Omics Technologies for Clinical Diagnosis and Gene Therapy: Medical Applications in Human Genetics

Rare inherited disorders have become a major public health concern in recent years. Owing to a lack of resources, poorly planned primary and basic health care, and inadequate political structures, treatment, and management policies are daunting challenges in many countries. As a result, these diseases need particular attention, especially in less developed areas, where these disorders remain unnoticed. Similarly, the effect of such severe disorders on underprivileged populations is expected to be devastating. Identifying certain genetic markers can provide a valuable explanation for disease etiology, molecular characterization, and pathogenesis. In this chapter, we highlight the importance of next-generation sequencing to explore and recognize the role of novel causative genes in developing successful treatments for the most prevalent rare genetic disorders. DNA methylation and transcriptome markers have been shown to aid in the prediction of common diseases; however, this has not been tested on rare genetic disorders. Since the rate of rare inherited disorders is higher in developing countries, we believe that these populations can provide us with much stronger clues for the genetic and environmental association. These markers, along with other parameters, can be used to systematically build machine learning models to improve risk prediction; this approach has the potential to reshape how we predict disease risk and save many lives around the world.

Rare genetic disorders affect a significant proportion of the global population. A large number of these patients are either misdiagnosed or remain undiagnosed which can have potentially adverse effects, including failure to provide anticipatory prognosis and identify potential treatment. With the completion of HGP, genetic testing has fast grown into a diagnostic discipline introducing new and costeffective diagnostic tests with reasonable accuracy and specificity. NGS technologies, in particular, changed the field of genetic diagnosis by sequencing the entire genome or subset thereof in a single test and accomplishing diagnosis of virtually all diseases, either congenital or late-onset. These technologies have opened up new opportunities and unique challenges. This chapter discusses the importance of genetic testing, its scope, various technologies and approaches and, finally, the opportunities and challenges accompanying the new age genetic tests.

Pre-implantation genetic diagnosis (PGD) is a practical alternate evolving approach to prenatal diagnosis and termination of pregnancies in families with a high risk of Mendelian monogenetic and polygenetic disorders. Pre-implantation genetic diagnosis testing is continuing to extend immensely, along with a novel genetic analysis and in vitro fertilization approaches are in practice in the medical field throughout the world. However, PGD is regarded as ethically sensitive because repetitive termination of pregnancy causes huge psychological effects on the couples, and also because the low rate of pregnancy and birth makes it unreliable compared to prenatal testing. But it is also helpful in achieving additional goals e.g., improved embryo and gender selection, overcoming the chances of birth of a child with an unknown genetic defect, better understanding of epigenomic regulations and reduction in the monetary burden of society. This chapter focuses on PGD, its procedure, utility and advantages, goals and objectives and the various issues surrounding it. We also discuss the future of this technology at the end of the chapter

In this chapter, we have focused on the journey of sorting genes and the connotation of genetic counseling started. In a literal sense, we will understand how genetic counseling could contribute to identifying pathogenicity and penetrance of genetic mutation/s in high-risk individual/s or populations. Great strides have been achieved in terms of diagnosis, management, and treatment of various genetic disorders due to rapid advancements in genetic research. The national Thalassemia Prevention Program of Cyprus has been one of the earliest and most celebrated successes in lowering the disease burden and improving life quality and survival rate in patients. The knowledge regarding gene/s and variant/s is quite instrumental for making important reproductive decisions and therapeutic interventions for both rare and common disorders. We also touch upon the associated ethical issues and challenges. 

 Genome-wide association studies (GWAS) are designed to find associations between genomic variants and a phenotype, usually a complex multifactorial disease. The idea for association studies in a large cohort was floated after linkage analysis, which proved extremely successful in the identification of causative genes for rare disorders, but it did not come up to expectations in the case of common complex disorders where causative alleles are less frequently aggregated in families. Ever since their advent in 2005, GWAS have transformed gene identification ventures in complex disease genetics over the past fifteen years, giving rise to several powerful associations for complex traits and disorders. Association studies are based on the “common disease common variant” hypothesis which assumes that genomic variation with low penetrance and high population frequency are involved in the causation of common complex disorders. Although GWAS, complemented with the downstream functional assessment of the variants, have been successful in identifying novel disease-causing genes and biological mechanisms, the field has also received intense criticism over the years, especially its failure in tracing the so-called ‘missing heritability’. Therefore, further functional studies are mandatory to precisely establish a link between risk alleles and a phenotype. This chapter broadly covers an introduction of GWAS, their successes and limitations, and various important factors affecting the design and results, followed by challenges in the post-GWAS era.

Regenerative medicine (RM) is defined as a replacement and revival of human cells, tissues, or organs to reinstate or reconstruct their normal physiology. RM is regarded as a solution to provide healthy substitutes for a malfunctioning/failed organ or a tissue. It is emerging as the suitable substitute for organ transplantation. Transplantation seems impractical due to the limited availability of donors as significant disparities lie between the number of patients that require transplantation and the availability of organs from the donor, so there was a gap created. Therefore, to comply with these needs, RM has emerged as a new science to create biological replacements and exploit the body's ability of regeneration to recover and sustain normal function in diseased and damaged tissues. This chapter overviews RM in terms of adopted strategies, its clinical applications in organ engineering along with inherited challenges and their plausible solutions.

Multiomics also described as integrative omics is an analytical approach that combines data from multiple ‘omics’ approaches including genomics, transcriptomics, proteomics, metabolomics, epigenomics, metagenomics and Meta transcriptomics to answer the complex biological processes involved in rare genetic disorders. This omics approach is particularly helpful since it identifies biomarkers of disease progression and treatment progress by collective characterization and quantification of pools of biological molecules within and among the various types of cells to better understand and categorize the Mendelian and non- Mendelian forms of rare diseases. As compared to studies of a single omics type, multi-omics offers the opportunity to understand the flow of information that underlies the disease. A range of omics software and databases, for example WikiPathways, MixOmics, MONGKIE, GalaxyP, GalaxyM, CrossPlatform Commander, and iCluster are used for multi-omics data exploration and integration in rare disease analysis. Recent advances in the field of genetics and translational research have opened new treatment avenues for patients. The innovation in the next generation sequencing and RNA sequencing has improved the ability from diagnostics to detection of molecular alterations like gene mutations in specific disease types. In this chapter, we provide an overview of such omics technologies and focus on methods for their integration across multiple omics layers. The scrupulous understanding of rare genetic disorders and their treatment at the molecular level led to the concept of a personalized approach, which is one of the most significant advancements in modern research which enable researchers to better comprehend the flow of knowledge which underpins genetic diseases.

The advent of bioinformatics and integrated biology approaches has given rise to new avenues of diagnostic and therapeutic regimes. Living systems have been explored to identify disease-associated biomarkers that facilitate the early diagnosis of perilous medical conditions. Likewise, gene networks are pondered upon to obtain better insights into biochemical systems that can assist in the prediction and testing of the effects of various interactions within the systems. Genomics and proteomics-based approaches are being explored to facilitate the early diagnosis of cancers, shifting the paradigm towards noninvasive diagnostic alternatives. Bioinformatics has also fueled pharmacogenomics and pharmacogenetics-based strategies that have in turn contributed to the development of personalized medications. Similarly, the reverse vaccinology approach has emerged as a prominent option to combat deadly pathogens that were otherwise unrestrainable. This chapter highlights the fruits of integrated bioinformatics in diagnosing and treating detrimental conditions.

Interpretation of molecular differences and intricacy at multiple stages, for instance proteome, genome, epigenome, metabolome, and transcriptome is needed for a thorough understanding of disease and human health. Biology has been reliant on data produced at these stages, which is collectively referred to as multi-omics data, after the emergence of sequencing techniques. Among all the aspects of biology, rapid development in high-throughput data initiation has enabled to carry out research on multi-omics systems biology. Metabolomics, proteomics, and transcriptomics data can provide answers to the targeted biological queries about the expression of metabolites, proteins, and transcripts, independently. A concise summary of multi-omics data sources, challenges in datasets integration, and visualization portals is also discussed.

Recent advances are nowadays providing opportunities to examine the complexities of organs and organisms at the single-cell level. The conventional cellbased analysis mainly examines the cellular processes from the bulk of cells but singlecell omics provides a more detailed insight into individual cell phenotypes, thus giving a link between the phenotype and genotype of cells. Single-cell analysis can be performed at genome, epigenome, transcriptome, proteome and metabolome levels and thus makes it possible to come across mechanisms not seen during the sequencing of bulk tissues. Researchers need to isolate single cells before the initiation of single-cell analysis. For this, various strategies like FACS, MACS, LCM, micro-manipulation and micro-fluids are used for cell isolation depending upon their physical properties and cellular biological characteristics. The analysis of single-cell data at multiple levels gives us an unusual view of multilevel transformation at the single-cell level and thus providing a better chance to discover novel biological processes. High throughput analysis of single cells at genome, transcriptome and proteome levels provides unique and important insights into cell variability and diverse processes like development, genetic expressions and severity of different symptoms in disease pathogenesis.

The ongoing development in new genotyping methods necessitates an understanding of their potential benefits and limits in terms of pharmacogenomics utility. We give an overview of technologies that can be used in pharmacogenomics research and clinical practice in this chapter. The Human Genome Project’s completion has paved the way for the development of clinical instruments for patient evaluation. Pharmacogenomics may enable the identification of patients who are most likely to benefit from a specific drug, as well as those for whom the expense and risk are greater than the advantages. Both drug therapy’s safety and efficacy may improve. In the future, genotyping may be used to tailor drug treatment for large groups of individuals, lowering drug treatment costs and improving therapeutic efficacy and overall health.

Bone healing and formation are under the control of growth factors. Among these factors, bone morphogenetic proteins (BMPs) have a vital role in bone and cartilage maintenance and formation. BMP itself belongs to the superfamily of transforming growth factor β (TGFβ). Although, the use of recombinant BMPs has no significant association with the treatment of bone fractures, arthroplasty, pseudoarthrosis or other bone-related diseases. Recent advancements in genetic engineering have led to the foundation of gene therapy. Gene therapy uses genes to be incorporated in the living cells to replace defective genes or manipulate gene expression for therapeutic purposes. Gene therapy is emerging for the treatment of diseases with approval in Europe where it is in the marketing surveillance phase (Phase IV Clinical trial). This technique has also been tested for the incorporation of osteogenic genes in stem cells for repairing spinal fusion and recovering defects in bones in preclinical models. Therefore, gene therapy has the potential for the treatment of different diseases and has the advantage over the use of recombinant proteins. In this chapter, we have discussed gene therapy, its mechanism, delivery system and its use in tissue engineering (soft and bone tissue) for clinical application.

 A limiting factor for the identification of disease mechanisms and development of new therapies has been the access to a model system/s that can faithfully recapitulate key features of the disease and more precise clinical translations of new treatments. Stem cells in this regard are very promising, but the ethical issues related to totipotent embryonic stem cells and functional constraints to unipotent somatic stem cells have led to focus on induced pluripotent stem cells to avoid both functional and ethical constraints. The introduction of human Induced Pluripotent Stem Cell (iPSC) technology provides a model system to replicate diseases in humans. In this technology, human somatic cells can be “reprogrammed” by the transgene expression of four transcription factors into stem cells called iPSC. In this chapter, it will be discussed how iPSCs can be used for disease modelling, drug discovery and regenerative medicine.

 Hemoglobinopathies are a group of inherited blood disorders characterized by compromised hemoglobin function. Hemoglobin is a 64kDa protein, consisting of four globin polypeptides each containing one heme molecule; blood acquires its red color from this heme molecule. Two of the four polypeptide chains are α-globin chains, whereas the other two are β and γ chains during adult and fetal life, respectively. Hemoglobin carries oxygen to respirating cells and tissues in vertebrates and defects in genes encoding this protein result in a variety of disorders, ranging from mild asymptomatic to severe fatal phenotypes. This chapter reviews various hemoglobinopathies underlying mutations in globin genes. We also provide a brief note of the traditional and contemporary diagnostic approaches and screening, both prenatal and postnatal, with a specific focus on recent advances in this regard. We have summarized various therapeutic strategies, from transfusion and iron chelation to CRISPR-driven genome editing aimed at reactivating fetal hemoglobin in adults. The chapter concludes with a brief account of the future challenges and prospects for developing a therapy for hemoglobinopathies a clinical reality

Metabolic Syndromes (MetS) are recognized as a cluster of risk factors which are known to increase the likelihood of obesity, type 2 diabetes (T2D) and cardiovascular disorders (CVDs). It is significant to understand disease pathology in order to discover a pathological mechanism leading to the development of MetS. Elevated triglycerides, increased blood pressure, hyperglycemia (increased blood glucose levels), low levels of High-density lipoprotein (HDL) cholesterol and elevated waist circumference are key parameters in diagnosing MetS. Various therapeutic interventions have been developed for treating metabolic diseases like polypills which are commonly known as combination pills, along with the fixed dose combinations. In addition to pharmacological handling, surgical treatment is also showing success in treating MetS such as Bariatric treatment. With the emerging experimental techniques, gene therapy allows the replacement of a defective gene with a healthy one, which may eventually reverse the disease. Leptin Gene Therapy, ZFN Gene Editing, CRISPR/ Cas9 genome editing are different platforms of gene therapy which are showing promising results in treating the metabolic disease. Novel experimental approaches and pharmacological treatments can provide a better insight into metabolic syndrome and its related complications, thereby reducing its global burden.

Intellectual disability (ID) is caused by the disruption of neurodevelopmental processes. Its diagnosis and severity are defined in terms of an Intelligence Quotient score of <70. ID has diverse presentations and clinical overlaps with other cognitive disorders such as autism spectrum disorder and microcephaly. ID has a diverse etiology encompassing both environmental and genetic insults to the developing brain. The precise diagnosis is challenging but crucial for prognosis and risk assessment for future pregnancies. The suspected cases of genetic ID often follow a strategic series of tests for diagnosis. There is no effective cure for this disorder except in the cases of early diagnosed metabolic disorders. The available therapies are mostly aimed at easing the symptoms and improving the quality of life.

Intellectual disabilities (ID) are among the most common genetic disabilities worldwide. Over the last two decades, ID has especially drawn special scientific interest being the key to understanding normal brain development, growth, and functioning. Here, we discuss two intellectual disabilities to better understand the emerging trends in disease diagnosis as well as the therapies available for their management. Primary microcephaly (MCPH) is a monogenic genetic disorder with twenty-eight loci (MCPH1-MCPH28) mapped so far with all the causative genes being elucidated as well. The role of these genes in disease prognosis along with their association with various MCPH-linked phenotypes plays an important role in the molecular diagnosis of the disease. As there is no cure/treatment yet available to enlarge a congenitally small brain, management modalities in use include physical, speech and occupational therapies as well as psychological and genetic counselling to not only reduce the incidence of the disorder but also to help families cope better. The second intellectual disability being discussed here is schizophrenia which is a multifactorial disorder owing to its complex and extremely heterogeneous etiology. Although various environmental factors play an important role, the genetic factors have been identified to play the most pivotal role in disease presentation as to date, 19 loci (SCZD1-SCZD19) have been linked to schizophrenia. However, underlying genes for only six of these loci have been mapped along with 10 other genes that are either linked to schizophrenia or show susceptibility to it. Diagnosis of schizophrenia needs careful consideration and various tests and tools currently employed for complete diagnosis have been discussed here. The management options for schizophrenia include pharmacological, non-pharmacological and intracranial therapies. These disorders shed light on the important role omics technologies have played not only in better understanding of the disease prognosis but also assisting in disease diagnosis and treatment modalities too.