Thalassemia is an inherited blood disorder, which main consequence is a reduced production of hemoglobin leading to mild to severe anemia. Since the diagnosis of one of its forms, ß thalassemia, as mentioned in our recommended lecture for this month[1], has proven challenging with Non-Invasive Prenatal Testing (NIPT) techniques, the authors propose a new assay utilizing cell-free DNA (cfDNA) from maternal plasma with Non-Invasive Prenatal Haplotyping (NIPH). This method is proposed to determine paternal inheritance in pregnancies at risk for β-thalassemia using targeted next-generation sequencing (NGS).
Since the discovery of the presence of fetal genetic material in maternal blood almost 30 years ago, new tools for non-invasive prenatal testing are continuously being developed as a way to avoid the risks related to invasive techniques such as amniocentesis and chorionic villi sampling, some of them having reached clinical practice. However, for others, especially in the case of monogenic disorders, such as ß thalassemia, the technique appears to be technically challenging. This is due to the fact that the maternal genome and the fetal genome share homology and can only be differentiated by single-point variant differences.
Like for other genetic diseases, geography is an important discriminant, with specificities related to well identified areas. The present study, that was conducted in Cyprus, focusses on a population where the carrier frequency for ß thalassemia reaches 12%. The pathogenic variants are located on the HBB gene. In this case, NGS, a high throughput methodology able to analyze a high number of SNVs on the ß-globin gene cluster appears to potentially reduce the risk of misdiagnosis thus providing high diagnostic accuracy and specificity.
The study involved 96 random samples to identify the SNVs and designed a custom NGS genotyping panel with 169 SNVs, including 15 common pathogenic ones. The panel’s efficiency was evaluated on cfDNA samples from 17 at-risk families, demonstrating a 100% success rate in diagnostic interpretation.
Although this study well underlines the complexity and limits of such a diagnostic approach, it demonstrates that effective noninvasive solutions can be developed to detect the paternally inherited haplotype with a high sensitivity (88.8%) and specificity (98.8%).
We at 4bases have developed the THALASSEMIA panel, a novel NGS Medical Device for the molecular profiling of variants causing alpha and beta Thalassemia.
THALASSEMIA panel is a kit for the analysis of the HBB, HBA1, HBA2, HBD, HBG1, HBG2, HS-40 and LCRB genes through a molecular protocol based on NGS technologies. The kit is validated for germline analysis (SNPs, indels) of DNA extracted from blood samples.
[1] Byrou S, Brouwer RWW, Tomazou M, Tamana S, Kountouris P, Lederer CW, Petrou M, Ozgur Z, den Dekker X, Azmani Z, Christou S, Makariou C, Kleanthous M, IJcken WFJV, Papasavva T. Non-Invasive Determination of the Paternal Inheritance in Pregnancies at Risk for β-Thalassaemia by Analyzing Cell-Free Fetal DNA Using Targeted Next-Generation Sequencing. Int J Mol Sci. 2025 Jan 10;26(2):570. doi: 10.3390/ijms26020570. PMID: 39859286; PMCID: PMC11765003.
