bims-scepro Biomed News
on Stem cell proteostasis
Issue of 2024–12–22
eleven papers selected by
William Grey, University of York
  1. Understanding human oncogene function and cooperativity in myeloid malignancy using iPSCs.
  2. Direct Interaction Between CD34+ Hematopoietic Stem Cells and Mesenchymal Stem Cells Reciprocally Preserves Stemness.
  3. Protocol for CRISPR-Cas9-mediated induction of KMT2A rearrangements in cell line and umbilical cord blood hematopoietic stem and progenitor cells.
  4. Transduction of Human Fetal Liver Hematopoietic CD34+ Stem and Progenitor Cells into a Cell Line by Enhancing Telomerase Activity.
  5. Progressive chromatin rewiring by ETO2::GLIS2 revealed in a human iPSC model of pediatric leukemia initiation.
  6. Immunocompetent mouse models of multiple myeloma.
  7. Functions of the native NPM1 protein and its leukemic mutant.
  8. Increased CSN5 expression enhances the sensitivity to lenalidomide in multiple myeloma cells.
  9. Hexamethylene amiloride induces lysosome-mediated cell death in multiple myeloma through transcription factor E3.
  10. Heme promotes venetoclax resistance in multiple myeloma through MEK-ERK signaling and purine biosynthesis.
  11. The ETS domain-containing hematopoietic transcription factor PU.1 mediates the induction of arachidonate 5-lipoxygenase by multi-walled carbon nanotubes in macrophages in vitro.
  1. Exp Hematol. 2024 Dec 12. pii: S0301-472X(24)00561-7. [Epub ahead of print] 104697
    Understanding human oncogene function and cooperativity in myeloid malignancy using iPSCs.
    Martina Sarchi, Sergei Doulatov.
      Myeloid malignancies are a spectrum of clonal disorders driven by genetic alterations that cooperatively confer aberrant self-renewal and differentiation of hematopoietic stem and progenitor cells (HSPCs). Induced pluripotent stem cells (iPSCs) can be differentiated into HSPCs and have been widely explored for modeling hematological disorders and cell therapies. More recently, iPSCs models have been applied to study the origins and pathophysiology of myeloid malignancies, motivated by the appreciation for the differences in human oncogene function and the need for genetically defined models that recapitulate leukemia development. In this review, we will provide a broad overview of the rationale, the challenges, practical aspects, history, and recent advances of iPSC models for modeling myeloid neoplasms. We will focus on the insights into the previously unknown aspects of human oncogene function and cooperativity gained through the use of these models. It is now safe to say that iPSC models are a mainstay of leukemia modeling "toolbox" alongside primary human cells from normal and patient sources.
    DOI:  https://doi.org/10.1016/j.exphem.2024.104697
  2. Cancers (Basel). 2024 Nov 27. pii: 3972. [Epub ahead of print]16(23):
    Direct Interaction Between CD34+ Hematopoietic Stem Cells and Mesenchymal Stem Cells Reciprocally Preserves Stemness.
    Rémi Safi, Tala Mohsen-Kanson, Farah Kouzi, Jamal El-Saghir, Vera Dermesrobian, Inés Zugasti, Kazem Zibara, Pablo Menéndez, Hiba El Hajj, Marwan El-Sabban.
       BACKGROUND/OBJECTIVES: A specialized microenvironment in the bone marrow, composed of stromal cells including mesenchymal stem cells (MSCs), supports hematopoietic stem cell (HSC) self-renewal, and differentiation bands play an important role in leukemia development and progression. The reciprocal direct interaction between MSCs and CD34+ HSCs under physiological and pathological conditions is yet to be fully characterized.
    METHODS: Here, we established a direct co-culture model between MSCs and CD34+ HSCs or MSCs and acute myeloid leukemia cells (THP-1, Molm-13, and primary cells from patients) to study heterocellular communication.
    RESULTS: Following MSCs-CD34+ HSCs co-culture, the expression of adhesion markers N-Cadherin and connexin 43 increased in both cell types, forming gap junction channels. Moreover, the clonogenic potential of CD34+ HSCs was increased. However, direct contact of acute myeloid leukemia cells with MSCs reduced the expression levels of connexin 43 and N-Cadherin in MSCs. The impairment in gap junction formation may potentially be due to a defect in the acute myeloid leukemia-derived MSCs. Interestingly, CD34+ HSCs and acute myeloid leukemia cell lines attenuated MSC osteoblastic differentiation upon prolonged direct cell-cell contact.
    CONCLUSIONS: In conclusion, under physiological conditions, connexin 43 and N-Cadherin interaction preserves stemness of both CD34+ HSCs and MSCs, a process that is compromised in acute myeloid leukemia, pointing to the possible role of gap junctions in modulating stemness.
    Keywords:  CD34+ hematopoietic stem cells; N-Cadherin; acute myeloid leukemia; bone marrow; connexin 43; gap junction; heterocellular interaction; mesenchymal stem cells; microenvironment
    DOI:  https://doi.org/10.3390/cancers16233972
  3. STAR Protoc. 2024 Dec 17. pii: S2666-1667(24)00646-4. [Epub ahead of print]6(1): 103481
    Protocol for CRISPR-Cas9-mediated induction of KMT2A rearrangements in cell line and umbilical cord blood hematopoietic stem and progenitor cells.
    Tamara Benz, Patrizia Larghero, Claus Meyer, Marcel Müller, Dörthe Brüggmann, Anna-Elisabeth Hentrich, Frank Louwen, Estelle Erkner, Rahel Fitzel, Corina Schneidawind, Rolf Marschalek.
      KMT2A rearrangements are associated with a poor clinical outcome in infant, pediatric, and adult acute lymphoblastic and myeloid leukemia. Here, we present a protocol to reconstruct chromosomal translocations with different partner genes of KMT2A in vitro. We describe steps for patient-specific single guide RNA (sgRNA) design, optimized sgRNA in vitro transcription, detailed purification of hematopoietic stem and progenitor cells (HSPCs) from umbilical cord blood (UCB), and CRISPR-Cas9 editing of the test cell line K562 as well as UCB HSPCs. The provided methodology is donor independent.
    Keywords:  CRISPR; Cancer; Genetics; Stem Cells
    DOI:  https://doi.org/10.1016/j.xpro.2024.103481
  4. Int J Hematol Oncol Stem Cell Res. 2024 Oct 01. 18(4): 330-343
    Transduction of Human Fetal Liver Hematopoietic CD34+ Stem and Progenitor Cells into a Cell Line by Enhancing Telomerase Activity.
    Rashmi Bhardwaj, Lalit Kumar, Deepika Chhabra, Atul Sharma, Sujata Mohanty, Narinder Mehra, Vinod Kochupillai.
      Background: Human fetal liver hematopoietic stem cells have proven potential as therapeutics but lack extensive research due to their limited supply. Even in vitro expanded fetal liver hematopoietic stem cells enter senescence or lose their self-renewal capacity after a few days in culture. The present study aimed to obtain a homogeneous and persistent supply of hematopoietic stem cells from the fetal liver by establishing a cell line through immortalization of cells by enhancing telomerase activity. Materials and Methods: Human fetal liver hematopoietic CD34+ stem and progenitor cells were transformed and immortalized using retroviruses carrying the human telomerase (hTERT) gene. Following transduction, telomerase activity was assessed using the TRAP assay and telomere length was examined by Southern blotting in transduced cells. Their characterization was conducted using flowcytometry to analyze the CD34+ population of hematopoietic stem cells and their colony forming potential using colony forming unit (CFU) assay. Results: After transduction with hTERT, the life span of human fetal liver hematopoietic CD34+ stem and progenitor cells were extended to 80 population doublings, without any change in cell morphology or population doubling times. Constitutive hTERT expression enhanced the replicative capacity and prevented terminal differentiation of CD34+ fetal liver hematopoietic stem and progenitor cells (FLHSPCs). Moreover, hTERT-transduced stem cells maintained their telomere length and telomerase activity. Conclusion: By introducing telomerase activity into hematopoietic stem and progenitor cells, their lifespan can be extended while maintaining stemness. These modified cells hold promise for in vitro research focused on studying hematopoietic stem cells derived from fetal liver.
    Keywords:  Fetal liver; Hematopoietic stem cells; Immortalization; Telomerase activity, human telomerase reverse transcriptase (hTERT)
    DOI:  https://doi.org/10.18502/ijhoscr.v18i4.16758
  5. Blood. 2024 Dec 10. pii: blood.2024024505. [Epub ahead of print]
    Progressive chromatin rewiring by ETO2::GLIS2 revealed in a human iPSC model of pediatric leukemia initiation.
    Fabien Boudia, Marie Baille, Loelia Babin, Zakia Aid, Elie Robert, Julie Riviere, Klaudia Galant, Verónica Alonso-Pérez, Laura Anselmi, Brahim Arkoun, Nassera Abermil, Christophe Marzac, Salvatore Nicola Bertuccio, Alexia Regnault de Premesnil, Cecile K Lopez, Alexandre Eeckhoutte, Audrey Naimo, Betty Leite, Cyril Catelain, Christophe Metereau, Patrick Gonin, Nathalie Gaspar, Juerg Schwaller, Olivier A Bernard, Hana Raslova, Murielle Gaudry, Antonin Marchais, Hélène Lapillonne, Arnaud Petit, Francoise Pflumio, Marie-Laure Arcangeli, Erika Brunet, Thomas Mercher.
      Pediatric acute myeloid leukemia frequently harbor fusion oncogenes associated with poor prognosis, including KMT2A, NUP98 and GLIS2 rearrangements. While murine models have demonstrated their leukemogenic activities, the steps from a normal human cell to leukemic blasts remain unclear. Here, we precisely reproduced the inversion of chromosome 16 resulting in ETO2::GLIS2 fusion in human induced pluripotent stem cells (iPSC). IPSC-derived ETO2::GLIS2-expressing hematopoietic cells showed differentiation alterations in vitro and efficiently induced in vivo development of leukemia that closely phenocopied human acute megakaryoblastic leukemia (AMKL) reflected by flow cytometry and single cell transcriptomes. Comparison of iPS-derived cells with patient-derived cells revealed altered chromatin accessibility at early and later bona fide leukemia stages with aberrantly higher accessibility and expression of the osteogenic homeobox factor DLX3 that preceded increased accessibility to ETS factors. DLX3 overexpression in normal CD34+ cells increased accessibility to ETS motifs and reduced accessibility to GATA motifs. A DLX3 transcriptional module was globally enriched in both ETO2::GLIS2 AMKL and some aggressive pediatric osteosarcoma. Importantly, DLX3 knock-out abrogated leukemia initiation in this ETO2::GLIS2 iPSC model. Collectively, characterization of a novel human iPSC-derived AMKL model revealed hijacking of the osteogenic homeobox transcription factor DLX3 as an essential early step in chromatin changes and leukemogenesis driven by the ETO2::GLIS2 fusion oncogene.
    DOI:  https://doi.org/10.1182/blood.2024024505
  6. Semin Hematol. 2024 Nov 16. pii: S0037-1963(24)00127-6. [Epub ahead of print]
    Immunocompetent mouse models of multiple myeloma.
    Peter Leif Bergsagel, Marta Chesi.
      Immunocompetent murine models of multiple myeloma are critical for understanding the pathogenesis of multiple myeloma and for the development of novel immunotherapeutics. Different models are available in Balb/c and C57Bl strains, each with different advantages and disadvantages. The availability of many transplantable cell lines allows for the conduct of experiments with large cohorts of mice bearing identical tumors, while cell lines that grow in vitro can be used for genetic manipulations. The introduction of human CRBN into these models allows for the study of IMiDs and cereblon based PROTACs in mice. New genetically engineered models based on germinal center cell activation of Nsd2 or Ccnd1 together with constitutive NFkB are being developed to model some of the important genetic subtypes of human multiple myeloma.
    Keywords:  Human CRBN; IMiDs; Immunotherapy; MYC; Mouse models; Multiple myeloma
    DOI:  https://doi.org/10.1053/j.seminhematol.2024.11.003
  7. Leukemia. 2024 Dec 17.
    Functions of the native NPM1 protein and its leukemic mutant.
    Brunangelo Falini, Daniele Sorcini, Vincenzo Maria Perriello, Paolo Sportoletti.
      The nucleophosmin (NPM1) gene encodes for the most abundant nucleolar protein. Thanks to its property to act as histone chaperone and to shuttle between the nucleus and cytoplasm, the NPM1 protein is involved in multiple cellular function that are here extensively reviewed and include the formation of the nucleolus through liquid-liquid phase separation, regulation of ribosome biogenesis and transport, control of DNA repair and centrosome duplication as well as response to nucleolar stress. NPM1 is mutated in about 30-35% of adult acute myeloid leukemia (AML). Due to its unique biological and clinical features, NPM1-mutated AML is regarded as a distinct leukemia entity in the WHO 5th edition and ICC classifications of myeloid malignancies. The NPM1 mutant undergoes changes at the C-terminus of the protein that leads to its delocalization in the cytoplasm of the leukemic cells. Here, we focus also on its biological functions discussing the murine models of NPM1 mutations and the various mechanisms that occur at cytoplasmic and nuclear levels to promote and maintain NPM1-mutated AML.
    DOI:  https://doi.org/10.1038/s41375-024-02476-4
  8. iScience. 2024 Dec 20. 27(12): 111399
    Increased CSN5 expression enhances the sensitivity to lenalidomide in multiple myeloma cells.
    Takumi Yamamoto, Arisu Furukawa, Yue Zhou, Nobuaki Kono, Shojiro Kitajima, Hiroto Ohguchi, Yawara Kawano, Shingo Ito, Norie Araki, Sumio Ohtsuki, Takeshi Masuda.
      Lenalidomide (LEN) is commonly used as an effective therapeutic agent for multiple myeloma (MM). However, in some patients, primary resistance to LEN is observed, the mechanisms of which remain poorly understood. In this study, we combined a LEN sensitivity assay with proteomics data from 15 MM cell lines to identify protein expression profiles associated with primary LEN resistance. Our findings revealed that CSN5 expression is lower in LEN-resistant cell lines than in LEN-sensitive lines. Moreover, we established that CSN5 is degraded via the cullin-RING ubiquitin ligase (CRL)-mediated ubiquitin-proteasome pathway through ubiquitination at lysine 194. Our data suggest that reduced CSN5 expression leads to abnormalities in the ubiquitination cycle of CRL4A, resulting in the inhibition of LEN-mediated degradation of IKZF1 and IKZF3. These findings delineate an additional mechanism of LEN resistance in MM cells and may contribute to the development of alternative therapeutic strategies to overcome LEN resistance.
    Keywords:  Cancer; Cell biology; Molecular biology; Proteomics
    DOI:  https://doi.org/10.1016/j.isci.2024.111399
  9. Cell Death Discov. 2024 Dec 18. 10(1): 505
    Hexamethylene amiloride induces lysosome-mediated cell death in multiple myeloma through transcription factor E3.
    Nianhui Yang, Zexuan Dong, Weihao Xiao, Suqi Deng, Yizhen Li, Lei Hua, Yue Li, Yingying Wu, Kexiu Huang, Wei Zhou, Hua Wang, Yonghua Li, Juan Du, Hui Zeng.
      Multiple myeloma (MM) is the second common hematological malignancy characterized by the abnormal proliferation of plasma cells. Although advances in the past decades have led to improved outcomes and longer survival, MM remains largely incurable. New targets and targeted therapy may help to achieve better outcomes. Proton exporter NHE1 is highly expressed by tumor cells to maintain pH gradient for their survival and its inhibitor Hexamethylene amiloride (HA) has been demonstrated anti-tumor effect. However, whether HA could inhibit MM remains unknown. In this study, we firstly demonstrated that elevated expression level of NHE1 is associated with poor prognosis of MM. Moreover, the NHE1 inhibitor HA inhibited growth and induced apoptosis effectively in both MM cell lines and primary bone marrow cells from MM patients. Mechanistically, inhibitory effect was achieved partially through TFE3-mediated lysosomal production. With a MM xenograft mouse model, we verified that HA has a significant anti MM effect in vivo. Importantly, HA induced apoptosis of the carfilzomib-resistant MM cells and enhanced the effect of carfilzomib in MM. In summary, we demonstrated that NHE1 inhibitor HA can effectively inhibit MM growth both in vitro and in vivo, providing a new therapeutic strategy for improved outcome of de novo and resistant MM.
    DOI:  https://doi.org/10.1038/s41420-024-02269-9
  10. Blood. 2024 Dec 18. pii: blood.2024025690. [Epub ahead of print]
    Heme promotes venetoclax resistance in multiple myeloma through MEK-ERK signaling and purine biosynthesis.
    Remya Nair, An Vu, Abigail K Freer, Karanpreet S Bhatia, Dongxue Wang, Milan R Savani, Shannon M Matulis, Sagar Lonial, David L Jaye, Lawrence H Boise, Seung-Yong Seo, Timothy William Corson, Ajay K Nooka, Shruti Bhatt, Samuel K McBrayer, Vikas A Gupta, Xin Hu, Benjamin G Barwick, Amit R Reddi, Mala Shanmugam.
      We previously demonstrated that reduced intrinsic electron transport chain (ETC) activity predicts and promotes sensitivity to the BCL-2 antagonist, venetoclax (Ven) in multiple myeloma (MM). Heme, an iron-containing prosthetic group, and metabolite is fundamental to maintaining ETC activity. Interrogation of the CD2 subgroup of MM from the CoMMpass trial (NCT01454297), which can be used as a proxy for Ven-sensitive MM (VS MM), shows reduced expression of the conserved heme biosynthesis pathway gene signature. Consistent with this, we identified that VS MM exhibit reduced heme biosynthesis and curiously elevated hemin (oxidized heme) uptake. Supplementation with hemin or protoporphyrin IX (heme lacking iron) promotes Ven resistance while targeting ferrochetalase, the penultimate enzyme involved in heme biosynthesis, increases Ven sensitivity in cell lines and primary MM cells. Mechanistically, heme-mediated activation of pro-survival RAS-RAF-MEK signaling and metabolic rewiring, increasing de novo purine synthesis, were found to contribute to heme-induced Ven resistance. Co-targeting BCL-2 and MCL-1 suppresses heme-induced Ven resistance. Interrogation of the MMRF CoMMpass study of patients shows increased purine and pyrimidine biosynthesis to corelate with poor progression free survival and overall survival. Elevated heme and purine biosynthesis gene signatures were also observed in matched relapse refractory MM, underscoring the relevance of heme metabolism in therapy refractory MM. Overall, our findings reveal for the first time a role for extrinsic heme, a physiologically relevant metabolite, in modulating proximity to the apoptotic threshold with translational implications for BCL-2 antagonism in MM therapy.
    DOI:  https://doi.org/10.1182/blood.2024025690
  11. Arch Toxicol. 2024 Dec 17.
    The ETS domain-containing hematopoietic transcription factor PU.1 mediates the induction of arachidonate 5-lipoxygenase by multi-walled carbon nanotubes in macrophages in vitro.
    Chol Seung Lim, Ja Kook Gu, Qiang Ma.
      Exposure to fibrogenic multi-walled carbon nanotubes (MWCNTs) induces the production of proinflammatory lipid mediators (LMs) in myeloid cells to instigate inflammation. The molecular underpinnings of LM production in nanotoxicity remain unclear. Here we report that PU.1, an ETS domain-containing master regulator of hematopoiesis, critically regulates the induction of arachidonate 5-lypoxygenase (Alox5) and the production of LMs. MWCNTs (Mitsui-7) at 2.5 or 10 µg/mL induced the expression of Alox5 in murine and human macrophages at both mRNA and protein levels, accompanied by marked elevation of chemotactic LM leukotriene B4 (LTB4). Induction is comparable to those by potent M1 inducers. Carbon black, an amorphous carbon material control, did not increase Alox5 expression or LTB4 production at equivalent doses. MWCNTs induced the expression of a heterologous luciferase reporter under the control of the murine Alox5 promoter. Deletional analysis of the 2 kb promoter uncovered multiple inhibitory and activating activities. The proximal 250 bp region had the largest activation that was further increased by MWCNTs. The Alox5 promoter contains four PU box-like enhancers. PU.1 bond to each of the enhancers constitutively, which was further increased by MWCNTs. Knockdown of PU.1 using specific small hairpin-RNA blocked the basal and induced expression of Alox5 and the production of LTB4 as well as prostaglandin E2. The results demonstrate a critical role of PU.1 in mediating MWCNTs-induced expression of Alox5 and production of proinflammatory LMs, revealing a molecular framework where the hematopoietic transcription factor PU.1 is activated to orchestrate multiple proinflammatory responses to sterile particulates.
    Keywords:  Alox5; Inflammation; MWCNT; Macrophage; PU.1
    DOI:  https://doi.org/10.1007/s00204-024-03925-w
This page is being maintained by Thomas Krichel. It was last updated on 2024‒12‒22 at 10:03.