AUTHOR=Marsh-Wakefield Felix , Kruzins Annabel , McGuire Helen M. , Yang Shihong , Bryant Christian , Fazekas de St. Groth Barbara , Nassif Najah , Byrne Scott N. , Gibson John , Brown Christina , Larsen Stephen , McCulloch Derek , Boyle Richard , Clark Georgina , Joshua Douglas , Ho Phoebe Joy , Vuckovic Slavica 

TITLE=Mass Cytometry Discovers Two Discrete Subsets of CD39−Treg Which Discriminate MGUS From Multiple Myeloma

JOURNAL=Frontiers in Immunology

VOLUME=Volume 10 - 2019

YEAR=2019

URL=https://www.frontiersin.org/journals/immunology/articles/10.3389/fimmu.2019.01596

DOI=10.3389/fimmu.2019.01596

ISSN=1664-3224

ABSTRACT=Multiple Myeloma (MM) is preceded by the clinically stable condition monoclonal gammopathy of undetermined significance (MGUS), which is defined by the presence of a monoclonal protein and less than 10% malignant plasma cell infiltration in the bone marrow (BM). Critical immune events that discriminate MGUS from newly diagnosed MM (NDMM) patients remain unknown, but may involve changes in the regulatory T cell (Treg) compartment that favour myeloma growth. To address this possibility, we used mass cytometry to interrogate the distribution of multiple subsets within CD25+CD27low/negTreg in matched BM and peripheral blood (PB) of MGUS and NDMM patients. Both mass cytometry and flow cytometry confirmed a dominance of CD39-Treg within the Treg compartment in BM and PB of NDMM patients compared to an even distribution of CD39-Treg and CD39+Treg in MGUS patients. Unsupervised clustering displayed a phenotypic organisation of Treg into 25 metaclusters that confirmed Treg heterogeneity and identified two subsets which emerged exclusively within CD39-Treg of NDMM patients. One subset was found in both BM and PB that phenotypically resembled activated Treg based on CD45RO, CD49d, and CD62L expression; another subset resembled BM-resident Treg based on its tissue-resident phenotype characterised by CD69 expression, a lack of CD62L and CD49d expression and restricted location within the BM. PD-1 and TIGIT coexpressed on activated Treg, while PD-1 was expressed at higher levels on BM-resident Treg then on activated Treg. Both subsets produce Perforin and Granzyme B. In conclusion, the use of mass cytometry discovered two discrete subsets of CD39-Treg which may be permissive of myeloma growth. Understanding the regulatory properties of these subsets may also advance MGUS and MM diagnosis, prognosis, and therapeutic implications for MM patients.