Next-generation sequencing (NGS), also known as high-throughput or massively parallel sequencing, represents a technological revolution in sequencing millions of genetic fragments, whether intact or disintegrated.
In the past, using the Sanger sequencing method, it took years to sequence the entire human genome. However, NGS has drastically accelerated this process, reducing the time to just a few days and significantly lowering costs. This advancement provides access to well-curated genetic and genomic data, enhancing the precision of modern medicine.
NGS has transformed both clinical medicine and biomedical research and is utilized across various medical fields:
- Oncology: NGS can detect low levels of circulating tumor DNA in the bloodstream, aiding in the identification and treatment of cancer.
- Rare Diseases: NGS can help diagnose rare monogenic conditions.
- Infectious Diseases: NGS can identify genetic factors that contribute to infectious diseases.
- Prenatal Diagnostics: NGS is used to diagnose conditions in prenatal settings.
- Precision Medicine: NGS identifies molecular markers for disease subtypes, enabling tailored medical interventions.
- Non-invasive Diagnostics: NGS facilitates the development of non-invasive diagnostic approaches, such as liquid biopsies, to monitor disease progression.
Studying Evolutionary History
NGS allows researchers to explain evolutionary relationships and the genetic composition of infectious agents, as demonstrated by SARS-CoV-2 studies during the COVID-19 pandemic. NGS revealed that SARS-CoV-2 has a positive-sense single-stranded RNA genome. It helped assemble the COVID-19 genome, design reverse transcriptase PCRs specific to SARS-CoV-2, and understand transmission, mutations, and virulence. While technologies like RT-PCR and microarrays were also employed during COVID-19 research, NGS provided more reliable results
Forward and Reverse Phenotyping
Traditionally, genetic diseases were studied using the forward genetic method, where research starts with the phenotype and moves towards the genotype. NGS enables reverse phenotyping, where studies begin with genes and progress to phenotypic outcomes. This approach has been especially beneficial in studying rare diseases, using a genotype-first method to uncover new phenotypes.
Drug and Antibiotic Applications
Accurate drug and antibiotic regimens are critical for maintaining health during infections. While other diagnostic techniques exist, NGS provides an unbiased and faster diagnosis. For instance, NGS made it possible to identify the gene-specific Streptococcus canis in synovial fluid and tissue cultures. in synovial fluid and tissue cultures.
Diagnosing Diseases and Infections
In a 2008 case, three individuals died of an unknown disease after receiving organ transplants from a single deceased donor. NGS analysis revealed the causative agent to be an Old-World arenavirus related to lymphocytic choriomeningitis virus, transmitted during organ transplantation. Numerous similar cases of infectious diseases have been diagnosed through NGS, demonstrating its expanding applications in medicine.
Clinical Trials
NGS helps identify patients suited for clinical trials. There are two types of trials:
- Umbrella Trials: Involve patients with the same disease or cancer, defined morphologically, but with different genetic mutations.
- Bucket Trials: Include patients with varying diseases, cancers, or tumors that share the same genetic alterations.
Cancer biopsies often undergo NGS to identify genetic abnormalities, which can then guide targeted therapies.
Cell-Free Rapid Diagnostics
Certain parts of the body are difficult to access for therapy or diagnostics. Liquid biopsy (cell-free circulating DNA) allows for diagnosis in such areas. Some DNA circulates outside of cells, and NGS can analyze this DNA to identify its origin, aiding in cancer detection and non-invasive prenatal studies.
Diagnosing Mutations
After RNA and whole exome sequencing, NGS can be used for targeted therapy by profiling genetic mutations. In cases of clonal evolution caused by mutation heterogeneity, such as in leukemia, identifying mutations can be challenging. NGS addresses this challenge by identifying mutations through targeted panel sequencing.
Drug Targeting
NGS can identify genetic biomarkers, facilitating the development of targeted molecular therapies and vaccines. Clinicians use this information to improve treatment outcomes and increase patient survivability.
Prenatal Care
Many diseases manifest at birth. NGS can identify monogenic disorders, and targeted panel screening can help prevent hemolytic conditions. Early diagnosis of other diseases can lead to timely therapeutic interventions.
Discovering the Unknown
Many infectious diseases remain undiagnosed due to the lack of advanced diagnostic techniques. For instance, McArdle disease, caused by a deficiency in the PYGM-encoded enzyme myophosphorylase, went undiagnosed in North America due to miscalculated prevalence. Similarly, distal myopathies have been difficult to diagnose. NGS now offers a more comprehensive method for detecting such rare and unidentified genetic disorders..
