Evaluation of Next-Generation Sequencing and Crystal Digital PCR for Chimerism Monitoring of Post-Allogeneic Hematopoietic Stem Cell Transplantation

• Published in: Biology of blood and marrow transplantation 2021-01, Vol.27 (1), p.89.e1-89.e10
• Main Authors: Pedini, Pascal, Cherouat, Nicem, Basire, Agnes, Simon, Sophie, Budon, Laurène, Pourtein, Monique, Grondin, Sandra, Moskovtchenko, Philippe, Chiaroni, Jacques, Michel, Gérard, Frassati, Coralie, Picard, Christophe
• Format: Article
• Language: English
• Subjects: Chimerism; Crystal digital PCR; Hematopoietic stem cell transplantation; Life Sciences; NGS
• ISSN: 2666-6367; 1083-8791; 2666-6367; 1523-6536
• DOI: 10.1016/j.bbmt.2020.09.023

•Quantification of chimerism by crystal digital PCR (cdPCR)and next-generation sequencing (NGS) Devyser are sensitive and accurate methods.•There are correlation and concordance between cdPCR or NGS Devyser and ddPCR.•cdPCR and NGS Devyser allow genotyping and monitoring in a single technique.•ADVYSER (Devyser NGS) is a complete software suite for interpreting genotyping and monitoring.•Whereas NGS Devyser is designed for large series, cdPCR is sufficiently flexible for control and emergency situations. Hematopoietic stem cell transplantation (HSCT) is a curative treatment for most hematologic diseases. To evaluate the level of donor engraftment, chimerism must be carefully monitored after HSCT. Short tandem repeats, quantitative PCR (qPCR), and, more recently, digital PCR (dPCR) are widely used to determine the proportions of donor and recipient cells after HSCT. The screening and quantification of chimerism have been evaluated by 2 new methods: a ready-to-use next-generation sequencing (NGS)-based method using the Devyser ChimerismNGS kit and an original combination of the Stilla crystal digital PCR (cdPCR) platform with 3-color multiplexing capacity using GenDX KMRtrack reagents. The genotyping of 4 HSCT pairs by cdPCR using 11 triplex mixes of the GenDX KMRtype kit was consistent at 98.8% with qPCR. Informative samples (n = 20) from 6 donor-recipient pairs and 1 external proficiency test demonstrated the reliability of the results (0.1% to 50%) for the 2 methods. The methods are also highly sensitive (0.1%) and accurate. The chimerism values of the 2 methods are correlated and concordant with those of the reference methods. In addition, the ADVYSER software (Devyser) is user-friendly and well adapted to chimerism monitoring. In conclusion, these 2 innovative methods are easy to perform and user-friendly in all molecular, hematology, and immunogenetic laboratories and allow the genotyping and monitoring of chimerism with high performance and sensitivity.