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Nature Dec 2022Higher-order chromatin structure is important for the regulation of genes by distal regulatory sequences. Structural variants (SVs) that alter three-dimensional (3D)...
Higher-order chromatin structure is important for the regulation of genes by distal regulatory sequences. Structural variants (SVs) that alter three-dimensional (3D) genome organization can lead to enhancer-promoter rewiring and human disease, particularly in the context of cancer. However, only a small minority of SVs are associated with altered gene expression, and it remains unclear why certain SVs lead to changes in distal gene expression and others do not. To address these questions, we used a combination of genomic profiling and genome engineering to identify sites of recurrent changes in 3D genome structure in cancer and determine the effects of specific rearrangements on oncogene activation. By analysing Hi-C data from 92 cancer cell lines and patient samples, we identified loci affected by recurrent alterations to 3D genome structure, including oncogenes such as MYC, TERT and CCND1. By using CRISPR-Cas9 genome engineering to generate de novo SVs, we show that oncogene activity can be predicted by using 'activity-by-contact' models that consider partner region chromatin contacts and enhancer activity. However, activity-by-contact models are only predictive of specific subsets of genes in the genome, suggesting that different classes of genes engage in distinct modes of regulation by distal regulatory elements. These results indicate that SVs that alter 3D genome organization are widespread in cancer genomes and begin to illustrate predictive rules for the consequences of SVs on oncogene activation.
Topics: Humans; Chromatin; Gene Rearrangement; Genomic Structural Variation; Neoplasms; Oncogenes; Oncogene Proteins; Chromosomes, Human; Cell Line, Tumor; Enhancer Elements, Genetic; Models, Genetic
PubMed: 36477537
DOI: 10.1038/s41586-022-05504-4 -
Nature Reviews. Cancer Sep 2023The genomic simplicity of differentiated cancers derived from thyroid follicular cells offers unique insights into how oncogenic drivers impact tumour phenotype.... (Review)
Review
The genomic simplicity of differentiated cancers derived from thyroid follicular cells offers unique insights into how oncogenic drivers impact tumour phenotype. Essentially, the main oncoproteins in thyroid cancer activate nodes in the receptor tyrosine kinase-RAS-BRAF pathway, which constitutively induces MAPK signalling to varying degrees consistent with their specific biochemical mechanisms of action. The magnitude of the flux through the MAPK signalling pathway determines key elements of thyroid cancer biology, including differentiation state, invasive properties and the cellular composition of the tumour microenvironment. Progression of disease results from genomic lesions that drive immortalization, disrupt chromatin accessibility and cause cell cycle checkpoint dysfunction, in conjunction with a tumour microenvironment characterized by progressive immunosuppression. This Review charts the genomic trajectories of these common endocrine tumours, while connecting them to the biological states that they confer.
Topics: Humans; Thyroid Epithelial Cells; Thyroid Neoplasms; Signal Transduction; Receptor Protein-Tyrosine Kinases; Oncogene Proteins; Proto-Oncogene Proteins B-raf; Tumor Microenvironment
PubMed: 37438605
DOI: 10.1038/s41568-023-00598-y -
Cancer Discovery Apr 2022NUP98 fusion oncoproteins (FO) are drivers in pediatric leukemias and many transform hematopoietic cells. Most NUP98 FOs harbor an intrinsically disordered region from...
UNLABELLED
NUP98 fusion oncoproteins (FO) are drivers in pediatric leukemias and many transform hematopoietic cells. Most NUP98 FOs harbor an intrinsically disordered region from NUP98 that is prone to liquid-liquid phase separation (LLPS) in vitro. A predominant class of NUP98 FOs, including NUP98-HOXA9 (NHA9), retains a DNA-binding homeodomain, whereas others harbor other types of DNA- or chromatin-binding domains. NUP98 FOs have long been known to form puncta, but long-standing questions are how nuclear puncta form and how they drive leukemogenesis. Here we studied NHA9 condensates and show that homotypic interactions and different types of heterotypic interactions are required to form nuclear puncta, which are associated with aberrant transcriptional activity and transformation of hematopoietic stem and progenitor cells. We also show that three additional leukemia-associated NUP98 FOs (NUP98-PRRX1, NUP98-KDM5A, and NUP98-LNP1) form nuclear puncta and transform hematopoietic cells. These findings indicate that LLPS is critical for leukemogenesis by NUP98 FOs.
SIGNIFICANCE
We show that homotypic and heterotypic mechanisms of LLPS control NUP98-HOXA9 puncta formation, modulating transcriptional activity and transforming hematopoietic cells. Importantly, these mechanisms are generalizable to other NUP98 FOs that share similar domain structures. These findings address long-standing questions on how nuclear puncta form and their link to leukemogenesis. This article is highlighted in the In This Issue feature, p. 873.
Topics: Carcinogenesis; Cell Nucleus; Child; Homeodomain Proteins; Humans; Leukemia; Nuclear Pore Complex Proteins; Oncogene Proteins, Fusion; Retinoblastoma-Binding Protein 2
PubMed: 34903620
DOI: 10.1158/2159-8290.CD-21-0674 -
Cancer Science Jul 2020The Mdm2 oncoprotein and its association with p53 were discovered 30 years ago, and a cornucopia of activities and regulatory pathways have been associated with it. In... (Review)
Review
The Mdm2 oncoprotein and its association with p53 were discovered 30 years ago, and a cornucopia of activities and regulatory pathways have been associated with it. In this review, we will raise questions about Mdm2 and its cousin Mdm4 that we consider worth pursuing in future research, reaching from molecular structures and intracellular activities all the way to development, evolution, and cancer therapy. We anticipate that such research will not only close a few gaps in our knowledge but could add new dimensions to our current view. This compilation of questions contributes to the preparation for the 10th Mdm2 Workshop in Tokyo.
Topics: Aging; Animals; Biological Evolution; Biomarkers, Tumor; Cell Cycle Proteins; Cell Transformation, Neoplastic; Gene Expression Regulation; Humans; Molecular Targeted Therapy; Multigene Family; Mutation; Protein Binding; Protein Transport; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-mdm2; Signal Transduction; Stress, Physiological; Tumor Suppressor Protein p53
PubMed: 32335977
DOI: 10.1111/cas.14433 -
Blood Feb 2022Translocations involving the NUP98 gene produce NUP98-fusion proteins and are associated with a poor prognosis in acute myeloid leukemia (AML). MLL1 is a molecular...
Translocations involving the NUP98 gene produce NUP98-fusion proteins and are associated with a poor prognosis in acute myeloid leukemia (AML). MLL1 is a molecular dependency in NUP98-fusion leukemia, and therefore we investigated the efficacy of therapeutic blockade of the menin-MLL1 interaction in NUP98-fusion leukemia models. Using mouse leukemia cell lines driven by NUP98-HOXA9 and NUP98-JARID1A fusion oncoproteins, we demonstrate that NUP98-fusion-driven leukemia is sensitive to the menin-MLL1 inhibitor VTP50469, with an IC50 similar to what we have previously reported for MLL-rearranged and NPM1c leukemia cells. Menin-MLL1 inhibition upregulates markers of differentiation such as CD11b and downregulates expression of proleukemogenic transcription factors such as Meis1 in NUP98-fusion-transformed leukemia cells. We demonstrate that MLL1 and the NUP98 fusion protein itself are evicted from chromatin at a critical set of genes that are essential for the maintenance of the malignant phenotype. In addition to these in vitro studies, we established patient-derived xenograft (PDX) models of NUP98-fusion-driven AML to test the in vivo efficacy of menin-MLL1 inhibition. Treatment with VTP50469 significantly prolongs survival of mice engrafted with NUP98-NSD1 and NUP98-JARID1A leukemias. Gene expression analysis revealed that menin-MLL1 inhibition simultaneously suppresses a proleukemogenic gene expression program, including downregulation of the HOXa cluster, and upregulates tissue-specific markers of differentiation. These preclinical results suggest that menin-MLL1 inhibition may represent a rational, targeted therapy for patients with NUP98-rearranged leukemias.
Topics: Animals; Cell Line, Tumor; Gene Expression Regulation, Leukemic; Gene Rearrangement; Histone-Lysine N-Methyltransferase; Leukemia, Myeloid, Acute; Mice; Mice, Inbred C57BL; Myeloid-Lymphoid Leukemia Protein; Nuclear Pore Complex Proteins; Oncogene Proteins, Fusion; Protein Interaction Maps; Proto-Oncogene Proteins
PubMed: 34582559
DOI: 10.1182/blood.2021012806 -
Nature Communications Jan 2022Efforts to therapeutically target EZH2 have generally focused on inhibition of its methyltransferase activity, although it remains less clear whether this is the central...
Efforts to therapeutically target EZH2 have generally focused on inhibition of its methyltransferase activity, although it remains less clear whether this is the central mechanism whereby EZH2 promotes cancer. In the current study, we show that EZH2 directly interacts with both MYC family oncoproteins, MYC and MYCN, and promotes their stabilization in a methyltransferase-independent manner. By competing against the SCF ubiquitin ligase to bind MYC and MYCN, EZH2 counteracts FBW7-mediated MYC(N) polyubiquitination and proteasomal degradation. Depletion, but not enzymatic inhibition, of EZH2 induces robust MYC(N) degradation and inhibits tumor cell growth in MYC(N) driven neuroblastoma and small cell lung carcinoma. Here, we demonstrate the MYC family proteins as global EZH2 oncogenic effectors and EZH2 pharmacologic degraders as potential MYC(N) targeted cancer therapeutics, pointing out that MYC(N) driven cancers may develop inherent resistance to the canonical EZH2 enzymatic inhibitors currently in clinical development.
Topics: Carcinoma, Small Cell; Cell Line, Tumor; Drug Delivery Systems; Enhancer of Zeste Homolog 2 Protein; Humans; N-Myc Proto-Oncogene Protein; Neuroblastoma; Oncogene Proteins; Small Cell Lung Carcinoma
PubMed: 35013218
DOI: 10.1038/s41467-021-27609-6 -
Cancer Cell Sep 2021Fusion-transcription factors (fusion-TFs) represent a class of driver oncoproteins that are difficult to therapeutically target. Recently, protein degradation has...
Fusion-transcription factors (fusion-TFs) represent a class of driver oncoproteins that are difficult to therapeutically target. Recently, protein degradation has emerged as a strategy to target these challenging oncoproteins. The mechanisms that regulate fusion-TF stability, however, are generally unknown. Using CRISPR-Cas9 screening, we discovered tripartite motif-containing 8 (TRIM8) as an E3 ubiquitin ligase that ubiquitinates and degrades EWS/FLI, a driver fusion-TF in Ewing sarcoma. Moreover, we identified TRIM8 as a selective dependency in Ewing sarcoma compared with >700 other cancer cell lines. Mechanistically, TRIM8 knockout led to an increase in EWS/FLI protein levels that was not tolerated. EWS/FLI acts as a neomorphic substrate for TRIM8, defining the selective nature of the dependency. Our results demonstrate that fusion-TF protein stability is tightly regulated and highlight fusion oncoprotein-specific regulators as selective therapeutic targets. This study provides a tractable strategy to therapeutically exploit oncogene overdose in Ewing sarcoma and potentially other fusion-TF-driven cancers.
Topics: Bone Neoplasms; Carrier Proteins; Cell Line, Tumor; Cell Proliferation; Cell Survival; Gene Knockout Techniques; HEK293 Cells; Humans; Microfilament Proteins; Nerve Tissue Proteins; Oncogene Proteins, Fusion; Protein Stability; Proteolysis; Proto-Oncogene Protein c-fli-1; RNA-Binding Protein EWS; Sarcoma, Ewing; Trans-Activators
PubMed: 34329586
DOI: 10.1016/j.ccell.2021.07.003 -
Nature Nov 2021The majority of oncogenic drivers are intracellular proteins, thus constraining their immunotherapeutic targeting to mutated peptides (neoantigens) presented by...
The majority of oncogenic drivers are intracellular proteins, thus constraining their immunotherapeutic targeting to mutated peptides (neoantigens) presented by individual human leukocyte antigen (HLA) allotypes. However, most cancers have a modest mutational burden that is insufficient to generate responses using neoantigen-based therapies. Neuroblastoma is a paediatric cancer that harbours few mutations and is instead driven by epigenetically deregulated transcriptional networks. Here we show that the neuroblastoma immunopeptidome is enriched with peptides derived from proteins that are essential for tumourigenesis and focus on targeting the unmutated peptide QYNPIRTTF, discovered on HLA-A*24:02, which is derived from the neuroblastoma dependency gene and master transcriptional regulator PHOX2B. To target QYNPIRTTF, we developed peptide-centric chimeric antigen receptors (CARs) using a counter-panning strategy with predicted potentially cross-reactive peptides. We further hypothesized that peptide-centric CARs could recognize peptides on additional HLA allotypes when presented in a similar manner. Informed by computational modelling, we showed that PHOX2B peptide-centric CARs also recognize QYNPIRTTF presented by HLA-A*23:01 and the highly divergent HLA-B*14:02. Finally, we demonstrated potent and specific killing of neuroblastoma cells expressing these HLAs in vitro and complete tumour regression in mice. These data suggest that peptide-centric CARs have the potential to vastly expand the pool of immunotherapeutic targets to include non-immunogenic intracellular oncoproteins and widen the population of patients who would benefit from such therapy by breaking conventional HLA restriction.
Topics: Animals; Antigens, Neoplasm; Cell Line; Cell Line, Tumor; Cross Reactions; Cross-Priming; Female; HLA Antigens; Homeodomain Proteins; Humans; Immunotherapy; Interferon-gamma; Mice; Neoplasms; Oncogene Proteins; Receptors, Chimeric Antigen; T-Lymphocytes; Transcription Factors
PubMed: 34732890
DOI: 10.1038/s41586-021-04061-6 -
Nature Cell Biology Feb 2023The EWS-FLI1 fusion oncoprotein deregulates transcription to initiate the paediatric cancer Ewing sarcoma. Here we used a domain-focused CRISPR screen to implicate the...
The EWS-FLI1 fusion oncoprotein deregulates transcription to initiate the paediatric cancer Ewing sarcoma. Here we used a domain-focused CRISPR screen to implicate the transcriptional repressor ETV6 as a unique dependency in this tumour. Using biochemical assays and epigenomics, we show that ETV6 competes with EWS-FLI1 for binding to select DNA elements enriched for short GGAA repeat sequences. Upon inactivating ETV6, EWS-FLI1 overtakes and hyper-activates these cis-elements to promote mesenchymal differentiation, with SOX11 being a key downstream target. We show that squelching of ETV6 with a dominant-interfering peptide phenocopies these effects and suppresses Ewing sarcoma growth in vivo. These findings reveal targeting of ETV6 as a strategy for neutralizing the EWS-FLI1 oncoprotein by reprogramming of genomic occupancy.
Topics: Child; Humans; Sarcoma, Ewing; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; RNA-Binding Protein EWS; Proto-Oncogene Protein c-fli-1; Oncogene Proteins, Fusion
PubMed: 36658219
DOI: 10.1038/s41556-022-01060-1 -
Nature Communications Feb 2022Targeted protein degradation allows targeting undruggable proteins for therapeutic applications as well as eliminating proteins of interest for research purposes. While...
Targeted protein degradation allows targeting undruggable proteins for therapeutic applications as well as eliminating proteins of interest for research purposes. While several degraders that harness the proteasome or the lysosome have been developed, a technology that simultaneously degrades targets and accelerates cellular autophagic flux is still missing. In this study, we develop a general chemical tool and platform technology termed AUTOphagy-TArgeting Chimera (AUTOTAC), which employs bifunctional molecules composed of target-binding ligands linked to autophagy-targeting ligands. AUTOTACs bind the ZZ domain of the otherwise dormant autophagy receptor p62/Sequestosome-1/SQSTM1, which is activated into oligomeric bodies in complex with targets for their sequestration and degradation. We use AUTOTACs to degrade various oncoproteins and degradation-resistant aggregates in neurodegeneration at nanomolar DC values in vitro and in vivo. AUTOTAC provides a platform for selective proteolysis in basic research and drug development.
Topics: Autophagy; Cell Line, Tumor; HeLa Cells; Humans; Lysosomes; Oncogene Proteins; Protein Aggregates; Protein Binding; Protein Folding; Proteolysis; Proteostasis; Signal Transduction
PubMed: 35173167
DOI: 10.1038/s41467-022-28520-4