Sequencing of the genome, or parts of it, is becoming an increasingly common practice in Diagnostics. What are the advantages and opportunities?

We discuss this with Dr. Giovanni Cazzaniga, researcher of Medical Genetics at the Department of Medicine of the University of Milan Bicocca and head of the laboratory of hemato-oncological diagnostics Tettamanti, belonging to the Pediatric Clinic of the Fondazione Monza e Brianza per il Bambino e la sua Mamma (MBBM).

Dr. Cazzaniga, please explain what Next-Generation Sequencing (NGS) consists of.

It’s a molecular technique that’s been used in research for about 15 years but has only recently begun to be used in Diagnostics. It offers the great advantage of being able to perform diagnostic analysis that’s not restricted to what is already known. As the name implies, it involves sequencing the DNA or RNA of certain cells in a patient and potentially discovering something new or confirming something already known. We then speak of Next-Generation because the First-Generation comes from the late ’90s and had many limitations, both in the number of samples that could be sequenced and in the length of the sequences, but those limits that have now been exceeded.

As an example, with First-Generation Sequencing it was possible to sequence up to 1000 bases (i.e. the bricks that make up our DNA) in a few hours. Next-Generation Sequencing allows you to sequence the entire human genome (3.2 billion “bricks”) in less than a day. The challenge is no longer producing the data but analyzing it.

Moreover, with Next-Generation Sequencing we can identify single mutations, small variations of a few bases or even structural rearrangements of even very large portions of genes. All this has opened up the possibility of running studies that until a few years ago were unthinkable“.

What does your lab work on?

The laboratory in which I work deals with hemato-oncology, more specifically with Acute Lymphoblastic Leukemia (ALL) in pediatric ages. Acute Lymphoblastic Leukemia (ALL) is the most common leukemia in children (S4L has published on this; here for more information). In addition, we’re the reference laboratory for molecular analysis for all the centers of the Italian Association of Pediatric Hematology and Oncology (AIEOP) that treat about 400 children who contract ALL every year. We don’t limit ourselves to diagnosing ALL for pediatric patients, but we also provide diagnostic services for adult hematologic-oncologic patients.

Returning to pediatric patients, the diagnostic laboratory, using Next-Generation Sequencing, identifies rearrangements of T-cell receptor (T-cell immunoglobulin) genes. These rearrangements are specific to the individual cell of the leukemic patient, different in each patient, and are markers of Minimal Residual Disease (MRD). We monitor these rearrangements over time in order to verify that the patient is responding well to treatment. Therefore, if the Minimal Residual Disease doesn’t advance, it means that the patient responds well to treatment, but if there is an advance in Minimal Residual Disease, then the Doctor treating the child must alter the treatment protocol“.

What advantages has Next-Generation Sequencing brought to your lab?

It’s essentially simplified the procedure and shortened the time for both diagnosis and reporting. Previously, we had to do rather elaborate and lengthy diagnostic investigations. Now, thanks to Next-Generation Sequencing, we can identify the rearrangements of each of the 400 children who get leukemia in Italy every year, with a single test at the onset of the disease and then monitor the Minimal Residual Disease several times (over extended periods) with other techniques.

In addition to measuring Minimal Residual Disease, we use Next-Generation Sequencing to identify other genetic abnormalities, called fusion transcripts, for genes that confer a worse prognosis but for which there are drug treatments. These analyses are performed between Day #33 and Day #50 after the start of treatment so that the pediatric patient’s treatment can be best targeted. Today, these analyses are performed only for those children who don’t respond to conventional therapies, but it will be implemented for all patients as we’ve seen that it’s important to monitor all patients for known genetic abnormalities to anticipate and make more effective the personalization of treatments.”

In what other areas does your lab apply Next-Generation Sequencing?

Our laboratory is also open to service other hospitals besides San Gerardo Hospital. In fact, we deal with sequencing services for mutation screening in adults for myeloproliferative diseases and myelodysplasias”.

Returning to the diagnosis of ALL in children, can we say that this diagnostic and therapeutic approach is an example of personalized medicine?

“Certainly. Every child has characteristic genetic abnormalities that differentiate it and determine the therapy they’ll need to follow. This is possible, thanks to Next-Generation Sequencing, which provides us with the possibility of identifying new gene alterations, as well as confirming known abnormalities.

The data that is generated by following children in this detail is quite complicated to understand. The next step would be to apply Artificial Intelligence (AI) to data analysis, so as to make diagnosis faster and more reliable and help the Doctor choose the appropriate therapy.”

This post is also available in: Italiano


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