Molecular detection of minimal residual disease in multiple myeloma

Abstract

Despite the significantly higher complete remission rates and improved survival achieved in the last decade, multiple myeloma (MM) patients continue to relapse due to persistence of minimal residual disease (MRD). Generally, MRD refers to persistence of low levels of disease in the order of one tumour cell in ≥10 normal cells. Currently, molecular and immunophenotypic techniques are employed for MRD detection. This review focuses on MRD detection by molecular techniques, including allele-specific oligonucleotide polymerase chain reaction (ASO-PCR), next-generation sequencing (NGS) and digital PCR (dPCR), in addition to a brief description of, and comparison with, multiparameter flow cytometry. The basic principles, technical advantages and limitations, and the clinical impact of all three molecular techniques are reviewed and compared. They all have a sensitivity of at least 10 , among which ASO real-time quantitative PCR is the most well-standardized, and NGS carries the highest sensitivity and applicability, while dPCR is still under investigation. Furthermore, molecular MRD negativity is a favourable prognostic factor for survival of patients with MM. However, several challenges inherent to molecular detection of MRD still remain to be overcome, particularly false negativity and failure to detect extramedullary disease. Finally, detection of MRD from peripheral blood remains challenging.

Comparison of next-generation sequencing (NGS) and next-generation flow (NGF) for minimal residual disease (MRD) assessment in multiple myeloma

“…reaching the same sensitivity level and using the same marrow pull) should be performed to shed light into this issue, although an intrinsic false-negativity risk will always exist for both methodologies due to the patchy nature of the disease. As it was mentioned before, samples undergoing laboratory analysis are not the same, and a great variation can be detected from the first marrow pull (which is usually sent to the morphology laboratory) to the last one (which is distributed for molecular, cytogenetic, and immunophenotypic studies) 23 , 32 . This could be a feasible explanation for the 15 discordant cases in our cohort.…”

“…Recently, the EuroFlow consortium has developed a two-tube, eight-color flow assay that allows the simultaneous analysis of up to 10 million cells (next-generation flow, NGF) 22 ; NGF has been already approved by the IMWG as a reference method to detect and define immunophenotypic CRs in MM 16 . NGF reaches a sensitivity of 2 × 10 −6 , overcoming previous flow protocols (10 −4 –10 −5 ), but it is highly dependent on the precise identification of the pathologic immunophenotype, requiring a high level of expertise; in addition, the quality of the sample must be high and should be promptly processed 23 .…”

New Molecular Technologies for Minimal Residual Disease Evaluation in B-Cell Lymphoid Malignancies

“…Based on these evidences, several studies were performed to define the clinical impact of LymphoSIGHT ® NGS in ALL, MCL, CLL and MM for MRD analysis. Experiments were also carried out in the context of different clinical trials, with the aim to assess the outcome predictor value of this technology [ 58 , 60 , 61 , 63 , 64 , 65 , 66 , 67 , 68 , 69 , 70 , 71 , 72 , 73 , 74 ]. In particular, Martinez-Lopez et al assessed that NGS method in MM has applicability almost comparable to MFC, but is able to reach higher sensitivity, with MRD level by NGS correlating with time to progression and OS [ 58 ].…”

Comparison of NGS and MFC Methods: Key Metrics in Multiple Myeloma MRD Assessment

“…State-of-the-art methods for the highly sensitive and standardized detection of bone marrow (BM) MRD in multiple myeloma (MM) include next-generation sequencing (NGS) and multicolor flow cytometry (MFC). NGS, in the form of an ultradeep targeted sequencing assay, targets immunoglobulin heavy-and light-chain DNA sequences using consensus primers [1]. In MFC, achieved through a two-tube, eight-color antibody panel, the identification of malignant cells is based on an aberrant immunophenotype displayed by neoplastic MM cells compared to normal plasma cells [2].…”

“…In MFC, achieved through a two-tube, eight-color antibody panel, the identification of malignant cells is based on an aberrant immunophenotype displayed by neoplastic MM cells compared to normal plasma cells [2]. Based on optimal requirements, including sample quality and sample processing, both MM MRD detection methods have been described to reach a sensitivity of up to one tumor cell per 1,000,000 BM cells (10 −6 ) [1,3].…”