Contributions
Abstract: S133
Type: Oral Presentation
Presentation during EHA22: On Friday, June 23, 2017 from 11:30 - 11:45
Location: Room N103
Background
Acute Myeloid Leukemia (AML) frequently harbors chromosomal rearrangements involving the Mixed Lineage Leukemia (MLL) gene. More than 65 different MLL fusion genes exist and many of them have been described to act as strong cancer drivers. While critical effectors of several distinct MLL fusion proteins (MLL-FPs) were identified, it is not clear if transforming mechanisms are conserved across the entire family of MLL fusions.
Aims
Methods
Results
Characterization of the protein complexes nucleated by 7 MLL fusion proteins by affinity purification coupled to mass spectrometry (AP-MS) revealed a densely interconnected protein-protein interaction network of 963 proteins, comprising previously known MLL-interacting protein complexes (such as PRC2 or SWI/SNF), as well as a high number of new interaction partners of MLL. 128 proteins were found to interact with ≥5 of all 7 MLL-fusions. This subset of conserved MLL-interaction partners was highly enriched for proteins with function in chromatin metabolism and transcriptional control. Systematic functional investigation of the conserved MLL-fusion interactome using subtractive shRNA screens identified the methyltransferase SETD2 as a critical effector of MLL fusion proteins. Both RNAi-based suppression and CRISPR/Cas9-mediated mutagenesis of SETD2 induced myeloid differentiation and apoptosis in human and mouse MLL-rearranged cell lines, while having only modest effects on the proliferation of MLL-wild-type leukemia cells. Depletion of Setd2 in MLL-fusion-transformed mouse fetal liver cells resulted in loss of serial re-plating capacity in vitro and prolonged disease onset in vivo. Furthermore, knockdown of SETD2 caused a proliferative disadvantage in primary cells from AML patients with different MLL-rearrangements without affecting MLL-wild-type AML cells. We found that SETD2 was essential for efficient repair of DNA breaks, as SETD2-deficient leukemia cells showed increased levels of DNA damage and activation of p53, leading to the accumulation of mutations.
Conclusion
In summary, our data highlight the functional relevance of combined proteomic-genomic cellular screening to identify critical effectors of MLL-FPs. In addition, our study identifies a novel role for SETD2 in the maintenance of genomic integrity during initiation and progression of MLL-rearranged AML and establishes SETD2 as a therapeutic target in leukemia with low genomic complexity.
Session topic: 3. Acute myeloid leukemia - Biology
Keyword(s): 11q23, RNA interference (RNAi), Protein-protein interaction, MLL
Abstract: S133
Type: Oral Presentation
Presentation during EHA22: On Friday, June 23, 2017 from 11:30 - 11:45
Location: Room N103
Background
Acute Myeloid Leukemia (AML) frequently harbors chromosomal rearrangements involving the Mixed Lineage Leukemia (MLL) gene. More than 65 different MLL fusion genes exist and many of them have been described to act as strong cancer drivers. While critical effectors of several distinct MLL fusion proteins (MLL-FPs) were identified, it is not clear if transforming mechanisms are conserved across the entire family of MLL fusions.
Aims
Methods
Results
Characterization of the protein complexes nucleated by 7 MLL fusion proteins by affinity purification coupled to mass spectrometry (AP-MS) revealed a densely interconnected protein-protein interaction network of 963 proteins, comprising previously known MLL-interacting protein complexes (such as PRC2 or SWI/SNF), as well as a high number of new interaction partners of MLL. 128 proteins were found to interact with ≥5 of all 7 MLL-fusions. This subset of conserved MLL-interaction partners was highly enriched for proteins with function in chromatin metabolism and transcriptional control. Systematic functional investigation of the conserved MLL-fusion interactome using subtractive shRNA screens identified the methyltransferase SETD2 as a critical effector of MLL fusion proteins. Both RNAi-based suppression and CRISPR/Cas9-mediated mutagenesis of SETD2 induced myeloid differentiation and apoptosis in human and mouse MLL-rearranged cell lines, while having only modest effects on the proliferation of MLL-wild-type leukemia cells. Depletion of Setd2 in MLL-fusion-transformed mouse fetal liver cells resulted in loss of serial re-plating capacity in vitro and prolonged disease onset in vivo. Furthermore, knockdown of SETD2 caused a proliferative disadvantage in primary cells from AML patients with different MLL-rearrangements without affecting MLL-wild-type AML cells. We found that SETD2 was essential for efficient repair of DNA breaks, as SETD2-deficient leukemia cells showed increased levels of DNA damage and activation of p53, leading to the accumulation of mutations.
Conclusion
In summary, our data highlight the functional relevance of combined proteomic-genomic cellular screening to identify critical effectors of MLL-FPs. In addition, our study identifies a novel role for SETD2 in the maintenance of genomic integrity during initiation and progression of MLL-rearranged AML and establishes SETD2 as a therapeutic target in leukemia with low genomic complexity.
Session topic: 3. Acute myeloid leukemia - Biology
Keyword(s): 11q23, RNA interference (RNAi), Protein-protein interaction, MLL