T-cell acute lymphoblastic leukemia exploits a neural proinflammatory pathway to colonize the meninges

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Abstract

Infiltration of T-cell acute lymphoblastic leukemia (T-ALL) into the meninges worsens prognosis, underscoring the need to understand mechanisms driving meningeal involvement. Here, we show that T-ALL cells expressing CXCR3 exploit normal T-cell function to infiltrate the inflamed meninges. CXCR3 deletion hampered disease progression and extramedullary dissemination by reducing leukemic cell proliferation and migration. Conversely, forced expression of CXCR3 facilitated T-ALL trafficking to the meninges. We identified the ubiquitin-specific protease 7 as a key regulator of CXCR3 protein stability in T-ALL. Furthermore, we discovered elevated levels of CXCL10, a CXCR3 ligand, in the cerebrospinal fluid from T-ALL patients and leukemia-bearing mice. Our studies demonstrate that meningeal stromal cells, specifically pericytes and fibroblasts, induce CXCL10 expression in response to leukemia, and that loss of CXCL10 attenuated T-ALL influx into the meninges. Moreover, we report that leukemia-derived proinflammatory cytokines, TNFα, IL27 and IFNγ, induced CXCL10 in the meningeal stroma. Pharmacological inhibition or deletion of CXCR3 or CXCL10 reduced T-ALL cell migration and adhesion to meningeal stromal cells. Finally, we reveal that CXCR3 and CXCL10 upregulated VLA-4/VCAM-1 signaling, promoting cell-cell adhesion and thus T-ALL retention in the meninges. Our findings highlight the pivotal role of CXCR3-CXCL10 signaling in T-ALL progression and meningeal colonization.

Authors

Nitesh D. Sharma, Esra'a Keewan, Wojciech Ornatowski, Silpita Paul, Monique Nysus, Christopher C. Barnett, Julie Wolfson, Quiteria Jacquez, Bianca L. Myers, Huining Kang, Katherine E. Zychowski, Stuart S. Winter, Mignon L. Loh, Stephen P. Hunger, Eliseo F. Castillo, Tom Taghon, Christina Halsey, Tou Yia Vue, Nicholas Jones, Panagiotis Ntziachristos, Ksenia Matlawska-Wasowska

The hematopoietic stem cell MYB enhancer is essential and recurrently amplified during T-cell leukemogenesis

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Abstract

There is an urgent need to find targeted agents for T-cell acute lymphoblastic leukemia (T-ALL). NOTCH1 is the most frequently mutated oncogene in T-ALL, but clinical trials showed that pan-Notch inhibitors caused dose-limiting toxicities. Thus, we shifted our focus to ETS1, which is one of the transcription factors that most frequently co-bind Notch-occupied regulatory elements in the T-ALL context. To identify the most essential enhancers, we performed a genome-wide CRISPR interference screen of the strongest ETS1-dependent regulatory elements. The #1-ranked element is located in an intron of AHI1 that interacts with the MYB promoter and is amplified with MYB in ~8.5% of T-ALL patients. Using mouse models, we showed that this enhancer promotes self-renewal of hematopoietic stem cells and T-cell leukemogenesis, maintains early T-cell precursors, and restrains myeloid expansion with aging. We named this enhancer the hematopoietic stem cell MYB enhancer (H-Me). The H-Me shows limited activity and function in committed T-cell progenitors but is accessed during leukemogenesis. In one T-ALL context, ETS1 binds the ETS motif in the H-Me to recruit cBAF to promote chromatin accessibility and activation. ETS1 or cBAF degraders impaired H-Me function. Thus, we identified a targetable stem cell element that is co-opted for T-cell transformation.

Authors

Carea Mullin, Karena Lin, Elizabeth Choe, Cher Sha, Zeel Shukla, Koral Campbell, Anna C. McCarter, Annie Wang, Jannaldo Nieves-Salva, Sarah Khan, Theresa M. Keeley, Shannon Liang, Qing Wang, Ashley F. Melnick, Pearl Evans, Alexander C. Monovich, Ashwin Iyer, Rohan Kodgule, Yamei Deng, Felipe da Veiga Leprevost, Kelly R. Barnett, Petri Pölönen, Rami Khoriaty, Daniel Savic, David T. Teachey, Charles G. Mullighan, Marcin Cieslik, Alexey I. Nesvizhskii, Linda C. Samuelson, Morgan Jones, Qing Li, Russell J.H. Ryan, Mark Y. Chiang

Spinal α2δ-1 induces GluA3 degradation to regulate assembly of calcium-permeable AMPA receptors and pain hypersensitivity

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Abstract

The increased prevalence of GluA2-lacking, Ca2+-permeable AMPA receptors (CP-AMPARs) at spinal cord sensory synapses amplifies nociceptive transmission and maintains chronic neuropathic pain. Nerve injury–induced upregulation of α2δ-1 disrupts the assembly of GluA1/GluA2 heteromers, favoring the synaptic incorporation of GluA1 homotetramers in the spinal dorsal horn. Although GluA1-GluA3 subunits are broadly expressed, whether α2δ-1 regulates GluA3-containing AMPARs remains unknown. Here, we unexpectedly found that coexpression with α2δ-1—but not α2δ-2 or α2δ-3—diminished GluA3 AMPAR currents and protein levels, an effect blocked by pregabalin, an α2δ-1 C-terminus peptide, or proteasome inhibition. Both nerve injury and α2δ-1 overexpression reduced protein levels of GluA3 and GluA2/GluA3 heteromers in the spinal cord. Furthermore, α2δ-1 coexpression or nerve injury increased GluA3 ubiquitination, with Lys-861 at the C terminus of GluA3 identified as a key ubiquitination site mediating α2δ-1–induced GluA3 degradation. Additionally, intrathecal delivery of the Gria3 gene reversed nerve injury–induced nociceptive hypersensitivity and synaptic CP-AMPARs by restoring protein levels of GluA3 and GluA2/GluA3 heteromers in the spinal cord. These findings reveal that α2δ-1 promotes GluA1 homotetramer assembly and synaptic CP-AMPAR expression by driving ubiquitin-proteasomal degradation of GluA3, providing insights into the molecular mechanisms of neuropathic pain and the therapeutic actions of gabapentinoids.

Authors

Meng-Hua Zhou, Shao-Rui Chen, Daozhong Jin, Yuying Huang, Hong Chen, Guanxing Chen, Jiusheng Yan, Hui-Lin Pan

Targeting STING–induced immune evasion with nanoparticulate binary pharmacology improves tumor control in mice

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Abstract

Harnessing the stimulator of interferon genes (STING) signaling pathway to trigger innate immune responses has shown remarkable promise in cancer immunotherapy; however, overwhelming resistance to intratumoral STING monotherapy has been witnessed in clinical trials, and the underlying mechanisms remain to be fully explored. Herein, we show that pharmacological STING activation following the intratumoral injection of a non-nucleotide STING agonist (i.e., MSA-2) results in apoptosis of the cytolytic T cells, interferon-mediated overexpression of indoleamine 2,3-dioxygenase 1 (IDO1), and evasion from immune surveillance. We leverage a noncovalent chemical strategy for developing immunomodulatory binary nanoparticles (iBINP) that include both the STING agonist and an IDO1 inhibitor for treating immune-evasive tumors. This iBINP platform developed by dual prodrug engineering and subsequent nanoparticle assembly enables tumor-restricted STING activation and IDO1 inhibition, achieving immune activation while mitigating immune tolerance. A systemic treatment of preclinical models of colorectal cancer with iBINP resulted in robust antitumor immune responses, reduced infiltration of regulatory T cells, and enhanced activity of CD8+ T cells. Importantly, this platform exhibits great therapeutic efficacy by overcoming STING–induced immune evasion and controlling the progression of multiple tumor models. This study unveils the mechanisms by which STING monotherapy induces immunosuppression in the tumor microenvironment and provides a combinatorial strategy for advancing cancer immunotherapies.

Authors

Fanchao Meng, Hengyan Zhu, Shuo Wu, Bohan Li, Xiaona Chen, Hangxiang Wang

Selective targeting of HIV infected clones by cognate peptide stimulation and antiproliferative drugs

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Abstract

Clonal expansion of HIV infected CD4+ T cells is a barrier to HIV eradication. We previously described a marked reduction in the frequency of the most clonally expanded infected CD4+ T cells in an individual with elite control (ES24) after initiating chemoradiation for metastatic lung cancer with a regimen that included paclitaxel and carboplatin. We tested the hypothesis that this phenomenon was due to a higher susceptibility to the chemotherapeutic drugs of CD4+ T cell clones that were sustained by proliferation. We studied a CD4+ T cell clone with replication-competent provirus integrated into the ZNF721 gene, termed ZNF721i. We stimulated the clone with its cognate peptide and then exposed the cells to paclitaxel and/or carboplatin or the antiproliferative drug, mycophenolate mofetil. While treatment of cells with the cognate peptide alone led to a marked expansion of the ZNF721i clone, treatment with the cognate peptide followed by culture with either paclitaxel or mycophenolate mofetil abrogated this process. The drugs did not affect the proliferation of other CD4+ T cell clones that were not specific for the cognate peptide. This strategy of antigen-specific stimulation followed by treatment with an antiproliferative agent may lead to the selective elimination of clonally expanded HIV-infected cells.

Authors

Filippo Dragoni, Joel Sop, Isha Gurumurthy, Tyler P. Beckey, Kellie N. Smith, Francesco R. Simonetti, Joel N. Blankson

High 4E-BP-1 expression associates with chromosome 8 gain and CDK4/6 sensitivity in Ewing Sarcoma

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Abstract

Chromosome 8 (chr8) gains are common in cancer, but their contribution to tumor heterogeneity is largely unexplored. Ewing sarcoma (EwS) is defined by FET::ETS fusions with few other recurrent mutations to explain clinical diversity. In EwS, chr8 gains are the second most frequent alteration, making it an ideal model to study their relevance in an otherwise silent genomic context. We report that chr8 gain-driven expression patterns correlate with poor overall survival of EwS patients. This effect is mainly mediated by increased expression of the translation initiation factor binding protein 4E-BP1, encoded by EIF4EBP1 on chr8. Among all chr8-encoded genes, EIF4EBP1 expression showed the strongest association with poor survival and correlated with chr8 gains in EwS tumors. Similar findings emerged across multiple TCGA cancer entities. Multi-omics profiling revealed that 4E-BP1 orchestrates a pro-proliferative proteomic network. Silencing 4E-BP1 reduced proliferation, clonogenicity, spheroidal growth in vitro, and tumor growth in vivo. Drug screens demonstrated that high 4E-BP1 expression sensitizes EwS to pharmacological CDK4/6-inhibition. Chr8 gains and elevated 4E-BP1 emerge as prognostic biomarkers in EwS, with poor outcomes driven by 4E-BP1-mediated pro-proliferative networks that sensitize tumors to CDK4/6 inhibitors. Testing for chr8 gains may enhance risk stratification and therapy in EwS and other cancers.

Authors

Cornelius M. Funk, Anna C. Ehlers, Martin F. Orth, Karim Aljakouch, Jing Li, Tilman L.B. Hoelting, Rainer Will, Florian H. Geyer, A. Katharina Ceranski, Franziska Willis, Endrit Vinca, Shunya Ohmura, Roland Imle, Jana Siebenlist, Angelina Yershova, Maximilian M.L. Knott, Felina Zahnow, Ana Sastre, Javier Alonso, Felix Sahm, Heike Peterziel, Anna Loboda, Martin Schneider, Ana Banito, Gabriel Leprivier, Wolfgang Hartmann, Uta Dirksen, Olaf Witt, Ina Oehme, Stefan M. Pfister, Laura Romero-Pérez, Jeroen Krijgsveld, Florencia Cidre-Aranaz, Thomas G.P. Grünewald, Julian Musa

A positive allosteric modulator of the β₁AR with antagonist activity for catecholaminergic polymorphic ventricular tachycardia

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Abstract

Orthosteric β-blockers represent the leading pharmacological intervention for managing heart diseases owing to their ability to competitively antagonize β-adrenergic receptors (βARs). However, their use is often limited by the development of adverse effects such as fatigue, hypotension, and reduced exercise capacity, due in part to the nonselective inhibition of multiple βAR subtypes. These challenges are particularly problematic in treating catecholaminergic polymorphic ventricular tachycardia (CPVT), a disease characterized by lethal tachyarrhythmias directly triggered by cardiac β1AR activation. To identify small molecule allosteric modulators of the β1AR that could offer enhanced subtype specificity and robust functional antagonism of β1AR-mediated signaling, we conducted a DNA-encoded small molecule library screen and discovered Compound 11 (C11). C11 selectively potentiates the binding affinity of orthosteric agonists to the β1AR while potently inhibiting downstream signaling following β1AR activation. Moreover, C11 prevents agonist-induced spontaneous contractile activity, Ca2+ release events, and exercise-induced ventricular tachycardia in the CSQ2–/– murine model of CPVT. Collectively, our studies demonstrate that C11 belongs to an emerging class of allosteric modulators termed PAM-antagonists that positively modulate agonist binding but block downstream function. With unique pharmacological properties and selective functional antagonism of β1AR-mediated signaling, C11 represents a promising therapeutic candidate for the treatment of CPVT and other forms of cardiac disease associated with excessive β1AR activation.

Authors

Alyssa Grogan, Robin M. Perelli, Seungkirl Ahn, Haoran Jiang, Arun Jyothidasan, Damini Sood, Chongzhao You, David I. Israel, Alex Shaginian, Qiuxia Chen, Jian Liu, Jialu Wang, Jan Steyaert, Alem W. Kahsai, Andrew P. Landstrom, Robert J. Lefkowitz, Howard A. Rockman

Excessive collagen type VII mediates pleural fibrosis via increasing extracellular matrix stiffness

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Abstract

The interaction between cells and extracellular matrix (ECM) has been recognized in mechanism of fibrotic diseases. Collagen type VII (collagen VII) is an ECM component which plays an important role in cell-ECM interaction, particularly in cell anchoring and maintaining ECM integrity. Pleural mesothelial cells (PMCs) drive inflammatory reactions and ECM production in pleura. However, the role of collagen VII and PMCs in pleural fibrosis was poorly understood. In this study, collagen VII protein was found increase in pleura of patients with tuberculous pleural fibrosis. Investigation of cellular and animal models revealed that collagen VII began to increase at early stage in pleural fibrotic process. Increase of collagen VII occurred ahead of collagen I and α-SMA in PMCs and pleura of animal models. Inhibition of collagen VII by mesothelial cell-specific deletion of collagen VII gene (WT1-Cre+-COL7A1flox/flox) attenuated mouse experimental pleural fibrosis. At last, it was found that excessive collagen VII changed collagen conformation which resulted in elevation of ECM stiffness. Elevation of ECM stiffness activated integrin/PI3K-AKT/JUN signaling and promoted more ECM deposition, as well as mediated pleural fibrosis. In conclusion, excessive collagen VII mediated pleural fibrosis via increasing extracellular matrix stiffness.

Authors

Qian Li, Xin-Liang He, Shuai-Jun Chen, Qian Niu, Tan-Ze Cao, Xiao-Ling Cui, Zi-Heng Jia, He-De Zhang, Xiao Feng, Ye-Han Jiang, Li-Mei Liang, Pei-Pei Cheng, Shi-He Hu, Liang Xiong, Meng Wang, Hong Ye, Wan-Li Ma

Biallelic variants in ARHGAP19 cause a progressive inherited motor-predominant neuropathy

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Abstract

Charcot-Marie-Tooth Disease is a clinically and genetically heterogeneous group of hereditary neuropathies. Despite progress in genetic sequencing, around a quarter of patients remain unsolved. Here, we identify 16 recessive variants in the RhoGTPase activating protein 19 gene (ARHGAP19) causing motor-predominant neuropathy in 25 individuals from 20 unrelated families. The ARHGAP19 protein acts as a negative regulator of the RhoA GTPase. In vitro biochemical and cellular assays revealed that patient variants impair the GTPase-activating protein (GAP) activity of ARHGAP19 and reduce ARHGAP19 protein levels. Combined in vitro and in vivo studies reveal that human ARHGAP19, and conserved ARHGAP19 orthologs in Drosophila and Zebrafish, influence motoneuron morphology and promote locomotor capacity. Transcriptomic studies further demonstrate that ARHGAP19 regulates cellular pathways associated with motor proteins and the cell cycle. Taken together, our findings establish ARHGAP19 variants as a cause of inherited neuropathy acting through a loss-of-function mechanism.

Authors

Natalia Dominik, Stephanie Efthymiou, Christopher J. Record, Xinyu Miao, Renee Q. Lin, Jevin M. Parmar, Annarita Scardamaglia, Reza Maroofian, Simon A. Lowe, Gabriel N. Aughey, Abigail D. Wilson, Riccardo Curro, Ricardo P. Schnekenberg, Shahryar Alavi, Leif Leclaire, Yi He, Kristina Zhelcheska, Yohanns Bellaiche, Isabelle Gaugué, Mariola Skorupinska, Liedewei Van de Vondel, Sahar I. Da'as, Valentina Turchetti, Serdal Güngör, Gavin V. Monahan, Ehsan Ghayoor Karimiani, Yalda Jamshidi, Phillipa J. Lamont, Camila Armirola-Ricaurte, Haluk Topaloglu, Albena Jordanova, Mashaya Zaman, Selina H. Banu, Wilson Marques, Pedro J. Tomaselli, Busra Aynekin, Ali Cansu, Huseyin Per, Ayten Güleç, Javeria Raza Alvi, Tipu Sultan, Arif Khan, Giovanni Zifarelli, Shahnaz Ibrahim, Grazia M. S. Mancini, M.M. Motazacker, Esther Brusse, Vincenzo Lupo, Teresa Sevilla, A. Nazli Başak, Seyma Tekgul, Robin J. Palvadeau, Jonathan Baets, Yesim Parman, Arman Çakar, Rita Horvath, Tobias B. Haack, Jan-Hendrik Stahl, Kathrin Grundmann-Hauser, Joohyun Park, Stephan Zuchner, Nigel G. Laing, Lindsay A. Wilson, Alexander M. Rossor, James Polke, Fernanda Barbosa Figueiredo, André Pessoa, Fernando Kok, Antônio Rodrigues Coimbra-Neto, Marcondes C. Franca Jr, Gianina Ravenscroft, Sherifa A. Hamed, Wendy K. Chung, Alan M. Pittman, Daniel P. Osborn, Michael Hanna, Andrea Cortese, Mary M. Reilly, James E.C. Jepson, Nathalie Lamarche-Vane, Henry Houlden

Stimulated thyroid hormone synthesis machinery drives thyrocyte cell death independent of ER stress

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Abstract

It is now recognized that patients and animal models expressing genetically-encoded misfolded mutant thyroglobulin (TG, the protein precursor for thyroid hormone synthesis) exhibit dramatic swelling of the endoplasmic reticulum (ER) with ER stress and cell death in thyrocytes — seen both in homozygotes (with severe hypothyroidism) and heterozygotes (with subclinical hypothyroidism). The thyrocyte death phenotype is exacerbated upon thyroidal stimulation (by thyrotropin, TSH), as cell death is inhibited upon treatment with exogenous thyroxine. TSH stimulation might contribute to cytotoxicity by promoting ER stress, or by an independent mechanism. Here we’ve engineered knockout mice completely lacking Tg expression. Like other animals/patients with mutant TG, these animals rapidly develop severe goitrous hypothyroidism; however, thyroidal ER stress is exceedingly low — lower even than that seen in wildtype mice. Nevertheless, mice lacking TG exhibit abundant thyroid cell death, which depends upon renegade thyroidal iodination — it is completely suppressed in a genetic model lacking effective iodination, or in Tg-KO mice treated with propylthiouracil (iodination inhibitor), or iodide deficiency. Thyrocytes in culture are killed not in the presence of H2O2 alone, but rather upon peroxidase-mediated iodination, with cell death blocked by propylthiouracil. Thus, in the thyroid gland bearing Tg mutation(s), TSH-stimulated iodination activity triggers thyroid cell death.

Authors

Crystal Young, Xiaohan Zhang, Xiaofan Wang, Aaron P. Kellogg, Kevin Pena, August Z. Cumming, Xiao-Hui Liao, Dennis Larkin, Hao Zhang, Emma Mastroianni, Helmut Grasberger, Samuel Refetoff, Peter Arvan