bims-scepro Biomed News
on Stem cell proteostasis
Issue of 2024–07–28
nineteen papers selected by
William Grey, University of York
  1. DNA fork remodeling proteins, Zranb3 and Smarcal1, are uniquely essential for aging hematopoiesis.
  2. An NAD+-dependent metabolic checkpoint regulates hematopoietic stem cell activation and aging.
  3. Epidermal growth factor augments the self-renewal capacity of aged hematopoietic stem cells.
  4. Kmt2c restricts G-CSF-driven HSC mobilization and granulocyte production in a methyltransferase-independent manner.
  5. In vivo CRISPR/Cas9-mediated screen reveals a critical function of TFDP1 and E2F4 transcription factors in hematopoiesis.
  6. Microfluidic Sample Preparation for Multiplexed Single-Cell Proteomics Using a Nested Nanowell Chip.
  7. Targeting ferritinophagy impairs quiescent cancer stem cells in acute myeloid leukemia in vitro and in vivo models.
  8. Astrocytes control quiescent NSC reactivation via GPCR signaling-mediated F-actin remodeling.
  9. Immunophenotypic features of early haematopoietic and leukaemia stem cells.
  10. A global view of altered ligand-receptor interactions in bone marrow aging based on single-cell sequencing.
  11. Mechanistic insights into the developmental origin of pediatric hematological disorders.
  12. STAT3 in acute myeloid leukemia facilitates natural killer cell-mediated surveillance.
  13. Targeted Mass Spectrometry for Quantification of Receptor Tyrosine Kinase Signaling.
  14. Subunit-specific analysis of cohesin-mutant myeloid malignancies reveals distinct ontogeny and outcomes.
  15. A Multiplexed Single-Cell Proteomic Workflow Applicable to Drug Treatment Studies.
  16. Effective Prevention and Treatment of Acute Leukemias in Mice by Activation of Thermogenic Adipose Tissues.
  17. Unlocking the therapeutic potential of selective CDK7 and BRD4 Inhibition against multiple myeloma cell growth.
  18. A Tip-Based Workflow for Sensitive IMAC-Based Low Nanogram Level Phosphoproteomics.
  19. Protocol for in vitro co-culture, proliferation, and cell cycle analyses of patient-derived leukemia cells.
  1. Aging Cell. 2024 Jul 23. e14281
    DNA fork remodeling proteins, Zranb3 and Smarcal1, are uniquely essential for aging hematopoiesis.
    Saul Kushinsky, Matthew V Puccetti, Clare M Adams, Irina Shkundina, Nikkole James, Brittany M Mahon, Peter Michener, Christine M Eischen.
      Over a lifetime, hematopoietic stem and progenitor cells (HSPCs) are forced to repeatedly proliferate to maintain hematopoiesis, increasing their susceptibility to DNA damaging replication stress. However, the proteins that mitigate this stress, protect HSPC replication, and prevent aging-driven dysregulation are unknown. We report two evolutionarily conserved, ubiquitously expressed chromatin remodeling enzymes with similar DNA replication fork reversal biochemical functions, Zranb3 and Smarcal1, have surprisingly specialized roles in distinct HSPC populations. While both proteins actively mitigate replication stress and prevent DNA damage and breaks during lifelong hematopoiesis, the loss of either resulted in distinct biochemical and biological consequences. Notably, defective long-term HSC function, revealed with bone marrow transplantation, caused hematopoiesis abnormalities in young mice lacking Zranb3. Aging significantly worsened these hematopoiesis defects in Zranb3-deficient mice, including accelerating the onset of myeloid-biased hematopoietic dysregulation to early in life. Such Zranb3-deficient HSPC abnormalities with age were driven by accumulated DNA damage and replication stress. Conversely, Smarcal1 loss primarily negatively affected progenitor cell functions that were exacerbated with aging, resulting in a lymphoid bias. Simultaneous loss of both Zranb3 and Smarcal1 compounded HSPC defects. Additionally, HSPC DNA replication fork dynamics had unanticipated HSPC type and age plasticity that depended on the stress and Zranb3 and/or Smarcal1. Our data reveal both Zranb3 and Smarcal1 have essential HSPC cell intrinsic functions in lifelong hematopoiesis that protect HSPCs from replication stress and DNA damage in unexpected, unique ways.
    Keywords:  DNA replication stress; Smarcal1; Zranb3; aging; hematopoiesis; hematopoietic stem and progenitor cells (HSPC)
    DOI:  https://doi.org/10.1111/acel.14281
  2. Nat Aging. 2024 Jul 23.
    An NAD+-dependent metabolic checkpoint regulates hematopoietic stem cell activation and aging.
    Zehan Song, Sang Hee Park, Wei-Chieh Mu, Yufan Feng, Chih-Ling Wang, Yifei Wang, Marine Barthez, Ayane Maruichi, Jiayue Guo, Fanghan Yang, Anita Wong Lin, Kartoosh Heydari, Claudia C S Chini, Eduardo N Chini, Cholsoon Jang, Danica Chen.
      How hematopoietic stem cells (HSCs) maintain metabolic homeostasis to support tissue repair and regeneration throughout the lifespan is elusive. Here, we show that CD38, an NAD+-dependent metabolic enzyme, promotes HSC proliferation by inducing mitochondrial Ca2+ influx and mitochondrial metabolism in young mice. Conversely, aberrant CD38 upregulation during aging is a driver of HSC deterioration in aged mice due to dysregulated NAD+ metabolism and compromised mitochondrial stress management. The mitochondrial calcium uniporter, a mediator of mitochondrial Ca2+ influx, also supports HSC proliferation in young mice yet drives HSC decline in aged mice. Pharmacological inactivation of CD38 reverses HSC aging and the pathophysiological changes of the aging hematopoietic system in aged mice. Together, our study highlights an NAD+ metabolic checkpoint that balances mitochondrial activation to support HSC proliferation and mitochondrial stress management to enhance HSC self-renewal throughout the lifespan, and links aberrant Ca2+ signaling to HSC aging.
    DOI:  https://doi.org/10.1038/s43587-024-00670-8
  3. iScience. 2024 Jul 19. 27(7): 110306
    Epidermal growth factor augments the self-renewal capacity of aged hematopoietic stem cells.
    Vivian Y Chang, Yuwei He, Samantha Grohe, Morgan R Brady, Aldi Chan, Rucha S Kadam, Tiancheng Fang, Amara Pang, Katherine Pohl, Evelyn Tran, Michelle Li, Jenny Kan, Yurun Zhang, Josie J Lu, Joshua P Sasine, Heather A Himburg, Peibin Yue, John P Chute.
      Hematopoietic aging is associated with decreased hematopoietic stem cell (HSC) self-renewal capacity and myeloid skewing. We report that culture of bone marrow (BM) HSCs from aged mice with epidermal growth factor (EGF) suppressed myeloid skewing, increased multipotent colony formation, and increased HSC repopulation in primary and secondary transplantation assays. Mice transplanted with aged, EGF-treated HSCs displayed increased donor cell engraftment within BM HSCs and systemic administration of EGF to aged mice increased HSC self-renewal capacity in primary and secondary transplantation assays. Expression of a dominant negative EGFR in Scl/Tal1+ hematopoietic cells caused increased myeloid skewing and depletion of long term-HSCs in 15-month-old mice. EGF treatment decreased DNA damage in aged HSCs and shifted the transcriptome of aged HSCs from genes regulating cell death to genes involved in HSC self-renewal and DNA repair but had no effect on HSC senescence. These data suggest that EGFR signaling regulates the repopulating capacity of aged HSCs.
    Keywords:  Human physiology; cellular physiology; functional aspects of cell biology; molecular medicine; stem cells research
    DOI:  https://doi.org/10.1016/j.isci.2024.110306
  4. Cell Rep. 2024 Jul 23. pii: S2211-1247(24)00871-4. [Epub ahead of print]43(8): 114542
    Kmt2c restricts G-CSF-driven HSC mobilization and granulocyte production in a methyltransferase-independent manner.
    Helen C Wang, Ran Chen, Wei Yang, Yanan Li, Rohini Muthukumar, Riddhi M Patel, Emily B Casey, Elisabeth Denby, Jeffrey A Magee.
      Granulocyte colony-stimulating factor (G-CSF) is widely used to enhance myeloid recovery after chemotherapy and to mobilize hematopoietic stem cells (HSCs) for transplantation. Unfortunately, through the course of chemotherapy, cancer patients can acquire leukemogenic mutations that cause therapy-related myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML). This raises the question of whether therapeutic G-CSF might potentiate therapy-related MDS/AML by disproportionately stimulating mutant HSCs and other myeloid progenitors. A common mutation in therapy-related MDS/AML involves chromosome 7 deletions that inactivate many tumor suppressor genes, including KMT2C. Here, we show that Kmt2c deletions hypersensitize murine HSCs and myeloid progenitors to G-CSF, as evidenced by increased HSC mobilization and enhanced granulocyte production from granulocyte-monocyte progenitors (GMPs). Furthermore, Kmt2c attenuates the G-CSF response independently from its SET methyltransferase function. Altogether, the data raise concerns that monosomy 7 can hypersensitize progenitors to G-CSF, such that clinical use of G-CSF may amplify the risk of therapy-related MDS/AML.
    Keywords:  CP: Immunology; CP: Stem cell research
    DOI:  https://doi.org/10.1016/j.celrep.2024.114542
  5. Leukemia. 2024 Jul 23.
    In vivo CRISPR/Cas9-mediated screen reveals a critical function of TFDP1 and E2F4 transcription factors in hematopoiesis.
    Ngoc Tung Tran, Robin Graf, Ernesto Acevedo-Ochoa, Janine Trombke, Timm Weber, Thomas Sommermann, Claudia Salomon, Ralf Kühn, Klaus Rajewsky, Van Trung Chu.
      Hematopoiesis is a continuous process of blood cell production driven by hematopoietic stem and progenitor cells (HSPCs) in the bone marrow. Proliferation and differentiation of HSPCs are regulated by complex transcriptional networks. In order to identify transcription factors with key roles in HSPC-mediated hematopoietic reconstitution, we developed an efficient and robust CRISPR/Cas9-based in vivo genetic screen. Using this experimental system, we identified the TFDP1 transcription factor to be essential for HSPC proliferation and post-transplant hematopoiesis. We further discovered that E2F4, an E2F transcription factor, serves as a binding partner of TFDP1 and is required for HSPC proliferation. Deletion of TFDP1 caused downregulation of genes associated with the cell cycle, with around 50% of these genes being identified as direct targets of TFDP1 and E2F4. Thus, our study expands the transcriptional network governing hematopoietic development through an in vivo CRISPR/Cas9-based genetic screen and identifies TFDP1/E2F4 as positive regulators of cell cycle genes in HSPCs.
    DOI:  https://doi.org/10.1038/s41375-024-02357-w
  6. Methods Mol Biol. 2024 ;2823 141-154
    Microfluidic Sample Preparation for Multiplexed Single-Cell Proteomics Using a Nested Nanowell Chip.
    Junho Park, Tommy K Cheung, Ying Zhu.
      Mass spectrometry-based single-cell proteomics has undergone rapid progress and has become an active research area. However, because of the ultralow amount of proteins in single cells, it is still highly challenging to achieve efficient sample preparation and sensitive LC-MS detection. Here, we provide a detailed protocol for isobaric labeling-based single-cell proteomics relying on a microfluidic droplet-based sample processing technology. The protocol allows for processing both single cells and carrier samples in separate microchips using a commercially available platform (cellenONE) with high sample recovery and high throughput. We also provide an optimized LC-MS method for sensitive and robust data collection.
    Keywords:  Droplet; Mass spectrometry; Microfluidic; Multiplexed labeling; NanoPOTS; Sample preparation; Single-cell proteomics
    DOI:  https://doi.org/10.1007/978-1-0716-3922-1_10
  7. Sci Transl Med. 2024 Jul 24. 16(757): eadk1731
    Targeting ferritinophagy impairs quiescent cancer stem cells in acute myeloid leukemia in vitro and in vivo models.
    Clement Larrue, Sarah Mouche, Paolo Angelino, Maxime Sajot, Rudy Birsen, Olivier Kosmider, Thomas Mckee, François Vergez, Christian Recher, Véronique Mansat-De Mas, Qiong Gu, Jun Xu, Petros Tsantoulis, Jean-Emmanuel Sarry, Jerome Tamburini.
      Acute myeloid leukemia (AML) remains a challenging hematological malignancy with poor prognosis and limited treatment options. Leukemic stem cells (LSCs) contribute to therapeutic failure, relapse, and adverse outcome. This study investigates the role of quiescence and related molecular mechanisms in AML pathogenesis and LSC functions to identify potential therapeutic targets. Transcriptomic analysis revealed that the LSC-enriched quiescent cell population has a distinct gene signature with prognostic relevance in patients with AML. Mechanistically, quiescent blasts exhibit increased autophagic activity, which contributes to their sustained viability. Proteomic profiling uncovered differential requirements for iron metabolism between quiescent and cycling cells, revealing a unique dependence of quiescent cells on ferritinophagy, a selective form of autophagy mediated by nuclear receptor coactivator 4 (NCOA4), which regulates iron bioavailability. We evaluated the therapeutic potential of inhibiting NCOA4-mediated ferritinophagy using genetic knockdown and chemical inhibition approaches. In vitro assays showed that suppression of NCOA4 was toxic to leukemic blasts, particularly the CD34+CD38- LSC-enriched population, without affecting normal CD34+ hematopoietic progenitors. In vivo studies using murine patient-derived xenograft (PDX) models of AML confirmed that NCOA4 inhibition reduced tumor burden and impaired LSC viability and self-renewal, indicating a specific vulnerability of these cells to ferritinophagy disruption. Our findings underscore the role of NCOA4-mediated ferritinophagy in maintaining LSC quiescence and function and suggest that targeting this pathway may be an effective therapeutic strategy for AML. This study highlights the potential of NCOA4 inhibition to improve AML outcomes and paves the way for future research and clinical development.
    DOI:  https://doi.org/10.1126/scitranslmed.adk1731
  8. Sci Adv. 2024 Jul 26. 10(30): eadl4694
    Astrocytes control quiescent NSC reactivation via GPCR signaling-mediated F-actin remodeling.
    Kun-Yang Lin, Mahekta R Gujar, Jiaen Lin, Wei Yung Ding, Jiawen Huang, Yang Gao, Ye Sing Tan, Xiang Teng, Low Siok Lan Christine, Pakorn Kanchanawong, Yusuke Toyama, Hongyan Wang.
      The transitioning of neural stem cells (NSCs) between quiescent and proliferative states is fundamental for brain development and homeostasis. Defects in NSC reactivation are associated with neurodevelopmental disorders. Drosophila quiescent NSCs extend an actin-rich primary protrusion toward the neuropil. However, the function of the actin cytoskeleton during NSC reactivation is unknown. Here, we reveal the fine filamentous actin (F-actin) structures in the protrusions of quiescent NSCs by expansion and super-resolution microscopy. We show that F-actin polymerization promotes the nuclear translocation of myocardin-related transcription factor, a microcephaly-associated transcription factor, for NSC reactivation and brain development. F-actin polymerization is regulated by a signaling cascade composed of G protein-coupled receptor Smog, G protein αq subunit, Rho1 guanosine triphosphatase, and Diaphanous (Dia)/Formin during NSC reactivation. Further, astrocytes secrete a Smog ligand folded gastrulation to regulate Gαq-Rho1-Dia-mediated NSC reactivation. Together, we establish that the Smog-Gαq-Rho1 signaling axis derived from astrocytes, an NSC niche, regulates Dia-mediated F-actin dynamics in NSC reactivation.
    DOI:  https://doi.org/10.1126/sciadv.adl4694
  9. Int J Lab Hematol. 2024 Jul 24.
    Immunophenotypic features of early haematopoietic and leukaemia stem cells.
    Tom Reuvekamp, Costa Bachas, Jacqueline Cloos.
      Many tumours are organised in a hierarchical structure with at its apex a cell that can maintain, establish, and repopulate the tumour-the cancer stem cell. The haematopoietic stem cell (HSC) is the founder cell for all functional blood cells. Like HSCs, the leukaemia stem cells (LSC) are hypothesised to be the leukaemia-initiating cells, which have features of stemness such as self-renewal, quiescence, and resistance to cytotoxic drugs. Immunophenotypically, CD34+CD38- defines HSCs by adding lineage negativity and CD90+CD45RA-. At which stage of maturation the further differentiation is blocked, determines the type of leukaemia, and determines the immunophenotype of the LSC specific to the leukaemia type. No apparent LSC phenotype has been described in lymphoid leukaemia, and it is debated if a specific acute lymphocytic leukaemia-initiating cell is present, as all cells are capable of engraftment in a secondary mouse model. In chronic lymphocytic leukaemia, a B-cell clone is responsible for uncontrolled proliferation, not a specific LSC. In chronic and acute myeloid leukaemia, LSC is described as CD34+CD38- with the expression of a marker that is aberrantly expressed (LSC marker), such as CD45RA, CD123 or in the case of chronic myeloid leukaemia CD26. In acute myeloid leukaemia, the LSC load had prognostic relevance and might be a biomarker that can be used for monitoring and as an addition to measurable residual disease. However, challenges such as the CD34-negative immunophenotype need to be explored.
    Keywords:  immunophenotypic analysis; leukaemia; stem cell
    DOI:  https://doi.org/10.1111/ijlh.14348
  10. Comput Struct Biotechnol J. 2024 Dec;23 2754-2762
    A global view of altered ligand-receptor interactions in bone marrow aging based on single-cell sequencing.
    Wenbo Chen, Xin Chen, Lei Yao, Jing Feng, Fengyue Li, Yuxin Shan, Linli Ren, Chenjian Zhuo, Mingqian Feng, Shan Zhong, Chunjiang He.
      Altered cell-cell communication is a hallmark of aging, but its impact on bone marrow aging remains poorly understood. Based on a common and effective pipeline and single-cell transcriptome sequencing, we detected 384,124 interactions including 2575 ligand-receptor pairs and 16 non-adherent bone marrow cell types in old and young mouse and identified a total of 5560 significantly different interactions, which were then verified by flow cytometry and quantitative real-time PCR. These differential ligand-receptor interactions exhibited enrichment for the senescence-associated secretory phenotypes. Further validation demonstrated supplementing specific extracellular ligands could modify the senescent signs of hematopoietic stem cells derived from old mouse. Our work provides an effective procedure to detect the ligand-receptor interactions based on single-cell sequencing, which contributes to understand mechanisms and provides a potential strategy for intervention of bone marrow aging.
    Keywords:  Aging; Bone marrow; Cell-cell interaction; Ligand and receptor; Single-cell transcriptome
    DOI:  https://doi.org/10.1016/j.csbj.2024.06.020
  11. Exp Hematol. 2024 Jul 24. pii: S0301-472X(24)00442-9. [Epub ahead of print] 104583
    Mechanistic insights into the developmental origin of pediatric hematological disorders.
    Giuseppina Camiolo, Christopher G Mullen, Katrin Ottersbach.
      Embryonic and fetal hematopoietic stem and progenitor cells differ in some key properties from cells that are part of the adult hematopoietic system. These include higher proliferation and self-renewal capacity, different globin gene usage and differing lineage biases. While these evolved to cover specific requirements of embryonic development, they can have serious consequences for the pathogenesis of hematological malignancies that initiate pre-birth in fetal blood cells and may result in a particularly aggressive disease that does not respond well to treatments that have been designed for adult leukemias. This indicates that these infant/pediatric leukemias should be considered as developmental diseases, where a thorough understanding of their unique biology is essential for designing more effective therapies. In this review, we will highlight some of these unique fetal properties and detail the underlying molecular drivers of these phenotypes. We specifically focus on those that are pertinent to disease pathogenesis and that may therefore reveal disease vulnerabilities. We have also included an extensive description of the origins, phenotype and key molecular drivers of the main infant and pediatric leukemias that have a known prenatal origin. Importantly, successes in recent years in generating faithful models of these malignancies in which cellular origins, key drivers and potential vulnerabilities can be investigated have resulted in uncovering potential, new therapeutic avenues.
    DOI:  https://doi.org/10.1016/j.exphem.2024.104583
  12. Front Immunol. 2024 ;15 1374068
    STAT3 in acute myeloid leukemia facilitates natural killer cell-mediated surveillance.
    Agnieszka Witalisz-Siepracka, Clio-Melina Denk, Bernhard Zdársky, Lorenz Hofmann, Sophie Edtmayer, Theresa Harm, Stefanie Weiss, Kerstin Heindl, Manuel Hessenberger, Sabrina Summer, Sayantanee Dutta, Emilio Casanova, Gerald J Obermair, Balázs Győrffy, Eva Maria Putz, Heinz Sill, Dagmar Stoiber.
      Acute myeloid leukemia (AML) is a heterogenous disease characterized by the clonal expansion of myeloid progenitor cells. Despite recent advancements in the treatment of AML, relapse still remains a significant challenge, necessitating the development of innovative therapies to eliminate minimal residual disease. One promising approach to address these unmet clinical needs is natural killer (NK) cell immunotherapy. To implement such treatments effectively, it is vital to comprehend how AML cells escape the NK-cell surveillance. Signal transducer and activator of transcription 3 (STAT3), a component of the Janus kinase (JAK)-STAT signaling pathway, is well-known for its role in driving immune evasion in various cancer types. Nevertheless, the specific function of STAT3 in AML cell escape from NK cells has not been deeply investigated. In this study, we unravel a novel role of STAT3 in sensitizing AML cells to NK-cell surveillance. We demonstrate that STAT3-deficient AML cell lines are inefficiently eliminated by NK cells. Mechanistically, AML cells lacking STAT3 fail to form an immune synapse as efficiently as their wild-type counterparts due to significantly reduced surface expression of intercellular adhesion molecule 1 (ICAM-1). The impaired killing of STAT3-deficient cells can be rescued by ICAM-1 overexpression proving its central role in the observed phenotype. Importantly, analysis of our AML patient cohort revealed a positive correlation between ICAM1 and STAT3 expression suggesting a predominant role of STAT3 in ICAM-1 regulation in this disease. In line, high ICAM1 expression correlates with better survival of AML patients underscoring the translational relevance of our findings. Taken together, our data unveil a novel role of STAT3 in preventing AML cells from escaping NK-cell surveillance and highlight the STAT3/ICAM-1 axis as a potential biomarker for NK-cell therapies in AML.
    Keywords:  AML; ICAM-1; NK cells; STAT3; immunotherapy
    DOI:  https://doi.org/10.3389/fimmu.2024.1374068
  13. Methods Mol Biol. 2024 ;2823 253-267
    Targeted Mass Spectrometry for Quantification of Receptor Tyrosine Kinase Signaling.
    Jeffrey R Whiteaker, Lei Zhao, Jacob J Kennedy, Richard G Ivey, Amanda G Paulovich.
      Targeted proteomics enables sensitive and specific quantification of proteins and post-translational modifications. By coupling peptide immunoaffinity enrichment with targeted mass spectrometry, we have developed the methodology for multiplexed quantification of proteins and phosphosites involved in the RAS/MAPK signaling network. The method uses anti-peptide antibodies to enrich analytes and heavy stable isotope-labeled internal standards, spiked in at known concentrations. The enriched peptides are directly measured by multiple-reaction monitoring (MRM), a well-characterized quantitative mass spectrometry-based method. The analyte (light) peptide response is measured relative to the heavy standard. The method described provides quantitative measurements of phospho-signaling and is generally applicable to other phosphopeptides and sample types.
    Keywords:   LC–MS; Multiplex; Phospho-signaling; Quantitation; RAS/MAPK; Targeted proteomics
    DOI:  https://doi.org/10.1007/978-1-0716-3922-1_16
  14. Leukemia. 2024 Jul 20.
    Subunit-specific analysis of cohesin-mutant myeloid malignancies reveals distinct ontogeny and outcomes.
    Johann-Christoph Jann, Christopher B Hergott, Marisa Winkler, Yiwen Liu, Benjamin Braun, Anne Charles, Kevin M Copson, Shougat Barua, Manja Meggendorfer, Niroshan Nadarajah, Shai Shimony, Eric S Winer, Martha Wadleigh, Richard M Stone, Daniel J DeAngelo, Jacqueline S Garcia, Torsten Haferlach, R Coleman Lindsley, Marlise R Luskin, Maximilian Stahl, Zuzana Tothova.
      Mutations in the cohesin complex components (STAG2, RAD21, SMC1A, SMC3, and PDS5B) are recurrent genetic drivers in myelodysplastic neoplasm (MDS) and acute myeloid leukemia (AML). Whether the different cohesin subunit mutations share clinical characteristics and prognostic significance is not known. We analyzed 790 cohesin-mutant patients from the Dana-Farber Cancer Institute (DFCI) and the Munich Leukemia Laboratory (MLL), 390 of which had available outcome data, and identified subunit-specific clinical, prognostic, and genetic characteristics suggestive of distinct ontogenies. We found that STAG2 mutations are acquired at MDS stage and are associated with secondary AML, adverse prognosis, and co-occurrence of secondary AML-type mutations. In contrast, mutations in RAD21, SMC1A and SMC3 share features with de novo AML with better prognosis, and co-occurrence with de novo AML-type lesions. The findings show the heterogeneous nature of cohesin complex mutations, and inform clinical and prognostic classification, as well as distinct biology of the cohesin complex.
    DOI:  https://doi.org/10.1038/s41375-024-02347-y
  15. Methods Mol Biol. 2024 ;2823 1-10
    A Multiplexed Single-Cell Proteomic Workflow Applicable to Drug Treatment Studies.
    Colten D Eberhard, Benjamin C Orsburn.
      It is now well accepted that individual cells within a population will respond to treatment of the same drug in a heterogenous manner. Recent advances have allowed, for the first time, the quantitative analysis of the proteomes of single human cells by mass spectrometry. A major focus of many groups, including our own, has been to use this emerging technology to rapidly identify subpopulations of cells with unique drug response and adaptation methods. While the technology in single-cell proteomics today is progressing at a truly staggering rate, we will detail our current methods for applying highly multiplexed single-cell proteomics to drug treatment studies.
    Keywords:  Drug treatment studies; Pharmacology; Single-cell proteomics
    DOI:  https://doi.org/10.1007/978-1-0716-3922-1_1
  16. Adv Sci (Weinh). 2024 Jul 25. e2402332
    Effective Prevention and Treatment of Acute Leukemias in Mice by Activation of Thermogenic Adipose Tissues.
    Ruibo Chen, Tianran Cheng, Sisi Xie, Xiaoting Sun, Mingjia Chen, Shumin Zhao, Qingyan Ruan, Xiaolei Ni, Mei Rao, Xinyi Quan, Kaiwen Chen, Shiyue Zhang, Tao Cheng, Yuanfu Xu, Yuguo Chen, Yunlong Yang, Yihai Cao.
      Acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) are common hematological malignancies in adults. Despite considerable research advances, the development of standard therapies, supportive care, and prognosis for the majority of AML and ALL patients remains poor and the development of new effective therapy is urgently needed. Here, it is reported that activation of thermogenic adipose tissues (TATs) by cold exposure or β3-adrenergic receptor agonists markedly alleviated the development and progression of AML and ALL in mouse leukemia models. TAT activation (TATA) monotherapy substantially reduces leukemic cells in bone marrow and peripheral blood, and suppresses leukemic cell invasion, including hepatomegaly and splenomegaly. Notably, TATA therapy prolongs the survivals of AML- and ALL-bearing mice. Surgical removal of thermogenic brown adipose tissue (BAT) or genetic deletion of uncoupling protein 1 (UCP1) largely abolishes the TATA-mediated anti-leukemia effects. Metabolomic pathway analysis demonstrates that glycolytic metabolism, which is essential for anabolic leukemic cell growth, is severely impaired in TATA-treated leukemic cells. Moreover, a combination of TATA therapy with chemotherapy produces enhanced anti-leukemic effects and reduces chemotoxicity. These data provide a new TATA-based therapeutic paradigm for the effective treatment of AML, ALL, and likely other types of hematological malignancies.
    Keywords:  cancer therapy; glycolysis; leukemia; metabolism; thermogenic adipose tissue
    DOI:  https://doi.org/10.1002/advs.202402332
  17. Haematologica. 2024 Jul 25.
    Unlocking the therapeutic potential of selective CDK7 and BRD4 Inhibition against multiple myeloma cell growth.
    Yao Yao, Shuhui Deng, Jessica Fong Ng, Mei Yuan, Chandraditya Chakraborty, Vera JoyWeiler, Nikhil Munshi, Mariateresa Fulciniti.
      Multiple myeloma (MM) is a plasma cell malignancy considered incurable despite the recent therapeutic advances. Effective targeted therapies are therefore needed. Our previous studies proved that inhibiting CDK7 impairs the cell cycle and metabolic programs by disrupting E2F1 and MYC transcriptional activities, making it an appealing therapeutic target for MM. Given that CDK7 and BRD4 operate in two distinct regulatory axes in MM, we hypothesized that targeting these two complementary pathways simultaneously would lead to a deeper and more durable response. Indeed, combination therapy had superior activity against MM cell growth and viability, and induced apoptosis to a greater extent than single-agent therapy in both cell lines and patient cells. This synergistic activity was also observed in Waldenström's Macroglobulinemia (WM) cells and with other inhibitors of E2F1 activity. Dual inhibition effectively impaired the MYC and E2F transcriptional programs and MM tumor growth and progression in xenograft animal models, providing evidence for combination therapy's potential as a therapeutic strategy in MM and WM.
    DOI:  https://doi.org/10.3324/haematol.2024.285491
  18. Methods Mol Biol. 2024 ;2823 129-140
    A Tip-Based Workflow for Sensitive IMAC-Based Low Nanogram Level Phosphoproteomics.
    Chia-Feng Tsai, Chuan-Chih Hsu, Yi-Ting Wang, Hyeyoon Kim, Tao Liu.
      Analyzing the phosphoproteome at nanoscale poses a significant challenge, mainly due to the substantial sample loss from nonspecific surface adsorption during the enrichment of low stoichiometric phosphopeptides. Here, we describe a tandem tip-based phosphoproteomics sample preparation method capable of sequential sample cleanup and enrichment without the need for additional sample transfer, thereby minimizing sample loss. Integration of this method to our recently developed SOP (surfactant-assisted one-pot sample preparation) and iBASIL (improved boosting to amplify signal with isobaric labeling) approaches creates a streamlined workflow, enabling sensitive, high-throughput nanoscale phosphoproteomics measurements.
    Keywords:  Immobilized metal ion affinity chromatography (IMAC); Isobaric labeling; Mass spectrometry; Nanoscale; Phosphoproteome; Single cell; iBASIL
    DOI:  https://doi.org/10.1007/978-1-0716-3922-1_9
  19. STAR Protoc. 2024 Jul 20. pii: S2666-1667(24)00367-8. [Epub ahead of print]5(3): 103202
    Protocol for in vitro co-culture, proliferation, and cell cycle analyses of patient-derived leukemia cells.
    Jessica Parker, Sean Hockney, Carly Knill, David McDonald, Andrew Filby, Deepali Pal.
      Leukemia niche impacts quiescence; however, culturing patient-derived samples ex vivo is technically challenging. Here, we present a protocol for in vitro co-culture of patient-derived xenograft acute lymphoblastic leukemia (PDX-ALL) cells with human mesenchymal stem cells (MSCs). We describe steps for labeling PDX-ALL cells with CellTrace Violet dye to demonstrate MSC-primed PDX-ALL cycling. We then detail procedures to identify MSC-primed G0/quiescent PDX-ALL cells via Hoechst-33342/Pyronin Y live cell cycle analysis. For complete details on the use and execution of this protocol, please refer to Pal et al.1,2.
    Keywords:  Cell Differentiation; Cell-based Assays; Stem Cells
    DOI:  https://doi.org/10.1016/j.xpro.2024.103202
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