Research Article
Abstract
Cell cycle regulation is largely abnormal in cancers. Molecular understanding and therapeutic targeting of the aberrant cell cycle are essential. Here, we identified that an underappreciated serine/threonine kinase, cyclin-dependent kinase–like 3 (CDKL3), crucially drives rapid cell cycle progression and cell growth in cancers. With regard to mechanism, CDKL3 localizes in the nucleus and associates with specific cyclin to directly phosphorylate retinoblastoma (Rb) for quiescence exit. In parallel, CDKL3 prevents the ubiquitin-proteasomal degradation of cyclin-dependent kinase 4 (CDK4) by direct phosphorylation on T172 to sustain G1 phase advancement. The crucial function of CDKL3 in cancers was demonstrated both in vitro and in vivo. We also designed, synthesized, and characterized a first-in-class CDKL3-specific inhibitor, HZ1. HZ1 exhibits greater potency than CDK4/6 inhibitor in pan-cancer treatment by causing cell cycle arrest and overcomes acquired resistance to CDK4/6 inhibitor. In particular, CDKL3 has significant clinical relevance in colon cancer, and the effectiveness of HZ1 was demonstrated by murine and patient-derived cancer models. Collectively, this work presents an integrated paradigm of cancer cell cycle regulation and suggests CDKL3 targeting as a feasible approach in cancer treatment.
Authors
Haijiao Zhang, Jiahui Lin, Shaoqin Zheng, Lanjing Ma, Zhongqiu Pang, Hongyi Yin, Chengcheng Meng, Yinuo Wang, Qing Han, Xi Zhang, Zexu Li, Liu Cao, Lijun Liu, Teng Fei, Daming Gao, Liang Yang, Xueqiang Peng, Chen Ding, Shixue Wang, Ren Sheng
Abstract
Myostatin (MSTN) has long been recognized as a critical regulator of muscle mass. Recently, there has been increasing interest in its role in metabolism. In our study, we specifically knocked out MSTN in brown adipose tissue (BAT) from mice (MSTNΔUCP1) and found that the mice gained more weight than did controls when fed a high-fat diet, with progressive hepatosteatosis and impaired skeletal muscle activity. RNA-Seq analysis indicated signatures of mitochondrial dysfunction and inflammation in the MSTN-ablated BAT. Further studies demonstrated that Kruppel-like factor 4 (KLF4) was responsible for the metabolic phenotypes observed, whereas fibroblast growth factor 21 (FGF21) contributed to the microenvironment communication between adipocytes and macrophages induced by the loss of MSTN. Moreover, the MSTN/SMAD2/3-p38 signaling pathway mediated the expression of KLF4 and FGF21 in adipocytes. In summary, our findings suggest that brown adipocyte–derived MSTN regulated BAT thermogenesis via autocrine and paracrine effects on adipocytes or macrophages, ultimately regulating systemic energy homeostasis.
Authors
Hui Wang, Shanshan Guo, Huanqing Gao, Jiyang Ding, Hongyun Li, Xingyu Kong, Shuang Zhang, Muyang He, Yonghao Feng, Wei Wu, Kexin Xu, Yuxuan Chen, Hanyin Zhang, Tiemin Liu, Xingxing Kong
Abstract
Leukemia relapse is a major cause of death after allogeneic hematopoietic cell transplantation (allo-HCT). We tested the potential of targeting T cell (Tc) immunoglobulin and mucin-containing molecule 3 (TIM-3) for improving graft-versus-leukemia (GVL) effects. We observed differential expression of TIM-3 ligands when hematopoietic stem cells overexpressed certain oncogenic-driver mutations. Anti–TIM-3 Ab treatment improved survival of mice bearing leukemia with oncogene-induced TIM-3 ligand expression. Conversely, leukemia cells with low ligand expression were anti–TIM-3 treatment resistant. In vitro, TIM-3 blockade or genetic deletion in CD8+ Tc enhanced Tc activation, proliferation, and IFN-γ production while enhancing GVL effects, preventing Tc exhaustion, and improving Tc cytotoxicity and glycolysis in vivo. Conversely, TIM-3 deletion in myeloid cells did not affect allogeneic Tc proliferation and activation in vitro, suggesting that anti–TIM-3 treatment–mediated GVL effects are Tc induced. In contrast to anti–programmed cell death protein 1 (anti–PD-1) and anti–cytotoxic T lymphocyte–associated protein 4 (anti–CTLA-4) treatment, anti–TIM-3-treatment did not enhance acute graft-versus-host disease (aGVHD). TIM-3 and its ligands were frequently expressed in acute myeloid leukemia (AML) cells of patients with post–allo-HCT relapse. We decipher the connections between oncogenic mutations found in AML and TIM-3 ligand expression and identify anti–TIM-3 treatment as a strategy for enhancing GVL effects via metabolic and transcriptional Tc reprogramming without exacerbation of aGVHD. Our findings support clinical testing of anti–TIM-3 Ab in patients with AML relapse after allo-HCT.
Authors
Nana Talvard-Balland, Lukas M. Braun, Karen O. Dixon, Melissa Zwick, Helena Engel, Alina Hartmann, Sandra Duquesne, Livius Penter, Geoffroy Andrieux, Lukas Rindlisbacher, Andrea Acerbis, Jule Ehmann, Christoph Köllerer, Michela Ansuinelli, Andres Rettig, Kevin Moschallski, Petya Apostolova, Tilman Brummer, Anna L. Illert, Markus A. Schramm, Yurong Cheng, Anna Köttgen, Justus Duyster, Hans D. Menssen, Jerome Ritz, Bruce R. Blazar, Melanie Boerries, Annette Schmitt-Gräff, Nurefsan Sariipek, Peter Van Galen, Joerg M. Buescher, Nina Cabezas-Wallscheid, Heike L. Pahl, Erika L. Pearce, Robert J. Soiffer, Catherine J. Wu, Luca Vago, Burkhard Becher, Natalie Köhler, Tobias Wertheimer, Vijay K. Kuchroo, Robert Zeiser
Abstract
IgG4-related disease (IgG4-RD) is a systemic immune-mediated fibroinflammatory disease whose pathomechanisms remain poorly understood. Here, we identified gene variants in familial IgG4-RD and determined their functional consequences. All 3 affected members of the family shared variants of the transcription factor IKAROS, encoded by IKZF1, and the E3 ubiquitin ligase UBR4. The IKAROS variant increased binding to the FYN promoter, resulting in higher transcription of FYN in T cells. The UBR4 variant prevented the lysosomal degradation of the phosphatase CD45. In the presence of elevated FYN, CD45 functioned as a positive regulatory loop, lowering the threshold for T cell activation. Consequently, T cells from the affected family members were hyperresponsive to stimulation. When transduced with a low-avidity, autoreactive T cell receptor, their T cells responded to the autoantigenic peptide. In parallel, high expression of FYN in T cells biased their differentiation toward Th2 polarization by stabilizing the transcription factor JunB. This bias was consistent with the frequent atopic manifestations in patients with IgG4-RD, including the affected family members in the present study. Building on the functional consequences of these 2 variants, we propose a disease model that is not only instructive for IgG4-RD but also for atopic diseases and autoimmune diseases associated with an IKZF1 risk haplotype.
Authors
Qingxiang Liu, Yanyan Zheng, Ines Sturmlechner, Abhinav Jain, Maryam Own, Qiankun Yang, Huimin Zhang, Filippo Pinto e Vairo, Karen Cerosaletti, Jane H. Buckner, Kenneth J. Warrington, Matthew J. Koster, Cornelia M. Weyand, Jörg J. Goronzy
Abstract
Tolerance of mouse kidney allografts arises in grafts that develop regulatory tertiary lymphoid organs (rTLOs). Single-cell RNA-seq (scRNA-seq) data and adoptive transfer of alloreactive T cells after transplantation showed that cytotoxic CD8+ T cells are reprogrammed within the accepted graft to an exhausted/regulatory-like phenotype mediated by IFN-γ. Establishment of rTLOs was required because adoptive transfer of alloreactive T cells prior to transplantation results in kidney allograft rejection. Despite the presence of intragraft CD8+ cells with a regulatory phenotype, they were not essential for the induction and maintenance of kidney allograft tolerance since renal allotransplantation into CD8-KO recipients resulted in acceptance and not rejection. Analysis of scRNA-seq data from allograft kidneys and malignant tumors identified similar regulatory-like cell types within the T cell clusters and trajectory analysis showed that cytotoxic CD8+ T cells are reprogrammed into an exhausted/regulatory-like phenotype intratumorally. Induction of cytotoxic CD8+ T cell dysfunction of infiltrating cells appears to be a beneficial mechanistic pathway that protects the kidney allotransplant from rejection through a process we call “defensive tolerance.” This pathway has implications for our understanding of allotransplant tolerance and tumor resistance to host immunity.
Authors
Takahiro Yokose, Edward S. Szuter, Ivy Rosales, Michael T. Guinn, Andrew S. Liss, Taisuke Baba, David A. Ruddy, Michelle Piquet, Jamil Azzi, A. Benedict Cosimi, Paul S. Russell, Joren C. Madsen, Robert B. Colvin, Alessandro Alessandrini
Abstract
Reelin (RELN) is a secreted glycoprotein essential for cerebral cortex development. In humans, recessive RELN variants cause cortical and cerebellar malformations, while heterozygous variants were associated with epilepsy, autism, and mild cortical abnormalities. However, the functional effects of RELN variants remain unknown. We identified inherited and de novo RELN missense variants in heterozygous patients with neuronal migration disorders (NMDs) as diverse as pachygyria and polymicrogyria. We investigated in culture and in the developing mouse cerebral cortex how different variants impacted RELN function. Polymicrogyria-associated variants behaved as gain-of-function, showing an enhanced ability to induce neuronal aggregation, while those linked to pachygyria behaved as loss-of-function, leading to defective neuronal aggregation/migration. The pachygyria-associated de novo heterozygous RELN variants acted as dominant-negative by preventing WT RELN secretion in culture, animal models, and patients, thereby causing dominant NMDs. We demonstrated how mutant RELN proteins in vitro and in vivo predict cortical malformation phenotypes, providing valuable insights into the pathogenesis of such disorders.
Authors
Martina Riva, Sofia Ferreira, Kotaro Hayashi, Yoann Saillour, Vera P. Medvedeva, Takao Honda, Kanehiro Hayashi, Claire Altersitz, Shahad Albadri, Marion Rosello, Julie Dang, Malo Serafini, Frédéric Causeret, Olivia J. Henry, Charles-Joris Roux, Céline Bellesme, Elena Freri, Dragana Josifova, Elena Parrini, Renzo Guerrini, Filippo Del Bene, Kazunori Nakajima, Nadia Bahi-Buisson, Alessandra Pierani
The NR4A2/VGF pathway fuels inflammation-induced neurodegeneration via promoting neuronal glycolysis
Abstract
A disturbed balance between excitation and inhibition (E/I balance) is increasingly recognized as a key driver of neurodegeneration in multiple sclerosis (MS), a chronic inflammatory disease of the central nervous system. To understand how chronic hyperexcitability contributes to neuronal loss in MS, we transcriptionally profiled neurons from mice lacking inhibitory metabotropic glutamate signaling with shifted E/I balance and increased vulnerability to inflammation-induced neurodegeneration. This revealed a prominent induction of the nuclear receptor NR4A2 in neurons. Mechanistically, NR4A2 increased susceptibility to excitotoxicity by stimulating continuous VGF secretion leading to glycolysis-dependent neuronal cell death. Extending these findings to people with MS (pwMS), we observed increased VGF levels in serum and brain biopsies. Notably, neuron-specific deletion of Vgf in a mouse model of MS ameliorated neurodegeneration. These findings underscore the detrimental effect of a persistent metabolic shift driven by excitatory activity as a fundamental mechanism in inflammation-induced neurodegeneration.
Authors
Marcel S. Woo, Lukas C. Bal, Ingo Winschel, Elias Manca, Mark Walkenhorst, Bachar Sevgili, Jana K. Sonner, Giovanni Di Liberto, Christina Mayer, Lars Binkle-Ladisch, Nicola Rothammer, Lisa Unger, Lukas Raich, Alexandros Hadjilaou, Barbara Noli, Antonio L. Manai, Vanessa Vieira, Nina Meurs, Ingrid Wagner, Ole Pless, Cristina Cocco, Samuel B. Stephens, Markus Glatzel, Doron Merkler, Manuel A. Friese
Abstract
Tissue fibrosis remains unamenable to meaningful therapeutic interventions and is the primary cause of chronic graft failure after organ transplantation. Eukaryotic translation initiation factor (eIF4E), a key translational regulator, serves as convergent target of multiple upstream profibrotic signaling pathways that contribute to mesenchymal cell (MC) activation. Here, we investigate the role of MAP kinase–interacting serine/threonine kinase–induced (MNK-induced) direct phosphorylation of eIF4E at serine 209 (Ser209) in maintaining fibrotic transformation of MCs and determine the contribution of the MNK/eIF4E pathway to the pathogenesis of chronic lung allograft dysfunction (CLAD). MCs from patients with CLAD demonstrated constitutively higher eIF4E phosphorylation at Ser209, and eIF4E phospho-Ser209 was found to be critical in regulating key fibrogenic protein autotaxin, leading to sustained β-catenin activation and profibrotic functions of CLAD MCs. MNK1 signaling was upregulated in CLAD MCs, and genetic or pharmacologic targeting of MNK1 activity inhibited eIF4E phospho-Ser209 and profibrotic functions of CLAD MCs in vitro. Treatment with an MNK1/2 inhibitor (eFT-508) abrogated allograft fibrosis in an orthotopic murine lung-transplant model. Together these studies identify what we believe is a previously unrecognized MNK/eIF4E/ATX/β-catenin signaling pathway of fibrotic transformation of MCs and present the first evidence, to our knowledge, for the utility of MNK inhibitors in fibrosis.
Authors
Natalie M. Walker, Yuta Ibuki, A. Patrick McLinden, Keizo Misumi, Dylan C. Mitchell, Gabriel G. Kleer, Alison M. Lock, Ragini Vittal, Nahum Sonenberg, Amanda L. Garner, Vibha N. Lama
Abstract
Our study was to characterize sarcopenia in C57BL/6J mice using a clinically relevant definition to investigate the underlying molecular mechanisms. Aged male (23–32 months old) and female (27–28 months old) C57BL/6J mice were classified as non-, probable-, or sarcopenic based on assessments of grip strength, muscle mass, and treadmill running time, using 2 SDs below the mean of their young counterparts as cutoff points. A 9%–22% prevalence of sarcopenia was identified in 23–26 month-old male mice, with more severe age-related declines in muscle function than mass. Females aged 27–28 months showed fewer sarcopenic but more probable cases compared with the males. As sarcopenia progressed, a decrease in muscle contractility and a trend toward lower type IIB fiber size were observed in males. Mitochondrial biogenesis, oxidative capacity, and AMPK-autophagy signaling decreased as sarcopenia progressed in males, with pathways linked to mitochondrial metabolism positively correlated with muscle mass. No age- or sarcopenia-related changes were observed in mitochondrial biogenesis, OXPHOS complexes, AMPK signaling, mitophagy, or atrogenes in females. Our results highlight the different trajectories of age-related declines in muscle mass and function, providing insights into sex-dependent molecular changes associated with sarcopenia progression, which may inform the future development of novel therapeutic interventions.
Authors
Haiming L. Kerr, Kora Krumm, Barbara Anderson, Anthony Christiani, Lena Strait, Theresa Li, Brynn Irwin, Siyi Jiang, Artur Rybachok, Amanda Chen, Elizabeth Dacek, Lucas Caeiro, Gennifer E. Merrihew, James W. MacDonald, Theo K. Bammler, Michael J. MacCoss, Jose M. Garcia
Abstract
CD44 is associated with a high risk of metastasis, recurrence, and drug resistance in various cancers. Here we report that platelet endothelial aggregation receptor 1 (PEAR1) is a CD44 chaperone protein that protected CD44 from endocytosis-mediated degradation and enhances cleavage of the CD44 intracellular domain (CD44-ICD). Furthermore, we found that lysyl oxidase–like protein 2 (LOXL2), an endogenous ligand of PEAR1, bound to the PEAR1-EMI domain and facilitated the interaction between PEAR1 and CD44 by inducing PEAR1 Ser891 phosphorylation in a manner that was independent of its enzyme activity. Levels of PEAR1 protein and PEAR1 phosphorylation at Ser891 were increased in patients with triple-negative breast cancer (TNBC), were positively correlated with expression of LOXL2 and CD44, and were negatively correlated with overall survival. The level of PEAR1 Ser891 phosphorylation was identified as the best independent prognostic factor in TNBC patients. The prognostic efficacy of the combination of PEAR1 phosphorylation at Ser891 and CD44 expression was superior to that of PEAR1 phosphorylation at Ser891 alone. Blocking the interaction between LOXL2 and PEAR1 with monoclonal antibodies significantly inhibited TNBC metastasis, representing a promising therapeutic strategy for TNBC.
Authors
Yingzhi Shen, Jie Yan, Lin Li, Huiyan Sun, Lin Zhang, Guoming Li, Xinxia Wang, Ruoyan Liu, Xuefeng Wu, Baosan Han, Xueqing Sun, Junling Liu, Xuemei Fan
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ISSN: 0021-9738 (print), 1558-8238 (online)