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Mass cytometry analysis shows that a novel memory phenotype B cell is expanded in multiple myeloma.

ABSTRACT: It would be very beneficial if the status of cancers could be determined from a blood specimen. However, peripheral blood leukocytes are very heterogeneous between individuals, and thus high-resolution technologies are likely required. We used cytometry by time-of-flight and next-generation sequencing to ask whether a plasma cell cancer (multiple myeloma) and related precancerous states had any consistent effect on the peripheral blood mononuclear cell phenotypes of patients. Analysis of peripheral blood samples from 13 cancer patients, 9 precancer patients, and 9 healthy individuals revealed significant differences in the frequencies of the T-cell, B-cell, and natural killer-cell compartments. Most strikingly, we identified a novel B-cell population that normally accounts for 4.0% ± 0.7% (mean ± SD) of total B cells and is up to 13-fold expanded in multiple myeloma patients with active disease. This population expressed markers previously associated with both memory (CD27(+)) and naïve (CD24(lo)CD38(+)) phenotypes. Single-cell immunoglobulin gene sequencing showed polyclonality, indicating that these cells are not precursors to the myeloma, and somatic mutations, a characteristic of memory cells. SYK, ERK, and p38 phosphorylation responses, and the fact that most of these cells expressed isotypes other than IgM or IgD, confirmed the memory character of this population, defining it as a novel type of memory B cells.

SUBMITTER: Hansmann L

PROVIDER: S-EPMC4457663 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Mass cytometry analysis shows that a novel memory phenotype B cell is expanded in multiple myeloma.

Hansmann Leo L Blum Lisa L Ju Chia-Hsin CH Liedtke Michaela M Robinson William H WH Davis Mark M MM

Cancer immunology research 20150220 6

It would be very beneficial if the status of cancers could be determined from a blood specimen. However, peripheral blood leukocytes are very heterogeneous between individuals, and thus high-resolution technologies are likely required. We used cytometry by time-of-flight and next-generation sequencing to ask whether a plasma cell cancer (multiple myeloma) and related precancerous states had any consistent effect on the peripheral blood mononuclear cell phenotypes of patients. Analysis of periphe ...[more]

PMID: 25711758

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Project description:Multiple Myeloma (MM) is preceded by the clinically stable condition monoclonal gammopathy of undetermined significance (MGUS). Critical immune events that discriminate MGUS from newly diagnosed MM (ND)MM patients remain unknown, but may involve changes in the regulatory T cell (Treg) compartment that favor myeloma growth. To address this possibility, we used mass cytometry and the unsupervised clustering algorithm Flow self-organizing map (FlowSOM) to interrogate the distribution of multiple subsets within CD25+CD127low/negTreg in matched bone marrow (BM) and peripheral blood (PB) of MGUS and NDMM patients. Both mass cytometry and flow cytometry confirmed a trend toward prevalence of CD39-Treg within the Treg compartment in BM and PB of NDMM patients compared to CD39-Treg in MGUS patients. FlowSOM clustering displayed a phenotypic organization of Treg into 25 metaclusters that confirmed Treg heterogeneity. It identified two subsets which emerged within CD39-Treg of NDMM patients that were negligible or absent in CD39-Treg of MGUS patients. One subset was found in both BM and PB which phenotypically resembled activated Treg based on CD45RO, CD49d, and CD62L expression; another subset resembled BM-resident Treg based on its tissue-resident CD69+CD62L-CD49d- phenotype and restricted location within the BM. Both subsets co-expressed PD-1 and TIGIT, but PD-1 was expressed at higher levels on BM-resident Treg than on activated Treg. Within BM, both subsets had limited Perforin and Granzyme B production, whilst activated Treg in PB acquired high Perforin and Granzyme B production. In conclusion, the use of mass cytometry and FlowSOM clustering discovered two discrete subsets of CD39-Treg which are discordant in MGUS and NDMM patients and may be permissive of myeloma growth which warrants further study. Understanding the regulatory properties of these subsets may also advance MGUS and MM diagnosis, prognosis, and therapeutic implications for MM patients.

Project description:Daratumumab is a CD38-targeted human monoclonal antibody with direct anti-myeloma cell mechanisms of action. Flow cytometry in relapsed and/or refractory multiple myeloma (RRMM) patients treated with daratumumab revealed cytotoxic T-cell expansion and reduction of immune-suppressive populations, suggesting immune modulation as an additional mechanism of action. Here, we performed an in-depth analysis of the effects of daratumumab on immune-cell subpopulations using high-dimensional mass cytometry. Whole-blood and bone-marrow baseline and on-treatment samples from RRMM patients who participated in daratumumab monotherapy studies (SIRIUS and GEN501) were evaluated with high-throughput immunophenotyping. In daratumumab-treated patients, the intensity of CD38 marker expression decreased on many immune cells in SIRIUS whole-blood samples. Natural killer (NK) cells were depleted with daratumumab, with remaining NK cells showing increased CD69 and CD127, decreased CD45RA, and trends for increased CD25, CD27, and CD137 and decreased granzyme B. Immune-suppressive population depletion paralleled previous findings, and a newly observed reduction in CD38+ basophils was seen in patients who received monotherapy. After 2 months of daratumumab, the T-cell population in whole-blood samples from responders shifted to a CD8 prevalence with higher granzyme B positivity (P = 0.017), suggesting increased killing capacity and supporting monotherapy-induced CD8+ T-cell activation. High-throughput cytometry immune profiling confirms and builds upon previous flow cytometry data, including comparable CD38 marker intensity on plasma cells, NK cells, monocytes, and B/T cells. Interestingly, a shift toward cytolytic granzyme B+ T cells was also observed and supports adaptive responses in patients that may contribute to depth of response. © 2018 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of International Society for Advancement of Cytometry.

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Dataset Information

Mass cytometry analysis shows that a novel memory phenotype B cell is expanded in multiple myeloma.

Publications

Mass cytometry analysis shows that a novel memory phenotype B cell is expanded in multiple myeloma.

Similar Datasets

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