bims-scepro Biomed News
on Stem cell proteostasis
Issue of 2024‒10‒06
24 papers selected by
William Grey, University of York
  1. Hematopoietic and leukemic stem cells homeostasis: the role of bone marrow niche.
  2. The epigenetic state of the cell of origin defines mechanisms of leukemogenesis.
  3. Endocytosis as a critical regulator of hematopoietic stem cell fate -implications for hematopoietic stem cell and gene therapy.
  4. Essential role of Dhx16-mediated ribosome assembly in maintenance of hematopoietic stem cells.
  5. BST2 Facilitates Activation of Hematopoietic Stem Cells Through ERK Signaling.
  6. Obesity modulates hematopoietic stem cell fate decision via IL-1β induced p38/MAPK signaling pathway.
  7. Epitope prime editing shields hematopoietic cells from CD123 immunotherapy for acute myeloid leukemia.
  8. Linker histone regulates the myeloid versus lymphoid bifurcation of multipotent hematopoietic stem and progenitors.
  9. CD37 in acute myeloid leukemia: a novel surface target for drug delivery.
  10. Selective translation of nuclear mitochondrial respiratory proteins reprograms succinate metabolism in AML development and chemoresistance.
  11. Thioredoxin-interacting protein (TXNIP) is a substrate of the NEDD4-like E3 ubiquitin-protein ligase WWP1 in cellular redox state regulation of acute myeloid leukemia cells.
  12. Cellular taxonomy of the preleukemic bone marrow niche of acute myeloid leukemia.
  13. CRISPR/Cas9 editing of NKG2A improves the efficacy of primary CD33-directed chimeric antigen receptor natural killer cells.
  14. Peripheral human red blood cell development in human immune system mouse model with heme oxygenase-1 deficiency.
  15. Highly effective strategy for isolation of mononuclear cells from frozen cord blood.
  16. TMEM56 deficiency impairs the haem metabolism and cell cycle progression during human erythropoiesis.
  17. A New Era of Functional Experimentation in Human Hematopoiesis and Leukemia Research.
  18. PTPN11 Mutation Clonal Hierarchy in Acute Myeloid Leukemia.
  19. Temporal alterations of the nascent proteome in response to mitochondrial stress.
  20. RNA m5C oxidation by TET2 regulates chromatin state and leukaemogenesis.
  21. Tuft cells act as regenerative stem cells in the human intestine.
  22. Network pharmacology-based investigation and experimental validation of the mechanism of metformin in the treatment of acute myeloid leukemia.
  23. Precise Modulation of Protein Degradation by Smart PROTACs.
  24. Cholesterol binding to VCAM-1 promotes vascular inflammation.
  1. Explor Target Antitumor Ther. 2024 ;5(5): 1027-1055
    Hematopoietic and leukemic stem cells homeostasis: the role of bone marrow niche.
    Shaimaa Khattab, Manal El Sorady, Ashraf El-Ghandour, Giuseppe Visani, Pier Paolo Piccaluga.
      The bone marrow microenvironment (BMM) has highly specialized anatomical characteristics that provide a sanctuary place for hematopoietic stem cells (HSCs) that allow appropriate proliferation, maintenance, and self-renewal capacity. Several cell types contribute to the constitution and function of the bone marrow niche. Interestingly, uncovering the secrets of BMM and its interaction with HSCs in health paved the road for research aiming at better understanding the concept of leukemic stem cells (LSCs) and their altered niche. In fact, they share many signals that are responsible for interactions between LSCs and the bone marrow niche, due to several biological similarities between LSCs and HSCs. On the other hand, LSCs differ from HSCs in their abnormal activation of important signaling pathways that regulate survival, proliferation, drug resistance, invasion, and spread. Targeting these altered niches can help in better treatment choices for hematological malignancies and bone marrow disorders in general and acute myeloid leukemia (AML) in particular. Moreover, targeting those niches may help in decreasing the emergence of drug resistance and lower the relapse rate. In this article, the authors reviewed the most recent literature on bone marrow niches and their relations with either normal HSCs and AML cells/LSC, by focusing on pathogenetic and therapeutic implications.
    Keywords:  Hematopoietic stem cell; acute myeloid leukemia; bone marrow niche; drug resistance; immune system; precision medicine; targeted therapy
    DOI:  https://doi.org/10.37349/etat.2024.00262
  2. Leukemia. 2024 Oct 01.
    The epigenetic state of the cell of origin defines mechanisms of leukemogenesis.
    Zhiheng Li, Sara Fierstein, Mayuri Tanaka-Yano, Katie Frenis, Chun-Chin Chen, Dahai Wang, Marcelo Falchetti, Parker Côté, Christina Curran, Kate Lu, Tianxin Liu, Stuart Orkin, Hojun Li, Edroaldo Lummertz da Rocha, Shaoyan Hu, Qian Zhu, R Grant Rowe.
      Acute myeloid leukemia (AML) shows variable clinical outcome. The normal hematopoietic cell of origin impacts the clinical behavior of AML, with AML from hematopoietic stem cells (HSCs) prone to chemotherapy resistance in model systems. However, the mechanisms by which HSC programs are transmitted to AML are not known. Here, we introduce the leukemogenic MLL-AF9 translocation into defined human hematopoietic populations, finding that AML from HSCs is enriched for leukemic stem cells (LSCs) compared to AML from progenitors. By epigenetic profiling, we identify a putative inherited program from the normal HSC that collaborates with oncogene-driven programs to confer aggressive behavior in HSC-AML. We find that components of this program are required for HSC-AML growth and survival and identify RNA polymerase (RNAP) II-mediated transcription as a therapeutic vulnerability. Overall, we propose a mechanism as to how epigenetic programs from the leukemic cell of origin are inherited through transformation to impart the clinical heterogeneity of AML.
    DOI:  https://doi.org/10.1038/s41375-024-02428-y
  3. Stem Cell Res Ther. 2024 Sep 27. 15(1): 319
    Endocytosis as a critical regulator of hematopoietic stem cell fate -implications for hematopoietic stem cell and gene therapy.
    Prathibha Babu Chandraprabha, Saravanabhavan Thangavel.
      Hematopoietic stem cells (HSCs) have emerged as one of the most therapeutically significant adult stem cells, paving way for a range of novel curative regimens over decades. HSCs are transplanted, either directly or post restorative genetic engineering in order to repopulate a healthy hematopoietic homeostasis in patients with disorders affecting the blood and immune cells. Despite being an extensively studied system, the maintenance and expansion of functional HSCs ex vivo remains a major bottleneck. The challenge primarily stems from difficulties in reproducing HSC self-renewal divisions and gradual depletion of stemness characters, in vitro. Refining the in vitro culture can be particularly beneficial in the case of cord blood HSCs (CB-HSCs), as inadequate numbers in a single umbilical cord limits its therapeutic potential. In recent years, molecular dissection of HSC stemness has significantly improved in vitro hematopoietic stem and progenitor cells (HSPCs) culture. Despite such significant progress, lacunae exist in fully understanding all the underlying mechanisms and their interplay active in bona fide HSCs, and how it transforms when cells proliferate in culture. A new groundbreaking study titled "MYCT1 controls environmental sensing in human haematopoietic stem cells", published in Nature in June 2024, sheds light on this complex field. Through a series of experiments, including knock-down, overexpression, single-cell RNA sequencing, and transplantation, the study identifies a previously unknown role of the MYC target 1 (MYCT1) protein in HSC maintenance. This protein acts as a crucial regulator of human HSCs, with high expression in primitive HSCs and subsequently downregulated during ex vivo culture. The study reveals that MYCT1 plays a vital role in moderating endocytosis and environmental sensing in HSCs, processes thereby essential for maintaining HSC stemness and function. This commentary will discuss the implications of the new findings for cord blood expansion in cell therapies and HSPC culture for gene therapy applications, providing valuable insights for the field of hematopoietic regenerative medicine.
    Keywords:  Endocytosis; Ex vivo expansion; Gene-editing; HSPC gene therapy; MYCT1; Self-renewal
    DOI:  https://doi.org/10.1186/s13287-024-03927-6
  4. Leukemia. 2024 Sep 27.
    Essential role of Dhx16-mediated ribosome assembly in maintenance of hematopoietic stem cells.
    Zhigang Li, Jiankun Fan, Yalan Xiao, Wei Wang, Changlin Zhen, Junbing Pan, Weiru Wu, Yuanyuan Liu, Zhe Chen, Qinrong Yan, Hanqing Zeng, Shuyu Luo, Lun Liu, Zhanhan Tu, Xueya Zhao, Yu Hou.
      Hematopoietic stem cells (HSCs) are vital for the differentiation of all mature blood cells, with their homeostasis being tightly regulated by intrinsic and extrinsic factors. Alternative splicing, mediated by the spliceosome complex, plays a crucial role in regulating HSC homeostasis by increasing protein diversity. This study focuses on the ATP-dependent RNA helicase DHX16, a key spliceosome component, and its role in HSC regulation. Using conditional knockout mice, we demonstrate that loss of Dhx16 in the hematopoietic system results in significant depletion of hematopoietic stem and progenitor cells, bone marrow failure, and rapid mortality. Dhx16-deficient HSCs exhibit impaired quiescence, G2-M phase cell cycle arrest, reduced protein synthesis, abnormal ribosome assembly, increased apoptosis, and decreased self-renewal capacity. Multi-omics analysis identified intron 4 retention in Emg1 mRNA in Dhx16 knockout HSCs, leading to reduced EMG1 protein expression, disrupted ribosome assembly, and nucleolar stress, activating the p53 pathway. Overexpression of Emg1 in Dhx16-deficient HSCs partially restored ribosome assembly and HSC function, suggesting Emg1 as a potential therapeutic target for ribosomopathies. Our findings reveal the critical role of Dhx16 in HSC homeostasis through the regulation of alternative splicing and ribosome assembly, providing insights into the molecular mechanisms underlying hematopoietic diseases and potential therapeutic strategies.
    DOI:  https://doi.org/10.1038/s41375-024-02423-3
  5. Exp Hematol. 2024 Oct 01. pii: S0301-472X(24)00518-6. [Epub ahead of print] 104653
    BST2 Facilitates Activation of Hematopoietic Stem Cells Through ERK Signaling.
    Marcus A Florez, Apoorva Thatavarty, Duy T Le, Holly A Hill, Youngjae Jeong, Brian M Ho, Pawel Kus, Trisha K Wathan, Bailee N Kain, Shixia Huang, Dongsu Park, Katherine Y King.
      The proinflammatory cytokine interferon-gamma (IFNγ) is upregulated in a variety of infections and contributes to bone marrow failure through hematopoietic stem cell (HSC) activation and subsequent exhaustion. The cell surface protein, bone marrow stromal antigen 2 (BST2), is a key mediator of this process, as it is induced upon interferon stimulation and required for interferon-dependent HSC activation. To identify the mechanism by which BST2 promotes interferon-dependent HSC activation, we evaluated its role in niche localization, immune cell function, lipid raft formation, and intracellular signaling. Our studies indicated that knock out (KO) of BST2 in a murine model does not disrupt immune cell responses to interferon-inducing mycobacterial infection. Furthermore, intravital imaging studies indicate that BST2 KO does not disrupt localization of HSCs relative to endothelial or osteoblastic niches in the bone marrow. However, using imaging-based flow cytometry, we found that IFNγ treatment shifts the lipid raft polarity of WT but not Bst2-/- hematopoietic stem and progenitor cells (HSPCs). Furthermore, RNAseq analysis, reverse phase protein array and western blot analysis of HSPCs indicate that BST2 promotes ERK1/2 phosphorylation during IFNγ-mediated stress. Overall, we find that BST2 facilitates HSC division by promoting cell polarization and ERK activation, thus elucidating a key mechanism of interferon-dependent HSPC activation. These findings inform future approaches in the treatment of cancer and bone marrow failure.
    Keywords:  BST2; CD317; hematopoietic stem cells; inflammation; interferon; stem cell activation; tetherin
    DOI:  https://doi.org/10.1016/j.exphem.2024.104653
  6. Stem Cell Res Ther. 2024 Sep 29. 15(1): 336
    Obesity modulates hematopoietic stem cell fate decision via IL-1β induced p38/MAPK signaling pathway.
    Jinxiao Yan, Pan Zhang, Xiru Liu, Chengwei Pan, Guolin Shi, Penghui Ye, Xiaohang Zou, Xiang Li, Xinmin Zheng, Yu Liu, Hui Yang.
      BACKGROUND: Obesity is accompanied by inflammation, which significantly affects the homeostasis of the immune microenvironment. Hematopoietic stem cells (HSCs), residing primarily in the bone marrow, play a vital role in maintaining and producing diverse mature blood cell lineages for the adult hematopoietic and immune systems. However, how HSCs development is affected by obese-promoting inflammation, and the mechanism by which HSC hematopoietic potency is affected by inflammatory signals originating from the obese-promoting changes on bone marrow niche remain unclear. This study elucidates the relationship between obesity-promoting inflammation and HSC fate determination.METHODS: The obesity mice model was established by feeding C57BL/6J mice a high-fat diet (HFD) containing 60% kcal fat. After 6 weeks, HSCs were analyzed using flow cytometry and identified key inflammation cytokine. Transcriptome sequencing techniques were used to discern the distinct pathways in HSCs. Ultimately, confirming the biological mechanism of obesity-induced HSC fate changes via Anakinra blocking specific inflammatory signals.
    RESULTS: Obesity caused by HFD changed the physical and biochemical properties of the bone marrow niche. In the HFD mice, the population of long-term HSCs in the bone marrow was decreased and facilitated HSCs differentiation towards the myeloid lineage. In addition, HFD increased expression of the inflammatory factor IL-1β in the bone marrow, and a significantly increased expression of IL-1r1 and active p38/MAPK signaling pathway were detected in the HSCs. Inhibition of IL-1β further normalized the expression of genes in p38/MAPK pathway and reversed HSC fate.
    CONCLUSIONS: These findings have been demonstrated that the p38/MAPK signaling pathway in HSCs is activated by elevated levels of IL-1β within the HSC niche in obese models, thereby regulating HSC differentiation. It suggested a direct link between obesity-promoting inflammation and myeloid differentiation bias of HSCs in the HFD mice.
    Keywords:  Bone marrow niche; Hematopoietic stem cells; IL-1β; Inflammation; Obesity
    DOI:  https://doi.org/10.1186/s13287-024-03915-w
  7. Cell Stem Cell. 2024 Sep 26. pii: S1934-5909(24)00317-5. [Epub ahead of print]
    Epitope prime editing shields hematopoietic cells from CD123 immunotherapy for acute myeloid leukemia.
    Rui-Jin Ji, Guo-Hua Cao, Wei-Qiang Zhao, Mu-Yao Wang, Pan Gao, Yi-Zhou Zhang, Xue-Bin Wang, Hou-Yuan Qiu, Di-Di Chen, Xiao-Han Tong, Min Duan, Hao Yin, Ying Zhang.
      Acute myeloid leukemia (AML) is a malignant cancer characterized by abnormal differentiation of hematopoietic stem and progenitor cells (HSPCs). While chimeric antigen receptor T (CAR-T) cell immunotherapies target AML cells, they often induce severe on-target/off-tumor toxicity by attacking normal cells expressing the same antigen. Here, we used base editors (BEs) and a prime editor (PE) to modify the epitope of CD123 on HSPCs, protecting healthy cells from CAR-T-induced cytotoxicity while maintaining their normal function. Although BE effectively edits epitopes, complex bystander products are a concern. To enhance precision, we optimized prime editing, increasing the editing efficiency from 5.9% to 78.9% in HSPCs. Epitope-modified cells were resistant to CAR-T lysis while retaining normal differentiation and function. Furthermore, BE- or PE-edited HSPCs infused into humanized mice endowed myeloid lineages with selective resistance to CAR-T immunotherapy, demonstrating a proof-of-concept strategy for treating relapsed AML.
    Keywords:  CAR-T immunotherapy; CD123; HSPCs; acute myeloid leukemia; base editing; bystander products; epitope editing; hematopoietic stem and progenitor cell; on-target off-tumor toxicity; prime editing
    DOI:  https://doi.org/10.1016/j.stem.2024.09.003
  8. bioRxiv. 2024 Sep 17. pii: 2024.09.16.613227. [Epub ahead of print]
    Linker histone regulates the myeloid versus lymphoid bifurcation of multipotent hematopoietic stem and progenitors.
    Kutay Karatepe, Bruna Mafra de Faria, Jian Zhang, Xinyue Chen, Hugo Pinto, Dmitry Fyodorov, Esen Sefik, Michael Willcockson, Richard Flavell, Arthur Skoultchi, Shangqin Guo.
      Myeloid-biased differentiation of multipotent hematopoietic stem and progenitor cells (HSPCs) occurs with aging or exhaustion. The molecular mechanism(s) responsible for this fate bias remain unclear. Here we report that linker histone regulates HSPC fate choice at the lymphoid versus myeloid bifurcation. HSPCs expressing H1.0 from a doxycycline (dox) inducible transgene favor the lymphoid fate, display strengthened nucleosome organization and reduced chromatin accessibility at genomic regions hosting key myeloid fate drivers. The transcription factor Hlf is located in one of such regions, where chromatin accessibility and gene expression is reduced in H1.0 high HSPCs. Furthermore, H1.0 protein in HSPCs decreases in an aspartyl protease dependent manner, a process enhanced in response to interferon alpha (IFNα) signaling. Aspartyl protease inhibitors preserve endogenous H1.0 levels and promote the lymphoid fate of wild type HSPCs. Thus, our work uncovers a point of intervention to mitigate myeloid skewed hematopoiesis.
    DOI:  https://doi.org/10.1101/2024.09.16.613227
  9. Blood Adv. 2024 Sep 30. pii: bloodadvances.2024013590. [Epub ahead of print]
    CD37 in acute myeloid leukemia: a novel surface target for drug delivery.
    Erin Jeremy, Esthela Artiga, Sara Elgamal, Carolyn M Cheney, Dalen Eicher, Kevan Zalponik, Shelley Orwick, Hsiao-Yin Charlene Charlene Mao, Ronni Wasmuth, Bonnie K Harrington, Allison Mustonen, Peter Beshay, Patrick Halley, Carlos Castro, Katie E Williams, Zachary A Hing, Timothy L Chen, Christopher R Lucas, Nicholas J Vantangoli, Rosa Lapalombella, Nicole Renee Grieselhuber, Xiaokui M Mo, Erin Hertlein, Natarajan Muthusamy, Bethany L Mundy-Bosse, John C Byrd, Karilyn Larkin.
      Acute myeloid leukemia (AML) is the most common and lethal leukemia in adults. AML consists of many genetic subtypes which limits broad applicability of targeted therapy. We discovered that the hematopoietic restricted tetraspanin CD37 is expressed on all primary AML blasts and thus may represent a common therapeutic target for AML regardless of subtype. We demonstrate that the internalization properties of CD37 are distinct in AML blasts when compared to normal blood cells, and that CD37 rapidly accumulates inside AML blasts via dynamin-dependent endocytosis. Our work revealed that the clinically relevant anti-CD37 antibody drug conjugate (ADC) Debio 1562 (αCD37-DM1) is highly cytotoxic to AML blasts, but not normal hematopoietic stem cells. We found that αCD37-DM1 improved clinical outcomes and overall survival in multiple in vivo models of AML. Together, these data demonstrate that targeting CD37 with an ADC such as αCD37-DM1 is a feasible and promising therapeutic option for the treatment of AML.
    DOI:  https://doi.org/10.1182/bloodadvances.2024013590
  10. Cell Stem Cell. 2024 Sep 26. pii: S1934-5909(24)00322-9. [Epub ahead of print]
    Selective translation of nuclear mitochondrial respiratory proteins reprograms succinate metabolism in AML development and chemoresistance.
    Guoqiang Han, Manman Cui, Pengbo Lu, Tiantian Zhang, Rong Yin, Jin Hu, Jihua Chai, Jing Wang, Kexin Gao, Weidong Liu, Shuxin Yao, Ziyan Cao, Yanbing Zheng, Wen Tian, Rongxia Guo, Min Shen, Zheming Liu, Weiming Li, Shanshan Zhao, Xiangpeng Lin, Yuhui Zhang, Kehan Song, Yan Sun, Fuling Zhou, Haojian Zhang.
      Mitochondrial adaptations dynamically reprogram cellular bioenergetics and metabolism and confer key properties for human cancers. However, the selective regulation of these mitochondrial responses remains largely elusive. Here, inspired by a genetic screening in acute myeloid leukemia (AML), we identify RAS effector RREB1 as a translational regulator and uncover a unique translation control system for nuclear-encoded mitochondrial proteins in human cancers. RREB1 deletion reduces mitochondrial activities and succinate metabolism, thereby damaging leukemia stem cell (LSC) function and AML development. Replenishing complex II subunit SDHD rectifies these deficiencies. Notably, inhibition of complex II re-sensitizes AML cells to venetoclax treatment. Mechanistically, a short RREB1 variant binds to a conserved motif in the 3' UTRs and cooperates with elongation factor eEF1A1 to enhance protein translation of nuclear-encoded mitochondrial mRNAs. Overall, our findings reveal a unique translation control mechanism for mitochondrial adaptations in AML pathogenesis and provide a potential strategy for targeting this vulnerability of LSCs.
    Keywords:  RREB1; SDHD; acute myeloid leukemia; leukemia stem cell; mitochondria; succinate; translation; venetoclax
    DOI:  https://doi.org/10.1016/j.stem.2024.09.008
  11. Mol Oncol. 2024 Oct 04.
    Thioredoxin-interacting protein (TXNIP) is a substrate of the NEDD4-like E3 ubiquitin-protein ligase WWP1 in cellular redox state regulation of acute myeloid leukemia cells.
    Sara Giovannini, Yanan Li, Rosalba Pecorari, Claudia Fierro, Claudia Fiorilli, Federica Corigliano, Valeria Moriconi, Ji Zhou, Anna De Antoni, Artem Smirnov, Sara Rinalducci, Anna Maria Timperio, Massimiliano Agostini, Jinping Zhang, Yufang Shi, Eleonora Candi, Gerry Melino, Francesca Bernassola.
      The HECT-type E3 ubiquitin WWP1 (also known as NEDD4-like E3 ubiquitin-protein ligase WWP1) acts as an oncogenic factor in acute myeloid leukemia (AML) cells. WWP1 overexpression in AML confers a proliferative advantage to leukemic blasts (abnormal immature white blood cells) and counteracts apoptotic cell death and differentiation. In an effort to elucidate the molecular basis of WWP1 oncogenic activities, we identified WWP1 as a previously unknown negative regulator of thioredoxin-interacting protein (TXNIP)-mediated reactive oxygen species (ROS) production in AML cells. TXNIP inhibits the disulfide reductase enzymatic activity of thioredoxin (Trx), impairing its antioxidant function and, ultimately, leading to the disruption of cellular redox homeostasis. In addition, TXNIP restricts cell growth and survival by blocking glucose uptake and metabolism. Here, we found that WWP1 directly interacts with TXNIP, thus promoting its ubiquitin-dependent proteasomal proteolysis. As a result, accumulation of TXNIP in response to WWP1 inactivation in AML blasts reduces Trx activity and increases ROS production, hence inducing cellular oxidative stress. Increased ROS generation in WWP1-depleted cells culminates in DNA strand breaks and subsequent apoptosis. Coherently with TXNIP stabilization following WWP1 inactivation, we also observed an impairment of both glucose up-take and consumption. Hence, a contribution to the increased cell death observed in WWP1-depleted cells also possibly arises from the attenuation of glucose up-take and glycolytic flux resulting from TXNIP accumulation. Future studies are needed to establish whether TXNIP-dependent deregulation of redox homeostasis in WWP1-overexpressing blasts may affect the response of leukemic cells to chemotherapeutic drugs.
    Keywords:  HECT E3 ubiquitin ligases; ROS; acute myeloid leukemia; protein ubiquitination; redox homeostasis
    DOI:  https://doi.org/10.1002/1878-0261.13722
  12. Leukemia. 2024 Oct 02.
    Cellular taxonomy of the preleukemic bone marrow niche of acute myeloid leukemia.
    Chinmayee Goda, Rohan Kulkarni, Yaphet Bustos, Wenjun Li, Alexander Rudich, Ozlen Balcioglu, Sadie Chidester, Amog P Urs, Malith Karunasiri, Yzen Al-Marrawi, Erin Korn, Sanjay Kanna, Elizabeth A R Garfinkle, Nisarg Shah, Ashley Wooten, Bethany Mundy-Bosse, Lalit Sehgal, Bin Zhang, Guido Marcucci, Elaine R Mardis, Ramiro Garzon, Robert L Bowman, Aaron D Viny, Linde A Miles, Katherine E Miller, Adrienne M Dorrance.
      Leukemias arise from recurrent clonal mutations in hematopoietic stem/progenitor cells (HSPCs) that cause profound changes in the bone marrow microenvironment (BMM) favoring leukemic stem cell (LSC) growth over normal HSPCs. Understanding the cross talk between preleukemic mutated HSPCs and the BMM is critical to develop novel therapeutic strategies to prevent leukemogenesis. We hypothesize that preleukemic-LSCs (pLSCs) induce BMM changes critical for leukemogenesis. Using our AML-murine model, we performed single-cell RNA-sequencing of preleukemic BMM (pBMM) cells. We found normal HSC (nHSC)-regulating LepR+ mesenchymal stem cells, and endothelial cells were decreased, along with increases in CD55+ fibroblasts and pericytes. Preleukemic CD55+ fibroblasts had higher proliferation rates and decreased collagen expression, suggesting extracellular matrix remodeling during leukemogenesis. Importantly, co-culture assays found preleukemic CD55+ fibroblasts expanded pLSCs significantly over nHSCs. In conclusion, we have identified a distinct pBMM and a novel CD55+ fibroblast population that is expanded in pBMM that promote fitness of pLSCs over nHSCs.
    DOI:  https://doi.org/10.1038/s41375-024-02415-3
  13. Nat Commun. 2024 Sep 30. 15(1): 8439
    CRISPR/Cas9 editing of NKG2A improves the efficacy of primary CD33-directed chimeric antigen receptor natural killer cells.
    Tobias Bexte, Nawid Albinger, Ahmad Al Ajami, Philipp Wendel, Leon Buchinger, Alec Gessner, Jamal Alzubi, Vinzenz Särchen, Meike Vogler, Hadeer Mohamed Rasheed, Beate Anahita Jung, Sebastian Wolf, Raj Bhayadia, Thomas Oellerich, Jan-Henning Klusmann, Olaf Penack, Nina Möker, Toni Cathomen, Michael A Rieger, Katharina Imkeller, Evelyn Ullrich.
      Chimeric antigen receptor (CAR)-modified natural killer (NK) cells show antileukemic activity against acute myeloid leukemia (AML) in vivo. However, NK cell-mediated tumor killing is often impaired by the interaction between human leukocyte antigen (HLA)-E and the inhibitory receptor, NKG2A. Here, we describe a strategy that overcomes CAR-NK cell inhibition mediated by the HLA-E-NKG2A immune checkpoint. We generate CD33-specific, AML-targeted CAR-NK cells (CAR33) combined with CRISPR/Cas9-based gene disruption of the NKG2A-encoding KLRC1 gene. Using single-cell multi-omics analyses, we identified transcriptional features of activation and maturation in CAR33-KLRC1ko-NK cells, which are preserved following exposure to AML cells. Moreover, CAR33-KLRC1ko-NK cells demonstrate potent antileukemic killing activity against AML cell lines and primary blasts in vitro and in vivo. We thus conclude that NKG2A-deficient CAR-NK cells have the potential to bypass immune suppression in AML.
    DOI:  https://doi.org/10.1038/s41467-024-52388-1
  14. Blood Adv. 2024 Sep 30. pii: bloodadvances.2023011754. [Epub ahead of print]
    Peripheral human red blood cell development in human immune system mouse model with heme oxygenase-1 deficiency.
    Aditi Khatri Patel, Kyle Trageser, Hyunjin Kim, Wei Keat Lim, Christina Adler, Brace Porter, Min Ni, Yi Wei, Gurinder S Atwal, Parnian Bigdelou, Vikas Kulshreshtha, Dharani Ajithdoss, Jun Zhong, Naxin Tu, Lynn Macdonald, Andrew Murphy, Davor Frleta.
      A challenge for human immune system (HIS) mouse models has been the lack of human red blood cells (hRBCs) survival after engraftment of these immune-deficient mice with human CD34+ hematopoietic stem cells (HSCs). This limits the use of HIS models for preclinical testing of targets directed at hRBCs-related diseases. Even though human white blood cells can develop in the peripheral blood of these human HSC-engrafted mice, peripheral hRBCs are quickly phagocytosed by murine macrophages upon egress from the bone marrow (BM). Genetic ablation of murine myeloid cells results in severe pathology in resulting mice, rendering such an approach to increase hRBC survival in HIS mice impractical. Heme oxygenase-1 (HMOX-1) deficient mice have reduced macrophages due to toxic build-up of intracellular heme upon engulfment of red blood cells, but do not have an overall loss of myeloid cells. We took advantage of this observation and generated a HMOX-1-/- on a humanized M-CSF/SIRPa/CD47 Rag2-/- IL-2Rg-/- background. These mice have reduced murine macrophages but comparable level of murine myeloid cells to HMOX-1+/+ control mice in the same background. Injected hRBCs survive longer in HMOX-1-/- mice than in HMOX-1+/+ controls. Additionally, upon human HSC-engraftment, hRBCs can be observed in the peripheral blood of HMOX-1-/- humanized M-CSF/SIRPa/CD47 Rag2-/- IL-2Rg-/- mice and hRBC levels can be increased by treatment with human erythropoietin. Since hRBC are present in the peripheral blood of engrafted HMOX-1-/- mice, these mice have the potential to be used for hematological disease modeling, and to test therapeutic treatments for hRBC diseases in vivo.
    DOI:  https://doi.org/10.1182/bloodadvances.2023011754
  15. J Immunol Methods. 2024 Sep 27. pii: S0022-1759(24)00147-9. [Epub ahead of print] 113762
    Highly effective strategy for isolation of mononuclear cells from frozen cord blood.
    Guangzhao Li, Yunyan Zhao, Rongzhi Liu, Yabin Zhang, Yong Zhang, Wei Du, Yu Zhang.
      BACKGROUND AIMS: Cord blood mononuclear cells (CBMCs) comprise a variety of single-nucleated cells found in the cord blood, mainly consisting of monocytes and lymphocytes. They also include a smaller proportion of other cell types, such as hematopoietic stem and progenitor cells (HSPCs) and mesenchymal stromal cells (MSCs). CBMCs are vital for acquiring hematopoietic stem and progenitor cells, MSCs, and other immune cells, like natural killer cells. These cells are essential for supporting subsequent research and clinical applications. Although automated equipment for CBMC enrichment has shown promise, the high cost of these machines and the expense of disposable consumables limit their routine use. Furthermore, limited information is available on manual strategies for isolating CBMCs from cryopreserved cord blood. Therefore, we aimed to optimize the dilution buffer and refine the isolation procedure for CBMCs.METHODS: We enhanced the CBMC recovery rate from cryopreserved cord blood using an optimized dilution buffer and a modified isolation procedure.
    RESULTS: We achieved average recovery rates of 42.4 % and 54.3 % for CBMCs and CD34+ cells, respectively. Notably, all reagents used in the isolation procedure were of GMP-grade or pharmaceutical preparations, underscoring the potential clinical benefits of our strategy.
    DISCUSSION: We devised an optimized protocol suitable for routine research and clinical applications for enhanced recovery of CBMCs from cryopreserved cord blood units using an optimized dilution buffer and a modified isolation procedure.
    Keywords:  Cord blood mononuclear cells (CBMCs); Cryopreserved cord blood; Density gradient centrifugation; Dilution buffer; Enrichment
    DOI:  https://doi.org/10.1016/j.jim.2024.113762
  16. Br J Haematol. 2024 Sep 30.
    TMEM56 deficiency impairs the haem metabolism and cell cycle progression during human erythropoiesis.
    Jingwei Wang, Yingnan Zhang, Jinhua Liu, Jing Wu, Yipeng Liang, Changlu Xu, Jinfa Ma, Jing Liang, Yanhong Zhao, Xiaoru Zhang, Yue Li, Di Wang, Lingyue Zheng, Ding Wang, Xu Jin, Haoze Song, Xu Zhu, Qimei Cheng, Lexuan Lin, Jie Gao, Jingyuan Tong, Lihong Shi.
      TMEM56, a gene coding a transmembrane protein, is abundantly expressed in erythroid cells. Despite this, its role in erythropoiesis has not been well characterized. In this study, we sought to clarify the function of TMEM56 in erythroid development, focusing specifically on its involvement in haem biosynthesis and cell cycle progression. To do this, we used CD34+ haematopoietic stem cells derived from umbilical cord blood and differentiated them into erythroid cells in an ex vivo model. Our results indicate that the loss of TMEM56 disrupts haem biosynthesis and impairs erythroid differentiation. Furthermore, deletion of Tmem56 in the erythroid lineage in murine models using erythropoietin receptor (EpoR)-Cre revealed defects in erythroid progenitors within the bone marrow under both normal conditions and during haemolytic anaemia. These observations underscore the regulatory role of TMEM56 in maintaining erythroid lineage homeostasis. Taken together, our results unveil a previously unrecognized function of TMEM56 in erythroid differentiation and suggest its potential as an unfounded target for therapeutic strategies in the treatment of erythropoietic disorders.
    Keywords:  TMEM56; erythropoiesis; haem metabolism
    DOI:  https://doi.org/10.1111/bjh.19801
  17. Exp Hematol. 2024 Oct 01. pii: S0301-472X(24)00517-4. [Epub ahead of print] 104652
    A New Era of Functional Experimentation in Human Hematopoiesis and Leukemia Research.
    Thomas Köhnke, Yang Feng, Ravindra Majeti.
      Functional experimentation has laid the foundation for our understanding of hematopoietic and leukemic stem cells. Yet, most recently, a flurry of descriptive studies of primary human cells, fueled by rapid technological advances in sequencing technologies, have emerged. These increasing opportunities to describe at great detail have taken precedence over rigorously interrogating functional mediators of biology, particularly in primary human cells. Here, we argue that an improved toolset of gene editing and stem cell biology technologies will allow the field to expand beyond extensive descriptive studies to more functional studies.
    Keywords:  Functional Experimentation; Human Hematopoiesis; Leukemia Research
    DOI:  https://doi.org/10.1016/j.exphem.2024.104652
  18. bioRxiv. 2024 Sep 20. pii: 2024.09.18.612239. [Epub ahead of print]
    PTPN11 Mutation Clonal Hierarchy in Acute Myeloid Leukemia.
    Sydney Fobare, Chia Sharpe, Kate Quinn, Kinsey Bryant, Linde A Miles, Robert L Bowman, Carolyn Cheney, Casie Furby, Marissa Long, Kaytlynn Fyock, Ben Wronowski, James R Lerma, Allison Mullaney, Krzysztof Mrózek, Deedra Nicolet, Tom Sesterhenn, Megan E Johnstone, Shesh N Rai, Chandrashekhar Pasare, Nives Zimmermann, Andrew J Carroll, Richard M Stone, Eunice S Wang, Jonathan E Kolitz, Bayard L Powell, John P Perentesis, Ann-Kathrin Eisfeld, Erin Hertlein, John C Byrd.
      Mutations in protein tyrosine phosphatase non-receptor type 11 ( PTPN11 ) have been considered late acquired mutations in acute myeloid leukemia (AML) development. To interrogate the ontogeny of PTPN11 mutations, we utilized single-cell DNA sequencing and identified that PTPN11 mutations can occur as initiating events in some AML patients when accompanied by strong oncogenic drivers, commonly NPM1 mutations. The co-driver role of PTPN11 mutations was confirmed in a novel murine model that exhibits an AML phenotype with early expansion of a diverse set of variably differentiated myeloid cells that engrafted into immunodeficient and immunocompetent mice. This immune diversity was reconstituted from early precursor cells when engrafted into immunodeficient mice. Moreover, immune diversity was also observed in the blast component of patient samples with NPM1 and PTPN11 mutations, providing novel antigen targets for immune based approaches in this subset of AML that is resistant to multiple targeted therapies.
    DOI:  https://doi.org/10.1101/2024.09.18.612239
  19. Cell Rep. 2024 Oct 02. pii: S2211-1247(24)01154-9. [Epub ahead of print]43(10): 114803
    Temporal alterations of the nascent proteome in response to mitochondrial stress.
    Tomasz M Stępkowski, Vanessa Linke, Dorota Stadnik, Maciej Zakrzewski, Anna E Zawada, Remigiusz A Serwa, Agnieszka Chacinska.
      Under stress, protein synthesis is attenuated to preserve energy and mitigate challenges to protein homeostasis. Here, we describe, with high temporal resolution, the dynamic landscape of changes in the abundance of proteins synthesized upon stress from transient mitochondrial inner membrane depolarization. This nascent proteome was altered when global translation was attenuated by stress and began to normalize as translation was recovering. This transition was associated with a transient desynchronization of cytosolic and mitochondrial translation and recovery of cytosolic and mitochondrial ribosomal proteins. Further, the elongation factor EEF1A1 was downregulated upon mitochondrial stress, and its silencing mimicked the stress-induced nascent proteome remodeling, including alterations in the nascent respiratory chain proteins. Unexpectedly, the stress-induced alterations in the nascent proteome were independent of physiological protein abundance and turnover. In summary, we provide insights into the physiological and pathological consequences of mitochondrial function and dysfunction.
    Keywords:  CP: Cell biology; CP: Metabolism; EEF1A; EEF1A1; cellular stress; elongation factor; mass spectrometry; mitochondria; nascent chain; protein synthesis; proteomics; translation
    DOI:  https://doi.org/10.1016/j.celrep.2024.114803
  20. Nature. 2024 Oct 02.
    RNA m5C oxidation by TET2 regulates chromatin state and leukaemogenesis.
    Zhongyu Zou, Xiaoyang Dou, Ying Li, Zijie Zhang, Juan Wang, Boyang Gao, Yu Xiao, Yiding Wang, Lijie Zhao, Chenxi Sun, Qinzhe Liu, Xianbin Yu, Hao Wang, Juyeong Hong, Qing Dai, Feng-Chun Yang, Mingjiang Xu, Chuan He.
      Mutation of tet methylcytosine dioxygenase 2 (encoded by TET2) drives myeloid malignancy initiation and progression1-3. TET2 deficiency is known to cause a globally opened chromatin state and activation of genes contributing to aberrant haematopoietic stem cell self-renewal4,5. However, the open chromatin observed in TET2-deficient mouse embryonic stem cells, leukaemic cells and haematopoietic stem and progenitor cells5 is inconsistent with the designated role of DNA 5-methylcytosine oxidation of TET2. Here we show that chromatin-associated retrotransposon RNA 5-methylcytosine (m5C) can be recognized by the methyl-CpG-binding-domain protein MBD6, which guides deubiquitination of nearby monoubiquitinated Lys119 of histone H2A (H2AK119ub) to promote an open chromatin state. TET2 oxidizes m5C and antagonizes this MBD6-dependent H2AK119ub deubiquitination. TET2 depletion thereby leads to globally decreased H2AK119ub, more open chromatin and increased transcription in stem cells. TET2-mutant human leukaemia becomes dependent on this gene activation pathway, with MBD6 depletion selectively blocking proliferation of TET2-mutant leukaemic cells and largely reversing the haematopoiesis defects caused by Tet2 loss in mouse models. Together, our findings reveal a chromatin regulation pathway by TET2 through retrotransposon RNA m5C oxidation and identify the downstream MBD6 protein as a feasible target for developing therapies specific against TET2 mutant malignancies.
    DOI:  https://doi.org/10.1038/s41586-024-07969-x
  21. Nature. 2024 Oct 02.
    Tuft cells act as regenerative stem cells in the human intestine.
    Lulu Huang, Jochem H Bernink, Amir Giladi, Daniel Krueger, Gijs J F van Son, Maarten H Geurts, Georg Busslinger, Lin Lin, Harry Begthel, Maurice Zandvliet, Christianne J Buskens, Willem A Bemelman, Carmen López-Iglesias, Peter J Peters, Hans Clevers.
      In mice, intestinal tuft cells have been described as a long-lived, postmitotic cell type. Two distinct subsets have been identified: tuft-1 and tuft-2 (ref. 1). By combining analysis of primary human intestinal resection material and intestinal organoids, we identify four distinct human tuft cell states, two of which overlap with their murine counterparts. We show that tuft cell development depends on the presence of Wnt ligands, and that tuft cell numbers rapidly increase on interleukin-4 (IL-4) and IL-13 exposure, as reported previously in mice2-4. This occurs through proliferation of pre-existing tuft cells, rather than through increased de novo generation from stem cells. Indeed, proliferative tuft cells occur in vivo both in fetal and in adult human intestine. Single mature proliferating tuft cells can form organoids that contain all intestinal epithelial cell types. Unlike stem and progenitor cells, human tuft cells survive irradiation damage and retain the ability to generate all other epithelial cell types. Accordingly, organoids engineered to lack tuft cells fail to recover from radiation-induced damage. Thus, tuft cells represent a damage-induced reserve intestinal stem cell pool in humans.
    DOI:  https://doi.org/10.1038/s41586-024-07952-6
  22. Eur J Med Res. 2024 Sep 30. 29(1): 475
    Network pharmacology-based investigation and experimental validation of the mechanism of metformin in the treatment of acute myeloid leukemia.
    Shaoyu Liu, Mingming Xu, Zhuofan Yang, Yangzi Li, Depei Wu, Xiaowen Tang.
      Metformin, a widely used anti-diabetic agent, has shown significant anti-cancer properties as reported in in various cancers, including acute myeloid leukemia. However, the detailed mechanisms by which metformin influences acute myeloid leukemia remain unrevealed. Employing a synergistic approach of network pharmacology and experimental validation, this study systematically identifies and analyzes potential metformin targets and AML-related genes. These findings are then cross-referenced with biomedical databases to construct a target-gene network, providing insights into metformin's pharmacodynamics in AML treatment. Protein-Protein Interaction (PPI), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses are utilized. Results show metformin's effectiveness in inhibiting AML cell proliferation and inducing apoptosis through the AKT/HIF1A/PDK1 signaling pathway. This research provides insights into metformin's clinical application in AML treatment.
    Keywords:  AML; Apoptosis; HIF1A; Metformin; Network pharmacology
    DOI:  https://doi.org/10.1186/s40001-024-02022-z
  23. Chembiochem. 2024 Oct 04. e202400682
    Precise Modulation of Protein Degradation by Smart PROTACs.
    Junfei Cheng, Guoqiang Dong, Wei Wang, Chunquan Sheng.
      Proteolysis-targeting chimera (PROTAC) has emerged as an attractive therapeutic modality in drug discovery. PROTACs are bifunctional molecules that effectively bridge proteins of interest (POIs) with E3 ubiquitin ligases, such that, the target proteins are tagged with ubiquitin and subsequently degraded via the proteasome. Despite significant progress in the field of targeted protein degradation (TPD), the application of conventional PROTAC degraders still faces significant challenges, including systemic toxicity induced by non-tissue-specific targeting. To address this issue, a variety of smart PROTACs that can be activated by specific stimuli, have been developed for achieving conditional and spatiotemporal modulation of protein levels. Here, on the basis of our contributions, we overview recent advances of smart PROTACs, including tumor microenvironment-, photo-, and X-ray radiation-responsive PROTACs, that enable controllable TPD. The design strategy, case studies, potential applications and challenges will be focused on.
    Keywords:  drug discovery; off-tissue toxicity; precision cancer therapy; smart PROTACs; targeted protein degradation
    DOI:  https://doi.org/10.1002/cbic.202400682
  24. bioRxiv. 2024 Sep 18. pii: 2024.09.17.613543. [Epub ahead of print]
    Cholesterol binding to VCAM-1 promotes vascular inflammation.
    John P Kennelly, Xu Xiao, Yajing Gao, Sumin Kim, Soon-Gook Hong, Miranda Villanueva, Alessandra Ferrari, Lauri Vanharanta, Alexander Nguyen, Rohith T Nagari, Nikolas R Burton, Marcus J Tol, Andrew P Becker, Min Jae Lee, Elina Ikonen, Keriann M Backus, Julia J Mack, Peter Tontonoz.
      Hypercholesterolemia has long been implicated in endothelial cell (EC) dysfunction, but the mechanisms by which excess cholesterol causes vascular pathology are incompletely understood. Here we used a cholesterol-mimetic probe to map cholesterol-protein interactions in primary human ECs and discovered that cholesterol binds to and stabilizes the adhesion molecule VCAM-1. We show that accessible plasma membrane (PM) cholesterol in ECs is acutely responsive to inflammatory stimuli and that the nonvesicular cholesterol transporter Aster-A regulates VCAM-1 stability in activated ECs by controlling the size of this pool. Deletion of Aster-A in ECs increases VCAM-1 protein, promotes immune cell recruitment to vessels, and impairs pulmonary immune homeostasis. Conversely, depleting cholesterol from the endothelium in vivo dampens VCAM-1 induction in response to inflammatory stimuli. These findings identify cholesterol binding to VCAM-1 as a key step during EC activation and provide a biochemical explanation for the ability of excess membrane cholesterol to promote immune cell recruitment to the endothelium.
    DOI:  https://doi.org/10.1101/2024.09.17.613543
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