Single-Cell Sequencing and Methylation

Summary

Chapter 1: Understanding Diseases from Single-Cell Sequencing and Methylation

This introductory chapter defines clinical single-cell biomedicine as an emerging discipline that integrates single-cell RNA and DNA sequencing, proteomics, and functional data with clinical phenomes, therapeutic responses, and prognosis. It highlights the book's aim to review the roles of single-cell sequencing and methylation in various diseases, exploring disease-specific alterations. The chapter emphasizes potential applications of these methodologies, particularly in pulmonary diseases, and discusses the importance of understanding signaling pathways and standardizing single-cell preparation.

Chapter 2: Methods for Single-Cell Isolation and Preparation

This chapter underscores the critical importance of accurate and reliable cell capture for single-cell sequencing. It provides an overview of current single-cell isolation methods, addressing key parameters such as sample compatibility, cell viability, purity, throughput, and isolation efficiency. The authors discuss various technologies and the considerations needed to avoid incorrect or biased data, ensuring successful experimental design for downstream analysis.

Chapter 3: Single-Cell Sequencing of T cell Receptors: A Perspective on the Technological Development and Translational Application

This chapter systematically describes the value of single-cell sequencing in investigating T cell receptors (TCRs) and their transcriptional profiles. It discusses the technological advancements in TCR sequencing and their translational and clinical applications, particularly for immune-therapy in cancer and autoimmune diseases. The authors prospect the importance of these developments for understanding the immunological response at a high resolution.

Chapter 4: DNA Methylation in Pulmonary Fibrosis

This chapter demonstrates how global methylation patterns and specific gene methylation (e.g., Thy-1, COX-2, p14ARF, and PTGER2) are associated with the development of pulmonary fibrosis. It discusses how these methylation patterns can serve as disease-specific biomarkers to predict the occurrence and progression of the disease. The potential of DNA methylation inhibitors as an emerging anti-fibrosis therapy is also explored.

Chapter 5: Methylation of Inflammatory Cells in Lung Diseases

This chapter uses bioinformatics to address how altered DNA methylation in inflammatory cells, such as lymphocytes and macrophages, can downregulate the gene expression of inflammatory mediators, thereby initiating lung diseases. It highlights the need to identify and validate the specificity and regulatory mechanisms of inflammatory cell epigenetics. The chapter also points out the epigenetic heterogeneity among T cell subsets and different gene regions in lung diseases and cancers.

Chapter 6: Research Advances on DNA Methylation in Idiopathic Pulmonary Fibrosis

This chapter reviews the progress in understanding DNA methylation in idiopathic pulmonary fibrosis (IPF), a chronic lung disease with a poor prognosis. It describes how DNA methylation affects gene expression, facilitates fibroblastic foci formation, and contributes to lung fibrosis. The potential of using DNA methylation as a biomarker for prognosis and as a therapeutic target for IPF is also discussed.

Chapter 7: DNA Methylation in Chronic Obstructive Pulmonary Disease

This chapter provides an overview of the role of DNA methylation in the etiology, pathogenesis, pathophysiological changes, and complications of Chronic Obstructive Pulmonary Disease (COPD). It discusses how aberrant methylation of target genes, influenced by environmental factors like smoking, contributes to COPD development. The clarification of these mechanisms aims to provide new disease-specific biomarkers and targets for early diagnosis and therapy.

Chapter 8: The Role of RASSF1 Methylation in Lung Carcinoma

This chapter focuses on the DNA methylation of the RASSF1 (RAS-Association Domain Family 1) tumor suppressor gene and its clinical significance in lung carcinoma. It summarizes how the inactivation of RASSF1A through promoter hypermethylation is a frequent event in lung cancer, contributing to carcinogenesis mainly via the Hippo signaling pathway. The potential of RASSF1A methylation as a biomarker for early diagnosis, prognosis, and new therapeutic strategies is also explored.

Chapter 9: Single Cell Sequencing: A New Dimension in Cancer Diagnosis and Treatment

This chapter reviews the recent developments in single-cell sequencing technologies and their applications in cancer research, emphasizing the complexity and heterogeneity of cancer. It discusses how insights from single-cell sequencing can be used to understand tumor microenvironments, primary and acquired drug resistance, and develop novel diagnostic and therapeutic approaches. The chapter also addresses the challenges in clinical application, such as the need for workflow optimization, standardized protocols, and comprehensive databases.

Chapter 10: The Role of Methylation in the CpG Island of the ARHI Promoter Region in Cancers

This chapter overviews the importance of Aplasia Ras homologue member I (ARHI, DIRAS3) methylation and its expression phenomes in various cancers like ovarian, breast, and liver cancer. It explains that ARHI is a maternally imprinted tumor suppressor and aberrant DNA methylation of its paternal allele is a primary inhibitor of its expression. The varying roles of methylation in the ARHI promoter's CpG islands across different cancer types offer new insights into tumorigenesis and progression.

Chapter 11: Clinical Significance of P16 Gene Methylation in Lung Cancer

This chapter highlights the role of p16 (CDKN2A) gene promoter methylation, a common epigenetic change, in the progression of lung cancer, particularly non-small cell lung cancer (NSCLC). It discusses how p16 methylation is associated with poor prognosis and therapeutic resistance and reviews its frequency and mechanisms in lung cancer, often linked to smoking and air pollution. The potential of p16 methylation as a biomarker for early detection, diagnosis, and prognosis of lung cancer is evaluated.

Chapter 12: Application of Single-Cell RNA Sequencing in Pancreatic Cancer and the Endocrine Pancreas

This chapter reviews the application of single-cell RNA sequencing (scRNA-seq) in analyzing cell heterogeneity in the pancreas and pancreatic cancer. It discusses how scRNA-seq aids in cell-type-specific molecule identification, understanding interactions between cancer cells and the stromal microenvironment, and characterizing cancer stem cells and circulating tumor cells. The chapter highlights discoveries related to pancreatic islet cells, type 2 diabetes, and pancreatic ductal adenocarcinoma (PDAC) using these technologies.

Chapter 13: Single-Cell Sequencing in Genitourinary Malignancies

This chapter highlights the use of single-cell sequencing (SCS) in understanding genitourinary (GU) malignancies such as renal cell carcinoma (RCC), bladder cancer, and prostate cancer. It illustrates how SCS helps characterize tumor heterogeneity, the tumor microenvironment, cancer stem cells (CSCs), circulating tumor cells (CTCs), and clonal evolution. The translational applications of SCS in GU malignancies for diagnostic, prognostic, and treatment-related approaches are discussed, emphasizing its role in precision-based care.

Chapter 14: PI3K Isoform-Selective Inhibitors in Cancer

This chapter provides an overview of the PI3K signaling pathway, its role in cellular functions like growth and apoptosis, and its frequent dysregulation in cancer through mutations in PI3K isoforms. It discusses the development and application of PI3K isoform-selective inhibitors as targeted cancer therapies, both as monotherapy and in combination with other pathway inhibitors. The chapter covers the impact of these inhibitors on various cancers, including breast, prostate, leukemia, and head and neck cancer.

Chapter 15: Single Cell Sequencing in Cancer Diagnostics

This chapter discusses the current possibilities and practical advice for implementing single-cell sequencing (SCS) in clinical cancer diagnostics to delineate tumor clonality and heterogeneity. It contrasts SCS with bulk sequencing, highlighting SCS's ability to overcome the limitations of averaged signals and identify distinct cell populations crucial for disease progression and therapeutic resistance. The chapter reviews various SCS technologies (plate-based and droplet-based for RNA and DNA) and their relevance for precision diagnostics, mutagenesis understanding, and addressing therapeutic resistance.

Chapter 16: Single Cell RNA Sequencing in Human Disease: Renal, Pancreatic, and Viral Diseases

This chapter explores the utility of single-cell RNA sequencing (scRNA-seq) in understanding cellular compositions, discovering heterogeneity in cellular expression patterns, and uncovering clues for developing targeted therapies in renal, pancreatic, and viral diseases. It highlights how scRNA-seq advances the study of these conditions by providing high-resolution transcriptomic data, which is crucial for understanding complex molecular signaling patterns. The chapter emphasizes the growing availability of whole tissue cellular atlases in health and disease as a valuable resource derived from such studies.

Chapter 17: Single-Cell Sequencing in Human Genital Infections

This chapter reviews the applications of single-cell sequencing in studying human genital infections, including sexually transmitted infections (STIs), urinary tract infections (UTIs), and vaginal infections. It discusses how this technology helps investigate drug-resistant clones, cell-to-cell variations, acquired drug resistance mutations, and the transcriptional diversity of pathogens across different infection stages. The limitations and challenges, such as difficulties in microbial culturing and data analysis, are also addressed.

Chapter 18: Emerging Strategies for Therapeutic Antibody Discovery from Human B Cells

This chapter reviews various approaches for discovering therapeutic monoclonal antibodies from human B cells, highlighting challenges such as the rarity of B cells encoding therapeutic antibodies and their short ex vivo lifespan. It discusses traditional methods like synthetic libraries and hybridoma technology, as well as next-generation microfluidic technologies that improve throughput and maintain native antibody chain pairing. The chapter emphasizes methods for high-throughput discovery from the human B-cell repertoire, including paired Ig sequencing and native library screening.

Chapter 19: Values of Single-Cell RNA Sequencing in Development of Cerebral Cortex

This chapter highlights single-cell RNA sequencing (scRNA-seq) as a powerful tool for exploring the complexity, cell clusters, and specific functions of brain cells, particularly in the context of cerebral cortex development and neurological diseases. It discusses how scRNA-seq helps define the heterogeneity and transcriptomic changes of single neurons during dynamic development, differentiation, and in pathogenesis. The chapter also addresses challenges, such as sample interference, and looks forward to integrating transcriptomic profiles with molecular and functional phenomes for a deeper understanding of brain development and diseases.