Repeated Bone Marrow Aspiration At the End of Induction Therapy: Implications for Treatment Stratification in Paediatric Acute Lymphoblastic Leukaemia
Authors: Zuna Jan , Arens Mari , Koehler Rolf , Panzer-Grümayer Renate , Möricke Anja , Bartram Claus R , Joas Ruth , Fronkova Eva , Cario Gunnar , Stanulla Martin , Zimmermann Martin , Trka Jan , Stary Jan , Attarbaschi Andishe , Mann Georg , Schrappe Martin , Schrauder André 
Published: October 2019

Abstract
Abstract 756FN2
Bone marrow (BM) aspiration at the end of induction therapy plays a crucial role for the evaluation of remission and the minimal residual disease (MRD), both critical for treatment stratification in modern treatment protocols for paediatric acute lymphoblastic leukaemia (ALL). However, the aspiration is repeated in 15–20% of patients, either due to non-representative morphology or to insufficient material needed for MRD analysis.
We prospectively analysed 320 paediatric ALL patients treated according to ALL-BFM 2000 (n=301) or ALL IC-BFM 2002 (n=19) protocols with repeated BM aspiration at the end of induction therapy, on treatment day 33. Fourteen patients had more than one re-puncture. The median follow-up was 69 months, 45 (14%) patients had an event (relapse/death). The cause for the repeated BM aspiration was non-representative morphology (32%), insufficient material for MRD analysis (33%) or both (35% cases). In order to evaluate prognostic significance of the re-punctures and to determine which of the repeated samples should be used for the final treatment stratification we analysed MRD levels and MRD stratification, morphology, leukocyte count (WBC) and the length of treatment delay caused by waiting for the repeated aspiration. MRD data were collected and interpreted according to the EuroMRD guidelines in one central reference laboratory per each participating country. Morphology was evaluated centrally using an own scoring system (with a max value of 26 points).
Treatment delay between the original and the last aspiration was one-third longer in patients with subsequent event compared to patients remaining in complete remission (CR) (median 8 (range 2 – 21) vs. 6 (1 - 28) days, respectively; p=0.020). Patients with a subsequent event had significantly higher WBC at the time of the last repeated BM aspiration, compared to patients without event (p=0.019), while there was no difference relative to the original aspiration (p=0.9).
Analysis of the BM morphology at the original aspiration showed no significant difference between patients with an event vs. those in CR. However, the repeated aspiration of patients with a subsequent event had significantly better morphology (median 18.5/26 vs. 15/26 points, p=0.0012) mainly due to higher cellularity (p=0.003) and number of megakaryocytes (p=0.048).
MRD levels were identical or decreased in 88% and increased in 12% of cases comparing the original aspiration to the repeated aspiration. In 63 patients (20%) the different MRD levels would lead to different treatment stratification. Higher MRD was associated with treatment failure; the best predictive values for subsequent event were obtained using the MRD results of the original aspiration (p=3.1e-07) or the highest of the detected MRD levels (p=6.0e-07). The last aspiration before proceeding with treatment had the lowest, though still a highly significant predictive value (p=8.6e-06). Corresponding results are obtained when MRD levels are substituted by final MRD risk stratification into standard, medium or high risk (p<0.0001, p=0.0005 and p=0.0008 for the prediction of treatment failure using the MRD level in the original, the highest and the last aspiration, respectively).
In conclusion, our data show that the original BM aspiration – independently of the quality of morphology – is not inferior for MRD treatment stratification, and that it actually has the best predictive value. In cases where sample quality precludes MRD analysis, the repeated sample with the highest MRD level should be used for stratification in order to not underestimate the putative risk of treatment failure.
Longer treatment delay caused by waiting for a more representative sample seems to worsen the outcome. Notion that patients with a subsequent event need more time for BM regeneration is not justified, as their cellularity, overall morphology and also WBC before proceeding with treatment are better than in patients remaining in long-term CR.
Any decision to perform a re-puncture at the end of induction therapy due to a non-representative morphology should be critically weighed. If possible, any unnecessary prolongation of treatment delay should be avoided unless being inevitable for other reasons, and therapy should be continued as soon as possible.

Support:
Deutsche Krebshilfe, Germany (Projects 50–2698 Schr1; 50–2722 BA6/7); St. Anna Kinderkrebsforschung, Austria; MSM0021620813; IGA NS/1000-4.

Disclosures:
No relevant conflicts of interest to declare.