Mutations in unfolded protein response regulator ATF6 cause hearing and vision loss syndrome

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Abstract

Activating transcription factor 6 (Atf6) is a key regulator of the unfolded protein response (UPR) and is important for endoplasmic reticulum (ER) function and protein homeostasis in metazoan cells. Patients carrying loss-of-function ATF6 disease alleles develop the cone dysfunction disorder, achromatopsia. The impact of loss of ATF6 function on other cell types, organs, and diseases in people remains unclear. Here, we reported that progressive sensorineural hearing loss was a notable complaint in some patients carrying ATF6 disease alleles and that Atf6-/- mice also showed progressive auditory deficits affecting both genders. In mice with hearing deficits, we found disorganized stereocilia on hair cells and focal loss of outer hair cells. Transcriptomic analysis of Atf6-/- cochleae revealed marked induction of UPR, especially through the PERK arm. These findings identify ATF6 as an essential regulator of cochlear health and function. Furthermore, they supported that ATF6 inactivation in people causes progressive sensorineural hearing loss as part of a blindness-deafness genetic syndrome targeting hair cells and cone photoreceptors. Lastly, our genetic findings support ER stress as an important pathomechanism underlying cochlear damage and hearing loss with clinical implications for patient lifestyle modifications that minimize environmental/physiologic sources of ER stress to the ear.

Authors

Eun-Jin Lee, Kyle Kim, Monica Sophia Diaz-Aguilar, Hyejung Min, Eduardo Chavez, Korina J. Steinbergs, Lance A. Safarta, Guirong Zhang, Allen F. Ryan, Jonathan H. Lin

G-CSF resistance of ELANE mutant neutropenia depends on SERF1 containing truncated neutrophil elastase aggregates

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Abstract

Severe congenital neutropenia (SCN) is frequently associated with dominant point mutations in ELANE, the gene encoding neutrophil elastase (NE). Chronic administration of granulocyte colony-stimulating factor (G-CSF) is a first-line treatment of ELANE-mutant (ELANEmut) SCN. However, some ELANEmut patients including patients with ELANE start codon mutations do not respond to G-CSF. Here, through directed granulopoiesis of gene-edited isogenic normal and patient-derived iPSCs, we demonstrate that ELANE start codon mutations suffice to induce G-CSF resistant granulocytic precursor cell death and refractory SCN. ELANE start codon mutated neutrophil precursors express predominantly nuclear N-terminal truncated alternate NE. Unlike G-CSF sensitive ELANE mutations that induce endoplasmic reticulum and unfolded protein response stress, we found that the mutation of the ELANE translation initiation codon resulted in NE aggregates and activated pro-apoptotic aggrephagy as determined by downregulated BAG1 expression, decreased BAG1/BAG3 ratio, NE co-localization with BAG3, and localized expression of autophagic LC3B. We found that SERF1, an RNA-chaperone protein, known to localize in misfolded protein aggregates in neurodegenerative diseases, was highly upregulated and interacted with cytoplasmic NE of mutant neutrophil precursors. Silencing of SERF1 enhanced survival and differentiation of iPSC-derived neutrophil precursors, restoring their responsiveness to G-CSF. These observations provide a mechanistic insight of G-CSF-resistant ELANEmut SCN, revealing targets for therapeutic intervention.

Authors

Ramesh C. Nayak, Sana Emberesh, Lisa Trump, Ashley Wellendorf, Abhishek Singh, Brice Korkmaz, Marshall S. Horwitz, Kasiani C. Myers, Theodosia A. Kalfa, Carolyn Lutzko, Jose A. Cancelas

An inducible RIPK3-driven necroptotic system enhances cancer cell-based immunotherapy and ensures safety

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Abstract

Recent progress in cancer cell-based therapies has led to effective targeting and robust immune responses against cancer. However, the inherent safety risks of using live cancer cells necessitate the creation of an optimized safety switch without hindering the efficacy of immunotherapy. The existing safety switches typically induce tolerogenic cell death, potentially leading to an immunosuppressive tumor immune microenvironment (TIME), which is counterproductive to the goals of immunotherapy. Here, we developed and characterized an inducible RIPK3-driven necroptotic system that serves as a dual function of safety switch as well as inducing immunogenic cell death which in turn stimulates antitumor immune responses. We showed that activating RIPK3 safety switch triggered immunogenic responses marked by an increased release of adenosine triphosphate (ATP) and damage-associated molecular patterns (DAMPs). Compared to other existing safety switches, incorporating RIPK3 system inhibited tumor growth, improved survival outcomes in tumor-bearing mice, and fostered long-term antitumor immunity. Moreover, RIPK3 system reinvigorated the TIME by promoting dendritic cell (DC) maturation, polarizing the macrophages towards the M1 phenotype, and reducing the exhaustion of CD4+ and CD8+ T lymphocytes. Our study highlights the dual role of RIPK3-driven necroptotic system in improving the safety and efficacy of cancer cell-based therapy, with broader implications for cellular therapies.

Authors

Kok-Siong Chen, Sarah Manoury-Battais, Nobuhiko Kanaya, Ioulia Vogiatzi, Paulo Borges, Sterre J. Kruize, Yi-Ching Chen, Laura Y. Lin, Filippo Rossignoli, Natalia Claire Mendonca, Khalid Shah

Tumor cell-derived spermidine promotes a pro-tumorigenic immune microenvironment in glioblastoma via CD8⁺ T cell inhibition

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Abstract

The glioblastoma (GBM) microenvironment is enriched in immunosuppressive factors that potently interfere with the function of cytotoxic T lymphocytes. Cancer cells can directly impact the immune system, but the mechanisms driving these interactions are not completely clear. Here we demonstrate that the polyamine metabolite spermidine (SPD) is elevated in the GBM tumor microenvironment. Exogenous administration of SPD drives tumor aggressiveness in an immune-dependent manner in pre-clinical mouse models via reduction of CD8+ T cell frequency and reduced cytotoxic function. Knockdown of ornithine decarboxylase, the rate-limiting enzyme in spermidine synthesis, did not impact cancer cell growth in vitro but did result in extended survival. Furthermore, glioblastoma patients with a more favorable outcome had a significant reduction in spermidine compared to patients with a poor prognosis. Our results demonstrate that spermidine functions as a cancer cell-derived metabolite that drives tumor progression by reducing CD8+ T cell number and function.

Authors

Kristen E. Kay, Juyeun Lee, Ellen S. Hong, Julia Beilis, Sahil Dayal, Emily R. Wesley, Sofia Mitchell, Sabrina Z. Wang, Daniel J. Silver, Josephine Volovetz, Sarah Johnson, Mary McGraw, Matthew Grabowski, Tianyao Lu, Lutz Freytag, Vinod K. Narayana, Saskia Freytag, Sarah A. Best, James R. Whittle, Zeneng Wang, Ofer Reizes, Jennifer S. Yu, Stanley L. Hazen, J. Mark Brown, Defne Bayik, Justin Lathia

Low tristetraprolin expression activates phenotypic plasticity and primes transition to lethal prostate cancer in mice

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Abstract

Phenotypic plasticity is a hallmark of cancer and increasingly realized as a mechanism of resistance to androgen receptor (AR)-targeted therapy. Now that many prostate cancer (PCa) patients are treated upfront with AR-targeted agents, it’s critical to identify actionable mechanisms that drive phenotypic plasticity, to prevent the emergence of resistance. We showed that loss of tristetraprolin (TTP, gene ZFP36) increased NF-κB activation, and was associated with higher rates of aggressive disease and early recurrence in primary PCa. We also examined the clinical and biological impact of ZFP36 loss with co-loss of PTEN, a known driver of PCa. Analysis of multiple independent primary PCa cohorts demonstrated that PTEN and ZFP36 co-loss was associated with increased recurrence risk. Engineering prostate-specific Zfp36 deletion in vivo, induced prostatic intraepithelial neoplasia, and, with Pten co-deletion, resulted in rapid progression to castration-resistant adenocarcinoma. Zfp36 loss altered the cell state driven by Pten loss, demonstrated by enrichment of EMT, inflammation, TNFα/NF-κB, IL6-JAK/STAT3 gene sets. Additionally, our work revealed that ZFP36 loss also induced enrichment of multiple gene sets involved in mononuclear cell migration, chemotaxis, and proliferation. Use of the NF-κB inhibitor, dimethylaminoparthenolide (DMAPT) induced marked therapeutic responses in tumors with PTEN and ZFP36 co-loss and reversed castration resistance.

Authors

Katherine L. Morel, Beatriz Germán, Anis A. Hamid, Jagpreet S. Nanda, Simon Linder, Andries M. Bergman, Henk van der Poel, Ingrid Hofland, Elise M. Bekers, Shana Y. Trostel, Deborah L. Burkhart, Scott Wilkinson, Anson T. Ku, Minhyung Kim, Jina Kim, Duanduan Ma, Jasmine T. Plummer, Sungyong You, Xiaofeng A. Su, Wilbert Zwart, Adam G. Sowalsky, Christopher J. Sweeney, Leigh Ellis

Rapid response of lichen planus to baricitinib associated with suppression of cytotoxic CXCL13⁺ CD8⁺ T-cells

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Abstract

BACKGROUND. Cutaneous lichen planus (LP) is a recalcitrant, difficult-to-treat, inflammatory skin disease characterized by pruritic, flat-topped, violaceous papules on the skin. Baricitinib is an oral Janus kinase (JAK) 1/2 inhibitor that interrupts the signaling pathway of interferon gamma (IFN)-γ, a cytokine implicated in the pathogenesis of LP. METHODS. In this phase II trial, twelve patients with cutaneous LP received baricitinib 2 mg daily for 16 weeks, accompanied by in-depth spatial, single-cell, and bulk transcriptomic profiling of pre- and post-treatment samples. RESULTS. An early and sustained clinical response was seen, with 83.3% of patients responsive at week 16. Our molecular data identified a unique, oligoclonal IFN-γ, CD8+, CXCL13+ cytotoxic T-cell population in LP skin and demonstrated a rapid decrease in IFN signature within 2 weeks of treatment, most prominently in the basal layer of the epidermis. CONCLUSION. This study demonstrates the efficacy and molecular mechanisms of JAK inhibition in LP. TRIAL REGISTRATION. NCT05188521. ROLE OF FUNDING SOURCE. Eli Lilly, Appignani Benefactor Funds, 5P30AR075043, Mayo Clinic Clinical Trials Stimulus Funds.

Authors

Angelina S. Hwang, Jacob A. Kechter, Tran H. Do, Alysia N. Hughes, Nan Zhang, Xing Li, Rachael Bogle, Caitlin M. Brumfiel, Meera H. Patel, Blake Boudreaux, Puneet Bhullar, Shams Nassir, Miranda L. Yousif, Alyssa L. Stockard, Zachary Leibovit-Reiben, Ewoma Ogbaudu, David J. DiCaudo, Jennifer Fox, Mehrnaz Gharaee-Kermani, Xianying Xing, Samantha Zunich, Emily Branch, J. Michelle Kahlenberg, Allison C. Billi, Olesya Plazyo, Lam C. Tsoi, Mark R. Pittelkow, Johann E. Gudjonsson, Aaron R. Mangold

Complement-producing maternal microchimeric cells override infection susceptibility in complement-deficient murine offspring

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Abstract

Authors

Giang Pham, Raymond E. Diep, Lucien H. Turner, David B. Haslam, Sing Sing Way

Single-nuclei transcriptomics reveals TBX5-dependent targets in a patient with Holt-Oram syndrome

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Abstract

Authors

Jeffrey D. Steimle, Yi Zhao, Fansen Meng, Mikaela E. Taylor, Diwakar Turaga, Iki Adachi, Xiao Li, James F. Martin

Nicotinamide and pyridoxine stimulate muscle stem cell expansion and enhance regenerative capacity during aging

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Abstract

Skeletal muscle relies on resident muscle stem cells (MuSCs) for growth and repair. Aging and muscle diseases impair MuSC function, leading to stem cell exhaustion and regenerative decline that contribute to the progressive loss of skeletal muscle mass and strength. In the absence of clinically available nutritional solutions specifically targeting MuSCs, we used a human myogenic progenitor (hMP) high-content imaging screen of natural molecules from food to identify nicotinamide (NAM) and pyridoxine (PN) as bioactive nutrients that stimulate MuSCs and have history of safe human use. NAM and PN synergize via CK1-mediated cytoplasmic β-catenin activation and AKT signaling to promote amplification and differentiation of MuSCs. Oral treatment with a combination of NAM/PN accelerates muscle regeneration in vivo by stimulating MuSCs, increases muscle strength during recovery, and overcomes MuSC dysfunction and regenerative failure during aging. Levels of NAM and bioactive PN spontaneously decline during aging in model organisms and inter-independently associate with muscle mass and walking speed in a human cohort of 186 aged people. Collectively, our results establish NAM/PN as a new nutritional intervention that stimulates MuSCs, enhances muscle regeneration, and alleviates age-related muscle decline with a direct opportunity for clinical translation.

Authors

Sara Ancel, Joris Michaud, Eugenia Migliavacca, Charline Jomard, Aurélie Fessard, Pauline Garcia, Sonia Karaz, Sruthi Raja, Guillaume E. Jacot, Thibaut Desgeorges, José-Luis Sánchez-García, Loic Tauzin, Yann Ratinaud, Benjamin Brinon, Sylviane Métairon, Lucas Pinero, Denis Barron, Stephanie Blum, Leonidas G. Karagounis, Ramin Heshmat, Afshin Ostovar, Farshad Farzadfar, Isabella Scionti, Rémi Mounier, Julien Gondin, Pascal Stuelsatz, Jerome N. Feige

Inflammatory crosstalk impairs phagocytic receptors and aggravates atherosclerosis in clonal hematopoiesis in mice

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Abstract

Clonal hematopoiesis (CH) increases inflammasome-linked atherosclerosis but the mechanisms by which CH mutant cells transmit inflammatory signals to non-mutant cells are largely unknown. To address this question we transplanted 1.5% Jak2VF bone marrow (BM) cells with 98.5% WT BM cells into hyperlipidemic Ldlr–/– mice. Low allele burden (LAB) mice showed accelerated atherosclerosis with increased features of plaque instability, decreased levels of macrophage phagocytic receptors MERTK and TREM2, and increased neutrophil extracellular traps (NETs). These changes were reversed when Jak2VF BM was transplanted with Il1r1–/– BM. LAB mice with non-cleavable MERTK in WT BM showed improvements in necrotic core and fibrous cap formation and reduced NETs. An agonistic TREM2 antibody (4D9) markedly increased fibrous caps in both control and LAB mice eliminating the difference between groups. Mechanistically, 4D9 increased TREM2+PDGFB+ macrophages and PDGF receptor-α positive fibroblast-like cells in the cap region. TREM2 and PDGFB mRNA levels were positively correlated in human carotid plaques and co-expressed in macrophages. In summary, low frequency Jak2VF mutations promote atherosclerosis via IL-1 signaling from Jak2VF to WT macrophages and neutrophils promoting cleavage of phagocytic receptors and features of plaque instability. Therapeutic approaches that stabilize MERTK or TREM2 could promote plaque stabilization especially in CH- and inflammasome-driven atherosclerosis.

Authors

Wenli Liu, Brian D. Hardaway, Eunyoung Kim, Jessica Pauli, Justus Leonard Wettich, Mustafa Yalcinkaya, Cheng-Chieh Hsu, Tong Xiao, Muredach P. Reilly, Ira Tabas, Lars Maegdefessel, Kai Schlepckow, Haass Christian, Nan Wang, Alan R. Tall