-Author name in bold denotes the presenting author
-Asterisk * with author name denotes a Non-ASH member
Clinically Relevant Abstract denotes an abstract that is clinically relevant.

PhD Trainee denotes that this is a recommended PHD Trainee Session.

Ticketed Session denotes that this is a ticketed session.

43 Epigenetic Resistance to Menin-MLL1 Inhibition Is Driven By Loss of the Non-Canonical Polycomb Repressive Complex 1.1 in NUP98-Rearranged AML

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, Translational Research, drug development, Diseases, Therapies, Myeloid Malignancies, Biological Processes, molecular biology
Saturday, December 10, 2022: 9:30 AM

Emily B. Heikamp, MD, PhD1*, Jill A Henrich1,2*, Hannah E Rice1*, Sanisha Mahendra-Rajah1* and Scott A. Armstrong, MD, PhD1

1Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, MA
2University of Pennsylvania, Philadelphia, PA

Chromosomal translocations producing oncogenic fusion proteins are common drivers of many types of cancer, including leukemia. Translocations involving the Nucleoporin 98 (NUP98) gene produce NUP98-fusion proteins and are associated with a dire prognosis in acute myeloid leukemia (AML). NUP98-fusion proteins interact with the mixed-lineage leukemia (MLL1) chromatin modifying enzyme, and MLL1 is a molecular dependency in NUP98-rearranged (NUP98-r) AML. Our group recently showed that targeting the interaction between MLL1 and its binding partner Menin is therapeutically effective in NUP98-r AML models. Menin-MLL1 inhibition using a small molecule VTP50469 simultaneously represses pro-leukemogenic genes and upregulates markers of myeloid differentiation.

Mechanistically, we showed that disruption of the Menin-MLL1 interaction using VTP50469 leads to eviction of NUP98-fusion proteins from the transcriptional start site (TSS) of pro-leukemogenic genes. However, it remains unclear whether loss of the NUP98-fusion-Menin-MLL1 complex is sufficient to silence pro-leukemogenic gene expression and induce differentiation. We hypothesized that differentiation must be enforced through epigenetic silencing of pro-leukemogenic genes by a repressive chromatin complex.

We performed a bespoke CRISPR screen targeting 616 epigenetic regulators in NUP98-r leukemia cells treated with the Menin-MLL1 inhibitor VTP50469. Our results revealed that the non-canonical polycomb repressive complex 1.1 (PRC1.1)—a dynamic, repressive chromatin regulatory complex with an established role in lineage specification and cancer—are required for Menin-MLL1 inhibitors to inhibit cell proliferation. Loss of each PRC1.1 complex component resulted in a competitive growth advantage in our CRISPR screen, and this effect was enhanced in cells treated with VTP50469, suggesting that loss of PRC1.1 may mediate resistance to Menin-MLL1 inhibition.

To validate our screen and further define the role of PRC1.1 in NUP98-r leukemia, we used CRISPR/Cas9 to genetically inactivate each component of the PRC1.1 complex. We found that loss of PRC1.1 components resulted in a competitive growth advantage of NUP98-r leukemia cells, and fitness of PRC1.1-deficient leukemia cells was enhanced by treatment with VTP50469.

Next, we determined how PRC1.1 localization and activity changes upon eviction of NUP98-fusion proteins from chromatin using Menin-MLL1 inhibitors. We found that PRC1.1 components colocalize with the NUP98-Menin-MLL1 complex at the TSS of pro-leukemogenic genes whose expression is significantly downregulated upon treatment with VTP50469 such as MEIS1, EYA1, and PBX3. Menin-MLL inhibition resulted in increased PRC1.1 activity at these loci, as measured by the repressive histone mark H2aK119Ub. These results suggest that the NUP98-fusion-Menin-MLL1 complex and PRC1.1 dynamically compete for transcriptional control of pro-leukemogenic gene expression in NUP98-r leukemia.

To confirm these findings in a different model, we engineered a system in which NUP98-fusion proteins can be rapidly and inducibly degraded using heterobifunctional small molecules such as dTAG-13. We tagged NUP98-fusion proteins with a modified version of FKBP and used these constructs to transform mouse lineage negative, Sca-1 +, cKit+ (LSK) cells. We found that degradation of NUP98-fusion proteins resulted in rapid changes in transcription at key pro-leukemogenic loci upon dTAG-13 treatment. Genes which lost active transcription were bound by PRC1.1, and degradation of NUP98-fusion proteins led to accumulation of this repressive complex, increased H2aK119Ub, and loss of activating histone marks such as H3K27Ac.

In summary, these results demonstrate that the NUP98-fusion-Menin-MLL complex and PRC1.1 dynamically compete for transcriptional control of developmentally regulated, stem-cell associated genes. Loss of PRC1.1 may promote NUP98-fusion protein-driven leukemogenesis and mediates resistance to Menin-MLL1 inhibition by failing to epigenetically silence genes that are essential for maintaining an undifferentiated, stem cell-like state.

Disclosures: Armstrong: C4 Therapeutics: Consultancy, Other: SHareholder; Accent Therapeutics: Consultancy, Other: Shareholder; Novartis: Research Funding; Twenty eight-seven Therapeutics: Consultancy, Other: Shareholder; Mana Therapeutics: Consultancy, Other: Shareholder; -: Patents & Royalties: MENIN inhibition WO/2017/132398A1; Janssen: Research Funding; Cyteir Therapeutics: Consultancy, Other: Shareholder; Imago Biosciences: Consultancy, Other: Shareholder; Syndax: Research Funding; Neomorph Inc: Consultancy.

Previous Abstract | Next Abstract >>
*signifies non-member of ASH