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In this series of posts on precision medicine, we have constantly mentioned DNA, DNA sequencing, genome sequencing, or massive sequencing (NGS), but what does this concept really mean? What is DNA sequencing Simply put, sequencing is the technique that converts a biological sample (such as a biopsy, blood, or saliva) into data that can be analyzed by a computer. We go from having a piece of tissue to obtaining a text file that contains the "instruction manual" of that person's cells, that is, the exact sequence of their DNA. This sequence, which we can think of as a plain text file, is what we will use in the methods and procedures for genomic testing for precision medicine that we discussed in our previous article. This time we will talk about the technical differences we can find to sequence that genomic material. This will enable us to analyze it and find patterns or variants that help us advance in precision medicine. DNA, or deoxyribonucleic acid, is the molecule that contains all living beings' genetic information. It consists of four nucleotides, each represented by a letter: A (adenine), T (thymine), C (cytosine), and G (guanine). These letters are organized in a double helix structure, where each nucleotide on one strand pairs with its complementary nucleotide on the other: A with T and C with G, forming the so-called base pairs. Deciphering the sequence of these letters in DNA allows us to understand how genes work, how they are regulated, and how they influence health and disease. The ability to sequence DNA has led to great advances in the knowledge of genome organization and function. Thanks to these techniques, scientists have identified genes responsible for hereditary diseases, developed targeted therapies in precision medicine, and reconstructed species evolution over time. But not all sequencing technologies are the same: for example, depending on the technique used, longer or shorter DNA fragments can be obtained, which influences the accuracy and usefulness of the analysis. Below, we will explore the different generations of sequencing techniques. First-generation sequencing: the Sanger method Historically, the most used method for sequencing DNA has been the one developed by Sanger and his team. In this procedure, the DNA molecule whose sequence is to be determined is converted into single strands, which are used as templates to synthesize a series of complementary strands. Each of these strands randomly ends in a different specific nucleotide. This produces a series of DNA fragments separated electrophoretically, and whose analysis reveals the DNA sequence. In the first step of this reaction, DNA is heated to denature, forming single strands. The single-stranded DNA is mixed with primers that hybridize to the 3' end of this DNA. The single-stranded DNA sample bound to the primer is distributed in four tubes. In the next step, DNA polymerase and the four deoxyribonucleotide triphosphates (dATP, dCTP, dGTP, and dTTP) are added to each tube. In addition, each tube also receives a small amount of a modified deoxyribonucleotide, called a dideoxynucleotide (ddATP, ddCTP, ddGTP, ddTTP). Dideoxynucleotides have a 3'-H group instead of a 3'-OH group. To analyze the sequence, one of the deoxyribonucleotides or the primer is radioactively labeled. DNA polymerase is added to each tube, and the primer is elongated in the 5'-3' direction, forming a complementary strand to the template. During DNA synthesis, DNA polymerase occasionally inserts a dideoxynucleotide instead of a deoxyribonucleotide into the growing DNA chain. Since the dideoxynucleotide lacks a 3'-OH group, it cannot form a 3' bond with any other nucleotide, and DNA synthesis stops. 🧬 For example, in the tube to which ddATP has been added, the polymerase inserts ddATP instead of dATP,causing chainn elongation to stop. In the other tubes, the reactions end in a C, a G, or a T respectively. The DNA fragments from each reaction tube (one for each dideoxynucleotide) are separated in adjacent lanes by gel electrophoresis. The result is a series of bands that form a ladder pattern that is visualized by exposing the film to the gel. The nucleotide sequence is read directly from the base to the top of the gel, which corresponds to the 5'-3' sequence of the DNA strand complementary to the template. DNA sequencing in large-scale genome sequencing projects has been automated and uses machines that can sequence several hundred thousand nucleotides each day. In this procedure, each of the four dideoxynucleotide analogs is labeled with a fluorescent dye of a different color. This is so that the chains that end in adenosine are labeled with one color, those ending in cytosine with another, and so on. All four labeled dideoxynucleotides are added to the same tube. After primer extension by DNA polymerase, the reaction products are loaded into one lane of a gel. The gel is scanned by a laser, causing each band to emit fluorescence of a different color. The sequencing machine has a detector that reads the color of each band and determines whether it represents an A, a T, a C, or a G. This data is represented as colored peaks, each corresponding to a nucleotide in the sequence. Image of the Sanger sequencing flow. Source 🧬 This is the simplest and most universal sequencing method. In second- and third-generation sequencers, sequencing techniques differ among different manufacturers, so we won't go into detail here (you can breathe easy). From traditional sequencing to second-generation sequencing (NGS): the great technological leap Before the advent of Next-Generation Sequencing (NGS), DNA sequencing was performed using the aforementioned Sanger method, which is ultimately a slow and expensive process that allows reading relatively short DNA fragments sequentially. The arrival of the second generation of sequencers marked a before and after, as it introduced the possibility of sequencing millions of DNA fragments simultaneously (massive parallel sequencing). This allowed the DNA of entire organisms to be deciphered in record time, accelerating the development of projects such as the Human Genome Project and making genetic sequencing more accessible. The great feature of NGS is that it fragments DNA into small pieces called short reads, typically measuring between 50 and 600 base pairs (bp), which will be copied many times to amplify the material to be read. Subsequently, these pieces are computationally reconstructed to obtain the complete sequence, in what we call pipelines or bioinformatics workflows. 🧬 This works very well for identifying point mutations and small changes in DNA and giving themmeaning. For example,s variants that can influence hereditary diseases. However, when it comes to analyzing complex regions of DNA, such as highly repetitive ones or those containing large rearrangements, reconstructing the complete sequence can be complicated and prone to errors. Third-generation sequencing: longer reads and real-time analysis While NGS remains the most widely used technology today, third-generation sequencers have brought significant improvements. Instead of splitting DNA into small fragments, these new technologies allow reading much longer fragments, ranging from 10,000 to 100,000 base pairs (long reads), making genome assembly easier and identifying complex structural changes. Another key point is that third-generation sequencing is performed in real-time and does not require prior amplification of DNA, reducing errors introduced by genetic duplication. Additionally, these techniques can detect epigenetic modifications, such as DNA methylation, without the need for additional steps in the process. 🧬 This has significant implications for studies of diseases like cancer, where the regulation of gene expression, that is, which genes are "activated" and when they do, plays a fundamental role. Short reads or long reads? Which is better? The choice between second- and third-generation technologies depends on the specific application. While short reads (NGS) are extremely accurate and allow analyzing large volumes of data at low cost, long reads offer greater capacity to detect structural variants and assemble complex genomes without complicated computational reconstructions. Short reads (NGS, second generation) ✅ High precision for detecting point mutations and small variations. ✅ Lower cost per sequence base. ✅ Ideal for studies of genetic diseases, cancer, and transcriptomics. ❌ Difficulties in assembling complete genomes due to fragmentation. ❌ Limitations to detecting large structural variants. Long reads (third generation) ✅ Long reads that allow assembling complete genomes more easily. ✅ Ability to detect structural variants and repeats in DNA. ✅ Does not require amplification, avoiding biases in the process. ❌ Higher error rate compared to NGS (although it can be corrected with additional coverage). ❌ Higher cost per sequencing. Trends in the use of these technologies for DNA sequencing In recent years, third-generation sequencing has gained ground, especially in studies where it is essential to have a complete view of the genome without interruptions. For example, long-read sequencing has been key to identifying structural variants in diseases such as autism and certain forms of epilepsy (Chaisson et al., 2019). Additionally, in 2022, the Telomere-to-Telomere (T2T) Consortium managed to sequence the entire human genome without gaps for the first time, thanks to the combination of third-generation technologies (Nurk et al., 2022). However, NGS remains the most widely used option in hospitals and diagnostic laboratories, due to its low cost and high precision in detecting individual mutations. In many research cases, the current trend is to combine both technologies in hybrid studies, where short reads offer precision and long reads allow resolving complex regions of DNA. In the clinical setting, third-generation sequencing is still received with some skepticism, which makes sense if we think that an error in sequencing can lead to a diagnostic error for a patient, and therefore directly affect their health. Sequencing technologies are constantly evolving, and it is likely that in the coming years we will see even greater integration between second- and third-generation techniques. It is foreseeable that third-generation sequencing will become widespread both in biomedical research and in the clinic as costs decrease and accuracy improves. Third generation will become increasingly common in hospitals. Regardless of the technology used, the impact of DNA sequencing on personalized medicine, the identification of rare diseases, and cancer research will continue to grow, bringing us ever closer to treatments specifically designed for each person's profile. Cyber Security Cloud Connectivity & IoT IA & Data Healthcare's digital transformation: challenges, needs, and benefits December 18, 2024

We have talked a lot in previous articles about genomic testing and its importance in the revolution of personalized precision medicine. But what are they? What exactly are we talking about? Genomic tests, sometimes also called genetic tests if we are talking about the study of inherited diseases, are those that sequence (“read”) the patient's genome, or a part of it, to identify biomarkers; i.e. specific signals within that genomic sequence, that are related to diseases. How useful is this? Well, knowing the presence or absence of these biomarkers can help doctors to detect the risk of suffering from certain diseases, such as hereditary diseases, or the specific genetic variation related to a disease that is already being suffered, as in the case of oncological diseases. Genomic tests sequence the genome to identify biomarkers that help detect risks for certain diseases. In this last case, knowing which mutation or genetic variation is causing the cancer will help the physician to decide which treatment has the best chance of success, thus personalizing the therapy to the patient. In this article we will find out what genomic tests are, what they mean for us as patients, and what are the most common types of tests. What is genomic or genetic testing? Genomic or genetic testing begins with taking a sample from the patient. In germline genetic testing or genomic testing, i.e., with markers present in all cells of the body, as in diseases inherited from the parents, samples are usually obtained from saliva or blood. This is due to the ease of obtaining this type of sample, with minimally invasive procedures such as a blood draw or saliva swab extraction. Why is blood still used if DNA can be obtained from saliva by simply “swabbing” a swab? Among other reasons, it must be taken into account that in the mouth we have an enormous amount of symbiont or commensal bacteria, which coexist with us and which have their own genetic material. Therefore, they can contaminate to some extent the result of the sequencing of that sample, lowering its quality. Knowing which genetic mutation or variation is causing cancer will help the physician decide which treatment has the best chance of success, thus personalizing the therapy to the patient. This does not mean that the results are useless, but for certain genetic markers that need to be detected with very high precision, the quality may not be high enough for the physician to consider the results as decisive. In the case of looking for genomic variations not in the germline, but in the somatic line, the sampling process is slightly different. Somatic mutations are those inherent to oncological diseases, for example, since the biomarkers we are looking for will be found in some cells of the body and not in others. That is to say, in a tumor cancer, for instance, we will look for the mutation or genomic variation in the tumor cells (“diseased”) in relation to the rest of the body cells (“healthy” or control cells). We will then normally need two samples, one as described above from saliva, or healthy tissue, and another from the tumor biopsy. This process is invasive and, depending on the patient's state of health and the location of the tumor, can be dangerous. Transforming biological samples into data There are liquid biopsies to minimize these risks, in which, through a blood sample, circulating tumor DNA is located in the bloodstream. Although this type of biopsy is not currently useful for all oncological diseases, it is being researched and developed at a dizzying pace due to its enormous practical usefulness in improving the quality of care. Samples are generally obtained from saliva or blood, due to the ease of obtaining this type of sample, with minimally invasive procedures. Once the sample of interest has been obtained, the genetic material of interest (DNA or RNA) will be extracted and prepared for the chosen sequencing technique. The prepared sample will then be introduced into the so-called sequencers, which will be the machines in charge of converting these biological samples into data, into text. This text will be used by the bioinformaticians to search for genetic variations compared to a reference sequence of a human genome, or of the control sample in the case of somatic mutations and give them meaning. Those mutations that have clinical weight in relation to the test requested by the physician will normally be communicated to the physician through the production of a report. From this point on, it will be the specialist physician, who has or should have all the information concerning the patient, who will assess the value of these detected variations and make the best decisions for the patient's health. IA & Data Omics sciences: redefining health October 29, 2024 Genetic testing types for precision medicine In this post we will focus on molecular tests related to the study of DNA sequencing. Apart from these, we can have other types of tests related to genetics within the world of precision medicine, including: Biochemical tests to measure protein activity. Expression tests to study which genes are activated. Chromosomal tests that look at changes in genetic material at the chromosomal level, on a large scale. Molecular tests, which are those that look for the aforementioned genetic variations, include whole genome sequencing (WGS) and whole exome sequencing (WES) and targeted sequencing, as in panels. Within healthcare practice, this translates into the following highlighted tests: Neonatal screening This is the most common genetic test; we are all familiar with the heel prick test. Through this systematic test, newborns can be screened for the possibility of suffering from some type of genetic disease before the adults around them can see the symptoms. ■ It may be too late in diseases such as glutaric aciduria type I, by the time the baby shows symptoms, and can have lifelong consequences. On the other hand, if detected early, the onset of symptoms can be prevented “simply” by adjusting the diet. Carrier screening or carrier-screening This type of testing is often used to help future parents to know if they are carriers of certain diseases, thus knowing if there is a risk of transmitting them to their offspring. ■ These tests are performed on people who have a family history and therefore have a high chance of being transmitters of inherited diseases. Prenatal diagnosis These are tests performed on the fetus to detect early modifications in its genes and chromosomes, so that both physicians and parents can make an informed decision. This type of test is performed, as before, in cases in which the parents may be carriers of hereditary diseases due to family history. However, given the universalization of these tests, they are also usually performed in pregnancies suspected of being at risk or that have presented some type of problem during their development. ■ In this case, the sample is obtained either by amniocentesis or by chorionic villus sampling (placental sampling). Genetic predisposition testing Panels sequence very specific parts of the genome as they target specific genes, which can be between one gene, or a few, to several thousand. They are used when it is known in which gene or genes the mutation should be sought. The best-known case might be the breast and ovarian cancer panel, which includes the BRCA1 and BRCA2 genes as the main indicators of the risk of developing breast and ovarian cancer in the future. But these are not the only genes that may be involved in this type of cancer; most current tests already sequence around 30 genes. Why perform a panel and not a WGS or WES? Well, because by reducing the size of the genetic material to be sequenced the technique is more cost-effective and the data produced are much easier for professionals to handle, so that waiting times for results are reduced. ■ These panels can be used both to determine genetic predisposition and to help determine patient treatment. Conclusion Genetic testing has revolutionized modern medicine by enabling early detection of inherited diseases and prediction of genetic predispositions. From testing for prospective parents, prenatal diagnostics to genetic predisposition panels, these tools provide key information to make informed and personalized health decisions. The universalization and continuous improvement of these tests ensure that more and more people can benefit from more accurate and effective medical care. Genetics thereby opens a window into the future of preventive and personalized medicine. With the development of more affordable and accurate tests thanks to digital technologies, more and more people will be able to know their risks and take proactive measures to maintain their health. In addition, the integration of genetics into clinical practice is not only improving the diagnosis and treatment of diseases but also providing a more holistic and patient-centered approach.

In recent years the term '–omics' has been gaining popularity in scientific and medical circles. The omics sciences, a category that includes genomics, proteomics, transcriptomics, metabolomics... among others, represent a comprehensive approach to the study of biological components within a complex biological system such as, for example, a person. Among all of them, genomic science stands out as one of the most revolutionary areas, laying the groundwork for major medical advances in the world of precision medicine that, assisted by the processing of large amounts of data and AI, improves the ability to prevent, detect and treat diseases in a personalized way, based on the individual genetic characteristics of each person. What are the omics sciences? The concept of omics derives from the suffix '-oma', which refers to the complete set of biological components of a specific system. For example, the 'genome' refers to the complete set of genes of an organism, while the 'proteome' refers to all the proteins expressed ('made') by the cells or tissues of an organism. The omics sciences, therefore, are the study of these assemblies as a whole, rather than examining each component (a gene, a protein...) in isolation. All these sciences have in common an integrative and systemic approach, seeking to understand not only the individual parts of an organism, but how they all interact to influence its health and development. This holistic approach is fundamental to modern medicine, as it allows for a more precise and detailed understanding of the biological mechanisms that affect people's health. Genomics: the heart of the omics sciences Of all the omics sciences, genomics is perhaps the best known and most fundamental. It focuses on the study of DNA, the genetic material that contains all the necessary information for the development, functioning and reproduction of living beings. It could be said that DNA is the cell's instruction manual. Our genome is all that DNA, including all its genes (which are nothing more than functional units of DNA). What is genomics? Genomics encompasses the study of the structure, function, evolution and mapping of the genome. The terms 'genetics' and 'genomics' are often confused, but have important differences: Genetics is the study of individual genes and how they are inherited from generation to generation, focusing on how specific traits (such as eye color, or predisposition to a disease) are passed down through families. Genomics refers to the study of the entire genome and its interaction with the environment. Whereas traditional genetics usually studies one or a few genes at a time, genomics allows scientists to study all the genes of an organism simultaneously, along with their interactions. ✅ The key tool in genomics is DNA sequencing, a process that makes it possible to determine the exact order of nucleotides (the basic units of DNA) in an organism's genome. Early sequencing technologies, such as the Sanger method, were slow and expensive, but advances in recent decades have led to the development of next-generation sequencing (NGS) technologies, which are much faster and more efficient.. Other omics sciences and their relationship to precision medicine While genomics is the best known and most applied omics science, the other omics disciplines also play a crucial role in precision medicine. The combination of different omics approaches allows for a comprehensive view of the organism, which can reveal complex interactions that are not evident when studying only one layer of information. Proteomics Proteomics is the study of the complete set of proteins expressed by a genome, and is essential for understanding biological function in the context of health and disease. Although genes provide the instructions for creating proteins, it is proteins that carry out most cellular functions. Proteomics can help identify biomarkers of disease, i.e., proteins that change in quantity or structure in response to a disease, in precision medicine. These biomarkers can be used for early diagnosis, treatment monitoring or even as therapeutic targets. ✅ It has been identified, for example, that certain levels of specific proteins can predict the progression of neurodegenerative diseases such as Alzheimer's or Parkinson's, allowing earlier and more personalized intervention. Transcriptomics Transcriptomics focuses on the study of RNA, the molecule that transcribes DNA information for use in protein synthesis. Let's say it transcribes that instruction manual that is DNA into tools that can be used to build the protein in question. Analysis of RNA transcripts can provide dynamic insight into how genes are expressed (i.e. 'turned on') in different contexts, such as the development of a disease or response to a treatment. ✅ In the field of oncology, for example, transcriptomics has made it possible to identify specific genetic signatures that can predict which patients will respond best to certain treatments. This can help physicians avoid ineffective or unnecessary treatments and direct patients to the most appropriate therapies. Epigenómica Epigenomics deals with changes in gene expression ('activation' of some genes or others) that are not caused by alterations in the DNA sequence, but by chemical modifications in the DNA or in the associated proteins, such as DNA methylation. These epigenetic modifications can be influenced by environmental factors, such as diet, stress, exposure to toxins, ... In precision medicine, epigenomics enables a deeper understanding of how the environment interacts with the genome to influence health. ✅ Epigenetic studies, for example, have identified DNA methylation patterns that can predict response to certain drugs or the risk of developing cancer. The future of omics and precision medicine The future of omics and precision medicine is extremely promising. Due to the rapid evolution of sequencing and data analysis technologies, it will soon be possible to integrate multiple layers of omics information into a single clinical analysis, providing physicians with a complete picture of a patient's health status. The development of artificial intelligence and machine learning tools is also making it possible to analyze the vast datasets generated by the omics sciences in a way that supports the clinician's decision making. They will make it possible to identify complex patterns that would be impossible to detect using traditional methods, which will lead to even more precise personalization of medical treatments. IA & Data Artificial Intelligence in hospital emergency rooms to improve patient care October 23, 2023

Nowadays we have all heard about Big Data, Artificial Intelligence, algorithms... and these are terms that we usually relate to the world of business or new technologies. But what if they could improve our health? This is where precision medicine comes into play. And what is it? The most commonly used definition is as follows: Precision medicine is the concept of customizing the treatment and prevention of diseases by considering differences in genetic, environmental or even lifestyle factors specific to groups of people. This means that precision medicine is an emerging clinical approach based on providing the patient with the most personalized therapy possible. To achieve this personalized therapy, the medical professional's decision making will be based on clinical, genomic, environmental and socioeconomic data, thereby building a patient profile in the greatest possible detail. This complete patient profile is what would be called a phenotype. Customized medicine to preserve your well-being Does this mean that the therapy will be specifically designed for me as a patient? Not exactly. It means that the medical professional will be able to find the therapy that best fits your patient profile, your phenotype. ✅ This small difference is what separates the terms customized medicine from precision medicine, and it is what makes the term precision medicine more accurate, although the two tend to be used interchangeably. Precision medicine methods identify patient phenotypes with less common responses to treatment or unique health needs. We already know that the same treatment, for example, the same drug, does not affect us all equally. Who doesn't know someone who has had to adjust the dosage of a drug, and adjust it (always following medical advice) until they find the one that works for them? In this simple example, thanks to precision medicine, the doctor will be able to give a more effective prescription from the beginning of the treatment. Data, data and more data for a better treatment (and even to avoid it) Another quite common case is that of cancer treatments. There are very aggressive types of tumors that need to be treated quickly, but for which there may be as many as 8 different treatments. How does the oncologist know which treatment will work best for a particular patient? By knowing the specific mutation of the cancer diagnosed in that patient, which only requires a blood sample and a genetic panel, and by knowing the patient's medical history. Data, data, data. The prevention and early detection of diseases is one of the major objectives of this medical approach. But what's more, precision medicine not only helps to provide better treatment, but it also sometimes helps to prevent that treatment from ever becoming necessary. ✅ We can find out whether the patient has a certain predisposition to diseases, such as cancer, by knowing both the phenotype and the genotype (set of genetic characteristics), and thus prevent the disease. The role of AI in precision medicine If we add to this knowledge, to the data we have, the generation of AI algorithms, we can have in our hands mathematical models for disease prevention. AI takes advantage of calculation and inference methods to generate ideas, allows the system to reason and learn, and enhances decision making by doctors. AI generates ideas, reasons, learns and empowers medicine. All of this sounds futuristic or perhaps even exclusive, but the truth is that precision medicine is here and it is here to stay. The final push for precision medicine has been the rise of genomic testing. The cost of sequencing, of reading our genome, has dropped from hundreds of millions of euros to hundreds of euros, which has largely contributed to the fact that tests that were previously exclusively for research are now normal. It is worth noting that, for example, there are panels such as those for the BRCA genes that are now routine and make it possible to diagnose predisposition to breast cancer. In addition, health systems are incorporating more and more genomic tests into their portfolio of services, and national and European initiatives are being carried out to facilitate the integration of genomic data with the rest of the clinical data. Healthcare systems are incorporating more and more genomic tests into their portfolio of services. In conclusion, thanks to precision medicine, and the inclusion of your genetic data in your medical record, your trusted doctor will be able to treat you in the most efficient and safe way, with the single purpose of preserving your health and the health of those around you. AI of Things Artificial Intelligence in hospital emergency rooms to improve patient care October 23, 2023