bims-scepro 2025-03-09 papers

bims-scepro

Biomed News

on Stem cell proteostasis

Issue of 2025–03–09
nineteen papers selected by
William Grey, University of York

Mitochondria-enriched hematopoietic stem cells exhibit elevated self-renewal capabilities, thriving within the context of aged bone marrow.
Restoring mitochondrial function promotes hematopoietic reconstitution from cord blood following cryopreservation-related functional decline.
Leukemia confers a durable imprint on healthy hematopoietic stem and progenitor cells.
Bone marrow sympathetic neuropathy is a hallmark of hematopoietic malignancies and it involves severe ultrastructural damage.
Hematopoietic stem cells undergo bidirectional fate transitions in vivo.
Metabolic alterations in HSCs during aging and leukemogenesis.
DNMT3A regulates murine megakaryocyte-biased hematopoietic stem cell fate decisions.
Feeder-cell-free system for ex vivo production of natural killer cells from cord blood hematopoietic stem and progenitor cells.
BCL11A-deficient human erythropoiesis is impaired in vitro and after xenotransplantation into mice.
Ribosome Profiling Reveals Translational Reprogramming via mTOR Activation in Omacetaxine Resistant Multiple Myeloma.
Peptide-enabled ribonucleoprotein delivery for CRISPR engineering (PERC) in primary human immune cells and hematopoietic stem cells.
High-dose radiation preferentially induces the clonal expansion of hematopoietic progenitor cells over mature T and B cells in mouse bone marrow.
In vitro differentiation of common lymphoid progenitor cells into B cell using stromal cell free culture system.
Structural basis for the midnolin-proteasome pathway and its role in suppressing myeloma.
WTAP-mediated m6A methylation of PHF19 facilitates cell cycle progression by remodeling the accessible chromatin landscape in t(8;21) AML.
Discovery of Novel, Potent, Orally Bioavailable and Efficacious, Hypoxia-Inducible Factor Prolyl Hydroxylase Inhibitors for Hematopoietic Stem Cell Mobilization.
Clofarabine-loaded aptamer-conjugated biodegradable nanoparticle successfully targeted CD117 overexpressed HL60 cells and potentially induced apoptosis.
Leveraging efficiency metrics for the optimization of CELMoDs™ as cereblon-based molecular glue degraders.
Targeting the RBM39-MEK5 axis synergizes with bortezomib to inhibit the malignant growth of multiple myeloma.

Nat Aging. 2025 Mar 06.

Mitochondria-enriched hematopoietic stem cells exhibit elevated self-renewal capabilities, thriving within the context of aged bone marrow.

Haruhito Totani, Takayoshi Matsumura, Rui Yokomori, Terumasa Umemoto, Yuji Takihara, Chong Yang, Lee Hui Chua, Atsushi Watanabe, Takaomi Sanda, Toshio Suda.

The aging of hematopoietic stem cells (HSCs) substantially alters their characteristics. Mitochondria, essential for cellular metabolism, play a crucial role, and their dysfunction is a hallmark of aging-induced changes. The impact of mitochondrial mass on aged HSCs remains incompletely understood. Here we demonstrate that HSCs with high mitochondrial mass during aging are not merely cells that have accumulated damaged mitochondria and become exhausted. In addition, these HSCs retain a high regenerative capacity and remain in the aging bone marrow. Furthermore, we identified GPR183 as a distinct marker characterizing aged HSCs through single-cell analysis. HSCs marked by GPR183 were also enriched in aged HSCs with high mitochondrial mass, possessing a high capacity of self-renewal. These insights deepen understanding of HSC aging and provide additional perspectives on the assessment of aged HSCs, underscoring the importance of mitochondrial dynamics in aging.

DOI: https://doi.org/10.1038/s43587-025-00828-y
J Clin Invest. 2025 Mar 04. pii: e183607. [Epub ahead of print]

Restoring mitochondrial function promotes hematopoietic reconstitution from cord blood following cryopreservation-related functional decline.

Yaojin Huang, Xiaowei Xie, Mengyao Liu, Yawen Zhang, Junye Yang, Wenling Yang, Yu Hu, Saibing Qi, Yahui Feng, Guojun Liu, Shihong Lu, Xuemei Peng, Jinhui Ye, Shihui Ma, Jiali Sun, Lu Wang, Linping Hu, Lin Wang, Xiaofan Zhu, Hui Cheng, Zimin Sun, Junren Chen, Fang Dong, Yingchi Zhang, Tao Cheng.

  Umbilical cord blood (UCB) showcases substantial roles in hematopoietic stem cells (HSCs) transplantation and regenerative medicine. UCB is usually cryopreserved for years before use. Whether and how cryopreservation affects its function remain unclear. We constructed single-cell transcriptomic profile of CD34+ hematopoietic stem and progenitor cells (HSPCs) and mononuclear cells (MNCs) from fresh and cryopreserved UCB stored for 1-, 5-, 10-, and 19- years. Compared to fresh UCB, cryopreserved HSCs and multipotent progenitors (MPPs) exhibited more active cell cycle and lower HSC/MPP signature gene expressions. Hematopoietic reconstitution of cryopreserved HSPCs gradually decreased during the first 5 years but stabilized thereafter, aligning with the negative correlation between clinical neutrophil engraftment and cryopreservation duration of UCB. Cryopreserved HSPCs also showed reduced megakaryocyte generation. In contrast, cryopreserved natural killer (NK) cells and T cells maintained cytokine production and cytotoxic ability comparable to fresh cells. Mechanistically, cryopreserved HSPCs exhibited elevated reactive oxygen species, reduced ATP synthesis, and abnormal mitochondrial distribution, which collectively led to attenuated hematopoietic reconstitution. These effects could be ameliorated by sulforaphane. Together, we elucidated the negative impact of cryopreservation on UCB HSPCs and provided sulforaphane as a mitigation strategy, broadening the temporal window and scope for clinical applications of cryopreserved UCB. .

Keywords:  Hematology; Hematopoietic stem cells; Stem cells

DOI:  https://doi.org/10.1172/JCI183607
Cancer Lett. 2025 Feb 26. pii: S0304-3835(25)00154-5. [Epub ahead of print]616 217590

Leukemia confers a durable imprint on healthy hematopoietic stem and progenitor cells.

Ding-Wen Chen, Julie M Schrey, Eric K Wafula, Jian-Meng Fan, Sarah E Adams, Deanne M Taylor, Peter Kurre.

Recent models of infection and experimental inflammation reveal that hematopoietic stem and progenitor cells (HSPCs) can generate a memory of the exposure. While the acute inflammatory activity rapidly resolves, cells acquire a heightened capacity to respond to subsequent stimulation. Inflammation is also a constitutive feature of cancer, including hematologic malignancies. Here, we adapt a translationally relevant model of acute myeloid leukemia (AML) to determine if inflammation in the bone marrow (BM) niche durably reprograms resident healthy HSPCs. To simulate the onset of malignancy and the subsequent remission, we generated hematopoietic chimera composed of healthy HSPCs and HSPCs bearing an inducible oncogenic human MLL-AF9 translocation expression cassette, a validated model of AML. Results show that the exposure to AML blasts in the BM leaves healthy HSPCs with transcriptomic changes and a shift to glycolytic metabolism during experimental remission. A secondary challenge of AML-experienced animals results in gene expression changes in inflammatory and metabolic pathways. These modified responses coincide with altered chromatin accessibility in AML-experienced HSPCs. Altogether, our observations provide first evidence for the durable inflammatory reprogramming of healthy HSPCs in the cancer microenvironment.

DOI: https://doi.org/10.1016/j.canlet.2025.217590
Exp Hematol Oncol. 2025 Mar 05. 14(1): 31

Bone marrow sympathetic neuropathy is a hallmark of hematopoietic malignancies and it involves severe ultrastructural damage.

Aurora Bernal, Vincent Cuminetti, Marc Serulla, Adrian Florit, Joanna Konieczny, Golnaz Golnarnik, Yimeng Chen, Marc Ferré, Samuel Geiseler, Anders Vik, Randi Olsen, Lorena Arranz.

  The hematopoietic stem cell (HSC) niche in the bone marrow (BM) supports HSC function, fate and numbers [1]. Sympathetic fibres innervate the BM and are components of the hematopoietic stem and progenitor cell (HSPC) niche [2]. Neuropathy of the HSPC niche is present and essential for disease development in experimental models of JAK2V617F+ myeloproliferative neoplasms (MPN) and MLL-AF9+ acute myeloid leukemia (AML), and it is present in the BM of human MPN and AML patients [3-6]. Neuropathy contributes to mutant HSC expansion and represents an effective therapeutic target to block disease progression in JAK2V617F+ MPN mice [3]. The sympathomimetic agonist mirabegron restored nestin+ cells and reduced reticulin fibrosis in MPN patients [7]. Here, we show that neuropathy of the HSPC niche emerges in two additional experimental models of hematological disease including pre-leukemic myelopoiesis driven by NRASG12D and lymphoma/lymphoblastic leukemia driven by p53 deletion. Neuropathy involves severe ultrastructural damage in NRASG12D+ mice and AML patients as shown by electron microscopy. When further reinforced chemically, neuropathy has a profound impact on the experimental NRASG12D mouse model, promoting myeloid bias, reducing HSPC numbers and inducing changes in the stem cell microenvironment that include reduced numbers of mesenchymal stromal cells (MSC) and increased presence of morphologically abnormal blood vessels in BM. Together, BM neuropathy is a prevalent factor in hematopoietic malignancies that involves important degradation of sympathetic fibres and contributes to disease in a different manner depending on the driver mutation. This should be taken in consideration in the clinic, given that chemotherapy induces neuropathy of the HSC niche [8] and it is the most frequent first line treatment for AML, acute lymphoblastic leukemia and MPN patients.

Keywords:  Hematological cancers; Peripheral nervous system; Stem cell niche; Sympathetic fibres; Transmission electron microscopy

DOI:  https://doi.org/10.1186/s40164-025-00614-x
bioRxiv. 2025 Feb 26. pii: 2025.02.23.639689. [Epub ahead of print]

Hematopoietic stem cells undergo bidirectional fate transitions in vivo.

Tsuyoshi Fukushima, Trine Ahn Kristiansen, Lai Ping Wong, Samuel Keyes, Yosuke Tanaka, Michael Mazzola, Ting Zhao, Lingli He, Masaki Yagi, Konrad Hochedlinger, Satoshi Yamazaki, Ruslan I Sadreyev, David T Scadden.

Transitions between subsets of differentiating hematopoietic cells are widely regarded as unidirectional in vivo. Here, we introduce clonal phylogenetic tracer (CP-tracer) that sequentially introduces genetic barcodes, enabling high-resolution analysis of ~100,000 subclones derived from ~500 individual hematopoietic stem cells (HSC). This revealed previously uncharacterized HSC functional subsets and identified bidirectional fate transitions between myeloid-biased and lineage-balanced HSC. Contrary to the prevailing view that the more self-renewing My-HSCs unidirectionally transition to balanced-HSCs, phylogenetic tracing revealed durable lineage bidirectionality with the transition favoring My-HSC accumulation over time1,2. Further, balanced-HSCs mature through distinct intermediates My-HSCs and lymphoid-biased-HSCs with lymphoid competence here shown by CRISPR/Cas9 screening to be dependent on the homeobox gene, Hhex. Hhex enables Ly-HSC differentiation, but its expression declines with age. These findings establish HSC plasticity and Hhex as a determinant of myeloid-lymphoid balance with each changing over time to favor the age-related myeloid bias of the elderly.

DOI: https://doi.org/10.1101/2025.02.23.639689
Physiology (Bethesda). 2025 Feb 28.

Metabolic alterations in HSCs during aging and leukemogenesis.

Yi-Hsuan Chiang, Stephan Emmrich, Nicola Vannini.

  Aging is a multifaceted process associated with a functional decline in cellular function over time, affecting all lifeforms. During the aging process, metabolism, a fundamental hallmark of life (1), is profoundly altered. In the context of hematopoiesis, the proper function of hematopoietic stem cells - at the apex of the blood system - is tightly linked to their energy metabolism, which in turn shapes hematopoietic output. Here, we review the latest developments in our understanding of the metabolic states and changes in aged hematopoietic stem cells, molecular players and pathways involved in aged hematopoietic stem cell metabolism, the consequences of perturbed metabolism on clonal hematopoiesis and leukemogenesis, and pharmacologic/ genetic strategies to reverse or rejuvenate altered metabolic phenotypes.

Keywords:  aging; clonal hematopoiesis; hematopoietic stem cell; leukemogenesis; metabolism

DOI:  https://doi.org/10.1152/physiol.00054.2024
Blood Adv. 2025 Mar 06. pii: bloodadvances.2024015061. [Epub ahead of print]

DNMT3A regulates murine megakaryocyte-biased hematopoietic stem cell fate decisions.

Sarah M Waldvogel, Virginia Camacho, Dandan Fan, Anna Guzman, Alejandra Garcia-Martell, Elmira Khabusheva, Jacey Rodriguez Pridgen, Josephine De La Fuente, Rachel E Rau, Ashlyn Laidman, Maria N Barrachina, Estelle Carminita, Justin Andrew Courson, Michael Williamson, Joanne Ino Hsu, Chun-Wei Chen, Jaime Reyes, Subhashree Pradhan, Rolando Rumbaut, Alan Burns, Benjamin Deneen, Jianzhong Su, Kellie R Machlus, Margaret A Goodell.

Hematopoietic stem cells (HSCs) are defined by their capacity to regenerate all main components of the peripheral blood, but individual HSCs exhibit a range of preferences for generating downstream cell types. Their propensities are thought to be epigenetically encoded, but few differential regulatory mechanisms have been identified. In this work, we explored the role of the DNA methyltransferase 3A (DNMT3A) in the megakaryocyte-biased HSC population, which is thought to reside at the top of the hematopoietic hierarchy. We demonstrate that heterozygous loss of DNMT3A (Dnmt3a+/-) in these megakaryocyte-biased HSCs has consequences distinct from the rest of the HSC pool. These megakaryocyte-biased HSCs become delayed in their lymphoid-repopulating ability but can ultimately regenerate all lineages. We further demonstrate that Dnmt3a+/- mice have increased numbers of megakaryocytes in the bone marrow. Analysis of DNA methylation differences between WT and Dnmt3a+/- HSC subsets, megakaryocyte-erythroid progenitors (MEP), and megakaryocytes revealed that DNA methylation is eroded in the mutants in a cell type-specific fashion. While transcriptional differences between the WT and Dnmt3a+/- megakaryocyte-biased HSCs are subtle, the pattern of DNA methylation loss in this HSC subset is almost completely different from that in non-megakaryocyte-biased HSCs. Together, our findings establish the role of epigenetic regulation in the fate of megakaryocyte-biased HSCs and their downstream progeny and suggest that the outcomes of DNMT3A loss might vary depending on the identity of the HSC that acquires the mutation.

DOI: https://doi.org/10.1182/bloodadvances.2024015061
Front Immunol. 2025 ;16 1531736

Feeder-cell-free system for ex vivo production of natural killer cells from cord blood hematopoietic stem and progenitor cells.

Marta Martin Corredera, Juliette Paillet, Pierre Gaudeaux, Tifanie Blein, Hanem Sadek, Pauline Rault, Asma Berriche, Jeanne Roche-Naude, Chantal Lagresle-Peyrou, Tayebeh-Shabi Soheili, Isabelle André, Ranjita Devi Moirangthem, Olivier Negre.

   Introduction: Natural Killer (NK) cells hold significant promise as therapeutic agents in immuno-oncology due to their ability to target and eliminate cancerous and infected cells without causing graft-versus-host disease or cytokine release syndrome. However, the limited availability of robust, scalable methods for generating clinical-grade NK cells remains a limiting factor to broader clinical application.
Methods: Here we report the development of a novel feeder-cell-free culture system optimized for producing NK cells from cord blood-derived CD34+ hematopoietic stem and progenitor cells (HSPCs). Our method eliminates the need for feeder cells while achieving high yields of NK cells that exhibit unique marker expression and cytotoxic functions. Cord blood CD34+ HSPCs were cultured in our established hDLL 4 culture system and generated large numbers of human T lymphoid progenitors (ProTcells) in 7 days. ProTcells were further cultured in a hDLL4-free, feeder-cell-free system for NK cell differentiation and supplemented with cytokines. Following a 7- or 14-day culture, this method produced highly pure NK cell populations (>90% CD3-CD56+).
Results: Flow and mass cytometric analysis confirmed the expression of activating receptors, transcription factors (ID2, T-bet) and cytotoxic molecules (perforin, granzyme A/B), all essential for ProT-NK cell functionality. These cells are in an immature state, indicated by the absence of maturation markers (CD16, KIRs). Functional assays demonstrated that these ProT-NK cells are capable of degranulation and cytokines production (TNFα) upon stimulation with K562 target cells and showed cytotoxicity against K562 cells superior to that of Peripheral Blood (PB)-NK. In NSG-Tg(hIL-15) mice, ProT-NK cells colonize bone marrow, the liver, and the spleen and persist and mature in bone marrow for at least 9 days post-injection. Compared to ProT-NK D21, ProT-NK D14 was superior in functional and homing potential. In vivo, an anti-tumor assay that uses a subcutaneous K562 model has demonstrated the anti-tumor potential of ProT-NK cells.
Discussion: Our ex vivo culture process supports scalable ProT-NK cell production in high yields, reducing dependency on feeder cells and mitigating contamination risks. Our findings demonstrate the feasibility of generating large, functional NK cell populations from HSPCs isolated from readily available cord blood sources and offer an efficient alternative to PB-NK cell therapies.

Keywords:  cord blood (CB); cytotoxicity; feeder-cell-free; hematopoietic stem and progenitor cells (HSPCs); immunotherapy; natural killer (NK)

DOI:  https://doi.org/10.3389/fimmu.2025.1531736
Blood Adv. 2025 Feb 28. pii: bloodadvances.2024015574. [Epub ahead of print]

BCL11A-deficient human erythropoiesis is impaired in vitro and after xenotransplantation into mice.

Yoonjeong Jang, Ruopeng Feng, Lance E Palmer, Thiyagaraj Mayuranathan, Yu Yao, Kalin Mayberry, Sheng Zhou, Jian Xu, Jeffrey Michael Gossett, Guolian Kang, Yong Cheng, Jonathan S Yen, Mitchell J Weiss.

Genetic depletion of the transcriptional repressor BCL11A in red blood cell precursors alleviates b-hemoglobinopathies by inducing the fetal g-globin genes. However, additional erythroid genes are regulated by BCL11A and the effects of its deficiency on erythropoiesis are insufficiently described. We discovered that Cas9 disruption of the BCL11A intron 2 erythroid enhancer in CD34+ hematopoietic stem and progenitor cells using a clinically approved strategy caused impaired expansion and apoptosis of erythroid precursors in vitro and reduced repopulation of the erythroid compartment after xenotransplantation into immunodeficient mice. Mutant colony-forming unit erythroid cells, proerythroblasts and basophilic erythroblasts exhibited dysregulation of 94 genes (> 2-fold change, FDR < 0.05), 25 of which are likely direct targets of BCL11A. Differentially expressed genes were associated with a range of biological pathways that impact cell expansion and survival. Our findings show that BCL11A regulates additional aspects of erythropoiesis beyond g-globin gene repression, with unknown clinical consequences.

DOI: https://doi.org/10.1182/bloodadvances.2024015574
Mol Cancer Res. 2025 Mar 06.

Ribosome Profiling Reveals Translational Reprogramming via mTOR Activation in Omacetaxine Resistant Multiple Myeloma.

Zachary J Walker, Katherine F Vaeth, Amber Baldwin, Denis J Ohlstrom, Lauren T Reiman, Kady A Dennis, Kate Matlin, Beau M Idler, Brett M Stevens, Neelanjan Mukherjee, Daniel W Sherbenou.

Protein homeostasis is critical to the survival of multiple myeloma (MM) cells. While this is targeted with proteasome inhibitors, mRNA translation inhibition has not entered trials. Recent work illustrates broad sensitivity MM cells to the translation inhibitor omacetaxine. We hypothesized that understanding how MM becomes resistant will lead to the development of drug combinations to prevent or delay relapse. We generated omacetaxine resistance in H929 and MM1S MM cell lines and compared them to parental lines. Resistant lines displayed decreased sensitivity to omacetaxine, with EC50 > 100 nM, compared to parental sensitivity of 24-54 nM. Since omacetaxine inhibits protein synthesis, we performed both RNA-sequencing and ribosome profiling (Ribo-seq) to identify shared and unique regulatory strategies of resistance. Transcripts encoding translation factors and containing Terminal OligoPyrimidine (TOP) sequence in their 5' UTR were translationally upregulated in both resistant cell lines. The mTOR pathway promotes the translation of TOP motif containing mRNAs. Indeed, mTOR inhibition with Torin 1 restored partial sensitivity to omacetaxine in both resistant cell lines. The combination was synergistic in omacetaxine naïve MM cell lines, and a combination effect was observed in vivo. Primary MM cells from patient samples were also sensitive to the combination. These results provide a rational approach for omacetaxine-based combination in patients with multiple myeloma, which have historically shown better responses to multi-agent regimens. Implications: Through the use of ribosome profiling, our findings indicate mTOR inhibition as a novel combination therapy for partnering with the translation inhibitor omacetaxine in the treatment of multiple myeloma.

DOI: https://doi.org/10.1158/1541-7786.MCR-24-0444
Nat Protoc. 2025 Mar 03.

Peptide-enabled ribonucleoprotein delivery for CRISPR engineering (PERC) in primary human immune cells and hematopoietic stem cells.

Srishti U Sahu, Madalena Castro, Joseph J Muldoon, Kunica Asija, Stacia K Wyman, Netravathi Krishnappa, Lorena de Oñate, Justin Eyquem, David N Nguyen, Ross C Wilson.

Peptide-enabled ribonucleoprotein delivery for CRISPR engineering (PERC) is a new approach for ex vivo genome editing of primary human cells. PERC uses a single amphiphilic peptide reagent to mediate intracellular delivery of the same pre-formed CRISPR ribonucleoprotein enzymes that are broadly used in research and therapeutics, resulting in high-efficiency editing of stimulated immune cells and cultured hematopoietic stem and progenitor cells (HSPCs). PERC facilitates nuclease-mediated gene knockout, precise transgene knock-in and base editing. The protocol involves mixing the CRISPR ribonucleoprotein enzyme with peptide and then incubating with cultured cells. For efficient transgene knock-in, adeno-associated virus (AAV) homology-directed repair template (HDRT) DNA may be included. In contrast to electroporation, PERC is appealing because it needs no dedicated hardware and has less impact on cell phenotype and viability. Because of the gentle nature of PERC, delivery can be performed multiple times without substantial impact to cell health or phenotype. Editing efficiencies can surpass 90% when using either Cas9 or Cas12a in primary T cells or HSPCs. After 3 h dedicated to reagent preparation, the PERC delivery step can be completed in 1 h, with the associated cell culture steps taking 3-7 d total. Because the protocol calls for only three readily available reagents (protein, RNA and peptide) and does not require dedicated hardware for any step, PERC demands no special expertise and is exceptionally straightforward to adopt. The inherent compatibility of PERC with established cell engineering pipelines makes the protocol appealing for rapid deployment in research and clinical settings.

DOI: https://doi.org/10.1038/s41596-025-01154-8
Stem Cell Reports. 2025 Feb 20. pii: S2213-6711(25)00027-X. [Epub ahead of print] 102423

High-dose radiation preferentially induces the clonal expansion of hematopoietic progenitor cells over mature T and B cells in mouse bone marrow.

Kengo Yoshida, Munechika Misumi, Kanya Hamasaki, Seishi Kyoizumi, Yasunari Satoh, Tatsuaki Tsuruyama, Arikuni Uchimura, Yoichiro Kusunoki.

  Radiation induces clonal hematopoiesis (CH) involving high-frequency somatic mutations in hematopoietic cells. However, the effects of radiation on clonal expansion of hematopoietic progenitor cells and lymphocytes remain elusive. Here, we investigate CH mutations and T cell receptor (TCR) and B cell receptor (BCR) sequences within the bone marrow cells of mice 18 months after irradiation (3 Gy) and age-matched controls. Two to six CH mutations were identified in the irradiated mice (N = 5), while only one of the four control mice carried a CH mutation. These CH mutations detected in the bone marrow were also identified in the splenic CD11b+ myeloid cell population. Meanwhile, the cumulative size of the ten largest TCR and BCR clones, as well as their clonality, did not differ significantly between irradiated and control mice. Our findings suggest that radiation preferentially induces clonal expansion of hematopoietic progenitor cells over mature lymphocytes in the bone marrow.

Keywords:  BCR; Mouse; TCR; clonal hematopoiesis; ionizing radiation; somatic mutation

DOI:  https://doi.org/10.1016/j.stemcr.2025.102423
BMC Immunol. 2025 Mar 04. 26(1): 12

In vitro differentiation of common lymphoid progenitor cells into B cell using stromal cell free culture system.

Pan Yang, Xiaoling Chen, Hao Wen, Meiling Yu, Haili Yu, Li Wang, Liang Gong, Lintao Zhao.

  The OP9 culture system is an important in vitro model for B cell development. However, the complex nature of operations and the intrinsic variability of stromal cell functionality, which can be influenced by factors such as radiation exposure or contamination, pose considerable challenges to their wider application. Currently, there exists a paucity of studies documenting in vitro B cell differentiation culture systems that exclude stromal cells, and the experimental methodologies available for reference remain limited. This report elucidates a robust stromal cell-free culture system. Specifically, bovine serum albumin (BSA) or fetal bovine serum (FBS), in conjunction with interleukin-7 (IL-7), Flt3 ligand (Flt3L), and stem cell factor (SCF), were incorporated into X-VIVO15 medium. This system proficiently facilitates the directed differentiation of common lymphoid progenitor cells (CLP), defined as lineage-CD127 + CD117lowsca-1lowCD135+, into B lymphocytes in vitro, achieving an amplification factor of up to one hundredfold.We examined the roles of IL-7, Flt3L and SCF in differentiation of B cells from CLP in this culture system. Our findings indicate that IL-7 is a pivotal cytokine essential for B cell differentiation in vitro, demonstrating a notable synergistic impact when combined with SCF and FLT3L. Moreover, this system is capable of supporting the differentiation of hematopoietic stem cells (HSCs) and lymphoid-primed multipotent progenitor cells (LMPPs) into B cells in vitro. The findings substantiated the efficacy of the culture system in investigating the in vitro differentiation of bone marrow-derived progenitor cells into B cells and elucidated the specific roles of BSA, FBS and three cytokines (IL-7, FLT3L and SCF) in promoting efficient B lineage differentiation.

Keywords:  B cell; Common lymphoid progenitor cells; Vitro differentiation; stromal cell free culture system

DOI:  https://doi.org/10.1186/s12865-025-00694-8
bioRxiv. 2025 Feb 23. pii: 2025.02.22.639686. [Epub ahead of print]

Structural basis for the midnolin-proteasome pathway and its role in suppressing myeloma.

Christopher Nardone, Jingjing Gao, Hyuk-Soo Seo, Julian Mintseris, Lucy Ort, Matthew C J Yip, Milen Negasi, Anna K Besschetnova, Nolan Kamitaki, Steven P Gygi, Sirano Dhe-Paganon, Nikhil Munshi, Mariateresa Fulciniti, Michael E Greenberg, Sichen Shao, Stephen J Elledge, Xin Gu.

The midnolin-proteasome pathway degrades many nuclear proteins without ubiquitination, but how it operates mechanistically remains unclear. Here, we present structures of the midnolin-proteasome complex, revealing how established proteasomal components are repurposed to enable a unique form of proteolysis. While the proteasomal subunit PSMD2/Rpn1 binds to ubiquitinated or ubiquitin-like proteins, we discover that it also interacts with the midnolin nuclear localization sequence, elucidating how midnolin's activity is confined to the nucleus. Likewise, PSMD14/Rpn11, an enzyme that normally cleaves ubiquitin chains, surprisingly functions non-enzymatically as a receptor for the midnolin ubiquitin-like (Ubl) domain, positioning the substrate-binding Catch domain directly above the proteasomal entry site to guide substrates into the proteasome. Moreover, we demonstrate that midnolin downregulation is critical for the survival of myeloma cells by promoting the expression of its transcription factor substrate IRF4. Our findings uncover the mechanisms underlying the midnolin-proteasome pathway and midnolin downregulation as a driver of multiple myeloma.

DOI: https://doi.org/10.1101/2025.02.22.639686
Oncogene. 2025 Mar 04.

WTAP-mediated m6A methylation of PHF19 facilitates cell cycle progression by remodeling the accessible chromatin landscape in t(8;21) AML.

Yu-Qing Li, Di Liu, Li-Li Wang, Yang-Liu Shao, Hui-Sheng Zhou, Ya-Lei Hu, Kai-Li Min, Chun-Ji Gao, Dai-Hong Liu, Jie Zhou, Ji Lin, Xiao-Ning Gao.

Wilms' tumor 1-associated protein (WTAP) is a key N6-methyladenosine (m6A) methyltransferase that is upregulated in t(8;21) acute myeloid leukemia (AML) under hypoxia inducible factor 1α-mediated transcriptional activation, promoting leukemogenesis through transcriptome-wide m6A modifications. However, the specific substrates and intrinsic regulatory mechanisms of WTAP are not well understood. Here, we provide evidence that PHD finger protein 19 (PHF19) overexpression is regulated by WTAP-mediated m6A modification and promotes cell cycle progression by altering chromatin accessibility. At the same time, high expression of PHF19 and WTAP in t(8;21) AML patients indicates a worse prognosis. Furthermore, inhibition of PHF19 expression significantly suppresses the growth of t(8;21) AML cells in both in vitro and in vivo. Mechanistically, WTAP enhances the stability of PHF19 mRNA by binding to m6A sites in the 3'-untranslated region, thereby upregulating PHF19 expression. Conversely, WTAP suppression reduces m6A modification levels on the PHF19 transcript, leading to increased instability. Knockdown of PHF19 precipitates loss of H3K27 trimethylation and enhanced chromatin accessibility, ultimately resulting in upregulated expression of genes involved in the cell cycle and DNA damage checkpoints. Therefore, WTAP/m6A-dependent PHF19 upregulation accelerates leukemia progression by coordinating m6A modification and histone methylation, establishing its status as a novel therapeutic target for t(8;21) AML.

DOI: https://doi.org/10.1038/s41388-025-03329-9
J Med Chem. 2025 Mar 06.

Discovery of Novel, Potent, Orally Bioavailable and Efficacious, Hypoxia-Inducible Factor Prolyl Hydroxylase Inhibitors for Hematopoietic Stem Cell Mobilization.

Min Wu, Dennis C Koester, Gail Walkinshaw, Danny Ng, Xiaoti Zhou, Angel Ho, Jenny Tsao, Michael Barnes, Mitchell C Brenner, Suzanne Spong, Grace Nelson, David C Gervasi, Elena Vaisberg, Mark Sternlicht, Parmjeet Sidhu, Jack Lin, Mohamed Ibrahim, Michael D Thompson, James Chou, Gerardo Pangilinan, Om Makwana, Zhihua Wei, Pierre E Signore, Ughetta Del Balzo, Ute Hoch, Savithri Ramurthy.

Hematopoietic stem cell (HSC) mobilization is often difficult to achieve in patients suffering from multiple myeloma and non-Hodgkin's lymphoma. Granulocyte-colony stimulating factor (G-CSF) therapy alone has often not led to the desired outcomes. Herein, we describe the discovery of 7-cyclohexyl-4-hydroxy-8-oxo-N-(pyridazin-4-ylmethyl)-7,8-dihydro-2,7-naphthyridine-3-carboxamide 13, a hypoxia-inducible factor prolyl hydroxylase (HIF-PH) inhibitor, which was discovered by focusing on drug-like properties. Building on a previous discovery that HIF-PH inhibitors can enhance HSC mobilization in combination with G-CSF, we optimized 13 to exhibit high PHD2 potency, improved solubility, and an optimized PK profile. 13 was effective at enhancing G-CSF-induced HSC mobilization in mice at a dose of 2 mg/kg.

DOI: https://doi.org/10.1021/acs.jmedchem.4c02889
Heliyon. 2025 Feb 28. 11(4): e42450

Clofarabine-loaded aptamer-conjugated biodegradable nanoparticle successfully targeted CD117 overexpressed HL60 cells and potentially induced apoptosis.

Manisheeta Ray, Ashique Al Hoque, Saptarshi Chatterjee, Sourav Adhikary, Samrat Paul, Biswajit Mukherjee, Amitava Bhattacharya.

  Acute Myeloid Leukemia (AML) is a rapidly progressing malignancy characterized by the proliferation of abnormal neutrophils, leading to severe symptoms and complications. Current widely used treatment options include chemotherapy and radiotherapy, which often result in suffering from systemic toxicity and drug resistance. To mitigate systemic toxicity and off-target side effects, a targeted therapeutic strategy is one of the remarkably successful options. For targeting AML cells, we have chosen a single-strand DNA aptamer (Apt), which is specific for the biomarker CD117, overexpressing AML cells. This study introduces explicitly a novel therapeutic approach employing aptamer-conjugated clofarabine-loaded PLGA nanoparticles (Apt-CNP) targeting the CD117 receptor on HL60 leukemia cells. Clofarabine, a potent nucleoside analogue, disrupts DNA synthesis and induces cancer cell death but is limited by its toxicity and resistance. Encapsulation in PLGA nanoparticles enables sustained drug release, maintaining therapeutic concentrations and potentially reducing drug resistance. Our findings demonstrate that Apt-CNP effectively targets HL60 leukemia cells, thereby improving drug delivery and reducing adverse effects on healthy cells. This targeted approach may open a new avenue for more specific drug delivery to mobile and floated blood cells, including AML (HL60 leukemia) cells, and overcome the limitations of traditional AML treatments.

Keywords:  Acute myeloid leukemia (AML); Biomarker CD117; DNA aptamer; Nanoparticles; Targeted delivery

DOI:  https://doi.org/10.1016/j.heliyon.2025.e42450
RSC Med Chem. 2025 Mar 03.

Leveraging efficiency metrics for the optimization of CELMoDs™ as cereblon-based molecular glue degraders.

Lei Jia, Jennifer R Riggs, Dahlia R Weiss, Brian L Claus, Veerabahu Shanmugasundaram, Stephen R Johnson, Christoph W Zapf.

Efficiency metrics are useful in medicinal chemistry to track small molecule progress in lead optimization (LO). Molecular glue degraders are small molecules that mediate targeted protein degradation by chemically inducing proximity between an E3 ligase and a protein target. The potency and depth of protein degradation are important factors in identifying molecular glue drug candidates. We developed degradation efficiency metrics based on both potency and depth of degradation to track lead optimization objectives. We applied these efficiency metrics retrospectively to track optimization of a clinical molecular glue degrader series, resulting in the identification of Golcadomide (CC-99282). This work illustrates that efficiency metrics are beneficial for the identification of molecular glue drug candidates.

DOI: https://doi.org/10.1039/d4md00870g
Blood Adv. 2025 Mar 06. pii: bloodadvances.2025015815. [Epub ahead of print]

Targeting the RBM39-MEK5 axis synergizes with bortezomib to inhibit the malignant growth of multiple myeloma.

Jia Liu, Zilu Zhang, Wenbin Xu, Mingyuan Jia, Xinyi Zeng, Chengyu Wu, Ze Fu, Xiaoguang Xu, Chenjing Ye, Chao Wu, Hanzhang Xu, Hu Lei, Ying-Li Wu, Yan Hua.

Aberrant alternative splicing is one of the hallmarks of cancer and is potentially based on upregulated expression of splicing factors in some types of cancer. Our previous study suggested that the splicing factor RBM39 is significantly upregulated in multiple myeloma (MM) and that its upregulation is positively associated with poor prognosis. Here, we further demonstrate that the survival and proliferation of MM cells rely on RBM39 and that RBM39 knockdown inhibits the malignant growth of MM. Indisulam, a "molecular glue" that mediates the proteasomal degradation of RBM39, has potent suppressive effects on MM both in vitro and in vivo. Deletion of RBM39 results in extensively altered splicing, with mis-splicing of MEK5 verified to inhibit the malignant growth of MM. Full-length MEK5 plays a vital role in maintaining MM cell survival, whereas aberrant MEK5 isoforms with exon loss exhibit loss of function and a propensity for proteasomal degradation. Targeting RBM39 or MEK5 synergistically increases the cytotoxicity of bortezomib in MM cells via the inhibition of p65. Our study validates the specific mechanism of RBM39 in MM, providing an approach for broader targeting and optimized therapeutic strategies for MM.

DOI: https://doi.org/10.1182/bloodadvances.2025015815