Assessment of Minimal Residual Disease by Next Generation Sequencing in Peripheral Blood as a Complementary Tool for Personalized Transplant Monitoring in Myeloid Neoplasms.

Paula Aguirre-Ruiz,Felipe Prosper,María Calasanz,María Larráyoz,Iria Vázquez,María Zudaire,Eva Bandrés,María Viguria,Margarita Redondo,Beñat Ariceta,Patricia Arnedo,Zuriñe Blasco-Iturri,Amagoia Mañú,Axier Jauregui,Marta Fernández-Mercado,Almudena Aguilera-Diaz,María Mateos

doi:10.3390/jcm9123818

Abstract

Patients with myeloid neoplasms who relapsed after allogenic hematopoietic stem cell transplant (HSCT) have poor prognosis. Monitoring of chimerism and specific molecular markers as a surrogate measure of relapse is not always helpful; therefore, improved systems to detect early relapse are needed. We hypothesized that the use of next generation sequencing (NGS) could be a suitable approach for personalized follow-up post-HSCT. To validate our hypothesis, we analyzed by NGS, a retrospective set of peripheral blood (PB) DNA samples previously evaluated by high-sensitive quantitative PCR analysis using insertion/deletion polymorphisms (indel-qPCR) chimerism engraftment. Post-HCST allelic burdens assessed by NGS and chimerism status showed a similar time-course pattern. At time of clinical relapse in 8/12 patients, we detected positive NGS-based minimal residual disease (NGS-MRD). Importantly, in 6/8 patients, we were able to detect NGS-MRD at time points collected prior to clinical relapse. We also confirmed the disappearance of post-HCST allelic burden in non-relapsed patients, indicating true clinical specificity. This study highlights the clinical utility of NGS-based post-HCST monitoring in myeloid neoplasia as a complementary specific analysis to high-sensitive engraftment testing. Overall, NGS-MRD testing in PB is widely applicable for the evaluation of patients following HSCT and highly valuable to personalized early treatment intervention when mixed chimerism is detected.

Highlights

Allogenic hematopoietic stem cell transplant (HSCT) is a potentially curative treatment in patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS), reducing risk of relapse and improving overall survival [1,2,3]; clinical outcomes still vary among patients [4,5,6,7]
Comparison of 4 peripheral blood (PB) and bone marrow (BM) samples at follow-up times showed high correlation (R2 = 0.9978; p-value < 0.0001) (Supplementary Materials Figure S2). These results showed similar sensitivity of next generation sequencing (NGS) on PB and BM samples both for the diagnosis and follow-up, confirming that PB samples are suitable for molecular testing when BM is not available
We have focused on patients with low levels of mixed chimerism (MC) in hope that close monitoring and NGS-based minimal residual disease (NGS-minimal residual disease (MRD)) detection could help to take specific clinical decisions such as better timing for the initiation of antineoplastic treatment

Summary

Introduction

Allogenic hematopoietic stem cell transplant (HSCT) is a potentially curative treatment in patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS), reducing risk of relapse and improving overall survival [1,2,3]; clinical outcomes still vary among patients [4,5,6,7]. Newer techniques to analyze chimerism with higher sensitivity (0.01–0.1%) have relatively recently emerged, such as quantitative PCR analysis using insertion/deletion polymorphisms (indel-qPCR) and droplet-digital PCR (ddPCR) [15,16,17]. These assays do not detect the presence of disease, but rather they offer a percentage of recipient’s DNA as a surrogate measure for recurrence. This lack of specificity is problematic in chimerism assays, showing high sensitivity, as non-malignant recipient cell lineages may be present in various sample types without representing disease relapse [18]. To maximize sensitivity and specificity, assays such as reverse transcriptase polymerase chain reaction (RT-PCR) may be applied to follow-up specific genetic alterations [19]; this is a major limitation in a disease characterized by a striking broad array of different potential oncogenic events across a notable number of genes

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