Research Article
Abstract
Although first-line epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) therapy is effective for treating EGFR-mutant non–small cell lung cancer (NSCLC), it is now understood that drug-tolerant persister (DTP) cells escaping from initial treatment eventually drives drug resistance. Here, through integration of metabolomics and transcriptomics, we found that the neurotransmitter acetylcholine (ACh) was specifically accumulated in DTP cells, and demonstrated that treatment with EGFR-TKI heightened the expression of the rate-limiting enzyme choline acetyltransferase (ChAT) in ACh biosynthesis via YAP mediation. Genetic and pharmacological manipulation of ACh biosynthesis or ACh signaling could predictably regulate the extent of DTP formation in vitro and in vivo. Strikingly, pharmacologically targeting ACh/M3R signaling with an FDA-approved drug, darifenacin, retarded tumor relapse in vivo. Mechanistically, upregulated ACh metabolism mediated drug tolerance in part through activating WNT signaling via ACh muscarinic receptor 3 (M3R). Importantly, we showed that aberrant ACh metabolism in patients with NSCLC played a potential role in predicting EGFR-TKI response rate and progression-free survival. Our study therefore defines a therapeutic strategy — targeting the ACh/M3R/WNT axis — for manipulating EGFR TKI drug tolerance in the treatment of NSCLC.
Authors
Meng Nie, Na Chen, Huanhuan Pang, Tao Jiang, Wei Jiang, Panwen Tian, LiAng Yao, Yangzi Chen, Ralph J. DeBerardinis, Weimin Li, Qitao Yu, Caicun Zhou, Zeping Hu
Abstract
Initiation and maintenance of transcriptional states are critical for controlling normal tissue homeostasis and differentiation. The cyclin dependent kinases CDK8 and CDK19 (Mediator kinases) are regulatory components of Mediator, a highly conserved complex that orchestrates enhancer-mediated transcriptional output. While Mediator kinases have been implicated in the transcription of genes necessary for development and growth, its function in mammals has not been well defined. Using genetically defined models and pharmacological inhibitors, we showed that CDK8 and CDK19 function in a redundant manner to regulate intestinal lineage specification in humans and mice. The Mediator kinase module bound and phosphorylated key components of the chromatin remodeling complex switch/sucrose non-fermentable (SWI/SNF) in intestinal epithelial cells. Concomitantly, SWI/SNF and MED12-Mediator colocalized at distinct lineage-specifying enhancers in a CDK8/19–dependent manner. Thus, these studies reveal a transcriptional mechanism of intestinal cell specification, coordinated by the interaction between the chromatin remodeling complex SWI/SNF and Mediator kinase.
Authors
Marius V. Dannappel, Danxi Zhu, Xin Sun, Hui Kheng Chua, Marle Poppelaars, Monica Suehiro, Subash Khadka, Terry C.C. Lim Kam Sian, Dhanya Sooraj, Melissa Loi, Hugh Gao, Daniel Croagh, Roger J. Daly, Pouya Faridi, Thomas G. Boyer, Ron Firestein
Abstract
CLN1 disease, also called infantile neuronal ceroid lipofuscinosis (NCL) or infantile Batten disease, is a fatal neurodegenerative lysosomal storage disorder resulting from mutations in the CLN1 gene encoding the soluble lysosomal enzyme palmitoyl-protein thioesterase 1 (PPT1). Therapies for CLN1 disease have proven challenging because of the aggressive disease course and the need to treat widespread areas of the brain and spinal cord. Indeed, gene therapy has proven less effective for CLN1 disease than for other similar lysosomal enzyme deficiencies. We therefore tested the efficacy of enzyme replacement therapy (ERT) by administering monthly infusions of recombinant human PPT1 (rhPPT1) to PPT1-deficient mice (Cln1–/–) and CLN1R151X sheep to assess how to potentially scale up for translation. In Cln1–/– mice, intracerebrovascular (i.c.v.) rhPPT1 delivery was the most effective route of administration, resulting in therapeutically relevant CNS levels of PPT1 activity. rhPPT1-treated mice had improved motor function, reduced disease-associated pathology, and diminished neuronal loss. In CLN1R151X sheep, i.c.v. infusions resulted in widespread rhPPT1 distribution and positive treatment effects measured by quantitative structural MRI and neuropathology. This study demonstrates the feasibility and therapeutic efficacy of i.c.v. rhPPT1 ERT. These findings represent a key step toward clinical testing of ERT in children with CLN1 disease and highlight the importance of a cross-species approach to developing a successful treatment strategy.
Authors
Hemanth R. Nelvagal, Samantha L. Eaton, Sophie H. Wang, Elizabeth M. Eultgen, Keigo Takahashi, Steven Q. Le, Rachel Nesbitt, Joshua T. Dearborn, Nicholas Siano, Ana C. Puhl, Patricia I. Dickson, Gerard Thompson, Fraser Murdoch, Paul M. Brennan, Mark Gray, Stephen N. Greenhalgh, Peter Tennant, Rachael Gregson, Eddie Clutton, James Nixon, Chris Proudfoot, Stefano Guido, Simon G. Lillico, C. Bruce A. Whitelaw, Jui-Yun Lu, Sandra L. Hofmann, Sean Ekins, Mark S. Sands, Thomas M. Wishart, Jonathan D. Cooper
Abstract
The etiology of type 1 diabetes has polygenic and environmental determinants that lead to autoimmune responses against pancreatic β cells and promote β cell death. The autoimmunity is considered silent without metabolic consequences until late preclinical stages,and it remains unknown how early in the disease process the pancreatic β cell is compromised. To address this, we investigated preprandial nonfasting and postprandial blood glucose concentrations and islet autoantibody development in 1,050 children with high genetic risk of type 1 diabetes. Pre- and postprandial blood glucose decreased between 4 and 18 months of age and gradually increased until the final measurements at 3.6 years of age. Determinants of blood glucose trajectories in the first year of life included sex, body mass index, glucose-related genetic risk scores, and the type 1 diabetes–susceptible INS gene. Children who developed islet autoantibodies had early elevations in blood glucose concentrations. A sharp and sustained rise in postprandial blood glucose was observed at around 2 months prior to autoantibody seroconversion, with further increases in postprandial and, subsequently, preprandial values after seroconversion. These findings show heterogeneity in blood glucose control in infancy and early childhood and suggest that islet autoimmunity is concurrent or subsequent to insults on the pancreatic islets.
Authors
Katharina Warncke, Andreas Weiss, Peter Achenbach, Thekla von dem Berge, Reinhard Berner, Kristina Casteels, Lidia Groele, Konstantinos Hatzikotoulas, Angela Hommel, Olga Kordonouri, Helena Elding Larsson, Markus Lundgren, Benjamin A. Marcus, Matthew D. Snape, Agnieszka Szypowska, John A. Todd, Ezio Bonifacio, Anette-G. Ziegler, for the GPPAD and POInT Study Groups
Abstract
Preexisting lung-restricted autoantibodies (LRAs) are associated with a higher incidence of primary graft dysfunction (PGD), although it remains unclear whether LRAs can drive its pathogenesis. In syngeneic murine left lung transplant recipients, preexisting LRAs worsened graft dysfunction, which was evident by impaired gas exchange, increased pulmonary edema, and activation of damage-associated pathways in lung epithelial cells. LRA-mediated injury was distinct from ischemia-reperfusion injury since deletion of donor nonclassical monocytes and host neutrophils could not prevent graft dysfunction in LRA-pretreated recipients. Whole LRA IgG molecules were necessary for lung injury, which was mediated by the classical and alternative complement pathways and reversed by complement inhibition. However, deletion of Fc receptors in donor macrophages or mannose-binding lectin in recipient mice failed to rescue lung function. LRA-mediated injury was localized to the transplanted lung and dependent on IL-1β–mediated permeabilization of pulmonary vascular endothelium, which allowed extravasation of antibodies. Genetic deletion or pharmacological inhibition of IL-1R in the donor lungs prevented LRA-induced graft injury. In humans, preexisting LRAs were an independent risk factor for severe PGD and could be treated with plasmapheresis and complement blockade. We conclude that preexisting LRAs can compound ischemia-reperfusion injury to worsen PGD for which complement inhibition may be effective.
Authors
Wenbin Yang, Emily Jeong Cerier, Félix L. Núñez-Santana, Qiang Wu, Yuanqing Yan, Chitaru Kurihara, Xianpeng Liu, Anjana Yeldandi, Nigar Khurram, Diego Avella-Patino, Haiying Sun, G.R. Scott Budinger, Daniel Kreisel, Thalachallour Mohanakumar, Emilia Lecuona, Ankit Bharat
Abstract
The mTORC1 pathway coordinates nutrient and growth factor signals to maintain organismal homeostasis. Whether nutrient signaling to mTORC1 regulates stem cell function remains unknown. Here, we show that SZT2 — a protein required for mTORC1 downregulation upon nutrient deprivation — is critical for hematopoietic stem cell (HSC) homeostasis. Ablation of SZT2 in HSCs decreased the reserve and impaired the repopulating capacity of HSCs. Furthermore, ablation of both SZT2 and TSC1 — 2 repressors of mTORC1 on the nutrient and growth factor arms, respectively — led to rapid HSC depletion, pancytopenia, and premature death of the mice. Mechanistically, loss of either SZT2 or TSC1 in HSCs led to only mild elevation of mTORC1 activity and reactive oxygen species (ROS) production. Loss of both SZT2 and TSC1, on the other hand, simultaneously produced a dramatic synergistic effect, with an approximately 10-fold increase of mTORC1 activity and approximately 100-fold increase of ROS production, which rapidly depleted HSCs. These data demonstrate a critical role of nutrient mTORC1 signaling in HSC homeostasis and uncover a strong synergistic effect between nutrient- and growth factor–mediated mTORC1 regulation in stem cells.
Authors
Na Yin, Gang Jin, Yuying Ma, Hanfei Zhao, Guangyue Zhang, Ming O. Li, Min Peng
Abstract
Food allergies are a leading cause of anaphylaxis, and allergen-specific immune responses in both the innate and the adaptive immune system play key roles in its pathogenesis. We conducted a comprehensive phenotypic and functional investigation of immune cell responses from nonallergic (NA) and peanut allergic (PA) participants cultured with media alone or peanut protein and found, surprisingly, that NK cell activation was strongly associated with the immune response to allergen in PA participants. Peanut-responsive NK cells manifested a distinct expression pattern in PA participants compared with NA participants. Allergen-activated NK cells expressed both Th2 and immune regulatory cytokines, hinting at a potential functional role in mediating and regulating the Th2 allergic response. Depletion of CD3+ T cells attenuated the response of NK cells to peanut-allergen stimulation, suggesting that peanut-responsive NK cells are T cell dependent. We also showed that oral immune therapy was associated with decreased NK responses to peanut allergen stimulation in vitro. These results demonstrate that NK cells are associated with the food-allergic immune response, and the magnitude of this mobilized cell population suggests that they play a functional role in allergic immunity.
Authors
Xiaoying Zhou, Wong Yu, Diane M. Dunham, Jackson P. Schuetz, Catherine A. Blish, Rosemarie H. DeKruyff, Kari C. Nadeau
Abstract
B cell depletion in patients with relapsing-remitting multiple sclerosis (RRMS) markedly prevents new MRI-detected lesions and disease activity, suggesting the hypothesis that altered B cell function leads to the activation of T cells driving disease pathogenesis. Here, we performed comprehensive analyses of CD40 ligand– (CD40L-) and IL-21–stimulated memory B cells from patients with MS and healthy age-matched controls, modeling the help of follicular helper T cells (Tfh cells), and found a differential gene expression signature in multiple B cell pathways. Most striking was the impaired TIGIT expression on MS-derived B cells mediated by dysregulation of the transcription factor TCF4. Activated circulating Tfh cells (cTfh cells) expressed CD155, the ligand of TIGIT, and TIGIT on B cells revealed their capacity to suppress the proliferation of IL-17–producing cTfh cells via the TIGIT/CD155 axis. Finally, CCR6+ cTfh cells were significantly increased in patients with MS, and their frequency was inversely correlated with that of TIGIT+ B cells. Together, these data suggest that the dysregulation of negative feedback loops between TIGIT+ memory B cells and cTfh cells in MS drives the activated immune system in this disease.
Authors
Hiromitsu Asashima, Pierre-Paul Axisa, Thi Hong Giang Pham, Erin E. Longbrake, William E. Ruff, Nikhil Lele, Inessa Cohen, Khadir Raddassi, Tomokazu S. Sumida, David A. Hafler
Abstract
Infantile (fetal and neonatal) megakaryocytes (Mks) have a distinct phenotype consisting of hyperproliferation, limited morphogenesis, and low platelet production capacity. These properties contribute to clinical problems that include thrombocytopenia in neonates, delayed platelet engraftment in recipients of cord blood stem cell transplants, and inefficient ex vivo platelet production from pluripotent stem cell–derived Mks. The infantile phenotype results from deficiency of the actin-regulated coactivator, MKL1, which programs cytoskeletal changes driving morphogenesis. As a strategy to complement this molecular defect, we screened pathways with the potential to affect MKL1 function and found that DYRK1A inhibition dramatically enhanced Mk morphogenesis in vitro and in vivo. Dyrk1 inhibitors rescued enlargement, polyploidization, and thrombopoiesis in human neonatal Mks. Mks derived from induced pluripotent stem cells responded in a similar manner. Progenitors undergoing Dyrk1 inhibition demonstrated filamentous actin assembly, MKL1 nuclear translocation, and modulation of MKL1 target genes. Loss-of-function studies confirmed MKL1 involvement in this morphogenetic pathway. Expression of Ablim2, a stabilizer of filamentous actin, increased with Dyrk1 inhibition, and Ablim2 knockdown abrogated the actin, MKL1, and morphogenetic responses to Dyrk1 inhibition. These results delineate a pharmacologically tractable morphogenetic pathway whose manipulation may alleviate clinical problems associated with the limited thrombopoietic capacity of infantile Mks.
Authors
Kamaleldin E. Elagib, Ashton Brock, Cara M. Clementelli, Goar Mosoyan, Lorrie L. Delehanty, Ranjit K. Sahu, Alexandra Pacheco-Benichou, Corinne Fruit, Thierry Besson, Stephan W. Morris, Koji Eto, Chintan Jobaliya, Deborah L. French, Paul Gadue, Sandeep Singh, Xinrui Shi, Fujun Qin, Robert Cornelison, Hui Li, Camelia Iancu-Rubin, Adam N. Goldfarb
Abstract
Evidence suggests that increased microRNA-155 (miR-155) expression in immune cells enhances antitumor immune responses. However, given the reported association of miR-155 with tumorigenesis in various cancers, a debate is provoked on whether miR-155 is oncogenic or tumor suppressive. We aimed to interrogate the impact of tumor miR-155 expression, particularly that of cancer cell–derived miR-155, on antitumor immunity in breast cancer. We performed bioinformatic analysis of human breast cancer databases, murine experiments, and human specimen examination. We revealed that higher tumor miR-155 levels correlate with a favorable antitumor immune profile and better patient outcomes. Murine experiments demonstrated that miR-155 overexpression in breast cancer cells enhanced T cell influx, delayed tumor growth, and sensitized the tumors to immune checkpoint blockade (ICB) therapy. Mechanistically, miR-155 overexpression in breast cancer cells upregulated their CXCL9/10/11 production, which was mediated by SOCS1 inhibition and increased phosphorylated STAT1 (p-STAT1)/p-STAT3 ratios. We further found that serum miR-155 levels in breast cancer patients correlated with tumor miR-155 levels and tumor immune status. Our findings suggest that high serum and tumor miR-155 levels may be a favorable prognostic marker for breast cancer patients and that therapeutic elevation of miR-155 in breast tumors may improve the efficacy of ICB therapy via remodeling the antitumor immune landscape.
Authors
Junfeng Wang, Quanyi Wang, Yinan Guan, Yulu Sun, Xiaozhi Wang, Kaylie Lively, Yuzhen Wang, Ming Luo, Julian A. Kim, E. Angela Murphy, Yongzhong Yao, Guoshuai Cai, Daping Fan
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ISSN: 0021-9738 (print), 1558-8238 (online)