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46 Erythroid/Megakaryocytic Differentiation Confers BCL-XL Dependency and Venetoclax Resistance in Acute Myeloid Leukemia

Program: Oral and Poster Abstracts
Type: Oral
Session: 604. Molecular Pharmacology and Drug Resistance: Myeloid Neoplasms: Therapeutic Vulnerabilities Triggering Mechanisms of Drug Resistance and Sensitivity in Myeloid Neoplasms
Hematology Disease Topics & Pathways:
Research, Acute Myeloid Malignancies, AML, apoptosis, Translational Research, Non-Biological therapies, Diseases, Therapies, Myeloid Malignancies, Biological Processes
Saturday, December 10, 2022: 10:15 AM

Olli Dufva1*, Heikki Kuusanmäki, PhD2,3, Markus Vaha-Koskela, PhD4*, Aino-Maija Leppä, MSc(Tech)5*, Jani Huuhtanen6,7*, Ida Vänttinen, MSc2*, Petra Johanna Nygren1*, Jay Klievink, MSc1*, Jonas Bouhlal, BSc6*, Petri Pölönen, PhD8*, Qi Zhang, PhD9*, Shady Adnan Awad, MD, PhD6*, Cristina Mancebo10*, Joseph Saad, MSc5*, Juho J. Miettinen, PhD11*, Komal Kumar Javarappa, PhD12, Sofia Aakko13*, Tanja Ruokoranta, MSc2*, Samuli Eldfors, PhD5, Merja Heinäniemi, PhD14*, Ulla Wartiovaara-Kautto, MD, PhD15, Marina Konopleva, MD, PhD16, Mikko A Keränen, MD, PhD17*, Kimmo Porkka, MD, PhD15*, Krister Wennerberg18,19*, Mika Kontro, MD, PhD20*, Caroline A. Heckman, PhD5 and Satu Mustjoki, MD, PhD1

1Hematology Research Unit Helsinki, University of Helsinki and Helsinki University Hospital Comprehensive Cancer Center, Helsinki, Finland
2Institute for Molecular Medicine Finland (FIMM), HiLIFE, University of Helsinki, Helsinki, Finland
3Biotech Research & Innovation Centre (BRIC), University of Copenhagen, Copenhagen, Denmark
4Institute for Molecular Medicine Finland, Helsinki Institute of Life Science, University of Helsinki, Finland, Helsinki, Finland
5Institute for Molecular Medicine Finland (FIMM), Helsinki Institute of Life Science, University of Helsinki, Helsinki, Finland
6Hematology Research Unit Helsinki, Department of Hematology, University of Helsinki and Helsinki University Hospital Comprehensive Cancer Center, Helsinki, Finland
7Department of Computer Science, Aalto University School of Science, Espoo, Finland
8Department of Pathology, St. Jude Children's Research Hospital, Memphis, TN
9Department of Leukemia, University of Texas M.D. Anderson Cancer Center, Houston, TX
10Institute for Molecular Medicine Finland, Helsinki Institute of Life Science, Helsinki, Finland
11University of Helsinki, Institute for Molecular Medicine Finland (FIMM), Helsinki Institute of Life Science, Helsinki, Finland
12Institute for Molecular Medicine Finland (FIMM), Helsinki Institute of Life Science, University of Helsinki, Helsinki, Helsinki, Finland
13Institute of Molecular Medicine Finland (FIMM), Helsinki, Finland
14Institute of Biomedicine, School of Medicine, University of Eastern Finland, Kuopio, Finland
15Department of Hematology, Helsinki University Hospital Comprehensive Cancer Center and University of Helsinki, Helsinki, Finland
16Department of Leukemia, The University of Texas M.D. Anderson Cancer Center, Houston, TX
17Department of Hematology, Helsinki University Hospital Comprehensive Cancer Center, University of Helsinki, Helsinki, Finland
18Institute for Molecular Medicine Finland (FIMM), University of Helsinki, Helsinki, Finland
19Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Copenhagen, Denmark
20Hematology Research Unit Helsinki, University of Helsinki and Department of Hematology, Helsinki University Hospital Comprehensive Cancer Centre, Helsinki University Hospital and University of Helsinki, Helsinki, Finland

Myeloid neoplasms with erythroid or megakaryocytic differentiation include pure erythroid leukemia (PEL), myelodysplastic syndrome (MDS) with erythroid features, and acute megakaryoblastic leukemia (FAB M7) and are characterized by poor prognosis and limited treatment options. Hypothesizing that the distinct lineage of acute myeloid leukemia (AML) with erythroid or megakaryocytic differentiation may confer specific vulnerabilities, we sought to identify selective dependencies in these rare subtypes in comparison with other AML types. We show that AML cells with erythroid or megakaryocytic differentiation depend on the anti-apoptotic protein BCL-XL, rather than BCL-2, using a combination of ex vivo drug sensitivity and resistance testing, genetic perturbation, transcriptomic profiling and mouse xenograft experiments.

To identify druggable dependencies in erythroid and megakaryoblastic leukemias, we screened over 500 drugs across a 10,000-fold concentration range in four erythroid and two megakaryoblastic AML cell lines and used other AML cell lines as controls (Figure A). The high-throughput screens identified BCL-XL-selective inhibitors including A-1331852 as highly effective against erythroid and megakaryoblastic leukemia cell lines in comparison with 15 cell lines representing other AML types (Figure B). In contrast, the erythroid/megakaryoblastic AML subtypes were resistant to the BCL-2 inhibitor venetoclax used clinically in the treatment of AML (Figure A).

To confirm the on-target mechanism of BCL-XL inhibition, we next evaluated the genetic dependencies of erythroid and megakaryoblastic leukemias using public genome-scale CRISPR-Cas9 and RNAi screening data from 5 erythroid, 3 megakaryoblastic, and 15 other AML cell lines. This analysis demonstrated striking essentiality of BCL2L1 encoding BCL-XL, but not BCL2 or MCL1, for the survival of erythroid and megakaryoblastic leukemia cell lines. Single-cell and bulk transcriptomics of patient samples with erythroid and megakaryoblastic leukemias identified high BCL2L1 expression compared to other subtypes of AML and other hematological malignancies, providing a basis for the observed functional dependencies.

BCL-XL inhibition effectively killed bone marrow cells of 5 out of 8 AML patient samples with erythroid or megakaryocytic differentiation ex vivo, with IC50 ranging from 0.6 to 55 nM. In contrast, only one out of 40 other AML patient samples was sensitive (IC50 < 60 nM) to BCL-XL inhibition. To better understand the drug sensitivity and gene expression profiles of the blasts, we performed integrated single-cell transcriptomics and ex vivo flow cytometry-based drug sensitivity profiling on mononuclear cells from two patients with erythroid/megakaryocytic AML. The blasts showed enriched BCL2L1 expression compared to other cell populations, providing a basis for the increased sensitivity of the blasts to BCL-XL inhibition and venetoclax resistance observed also at the single-cell resolution.

In a mouse xenograft model using the HEL erythroleukemia cell line, treatment with the BCL-XL inhibitor A-1331852 effectively reduced tumor burden. To find treatment regimens enabling long-term disease control, we explored potential combination treatments by testing 6 other compounds with the BCL-XL inhibitor A-1331852 in 8x8 concentration matrices. Combining BCL-XL inhibition with the JAK inhibitor ruxolitinib, the BCL-2 inhibitor venetoclax, or the hypomethylating agent azacitidine showed potential for more durable responses both in a 3-day ex vivo assay as well as long-term treatments spanning over a month.

Collectively, our results suggest targeting BCL-XL as a potential therapy option in erythroid and megakaryoblastic leukemias and highlight an AML subgroup with potentially reduced sensitivity to venetoclax-based treatments.

Disclosures: Kuusanmäki: Faron: Consultancy; AbbVie: Research Funding. Konopleva: Eli Lilly: Consultancy, Patents & Royalties, Research Funding; Cellectis: Consultancy, Other: Grant support, Research Funding; Calithera: Other: Grant Support, Research Funding; Ablynx: Other: Grant support, Research Funding; Agios: Other: grant support, Research Funding; Ascentage: Other: grant support, Research Funding; AstraZeneca: Other: grant support, Research Funding; Rafael Pharmaceutical: Other: grant support, Research Funding; Sanofi: Other: grant support, Research Funding; Novartis: Patents & Royalties, Research Funding; Reata Pharmaceuticals: Current equity holder in private company, Patents & Royalties; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Kisoji: Consultancy, Honoraria; Forty-Seven: Consultancy, Honoraria, Other: Grant support; Amgen: Consultancy; F. Hoffman La Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Grant support, Research Funding; Stemline Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genentech: Consultancy, Other: grant support, Research Funding; AbbVie: Consultancy, Other: grant support, Research Funding. Porkka: Pfizer: Honoraria; Celgene/Bristol-Myers Squibb: Research Funding; Incyte: Research Funding; Pfizer: Research Funding; Novartis: Research Funding; Novartis: Honoraria; Incyte: Honoraria; Bristol-Myers Squibb: Honoraria; Astellas: Honoraria; AbbVie: Honoraria. Kontro: Novartis: Membership on an entity's Board of Directors or advisory committees; Faron Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Astellas Pharma: Consultancy; Pfizer: Membership on an entity's Board of Directors or advisory committees. Heckman: Celgene: Research Funding; Kronos Bio: Research Funding; Novartis: Research Funding; Oncopeptides: Research Funding; Orion: Research Funding; IMI2 projects HARMONY and HARMONY PLUS: Research Funding; WntResearch: Research Funding. Mustjoki: Pfizer: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

*signifies non-member of ASH