Patterns of Individual Tumor Response Testing (ITRT) and Bone Marrow Minimal Residual Disease (MRD) On Day 15 of Therapy in Childhood Precursor-B-Lineage Acute Lymphoblastic Leukemia (ALL): MRD and Ex Vivo Drug Resistance Have Similar Predictive Value of Relapse.
Authors: Styczynski Jan , Jaworska-Posadzy Anna , Kubicka Malgorzata , Debski Robert , Kurylo-Rafinska Beata , Kolodziej Beata , Pogorzala Monika , Piatkowska Magdalena , Czyzewski Krzysztof , Wysocki Mariusz 
Published: October 2019

Abstract
Abstract 2448

Introduction:
The speed of blast clearance during therapy is a major prognostic factor of outcome in childhood acute lymphoblastic leukemia (ALL). Blast count in the peripheral blood on day 8, or in the bone marrow on day 15 and day 33, have been widely used to deliver risk-directed therapy. Another approach to measure the speed of leukemia clearance is the detection of minimal residual disease during induction therapy, as well as at days 33 and 78 of therapy. In vitro measurements of drug resistance (called recently as ITRT, individual tumor resistance testing) in leukemic cells obtained at diagnosis have been of prognostic significance in the prediction of clinical outcome in selected groups of patients.

Objective:
The analysis of the prognostic impact of (A) residual disease (MRD) at day 15 of induction therapy; (B) in vitro drug resistance at diagnosis (ITRT), (C) correlation of MRD and ITRT, and (D) multivariate analysis of prognostic role of MRD, ITRT, initial factors and initial therapy response to the risk of relapse.

Patients and Methods:
A total number of 87 children (aged 1–18 years) diagnosed for pre-B-ALL, treated either with ALL-BMF-90 or ALL-IC-2002 protocol were included into the study. ITRT was tested at diagnosis by the MTT assay. Residual disease at day 15 was measured by flow cytometry and determined for cut-off value BML15<0.5%. The median follow-up was 8.9 yrs (range, 0–11.5). Following drugs were tested: prednisolone, dexamethasone, vincristine, L-asparaginase, daunorubicin, doxorubicin, etoposide and cytarabine. PVA score was determined as combined ITRT profile to prednisolone, vincristine and L-asparaginase.

Results:
(A) The overall pDFS was 0.721±0.052 and the mean survival 9.1 yrs (95%CI=8.2–9.9). Patients with BML15<0.5% had pDFS=0.816±0.055, while those with BML15>0.5% had pDFS=0.542±0.098 (p=0.009, log-rank). The risk of relapse in BML15-positive patients was 3.0-fold higher (1.3–7.1, p=0.013). (B) pDFS was significantly better for patients with sensitive ITRT profile to: PVA (1.00±0.00 vs 0.61±0.06, p=0.002), prednisolone (0.89±0.05 vs 0.54±0.08, p=00002), vincristine (0.84±0.06 vs 0.61±0.08, p=0.035), daunorubicin (0.094±0.04 vs 0.51±0.08, p=0.00002), and L-asparaginase (0.84±0.06 vs 0.59±0.08, p=0.009). In multivariate analysis in Cox model, the prognostic value was retained only for ITRT for prednisolone (p=0.013, HR=0.08, 95%CI=0.01–0.6) and daunorubicin (p=0.004, HR=0.05, 95%CI=0.01–0.4), while ITRT for PVA score was below of significance (p=0.068, HR=0.03, 95%CI=0.01–1.3). (C) Patients with MRD-positive ALL at day 15 (BML15>0.5%) had higher ITRT for following drugs: doxorubicin (p=0.005, RR=1.8, Mann-Whitney U test), L-asparaginase (p=0.029, RR=3.2), and etoposide (p=0.055, RR=4.1), while no differences were found for other drugs. In multivariate logistic regression, the significance impact to development of BML15>0.5% was found for doxorubicin (p=0.035, OR=0.33) and etoposide (p=0.048, OR=0.14). (D) In multivariate analysis in Cox model for relapse risk, three factors had predictive value: BML15>0.5% (p=0.010, OR=3.3, 95%CI=1.3–8.2), ITRT for prednisolone (p=0.012, OR=4.4, 95%CI=1.4–13) and ITRT for daunorubicin (p=0.018, OR=5.9, 95%CI=1.4–26), while age, prednisolone-poor-response at day 8, BM response at day 15, BM response at day 33, and BCR-ABL rearrangement had no significant value.

Conclusions:
Patients with residual disease at day 15 had 3-fold higher risk of relapse. Patients with resistant ITRT profile to prednisolone and daunorubicin had respectively 12- and 20-fold higher risk of relapse. Presence of residual blasts at day 15 correlates with ITRT to etoposide and doxorubicin. Finally, persistence of blast in marrow at day 15 (BML15>0.5%) and ex vivo drug resistance (ITRT) to prednisolone and to daunorubicin were the strongest prognostic factors predicting relapse in childhood ALL.

Disclosures:
No relevant conflicts of interest to declare.