Cell-Free DNA in Pediatric Solid Organ Transplantation Using a New Detection Method of Separating Donor-Derived from Recipient Cell-Free DNA

Evgenia Preka, Drew Ellershaw, Natalie Chandler, Helena Ahlfors, Helen Spencer, Lyn S. Chitty, Matthew J. Fenton, Stephen D. Marks

Research output: Contribution to journalArticleAcademicpeer-review

5 Citations (Scopus)

Abstract

BACKGROUND: The use of cell-free DNA (cfDNA) as a noninvasive biomarker to detect allograft damage is expanding rapidly. However, quantifying the low fraction of donor-derived cfDNA (ddcfDNA) is challenging and requires a highly sensitive technique. ddcfDNA detection through unique donor single nucleotide polymorphisms (SNPs) is a recent new approach, however there are limited data in pediatric solid organ transplant (SOT) recipients. METHODS: We developed an assay using a combination of 61 SNPs to quantify the ddcfDNA accurately using a custom R script to model for both the patient and donor genotypes requiring only a single sample from the allograft recipient. Performance of the assay was validated using genomic DNA (gDNA), cfDNA and donor samples where available. RESULTS: The R "genotype-free" method gave results comparable to when using the known donor genotype. applicable to both related and unrelated pairs and can reliably measure ddcfDNA (limit of blank, below 0.12%; limit of detection, above 0.25%; limit of quantification 0.5% resulting in 84% accuracy). 159 pediatric SOT recipients (kidney, heart, and lung) were tested without the need for donor genotyping. Serial sampling was obtained from 82 patients. CONCLUSION: We have developed and validated a new assay to measure the fraction of ddcfDNA in the plasma of pediatric SOT recipients. Our method can be applicable in any donor-recipient pair without the need for donor genotyping and can provide results in 48 h at a low cost. Additional prospective studies are required to demonstrate its clinical validity in a large cohort of pediatric SOT recipients.
Original languageEnglish
Pages (from-to)1300-1309
Number of pages10
JournalClinical Chemistry
Volume66
Issue number10
DOIs
Publication statusPublished - 1 Oct 2020

Keywords

  • cell free DNA
  • donor-derived cell-free DNA
  • molecular diagnostics
  • noninvasive biomarker
  • pediatric solid organ transplantation

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