Combining Measurements of Plasma Cell Apoptosis and Proliferation in Multiple Myeloma Identifies Patients with Poor Survival.
Authors: Kumar Shaji , Timm Michael , Lacy Martha Q. , Dispenzieri Angela , Hayman Suzanne R. , Gertz Morie A. , Rajkumar S. Vincent , Greipp Philip R. , Witzig Thomas E. 
Published: April 2020

Abstract
Background: Multiple Myeloma (MM) is characterized by accumulation of malignant plasma cells with relatively low proliferative and apoptotic rates. The proliferative rate of plasma cells, measured in terms of % plasma cells labeled by bromodeoxyuridine (BrdU) using slide based fluorescence microscopy (plasma cell labeling index or PCLI) has been shown to be a powerful prognostic factor in MM. We have previously shown that the % bone marrow plasma cells in apoptosis (PCAP) determined by a flow cytometry method that uses Annexin V staining or 7-amino Actinomycin D (7-AAD) correlates with disease stage in MM. The goal of this study was to examine the prognostic value of the PCAP.
Methods: We studied 145 patients with MM including 74 with newly diagnosed MM, 40 with relapsed MM, and 31 with MM on treatment in whom simultaneous measurements of % plasma cell apoptosis and PCLI were available. The PCLI is expressed as the percentage of cytoplasmic immunoglobulin positive plasma cells that have taken up BrdU. The PCLI was characterized as high when &gt;= 1%. Mononuclear cells isolated from bone marrow aspirate, following red cell lysis with ACK solution, are incubated with CD38-PE and either CD138-FITC or CD45-FITC. This is followed by incubation with 7-AAD and the plasma cells are gated using the CD38/CD45 staining pattern. The Annexin method used a similar strategy, with Annexin V in place of 7-AAD. The percentage of plasma cells in apoptosis (PCAP) was categorized as high when &gt;5%, the median value.
Results: The median follow up for the entire group was 34 months (range, 0.1 mo to 9 years) and 112 (77%) had died at the time of analysis. Forty seven patients (32%) had a high PCLI (&gt;=1%). The median overall survival for the patients with high PCLI was 14.6 months versus 42.3 months for those with a low PCLI; P = 0.0005 (log rank test). Seventy-five (51.7%) patients had a high PCAP. The median overall survival for those patients was 28.9 months versus 40.2 months for those with a low PCAP; P = 0.078. We next examined if there was any correlation between a high PCAP and high PCLI and found none; P = NS by (Fisher’s exact test). Additionally in a multivariate Cox model using both PCLI and PCAP, both were independently prognostic for overall survival. We then developed an index by adding PCLI to log transformed PCAP (PCLI + log PCAP). Using a cutoff of 1.5 for the new index we were able identify a group of patients with poor survival. The 59 patients with a high value for the new index had a median survival of 19.3 months vs 46.1 months for the rest (P &lt; 0.0001, log rank test).
Conclusion: The measurement of plasma cell apoptosis has prognostic value in patients with MM. More importantly, we have shown that by combining measures of two important biological characteristics of the plasma cell, namely proliferation and apoptosis, patients with a poor prognosis can be identified. Such a prognostication strategy can help stratify patients in clinical trials as well as shed light on the disease biology.