Fast-Track Measurable Residual Disease Detection By Multiparametric Flow Cytometry in Acute Myeloid Leukemia before Allogeneic Hematopoietic Cell Transplantation

Introduction

In acute myeloid leukemia (AML), measurable residual disease (MRD) becomes more and more important for individualized treatment decisions. The MRD status at the time of allogeneic hematopoietic cell transplantation (alloHCT) provides prognostic information beyond baseline risk features. Peri-transplant treatment strategies and conditioning intensity can be stratified according MRD-levels. However, therapeutic interventions (e.g. conditioning intensity) usually require the prompt availability of reliable information on the MRD status. We assessed the prognostic value of a standardized, reproducible, and fast turnover multiparametric flow cytometry (MFC) approach that has been validated at the end of intensive induction therapy to monitor MRD before alloHCT (Röhnert, Leukemia 2022). This approach allows for the simultaneous detection of MRD by the leukemia-associated immunophenotype (LAIP) and the different from normal (DfN) method.

Methods

Between October 2016 and March 2020, bone marrow (BM) aspirates of patients (pts) with AML were analyzed for the presence of MRD immediately before the start of conditioning. therapy with the aim of analyzing to predictive value of this fast-track parameter. Pts receiving sequential regimens (re-induction chemotherapy followed by reduced-intensity conditioning) were excluded.

Briefly, the LAIP-based DfN analysis is based on a hierarchical gating strategy with fixed gates and encompasses the antigens CD34, CD117, HLA-DR (backbone markers), CD13, CD33 (myeloid markers), CD7 and CD56 (lymphoid markers). Within less than 5 minutes, the analysis checks for the presence of the aberrant categories i) deficiency of myeloid markers and ii) cross-lineage expression of lymphoid markers. Reference values for 32 immunophenotypically defined populations were established on BM aspirates of 90 leukemia free controls. An aberrant category already detectable at diagnosis on ≥10% of myeloid progenitors/ monocytes was termed aberrant LAIP (aLAIP) and a de-novo appearance (after chemotherapy) post-treatment DfN (ptDfN). In conclusion this approach is very fast and provides high reproducibility.

Results

We identified 58 pts with a median follow-up of 33.3 months after alloHCT (Table 1 with pts characteristics). Response assessment was performed in median 8 days before alloHCT. Most of the patients were in morphologic complete remission (CR n=37). A subset of 21 patients had refractory disease with a median BM blast count of 6.5%.

Samples at AML diagnosis were available for 48/58 (83%) pts, of whom 34/48 (71%) showed at least one aLAIP at initial diagnosis. Before start of conditioning, 44/58 (76%) patients were MRD^pos. The proportion of MRD^pos in responders was lower compared to non-responders: 24/37 (65%) vs. 20/21 (95%) (p=0.0106). MRD^pos was designated by aLAIP^only, ptDfN^only and aLAIP+ptDfN in 18%, 48% and 34% of cases, respectively. Noteworthy, 8 of 21 pts classified MRD^pos by ptDfN^only had no sample available at diagnosis and therefore could not be assessed for aLAIP persistence.

Amongst 37 pts in CR before alloHCT, 12/24 (50%) of MRD^pos pts relapsed compared to 3/13 (23%) of MRD^neg pts. TRM in these patients did not differ (13% vs. 8%).

In a univariable Cox regression analysis, the 1-year Overall survival (OS) rate of MRD^pos cohort was significantly shorter compared to MRD^neg pts (64% vs. 93%, Hazard ratio (HR) 10.5, p=0.004). MRD was detected by ptDfN^only in 23/44 (52%) of MRD^pos pts. The OS of these pts was comparable to MRD^pos by aLAIP with or without ptDfN (Figure 1). As expected, the MRD^pos cohort was enriched for non-responders. However, when the analysis was restricted to pts in CR (n=37), MRD status preserved its prognostic significance (HR 7.9, p=0.019). And even in the subpopulation of pts in first CR, MRD^pos was associated with a significantly reduced OS compared to MRD^neg (HR 6.3, p=0.043). Interestingly, a single non-responder was classified as MRD^neg and showed a long-term OS.

Conclusions

Our fast-track LAIP-based DfN approach provides relevant prognostic information prior to conditioning for allo HCT. On top of the classical LAIP approach, the ptDfN analysis gives additional prognostic information. The LAIP-based DfN approach might be a suitable tool for fast-track and reliable MRD assessment before starting the preparative regimen for alloHCT and might help to guide potential treatment modifications.

Röllig:AbbVie Inc.: Consultancy, Honoraria, Research Funding; Amgen Inc.: Honoraria; Bristol Myers Squibb: Consultancy, Honoraria; Celgene Corporation: Consultancy, Honoraria; Jazz Pharmaceuticals: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Research Funding; Pfizer Inc.: Consultancy, Honoraria, Research Funding; F. Hoffmann-La Roche Ltd: Consultancy, Honoraria; Servier: Consultancy. Schetelig:Abbvie: Honoraria, Membership on an entity's Board of Directors or advisory committees.