Session: 501. Hematopoietic Stem and Progenitor Cells and Hematopoiesis: Basic and Translational: Poster III
Hematology Disease Topics & Pathways:
Research, Translational Research
The rationale of the current research was to identify at diagnosis the leukemic HSPC clones that could promote AML progression, aiming to therapeutically target them at an early disease stage, thus preventing relapse. To that end, this study explored the genetic composition, chemoresistance propensity and leukemogenic potential of single HSPCs derived from AML patients.
Single leukemic cells (SLCs) obtained from AML patients at diagnosis and relapse were sorted into 4 HSPC subpopulations (HSC-MPP, MLP, CMP-MEP and GMP). Additionally, single HSPC-derived subclones were generated. Both SLCs and the subclones were analyzed for their chemoresistance, leukemia-associated mutations and cell lineage, using next generation sequencing, immunophenotyping, and lineage analyses with phylogenetic tree reconstruction. An NSG-mouse model was used in chemoresistance screening and functional assays to assess HSPC leukemogenic potential. HSPCs obtained from healthy donors (HD) were employed as control.
The most prevalent components of the AML-HSPC subpopulations identified at diagnosis were MLP33-/+-like cells of the lymphoid lineage and GMP33+-like cells of the myeloid linage, while HSC-MPP33-like cells appeared to be the major component of HD-HSPCs. Notably, either GMP-like (granulocyte/monocyte progenitor) or MLP-like (multipotent lymphoid progenitor) HSPC subpopulation was also shown to be most frequent in all the samples obtained at relapse, suggesting significant involvement of these HSPC subsets in AML progression.
Ex-vivo exposure of HSPC subpopulations to various concentrations of chemotherapy agents, cytarabine (Ara-C) or daunorubicin, displayed diverse response. The viability of the GMP33+-like fraction obtained at diagnosis was unaffected by these agents, irrespective of their concentration. Conversely, the diagnosis-derived MLP33+-like fraction was found to be chemo-sensitive. Moreover, the sensitivity of the latter cell fraction to these agents was different, with the cells derived from 2/10 AML patients being sensitive to Ara-C and not to daunorubicin, while cells obtained from another 2/10 patients demonstrated reverse results. In samples derived from additional 3/10 patients, GMP-like and MLP-like fractions appeared to be sensitive to both agents.
Phylogenetic tree reconstruction combined with single cell/subclone genetic analyses demonstrated 2 distinct clusters present at diagnosis and relapse in all the evaluated AML patients. In 2 patients, the same GMP-like and MLP-like subpopulations were detected both at diagnosis and relapse, suggesting their contribution to disease progression, while in an additional patient the relapse clearly originated from the MLP-like subpopulation only. The leukemogenic potential of chemoresistant GMP-like and MLP-like subpopulations was further confirmed in an NSG mouse model.
The comprehensive strategy, combining single-cell analysis and phylogenetic assessment of AML-HSPC subpopulations, employed in this study, enabled advanced functional and genomic profiling of leukemic HSPCs, that unraveled their inter- and intra-patient heterogeneity. The presence of GMP-like and/or MLP-like subpopulations at AML diagnosis, their identified chemoresistance propensity and occurrence at relapse, strongly point to the involvement of these HSPC fractions in relapse initiation. Overall, these findings highlight the need for personalized therapy targeting specific HSPC subsets as a therapeutic approach to prevent AML recurrence.
Disclosures: Zuckerman: Orgenesis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; BioSight Ltd: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Cellect Biotechnology: Honoraria, Speakers Bureau; Janssen: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau.
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