In the last decade, a variety of new technologies have emerged that permit dissection of the molecular and cellular components of the immune system at an unprecedented level of depth and complexity. Availability of these new tools is especially important in the diagnostic approach to human disorders of the immune system. The very large number of variants in genes that may impact on immune system development and function is responsible for remarkable heterogeneity in the ability of the immune system of single individuals to react to antigenic challenges. Natural exposure to a large number of antigens (including pathogens, but also the commensal flora) introduces additional variability, making the study of immune system physiology and pathology extraordinarily complex. The advent of new technologies that may challenge this level of complexity offers new insights to the study of human immune disorders, and of Primary Immune deficiencies (PIDs) in particular.
Next generation sequencing (NGS) is highly powerful approach to study the human genome by means of whole exome or whole genome sequencing (WES, WGS). Until recently, the discovery of PID genes required collection of multiple families in which affected individuals shared the same phenotype. Linkage analysis was used to define the region of the genome that could possibly include the disease causing gene. Positional cloning, or in some cases the candidate gene approach, were then used to identify the actual PID-causing gene. By contrast, NGS techniques permit detailed annotation of genetic variants in single individuals, and have in fact allowed identification of novel PID genes, even through the study of single patients or families. Use of bioinformatics tools that are continuously updated and empowered by experimental findings, may help predict which genes and pathways are most likely to be affected in patients with inborn errors of immunity presenting with a defined phenotype. The connectome is one such tool that has already shown its promise. NGS can also be used to obtain very detailed information on the composition and diversity of T and B cell antigen receptor repertoire. Using this approach, specific abnormalities of antigen receptor repertoire composition and diversity can be identified in patients with PIDs. Moreover, it becomes possible to investigate how exposure to specific challenges (vaccines) or use of therapeutic approaches (stem cell transplantation, immune suppression, etc.) may shape the immune repertoire in patients with PIDs. Coupled with single cell cloning, the study of B cell receptor specificities permits to define mechanisms that determine autoimmunity in several forms of PID. At a more global level, the study of phenotypic heterogeneity of hematopoietic and immune cell populations has been improved with the advent of standardized panels of autoantibodies using multicolor flow cytometry (as shown by the EUROFLOW panel to study PIDs), or by development of new techniques, such as CyTOF, that have dramatically expanded the number of markers that can be used to identify single populations. Induced pluripotent stem cells represent a novel platform for PID modeling, and are especially important in the study of extremely are conditions and of diseases that affect tissues that are hardly accessible in patients, such as the central nervous system. Finally, evolution in gene editing has already permitted rapid generation of cellular and animal models with defined gene mutations, and may provide hope for the future development of novel and safer approaches to gene therapy. In this Topic, several experts illustrate how technological advances are revolutionizing the study of human PIDs.
In the last decade, a variety of new technologies have emerged that permit dissection of the molecular and cellular components of the immune system at an unprecedented level of depth and complexity. Availability of these new tools is especially important in the diagnostic approach to human disorders of the immune system. The very large number of variants in genes that may impact on immune system development and function is responsible for remarkable heterogeneity in the ability of the immune system of single individuals to react to antigenic challenges. Natural exposure to a large number of antigens (including pathogens, but also the commensal flora) introduces additional variability, making the study of immune system physiology and pathology extraordinarily complex. The advent of new technologies that may challenge this level of complexity offers new insights to the study of human immune disorders, and of Primary Immune deficiencies (PIDs) in particular.
Next generation sequencing (NGS) is highly powerful approach to study the human genome by means of whole exome or whole genome sequencing (WES, WGS). Until recently, the discovery of PID genes required collection of multiple families in which affected individuals shared the same phenotype. Linkage analysis was used to define the region of the genome that could possibly include the disease causing gene. Positional cloning, or in some cases the candidate gene approach, were then used to identify the actual PID-causing gene. By contrast, NGS techniques permit detailed annotation of genetic variants in single individuals, and have in fact allowed identification of novel PID genes, even through the study of single patients or families. Use of bioinformatics tools that are continuously updated and empowered by experimental findings, may help predict which genes and pathways are most likely to be affected in patients with inborn errors of immunity presenting with a defined phenotype. The connectome is one such tool that has already shown its promise. NGS can also be used to obtain very detailed information on the composition and diversity of T and B cell antigen receptor repertoire. Using this approach, specific abnormalities of antigen receptor repertoire composition and diversity can be identified in patients with PIDs. Moreover, it becomes possible to investigate how exposure to specific challenges (vaccines) or use of therapeutic approaches (stem cell transplantation, immune suppression, etc.) may shape the immune repertoire in patients with PIDs. Coupled with single cell cloning, the study of B cell receptor specificities permits to define mechanisms that determine autoimmunity in several forms of PID. At a more global level, the study of phenotypic heterogeneity of hematopoietic and immune cell populations has been improved with the advent of standardized panels of autoantibodies using multicolor flow cytometry (as shown by the EUROFLOW panel to study PIDs), or by development of new techniques, such as CyTOF, that have dramatically expanded the number of markers that can be used to identify single populations. Induced pluripotent stem cells represent a novel platform for PID modeling, and are especially important in the study of extremely are conditions and of diseases that affect tissues that are hardly accessible in patients, such as the central nervous system. Finally, evolution in gene editing has already permitted rapid generation of cellular and animal models with defined gene mutations, and may provide hope for the future development of novel and safer approaches to gene therapy. In this Topic, several experts illustrate how technological advances are revolutionizing the study of human PIDs.