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Sahoo SS, Khiami M, Wlodarski MW. Inducible pluripotent stem cell models to study bone marrow failure and MDS predisposition syndromes. Exp Hematol 2025; 143:104669. [PMID: 39491640 DOI: 10.1016/j.exphem.2024.104669] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/02/2024] [Revised: 10/24/2024] [Accepted: 10/26/2024] [Indexed: 11/05/2024]
Abstract
Induced pluripotent stem cells (iPSCs) have emerged as powerful tools for in vitro modeling of bone marrow failure (BMF) syndromes and hereditary conditions predisposing to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). This review synthesizes recent advances in iPSC-based disease modeling for various inherited BMF/MDS disorders, including Fanconi anemia, dyskeratosis congenita, Diamond Blackfan anemia syndrome, Shwachman-Diamond syndrome, and severe congenital neutropenia as well as GATA2, RUNX1, ETV6, ANKRD26, SAMD9, SAMD9L, and ADH5/ALDH2 syndromes. Although the majority of these iPSC lines are derived from patient cells, some are generated by introducing patient-specific mutations into healthy iPSC backgrounds, offering complementary approaches to disease modeling. The review highlights the ability of iPSCs to recapitulate key disease phenotypes, such as impaired hematopoietic differentiation, telomere dysfunction, and defects in DNA repair or ribosome biogenesis. We discuss how these models have enhanced our understanding of disease pathomechanisms, hematopoietic defects, and potential therapeutic approaches. Challenges in generating and maintaining disease-specific iPSCs are examined, particularly for disorders involving DNA repair. We emphasize the necessity of creating isogenic controls to elucidate genotype-phenotype relationships. Furthermore, we address limitations of current iPSC models, including genetic variability among iPSC clones derived from the same patient, and difficulties in achieving robust engraftment of iPSC-derived hematopoietic progenitor cells in mouse transplantation models. The review also explores future directions, including the potential of iPSC models for drug discovery and personalized medicine approaches. This review underscores the significance of iPSC technology in advancing our understanding of inherited hematopoietic disorders and its potential to inform novel therapeutic strategies.
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Affiliation(s)
- Sushree S Sahoo
- Department of Hematology, St. Jude Children's Research Hospital, Memphis, TN
| | - Majd Khiami
- Department of Hematology, St. Jude Children's Research Hospital, Memphis, TN
| | - Marcin W Wlodarski
- Department of Hematology, St. Jude Children's Research Hospital, Memphis, TN.
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2
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Mohammadhosseini M, Enright T, Duvall A, Chitsazan A, Lin HY, Ors A, Davis BA, Nikolova O, Bresciani E, Diemer J, Craft K, Menezes AC, Merguerian M, Chong S, Calvo KR, Deuitch NT, Glushakow-Smith S, Gritsman K, Godley LA, Horwitz MS, Keel S, Castilla LH, Demir E, Mohammed H, Liu P, Agarwal A. Targeting the CD74 signaling axis suppresses inflammation and rescues defective hematopoiesis in RUNX1-familial platelet disorder. Sci Transl Med 2025; 17:eadn9832. [PMID: 39772771 PMCID: PMC11912227 DOI: 10.1126/scitranslmed.adn9832] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2024] [Revised: 08/11/2024] [Accepted: 11/03/2024] [Indexed: 01/30/2025]
Abstract
Familial platelet disorder (FPD) is associated with germline RUNX1 mutations, establishing a preleukemic state and increasing the risk of developing leukemia. Currently, there are no intervention strategies to prevent leukemia progression. Single-cell RNA sequencing (n = 10) combined with functional analysis of samples from patients with RUNX1-FPD (n > 75) revealed that FPD hematopoietic stem and progenitor cells (HSPCs) displayed increased myeloid differentiation and suppressed megakaryopoiesis because of increased activation of prosurvival and inflammatory pathways. Bone marrow from patients with RUNX1-FPD contained an elevated cytokine milieu, exerting chronic inflammatory stress on HSPCs. RUNX1-FPD HSPCs were myeloid biased, had increased self-renewal, and were resistant to inflammation-mediated exhaustion. The bone marrow from patients with RUNX1-FPD showed high transcript and protein expression of CD74 at the preleukemic stage compared with that of healthy controls, which remained high upon patient transformation into leukemia. Further, CD74-mediated signaling was exaggerated in RUNX1-FPD HSPCs compared with healthy controls, leading to the activation of mTOR and JAK/STAT pathways with increased cytokine production. Genetic and pharmacological targeting of CD74 with ISO-1 and its downstream targets JAK1/2 and mTOR reversed RUNX1-FPD differentiation defects in vitro and in vivo and reduced inflammation. Our results highlight that inflammation is an early event in RUNX1-FPD pathogenesis, and CD74 signaling is one of the drivers of this inflammation. The repurposing of JAK1/2i (ruxolitinib) and mTORi (sirolimus) and promoting the advancement of CD74 inhibitors in clinical settings as an early intervention strategy would be beneficial to improve the phenotype of patients with RUNX1-FPD and prevent myeloid progression.
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Affiliation(s)
- Mona Mohammadhosseini
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
- Division of Oncological Sciences, Oregon Health & Science University, Portland, OR 97239, USA
| | - Trevor Enright
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
- Cancer Early Detection Advanced Research Center, Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97201, USA
| | - Adam Duvall
- Department of Medicine, University of Chicago, Chicago, IL 60637, USA
| | - Alex Chitsazan
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
- Cancer Early Detection Advanced Research Center, Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97201, USA
| | - Hsin-Yun Lin
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
- Division of Oncological Sciences, Oregon Health & Science University, Portland, OR 97239, USA
| | - Aysegul Ors
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
- Cancer Early Detection Advanced Research Center, Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97201, USA
| | - Brett A Davis
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
| | - Olga Nikolova
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
- Division of Oncological Sciences, Oregon Health & Science University, Portland, OR 97239, USA
| | - Erica Bresciani
- Oncogenesis and Development Section, Division of Intramural Research, National Human Genome Research Institute, Bethesda, MD 20894, USA
| | - Jamie Diemer
- Oncogenesis and Development Section, Division of Intramural Research, National Human Genome Research Institute, Bethesda, MD 20894, USA
| | - Kathleen Craft
- Oncogenesis and Development Section, Division of Intramural Research, National Human Genome Research Institute, Bethesda, MD 20894, USA
| | - Ana Catarina Menezes
- Oncogenesis and Development Section, Division of Intramural Research, National Human Genome Research Institute, Bethesda, MD 20894, USA
| | - Matthew Merguerian
- Oncogenesis and Development Section, Division of Intramural Research, National Human Genome Research Institute, Bethesda, MD 20894, USA
- Department of Pediatrics, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Shawn Chong
- Oncogenesis and Development Section, Division of Intramural Research, National Human Genome Research Institute, Bethesda, MD 20894, USA
| | - Katherine R Calvo
- Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, MD 20892, USA
| | - Natalie T Deuitch
- Oncogenesis and Development Section, Division of Intramural Research, National Human Genome Research Institute, Bethesda, MD 20894, USA
| | | | - Kira Gritsman
- Albert Einstein College of Medicine, Bronx, NY 10461, USA
| | - Lucy A Godley
- Division of Hematology/Oncology, Department of Medicine, Robert H. Lurie Comprehensive Cancer Center, Northwestern University, Chicago, IL 60611, USA
| | - Marshall S Horwitz
- Department of Laboratory Medicine & Pathology, University of Washington School of Medicine, Seattle, WA 98195, USA
| | - Sioban Keel
- Department of Medicine, University of Washington School of Medicine, Seattle, WA 98195, USA
| | - Lucio H Castilla
- Department of Molecular, Cell, and Cancer Biology, University of Massachusetts Chan Medical School, Worcester, MA 01655, USA
| | - Emek Demir
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
- Division of Oncological Sciences, Oregon Health & Science University, Portland, OR 97239, USA
- Cancer Early Detection Advanced Research Center, Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97201, USA
| | - Hisham Mohammed
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
- Cancer Early Detection Advanced Research Center, Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97201, USA
- Department of Molecular and Medical Genetics, Oregon Health & Science University, Portland, OR 97201, USA
| | - Paul Liu
- Oncogenesis and Development Section, Division of Intramural Research, National Human Genome Research Institute, Bethesda, MD 20894, USA
| | - Anupriya Agarwal
- Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA
- Division of Oncological Sciences, Oregon Health & Science University, Portland, OR 97239, USA
- Cancer Early Detection Advanced Research Center, Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97201, USA
- Department of Molecular and Medical Genetics, Oregon Health & Science University, Portland, OR 97201, USA
- Department of Cell, Developmental, and Cancer Biology, Oregon Health & Science University, Portland, OR 97239, USA
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3
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Tanaka Y, Nakanishi Y, Furuhata E, Nakada KI, Maruyama R, Suzuki H, Suzuki T. FLI1 is associated with regulation of DNA methylation and megakaryocytic differentiation in FPDMM caused by a RUNX1 transactivation domain mutation. Sci Rep 2024; 14:14080. [PMID: 38890442 PMCID: PMC11189521 DOI: 10.1038/s41598-024-64829-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2024] [Accepted: 06/13/2024] [Indexed: 06/20/2024] Open
Abstract
Familial platelet disorder with associated myeloid malignancies (FPDMM) is an autosomal dominant disease caused by heterozygous germline mutations in RUNX1. It is characterized by thrombocytopenia, platelet dysfunction, and a predisposition to hematological malignancies. Although FPDMM is a precursor for diseases involving abnormal DNA methylation, the DNA methylation status in FPDMM remains unknown, largely due to a lack of animal models and challenges in obtaining patient-derived samples. Here, using genome editing techniques, we established two lines of human induced pluripotent stem cells (iPSCs) with different FPDMM-mimicking heterozygous RUNX1 mutations. These iPSCs showed defective differentiation of hematopoietic progenitor cells (HPCs) and megakaryocytes (Mks), consistent with FPDMM. The FPDMM-mimicking HPCs showed DNA methylation patterns distinct from those of wild-type HPCs, with hypermethylated regions showing the enrichment of ETS transcription factor (TF) motifs. We found that the expression of FLI1, an ETS family member, was significantly downregulated in FPDMM-mimicking HPCs with a RUNX1 transactivation domain (TAD) mutation. We demonstrated that FLI1 promoted binding-site-directed DNA demethylation, and that overexpression of FLI1 restored their megakaryocytic differentiation efficiency and hypermethylation status. These findings suggest that FLI1 plays a crucial role in regulating DNA methylation and correcting defective megakaryocytic differentiation in FPDMM-mimicking HPCs with a RUNX1 TAD mutation.
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Affiliation(s)
- Yuki Tanaka
- Laboratory for Cellular Function Conversion Technology, RIKEN Center for Integrative Medical Sciences (IMS), RIKEN Yokohama Campus, 1-7-22 Suehiro-Cho, Tsurumi-Ku, Yokohama City, Kanagawa, 230-0045, Japan
| | - Yuri Nakanishi
- Laboratory for Cellular Function Conversion Technology, RIKEN Center for Integrative Medical Sciences (IMS), RIKEN Yokohama Campus, 1-7-22 Suehiro-Cho, Tsurumi-Ku, Yokohama City, Kanagawa, 230-0045, Japan
| | - Erina Furuhata
- Laboratory for Cellular Function Conversion Technology, RIKEN Center for Integrative Medical Sciences (IMS), RIKEN Yokohama Campus, 1-7-22 Suehiro-Cho, Tsurumi-Ku, Yokohama City, Kanagawa, 230-0045, Japan
| | - Ken-Ichi Nakada
- Laboratory for Cellular Function Conversion Technology, RIKEN Center for Integrative Medical Sciences (IMS), RIKEN Yokohama Campus, 1-7-22 Suehiro-Cho, Tsurumi-Ku, Yokohama City, Kanagawa, 230-0045, Japan
- Graduate School of Medical Life Science, Yokohama City University, 1-7-29 Suehiro-Cho, Tsurumi-Ku, Yokohama City, Kanagawa, 230-0045, Japan
| | - Rino Maruyama
- Laboratory for Cellular Function Conversion Technology, RIKEN Center for Integrative Medical Sciences (IMS), RIKEN Yokohama Campus, 1-7-22 Suehiro-Cho, Tsurumi-Ku, Yokohama City, Kanagawa, 230-0045, Japan
- Graduate School of Medical Life Science, Yokohama City University, 1-7-29 Suehiro-Cho, Tsurumi-Ku, Yokohama City, Kanagawa, 230-0045, Japan
| | - Harukazu Suzuki
- Laboratory for Cellular Function Conversion Technology, RIKEN Center for Integrative Medical Sciences (IMS), RIKEN Yokohama Campus, 1-7-22 Suehiro-Cho, Tsurumi-Ku, Yokohama City, Kanagawa, 230-0045, Japan
| | - Takahiro Suzuki
- Laboratory for Cellular Function Conversion Technology, RIKEN Center for Integrative Medical Sciences (IMS), RIKEN Yokohama Campus, 1-7-22 Suehiro-Cho, Tsurumi-Ku, Yokohama City, Kanagawa, 230-0045, Japan.
- Department of Obstetrics & Gynecology, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-Ku, Tokyo, 113-8421, Japan.
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Jiang YZ, Hu LY, Chen MS, Wang XJ, Tan CN, Xue PP, Yu T, He XY, Xiang LX, Xiao YN, Li XL, Ran Q, Li ZJ, Chen L. GATA binding protein 2 mediated ankyrin repeat domain containing 26 high expression in myeloid-derived cell lines. World J Stem Cells 2024; 16:538-550. [PMID: 38817334 PMCID: PMC11135246 DOI: 10.4252/wjsc.v16.i5.538] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/26/2023] [Revised: 03/12/2024] [Accepted: 04/12/2024] [Indexed: 05/24/2024] Open
Abstract
BACKGROUND Thrombocytopenia 2, an autosomal dominant inherited disease characterized by moderate thrombocytopenia, predisposition to myeloid malignancies and normal platelet size and function, can be caused by 5'-untranslated region (UTR) point mutations in ankyrin repeat domain containing 26 (ANKRD26). Runt related transcription factor 1 (RUNX1) and friend leukemia integration 1 (FLI1) have been identified as negative regulators of ANKRD26. However, the positive regulators of ANKRD26 are still unknown. AIM To prove the positive regulatory effect of GATA binding protein 2 (GATA2) on ANKRD26 transcription. METHODS Human induced pluripotent stem cells derived from bone marrow (hiPSC-BM) and urothelium (hiPSC-U) were used to examine the ANKRD26 expression pattern in the early stage of differentiation. Then, transcriptome sequencing of these iPSCs and three public transcription factor (TF) databases (Cistrome DB, animal TFDB and ENCODE) were used to identify potential TF candidates for ANKRD26. Furthermore, overexpression and dual-luciferase reporter experiments were used to verify the regulatory effect of the candidate TFs on ANKRD26. Moreover, using the GENT2 platform, we analyzed the relationship between ANKRD26 expression and overall survival in cancer patients. RESULTS In hiPSC-BMs and hiPSC-Us, we found that the transcription levels of ANKRD26 varied in the absence of RUNX1 and FLI1. We sequenced hiPSC-BM and hiPSC-U and identified 68 candidate TFs for ANKRD26. Together with three public TF databases, we found that GATA2 was the only candidate gene that could positively regulate ANKRD26. Using dual-luciferase reporter experiments, we showed that GATA2 directly binds to the 5'-UTR of ANKRD26 and promotes its transcription. There are two identified binding sites of GATA2 that are located 2 kb upstream of the TSS of ANKRD26. In addition, we discovered that high ANKRD26 expression is always related to a more favorable prognosis in breast and lung cancer patients. CONCLUSION We first discovered that the transcription factor GATA2 plays a positive role in ANKRD26 transcription and identified its precise binding sites at the promoter region, and we revealed the importance of ANKRD26 in many tissue-derived cancers.
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Affiliation(s)
- Yang-Zhou Jiang
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Lan-Yue Hu
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Mao-Shan Chen
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Xiao-Jie Wang
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Cheng-Ning Tan
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Pei-Pei Xue
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Teng Yu
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Xiao-Yan He
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Li-Xin Xiang
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Yan-Ni Xiao
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Xiao-Liang Li
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Qian Ran
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Zhong-Jun Li
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Li Chen
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China.
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Lee BC, Zhou Y, Bresciani E, Ozkaya N, Dulau-Florea A, Carrington B, Shin TH, Baena V, Syed ZA, Hong SG, Zhen T, Calvo KR, Liu P, Dunbar CE. A RUNX1-FPDMM rhesus macaque model reproduces the human phenotype and predicts challenges to curative gene therapies. Blood 2023; 141:231-237. [PMID: 36322931 PMCID: PMC9936307 DOI: 10.1182/blood.2022018193] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2022] [Revised: 10/14/2022] [Accepted: 10/19/2022] [Indexed: 11/06/2022] Open
Abstract
Germ line loss-of-function heterozygous mutations in the RUNX1 gene cause familial platelet disorder with associated myeloid malignancies (FPDMM) characterized by thrombocytopenia and a life-long risk of hematological malignancies. Although gene therapies are being considered as promising therapeutic options, current preclinical models do not recapitulate the human phenotype and are unable to elucidate the relative fitness of mutation-corrected and RUNX1-heterozygous mutant hematopoietic stem and progenitor cells (HSPCs) in vivo long term. We generated a rhesus macaque with an FPDMM competitive repopulation model using CRISPR/Cas9 nonhomologous end joining editing in the RUNX1 gene and the AAVS1 safe-harbor control locus. We transplanted mixed populations of edited autologous HSPCs and tracked mutated allele frequencies in blood cells. In both animals, RUNX1-edited cells expanded over time compared with AAVS1-edited cells. Platelet counts remained below the normal range in the long term. Bone marrows developed megakaryocytic dysplasia similar to human FPDMM, and CD34+ HSPCs showed impaired in vitro megakaryocytic differentiation, with a striking defect in polyploidization. In conclusion, the lack of a competitive advantage for wildtype or control-edited HSPCs over RUNX1 heterozygous-mutated HSPCs long term in our preclinical model suggests that gene correction approaches for FPDMM will be challenging, particularly to reverse myelodysplastic syndrome/ acute myeloid leukemia predisposition and thrombopoietic defects.
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Affiliation(s)
- Byung-Chul Lee
- Translational Stem Cell Biology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD
| | - Yifan Zhou
- Translational Stem Cell Biology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD
- Wellcome-MRC Cambridge Stem Cell Institute, University of Cambridge, Puddicombe Way, Cambridge, United Kingdom
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom
| | - Erica Bresciani
- Oncogenesis and Development Section, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD
| | - Neval Ozkaya
- Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, NY
| | - Alina Dulau-Florea
- Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, MD
| | - Blake Carrington
- Zebrafish Core, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD
| | - Tae-Hoon Shin
- Translational Stem Cell Biology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD
- Department of Laboratory Animal Medicine, College of Veterinary Medicine, Jeju National University, Jeju, Republic of Korea
| | - Valentina Baena
- Electron Microscopy Core, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD
| | - Zulfeqhar A. Syed
- Electron Microscopy Core, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD
| | - So Gun Hong
- Translational Stem Cell Biology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD
| | - Tao Zhen
- Oncogenesis and Development Section, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD
| | - Katherine R. Calvo
- Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, MD
| | - Paul Liu
- Oncogenesis and Development Section, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD
| | - Cynthia E. Dunbar
- Translational Stem Cell Biology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD
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6
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Lee K, Ahn HS, Estevez B, Poncz M. RUNX1-deficient human megakaryocytes demonstrate thrombopoietic and platelet half-life and functional defects. Blood 2023; 141:260-270. [PMID: 36219879 PMCID: PMC9936297 DOI: 10.1182/blood.2022017561] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2022] [Revised: 09/16/2022] [Accepted: 09/23/2022] [Indexed: 01/24/2023] Open
Abstract
Heterozygous defects in runt-related transcription factor 1 (RUNX1) are causative of a familial platelet disorder with associated myeloid malignancy (FPDMM). Because RUNX1-deficient animal models do not mimic bleeding disorder or leukemic risk associated with FPDMM, development of a proper model system is critical to understanding the underlying mechanisms of the observed phenotype and to identifying therapeutic interventions. We previously reported an in vitro megakaryopoiesis system comprising human CD34+ hematopoietic stem and progenitor cells that recapitulated the FPDMM quantitative megakaryocyte defect through a decrease in RUNX1 expression via a lentiviral short hairpin RNA strategy. We now show that shRX-megakaryocytes have a marked reduction in agonist responsiveness. We then infused shRX-megakaryocytes into immunocompromised NOD scid gamma (NSG) mice and demonstrated that these megakaryocytes released fewer platelets than megakaryocytes transfected with a nontargeting shRNA, and these platelets had a diminished half-life. The platelets were also poorly responsive to agonists, unable to correct thrombus formation in NSG mice homozygous for a R1326H mutation in von Willebrand Factor (VWFR1326H), which switches the species-binding specificity of the VWF from mouse to human glycoprotein Ibα. A small-molecule inhibitor RepSox, which blocks the transforming growth factor β1 (TGFβ1) pathway and rescued defective megakaryopoiesis in vitro, corrected the thrombopoietic defect, defects in thrombus formation and platelet half-life, and agonist response in NSG/VWFR1326H mice. Thus, this model recapitulates the defects in FPDMM megakaryocytes and platelets, identifies previously unrecognized defects in thrombopoiesis and platelet half-life, and demonstrates for the first time, reversal of RUNX1 deficiency-induced hemostatic defects by a drug.
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Affiliation(s)
- Kiwon Lee
- Division of Hematology, Children’s Hospital of Philadelphia, Philadelphia, PA
| | - Hyun Sook Ahn
- Division of Hematology, Children’s Hospital of Philadelphia, Philadelphia, PA
| | - Brian Estevez
- Division of Hematology, Children’s Hospital of Philadelphia, Philadelphia, PA
| | - Mortimer Poncz
- Division of Hematology, Children’s Hospital of Philadelphia, Philadelphia, PA
- Department of Pediatrics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA
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7
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Zhang S, Qu K, Lyu S, Hoyle DL, Smith C, Cheng L, Cheng T, Shen J, Wang ZZ. PEAR1 is a potential regulator of early hematopoiesis of human pluripotent stem cells. J Cell Physiol 2023; 238:179-194. [PMID: 36436185 DOI: 10.1002/jcp.30924] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2022] [Revised: 10/17/2022] [Accepted: 10/28/2022] [Indexed: 11/28/2022]
Abstract
Hemogenic endothelial (HE) cells are specialized endothelial cells to give rise to hematopoietic stem/progenitor cells during hematopoietic development. The underlying mechanisms that regulate endothelial-to-hematopoietic transition (EHT) of human HE cells are not fully understand. Here, we identified platelet endothelial aggregation receptor-1 (PEAR1) as a novel regulator of early hematopoietic development in human pluripotent stem cells (hPSCs). We found that the expression of PEAP1 was elevated during hematopoietic development. A subpopulation of PEAR1+ cells overlapped with CD34+ CD144+ CD184+ CD73- arterial-type HE cells. Transcriptome analysis by RNA sequencing indicated that TAL1/SCL, GATA2, MYB, RUNX1 and other key transcription factors for hematopoietic development were mainly expressed in PEAR1+ cells, whereas the genes encoding for niche-related signals, such as fibronectin, vitronectin, bone morphogenetic proteins and jagged1, were highly expressed in PEAR1- cells. The isolated PEAR1+ cells exhibited significantly greater EHT capacity on endothelial niche, compared with the PEAR1- cells. Colony-forming unit (CFU) assays demonstrated the multilineage hematopoietic potential of PEAR1+ -derived hematopoietic cells. Furthermore, PEAR1 knockout in hPSCs by CRISPR/Cas9 technology revealed that the hematopoietic differentiation was impaired, resulting in decreased EHT capacity, decreased expression of hematopoietic-related transcription factors, and increased expression of niche-related signals. In summary, this study revealed a novel role of PEAR1 in balancing intrinsic and extrinsic signals for early hematopoietic fate decision.
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Affiliation(s)
- Shuo Zhang
- State Key Laboratory of Experimental Hematology, National Clinical Research Center for Blood Diseases, Haihe Laboratory of Cell Ecosystem, Institute of Hematology & Blood Diseases Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Tianjin, China.,Division of Hematology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.,School of Traditional Chinese Medicine, Beijing University of Chinese Medicine, Beijing, China
| | - Kengyuan Qu
- State Key Laboratory of Experimental Hematology, National Clinical Research Center for Blood Diseases, Haihe Laboratory of Cell Ecosystem, Institute of Hematology & Blood Diseases Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Tianjin, China.,Department of Histology and Embryology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, China
| | - Shuzhen Lyu
- State Key Laboratory of Experimental Hematology, National Clinical Research Center for Blood Diseases, Haihe Laboratory of Cell Ecosystem, Institute of Hematology & Blood Diseases Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Tianjin, China
| | - Dixie L Hoyle
- Division of Hematology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| | - Cory Smith
- Division of Hematology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| | - Linzhao Cheng
- Division of Hematology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.,Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, China
| | - Tao Cheng
- State Key Laboratory of Experimental Hematology, National Clinical Research Center for Blood Diseases, Haihe Laboratory of Cell Ecosystem, Institute of Hematology & Blood Diseases Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Tianjin, China.,Center for Stem Cell Medicine, Chinese Academy of Medical Sciences, Tianjin, China.,Department of Stem Cell & Regenerative Medicine, Peking Union Medical College, Tianjin, China
| | - Jun Shen
- State Key Laboratory of Experimental Hematology, National Clinical Research Center for Blood Diseases, Haihe Laboratory of Cell Ecosystem, Institute of Hematology & Blood Diseases Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Tianjin, China
| | - Zack Z Wang
- Division of Hematology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
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8
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Nations CC, Pavani G, French DL, Gadue P. Modeling genetic platelet disorders with human pluripotent stem cells: mega-progress but wanting more on our plate(let). Curr Opin Hematol 2021; 28:308-314. [PMID: 34397590 PMCID: PMC8371829 DOI: 10.1097/moh.0000000000000671] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
PURPOSE OF REVIEW Megakaryocytes are rare hematopoietic cells that play an instrumental role in hemostasis, and other important biological processes such as immunity and wound healing. With the advent of cell reprogramming technologies and advances in differentiation protocols, it is now possible to obtain megakaryocytes from any pluripotent stem cell (PSC) via hematopoietic induction. Here, we review recent advances in PSC-derived megakaryocyte (iMK) technology, focusing on platform validation, disease modeling and current limitations. RECENT FINDINGS A comprehensive study confirmed that iMK can recapitulate many transcriptional and functional aspects of megakaryocyte and platelet biology, including variables associated with complex genetic traits such as sex and race. These findings were corroborated by several pathological models in which iMKs revealed molecular mechanisms behind inherited platelet disorders and assessed the efficacy of novel pharmacological interventions. However, current differentiation protocols generate primarily embryonic iMK, limiting the clinical and translational potential of this system. SUMMARY iMK are strong candidates to model pathologic mutations involved in platelet defects and develop innovative therapeutic strategies. Future efforts on generating definitive hematopoietic progenitors would improve current platelet generation protocols and expand our capacity to model neonatal and adult megakaryocyte disorders.
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Affiliation(s)
- Catriana C Nations
- Department of Cell and Molecular Biology, University of Pennsylvania Perelman School of Medicine
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia
| | - Giulia Pavani
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia
| | - Deborah L French
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia
- Department of Pathology and Laboratory Medicine, University of Pennsylvania Perelman School of Medicine and Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, USA
| | - Paul Gadue
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia
- Department of Pathology and Laboratory Medicine, University of Pennsylvania Perelman School of Medicine and Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, USA
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9
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Li Y, Yang W, Devidas M, Winter SS, Kesserwan C, Yang W, Dunsmore KP, Smith C, Qian M, Zhao X, Zhang R, Gastier-Foster JM, Raetz EA, Carroll WL, Li C, Liu PP, Rabin KR, Sanda T, Mullighan CG, Nichols KE, Evans WE, Pui CH, Hunger SP, Teachey DT, Relling MV, Loh ML, Yang JJ. Germline RUNX1 variation and predisposition to childhood acute lymphoblastic leukemia. J Clin Invest 2021; 131:147898. [PMID: 34166225 PMCID: PMC8409579 DOI: 10.1172/jci147898] [Citation(s) in RCA: 26] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2021] [Accepted: 06/22/2021] [Indexed: 12/31/2022] Open
Abstract
Genetic alterations in the RUNX1 gene are associated with benign and malignant blood disorders, particularly of megakaryocyte and myeloid lineages. The role of RUNX1 in acute lymphoblastic leukemia (ALL) is less clear, particularly how germline genetic variation influences the predisposition to this type of leukemia. Sequencing 4,836 children with B-ALL and 1,354 cases of T-ALL, we identified 31 and 18 germline RUNX1 variants, respectively. RUNX1 variants in B-ALL consistently showed minimal damaging effects. By contrast, 6 T-ALL-related variants result in drastic loss of RUNX1 activity as a transcription activator in vitro. Ectopic expression of dominant-negative RUNX1 variants in human CD34+ cells repressed differentiation into erythroid, megakaryocytes, and T cells, while promoting myeloid cell development. Chromatin immunoprecipitation sequencing of T-ALL models showed distinctive patterns of RUNX1 binding by variant proteins. Further whole genome sequencing identified JAK3 mutation as the most frequent somatic genomic abnormality in T-ALL with germline RUNX1 variants. Co-introduction of RUNX1 variant and JAK3 mutation in hematopoietic stem and progenitor cells in mice gave rise to T-ALL with early T-cell precursor phenotype. Taken together, these results indicated that RUNX1 is an important predisposition gene for T-ALL and pointed to novel biology of RUNX1-mediated leukemogenesis in the lymphoid lineages.
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Affiliation(s)
- Yizhen Li
- Department of Pharmaceutical Sciences and
| | | | - Meenakshi Devidas
- Department of Global Pediatric Medicine, St. Jude Children’s Research Hospital, Memphis, Tennessee, USA
| | - Stuart S. Winter
- Children’s Minnesota Research Institute, Children’s Minnesota, Minneapolis, Minnesota, USA
| | - Chimene Kesserwan
- Center for Cancer Research, Genetics Branch, National Cancer Institute, Bethesda, Maryland, USA
| | | | - Kimberly P. Dunsmore
- Children’s Hematology and Oncology, Carilion Clinic and Virginia Tech Carilion School of Medicine, Roanoke, Virginia, USA
| | | | - Maoxiang Qian
- Institute of Pediatrics and Department of Hematology and Oncology, Children’s Hospital of Fudan University, Institutes of Biomedical Sciences, Shanghai, China
| | - Xujie Zhao
- Department of Pharmaceutical Sciences and
| | | | | | - Elizabeth A. Raetz
- Division of Pediatric Hematology and Oncology, Perlmutter Cancer Center, New York University Langone Health, New York, New York, USA
| | - William L. Carroll
- Division of Pediatric Hematology and Oncology, Perlmutter Cancer Center, New York University Langone Health, New York, New York, USA
| | - Chunliang Li
- Tumor Cell Biology, St. Jude Children’s Research Hospital, Memphis, Tennessee, USA
| | - Paul P. Liu
- Oncogenesis and Development Section, National Human Genome Research Institute, NIH, Bethesda, Maryland, USA
| | - Karen R. Rabin
- Texas Children’s Cancer and Hematology Centers, Baylor College of Medicine, Houston, Texas, USA
| | - Takaomi Sanda
- Cancer Science Institute of Singapore, and
- Department of Medicine, Yong Loo Lin School of Medicine, National University of Singapore, Singapore
| | | | | | - William E. Evans
- Department of Pharmaceutical Sciences and
- Hematological Malignancies Program, St. Jude Children’s Research Hospital, Memphis, Tennessee, USA
| | - Ching-Hon Pui
- Department of Oncology, and
- Hematological Malignancies Program, St. Jude Children’s Research Hospital, Memphis, Tennessee, USA
| | - Stephen P. Hunger
- Department of Pediatrics and Center for Childhood Cancer Research, Children’s Hospital of Philadelphia and Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | - David T. Teachey
- Department of Pediatrics and Center for Childhood Cancer Research, Children’s Hospital of Philadelphia and Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | - Mary V. Relling
- Department of Pharmaceutical Sciences and
- Hematological Malignancies Program, St. Jude Children’s Research Hospital, Memphis, Tennessee, USA
| | - Mignon L. Loh
- Department of Pediatrics, Benioff Children’s Hospital and the Helen Diller Family Comprehensive Cancer Center, UCSF, San Francisco, California, USA
| | - Jun J. Yang
- Department of Pharmaceutical Sciences and
- Department of Oncology, and
- Hematological Malignancies Program, St. Jude Children’s Research Hospital, Memphis, Tennessee, USA
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10
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Borst S, Nations CC, Klein JG, Pavani G, Maguire JA, Camire RM, Drazer MW, Godley LA, French DL, Poncz M, Gadue P. Study of inherited thrombocytopenia resulting from mutations in ETV6 or RUNX1 using a human pluripotent stem cell model. Stem Cell Reports 2021; 16:1458-1467. [PMID: 34019812 PMCID: PMC8190596 DOI: 10.1016/j.stemcr.2021.04.013] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2020] [Revised: 04/20/2021] [Accepted: 04/21/2021] [Indexed: 12/29/2022] Open
Abstract
Inherited thrombocytopenia results in low platelet counts and increased bleeding. Subsets of these patients have monoallelic germline mutations in ETV6 or RUNX1 and a heightened risk of developing hematologic malignancies. Utilizing CRISPR-Cas9, we compared the in vitro phenotype of hematopoietic progenitor cells and megakaryocytes derived from induced pluripotent stem cell (iPSC) lines harboring mutations in either ETV6 or RUNX1. Both mutant lines display phenotypes consistent with a platelet-bleeding disorder. Surprisingly, these cellular phenotypes were largely distinct. The ETV6-mutant iPSCs yield more hematopoietic progenitor cells and megakaryocytes, but the megakaryocytes are immature and less responsive to agonist stimulation. On the contrary, RUNX1-mutant iPSCs yield fewer hematopoietic progenitor cells and megakaryocytes, but the megakaryocytes are more responsive to agonist stimulation. However, both mutant iPSC lines display defects in proplatelet formation. Our work highlights that, while patients harboring germline ETV6 or RUNX1 mutations have similar clinical phenotypes, the molecular mechanisms may be distinct.
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Affiliation(s)
- Sara Borst
- Department of Cell and Molecular Biology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104, USA; Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, CTRB 5012, 3501 Civic Center Boulevard, Philadelphia, PA 19104, USA
| | - Catriana C Nations
- Department of Cell and Molecular Biology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104, USA; Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, CTRB 5012, 3501 Civic Center Boulevard, Philadelphia, PA 19104, USA
| | - Joshua G Klein
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, CTRB 5012, 3501 Civic Center Boulevard, Philadelphia, PA 19104, USA
| | - Giulia Pavani
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, CTRB 5012, 3501 Civic Center Boulevard, Philadelphia, PA 19104, USA
| | - Jean Ann Maguire
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, CTRB 5012, 3501 Civic Center Boulevard, Philadelphia, PA 19104, USA
| | - Rodney M Camire
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, CTRB 5012, 3501 Civic Center Boulevard, Philadelphia, PA 19104, USA; Department of Pediatrics, University of Pennsylvania Perelman School of Medicine and Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Michael W Drazer
- Section of Hematology/Oncology, Departments of Medicine and Human Genetics, The University of Chicago, Chicago, IL 60637, USA
| | - Lucy A Godley
- Section of Hematology/Oncology, Departments of Medicine and Human Genetics, The University of Chicago, Chicago, IL 60637, USA
| | - Deborah L French
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, CTRB 5012, 3501 Civic Center Boulevard, Philadelphia, PA 19104, USA; Department of Pathology and Laboratory Medicine, University of Pennsylvania Perelman School of Medicine and Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Mortimer Poncz
- Department of Pediatrics, University of Pennsylvania Perelman School of Medicine and Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Paul Gadue
- Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, CTRB 5012, 3501 Civic Center Boulevard, Philadelphia, PA 19104, USA; Department of Pathology and Laboratory Medicine, University of Pennsylvania Perelman School of Medicine and Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
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11
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Restoring RUNX1 deficiency in RUNX1 familial platelet disorder by inhibiting its degradation. Blood Adv 2021; 5:687-699. [PMID: 33560381 DOI: 10.1182/bloodadvances.2020002709] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2020] [Accepted: 12/16/2020] [Indexed: 12/15/2022] Open
Abstract
RUNX1 familial platelet disorder (RUNX1-FPD) is an autosomal dominant disorder caused by a monoallelic mutation of RUNX1, initially resulting in approximately half-normal RUNX1 activity. Clinical features include thrombocytopenia, platelet functional defects, and a predisposition to leukemia. RUNX1 is rapidly degraded through the ubiquitin-proteasome pathway. Moreover, it may autoregulate its expression. A predicted kinetic property of autoregulatory circuits is that transient perturbations of steady-state levels result in continued maintenance of expression at adjusted levels, even after inhibitors of degradation or inducers of transcription are withdrawn, suggesting that transient inhibition of RUNX1 degradation may have prolonged effects. We hypothesized that pharmacological inhibition of RUNX1 protein degradation could normalize RUNX1 protein levels, restore the number of platelets and their function, and potentially delay or prevent malignant transformation. In this study, we evaluated cell lines, induced pluripotent stem cells derived from patients with RUNX1-FPD, RUNX1-FPD primary bone marrow cells, and acute myeloid leukemia blood cells from patients with RUNX1 mutations. The results showed that, in some circumstances, transient expression of exogenous RUNX1 or inhibition of steps leading to RUNX1 ubiquitylation and proteasomal degradation restored RUNX1 levels, thereby advancing megakaryocytic differentiation in vitro. Thus, drugs retarding RUNX1 proteolytic degradation may represent a therapeutic avenue for treating bleeding complications and preventing leukemia in RUNX1-FPD.
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12
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The Emerging Role of Hematopathologists and Molecular Pathologists in Detection, Monitoring, and Management of Myeloid Neoplasms with Germline Predisposition. Curr Hematol Malig Rep 2021; 16:336-344. [PMID: 34028637 DOI: 10.1007/s11899-021-00636-2] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 03/19/2021] [Indexed: 12/12/2022]
Abstract
PURPOSE OF REVIEW Awareness, widespread availability, and routine use of sequencing techniques in work-up of myelodysplastic syndromes and acute myeloid leukemia have facilitated increased recognition of these entities arising in a background of germline predisposition disorders (GPD). RECENT FINDINGS The latest revisions to the WHO classification of myeloid neoplasms incorporate "myeloid neoplasms with germline predisposition" as a separate entity due to the therapeutic implications of this diagnosis. It has become apparent that some of these entities have unique recognizable morphologic findings that can be challenging to interpret at time. Hence, much needs to be studied, posing a new layer of complexity to hematopathologists and oncologists. A thorough understanding of cytogenetic and molecular findings during disease evolution is essential. Consequently, hematopathologists and molecular pathologists play an increasing role in recognition of bone marrow morphologic features that help in recognition of underlying GPD, monitoring, and prompt identification of progression.
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13
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RUNX-1 haploinsufficiency causes a marked deficiency of megakaryocyte-biased hematopoietic progenitor cells. Blood 2021; 137:2662-2675. [PMID: 33569577 DOI: 10.1182/blood.2020006389] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2020] [Accepted: 01/17/2021] [Indexed: 12/18/2022] Open
Abstract
Patients with familial platelet disorder with a predisposition to myeloid malignancy (FPDMM) harbor germline monoallelic mutations in a key hematopoietic transcription factor, RUNX-1. Previous studies of FPDMM have focused on megakaryocyte (Mk) differentiation and platelet production and signaling. However, the effects of RUNX-1 haploinsufficiency on hematopoietic progenitor cells (HPCs) and subsequent megakaryopoiesis remains incomplete. We studied induced pluripotent stem cell (iPSC)-derived HPCs (iHPCs) and Mks (iMks) from both patient-derived lines and a wild-type (WT) line modified to be RUNX-1 haploinsufficient (RUNX-1+/-), each compared with their isogenic WT control. All RUNX-1+/- lines showed decreased iMk yield and depletion of an Mk-biased iHPC subpopulation. To investigate global and local gene expression changes underlying this iHPC shift, single-cell RNA sequencing was performed on sorted FPDMM and control iHPCs. We defined several cell subpopulations in the Mk-biased iHPCs. Analyses of gene sets upregulated in FPDMM iHPCs indicated enrichment for response to stress, regulation of signal transduction, and immune signaling-related gene sets. Immunoblot analyses in FPDMM iMks were consistent with these findings, but also identified augmented baseline c-Jun N-terminal kinase (JNK) phosphorylation, known to be activated by transforming growth factor-β1 (TGF-β1) and cellular stressors. These findings were confirmed in adult human CD34+-derived stem and progenitor cells (HSPCs) transduced with lentiviral RUNX1 short hairpin RNA to mimic RUNX-1+/-. In both iHPCs and CD34+-derived HSPCs, targeted inhibitors of JNK and TGF-β1 pathways corrected the megakaryopoietic defect. We propose that such intervention may correct the thrombocytopenia in patients with FPDMM.
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14
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Bąk A, Skonieczka K, Jaśkowiec A, Junkiert-Czarnecka A, Heise M, Pilarska-Deltow M, Potoczek S, Czyżewska M, Haus O. Searching for germline mutations in the RUNX1 gene among Polish patients with acute myeloid leukemia. Leuk Lymphoma 2021; 62:1749-1755. [PMID: 33563056 DOI: 10.1080/10428194.2021.1881503] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/22/2022]
Abstract
The aim of the study was the identification of constitutional RUNX1 mutations among AML patients. The study group included 100 patients of Polish origin, diagnosed with de novo AML. 14 out of 100 AML patients had together 17 RUNX1 mutations, three of which were found to be germline changes. The difference in germline mutation frequency between study and control groups was not statistically significant (p = 0.193), but the odds ratio was 7.215. In all patients with germline mutations, chromosome 7 aberrations were found. The difference in the frequency of chromosome 7 aberrations between the group of patients with and without germline mutations was statistically significant (p = 0.008, OR = 73.00). We showed a higher frequency of germline mutations of RUNX1 in AML patients than in the control group, which confirms the role of these mutations in the development of AML, and an association of germline mutations with aberrations of chromosome 7.
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Affiliation(s)
- Aneta Bąk
- Department of Clinical Genetics, Collegium Medicum, Bydgoszcz, Nicolaus Copernicus University, Toruń, Poland
| | - Katarzyna Skonieczka
- Department of Clinical Genetics, Collegium Medicum, Bydgoszcz, Nicolaus Copernicus University, Toruń, Poland
| | - Anna Jaśkowiec
- Department of Hematology, Blood Neoplasms and Bone Marrow Transplantation, Medical University, Wrocław, Poland
| | - Anna Junkiert-Czarnecka
- Department of Clinical Genetics, Collegium Medicum, Bydgoszcz, Nicolaus Copernicus University, Toruń, Poland
| | - Marta Heise
- Department of Clinical Genetics, Collegium Medicum, Bydgoszcz, Nicolaus Copernicus University, Toruń, Poland
| | - Maria Pilarska-Deltow
- Department of Clinical Genetics, Collegium Medicum, Bydgoszcz, Nicolaus Copernicus University, Toruń, Poland
| | - Stanisław Potoczek
- Department of Hematology, Blood Neoplasms and Bone Marrow Transplantation, Medical University, Wrocław, Poland
| | | | - Olga Haus
- Department of Clinical Genetics, Collegium Medicum, Bydgoszcz, Nicolaus Copernicus University, Toruń, Poland
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15
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Reilly A, Doulatov S. Induced pluripotent stem cell models of myeloid malignancies and clonal evolution. Stem Cell Res 2021; 52:102195. [PMID: 33592565 PMCID: PMC10115516 DOI: 10.1016/j.scr.2021.102195] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/18/2020] [Revised: 01/15/2021] [Accepted: 01/20/2021] [Indexed: 12/13/2022] Open
Abstract
Reprogramming of cells from patients with genetic disorders to pluripotency is a promising avenue to understanding disease biology. A number of induced pluripotent stem cell (iPSC) models of inherited monogenic blood disorders have been reported over the past decade. However, the application of iPSCs for modeling of hematological malignancies has only recently been explored. Blood malignancies comprise a spectrum of genetically heterogeneous disorders marked by the acquisition of somatic mutations and chromosomal aberrations. This genetic heterogeneity presents unique challenges for iPSC modeling, but also opportunities to capture genetically distinct states and generate models of stepwise progression from normal to malignant hematopoiesis. Here we briefly review the current state of this field, highlighting current models of acquired pre-malignant and malignant blood disorders and clonal evolution, and challenges including barriers to reprogramming and differentiation of iPSCs into bona fide hematopoietic stem cells.
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Affiliation(s)
- Andreea Reilly
- Division of Hematology, Department of Medicine, Department of Genome Sciences, Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98195, United States
| | - Sergei Doulatov
- Division of Hematology, Department of Medicine, Department of Genome Sciences, Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98195, United States.
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16
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Yang Y, Li T, Geng Y, Li J. [RUNX1 gene mutations are associated with adverse prognosis of patients with acute myeloidleukemia]. NAN FANG YI KE DA XUE XUE BAO = JOURNAL OF SOUTHERN MEDICAL UNIVERSITY 2020; 40:1601-1606. [PMID: 33243739 DOI: 10.12122/j.issn.1673-4254.2020.11.10] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Indexed: 11/24/2022]
Abstract
OBJECTIVE To explore the rate and distribution of Runt- related transcription factor 1 (RUNX1) gene mutations in patients with acute myeloid leukemia (AML) and the correlation of these mutations with the clinical characteristics and survival outcomes of the patients. METHODS The genomic DNA extracted from the bone marrow of 158 patients with newly diagnosed AML for PCR amplification of RUNX1 gene and sequence analysis to identify the mutations. The mutations of ASXL1, DNMT3A, TET2, FLT3, CEBPA, NPM1, IDH2, NRAS and c-KIT genes were also examined to analyze their association with RUNX1 gene mutations. RESULTS Among the 158 AML patients, 19 (12.0%) were found to have RUNX1 mutations in A166G (9 cases), A142T (6 cases) and A162L (4 cases). RUNX1 mutations were more frequent in elderly patients (P < 0.01) and in cases of AML subtypes M4 and M5, and were associated with more frequent CD36 and CD7 expression as compared with the wild type. RUNX1 mutations were more likely to occur in patients with normal karyotype or karyotypes associated with moderate prognostic risks, but the difference was not significant (P > 0.05). The patients with RUNX1 mutations had significantly lower complete remission (CR) rate and overall survival (OS) rate than those without the mutations (P < 0.05). RUNX1 mutations were not associated with gender, white blood cell count upon diagnosis, hemoglobin level, platelet count, bone marrow blast cell ratio or lactate dehydrogenase level (P > 0.05). CONCLUSIONS RUNX1 gene mutations are associated with an adverse prognosis of patients with AML.
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Affiliation(s)
- Yanli Yang
- Department of Hematology, First Affiliated Hospital of Bengbu Medical College, Bengbu 233000, China
| | - Tiantian Li
- Department of Hematology, First Affiliated Hospital of Bengbu Medical College, Bengbu 233000, China
| | - Yinghua Geng
- Department of Hematology, First Affiliated Hospital of Bengbu Medical College, Bengbu 233000, China
| | - Jun Li
- Department of Hematology, First Affiliated Hospital of Bengbu Medical College, Bengbu 233000, China
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17
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Donada A, Basso-Valentina F, Arkoun B, Monte-Mor B, Plo I, Raslova H. Induced pluripotent stem cells and hematological malignancies: A powerful tool for disease modeling and drug development. Stem Cell Res 2020; 49:102060. [PMID: 33142254 DOI: 10.1016/j.scr.2020.102060] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/02/2020] [Revised: 10/09/2020] [Accepted: 10/16/2020] [Indexed: 01/12/2023] Open
Abstract
The derivation of human pluripotent stem cell (iPSC) lines by in vitro reprogramming of somatic cells revolutionized research: iPSCs have been used for disease modeling, drug screening and regenerative medicine for many disorders, especially when combined with cutting-edge genome editing technologies. In hematology, malignant transformation is often a multi-step process, that starts with either germline or acquired genetic alteration, followed by progressive acquisition of mutations combined with the selection of one or more pre-existing clones. iPSCs are an excellent model to study the cooperation between different genetic alterations and to test relevant therapeutic drugs. In this review, we will describe the use of iPSCs for pathophysiological studies and drug testing in inherited and acquired hematological malignancies.
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Affiliation(s)
- A Donada
- INSERM, UMR1287, Université Paris Sud, Université Paris Saclay, Gustave Roussy, Equipe Labellisée LNCC, Villejuif, France
| | - F Basso-Valentina
- INSERM, UMR1287, Université Paris Sud, Université Paris Saclay, Gustave Roussy, Equipe Labellisée LNCC, Villejuif, France
| | - B Arkoun
- INSERM, UMR1287, Université Paris Sud, Université Paris Saclay, Gustave Roussy, Equipe Labellisée LNCC, Villejuif, France
| | - B Monte-Mor
- Brazilian National Cancer Institute, Rio de Janeiro, Brazil
| | - I Plo
- INSERM, UMR1287, Université Paris Sud, Université Paris Saclay, Gustave Roussy, Equipe Labellisée LNCC, Villejuif, France
| | - H Raslova
- INSERM, UMR1287, Université Paris Sud, Université Paris Saclay, Gustave Roussy, Equipe Labellisée LNCC, Villejuif, France.
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Abstract
PURPOSE OF REVIEW We review the ways in which stem cells are used in psychiatric disease research, including the related advances in gene editing and directed cell differentiation. RECENT FINDINGS The recent development of induced pluripotent stem cell (iPSC) technologies has created new possibilities for the study of psychiatric disease. iPSCs can be derived from patients or controls and differentiated to an array of neuronal and non-neuronal cell types. Their genomes can be edited as desired, and they can be assessed for a variety of phenotypes. This makes them especially interesting for studying genetic variation, which is particularly useful today now that our knowledge on the genetics of psychiatric disease is quickly expanding. The recent advances in cell engineering have led to powerful new methods for studying psychiatric illness including schizophrenia, bipolar disorder, and autism. There is a wide array of possible applications as illustrated by the many examples from the literature, most of which are cited here.
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Affiliation(s)
- Debamitra Das
- Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Kyra Feuer
- Predoctoral Training Program in Human Genetics, Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA
- Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Marah Wahbeh
- Predoctoral Training Program in Human Genetics, Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA
- Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Dimitrios Avramopoulos
- Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
- Department of Psychiatry, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
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19
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Olofsen PA, Touw IP. RUNX1 Mutations in the Leukemic Progression of Severe Congenital Neutropenia. Mol Cells 2020; 43:139-144. [PMID: 32041395 PMCID: PMC7057833 DOI: 10.14348/molcells.2020.0010] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2020] [Accepted: 01/08/2020] [Indexed: 12/19/2022] Open
Abstract
Somatic RUNX1 mutations are found in approximately 10% of patients with de novo acute myeloid leukemia (AML), but are more common in secondary forms of myelodysplastic syndrome (MDS) or AML. Particularly, this applies to MDS/AML developing from certain types of leukemia-prone inherited bone marrow failure syndromes. How these RUNX1 mutations contribute to the pathobiology of secondary MDS/AML is still unknown. This mini-review focusses on the role of RUNX1 mutations as the most common secondary leukemogenic hit in MDS/AML evolving from severe congenital neutropenia (SCN).
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Affiliation(s)
| | - Ivo P. Touw
- Department of Hematology, Erasmus MC, Rotterdam 3015 CN, The Netherlands
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20
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Choi IY, Lim H, Cho HJ, Oh Y, Chou BK, Bai H, Cheng L, Kim YJ, Hyun S, Kim H, Shin JH, Lee G. Transcriptional landscape of myogenesis from human pluripotent stem cells reveals a key role of TWIST1 in maintenance of skeletal muscle progenitors. eLife 2020; 9:e46981. [PMID: 32011235 PMCID: PMC6996923 DOI: 10.7554/elife.46981] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2019] [Accepted: 01/14/2020] [Indexed: 12/15/2022] Open
Abstract
Generation of skeletal muscle cells with human pluripotent stem cells (hPSCs) opens new avenues for deciphering essential, but poorly understood aspects of transcriptional regulation in human myogenic specification. In this study, we characterized the transcriptional landscape of distinct human myogenic stages, including OCT4::EGFP+ pluripotent stem cells, MSGN1::EGFP+ presomite cells, PAX7::EGFP+ skeletal muscle progenitor cells, MYOG::EGFP+ myoblasts, and multinucleated myotubes. We defined signature gene expression profiles from each isolated cell population with unbiased clustering analysis, which provided unique insights into the transcriptional dynamics of human myogenesis from undifferentiated hPSCs to fully differentiated myotubes. Using a knock-out strategy, we identified TWIST1 as a critical factor in maintenance of human PAX7::EGFP+ putative skeletal muscle progenitor cells. Our data revealed a new role of TWIST1 in human skeletal muscle progenitors, and we have established a foundation to identify transcriptional regulations of human myogenic ontogeny (online database can be accessed in http://www.myogenesis.net/).
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Affiliation(s)
- In Young Choi
- The Institute for Cell EngineeringJohns Hopkins University, School of MedicineBaltimoreUnited States
- Department of Medicine, Graduate SchoolKyung Hee UniversitySeoulRepublic of Korea
| | - Hotae Lim
- The Institute for Cell EngineeringJohns Hopkins University, School of MedicineBaltimoreUnited States
- College of Veterinary MedicineChungbuk National UniversityChungbukRepublic of Korea
| | - Hyeon Jin Cho
- Lieber Institute for Brain Development, Johns Hopkins Medical CampusBaltimoreUnited States
| | - Yohan Oh
- The Institute for Cell EngineeringJohns Hopkins University, School of MedicineBaltimoreUnited States
| | - Bin-Kuan Chou
- The Institute for Cell EngineeringJohns Hopkins University, School of MedicineBaltimoreUnited States
- Division of Hematology, Department of MedicineJohns Hopkins University, School of MedicineBaltimoreUnited States
| | - Hao Bai
- The Institute for Cell EngineeringJohns Hopkins University, School of MedicineBaltimoreUnited States
- Division of Hematology, Department of MedicineJohns Hopkins University, School of MedicineBaltimoreUnited States
| | - Linzhao Cheng
- Division of Hematology, Department of MedicineJohns Hopkins University, School of MedicineBaltimoreUnited States
| | - Yong Jun Kim
- Department of Pathololgy, College of MedicineKyung Hee UniversitySeoulRepublic of Korea
| | - SangHwan Hyun
- The Institute for Cell EngineeringJohns Hopkins University, School of MedicineBaltimoreUnited States
- College of Veterinary MedicineChungbuk National UniversityChungbukRepublic of Korea
| | - Hyesoo Kim
- The Institute for Cell EngineeringJohns Hopkins University, School of MedicineBaltimoreUnited States
- Department of NeurologyJohns Hopkins University, School of MedicineBaltimoreUnited States
| | - Joo Heon Shin
- Lieber Institute for Brain Development, Johns Hopkins Medical CampusBaltimoreUnited States
| | - Gabsang Lee
- The Institute for Cell EngineeringJohns Hopkins University, School of MedicineBaltimoreUnited States
- Department of NeurologyJohns Hopkins University, School of MedicineBaltimoreUnited States
- The Solomon H. Synder Department of NeuroscienceJohns Hopkins University, School of MedicineBaltimoreUnited States
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21
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Thom CS, Chou ST, French DL. Mechanistic and Translational Advances Using iPSC-Derived Blood Cells. JOURNAL OF EXPERIMENTAL PATHOLOGY 2020; 1:36-44. [PMID: 33768218 PMCID: PMC7990314 DOI: 10.33696/pathology.1.010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 12/03/2022]
Abstract
Human induced pluripotent stem cell (iPSC)-based model systems can be used to produce blood cells for the study of both hematologic and non-hematologic disorders. This commentary discusses recent advances that have utilized iPSC-derived red blood cells, megakaryocytes, myeloid cells, and lymphoid cells to model hematopoietic disorders. In addition, we review recent studies that have defined how microglial cells differentiated from iPSC-derived monocytes impact neurodegenerative disease. Related translational insights highlight the utility of iPSC models for studying pathologic anemia, bleeding, thrombosis, autoimmunity, immunodeficiency, blood cancers, and neurodegenerative disease such as Alzheimer's.
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Affiliation(s)
- Christopher S Thom
- Division of Neonatology, Children’s Hospital of Philadelphia, Philadelphia, PA, USA
| | - Stella T Chou
- Division of Hematology, Children’s Hospital of Philadelphia, Philadelphia, PA, USA
| | - Deborah L French
- Center for Cellular and Molecular Therapeutics, Children’s Hospital of Philadelphia, Philadelphia, PA, USA
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22
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Modeling Leukemia with Human Induced Pluripotent Stem Cells. Cold Spring Harb Perspect Med 2019; 9:cshperspect.a034868. [PMID: 31451537 DOI: 10.1101/cshperspect.a034868] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
The reprogramming of human somatic cells into induced pluripotent stem cells (iPSCs) a little over a decade ago raised exciting prospects to transform the study and potentially also the therapy of human diseases. iPSC models have now been created for a multitude of hematologic diseases, including malignancies. Here we discuss practical aspects of iPSC modeling of malignant diseases, review recent studies, and discuss the new opportunities that iPSC models offer, as well as their current limitations and prospects for future development.
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23
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Galera P, Dulau-Florea A, Calvo KR. Inherited thrombocytopenia and platelet disorders with germline predisposition to myeloid neoplasia. Int J Lab Hematol 2019; 41 Suppl 1:131-141. [PMID: 31069978 DOI: 10.1111/ijlh.12999] [Citation(s) in RCA: 41] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/02/2019] [Revised: 02/07/2019] [Accepted: 02/10/2019] [Indexed: 12/21/2022]
Abstract
Advances in molecular genetic sequencing techniques have contributed to the elucidation of previously unknown germline mutations responsible for inherited thrombocytopenia (IT). Regardless of age of presentation and severity of symptoms related to thrombocytopenia and/or platelet dysfunction, a subset of patients with IT are at increased risk of developing myeloid neoplasms during their life time, particularly those with germline autosomal dominant mutations in RUNX1, ANKRD26, and ETV6. Patients may present with isolated thrombocytopenia and megakaryocytic dysmorphia or atypia on baseline bone marrow evaluation, without constituting myelodysplasia (MDS). Bone marrow features may overlap with idiopathic thrombocytopenic purpura (ITP) or sporadic MDS leading to misdiagnosis. Progression to myelodysplastic syndrome/ acute myeloid leukemia (MDS/AML) may be accompanied by progressive bi- or pancytopenia, multilineage dysplasia, increased blasts, cytogenetic abnormalities, acquisition of bi-allelic mutations in the underlying gene with germline mutation, or additional somatic mutations in genes associated with myeloid malignancy. A subset of patients may present with MDS/AML at a young age, underscoring the growing concern for evaluating young patients with MDS/AML for germline mutations predisposing to myeloid neoplasm. Early recognition of germline mutation and predisposition to myeloid malignancy permits appropriate treatment, adequate monitoring for disease progression, proper donor selection for hematopoietic stem cell transplantation, as well as genetic counseling of the affected patients and their family members. Herein, we describe the clinical and diagnostic features of IT with germline mutations predisposing to myeloid neoplasms focusing on mutations involving RUNX1, ANKRD26, and ETV6.
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Affiliation(s)
- Pallavi Galera
- Department of Laboratory Medicine, Hematology Section, Clinical Center, National Institutes of Health (NIH), Bethesda, Maryland
| | - Alina Dulau-Florea
- Department of Laboratory Medicine, Hematology Section, Clinical Center, National Institutes of Health (NIH), Bethesda, Maryland
| | - Katherine R Calvo
- Department of Laboratory Medicine, Hematology Section, Clinical Center, National Institutes of Health (NIH), Bethesda, Maryland
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24
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Georgomanoli M, Papapetrou EP. Modeling blood diseases with human induced pluripotent stem cells. Dis Model Mech 2019; 12:12/6/dmm039321. [PMID: 31171568 PMCID: PMC6602313 DOI: 10.1242/dmm.039321] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022] Open
Abstract
Induced pluripotent stem cells (iPSCs) are derived from somatic cells through a reprogramming process, which converts them to a pluripotent state, akin to that of embryonic stem cells. Over the past decade, iPSC models have found increasing applications in the study of human diseases, with blood disorders featuring prominently. Here, we discuss methodological aspects pertaining to iPSC generation, hematopoietic differentiation and gene editing, and provide an overview of uses of iPSCs in modeling the cell and gene therapy of inherited genetic blood disorders, as well as their more recent use as models of myeloid malignancies. We also discuss the strengths and limitations of iPSCs compared to model organisms and other cellular systems commonly used in hematology research.
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Affiliation(s)
- Maria Georgomanoli
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA
| | - Eirini P Papapetrou
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA
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25
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26
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RUNX1-targeted therapy for AML expressing somatic or germline mutation in RUNX1. Blood 2019; 134:59-73. [PMID: 31023702 DOI: 10.1182/blood.2018893982] [Citation(s) in RCA: 75] [Impact Index Per Article: 12.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/25/2018] [Accepted: 04/16/2019] [Indexed: 12/22/2022] Open
Abstract
RUNX1 transcription factor regulates normal and malignant hematopoiesis. Somatic or germline mutant RUNX1 (mtRUNX1) is associated with poorer outcome in acute myeloid leukemia (AML). Knockdown or inhibition of RUNX1 induced more apoptosis of AML expressing mtRUNX1 versus wild-type RUNX1 and improved survival of mice engrafted with mtRUNX1-expressing AML. CRISPR/Cas9-mediated editing-out of RUNX1 enhancer (eR1) within its intragenic super-enhancer, or BET protein BRD4 depletion by short hairpin RNA, repressed RUNX1, inhibited cell growth, and induced cell lethality in AML cells expressing mtRUNX1. Moreover, treatment with BET protein inhibitor or degrader (BET-proteolysis targeting chimera) repressed RUNX1 and its targets, inducing apoptosis and improving survival of mice engrafted with AML expressing mtRUNX1. Library of Integrated Network-based Cellular Signatures 1000-connectivity mapping data sets queried with messenger RNA signature of RUNX1 knockdown identified novel expression-mimickers (EMs), which repressed RUNX1 and exerted in vitro and in vivo efficacy against AML cells expressing mtRUNX1. In addition, the EMs cinobufagin, anisomycin, and narciclasine induced more lethality in hematopoietic progenitor cells (HPCs) expressing germline mtRUNX1 from patients with AML compared with HPCs from patients with familial platelet disorder (FPD), or normal untransformed HPCs. These findings highlight novel therapeutic agents for AML expressing somatic or germline mtRUNX1.
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27
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Saha SK, Saikot FK, Rahman MS, Jamal MAHM, Rahman SMK, Islam SMR, Kim KH. Programmable Molecular Scissors: Applications of a New Tool for Genome Editing in Biotech. MOLECULAR THERAPY. NUCLEIC ACIDS 2019; 14:212-238. [PMID: 30641475 PMCID: PMC6330515 DOI: 10.1016/j.omtn.2018.11.016] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 05/28/2018] [Revised: 11/23/2018] [Accepted: 11/23/2018] [Indexed: 01/04/2023]
Abstract
Targeted genome editing is an advanced technique that enables precise modification of the nucleic acid sequences in a genome. Genome editing is typically performed using tools, such as molecular scissors, to cut a defined location in a specific gene. Genome editing has impacted various fields of biotechnology, such as agriculture; biopharmaceutical production; studies on the structure, regulation, and function of the genome; and the creation of transgenic organisms and cell lines. Although genome editing is used frequently, it has several limitations. Here, we provide an overview of well-studied genome-editing nucleases, including single-stranded oligodeoxynucleotides (ssODNs), transcription activator-like effector nucleases (TALENs), zinc-finger nucleases (ZFNs), and CRISPR-Cas9 RNA-guided nucleases (CRISPR-Cas9). To this end, we describe the progress toward editable nuclease-based therapies and discuss the minimization of off-target mutagenesis. Future prospects of this challenging scientific field are also discussed.
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Affiliation(s)
- Subbroto Kumar Saha
- Department of Stem Cell and Regenerative Biotechnology, Konkuk University, 120 Neungdong-Ro, Seoul 05029, Republic of Korea.
| | - Forhad Karim Saikot
- Department of Genetic Engineering and Biotechnology, Jashore University of Science and Technology, Jashore 7408, Bangladesh
| | - Md Shahedur Rahman
- Department of Genetic Engineering and Biotechnology, Jashore University of Science and Technology, Jashore 7408, Bangladesh
| | | | - S M Khaledur Rahman
- Department of Genetic Engineering and Biotechnology, Jashore University of Science and Technology, Jashore 7408, Bangladesh
| | - S M Riazul Islam
- Department of Computer Science and Engineering, Sejong University, 209 Neungdong-ro, Gwangjin-gu, Seoul 05006, South Korea
| | - Ki-Hyun Kim
- Department of Civil & Environmental Engineering, Hanyang University, 222 Wangsimni-Ro, Seoul 04763, Republic of Korea.
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28
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Churpek JE, Bresnick EH. Transcription factor mutations as a cause of familial myeloid neoplasms. J Clin Invest 2019; 129:476-488. [PMID: 30707109 DOI: 10.1172/jci120854] [Citation(s) in RCA: 45] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
The initiation and evolution of myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) are driven by genomic events that disrupt multiple genes controlling hematopoiesis. Human genetic studies have discovered germline mutations in single genes that instigate familial MDS/AML. The best understood of these genes encode transcription factors, such as GATA-2, RUNX1, ETV6, and C/EBPα, which establish and maintain genetic networks governing the genesis and function of blood stem and progenitor cells. Many questions remain unanswered regarding how genes and circuits within these networks function in physiology and disease and whether network integrity is exquisitely sensitive to or efficiently buffered from perturbations. In familial MDS/AML, mutations change the coding sequence of a gene to generate a mutant protein with altered activity or introduce frameshifts or stop codons or disrupt regulatory elements to alter protein expression. Each mutation has the potential to exert quantitatively and qualitatively distinct influences on networks. Consistent with this mechanistic diversity, disease onset is unpredictable and phenotypic variability can be considerable. Efforts to elucidate mechanisms and forge prognostic and therapeutic strategies must therefore contend with a spectrum of patient-specific leukemogenic scenarios. Here we illustrate mechanistic advances in our understanding of familial MDS/AML syndromes caused by germline mutations of hematopoietic transcription factors.
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Affiliation(s)
- Jane E Churpek
- Section of Hematology/Oncology and Center for Clinical Cancer Genetics, The University of Chicago, Chicago, Illinois, USA
| | - Emery H Bresnick
- UW-Madison Blood Research Program, Department of Cell and Regenerative Biology, Wisconsin Institutes for Medical Research, UW Carbone Cancer Center, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, USA
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29
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Elbadry MI, Espinoza JL, Nakao S. Disease modeling of bone marrow failure syndromes using iPSC-derived hematopoietic stem progenitor cells. Exp Hematol 2019; 71:32-42. [PMID: 30664904 DOI: 10.1016/j.exphem.2019.01.006] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2018] [Revised: 01/04/2019] [Accepted: 01/15/2019] [Indexed: 01/19/2023]
Abstract
The plasticity of induced pluripotent stem cells (iPSCs) with the potential to differentiate into virtually any type of cells and the feasibility of generating hematopoietic stem progenitor cells (HSPCs) from patient-derived iPSCs (iPSC-HSPCs) has many potential applications in hematology. For example, iPSC-HSPCs are being used for leukemogenesis studies and their application in various cell replacement therapies is being evaluated. The use of iPSC-HSPCs can now provide an invaluable resource for the study of diseases associated with the destruction of HSPCs, such as bone marrow failure syndromes (BMFSs). Recent studies have shown that generating iPSC-HSPCs from patients with acquired aplastic anemia and other BMFSs is not only feasible, but is also a powerful tool for understanding the pathogenesis of these disorders. In this article, we highlight recent advances in the application of iPSCs for disease modeling of BMFSs and discuss the discoveries of these studies that provide new insights in the pathophysiology of these conditions.
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Affiliation(s)
- Mahmoud I Elbadry
- Hematology/Respiratory Medicine, Faculty of Medicine, Institute of Medical Pharmaceutical and Health Sciences, Kanazawa University, Kanazawa, Japan; Department of Internal Medicine, Division of Hematology, Faculty of Medicine, Sohag University, Egypt
| | - J Luis Espinoza
- Department of Hematology and Rheumatology, Faculty of Medicine, Kindai University, Osaka, Japan
| | - Shinji Nakao
- Hematology/Respiratory Medicine, Faculty of Medicine, Institute of Medical Pharmaceutical and Health Sciences, Kanazawa University, Kanazawa, Japan.
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30
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31
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Kim H, Schaniel C. Modeling Hematological Diseases and Cancer With Patient-Specific Induced Pluripotent Stem Cells. Front Immunol 2018; 9:2243. [PMID: 30323816 PMCID: PMC6172418 DOI: 10.3389/fimmu.2018.02243] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2018] [Accepted: 09/10/2018] [Indexed: 12/13/2022] Open
Abstract
The advent of induced pluripotent stem cells (iPSCs) together with recent advances in genome editing, microphysiological systems, tissue engineering and xenograft models present new opportunities for the investigation of hematological diseases and cancer in a patient-specific context. Here we review the progress in the field and discuss the advantages, limitations, and challenges of iPSC-based malignancy modeling. We will also discuss the use of iPSCs and its derivatives as cellular sources for drug target identification, drug development and evaluation of pharmacological responses.
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Affiliation(s)
- Huensuk Kim
- Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Department of Cell, Developmental and Regenerative Biology, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Christoph Schaniel
- Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Department of Cell, Developmental and Regenerative Biology, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Department of Pharmacological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Mount Sinai Institute for Systems Biomedicine, Icahn School of Medicine at Mount Sinai, New York, NY, United States
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32
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Li Y, Jin C, Bai H, Gao Y, Sun S, Chen L, Qin L, Liu PP, Cheng L, Wang QF. Human NOTCH4 is a key target of RUNX1 in megakaryocytic differentiation. Blood 2018; 131:191-201. [PMID: 29101237 PMCID: PMC5757696 DOI: 10.1182/blood-2017-04-780379] [Citation(s) in RCA: 32] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/19/2017] [Accepted: 10/13/2017] [Indexed: 12/19/2022] Open
Abstract
Megakaryocytes (MKs) in adult marrow produce platelets that play important roles in blood coagulation and hemostasis. Monoallelic mutations of the master transcription factor gene RUNX1 lead to familial platelet disorder (FPD) characterized by defective MK and platelet development. However, the molecular mechanisms of FPD remain unclear. Previously, we generated human induced pluripotent stem cells (iPSCs) from patients with FPD containing a RUNX1 nonsense mutation. Production of MKs from the FPD-iPSCs was reduced, and targeted correction of the RUNX1 mutation restored MK production. In this study, we used isogenic pairs of FPD-iPSCs and the MK differentiation system to identify RUNX1 target genes. Using integrative genomic analysis of hematopoietic progenitor cells generated from FPD-iPSCs, and mutation-corrected isogenic controls, we identified 2 gene sets the transcription of which is either up- or downregulated by RUNX1 in mutation-corrected iPSCs. Notably, NOTCH4 expression was negatively controlled by RUNX1 via a novel regulatory DNA element within the locus, and we examined its involvement in MK generation. Specific inactivation of NOTCH4 by an improved CRISPR-Cas9 system in human iPSCs enhanced megakaryopoiesis. Moreover, small molecules known to inhibit Notch signaling promoted MK generation from both normal human iPSCs and postnatal CD34+ hematopoietic stem and progenitor cells. Our study newly identified NOTCH4 as a RUNX1 target gene and revealed a previously unappreciated role of NOTCH4 signaling in promoting human megakaryopoiesis. Our work suggests that human iPSCs with monogenic mutations have the potential to serve as an invaluable resource for discovery of novel druggable targets.
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Affiliation(s)
- Yueying Li
- Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China
| | - Chen Jin
- Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Hao Bai
- Division of Hematology, Department of Medicine and
- Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD; and
| | - Yongxing Gao
- Division of Hematology, Department of Medicine and
- Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD; and
| | - Shu Sun
- Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Lei Chen
- Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Lei Qin
- Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Paul P Liu
- Translational and Functional Genomics Branch, National Institutes of Health, National Human Genome Research Institute, Bethesda, MD
| | - Linzhao Cheng
- Division of Hematology, Department of Medicine and
- Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD; and
| | - Qian-Fei Wang
- Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China
- University of Chinese Academy of Sciences, Beijing, China
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33
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Borst S, Sim X, Poncz M, French DL, Gadue P. Induced Pluripotent Stem Cell-Derived Megakaryocytes and Platelets for Disease Modeling and Future Clinical Applications. Arterioscler Thromb Vasc Biol 2017; 37:2007-2013. [PMID: 28982668 PMCID: PMC5675007 DOI: 10.1161/atvbaha.117.309197] [Citation(s) in RCA: 26] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2017] [Accepted: 09/21/2017] [Indexed: 12/13/2022]
Abstract
Platelets, derived from megakaryocytes, are anucleate cytoplasmic discs that circulate in the blood stream and play major roles in hemostasis, inflammation, and vascular biology. Platelet transfusions are used in a variety of medical settings to prevent life-threatening thrombocytopenia because of cancer therapy, other causes of acquired or inherited thrombocytopenia, and trauma. Currently, platelets used for transfusion purposes are donor derived. However, there is a drive to generate nondonor sources of platelets to help supplement donor-derived platelets. Efforts have been made by many laboratories to generate in vitro platelets and optimize their production and quality. In vitro-derived platelets have the potential to be a safer, more uniform product, and genetic manipulation could allow for better treatment of patients who become refractory to donor-derived units. This review focuses on potential clinical applications of in vitro-derived megakaryocytes and platelets, current methods to generate and expand megakaryocytes from pluripotent stem cell sources, and the use of these cells for disease modeling.
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Affiliation(s)
- Sara Borst
- From the Department of Cell and Molecular Biology, Perelman School of Medicine (S.B., X.S.), Department of Pharmacology, Perelman School of Medicine (M.P.), and Department of Pathology and Laboratory Medicine (D.L.F., P.G.), University of Pennsylvania, Philadelphia; and Center for Cellular and Molecular Therapeutics (S.B., X.S., D.L.F., P.G.) and Division of Hematology (M.P.), Children's Hospital of Philadelphia, PA
| | - Xiuli Sim
- From the Department of Cell and Molecular Biology, Perelman School of Medicine (S.B., X.S.), Department of Pharmacology, Perelman School of Medicine (M.P.), and Department of Pathology and Laboratory Medicine (D.L.F., P.G.), University of Pennsylvania, Philadelphia; and Center for Cellular and Molecular Therapeutics (S.B., X.S., D.L.F., P.G.) and Division of Hematology (M.P.), Children's Hospital of Philadelphia, PA
| | - Mortimer Poncz
- From the Department of Cell and Molecular Biology, Perelman School of Medicine (S.B., X.S.), Department of Pharmacology, Perelman School of Medicine (M.P.), and Department of Pathology and Laboratory Medicine (D.L.F., P.G.), University of Pennsylvania, Philadelphia; and Center for Cellular and Molecular Therapeutics (S.B., X.S., D.L.F., P.G.) and Division of Hematology (M.P.), Children's Hospital of Philadelphia, PA
| | - Deborah L French
- From the Department of Cell and Molecular Biology, Perelman School of Medicine (S.B., X.S.), Department of Pharmacology, Perelman School of Medicine (M.P.), and Department of Pathology and Laboratory Medicine (D.L.F., P.G.), University of Pennsylvania, Philadelphia; and Center for Cellular and Molecular Therapeutics (S.B., X.S., D.L.F., P.G.) and Division of Hematology (M.P.), Children's Hospital of Philadelphia, PA
| | - Paul Gadue
- From the Department of Cell and Molecular Biology, Perelman School of Medicine (S.B., X.S.), Department of Pharmacology, Perelman School of Medicine (M.P.), and Department of Pathology and Laboratory Medicine (D.L.F., P.G.), University of Pennsylvania, Philadelphia; and Center for Cellular and Molecular Therapeutics (S.B., X.S., D.L.F., P.G.) and Division of Hematology (M.P.), Children's Hospital of Philadelphia, PA.
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Kanagal-Shamanna R, Loghavi S, DiNardo CD, Medeiros LJ, Garcia-Manero G, Jabbour E, Routbort MJ, Luthra R, Bueso-Ramos CE, Khoury JD. Bone marrow pathologic abnormalities in familial platelet disorder with propensity for myeloid malignancy and germline RUNX1 mutation. Haematologica 2017; 102:1661-1670. [PMID: 28659335 PMCID: PMC5622850 DOI: 10.3324/haematol.2017.167726] [Citation(s) in RCA: 60] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/17/2017] [Accepted: 06/20/2017] [Indexed: 01/20/2023] Open
Abstract
A subset of patients with familial platelet disorder with propensity to myeloid malignancy and germline RUNX1 mutation develops hematological malignancies, often myelodysplastic syndrome/acute myeloid leukemia, currently recognized in the 2016 WHO classification. Patients who develop hematologic malignancies are typically young, respond poorly to conventional therapy, and need allogeneic stem cell transplant from non-familial donors. Understanding the spectrum of bone marrow morphologic and genetic findings in these patients is critical to ensure diagnostic accuracy and develop criteria to recognize the onset of hematologic malignancies, particularly myelodysplastic syndrome. However, bone marrow features remain poorly characterized. To address this knowledge gap, we analyzed the clinicopathologic and genetic findings of 11 patients from 7 pedigrees. Of these, 6 patients did not develop hematologic malignancies over a 22-month follow-up period; 5 patients developed hematologic malignancies (3 acute myeloid leukemia; 2 myelodysplastic syndrome). All patients had thrombocytopenia at initial presentation. All 6 patients who did not develop hematologic malignancies showed baseline bone marrow abnormalities: low-for-age cellularity (n=4), dysmegakaryopoiesis (n=5), megakaryocytic hypoplasia/hyperplasia (n=5), and eosinophilia (n=4). Two patients had multiple immunophenotypic alterations in CD34-positive myeloblasts; 1 patient had clonal hematopoiesis. In contrast, patients who developed hematologic malignancies had additional cytopenia(s) (n=4), abnormal platelet granulation (n=5), bone marrow hypercellularity (n=4), dysplasia in ≥2 lineages including megakaryocytes (n=3) and acquired clonal genetic aberrations (n=5). In conclusion, our study demonstrated that specific bone marrow abnormalities and acquired genetic alterations may be harbingers of progression to hematological malignancies in patients with familial platelet disorder with germline RUNX1 mutation.
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Affiliation(s)
- Rashmi Kanagal-Shamanna
- Department of Hematopathology, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
| | - Sanam Loghavi
- Department of Hematopathology, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
| | - Courtney D DiNardo
- Department of Leukemia, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
| | - L Jeffrey Medeiros
- Department of Hematopathology, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
| | - Guillermo Garcia-Manero
- Department of Leukemia, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
| | - Elias Jabbour
- Department of Leukemia, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
| | - Mark J Routbort
- Department of Hematopathology, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
| | - Rajyalakshmi Luthra
- Department of Hematopathology, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
| | - Carlos E Bueso-Ramos
- Department of Hematopathology, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
| | - Joseph D Khoury
- Department of Hematopathology, The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA
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35
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Hayashi Y, Harada Y, Huang G, Harada H. Myeloid neoplasms with germ line RUNX1 mutation. Int J Hematol 2017; 106:183-188. [PMID: 28534116 DOI: 10.1007/s12185-017-2258-5] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2017] [Accepted: 05/16/2017] [Indexed: 01/23/2023]
Abstract
Familial platelet disorder with propensity to myeloid malignancies (FPD/AML) is an autosomal dominant disorder characterized by quantitative and/or qualitative platelet defects with a tendency to develop a variety of hematological malignancies. Heterozygous germ line mutations in the RUNX1 gene are responsible genetic events for FPD/AML. Notably, about half of individuals in the family with germ line mutations in RUNX1 develop overt hematological malignancies. The latency is also relatively long as an average age at diagnosis is more than 30 years. Similar to what is observed in sporadic hematological malignancies, acquired additional genetic events cooperate with inherited RUNX1 mutations to progress the overt malignant phase. Reflecting recent increased awareness of hematological malignancies with germ line mutations, FPD/AML was added in the revised WHO 2016 classification. In this review, we provide an update on FPD/AML with recent clinical and experimental findings.
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Affiliation(s)
- Yoshihiro Hayashi
- Laboratory of Oncology, School of Life Science, Tokyo University of Pharmacy and Life Sciences, 1432-1 Horinouchi, Hachioji, Tokyo, 192-0392, Japan.,Divisions of Pathology and Experimental Hematology and Cancer Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, 45229, USA
| | - Yuka Harada
- Department of Clinical Laboratory Medicine, Faculty of Health Science Technology, Bunkyo Gakuin University, Tokyo, 113-0023, Japan
| | - Gang Huang
- Divisions of Pathology and Experimental Hematology and Cancer Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, 45229, USA
| | - Hironori Harada
- Laboratory of Oncology, School of Life Science, Tokyo University of Pharmacy and Life Sciences, 1432-1 Horinouchi, Hachioji, Tokyo, 192-0392, Japan.
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36
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Ackermann M, Kuhn A, Kunkiel J, Merkert S, Martin U, Moritz T, Lachmann N. Ex vivo Generation of Genetically Modified Macrophages from Human Induced Pluripotent Stem Cells. Transfus Med Hemother 2017. [PMID: 28626364 DOI: 10.1159/000477129] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023] Open
Abstract
BACKGROUND Pluripotent stem cells, including induced pluripotent stem cells (iPSCs), have the capacity to differentiate towards all three germ layers and have been highlighted as an attractive cell source for the field of regenerative medicine. Thus, stable expression of therapeutic transgenes in iPSCs, as well as thereof derived progeny of hematopoietic lineage, may lay the foundation for innovative cell replacement therapies. METHODS We have utilized human iPSC lines genetically modified by lentiviral vector technology or targeted integration of reporter genes to evaluate transgene expression during hematopoietic specification and differentiation towards macrophages. RESULTS Use of lentiviral vectors equipped with an ubiquitous chromatin opening element (CBX3-UCOE) as well as zinc finger nuclease-mediated targeting of an expression cassette into the human adeno-associated virus integration site 1 (AAVS1) safe harbor resulted in stable transgene expression in iPSCs. When iPSCs were differentiated along the myeloid pathway into macrophages, both strategies yielded sustained transgene expression during the hematopoietic specification process including mature CD14+ and CD11b+ macrophages. CONCLUSION Combination of human iPSC technology with either lentiviral vector technology or designer nuclease-based genome editing allows for the generation of transgenic iPSC-derived macrophages with stable transgene expression which may be useful for novel cell and gene replacement therapies.
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Affiliation(s)
- Mania Ackermann
- JRG Translational Hematology, REBIRTH Cluster of Excellence, Hanover Medical School, Hanover, Germany.,Institute of Experimental Hematology, Hanover Medical School, Hanover, Germany
| | - Alexandra Kuhn
- Institute of Experimental Hematology, Hanover Medical School, Hanover, Germany.,RG Reprogramming and Gene Therapy, REBIRTH Cluster of Excellence, Hanover Medical School, Hanover, Germany
| | - Jessica Kunkiel
- Institute of Experimental Hematology, Hanover Medical School, Hanover, Germany.,RG Reprogramming and Gene Therapy, REBIRTH Cluster of Excellence, Hanover Medical School, Hanover, Germany
| | - Sylvia Merkert
- Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery (HTTG), REBIRTH-Cluster of Excellence, Hanover Medical School, Hanover, Germany.,Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), Hanover, Germany
| | - Ulrich Martin
- Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiothoracic, Transplantation and Vascular Surgery (HTTG), REBIRTH-Cluster of Excellence, Hanover Medical School, Hanover, Germany.,Biomedical Research in Endstage and Obstructive Lung Disease (BREATH), Member of the German Center for Lung Research (DZL), Hanover, Germany
| | - Thomas Moritz
- Institute of Experimental Hematology, Hanover Medical School, Hanover, Germany.,RG Reprogramming and Gene Therapy, REBIRTH Cluster of Excellence, Hanover Medical School, Hanover, Germany
| | - Nico Lachmann
- JRG Translational Hematology, REBIRTH Cluster of Excellence, Hanover Medical School, Hanover, Germany.,Institute of Experimental Hematology, Hanover Medical School, Hanover, Germany
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37
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Hematopoietic transcription factor mutations: important players in inherited platelet defects. Blood 2017; 129:2873-2881. [PMID: 28416505 DOI: 10.1182/blood-2016-11-709881] [Citation(s) in RCA: 47] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/17/2016] [Accepted: 01/26/2017] [Indexed: 01/19/2023] Open
Abstract
Transcription factors (TFs) are proteins that bind to specific DNA sequences and regulate expression of genes. The molecular and genetic mechanisms in most patients with inherited platelet defects are unknown. There is now increasing evidence that mutations in hematopoietic TFs are an important underlying cause for defects in platelet production, morphology, and function. The hematopoietic TFs implicated in patients with impaired platelet function and number include runt-related transcription factor 1, Fli-1 proto-oncogene, E-twenty-six (ETS) transcription factor (friend leukemia integration 1), GATA-binding protein 1, growth factor independent 1B transcriptional repressor, ETS variant 6, ecotropic viral integration site 1, and homeobox A11. These TFs act in a combinatorial manner to bind sequence-specific DNA within promoter regions to regulate lineage-specific gene expression, either as activators or repressors. TF mutations induce rippling downstream effects by simultaneously altering the expression of multiple genes. Mutations involving these TFs affect diverse aspects of megakaryocyte biology, and platelet production and function, culminating in thrombocytopenia and platelet dysfunction. Some are associated with predisposition to hematologic malignancies. These TF variants may occur more frequently in patients with inherited platelet defects than generally appreciated. This review focuses on alterations in hematopoietic TFs in the pathobiology of inherited platelet defects.
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38
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Schlegelberger B, Heller PG. RUNX1 deficiency (familial platelet disorder with predisposition to myeloid leukemia, FPDMM). Semin Hematol 2017. [PMID: 28637620 DOI: 10.1053/j.seminhematol.2017.04.006] [Citation(s) in RCA: 66] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
In this review, we discuss disease-causing alterations of RUNT-related transcription factor 1 (RUNX1), a master regulator of hematopoietic differentiation. Familial platelet disorder with predisposition to myeloid leukemia (FPDMM) typically presents with (1) mild to moderate thrombocytopenia with normal-sized platelets; (2) functional platelets defects leading to prolonged bleeding; and (3) an increased risk to develop myelodysplastic syndromes (MDS), acute myeloid leukemia (AML), or T-cell acute lymphoblastic leukemia (T-ALL). Hematological neoplasms in carriers of a germline RUNX1 mutation need additional secondary mutations or chromosome aberrations to develop. If a disease-causing mutation is known in the family, it is important to prevent hematopoietic stem cell transplantation from a sibling or other relative carrying the familial mutation. First experiments introducing a wild-type copy of RUNX1 into induce pluripotent stem cells (iPSC) lines from patients with FPDMM appear to demonstrate that by gene correction reversal of the phenotype may be possible.
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Affiliation(s)
| | - Paula G Heller
- Instituto de Investigaciones Médicas Alfredo Lanari, Universidad de Buenos Aires, IDIM-CONICET, Buenos Aires, Argentina
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39
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Mao GF, Goldfinger LE, Fan DC, Lambert MP, Jalagadugula G, Freishtat R, Rao AK. Dysregulation of PLDN (pallidin) is a mechanism for platelet dense granule deficiency in RUNX1 haplodeficiency. J Thromb Haemost 2017; 15:792-801. [PMID: 28075530 PMCID: PMC5378588 DOI: 10.1111/jth.13619] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2016] [Indexed: 01/01/2023]
Abstract
Essentials Platelet dense granule (DG) deficiency is a major abnormality in RUNX1 haplodeficiency patients. The molecular mechanisms leading to the platelet DG deficiency are unknown. Platelet expression of PLDN (BLOC1S6, pallidin), involved in DG biogenesis, is regulated by RUNX1. Downregulation of PLDN is a mechanism for DG deficiency in RUNX1 haplodeficiency. SUMMARY Background Inherited RUNX1 haplodeficiency is associated with thrombocytopenia and platelet dysfunction. Dense granule (DG) deficiency has been reported in patients with RUNX1 haplodeficiency, but the molecular mechanisms are unknown. Platelet mRNA expression profiling in a patient previously reported by us with a RUNX1 mutation and platelet dysfunction showed decreased expression of PLDN (BLOC1S6), which encodes pallidin, a subunit of biogenesis of lysosome-related organelles complex-1 (BLOC-1) involved in DG biogenesis. PLDN mutations in the pallid mouse and Hermansky-Pudlak syndrome-9 are associated with platelet DG deficiency. Objectives We postulated that PLDN is a RUNX1 target, and that its decreased expression leads to platelet DG deficiency in RUNX1 haplodeficiency. Results Platelet pallidin and DG levels were decreased in our patient. This was also observed in two siblings from a different family with a RUNX1 mutation. Chromatin immunoprecipitation and electrophoretic mobility shift assays with phorbol ester-treated human erythroleukemia (HEL) cells showed RUNX1 binding to RUNX1 consensus sites in the PLDN1 5' upstream region. In luciferase reporter studies, mutation of RUNX1 sites in the PLDN promoter reduced activity. RUNX1 overexpression enhanced and RUNX1 downregulation decreased PLDN1 promoter activity and protein expression. RUNX1 downregulation resulted in impaired handling of mepacrine and mislocalization of the DG marker CD63 in HEL cells, indicating impaired DG formation, recapitulating findings on PLDN downregulation. Conclusions These studies provide the first evidence that PLDN is a direct target of RUNX1 and that its dysregulation is a mechanism for platelet DG deficiency associated with RUNX1 haplodeficiency.
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Affiliation(s)
- G F Mao
- Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA, USA
| | - L E Goldfinger
- Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA, USA
- Department of Anatomy and Cell Biology, Temple University School of Medicine, Philadelphia, PA, USA
| | - D C Fan
- Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA, USA
| | - M P Lambert
- Division of Hematology, The Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
- Children's Hospital of Philadelphia and Department of Pediatrics, The Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA
| | - G Jalagadugula
- Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA, USA
| | - R Freishtat
- Department of Pediatrics, Children's National Medical Center, Washington, DC, USA
| | - A K Rao
- Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, PA, USA
- Department of Medicine, Temple University School of Medicine, Philadelphia, PA, USA
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40
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Abstract
RUNX1 is a member of the core-binding factor family of transcription factors and is indispensable for the establishment of definitive hematopoiesis in vertebrates. RUNX1 is one of the most frequently mutated genes in a variety of hematological malignancies. Germ line mutations in RUNX1 cause familial platelet disorder with associated myeloid malignancies. Somatic mutations and chromosomal rearrangements involving RUNX1 are frequently observed in myelodysplastic syndrome and leukemias of myeloid and lymphoid lineages, that is, acute myeloid leukemia, acute lymphoblastic leukemia, and chronic myelomonocytic leukemia. More recent studies suggest that the wild-type RUNX1 is required for growth and survival of certain types of leukemia cells. The purpose of this review is to discuss the current status of our understanding about the role of RUNX1 in hematological malignancies.
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41
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iPSCs and fibroblast subclones from the same fibroblast population contain comparable levels of sequence variations. Proc Natl Acad Sci U S A 2017; 114:1964-1969. [PMID: 28167771 DOI: 10.1073/pnas.1616035114] [Citation(s) in RCA: 48] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
Genome integrity of induced pluripotent stem cells (iPSCs) has been extensively studied in recent years, but it is still unclear whether iPSCs contain more genomic variations than cultured somatic cells. One important question is the origin of genomic variations detected in iPSCs-whether iPSC reprogramming induces such variations. Here, we undertook a unique approach by deriving fibroblast subclones and clonal iPSC lines from the same fibroblast population and applied next-generation sequencing to compare genomic variations in these lines. Targeted deep sequencing of parental fibroblasts revealed that most variants detected in clonal iPSCs and fibroblast subclones were rare variants inherited from the parental fibroblasts. Only a small number of variants remained undetectable in the parental fibroblasts, which were thus likely to be de novo. Importantly, the clonal iPSCs and fibroblast subclones contained comparable numbers of de novo variants. Collectively, our data suggest that iPSC reprogramming is not mutagenic.
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42
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RUNX1 and CBFβ Mutations and Activities of Their Wild-Type Alleles in AML. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2017; 962:265-282. [DOI: 10.1007/978-981-10-3233-2_17] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
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43
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Songdej N, Rao AK. Inherited platelet dysfunction and hematopoietic transcription factor mutations. Platelets 2017; 28:20-26. [PMID: 27463948 PMCID: PMC5628047 DOI: 10.1080/09537104.2016.1203400] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2016] [Revised: 05/16/2016] [Accepted: 05/30/2016] [Indexed: 01/19/2023]
Abstract
Transcription factors (TFs) are proteins that bind to specific DNA sequences and regulate expression of genes. The molecular and genetic mechanisms in most patients with inherited platelet dysfunction are unknown. There is now increasing evidence that mutations in hematopoietic TFs are an important underlying cause for the defects in platelet production, morphology, and function. The hematopoietic TFs implicated in the patients with impaired platelet function include Runt related TF 1 (RUNX1), Fli-1 proto-oncogene, ETS TF (FLI1), GATA-binding protein 1 (GATA1), and growth factor independent 1B transcriptional repressor (GFI1B). These TFs act in a combinatorial manner to bind sequence-specific DNA within a promoter region to regulate lineage-specific gene expression, either as activators or as repressors. TF mutations induce rippling downstream effects by simultaneously altering the expression of multiple genes. Mutations involving these TFs affect diverse aspects of megakaryocyte biology and platelet production and function, culminating in thrombocytopenia, platelet dysfunction, and associated clinical features. Mutations in TFs may occur more frequently in the patients with inherited platelet dysfunction than generally appreciated. This review focuses on the alterations in hematopoietic TFs in the pathobiology of inherited platelet dysfunction.
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Affiliation(s)
- Natthapol Songdej
- a Sol Sherry Thrombosis Research Center, and Hematology Section, Department of Medicine , Lewis Katz School of Medicine at Temple University , Philadelphia , PA , USA
| | - A Koneti Rao
- a Sol Sherry Thrombosis Research Center, and Hematology Section, Department of Medicine , Lewis Katz School of Medicine at Temple University , Philadelphia , PA , USA
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44
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Paes BCMF, Moço PD, Pereira CG, Porto GS, de Sousa Russo EM, Reis LCJ, Covas DT, Picanço-Castro V. Ten years of iPSC: clinical potential and advances in vitro hematopoietic differentiation. Cell Biol Toxicol 2016; 33:233-250. [PMID: 28039590 DOI: 10.1007/s10565-016-9377-2] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2016] [Accepted: 12/18/2016] [Indexed: 01/19/2023]
Abstract
Ten years have passed since the first publication announcing the generation of induced pluripotent stem cells (iPSCs). Issues related to ethics, immune rejection, and cell availability seemed to be solved following this breakthrough. The development of iPSC technology allows advances in in vitro cell differentiation for cell therapy purpose and other clinical applications. This review provides a perspective on the iPSC potential for cell therapies, particularly for hematological applications. We discuss the advances in in vitro hematopoietic differentiation, the possibilities to employ iPSC in hematology studies, and their potential clinical application in hematologic diseases. The generation of red blood cells and functional T cells and the genome editing technology applied to mutation correction are also covered. We highlight some of the requirements and obstacles to be overcome before translating these cells from research to the clinic, for instance, iPSC variability, genotoxicity, the differentiation process, and engraftment. Also, we evaluate the patent landscape and compile the clinical trials in the field of pluripotent stem cells. Currently, we know much more about iPSC than in 2006, but there are still challenges that must be solved. A greater understanding of molecular mechanisms underlying the generation of hematopoietic stem cells is necessary to produce suitable and transplantable hematopoietic stem progenitor cells from iPSC.
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Affiliation(s)
- Bárbara Cristina Martins Fernandes Paes
- Ribeirão Preto Medical School and Center for Cell-based Therapy (CTC), University of São Paulo, São Paulo, Brazil
- Regional Blood Center of Ribeirão Preto, Rua Tenente Catão Roxo, 2501, Ribeirão Preto, São Paulo, 14051-140, Brazil
| | - Pablo Diego Moço
- Ribeirão Preto Medical School and Center for Cell-based Therapy (CTC), University of São Paulo, São Paulo, Brazil
- Regional Blood Center of Ribeirão Preto, Rua Tenente Catão Roxo, 2501, Ribeirão Preto, São Paulo, 14051-140, Brazil
| | - Cristiano Gonçalves Pereira
- School of Economics, Business Administration and Accounting at Ribeirão Preto, University of São Paulo, São Paulo, Brazil
| | - Geciane Silveira Porto
- School of Economics, Business Administration and Accounting at Ribeirão Preto, University of São Paulo, São Paulo, Brazil
| | - Elisa Maria de Sousa Russo
- Regional Blood Center of Ribeirão Preto, Rua Tenente Catão Roxo, 2501, Ribeirão Preto, São Paulo, 14051-140, Brazil
- Ribeirão Preto Pharmaceutical Sciences School, University of São Paulo, São Paulo, Brazil
| | - Luiza Cunha Junqueira Reis
- Regional Blood Center of Ribeirão Preto, Rua Tenente Catão Roxo, 2501, Ribeirão Preto, São Paulo, 14051-140, Brazil
- Ribeirão Preto Pharmaceutical Sciences School, University of São Paulo, São Paulo, Brazil
| | - Dimas Tadeu Covas
- Ribeirão Preto Medical School and Center for Cell-based Therapy (CTC), University of São Paulo, São Paulo, Brazil
- Regional Blood Center of Ribeirão Preto, Rua Tenente Catão Roxo, 2501, Ribeirão Preto, São Paulo, 14051-140, Brazil
| | - Virginia Picanço-Castro
- Regional Blood Center of Ribeirão Preto, Rua Tenente Catão Roxo, 2501, Ribeirão Preto, São Paulo, 14051-140, Brazil.
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45
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Koeffler HP, Leong G. Preleukemia: one name, many meanings. Leukemia 2016; 31:534-542. [PMID: 27899806 PMCID: PMC5339433 DOI: 10.1038/leu.2016.364] [Citation(s) in RCA: 43] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2016] [Revised: 11/11/2016] [Accepted: 11/14/2016] [Indexed: 12/15/2022]
Abstract
Definition of preleukemia has evolved. It was first used to describe the myelodysplastic syndrome (MDS) with a propensity to progress to acute myeloid leukemia (AML). Individuals with germline mutations of either RUNX1, CEBPA, or GATA2 can also be called as preleukemic because they have a markedly increased incidence of evolution into AML. Also, alkylating chemotherapy or radiation can cause MDS/preleukemia, which nearly always progress to AML. More recently, investigators noted that AML patients who achieved complete morphological remission after chemotherapy often have clonal hematopoiesis predominantly marked by either DNMT3A, TET2 or IDH1/2 mutations, which were also present at diagnosis of AML. This preleukemic clone represents involvement of an early hematopoietic stem cells, which is resistant to standard therapy. The same clonal hematopoietic mutations have been identified in older ‘normal' individuals who have a modest increased risk of developing frank AML. These individuals have occasionally been said, probably inappropriately, to have a preleukemia clone. Our evolving understanding of the term preleukemia has occurred by advancing technology including studies of X chromosome inactivation, cytogenetics and more recently deep nucleotide sequencing.
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Affiliation(s)
- H P Koeffler
- Department of Hematology and Oncology, Cedars-Sinai Medical Center, Los Angeles, CA, USA.,Cancer Science Institute of Singapore, National University of Singapore, Singapore.,National University Cancer Institute of Singapore, National University Hospital, Singapore
| | - G Leong
- Cancer Science Institute of Singapore, National University of Singapore, Singapore
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46
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Ripperger T, Tawana K, Kratz C, Schlegelberger B, Fitzgibbon J, Steinemann D. Clinical utility gene card for: Familial platelet disorder with associated myeloid malignancies. Eur J Hum Genet 2016; 24:ejhg2015278. [PMID: 26813945 PMCID: PMC4970691 DOI: 10.1038/ejhg.2015.278] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/31/2015] [Revised: 11/18/2015] [Accepted: 12/08/2015] [Indexed: 01/16/2023] Open
Affiliation(s)
- Tim Ripperger
- Institute of Human Genetics, Hannover
Medical School, Carl-Neuberg-Str. 1, 30625
Hannover, Germany
| | - Kiran Tawana
- Centre for Haemato-Oncology, Barts Cancer
Institute, Queen Mary University of London, London,
UK
| | - Christian Kratz
- Department of Paediatric Haematology
& Oncology, Hannover Medical School, Hannover,
Germany
| | - Brigitte Schlegelberger
- Institute of Human Genetics, Hannover
Medical School, Carl-Neuberg-Str. 1, 30625
Hannover, Germany
| | - Jude Fitzgibbon
- Centre for Haemato-Oncology, Barts Cancer
Institute, Queen Mary University of London, London,
UK
| | - Doris Steinemann
- Institute of Human Genetics, Hannover
Medical School, Carl-Neuberg-Str. 1, 30625
Hannover, Germany
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How I diagnose and manage individuals at risk for inherited myeloid malignancies. Blood 2016; 128:1800-1813. [PMID: 27471235 DOI: 10.1182/blood-2016-05-670240] [Citation(s) in RCA: 125] [Impact Index Per Article: 13.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2016] [Accepted: 07/15/2016] [Indexed: 01/24/2023] Open
Abstract
Although inherited hematopoietic malignancies have been reported clinically since the early twentieth century, the molecular basis for these diseases has only recently begun to be elucidated. Growing utilization of next-generation sequencing technologies has facilitated the rapid discovery of an increasing number of recognizable heritable hematopoietic malignancy syndromes while also deepening the field's understanding of the molecular mechanisms that underlie these syndromes. Because individuals with inherited hematopoietic malignancies continue to be underdiagnosed and are increasingly likely to be encountered in clinical practice, clinicians need to have a high index of suspicion and be aware of the described syndromes. Here, we present the methods we use to identify, test, and manage individuals and families suspected of having a hereditary myeloid malignancy syndrome. Finally, we address the areas of ongoing research in the field and encourage clinicians and researchers to contribute and collaborate.
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48
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Daly ME. Transcription factor defects causing platelet disorders. Blood Rev 2016; 31:1-10. [PMID: 27450272 DOI: 10.1016/j.blre.2016.07.002] [Citation(s) in RCA: 29] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/25/2016] [Revised: 06/10/2016] [Accepted: 07/12/2016] [Indexed: 01/19/2023]
Abstract
Recent years have seen increasing recognition of a subgroup of inherited platelet function disorders which are due to defects in transcription factors that are required to regulate megakaryopoiesis and platelet production. Thus, germline mutations in the genes encoding the haematopoietic transcription factors RUNX1, GATA-1, FLI1, GFI1b and ETV6 have been associated with both quantitative and qualitative platelet abnormalities, and variable bleeding symptoms in the affected patients. Some of the transcription factor defects are also associated with an increased predisposition to haematologic malignancies (RUNX1, ETV6), abnormal erythropoiesis (GATA-1, GFI1b, ETV6) and immune dysfunction (FLI1). The persistence of MYH10 expression in platelets is a surrogate marker for FLI1 and RUNX1 defects. Characterisation of the transcription factor defects that give rise to platelet function disorders, and of the genes that are differentially regulated as a result, are yielding insights into the roles of these genes in platelet formation and function.
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Affiliation(s)
- Martina E Daly
- Department of Infection, Immunity and Cardiovascular Disease, University of Sheffield Medical School, Beech Hill Road, Sheffield, S10 2RX, UK.
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Gene correction in patient-specific iPSCs for therapy development and disease modeling. Hum Genet 2016; 135:1041-58. [PMID: 27256364 DOI: 10.1007/s00439-016-1691-5] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2016] [Accepted: 05/18/2016] [Indexed: 12/20/2022]
Abstract
The discovery that mature cells can be reprogrammed to become pluripotent and the development of engineered endonucleases for enhancing genome editing are two of the most exciting and impactful technology advances in modern medicine and science. Human pluripotent stem cells have the potential to establish new model systems for studying human developmental biology and disease mechanisms. Gene correction in patient-specific iPSCs can also provide a novel source for autologous cell therapy. Although historically challenging, precise genome editing in human iPSCs is becoming more feasible with the development of new genome-editing tools, including ZFNs, TALENs, and CRISPR. iPSCs derived from patients of a variety of diseases have been edited to correct disease-associated mutations and to generate isogenic cell lines. After directed differentiation, many of the corrected iPSCs showed restored functionality and demonstrated their potential in cell replacement therapy. Genome-wide analyses of gene-corrected iPSCs have collectively demonstrated a high fidelity of the engineered endonucleases. Remaining challenges in clinical translation of these technologies include maintaining genome integrity of the iPSC clones and the differentiated cells. Given the rapid advances in genome-editing technologies, gene correction is no longer the bottleneck in developing iPSC-based gene and cell therapies; generating functional and transplantable cell types from iPSCs remains the biggest challenge needing to be addressed by the research field.
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50
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Li W, Morrone K, Kambhampati S, Will B, Steidl U, Verma A. Thrombocytopenia in MDS: epidemiology, mechanisms, clinical consequences and novel therapeutic strategies. Leukemia 2015; 30:536-44. [PMID: 26500138 DOI: 10.1038/leu.2015.297] [Citation(s) in RCA: 36] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2015] [Accepted: 08/03/2015] [Indexed: 12/14/2022]
Abstract
Thrombocytopenia is commonly seen in myelodysplastic syndrome (MDS) patients, and bleeding complications are a major cause of morbidity and mortality. Thrombocytopenia is an independent factor for decreased survival and has been incorporated in newer prognostic scoring systems. The mechanisms of thrombocytopenia are multifactorial and involve a differentiation block of megakaryocytic progenitor cells, leading to dysplastic, hypolobated and microscopic appearing megakaryocytes or increased apoptosis of megakaryocytes and their precursors. Dysregulated thrombopoietin (TPO) signaling and increased platelet destruction through immune or nonimmune mechanisms are frequently observed in MDS. The clinical management of patients with low platelet counts remains challenging and approved chemotherapeutic agents such as lenalidomide and azacytidine can also lead to a transient worsening of thrombocytopenia. Platelet transfusion is the only supportive treatment option currently available for clinically significant thrombocytopenia. The TPO receptor agonists romiplostim and eltrombopag have shown clinical activity in clinical trials in MDS. In addition to thrombopoietic effects, eltrombopag can inhibit leukemic cell proliferation via TPO receptor-independent effects. Other approaches such as treatment with cytokines, immunomodulating drugs and signal transduction inhibitors have shown limited activity in selected groups of MDS patients. Combination trials of approved agents with TPO agonists are ongoing and hold promise for this important clinical problem.
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Affiliation(s)
- W Li
- Department of Medicine, Albert Einstein College of Medicine/Jacobi Medical Center, Bronx, NY, USA
| | - K Morrone
- Department of Pediatrics, Albert Einstein College of Medicine/Montefiore Medical Center, Bronx, NY, USA
| | - S Kambhampati
- Department of Medicine, University of Kansas Medical Center, Kansas City, KS, USA
| | - B Will
- Division of Hemato-Oncology, Department of Oncology, Albert Einstein College of Medicine, Bronx, NY, USA
| | - U Steidl
- Division of Hemato-Oncology, Department of Oncology, Albert Einstein College of Medicine, Bronx, NY, USA
| | - A Verma
- Division of Hemato-Oncology, Department of Oncology, Albert Einstein College of Medicine, Bronx, NY, USA
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