Fc receptor-like 2 (FCRL2) is a novel marker of low-risk CLL and refines prognostication based on IGHV mutation status
Authors: Lauren K. Shea, Kazuhito Honjo, David T. Redden, Edlue Tabengwa, Ran Li, Fu-Jun Li, Mikhail Shakhmatov, Nicholas Chiorazzi, Randall S. Davis
Journal: Blood Cancer Journal (2019)

Chronic lymphocytic leukemia (CLL) is the most prevalent leukemia in Western countries and is characterized by a progressive accumulation of CD5 + monoclonal B cells in the blood and bone marrow.The clinical course in CLL is remarkably heterogeneous; the monoclonal population may expand in a protracted fashion over many years or increase rapidly, leading to bulky infiltration of lymphoid organs, progressive cellular and humoral immunodeficiency, autoimmunity, and cytopenias 1,2 .Prognostication at the time of diagnosis has important implications for patient care and has been an active area of investigation over the past several decades.Various means of risk stratification have been developed and validated, including clinical stage 3,4 , IGHV mutation status 5 , cytogenetic abnormalities 6 , and expression of surface proteins such as CD38 5,7 and CD49d 8 .Composite prognostic indices, such as the German CLL Study Group prognostic score and the CLL-IPI, have also been proposed 9,10 .Despite an abundance of validated prognostic indicators, decisions to initiate treatment are still based on the presence of symptoms and/or cytopenias, and there is no evidence yet that preemptive treatment based on adverse prognostic factors improves overall survival (OS).Therefore, the search for reliable and widely available indicators that can be easily assayed at diagnosis and integrated to refine prognostication in CLL is ongoing.Our own studies have focused on a family of Fc receptor-like (FCRL1-5) molecules that have tyrosine-based signaling potential and preferential expression by B lymphocytes 11 .In healthy donor peripheral blood mononuclear cell samples, the family member designated FCRL2 identifies subsets of CD19 + CD27 + memory B cells and inhibits activation when co-ligated with the B-cell receptor, but has not been found to bind Ig 12 .Interestingly, in a series of 107 cryopreserved CLL samples, FCRL2 emerged as a powerful prognostic indicator 13 .FCRL2 expression was significantly elevated on CLL samples with mutated (M-CLL) as compared to unmutated (U-CLL) IGHV sequences, showed 94.4% concordance with IGHV mutation status, and was superior to nine other factors (including IGHV status) analyzed in predicting time to first treatment (TFT).Extended followup of our original 107-patient cohort showed that FCRL2 expression was predictive of OS as well as TFT (Fig. 1a).As expected, IGHV mutation status was also predictive of OS in our cohort.These encouraging results prompted us to pursue optimization of assaying FCRL2 expression to further its potential clinical use.We first set out to develop an FCRL2-specific monoclonal antibody (mAb) with superior reactivity and binding properties relative to our prior clone 7F2 13 .After institutional IACUC approval, BALB/c mice were hyperimmunized with BW5147 mouse T cell line retroviral transductants expressing FCRL2.Two separate fusions were performed and an extensive analysis of 92 FCRL2reactive hybridoma clones was undertaken.This screen led to the identification of 3E11, a mouse IgG1κ subclone that does not cross-react with the five other FCRL molecules expressed by human lymphocytes (Fig. S1).Blocking experiments indicated that 7F2 and 3E11 bind distinct epitopes on FCRL2 (Fig. S2).To pursue further characterization of the 3E11 clone, we directly conjugated