High prognostic value of measurable residual disease detection by flow cytometry in chronic lymphocytic leukemia patients treated with front-line fludarabine, cyclophosphamide, and rituximab, followed by three years of rituximab maintenance

Abstract

It has been postulated that monitoring measurable residual disease (MRD) could be used as a surrogate marker of progression-free survival (PFS) in chronic lymphocytic leukemia (CLL) patients after treatment with immunochemotherapy regimens. In this study, we analyzed the outcome of 84 patients at 3 years of follow-up after first-line treatment with fludarabine, cyclophosphamide and rituximab (FCR) induction followed by 36 months of rituximab maintenance thearpy. MRD was assessed by a quantitative four-color flow cytometry panel with a sensitivity level of 10−4. Eighty out of 84 evaluable patients (95.2%) achieved at least a partial response or better at the end of induction. After clinical evaluation, 74 patients went into rituximab maintenance and the primary endpoint was assessed in the final analysis at 3 years of follow-up. Bone marrow (BM) MRD analysis was performed after the last planned induction course and every 6 months in cases with detectable residual disease during the 36 months of maintenance therapy. Thirty-seven patients (44%) did not have detectable residual disease in the BM prior to maintenance therapy. Interestingly, 29 patients with detectable residual disease in the BM after induction no longer had detectable disease in the BM following maintenance therapy. After a median followup of 6.30 years, the median overall survival (OS) and PFS had not been reached in patients with either undetectable or detectable residual disease in the BM, who had achieved a complete response at the time of starting maintenance therapy. Interestingly, univariate analysis showed that after rituximab maintenance OS was not affected by IGHV status (mutated vs. unmutated OS: 85.7% alive at 7.2 years vs. 79.6% alive at 7.3 years, respectively). As per protocol, 15 patients (17.8%), who achieved a complete response and undetectable peripheral blood and BM residual disease after four courses of induction, were allowed to stop fludarabine and cyclophosphamide and complete two additional courses of rituximab and continue with maintenance therapy for 18 cycles. Surprisingly, the outcome in this population was similar to that observed in patients who received the full six cycles of the induction regimen. These data show that, compared to historic controls, patients treated with FCR followed by rituximab maintenance have high-quality responses with fewer relapses and improved OS. The tolerability of this regime is favorable. Furthermore, attaining an early undetectable residual disease status could shorten the duration of chemoimmunotherapy, reducing toxicities and preventing long-term side effects. The analysis of BM MRD after fludarabine-based induction could be a powerful predictor of post-maintenance outcomes in patients with CLL undergoing rituximab maintenance and could be a valuable tool to identify patients at high risk of relapse, influencing further treatment strategies. This trial is registered with EudraCT n. 2007-002733-36 and ClinicalTrials.gov Identifier: NCT00545714.

Genomic mutation profile in progressive chronic lymphocytic leukemia patients prior to first-line chemoimmunotherapy with FCR and rituximab maintenance (REM)

“…The patients were enrolled in the REM clinical trial. REM is a multicenter, non-randomized, prospective phase II clinical trial evaluating the overall response and PFS in patients with CLL with active progressive disease after first-line treatment with FCR, followed by rituximab maintenance every two months for three years in responding patients [15]. Samples were collected at the time of enrollment before treatment.…”

“…Samples were stained and lysed using a direct immunofluorescence technique as previously described [15]. In summary, sequential bone marrow (BM) and peripheral blood (PB) samples were collected in tubes containing K3 EDTA as anticoagulant.…”

“…immunofluorescence technique, as previously described [15]. The antibody combinations tested were CD22/CD23/CD19/CD5, CD81/CD22/CD19/CD5, CD20/CD38/CD19/CD5, CD20/CD79b/CD19/CD5 and sIgKappa/sIgLambda/CD19/CD5.…”

“…To ensure a lower limit of quantification of 0.01%, a minimum of 50 events were required. For ZAP70, CD38 and CD49d measurements see García-Marco et al 2019 [15].…”

“…In this study, we have taken a deep-targeted massive sequencing approach to analyze the impact of mutations in the most frequently mutated genes in a prospectively selected group of patients with CLL with active progressive disease who require treatment. All patients were enrolled in the REM (Rituximab En Mantenimiento [Rituximab in Maintenance]) clinical trial, which consisted of rituximab maintenance for 36 months after achieving at least a partial clinical response to front-line FCR treatment [15].…”