Application of allele-specific quantitative PCR using genomic DNA to monitor minimal residual disease based on mutant gene levels following allogeneic hematopoietic stem cell transplantation in patients with hematological malignancies: comparison of mutant levels with autologous DNA percentage by short tandem repeat-PCR

Abstract

Background: Quantitative evaluation of minimal residual disease (MRD) following hematopoietic stem cell transplantation (HSCT) is indispensable for patients with hematological malignancies. In addition to established MRD markers such as immunoglobulin and T-cell receptor gene rearrangements, fusion genes, or aberrantly expressed genes, single nucleotide mutations are considered one of the MRD markers that reflect the malignant cell clone.

Methods: We compared the quantity of mutant genes by allele-specific quantitative polymerase chain reaction (AS-qPCR) for single nucleotide mutations (TP53 410T>A and PTPN11 1508G>A) with the percentage of autologous DNA by short tandem repeat (STR)-PCR.

Results: Following HSCT, the quantity of mutant genes detected by AS-qPCR correlated with the percentage of autologous DNA assessed by the STR-PCR. Moreover, mutant DNAs were detected at a quantifiable level before relapse, whereas the percentage of autologous DNA was less than 5%, that is, complete chimerism.

Conclusions: The AS-qPCR approach for single nucleotide mutations was accurate and highly sensitive for monitoring pre-transplantation as well as post-transplantation MRD. AS-qPCR for single nucleotide mutation is suitable for monitoring MRD in patients who lack previously established MRD markers.