Publications by authors named "Takaomi Sanda"

64 Publications

Myeloma-specific superenhancers affect genes of biological and clinical relevance in myeloma.

Blood Cancer J 2021 Feb 12;11(2):32. Epub 2021 Feb 12.

Cancer Science Institute of Singapore, National University of Singapore, 14 Medical Drive, Centre for Translational Medicine, Singapore, 117599, Republic of Singapore.

Multiple myeloma (MM) is an aggressive plasma cell neoplasm characterized by genomic heterogeneity. Superenhancers (SEs) are defined as large clusters of enhancers in close genomic proximity, which regulate genes for maintaining cellular identity and promote oncogenic transcription to which cancer cells highly addicted. Here, we analyzed cis-regulatory elements in MM samples with H3K27ac ChIP-seq, to identify novel SE-associated genes involved in the myeloma pathogenesis. SEs and their associated genes in cancerous tissue were compared with the control samples, and we found SE analysis alone uncovered cell-lineage-specific transcription factors and well-known oncogenes ST3GAL6 and ADM. Using a transcriptional CDK7 inhibitor, THZ1, coupled with H3K27ac ChlP-seq, we identified MAGI2 as a novel SE-associated gene of myeloma cells. Elevated MAGI2 was related to myelomagenesis with gradual increased expression from MGUS, SMM to newly diagnosed and relapsed MM. High prevalence of MAGI2 was also associated with poor survival of MM patients. Importantly, inhibition of the SE activity associated with MAGI2 decreased MAGI2 expression, inhibited cell growth and induced cell apoptosis. Mechanistically, we revealed that the oncogenic transcription factor, MAF, directly bound to the SE region and activated gene transcription. In summary, the discoveries of these acquired SEs-associated genes and the novel mechanism by which they are regulated provide new insights into MM biology and MAGI2-MAF-SE regulatory circuit offer potential novel targets for disease treatment.
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http://dx.doi.org/10.1038/s41408-021-00421-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7881003PMC
February 2021

Super-enhancers for RUNX3 are required for cell proliferation in EBV-infected B cell lines.

Gene 2021 Mar 12;774:145421. Epub 2021 Jan 12.

Cancer Science Institute of Singapore, National University of Singapore, Singapore; Department of Paediatrics, Yong Loo Lin School of Medicine, National University of Singapore, Singapore; International Research Center for Medical Sciences, Kumamoto University, Japan. Electronic address:

Epstein-Barr virus nuclear antigens 2 (EBNA2) mediated super-enhancers, defined by in silico data, localize near genes associated with B cell transcription factors including RUNX3. However, the biological function of super-enhancer for RUNX3 gene (seR3) remains unclear. Here, we show that two seR3s, tandemly-located at 59- and 70-kb upstream of RUNX3 transcription start site, named seR3 -59h and seR3 -70h, are required for RUNX3 expression and cell proliferation in Epstein-Barr virus (EBV)-positive malignant B cells. A BET bromodomain inhibitor, JQ1, potently suppressed EBV-positive B cell growth through the reduction of RUNX3 and MYC expression. Excision of either or both seR3s by employing CRISPR/Cas9 system resulted in the decrease in RUNX3 expression and the subsequent suppression of cell proliferation and colony forming capability. The expression of MYC was also reduced when seR3s were deleted, probably due to the loss of trans effect of seR3s on the super-enhancers for MYC. These findings suggest that seR3s play a pivotal role in expression and biological function of both RUNX3 and MYC. seR3s would serve as a potential therapeutic target in EBV-related widespread tumors.
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http://dx.doi.org/10.1016/j.gene.2021.145421DOI Listing
March 2021

ESRRB regulates glucocorticoid gene expression in mice and patients with acute lymphoblastic leukemia.

Blood Adv 2020 07;4(13):3154-3168

Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, MA; and.

Synthetic glucocorticoids (GCs), such as dexamethasone and prednisone, remain key components of therapy for patients with lymphoid malignancies. For pediatric patients with acute lymphoblastic leukemia (ALL), response to GCs remains the most reliable prognostic indicator; failure to respond to GC correlates with poor event-free survival. To uncover GC resistance mechanisms, we performed a genome-wide, survival-based short hairpin RNA screen and identified the orphan nuclear receptor estrogen-related receptor-β (ESRRB) as a critical transcription factor that cooperates with the GC receptor (GR) to mediate the GC gene expression signature in mouse and human ALL cells. Esrrb knockdown interfered with the expression of genes that were induced and repressed by GR and resulted in GC resistance in vitro and in vivo. Dexamethasone treatment stimulated ESRRB binding to estrogen-related receptor elements (ERREs) in canonical GC-regulated genes, and H3K27Ac Hi-chromatin immunoprecipitation revealed increased interactions between GR- and ERRE-containing regulatory regions in dexamethasone-treated human T-ALL cells. Furthermore, ESRRB agonists enhanced GC target gene expression and synergized with dexamethasone to induce leukemic cell death, indicating that ESRRB agonists may overcome GC resistance in ALL, and potentially, in other lymphoid malignancies.
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http://dx.doi.org/10.1182/bloodadvances.2020001555DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7362368PMC
July 2020

Oncorequisite role of an aldehyde dehydrogenase in the pathogenesis of T-cell acute lymphoblastic leukemia.

Haematologica 2020 05 15. Epub 2020 May 15.

Cancer Science Institute of Singapore, National University of Singapore, Singapore;

Aldehyde dehydrogenases (ALDHs) are overexpressed in various types of cancers. One of the ALDH family genes, ALDH1A2, is aberrantly expressed in more than 50% of T-cell acute lymphoblastic leukemia (T-ALL) cases. However, its molecular function and role in T-ALL pathogenesis are largely unknown. ChIP-seq and RNA-seq analyses showed that the oncogenic transcription factor TAL1 and its regulatory partners bind to the intronic regulatory element of the ALDH1A2 gene, directly inducing a T-ALL-specific isoform with enzymatic activity. ALDH1A2 was preferentially expressed in the TAL1-positive T-ALL subgroup. In T-ALL cell lines, depletion of ALDH1A2 inhibited cell viability and induced apoptosis. Interestingly, gene expression and metabolomic profiling revealed that ALDH1A2 supported glycolysis and the TCA cycle, accompanied by NADH production, by affecting multiple metabolic enzymes to promote ATP production. Depletion of ALDH1A2 increased the levels of reactive oxygen species (ROS), while ROS levels were reduced by ALDH1A2 overexpression both in vitro and in vivo. Overexpression of ALDH1A2 accelerated tumor onset and increased tumor penetrance in a zebrafish T-ALL model. Taken together, our results indicate that ALDH1A2 protects against intracellular stress and promotes T-ALL cell metabolism and survival. ALDH1A2 overexpression enables leukemic clones to sustain a hyper-proliferative state driven by oncogenes.
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http://dx.doi.org/10.3324/haematol.2019.245639DOI Listing
May 2020

Feed-forward regulatory loop driven by IRF4 and NF-κB in adult T-cell leukemia/lymphoma.

Blood 2020 03;135(12):934-947

Cancer Science Institute of Singapore, National University of Singapore, Singapore.

Adult T-cell leukemia/lymphoma (ATL) is a highly aggressive hematological malignancy derived from mature CD4+ T-lymphocytes. Here, we demonstrate the transcriptional regulatory network driven by 2 oncogenic transcription factors, IRF4 and NF-κB, in ATL cells. Gene expression profiling of primary ATL samples demonstrated that the IRF4 gene was more highly expressed in ATL cells than in normal T cells. Chromatin immunoprecipitation sequencing analysis revealed that IRF4-bound regions were more frequently found in super-enhancers than in typical enhancers. NF-κB was found to co-occupy IRF4-bound regulatory elements and formed a coherent feed-forward loop to coordinately regulate genes involved in T-cell functions and development. Importantly, IRF4 and NF-κB regulated several cancer genes associated with super-enhancers in ATL cells, including MYC, CCR4, and BIRC3. Genetic inhibition of BIRC3 induced growth inhibition in ATL cells, implicating its role as a critical effector molecule downstream of the IRF4-NF-κB transcriptional network.
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http://dx.doi.org/10.1182/blood.2019002639DOI Listing
March 2020

Sphingosine 1-Phosphate Receptor 2 Induces Otoprotective Responses to Cisplatin Treatment.

Cancers (Basel) 2020 Jan 15;12(1). Epub 2020 Jan 15.

Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, 117600 Singapore, Singapore.

Ototoxicity is a major adverse effect of platinum-based chemotherapeutics and currently, there remains a lack of United States Food and Drug Administration-approved therapies to prevent or treat this problem. In our study, we examined the role of the sphingosine 1-phosphate receptor 2 (S1P) in attenuating cisplatin-induced ototoxicity in several different animal models and cell lines. We found that ototoxicity in S1P knockout mice is dependent on reactive oxygen species (ROS) production and that S1P receptor activation with a specific agonist, CYM-5478, significantly attenuates cisplatin-induced defects, including hair cell degeneration in zebrafish and prolonged auditory brainstem response latency in rats. We also evaluated the cytoprotective effect of CYM-5478 across different cell lines and showed that CYM-5478 protects neural-derived cell lines but not breast cancer cells against cisplatin toxicity. We show that this selective protection of CYM-5478 is due to its differential effects on key regulators of apoptosis between neural cells and breast cancer cells. Overall, our study suggests that targeting the S1P receptor represents a promising therapeutic approach for the treatment of cisplatin-induced ototoxicity in cancer patients.
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http://dx.doi.org/10.3390/cancers12010211DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7016659PMC
January 2020

ASCL1 is a MYCN- and LMO1-dependent member of the adrenergic neuroblastoma core regulatory circuitry.

Nat Commun 2019 12 9;10(1):5622. Epub 2019 Dec 9.

Cancer Science Institute of Singapore, National University of Singapore, Singapore, 117599, Singapore.

A heritable polymorphism within regulatory sequences of the LMO1 gene is associated with its elevated expression and increased susceptibility to develop neuroblastoma, but the oncogenic pathways downstream of the LMO1 transcriptional co-regulatory protein are unknown. Our ChIP-seq and RNA-seq analyses reveal that a key gene directly regulated by LMO1 and MYCN is ASCL1, which encodes a basic helix-loop-helix transcription factor. Regulatory elements controlling ASCL1 expression are bound by LMO1, MYCN and the transcription factors GATA3, HAND2, PHOX2B, TBX2 and ISL1-all members of the adrenergic (ADRN) neuroblastoma core regulatory circuitry (CRC). ASCL1 is required for neuroblastoma cell growth and arrest of differentiation. ASCL1 and LMO1 directly regulate the expression of CRC genes, indicating that ASCL1 is a member and LMO1 is a coregulator of the ADRN neuroblastoma CRC.
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http://dx.doi.org/10.1038/s41467-019-13515-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6901540PMC
December 2019

The enhancer RNA activates the oncogenic transcriptional program in T-cell acute lymphoblastic leukemia.

Blood 2019 07 10;134(3):239-251. Epub 2019 May 10.

Cancer Science Institute of Singapore, National University of Singapore, Singapore.

The oncogenic transcription factor TAL1 regulates the transcriptional program in T-ALL. ARID5B is one of the critical downstream targets of TAL1, which further activates the oncogenic regulatory circuit in T-ALL cells. Here, we elucidated the molecular functions of the noncoding RNA, ARID5B-inducing enhancer associated long noncoding RNA (), in T-ALL pathogenesis. We demonstrated that is specifically activated in T-ALL cases, and its expression is associated with enhancer activity. recruits mediator proteins to the enhancer, promotes enhancer-promoter interactions, and activates the expression of , thereby positively regulating the TAL1-induced transcriptional program and the oncogene. The TAL1 complex coordinately regulates the expression of Knockdown of inhibits cell growth and survival of T-ALL cells in culture and blocks disease progression in a murine xenograft model. Our results indicate that plays an oncogenic role as an enhancer RNA in T-ALL.
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http://dx.doi.org/10.1182/blood.2018874503DOI Listing
July 2019

Genome-Wide Association Study of Susceptibility Loci for T-Cell Acute Lymphoblastic Leukemia in Children.

J Natl Cancer Inst 2019 12;111(12):1350-1357

Background: Acute lymphoblastic leukemia (ALL) is the most common cancer in children and can arise in B or T lymphoid lineages. Although risk loci have been identified for B-ALL, the inherited basis of T-ALL is mostly unknown, with a particular paucity of genome-wide investigation of susceptibility variants in large patient cohorts.

Methods: We performed a genome-wide association study (GWAS) in 1191 children with T-ALL and 12 178 controls, with independent replication using 117 cases and 5518 controls. The associations were tested using an additive logistic regression model. Top risk variants were tested for effects on enhancer activity using luciferase assay. All statistical tests were two sided.

Results: A novel risk locus in the USP7 gene (rs74010351, odds ratio [OR] = 1.44, 95% confidence interval [CI] = 1.27 to 1.65, P = 4.51 × 10-8) reached genome-wide significance in the discovery cohort, with independent validation (OR = 1.51, 95% CI = 1.03 to 2.22, P = .04). The USP7 risk allele was overrepresented in individuals of African descent, thus contributing to the higher incidence of T-ALL in this race/ethnic group. Genetic changes in USP7 (germline variants or somatic mutations) were observed in 56.4% of T-ALL with TAL1 overexpression, statistically significantly higher than in any other subtypes. Functional analyses suggested this T-ALL risk allele is located in a putative cis-regulatory DNA element with negative effects on USP7 transcription. Finally, comprehensive comparison of 14 susceptibility loci in T- vs B-ALL pointed to distinctive etiology of these leukemias.

Conclusions: These findings indicate strong associations between inherited genetic variation and T-ALL susceptibility in children and shed new light on the molecular etiology of ALL, particularly commonalities and differences in the biology of the two major subtypes (B- vs T-ALL).
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http://dx.doi.org/10.1093/jnci/djz043DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6910193PMC
December 2019

Oncogenic transcriptional program driven by TAL1 in T-cell acute lymphoblastic leukemia.

Int J Hematol 2019 Jan 25;109(1):5-17. Epub 2018 Aug 25.

Cancer Science Institute of Singapore, National University of Singapore, Centre for Translational Medicine, 14 Medical Drive, #12-01, Singapore, 117599, Singapore.

TAL1/SCL is a prime example of an oncogenic transcription factor that is abnormally expressed in acute leukemia due to the replacement of regulator elements. This gene has also been recognized as an essential regulator of hematopoiesis. TAL1 expression is strictly regulated in a lineage- and stage-specific manner. Such precise control is crucial for the switching of the transcriptional program. The misexpression of TAL1 in immature thymocytes leads to a widespread series of orchestrated downstream events that affect several different cellular machineries, resulting in a lethal consequence, namely T-cell acute lymphoblastic leukemia (T-ALL). In this article, we will discuss the transcriptional regulatory network and downstream target genes, including protein-coding genes and non-coding RNAs, controlled by TAL1 in normal hematopoiesis and T-cell leukemogenesis.
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http://dx.doi.org/10.1007/s12185-018-2518-zDOI Listing
January 2019

Dependency on the TYK2/STAT1/MCL1 axis in anaplastic large cell lymphoma.

Leukemia 2019 03 21;33(3):696-709. Epub 2018 Aug 21.

Cancer Science Institute of Singapore, National University of Singapore, 117599, Singapore, Singapore.

TYK2 is a member of the JAK family of tyrosine kinases that is involved in chromosomal translocation-induced fusion proteins found in anaplastic large cell lymphomas (ALCL) that lack rearrangements activating the anaplastic lymphoma kinase (ALK). Here we demonstrate that TYK2 is highly expressed in all cases of human ALCL, and that in a mouse model of NPM-ALK-induced lymphoma, genetic disruption of Tyk2 delays the onset of tumors and prolongs survival of the mice. Lymphomas in this model lacking Tyk2 have reduced STAT1 and STAT3 phosphorylation and reduced expression of Mcl1, a pro-survival member of the BCL2 family. These findings in mice are mirrored in human ALCL cell lines, in which TYK2 is activated by autocrine production of IL-10 and IL-22 and by interaction with specific receptors expressed by the cells. Activated TYK2 leads to STAT1 and STAT3 phosphorylation, activated expression of MCL1 and aberrant ALCL cell survival. Moreover, TYK2 inhibitors are able to induce apoptosis in ALCL cells, regardless of the presence or absence of an ALK-fusion. Thus, TYK2 is a dependency that is required for ALCL cell survival through activation of MCL1 expression. TYK2 represents an attractive drug target due to its essential enzymatic domain, and TYK2-specific inhibitors show promise as novel targeted inhibitors for ALCL.
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http://dx.doi.org/10.1038/s41375-018-0239-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8076043PMC
March 2019

[Understanding of molecular pathogenesis of T-cell leukemia by super-enhancer profiling].

Authors:
Takaomi Sanda

Rinsho Ketsueki 2018;59(7):899-908

Cancer Science Institute of Singapore, National University of Singapore.

Super-enhancers comprise of clusters of enhancers that are typically defined by the ChIP-seq analysis for active histone marks. Although the biological significance of super-enhancers is still controversial, this concept is gaining prominence as useful characteristics of genes that play crucial roles in normal development and pathogenesis of cancer. In various cancer cells, super-enhancers are often associated with genes involved in carcinogenesis. For example, in T-cell acute lymphoblastic leukemia, the oncogenic transcription factor TAL1 and its regulatory partners (GATA3, RUNX1 and MYB) are regulated by super-enhancers; these genes are sensitive to transcriptional inhibition, for example, via the pharmacological approach using a small-molecule CDK7 inhibitor. This preferential inhibition of cancer genes can also be observed for other types of cancer. Based on these findings, we recently performed super-enhancer profiling combined with gene expression analysis in adult T-cell leukemia/lymphoma, which is a genetically complicated hematological malignancy, to identify critical genes responsible for the pathogenesis. This review article aims to discuss the concept of super-enhancers, their significance in biomedical research, and their potential utility in elucidating the molecular pathogenesis of cancer.
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http://dx.doi.org/10.11406/rinketsu.59.899DOI Listing
July 2019

Cyclin-dependent kinase 9 as a potential specific molecular target in NK-cell leukemia/lymphoma.

Haematologica 2018 12 3;103(12):2059-2068. Epub 2018 Aug 3.

Department of Hematology and Oncology, Nagoya City University Graduate School of Medical Sciences, Japan.

BAY 1143572 is a highly selective inhibitor of cyclin-dependent kinase 9/positive transcription elongation factor b. It has entered phase I clinical studies. Here, we have assessed the utility of BAY 1143572 for treating natural killer (NK) cell leukemias/lymphomas that have a poor prognosis, namely extranodal NK/T-cell lymphoma, nasal type and aggressive NK-cell leukemia, in a preclinical mouse model in vivo as well as in tissue culture models Seven NK-cell leukemia/lymphoma lines and primary aggressive NK-cell leukemia cells from two individual patients were treated with BAY 1143572 Primary tumor cells from an aggressive NK-cell leukemia patient were used to establish a xenogeneic murine model for testing BAY 1143572 therapy. Cyclin-dependent kinase 9 inhibition by BAY 1143572 resulted in prevention of phosphorylation at the serine 2 site of the C-terminal domain of RNA polymerase II. This resulted in lower c-Myc and Mcl-1 levels in the cell lines, causing growth inhibition and apoptosis. In aggressive NK-cell leukemia primary tumor cells, exposure to BAY 1143572 resulted in decreased Mcl-1 protein levels resulting from inhibition of RNA polymerase II C-terminal domain phosphorylation at the serine 2 site. Orally administering BAY 1143572 once per day to aggressive NK-cell leukemia-bearing mice resulted in lower tumor cell infiltration into the bone marrow, liver, and spleen, with less export to the periphery relative to control mice. The treated mice also had a survival advantage over the untreated controls. The specific small molecule targeting agent BAY1143572 has potential for treating NK-cell leukemia/lymphoma.
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http://dx.doi.org/10.3324/haematol.2018.191395DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6269314PMC
December 2018

JDP2: An oncogenic bZIP transcription factor in T cell acute lymphoblastic leukemia.

J Exp Med 2018 07 25;215(7):1929-1945. Epub 2018 Jun 25.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA

A substantial subset of patients with T cell acute lymphoblastic leukemia (T-ALL) develops resistance to steroids and succumbs to their disease. encodes a bZIP protein that has been implicated as a T-ALL oncogene from insertional mutagenesis studies in mice, but its role in human T-ALL pathogenesis has remained obscure. Here we show that is aberrantly expressed in a subset of T-ALL patients and is associated with poor survival. JDP2 is required for T-ALL cell survival, as its depletion by short hairpin RNA knockdown leads to apoptosis. Mechanistically, JDP2 regulates prosurvival signaling through direct transcriptional regulation of Furthermore, is one of few oncogenes capable of initiating T-ALL in transgenic zebrafish. Notably, thymocytes from transgenic zebrafish express high levels of and demonstrate resistance to steroids in vivo. These studies establish as a novel oncogene in high-risk T-ALL and implicate overexpression of as a mechanism of steroid resistance in -overexpressing cells.
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http://dx.doi.org/10.1084/jem.20170484DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6028512PMC
July 2018

Targeting General Transcriptional Machinery as a Therapeutic Strategy for Adult T-Cell Leukemia.

Molecules 2018 May 2;23(5). Epub 2018 May 2.

Cancer Science Institute of Singapore, National University of Singapore, Singapore 117599, Singapore.

Cancer cells are highly reliant on certain molecular pathways, which support their survival and proliferation. The fundamental concept of molecularly targeted therapy is to target a protein that is specifically deregulated or overexpressed in cancer cells. However, drug resistance and tumor heterogeneity are major obstacles in the development of specific inhibitors. Additionally, many driver oncogenes exert their oncogenic property via abnormal expression without having genetic mutations. Interestingly, recent accumulating evidence has demonstrated that many critical cancer genes are driven by a unique class of enhancers termed super-enhancers. Genes associated with super-enhancers are relatively more susceptible to the inhibition of general transcriptional machinery compared with genes that are regulated by typical enhancers. Cancer cells are more sensitive to treatment with small-molecule inhibitors of CDK7 or BRD4 than non-transformed cells. These findings proposed a novel strategy to identify functionally important genes as well as novel therapeutic modalities in cancer. This approach would be particularly useful for genetically complicated cancers, such as adult T-cell leukemia (ATL), whereby a large mutational burden is present, but the functional consequences of each mutation have not been well-studied. In this review, we discuss recent findings on super-enhancers, underlying mechanisms, and the efficacy of small-molecule transcriptional inhibitors in ATL.
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http://dx.doi.org/10.3390/molecules23051057DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6099935PMC
May 2018

Identification of novel lncRNAs regulated by the TAL1 complex in T-cell acute lymphoblastic leukemia.

Leukemia 2018 10 26;32(10):2138-2151. Epub 2018 Mar 26.

Cancer Science Institute of Singapore, National University of Singapore, Singapore, 117599, Singapore.

TAL1/SCL is one of the most prevalent oncogenes in T-cell acute lymphoblastic leukemia (T-ALL). TAL1 and its regulatory partners (GATA3, RUNX1, and MYB) positively regulate each other and coordinately regulate the expression of their downstream target genes in T-ALL cells. However, long non-coding RNAs (lncRNAs) regulated by these factors are largely unknown. Here we established a bioinformatics pipeline and analyzed RNA-seq datasets with deep coverage to identify lncRNAs regulated by TAL1 in T-ALL cells. Our analysis predicted 57 putative lncRNAs that are activated by TAL1. Many of these transcripts were regulated by GATA3, RUNX1, and MYB in a coordinated manner. We identified two novel transcripts that were activated in multiple T-ALL cell samples but were downregulated in normal thymocytes. One transcript near the ARID5B gene locus was specifically expressed in TAL1-positive T-ALL cases. The other transcript located between the FAM49A and MYCN gene locus was also expressed in normal hematopoietic stem cells and T-cell progenitor cells. In addition, we identified a subset of lncRNAs that were negatively regulated by TAL1 and positively regulated by E-proteins in T-ALL cells. This included a known lncRNA (lnc-OAZ3-2:7) located near the RORC gene, which was expressed in normal thymocytes but repressed in TAL1-positive T-ALL cells.
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http://dx.doi.org/10.1038/s41375-018-0110-4DOI Listing
October 2018

ARID5B as a critical downstream target of the TAL1 complex that activates the oncogenic transcriptional program and promotes T-cell leukemogenesis.

Genes Dev 2017 12 11;31(23-24):2343-2360. Epub 2018 Jan 11.

Cancer Science Institute of Singapore, National University of Singapore, 117599 Singapore.

The oncogenic transcription factor induces an aberrant transcriptional program in T-cell acute lymphoblastic leukemia (T-ALL) cells. However, the critical factors that are directly activated by TAL1 and contribute to T-ALL pathogenesis are largely unknown. Here, we identified () as a collaborating oncogenic factor involved in the transcriptional program in T-ALL. expression is down-regulated at the double-negative 2-4 stages in normal thymocytes, while it is induced by the TAL1 complex in human T-ALL cells. The enhancer located 135 kb upstream of the gene locus is activated under a superenhancer in T-ALL cells but not in normal T cells. Notably, ARID5B-bound regions are associated predominantly with active transcription. ARID5B and TAL1 frequently co-occupy target genes and coordinately control their expression. ARID5B positively regulates the expression of TAL1 and its regulatory partners. ARID5B also activates the expression of the oncogene Importantly, ARID5B is required for the survival and growth of T-ALL cells and forced expression of ARID5B in immature thymocytes results in thymus retention, differentiation arrest, radioresistance, and tumor formation in zebrafish. Our results indicate that ARID5B reinforces the oncogenic transcriptional program by positively regulating the TAL1-induced regulatory circuit and in T-ALL, thereby contributing to T-cell leukemogenesis.
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http://dx.doi.org/10.1101/gad.302646.117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5795782PMC
December 2017

Leukemia-Initiating Cells in T-Cell Acute Lymphoblastic Leukemia.

Front Oncol 2017 25;7:218. Epub 2017 Sep 25.

Cancer Science Institute of Singapore, National University of Singapore, Singapore, Singapore.

T-cell acute lymphoblastic leukemia (T-ALL) is a hematological malignancy characterized by the clonal proliferation of immature T-cell precursors. T-ALL has many similar pathophysiological features to acute myeloid leukemia, which has been extensively studied in the establishment of the cancer stem cell (CSC) theory, but the CSC concept in T-ALL is still debatable. Although leukemia-initiating cells (LICs), which can generate leukemia in a xenograft setting, have been found in both human T-ALL patients and animal models, the nature and origin of LICs are largely unknown. In this review, we discuss recent studies on LICs in T-ALL and the potential mechanisms of LIC emergence in this disease. We focus on the oncogenic transcription factors , and and highlight the significance of the transcriptional regulatory programs in normal hematopoietic stem cells and T-ALL.
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http://dx.doi.org/10.3389/fonc.2017.00218DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5627022PMC
September 2017

RUNX1 in T-ALL: tumor suppressive or oncogenic?

Authors:
Takaomi Sanda

Blood 2017 10;130(15):1686-1688

NATIONAL UNIVERSITY OF SINGAPORE.

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http://dx.doi.org/10.1182/blood-2017-08-802181DOI Listing
October 2017

Enhancer profiling identifies critical cancer genes and characterizes cell identity in adult T-cell leukemia.

Blood 2017 11 4;130(21):2326-2338. Epub 2017 Oct 4.

Cancer Science Institute of Singapore, National University of Singapore, Singapore.

A number of studies have recently demonstrated that super-enhancers, which are large cluster of enhancers typically marked by a high level of acetylation of histone H3 lysine 27 and mediator bindings, are frequently associated with genes that control and define cell identity during normal development. Super-enhancers are also often enriched at cancer genes in various malignancies. The identification of such enhancers would pinpoint critical factors that directly contribute to pathogenesis. In this study, we performed enhancer profiling using primary leukemia samples from adult T-cell leukemia/lymphoma (ATL), which is a genetically heterogeneous intractable cancer. Super-enhancers were enriched at genes involved in the T-cell activation pathway, including , and in both ATL and normal mature T cells, which reflected the origin of the leukemic cells. Super-enhancers were found at several known cancer gene loci, including , and , in multiple ATL samples, but not in normal mature T cells, which implicated those genes in ATL pathogenesis. A small-molecule CDK7 inhibitor, THZ1, efficiently inhibited cell growth, induced apoptosis, and downregulated the expression of super-enhancer-associated genes in ATL cells. Furthermore, enhancer profiling combined with gene expression analysis identified a previously uncharacterized gene, , that was associated with super-enhancers in all ATL samples, but not in normal T cells. Knockdown of induced apoptosis in ATL cell lines, whereas overexpression of this gene promoted cell growth. Our study provides a novel strategy for identifying critical cancer genes.
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http://dx.doi.org/10.1182/blood-2017-06-792184DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5701524PMC
November 2017

TAL1 as a master oncogenic transcription factor in T-cell acute lymphoblastic leukemia.

Exp Hematol 2017 09 24;53:7-15. Epub 2017 Jun 24.

Cancer Science Institute of Singapore, National University of Singapore, Singapore.

In hematopoietic cell development, the transcriptional program is strictly regulated in a lineage- and stage-specific manner that requires a number of transcription factors to work in a cascade or in a loop, in addition to interactions with nonhematopoietic cells in the microenvironment. Disruption of the transcriptional program alters the cellular state and may predispose cells to the acquisition of genetic abnormalities. Early studies have shown that proteins that promote cell differentiation often serve as tumor suppressors, whereas inhibitors of those proteins act as oncogenes in the context of acute leukemia. A prime example is T-cell acute lymphoblastic leukemia (T-ALL), a malignant disorder characterized by clonal proliferation of immature stage thymocytes. Although a relatively small number of genetic abnormalities are observed in T-ALL, these abnormalities are crucial for leukemogenesis. Many oncogenes and tumor suppressors in T-ALL are transcription factors that are required for normal hematopoiesis. The transformation process in T-ALL is efficient and orchestrated; the oncogene disrupts the transcriptional program directing T-cell differentiation and also uses its native ability as a master transcription factor in hematopoiesis. This imbalance in the transcriptional program is a primary determinant underlying the molecular pathogenesis of T-ALL. In this review, we focus on the oncogenic transcription factor TAL1 and the tumor-suppressor E-proteins and discuss the malignant cell state, the transcriptional circuit, and the consequence of molecular abnormalities in T-ALL.
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http://dx.doi.org/10.1016/j.exphem.2017.06.001DOI Listing
September 2017

Roles of the RUNX1 Enhancer in Normal Hematopoiesis and Leukemogenesis.

Adv Exp Med Biol 2017 ;962:139-147

Cancer Science Institute of Singapore, National University of Singapore, Singapore, 117599, Singapore.

Enhancers are regulatory elements in genomic DNA that contain specific sequence motifs that are bound by DNA-binding transcription factors. The activity of enhancers is tightly regulated in an integrated and combinatorial manner, thus yielding complex patterns of transcription in different tissues. Identifying enhancers is crucial to understanding the physiological and pathogenic roles of their target genes. The RUNX1 intronic enhancer, eR1, acts in cis to regulate RUNX1 gene expression in hematopoietic stem cells (HSCs) and hemogenic endothelial cells. RUNX1 and other hematopoietic transcription factors TAL1/SCL, GATA2, PU.1, LMO2 and LDB1 bind at this region. Interestingly, recent studies have revealed that this region is involved in a large cluster of enhancers termed a super-enhancer. The RUNX1 super-enhancer is observed in normal HSCs and T-cell acute lymphoblastic leukemia cells. In this review, we describe the discovery of eR1 and its roles in normal development and leukemogenesis, as well as its potential applications in stem cell research.
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http://dx.doi.org/10.1007/978-981-10-3233-2_10DOI Listing
September 2017

Anti-leukaemic activity of the TYK2 selective inhibitor NDI-031301 in T-cell acute lymphoblastic leukaemia.

Br J Haematol 2017 04 14;177(2):271-282. Epub 2017 Mar 14.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, USA.

Activation of tyrosine kinase 2 (TYK2) contributes to the aberrant survival of T-cell acute lymphoblastic leukaemia (T-ALL) cells. Here we demonstrate the anti-leukaemic activity of a novel TYK2 inhibitor, NDI-031301. NDI-031301 is a potent and selective inhibitor of TYK2 that induced robust growth inhibition of human T-ALL cell lines. NDI-031301 treatment of human T-ALL cell lines resulted in induction of apoptosis that was not observed with the JAK inhibitors tofacitinib and baricitinib. Further investigation revealed that NDI-031301 treatment uniquely leads to activation of three mitogen-activated protein kinases (MAPKs), resulting in phosphorylation of ERK, SAPK/JNK and p38 MAPK coincident with PARP cleavage. Activation of p38 MAPK occurred within 1 h of NDI-031301 treatment and was responsible for NDI-031301-induced T-ALL cell death, as pharmacological inhibition of p38 MAPK partially rescued apoptosis induced by TYK2 inhibitor. Finally, daily oral administration of NDI-031301 at 100 mg/kg bid to immunodeficient mice engrafted with KOPT-K1 T-ALL cells was well tolerated, and led to decreased tumour burden and a significant survival benefit. These results support selective inhibition of TYK2 as a promising potential therapeutic strategy for T-ALL.
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http://dx.doi.org/10.1111/bjh.14563DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5384871PMC
April 2017

The KDM3A-KLF2-IRF4 axis maintains myeloma cell survival.

Nat Commun 2016 Jan 5;7:10258. Epub 2016 Jan 5.

Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts 02215, USA.

KDM3A is implicated in tumorigenesis; however, its biological role in multiple myeloma (MM) has not been elucidated. Here we identify KDM3A-KLF2-IRF4 axis dependence in MM. Knockdown of KDM3A is toxic to MM cells in vitro and in vivo. KDM3A maintains expression of KLF2 and IRF4 through H3K9 demethylation, and knockdown of KLF2 triggers apoptosis. Moreover, KLF2 directly activates IRF4 and IRF4 reciprocally upregulates KLF2, forming a positive autoregulatory circuit. The interaction of MM cells with bone marrow milieu mediates survival of MM cells. Importantly, silencing of KDM3A, KLF2 or IRF4 both decreases MM cell adhesion to bone marrow stromal cells and reduces MM cell homing to the bone marrow, in association with decreased ITGB7 expression in MAF-translocated MM cell lines. Our results indicate that the KDM3A-KLF2-IRF4 pathway plays an essential role in MM cell survival and homing to the bone marrow, and therefore represents a therapeutic target.
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http://dx.doi.org/10.1038/ncomms10258DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4728406PMC
January 2016

Inhibition of Wild-Type p53-Expressing AML by the Novel Small Molecule HDM2 Inhibitor CGM097.

Mol Cancer Ther 2015 Oct 23;14(10):2249-59. Epub 2015 Jul 23.

Dana Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts.

The tumor suppressor p53 is a key regulator of apoptosis and functions upstream in the apoptotic cascade by both indirectly and directly regulating Bcl-2 family proteins. In cells expressing wild-type (WT) p53, the HDM2 protein binds to p53 and blocks its activity. Inhibition of HDM2:p53 interaction activates p53 and causes apoptosis or cell-cycle arrest. Here, we investigated the ability of the novel HDM2 inhibitor CGM097 to potently and selectively kill WT p53-expressing AML cells. The antileukemic effects of CGM097 were studied using cell-based proliferation assays (human AML cell lines, primary AML patient cells, and normal bone marrow samples), apoptosis, and cell-cycle assays, ELISA, immunoblotting, and an AML patient-derived in vivo mouse model. CGM097 potently and selectively inhibited the proliferation of human AML cell lines and the majority of primary AML cells expressing WT p53, but not mutant p53, in a target-specific manner. Several patient samples that harbored mutant p53 were comparatively unresponsive to CGM097. Synergy was observed when CGM097 was combined with FLT3 inhibition against oncogenic FLT3-expressing cells cultured both in the absence as well as the presence of cytoprotective stromal-secreted cytokines, as well as when combined with MEK inhibition in cells with activated MAPK signaling. Finally, CGM097 was effective in reducing leukemia burden in vivo. These data suggest that CGM097 is a promising treatment for AML characterized as harboring WT p53 as a single agent, as well as in combination with other therapies targeting oncogene-activated pathways that drive AML.
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http://dx.doi.org/10.1158/1535-7163.MCT-15-0429DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4596780PMC
October 2015

Oncogene regulation. An oncogenic super-enhancer formed through somatic mutation of a noncoding intergenic element.

Science 2014 Dec 13;346(6215):1373-7. Epub 2014 Nov 13.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02215, USA. Division of Pediatric Hematology-Oncology, Boston Children's Hospital, MA 02115, USA.

In certain human cancers, the expression of critical oncogenes is driven from large regulatory elements, called super-enhancers, that recruit much of the cell's transcriptional apparatus and are defined by extensive acetylation of histone H3 lysine 27 (H3K27ac). In a subset of T-cell acute lymphoblastic leukemia (T-ALL) cases, we found that heterozygous somatic mutations are acquired that introduce binding motifs for the MYB transcription factor in a precise noncoding site, which creates a super-enhancer upstream of the TAL1 oncogene. MYB binds to this new site and recruits its H3K27 acetylase-binding partner CBP, as well as core components of a major leukemogenic transcriptional complex that contains RUNX1, GATA-3, and TAL1 itself. Additionally, most endogenous super-enhancers found in T-ALL cells are occupied by MYB and CBP, which suggests a general role for MYB in super-enhancer initiation. Thus, this study identifies a genetic mechanism responsible for the generation of oncogenic super-enhancers in malignant cells.
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http://dx.doi.org/10.1126/science.1259037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4720521PMC
December 2014

Molecular rationale for the use of PI3K/AKT/mTOR pathway inhibitors in combination with crizotinib in ALK-mutated neuroblastoma.

Oncotarget 2014 Sep;5(18):8737-49

Departments of Pediatric Hematology/Oncology, Dana-Farber Cancer Institute, Boston, MA.

Mutations in the ALK tyrosine kinase receptor gene represent important therapeutic targets in neuroblastoma, yet their clinical translation has been challenging. The ALK(F1174L) mutation is sensitive to the ALK inhibitor crizotinib only at high doses and mediates acquired resistance to crizotinib in ALK-translocated cancers. We have shown that the combination of crizotinib and an inhibitor of downstream signaling induces a favorable response in transgenic mice bearing ALK(F1174L)/MYCN-positive neuroblastoma. Here, we investigated the molecular basis of this effect and assessed whether a similar strategy would be effective in ALK-mutated tumors lacking MYCN overexpression. We show that in ALK-mutated, MYCN-amplified neuroblastoma cells, crizotinib alone does not affect mTORC1 activity as indicated by persistent RPS6 phosphorylation. Combined treatment with crizotinib and an ATP-competitive mTOR inhibitor abrogated RPS6 phosphorylation, leading to reduced tumor growth and prolonged survival in ALK(F1174L)/MYCN-positive models compared to single agent treatment. By contrast, this combination, while inducing mTORC1 downregulation, caused reciprocal upregulation of PI3K activity in ALK-mutated cells expressing wild-type MYCN. Here, an inhibitor with potency against both mTOR and PI3K was more effective in promoting cytotoxicity when combined with crizotinib. Our findings should enable a more precise selection of molecularly targeted agents for patients with ALK-mutated tumors.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4226718PMC
http://dx.doi.org/10.18632/oncotarget.2372DOI Listing
September 2014

Targeting transcription regulation in cancer with a covalent CDK7 inhibitor.

Nature 2014 Jul 22;511(7511):616-20. Epub 2014 Jun 22.

1] Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA [2] Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.

Tumour oncogenes include transcription factors that co-opt the general transcriptional machinery to sustain the oncogenic state, but direct pharmacological inhibition of transcription factors has so far proven difficult. However, the transcriptional machinery contains various enzymatic cofactors that can be targeted for the development of new therapeutic candidates, including cyclin-dependent kinases (CDKs). Here we present the discovery and characterization of a covalent CDK7 inhibitor, THZ1, which has the unprecedented ability to target a remote cysteine residue located outside of the canonical kinase domain, providing an unanticipated means of achieving selectivity for CDK7. Cancer cell-line profiling indicates that a subset of cancer cell lines, including human T-cell acute lymphoblastic leukaemia (T-ALL), have exceptional sensitivity to THZ1. Genome-wide analysis in Jurkat T-ALL cells shows that THZ1 disproportionally affects transcription of RUNX1 and suggests that sensitivity to THZ1 may be due to vulnerability conferred by the RUNX1 super-enhancer and the key role of RUNX1 in the core transcriptional regulatory circuitry of these tumour cells. Pharmacological modulation of CDK7 kinase activity may thus provide an approach to identify and treat tumour types that are dependent on transcription for maintenance of the oncogenic state.
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http://dx.doi.org/10.1038/nature13393DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4244910PMC
July 2014

Ribonucleoprotein HNRNPA2B1 interacts with and regulates oncogenic KRAS in pancreatic ductal adenocarcinoma cells.

Gastroenterology 2014 Oct 3;147(4):882-892.e8. Epub 2014 Jul 3.

Departament de Biologia Cellular, Immunologia i Neurociències, Facultat de Medicina, IDIBAPS, Universitat de Barcelona, Barcelona, Spain. Electronic address:

Background & Aims: Development of pancreatic ductal adenocarcinoma (PDAC) involves activation of c-Ki-ras2 Kirsten rat sarcoma oncogene homolog (KRAS) signaling, but little is known about the roles of proteins that regulate the activity of oncogenic KRAS. We investigated the activities of proteins that interact with KRAS in PDAC cells.

Methods: We used mass spectrometry to demonstrate that heterogeneous nuclear ribonucleoproteins (HNRNP) A2 and B1 (encoded by the gene HNRNPA2B1) interact with KRAS G12V. We used co-immunoprecipitation analyses to study interactions between HNRNPA2B1 and KRAS in KRAS-dependent and KRAS-independent PDAC cell lines. We knocked down HNRNPA2B1 using small hairpin RNAs and measured viability, anchorage-independent proliferation, and growth of xenograft tumors in mice. We studied KRAS phosphorylation using the Phos-tag system.

Results: We found that interactions between HRNPA2B1 and KRAS correlated with KRAS-dependency of some human PDAC cell lines. Knock down of HNRNPA2B1 significantly reduced viability, anchorage-independent proliferation, and formation of xenograft tumors by KRAS-dependent PDAC cells. HNRNPA2B1 knock down also increased apoptosis of KRAS-dependent PDAC cells, inactivated c-akt murine thymoma oncogene homolog 1 signaling via mammalian target of rapamycin, and reduced interaction between KRAS and phosphatidylinositide 3-kinase. Interaction between HNRNPA2B1 and KRAS required KRAS phosphorylation at serine 181.

Conclusions: In KRAS-dependent PDAC cell lines, HNRNPA2B1 interacts with and regulates the activity of KRAS G12V and G12D. HNRNPA2B1 is required for KRAS activation of c-akt murine thymoma oncogene homolog 1-mammalian target of rapamycin signaling, interaction with phosphatidylinositide 3-kinase, and PDAC cell survival and tumor formation in mice. HNRNPA2B1 might be a target for treatment of pancreatic cancer.
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http://dx.doi.org/10.1053/j.gastro.2014.06.041DOI Listing
October 2014