activation of CMYB in B - lymphomas: the Case of Superactivating ALV 93
4.4 Insertional Activation Can Be Accompanied by Extensive Alterations of the Oncogene Structure and by Formation of an Oncogene - Transducing Virus: the Case of Chicken Erythroblastosis 94
4.5 Chicken Nephroblastomas Induced by MAV: Complex Model Suitable for High Throughput Oncogene Screening 96
4.6 Extension of Spectrum of MAV-Induced Tumors By Local Homeostasis Perturbation: Lung Sarcomas, Liver Carcinomas and the Industasis Phenomenon 100
4.7 Evidence of Multistage Cancerogenesis in Chicken Models 103
4.8 The Future of Models Based on Chicken Retroviruses 104
References 109
5 Sleeping Beauty Models of Cancer 119
5.1 Introduction 119
5.2 Resurrecting a DNA Fossil: the Sleeping Beauty Transposon System 120
5.3 Transposase/Transposon Structure 120
5.4 Molecular Characteristics of SB Transposition 121
5.4.1 Transposition Mechanism 121
5.4.2 Integration Site Bias 123
5.4.3 Local Hopping 124
5.4.4 Integration Site Mapping 124
5.5 Sleeping Beauty in Cancer Research 125
5.5.1 Modifications of Transposon Design 125
5.5.2 Initial Tumor Models Induced by Transposition 126
5.5.3 Identification of Driver Mutations in SB-Induced Tumors 128
5.6 Modifications to the SB System 129
5.7 Future Directions 131
5.7.1 Identifying Genes Involved in Metastasis 131
5.7.2 Identifying Genes Involved in Treatment Response 131
5.7.3 Identifying Cooperating Mutations 132
5.8 Challenges Remaining 132
References 133
6 Insertional Mutagenesis in Hematopoietic Cells: Lessons Learned from Adverse Events in Clinical Gene Therapy Trials 137
6.1 Introduction 137
6.2 Experience with X-Linked Severe Combined Immunodeficiency (SCID-X1) 141
6.3 Experience with Adenosine Deaminase Deficiency Severe Combined Immunodeficiency 145
6.4 Experience with Chronic Granulomatous Disease 146
6.5 Preliminary Data from Other Trials 148
6.5.1 Wiskott-Aldrich Syndrome (WAS) 148
6.5.2 X-Linked Adrenoleukodystrophy (ALD) 149
6.5.3 Thalassemia 149
6.6 Lessons Learned and the Way Forward 151
6.6.1 Advances in the Analysis of Integration Site Preference 151
6.6.2 Novel Model Systems to Characterize Insertional Mutagenesis 153
6.6.3 Disease Specific Effects Upon Insertional Mutagenesis 155
6.6.4 Modification of Vector Architecture to Minimize Insertional Mutagenesis 157
6.7 Summary 160
References 161
7 Bioinformatics of High-Throughput Insertional Mutagenesis 172
7.1 Sequence Analysis Pipeline and Database 172
7.1.1 Sequencing and Barcoding Technologies 172
7.2 Sequence Mapping 173
7.3 Target Gene Identification 174
7.4 Insertional Mutagenesis Database 175
7.5 Detection of Common Insertion Sites 176
7.5.1 Statistical Approaches of Common Insertion Sites Detection 176
7.6 Bias of Insertion Site Preference in Genome 177
7.7 Pathway and Network Analysis 179
7.8 Overview of Pathway and Network Analysis 180
7.9 Pathway and Network Analysis of Insertional Mutagenesis Screening Data 182
7.10 Integrative Discovery with Other Sources of HTP Data 186
7.11 Conclusions 189
References 190
Index 194