Diagnostic Pathology: Molecular Oncology

Mohammad A. Vasef, MD, is a Professor at the Department of Pathology and Director of MGP Fellowship Program at The University of New Mexico Health Sciences Center. He is also Medical Director of Molecular Diagnostics, and works for TriCore Reference Laboratories in Albuquerque, New Mexico. Aaron Auerbach, MD, MPH, is the senior hematopathologist at the Joint Pathology Center in Silver Spring, Maryland, where he delivers expertise in both hematopathology and gastrointestinal pathology. He is the author of the prior and upcoming editions of Diagnostic Pathology: Molecular Oncology (2019 and 2023), and the sole author of Diagnostic Pathology: Spleen (2022).

SECTION 1: INTRODUCTION 4 Structure of Chromosomes and Nucleic Acids Jeremy C. Wallentine, MD 8 DNA Replication, Transcription, and Translation Jeremy C. Wallentine, MD 12 Gene Mutations Jeremy C. Wallentine, MD SECTION 2: COMMON TECHNIQUES FOR ANALYSIS OF ALTERATIONS IN CHROMOSOMES AND DNA 18 Cytogenetics and FISH Mohammad A. Vasef, MD and David R. Czuchlewski, MD 22 Chromosomal Microarray Mohammad A. Vasef, MD and Sibel Kantarci, PhD 26 Amplification Methods Devon Chabot-Richards, MD 30 DNA High-Resolution Melting Curve Analysis Amer Mahmoud, MD, FCAP 32 Sequencing Technologies Eric Y. Loo, MD and Mohammad A. Vasef, MD 36 High-Throughput Methods in Molecular Pathology Rama Gullapalli, MD, PhD 38 DNA Methylation Analysis Devon Chabot-Richards, MD SECTION 3: MOLECULAR GENETIC TESTS IN HEMATOPOIETIC TUMORS 42 BCR::ABL1 Fusion Eric Y. Loo, MD and Mohammad A. Vasef, MD 46 PML::RARA Fusion Devon Chabot-Richards, MD 48 Reverse Transcription PCR for Myeloid Leukemia Transcripts Kristin Hunt Karner, MD 50 FLT3, NPM1, and CEBPA Mutations Devon Chabot-Richards, MD 54 JAK2 Mutations and Rearrangements Mohammad A. Vasef, MD 56 MPL Mutations Eric Y. Loo, MD and David R. Czuchlewski, MD 58 Calreticulin (CALR) Mutations Eric Y. Loo, MD and Mohammad A. Vasef, MD 60 CSF3R Mutations Amer Mahmoud, MD, FCAP 62 SETBP1 Mutations Amer Mahmoud, MD, FCAP 64 KIT Mutations Eric Y. Loo, MD and Mohammad A. Vasef, MD 68 IDH1 and IDH2 Mutations Eric Y. Loo, MD and David R. Czuchlewski, MD 70 Other Mutations and Gene Panel Testing in Myeloid Neoplasms Mohammad A. Vasef, MD and David R. Czuchlewski, MD 72 Immunoglobulin Heavy (IGH) Chain Gene Rearrangements Mohammad A. Vasef, MD and Anna P. Matynia, MD 76 T-Cell Receptor Gamma (TRG) and Delta (TRD) Chain Rearrangements Mohammad A. Vasef, MD and Anna P. Matynia, MD 80 T-Cell Receptor Beta (TRB) Chain Gene Rearrangements Mohammad A. Vasef, MD and Anna P. Matynia, MD 84 Southern Blot Analysis of Antigen Receptor Genes Mohammad A. Vasef, MD 86 EBV-Associated Human Neoplasms Mohammad A. Vasef, MD 94 MYD88 Mutation Giovanni Insuasti-Beltran, MD 96 NOTCH1 Mutation Giovanni Insuasti-Beltran, MD 98 EZH2 Mutation Amer Mahmoud, MD, FCAP SECTION 4: MOLECULAR GENETIC TESTS IN SOLID TUMORS, BONE AND SOFT TISSUE TUMORS, AND CNS TUMORS 104 ALK Rearrangements and Mutations Guanghua Wang, MD 106 ROS1 Rearrangements Mohammad A. Vasef, MD and David R. Czuchlewski, MD 108 RET Rearrangements Eric Y. Loo, MD and David R. Czuchlewski, MD 110 FISH for MET Amplifications Devon Chabot-Richards, MD 114 FISH for ERBB2 (HER2) Amplifications M. Carmen Frias Kletecka, MD and Kristin Hunt Karner, MD 116 KRAS Mutations Guanghua Wang, MD 118 BRAF Mutations Yongjie Zhou, MD, PhD and Guanghua Wang, MD 120 EGFR Mutations Guanghua Wang, MD 122 Targeted Hotspot Gene Panel Using Massively Parallel Sequencing (Next-Generation Sequencing) Mohammad A. Vasef, MD and David R. Czuchlewski, MD 124 Targeted Hotspot Gene Panel Table Mohammad A. Vasef, MD and David R. Czuchlewski, MD 130 MGMT Promoter Gene Methylation Assay Devon Chabot-Richards, MD SECTION 5: MOLECULAR PATHOLOGY OF MYELOID NEOPLASMS 134 Molecular Work-Up of Myeloid Neoplasms Mohammad A. Vasef, MD and David R. Czuchlewski, MD MYELOPROLIFERATIVE NEOPLASMS 136 Chronic Myeloid Leukemia, BCR::ABL1-Positive Mohammad A. Vasef, MD, Kaaren K. Reichard, MD, and David R. Czuchlewski, MD 144 Chronic Neutrophilic Leukemia Devon Chabot-Richards, MD and David R. Czuchlewski, MD 148 Polycythemia Vera Mohammad A. Vasef, MD and M. Carmen Frias Kletecka, MD 152 Primary Myelofibrosis Devon Chabot-Richards, MD, Kaaren K. Reichard, MD, and Daniel Babu, MD 156 Essential Thrombocythemia Giovanni Insuasti-Beltran, MD 160 Chronic Eosinophilic Leukemia, NOS Devon Chabot-Richards, MD, Kaaren K. Reichard, MD, and Daniel Babu, MD 162 Myeloproliferative Neoplasm, Unclassifiable Devon Chabot-Richards, MD, Kaaren K. Reichard, MD, and Daniel Babu, MD MASTOCYTOSIS 166 Mastocytosis Mohammad A. Vasef, MD MYELOID/LYMPHOID NEOPLASMS WITH EOSINOPHILIA AND GENE REARRANGEMENT 174 Myeloid/Lymphoid Neoplasms With Eosinophilia and Tyrosine Kinase Gene Fusions Devon Chabot-Richards, MD and David R. Czuchlewski, MD PREMALIGNANT CLONAL HEMATOPOIESIS AND CYTOPENIAS 178 VEXAS Syndrome Mohammad A. Vasef, MD and Michael M. Franklin, DO 182 Clonal Hematopoiesis and Premalignant Clonal Cytopenia Mohammad A. Vasef, MD MYELODYSPLASTIC SYNDROME (MDS) 184 Overview and Classification of Myelodysplastic Syndrome Devon Chabot-Richards, MD, David R. Czuchlewski, MD, and Carla S. Wilson, MD, PhD 188 Myelodysplastic Syndrome With Mutated SF3B1 Mohammad A. Vasef, MD and Carla S. Wilson, MD, PhD 192 Myelodysplastic Syndrome, NOS Kristin Hunt Karner, MD 200 Myelodysplastic Syndrome With del(5q) Giovanni Insuasti-Beltran, MD 204 Myelodysplastic Syndrome With TP53 Multihit Mutations Kristin Hunt Karner, MD 206 Myelodysplastic Syndrome/Acute Myeloid Leukemia Kristin Hunt Karner, MD 208 Pediatric Myelodysplastic Syndrome and Refractory Cytopenia of Childhood Devon Chabot-Richards, MD and Carla S. Wilson, MD, PhD MYELODYSPLASTIC/MYELOPROLIFERATIVE NEOPLASMS 214 Overview of Myelodysplastic/Myeloproliferative Neoplasms Devon Chabot-Richards, MD 216 Chronic Myelomonocytic Leukemia Kristin Hunt Karner, MD 220 Atypical Chronic Myeloid Leukemia Devon Chabot-Richards, MD 224 Juvenile Myelomonocytic Leukemia Devon Chabot-Richards, MD and Qian-Yun Zhang, MD, PhD 228 Myelodysplastic/Myeloproliferative Neoplasm, NOS Devon Chabot-Richards, MD and Kathryn Foucar, MD ACUTE MYELOID LEUKEMIA 232 Overview of Acute Myeloid Leukemia Mohammad A. Vasef, MD and Kaaren K. Reichard, MD 240 Acute Myeloid Leukemia With t(8;21)/RUNX1::RUNX1T1 Devon Chabot-Richards, MD, Kaaren K. Reichard, MD, and David R. Czuchlewski, MD 244 Acute Myeloid Leukemia With inv(16) or t(16;16)/CBFB::MYH11 Devon Chabot-Richards, MD, Kaaren K. Reichard, MD, and David R. Czuchlewski, MD 248 Acute Promyelocytic Leukemia With t(15;17)/PML::RARA Mohammad A. Vasef, MD 254 Acute Myeloid Leukemia With t(9;11)/MLLT3::KMT2A Devon Chabot-Richards, MD, Kaaren K. Reichard, MD, and David R. Czuchlewski, MD 258 Acute Myeloid Leukemia With t(6:9)/DEK::NUP214 Eric Y. Loo, MD and David R. Czuchlewski, MD 260 Acute Myeloid Leukemia With inv(3) or t(3;3)/GATA2; MECOM Eric Y. Loo, MD and David R. Czuchlewski, MD 262 Acute Myeloid Leukemia (Megakaryoblastic) With t(1;22)/RBM15::MRTFA Eric Y. Loo, MD and David R. Czuchlewski, MD 264 Acute Myeloid Leukemia With Myelodysplasia-Related Gene Mutations or Cytogenetic Abnormalities Kathryn Foucar, MD and Mohammad A. Vasef, MD 270 Acute Myeloid Leukemia, NOS Kristin Hunt Karner, MD 276 Myeloid Sarcoma Devon Chabot-Richards, MD 280 Myeloid Proliferations Associated With Down Syndrome Eric Y. Loo, MD and David R. Czuchlewski, MD BLASTIC PLASMACYTOID DENDRITIC CELL NEOPLASM 284 Blastic Plasmacytoid Dendritic Cell Neoplasm Mohammad A. Vasef, MD SECTION 6: MOLECULAR PATHOLOGY OF LYMPHOID NEOPLASMS 292 Overview of Lymphoid Neoplasms Nadine Aguilera, MD 296 Molecular Work-Up of Lymphoid Neoplasms Mohammad A. Vasef, MD and David R. Czuchlewski, MD DETERMINATION OF CLONALITY IN B-, T-, AND NK-CELL NEOPLASMS 298 Lymphoma-Associated Chromosomal Translocations Kristin Hunt Karner, MD PRECURSOR LYMPHOID NEOPLASMS 302 B-Lymphoblastic Leukemia/Lymphoma With Recurrent Genetic Abnormalities Kristin Hunt Karner, MD 306 B-Lymphoblastic Leukemia/Lymphoma, BCR::ABL1-Like (Ph-Like ALL) Joanna L. Conant, MD and Mohammad A. Vasef, MD 310 T-Lymphoblastic Leukemia/Lymphoma Mohammad A. Vasef, MD MATURE B-CELL NEOPLASMS 316 Small Lymphocytic Lymphoma/Chronic Lymphocytic Leukemia Devon Chabot-Richards, MD 322 B-Cell Prolymphocytic Leukemia Aaron Auerbach, MD, MPH 326 Splenic Marginal Zone Lymphoma Aaron Auerbach, MD, MPH and Roberto N. Miranda, MD 332 Hairy Cell Leukemia Mohammad A. Vasef, MD 336 Splenic B-Cell Lymphoma/Leukemia, Unclassifiable Nadine Aguilera, MD 344 Lymphoplasmacytic Lymphoma Aaron Auerbach, MD, MPH 350 Monoclonal Gammopathy of Undetermined Significance Nadine Aguilera, MD 354 Multiple Myeloma (Plasma Cell Myeloma) Nadine Aguilera, MD and Carla S. Wilson, MD, PhD 362 Follicular Lymphoma Aaron Auerbach, MD, MPH, C. Cameron Yin, MD, PhD, and John Schmieg, MD, PhD 370 Mantle Cell Lymphoma Aaron Auerbach, MD, MPH 376 Diffuse Large B-Cell Lymphoma Nadine Aguilera, MD 386 Burkitt Lymphoma Aaron Auerbach, MD, MPH and Carlos Bueso-Ramos, MD, PhD MATURE T- AND NK-CELL NEOPLASMS 394 T-Cell Prolymphocytic Leukemia Nadine Aguilera, MD 398 Chronic Lymphoproliferative Disorder of NK Cells Nadine Aguilera, MD 402 Aggressive NK-Cell Leukemia Nadine Aguilera, MD 406 Extranodal NK-/T-Cell Lymphoma Aaron Auerbach, MD, MPH and L. Jeffrey Medeiros, MD 410 Adult T-Cell Leukemia/Lymphoma Nadine Aguilera, MD 416 Intestinal T-Cell Lymphoma Aaron Auerbach, MD, MPH and Sa A. Wang, MD 424 Hepatosplenic T-Cell Lymphoma Aaron Auerbach, MD, MPH and Roberto N. Miranda, MD 428 Subcutaneous Panniculitis-Like T-Cell Lymphoma Aaron Auerbach, MD, MPH 432 Mycosis Fungoides/Sézary Syndrome Aaron Auerbach, MD, MPH and Sa A. Wang, MD 438 Primary Cutaneous CD30-Positive T-Cell Lymphoproliferative Disorders Aaron Auerbach, MD, MPH 442 Peripheral T-Cell Lymphoma, NOS Aaron Auerbach, MD, MPH 448 Nodal Follicular Helper T-Cell Lymphoma Aaron Auerbach, MD, MPH 456 Anaplastic Large Cell Lymphoma, ALK-Negative Mohammad A. Vasef, MD 462 Anaplastic Large Cell Lymphoma, ALK-Positive Mohammad A. Vasef, MD SECTION 7: MOLECULAR PATHOLOGY OF DENDRITIC CELL AND HISTIOCYTIC NEOPLASMS 470 Histiocytic Sarcoma Nadine Aguilera, MD 478 Langerhans Cell Histiocytosis Guanghua Wang, MD 484 Interdigitating Dendritic Cell Sarcoma Aaron Auerbach, MD, MPH and Sa A. Wang, MD 488 Follicular Dendritic Cell Sarcoma Aaron Auerbach, MD, MPH SECTION 8: MOLECULAR PATHOLOGY OF SOLID TUMORS HEAD AND NECK, INCLUDING SINONASAL TUMORS AND SALIVARY GLANDS 494 HPV-Associated Head and Neck Carcinomas Thèrése Bocklage, MD 500 Head and Neck Mucosal Squamous Cell Carcinoma, Non-HPV Related Thèrése Bocklage, MD 506 Important Alterations in Non-HPV-Related Head and Neck Squamous Cell Carcinoma Thèrése Bocklage, MD 508 Translocation-Specific Salivary Gland Tumors Thèrése Bocklage, MD 518 Translocation-Specific Salivary Gland Tumors (Continued) Thèrése Bocklage, MD 526 Nasopharyngeal EBV-Related Squamous Cell Carcinoma Thèrése Bocklage, MD 530 Other Head and Neck Tumors Thèrése Bocklage, MD THYROID 538 Papillary Thyroid Carcinoma Giovanni Insuasti-Beltran, MD 542 Follicular Thyroid Carcinoma Giovanni Insuasti-Beltran, MD 546 Poorly Differentiated Thyroid Carcinoma Von Samedi, MD, PhD 550 Anaplastic Thyroid Carcinoma Von Samedi, MD, PhD 554 Medullary Thyroid Carcinoma Giovanni Insuasti-Beltran, MD LUNG 560 Adenocarcinoma, Lung Jeremy C. Wallentine, MD 568 Squamous Cell Carcinoma, Lung Jeremy C. Wallentine, MD 572 Small Cell Neuroendocrine Carcinoma M. Carmen Frias Kletecka, MD and Samuel Reynolds, MD 576 Mesothelioma Mohammad A. Vasef, MD and Khalil Sheibani, MD, FCAP LUMINAL GASTROINTESTINAL TRACT 580 Gastric Adenocarcinoma M. Carmen Frias Kletecka, MD and Joshua Anspach Hanson, MD 584 Colorectal Adenocarcinoma and Precancerous Lesions M. Carmen Frias Kletecka, MD and Joshua Anspach Hanson, MD 590 Gastrointestinal Stromal Tumor M. Carmen Frias Kletecka, MD and Joshua Anspach Hanson, MD LIVER 598 HNF1A-Inactivated Hepatocellular Adenoma Giovanni Insuasti-Beltran, MD and Joshua Anspach Hanson, MD 602 ?-Catenin-Activated Hepatocellular Adenoma Giovanni Insuasti-Beltran, MD and Joshua Anspach Hanson, MD 606 Inflammatory Hepatocellular Adenoma Giovanni Insuasti-Beltran, MD and Joshua Anspach Hanson, MD 610 Hepatocellular Carcinoma Rama Gullapalli, MD, PhD PANCREATICOBILIARY 614 Pancreatic Ductal Adenocarcinoma Von Samedi, MD, PhD 618 Pancreatic Mucinous Cystic Neoplasm Von Samedi, MD, PhD 620 Pancreatic Intraductal Papillary Mucinous Neoplasm Von Samedi, MD, PhD 622 Cholangiocarcinoma Rama Gullapalli, MD, PhD TESTIS 624 Testicular Germ Cell Tumors Edgar Fischer, MD, PhD KIDNEY 630 Clear Cell Renal Cell Carcinoma Edgar Fischer, MD, PhD 634 Chromophobe Renal Cell Carcinoma Edgar Fischer, MD, PhD 636 Papillary Renal Cell Carcinoma Edgar Fischer, MD, PhD 638 TFE3-Rearranged and TFEB-Altered Renal Cell Carcinomas Edgar Fischer, MD, PhD 642 Wilms Tumor Marc Barry, MD 648 Clear Cell Sarcoma of Kidney Marc Barry, MD 650 Oncocytoma Marc Barry, MD 652 Rhabdoid Tumor of Kidney Marc Barry, MD BLADDER 654 Urothelial Carcinoma Marc Barry, MD ADRENAL 660 Pheochromocytoma/Paraganglioma M. Carmen Frias Kletecka, MD and Samuel Reynolds, MD 666 Adrenal Cortical Carcinoma M. Carmen Frias Kletecka, MD and Samuel Reynolds, MD 672 Neuroblastoma Giovanni Insuasti-Beltran, MD, Thèrése Bocklage, MD, and Samuel Reynolds, MD 680 Adrenal Cortical Adenoma M. Carmen Frias Kletecka, MD and Samuel Reynolds, MD PROSTATE 684 Prostatic Adenocarcinoma, Acinar Type and High-Grade Prostatic Intraepithelial Neoplasia Edgar Fischer, MD, PhD BREAST 688 ADH and DCIS (Dysplastic, Premalignant) David G. Hicks, MD 694 Ductal Carcinomas David G. Hicks, MD 700 Lobular Carcinoma David G. Hicks, MD 706 Invasive Ductal Carcinoma of No Special Type With Medullary Features Rania Bakkar, MD 710 Metaplastic Breast Carcinoma Rania Bakkar, MD 716 Phyllodes Tumors Rania Bakkar, MD 720 Basal-Like and Triple-Negative Breast Carcinomas David G. Hicks, MD and Susan C. Lester, MD, PhD CERVIX/VULVA/VAGINA 730 Preneoplastic Conditions, Cervix/Vulva/Vagina Nancy Joste, MD 734 Squamous Cell Carcinoma, Cervix/Vulva/Vagina Nancy Joste, MD 738 Adenocarcinoma, Cervix/Vulva/Vagina Nancy Joste, MD UTERUS 742 Endometrial Intraepithelial Neoplasia Lesley Lomo, MD 744 Uterine Endometrioid Carcinoma Lesley Lomo, MD 748 Uterine Serous Carcinoma Lesley Lomo, MD 750 Clear Cell Carcinoma, Uterus Lesley Lomo, MD 754 Uterine Sarcomas Thèrése Bocklage, MD OVARIES/FALLOPIAN TUBE 762 Serous Tumors of Ovary and Fallopian Tube Rania Bakkar, MD 768 Other Surface Epithelial Tumors of Ovary Rania Bakkar, MD 774 Sex Cord-Stromal Tumors of Ovary Rania Bakkar, MD 780 Germ Cell Tumor, Ovary Rania Bakkar, MD and Aaron Auerbach, MD, MPH SKIN 784 Premalignant Conditions, Skin David Cassarino, MD, PhD 788 Melanoma David Cassarino, MD, PhD 792 Squamous Cell Carcinoma, Skin David Cassarino, MD, PhD 796 Basal Cell Carcinoma, Skin David Cassarino, MD, PhD 800 Sebaceous Tumors David Cassarino, MD, PhD 802 Dermatofibroma and Dermatofibrosarcoma Protuberans David Cassarino, MD, PhD BONE 804 Overview of Molecular Pathology of Bone and Soft Tissue Tumors Thèrése Bocklage, MD 810 Osteosarcomas Thèrése Bocklage, MD 818 Ewing Sarcoma Thèrése Bocklage, MD 824 Other Small Round Blue Cell Sarcomas Thèrése Bocklage, MD 832 Giant Cell Tumor of Bone Thèrése Bocklage, MD 838 Intermediate and Malignant Cartilaginous Tumors of Bone Thèrése Bocklage, MD SOFT TISSUE 848 Intermediate and Malignant Myofibroblastic/Fibroblastic Tumors Thèrése Bocklage, MD 858 Liposarcomas Thèrése Bocklage, MD 868 Muscle Sarcomas Thèrése Bocklage, MD 878 Intermediate and Malignant Vascular Tumors Thèrése Bocklage, MD 886 Malignant Peripheral Nerve Sheath Tumor Thèrése Bocklage, MD and David R. Lucas, MD 892 Representative Genetic Findings in Bone and Soft Tissue Tumors Thèrése Bocklage, MD SARCOMAS OF UNCERTAIN DIFFERENTIATION 898 Rare Sarcomas of Uncertain Differentiation With Specific Molecular Alterations Thèrése Bocklage, MD 908 Rare Sarcomas of Uncertain Differentiation With Specific Molecular Alterations (Continued) Thèrése Bocklage, MD CNS 916 Glioblastoma, IDH Wildtype Elisabeth Rushing, MD and Mohammad A. Vasef, MD 920 Astrocytoma, IDH-Mutant Elisabeth Rushing, MD and Mohammad A. Vasef, MD 924 Pilocytic Astrocytoma Aaron Auerbach, MD, MPH, Karen SantaCruz, MD, and Elisabeth Rushing, MD 928 Oligodendroglioma, IDH Mutant, and 1p/19q Codeleted Aaron Auerbach, MD, MPH, Karen SantaCruz, MD, and Elisabeth Rushing, MD 932 Ependymal Tumors Mohammad A. Vasef, MD and Fausto J. Rodríguez, MD 938 Medulloblastoma Mohammad A. Vasef, MD and Fausto J. Rodríguez, MD 944 Choroid Plexus Tumors Mohammad A. Vasef, MD and Fausto J. Rodríguez, MD 948 Meningioma Mohammad A. Vasef, MD and Karen SantaCruz, MD 952 Retinoblastoma Mohammad A. Vasef, MD and Fausto J. Rodríguez, MD SECTION 9: QUALITY ASSURANCE AND REGULATORY ISSUES 958 Federal Agencies and Regulation of Laboratories Amer Mahmoud, MD, FCAP 960 FDA Regulations Yongjie Zhou, MD, PhD 966 Proficiency Testing and Accreditation Amer Mahmoud, MD, FCAP