Minimal residual disease monitoring and immune profiling using second generation flow cytometry in elderly multiple myeloma Running head: 2 generation flow MRD monitoring in elderly MM

The value of minimal residual disease (MRD) in multiple myeloma (MM) has been more frequently investigated in transplant-eligible than elderly patients. Since an optimal balance between treatment efficacy and toxicity is of utmost importance in elderly MM, sensitive MRD monitoring might be particularly valuable in this patient population. Here, we used 2 generation 8-color multiparameter-flow-cytometry (MFC) to monitor MRD in 162 transplant-ineligible MM patients enrolled in the PETHEMA/GEM2010MAS65 study, The transition from 1 to 2 generation MFC resulted in increased sensitivity, and allowed to identify three patient groups according to MRD levels: MRD-negative (<10; n=54, 34%), MRD-positive between <10 and ≥10 (n=20, 12%), and MRD-positive ≥10 (n=88, 54%). MRD status was an independent prognostic factor for time-to progression (-TTPHR:2.7; P=.007) and overall survival (-OSHR:3.1; P=.04) with significant benefit for MRD-negative patients (median TTP not reached, 70% OS at 3-years), and similar poorer outcomes for cases with MRD levels between <10 and ≥10 vs ≥10 (both median TTP of 15 months; 63% and 55% OS at 3-years). Furthermore, MRD-negativity significantly improved TTP of patients >75-years (HR:4.8; P<.001), and those with high-risk cytogenetics (HR:12.6; P=.01). Using 2 generation MFC, immune profiling concomitant to MRD monitoring also contributed to identify patients with poor, intermediate and favorable outcome (25%, 61% and 100% OS at 3-years; P=.01); the later patients being characterized by an increased compartment of mature B-cells. Our results show that similarly to transplant-candidates, MRD monitoring is one of the most relevant prognostic factors in elderly MM, irrespectively of patients’ age and cytogenetic risk. The trial is registered within ClinicalTrials.gov (number NCT01237249): https://clinicaltrials.gov/ct2/show/NCT01237249