PB1660 FLOW CYTOMETRY CHALLENGE FOR MEASURABLE RESIDUAL DISEASE MONITORING IN B-ACUTE LYMPHOBLASTIC LEUKEMIA (B-ALL) PATIENTS RECEIVING TARGETED IMMUNOTHERAPY
Authors: A. Yeguas Bermejo, A. Martín García-Sancho, N. Puig Morón, M. P. Leoz Allegretti, V. Higuero Saavedra, I. Aires Mejía, C. Casquero Hernández, L. López Corral, E. Pérez López, A. Á. Martín López, F. López Cadenas, A. Redondo Guijo, Á. Veiga Vaz, E. Sobejano Fuertes, M. Fonseca Santos, D. Presa Morales, R. Azibeiro Melchor, D. Rivera Delgado, J. M. Hernández-Rivas, M. González Díaz, M. B. Vidriales Vicente
Journal: HemaSphere (2019)

Background: Targeted immunotherapy (TI) with Blinatumomab (Bln) or Inotuzumab ozogamicin (IO) are a salvage option for the management of B-ALL patients with measurable residual disease (MRD), in which high rates of MRD negative can be achieved, allowing a bridge to transplantation and longer survival. However, some patients relapse with a phenotype lacking CD19 or CD22 expression can be found in blast cells. Those escape variants have also been described under CAR-T cells therapy, and different mechanism of escape have been proposed. This resistance mechanism has to be considered in flow cytometry immunophenotyping (FCM) analysis when monitoring MRD. Aims: Review the MRD monitoring by FCM in patients diagnosed of B-LLA treated with TI in our center that experience relapse. Methods: Two B-ALL patients treated in our center with TI experienced relapse. The FCM monitoring were performed in BMA samples, using 8-colour combinations of monoclonal antibodies adapted to the TI received and diagnosis phenotype. Stained cells were measured in FACSCanto II cytometer equipped with the FACSDiva software and analyzed with the Infinicyt software with a sensitivity (S) 10-4. Results: Between 2015 and 2018, 2 B-LLA patients relapsed after TI. One of them relapsed with the same phenotype previously detected, and the other patient experienced two relapses, the first one after Bln and then after IO. Interestingly, both relapses showed immunophenotypic changes (figure). The patient was a 72 years old woman that initially received treatment according to PETHEMA-LAL-OLD-07 protocol. Bone marrow aspiration (BMA) at the end of induction was in morphological complete response (CR) with MRD positive (0.12%). After 3rd consolidation cycle, MRD persisted (0.04%), so she started Bln and achieved MRD negative (S 10-4) after 1st cycle. However, after the 3rd cycle, a new CD19- blast cell population was detected (0.07%), with a lower expression of CD20, being the remaining markers similar to the diagnostic. In normal B cells (0.05%) the intensity of CD19 expression was preserved. Blast cells increased after 4th cycle (2.9%) with an associated morphological relapse (21%). She started IO treatment and reached CR and MRD negative (S 10-4) after 1st cycle. Treatment was stopped after the 4th cycle due to upper gastrointestinal bleeding, and in the BMA previous to the next cycle programmed, CD19- CD20dim CD34+ blast cells were detected again (0.55%). All blasts lost CD22 expression, and only 9% maintained the CD10 expression detected previously, being the majority CD10 negative. The patient was not candidate to any other therapy, she started palliative treatment and died 2 months later.Summary/Conclusion: In our experience, changes in immunophenotype of B-ALL blast cells are frequent after TI, due to loss of CD19 and CD22 expression. Therefore, FCM analysis in patients treated with TI demands a high degree of expertise and attention, to avoid the loss of detection of new populations that could emerge with different phenotype.