Pathogenic microdeletions/duplications (pCNVs) account for a significant perinatal morbidity and mortality and have a 1/100-200 prevalence in the general prenatal population. ^1,2 Cell-free DNA noninvasive prenatal testing (cfNIPT) shows significant limitations for comprehensive profiling of pCNVs below 7Mb in size. The cfDNA fragmentation and the maternal genomic background cause significant limitations for a reliable detection of the full fetal pCNV spectrum. ^3,4 

With a proof-of-concept study published in Prenatal Diagnosis Journal, we have shown the feasibility to isolate, with a proprietary laboratory workflow, intact circulating fetal extravillous trophoblasts (cEVTs) from pregnant women’s blood for non-invasive fetal copy number variant screening through single cell sequencing. ⁵ We have also described in Ultrasound in Obstetrics and Gynecology journal a paradigmatic case of diandric partial hydatidiform mole identified with this novel proprietary cell-based noninvasive prenatal testing (CB-NIPT) as a stand-alone early blood-based investigation. ⁶

Our technology represents therefore the promise for a true high‐resolution comprehensive genomic profiling for the detection of fetal pathogenic genomic abnormalities down to less than 1Mb in size, below the resolution of cell-free DNA screening. In the general population of pregnancies, this would allow the opportunity for a significant reduction of residual risk for clinically relevant pCNVs at early gestational weeks as well as improvement of women’s clinical management. 

Menarini Silicon Biosystems has developed this new workflow combining the enrichment of circulating fetal extravillous trophoblasts from maternal blood using proprietary ferro-fluid technology ( CELLSEARCH® system) and unique cell sorting capabilities with a high throughput single cell sequencing using Ampli1 WGA kit and Ampli1 LowPass for library preparation. 

The preliminary evidence derived from the proof-of-concept study is being further explored through a multicenter Italian clinical validation study in a large population of pregnancies enriched for fetal genomic abnormalities. The scope of the study is to demonstrate cbNIPT scientific validity in a large blinded prospective performance study comparing the fetal genomic result with chromosomal microarray and/or karyotype with that of cEVTs screening. 

Bibliography

1. Wapner RJ, Martin CL, Levy B, et al. Chromosomal microarray versus karyotyping for prenatal diagnosis. N Engl J Med. 2012; 367(23):2175‐2184
2. Srebniak MI, Joosten M, Knapen MFCM, et al. Frequency of submicroscopic chromosomal aberrations in pregnancies without increased risk for structural chromosomal aberrations: systematic review and meta‐analysis. Ultrasound Obstet Gynecol. 2018;51(4): 445‐452.
3. Zhao C, Tynan J, Ehrich M, et al. Detection of fetal subchromosomal abnormalities by sequencing circulating cell‐free DNA from maternal plasma. Clin Chem. 2015;61(4):608‐616.
4. Kucharik M, Gnip A, Hyblova M, et al. Non‐invasive prenatal testing (NIPT) by low coverage genomic sequencing: detection limits of screened chromosomal microdeletions. PLoS One. 2020;15(8): e0238245.
5. Doffini A, Forcato C, Mangano C, Lattuada D, Aversa R, Maranta C, Giovannone ED, Buson G, Bolognesi C, Maiocchi R, Dori M, Jamal L, Ahmad RB, Yeo GSH, Yeo TW, Saragozza S, Silipigni R, Serafini M, Biondi A, Perego S, Vergani P, Ferrazzi E, Ricciardi-Castagnoli P, Musci TJ, Grati FR. Isolation of single circulating trophoblasts from maternal circulation for noninvasive fetal copy number variant profiling. Prenat Diagn. 2023 Jan;43(1):14-27.
6. Mangano C, Doffini A, Forcato C, Boito S, Lattuada D, Giovannone ED, Buson G, Hyett J, Musci TJ, Grati FR. Hydatidiform mole identification using noninvasive single cell sequencing on circulating extravillous trophoblasts isolated from maternal blood. Ultrasound Obstet Gynecol. 2024 Feb 14. doi: 10.1002/uog.27615