MSACL 2018 US Abstract

Topic: Proteomics

Use of a MS-based Targeted Approach for Detection of Minimal Residual Disease (MRD) in Multiple Myeloma

Carlo Martins (Presenter)
Memorial Sloan Kettering Cancer Center

Bio: Carlo is a research fellow at Memorial Sloan Kettering Cancer Center, in the Department of Hematopathology. He had his first contact with proteomics during his PhD, studying rheumatic heart disease by 2D-DIGE, MALDI and ESI mass spectrometry approaches, at the School of Medicine from University of Sao Paulo, in Brazil. He also used proteomics to study food allergens in Brazilian fruits and tuberous roots. Then, he moved to Maryland, for a postdoctoral position at MedImmune (Astra Zeneca) to analyze the secreted proteins of bispecific antibody-producing cells in culture. In this project he used click chemistry, pulsed SILAC, and automation for both protein digestion and pre-fractionation processes.

Authorship: Carlo Martins(1); Maria Stella Ritorto(1); Sarah Huet(1); Janine Pichardo(1); San San Yi(1); Ola Landgren(1); Ahmet Dogan(1)
(1) Memorial Sloan Kettering Cancer Center

Short Abstract

Minimal residual disease (MRD) is the most important biomarker for management of patients with multiple myeloma. Here we developed a method to detect lower levels of peptides from variable light-chain of immunoglobulins produced in multiple myeloma, to better assess MRD. We used immunoglobulin enrichment and targeted proteomics to detect the peptides specific for each patient, to help individualized therapy.

Long Abstract

Introduction

Minimal residual disease (MRD) assessment has emerged as the most important prognostic and therapeutic biomarker for management of patients with multiple myeloma. Currently MRD is determined by a number of assays such as immunofixation, protein electrophoresis, flow cytometry. However these assays either lack sufficient sensitivity or require invasive bone marrow aspirates limiting clinical utility. To address these issues we developed a highly sensitive mass spectrometry based assay capable of detecting for patient’s clone specific unique immunoglobulin (Ig) peptides in small amounts of serum.

Methods

We used serum samples collected at diagnosis of multiple myeloma and after treatment, either classified as MRD+ or MRD- (MRD: Minimal Residual Disease). Variable regions of Ig light chain were sequenced by RACE PCR at diagnosis from neoplastic bone marrow plasma cells, and by De Novo sequencing from serum. To simplify the discovery and identification by LC-MS, we minimized the sample complexity using activated resins that can enrich for kappa or lambda light chains immunoglobulins prior to tryptic digestion.

Translational data (RACE-PCR) identified the unique peptides characteristic for the clone. Those unique peptides, specific for each patient, were successfully identified also by LC-MS in DDA mode and monitored after patient treatment in PRM mode.

LC: Ultimate 3000 nano-RSLC (Thermo)

MS: Q-Exactive Plus (Thermo)

Results

We detected constant and variable regions of immunoglobulins using both DDA and PRM approaches. Our results confirmed all peptides predicted by RACE-PCR in samples collected at the time of diagnosis, and also after treatment in most of the patients (all MRD+, and MRD- from patients who relapsed). Peptides detected by De Novo sequencing were also evaluated. The results indicated that our method is more sensitive than those currently used in the clinical setting.

Conclusions & Discussion

The clinical assay based on LC-MS analysis of peripheral blood serum described in this study overcomes the need for invasive bone marrow aspirates, and provides unprecedented sensitivity for monitoring for MRD in myeloma patients to help with individualized therapy.


References & Acknowledgements:


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