Issue published January 2, 2014 Previous issue | Next issue
- Volume 124, Issue 1
Go to section:
On the cover: Hippo signaling in alveolar rhabdomyosarcoma
-
Science in Medicine
×
Abstract
The ubiquitin proteasome system (UPS) is an essential metabolic constituent of cellular physiology that tightly regulates cellular protein concentrations with specificity and precision to optimize cellular function. Inhibition of the proteasome has proven very effective in the treatment of multiple myeloma, and this approach is being tested for utility in other malignancies. New pharmaceuticals targeting the proteasome itself or specific proximal pathways of the UPS are in development as antiproliferatives or immunomodulatory agents. In this article, we discuss the biology of UPS-targeting drugs, their use as therapy for neoplasia, and the state of clinical and preclinical development for emerging therapeutics.
Authors
Nathaniel M. Weathington, Rama K. Mallampalli
-
Review Series
×
Abstract
The field of epigenetics has exploded in the last two decades, with incredible advances in recent years driven by high-throughput sequencing studies. Cancer cells frequently exhibit marked changes in DNA methylation and histone modification during tumorigenesis and tumor progression. These changes in the cancer epigenome are thought to be important in initiating and maintaining malignancy, and pharmaceutical approaches targeting epigenome-modifying enzymes are an attractive therapeutic strategy. Early successes have been made with DNA-demethylating drugs in hematologic malignancies, and efforts are underway to target additional epigenetic regulators and a broader array of tumor types. The Reviews in this issue of the
JCI highlight ongoing efforts in this burgeoning field to translate our understanding of the cancer epigenome into successful interventional strategies in the clinic.Authors
Peter A. Jones
×Abstract
The Cancer Genome Atlas (TCGA) Research Network is an ambitious multi-institutional consortium effort aimed at characterizing sequence, copy number, gene (mRNA) expression, microRNA expression, and DNA methylation alterations in 30 cancer types. TCGA data have become an extraordinary resource for basic, translational, and clinical researchers and have the potential to shape cancer diagnostic and treatment strategies. DNA methylation changes are integral to all aspects of cancer genomics and have been shown to have important associations with gene expression, sequence, and copy number changes. This Review highlights the knowledge gained from DNA methylation alterations in human cancers from TCGA.
Authors
Daniel J. Weisenberger
×Abstract
The term epigenetics refers to stable patterns of gene expression that are seen during differentiation or X chromosome inactivation and are not dependent on dynamic changes in coding DNA. These gene expression states are encoded in the epigenome — a collection of marks on DNA or on histone tails that are established during embryogenesis. Genome-wide studies in aging cells and tissues have uncovered stochastic DNA methylation drift (gradual increases or decreases at specific loci) that reflects imperfect maintenance of epigenetic marks. Drift creates epigenetic mosaicism in aging stem cells that could potentially restrict their plasticity and worsen phenotypes such as stem cell exhaustion and focal proliferative defects that can lead to cancer.
Authors
Jean-Pierre Issa
×Abstract
Epigenetic enzymes are often dysregulated in human tumors through mutation, altered expression, or inappropriate recruitment to certain loci. The identification of these enzymes and their partner proteins has driven the rapid development of small-molecule inhibitors that target the cancer epigenome. Herein, we discuss the influence of aberrantly regulated histone deacetylases (HDACs) in tumorigenesis. We examine HDAC inhibitors (HDACis) targeting class I, II, and IV HDACs that are currently under development for use as anticancer agents following the FDA approval of two HDACis, vorinostat and romidepsin.
Authors
Alison C. West, Ricky W. Johnstone
×Abstract
The term epigenetics refers to the heritable changes in gene expression that are not associated with a change in the actual DNA sequence. Epigenetic dysregulation is linked to the pathogenesis of a number of malignancies and has been studied extensively in myelodysplastic syndromes and acute myeloid leukemia. DNA methylation is frequently altered in cancerous cells and likely results in transcriptional silencing of tumor suppressor genes. Re-expression of these genes by inhibition of the DNA methyltransferases has been successful in the treatment of benign and malignant disease. In this Review, we discuss the clinical development of demethylating agents in hematology, with a focus on azacitidine and decitabine.
Authors
Shyamala C. Navada, Juliane Steinmann, Michael Lübbert, Lewis R. Silverman
×Abstract
Drugs targeting the epigenome are new promising cancer treatment modalities; however, not all patients receive the same benefit from these drugs. In contrast to conventional chemotherapy, responses may take several months after the initiation of treatment to occur. Accordingly, identification of good pretreatment predictors of response is of great value. Many clinical parameters and molecular targets have been tested in preclinical and clinical studies with varying results, leaving room for optimization. Here we provide an overview of markers that may predict the efficacy of FDA- and EMA-approved epigenetic drugs.
Authors
Marianne B. Treppendahl, Lasse S. Kristensen, Kirsten Grønbæk
×Abstract
Epigenetic therapies may play a prominent role in the future management of solid tumors. This possibility is based on the clinical efficacy of existing drugs in treating defined hematopoietic neoplasms, paired with promising new data from preclinical and clinical studies that examined these agents in solid tumors. We suggest that current drugs may represent a targeted therapeutic approach for reprogramming solid tumor cells, a strategy that must be pursued in concert with the explosion in knowledge about the molecular underpinnings of normal and cancer epigenomes. We hypothesize that understanding targeted proteins in the context of their enzymatic and scaffolding functions and in terms of their interactions in complexes with proteins that are targets of new drugs under development defines the future of epigenetic therapies for cancer.
Authors
Nita Ahuja, Hariharan Easwaran, Stephen B. Baylin
×Abstract
Over the past several years, there has been rapidly expanding evidence of epigenetic dysregulation in cancer, in which histone and DNA modification play a critical role in tumor growth and survival. These findings have gained the attention of the drug discovery and development community, and offer the potential for a second generation of cancer epigenetic agents for patients following the approved “first generation” of DNA methylation (e.g., Dacogen, Vidaza) and broad-spectrum HDAC inhibitors (e.g., Vorinostat, Romidepsin). This Review provides an analysis of prospects for discovery and development of novel cancer agents that target epigenetic proteins. We will examine key examples of epigenetic dysregulation in tumors as well as challenges to epigenetic drug discovery with emerging biology and novel classes of drug targets. We will also highlight recent successes in cancer epigenetics drug discovery and consider important factors for clinical success in this burgeoning area.
Authors
Robert M. Campbell, Peter J. Tummino
-
Hindsight
×
Abstract
I devised a method for obtaining information on cancellous bone structure from iliac bone histomorphometry that led to the demonstration that architecture is an important component of bone strength and bone fragility. Furthermore, this method contributed to the recognition of the importance of changes in osteoclast and osteocyte apoptosis in response to estrogen deficiency and replacement.
Authors
A. Michael Parfitt
-
Commentaries
×
Abstract
Oxaliplatin, a commonly used chemotherapeutic agent, is associated with both acute and chronic neurotoxicity. Chronic sensory neuropathy can be dose limiting and may have detrimental effects on patients’ quality of life. Preclinical studies provide an understanding of the pathophysiology of chemotherapy-induced peripheral neuropathy (CIPN) and may be important for developing effective preventative interventions. In this issue of the
JCI , Coriat and colleagues used an animal model and a human pilot trial to evaluate the use of mangafodipir to reduce CIPN. Although many pilot clinical studies have reported promising data, larger clinical trials have repeatedly been unable to confirm these preliminary results. Thus, no agents are currently clinically recommended for the prevention of CIPN.Authors
Deirdre R. Pachman, Charles L. Loprinzi, Axel Grothey, Lauren E. Ta
×Abstract
The ability to suppress the immune system has lead to great advances in transplant technology and treatment of autoimmune diseases. Unfortunately, the immunosuppression of these patients has led to the rise of opportunistic infections by organisms that are recalcitrant to current prophylactic strategies. One such example is the increase of mucormycosis, an invasive infection caused by filamentous fungi of the order Mucorales. In this issue of the
JCI , Gebremariam and colleagues determined that spore coat homolog (CotH) proteins are required for angioinvasion and that these proteins are unique to Mucorales. Their findings provide a potential therapeutic target for prevention and treatment of mucormycosis.Authors
J. Andrew Alspaugh
×Abstract
Epidemiological studies have identified racial differences in susceptibility to numerous diseases, including several ocular and skin diseases characterized by increased vascular growth. In most cases, the specific mechanisms and genetic variants responsible for these differences have remained elusive. In this issue of the
JCI , Adini et al. explore a direct connection between skin pigmentation and susceptibility to angiogenic diseases and identify an extracellular matrix protein that is regulated by melanogenesis and potently modulates angiogenesis.Authors
Christopher D. Kontos
×Abstract
Loeys-Dietz syndrome (LDS) is a connective tissue disorder that is characterized by skeletal abnormalities, craniofacial malformations, and a high predisposition for aortic aneurysm. In this issue of the
JCI , Gallo et al. developed transgenic mouse strains harboring missense mutations in the genes encoding type I or II TGF-β receptors. These mice exhibited several LDS-associated phenotypes. Despite being functionally defective, the mutated receptors enhanced TGF-β signaling in vivo, inferred by detection of increased levels of phosphorylated Smad2. Aortic aneurysms in these LDS mice were ablated by treatment with the Ang II type 1 (AT1) receptor antagonist losartan. The results from this study will foster further interest into the potential therapeutic implications of AT1 receptor antagonists.Authors
Frank Davis, Debra L. Rateri, Alan Daugherty
×Abstract
Aniridia is a panocular disorder that severely affects vision in early life. Most cases are caused by dominantly inherited mutations or deletions of the
PAX6 gene, which encodes a transcription factor that is essential for the development of the eye and the central nervous system. In this issue of theJCI , Gregory-Evans and colleagues demonstrate that early postnatal topical administration of an ataluren-based formulation reverses congenital malformations in the postnatal mouse eye, providing evidence that manipulation ofPAX6 after birth may lead to corrective tissue remodeling. These findings offer hope that ataluren administration could be a therapeutic paradigm applicable to some major congenital eye defects.Authors
José-Alain Sahel, Katia Marazova
-
Research Articles
×
Abstract
Defective neurogenesis in the postnatal brain can lead to many neurological and psychiatric disorders, yet the mechanism behind postnatal neurogenesis remains to be investigated. Huntingtin-associated protein 1 (HAP1) participates in intracellular trafficking in neurons, and its absence leads to postnatal death in mice. Here, we used tamoxifen-induced (TM-induced) Cre recombination to deplete HAP1 in mice at different ages. We found that HAP1 reduction selectively affects survival and growth of postnatal mice, but not adults. Neurogenesis, but not gliogenesis, was affected in HAP1-null neurospheres and mouse brain. In the absence of HAP1, postnatal hypothalamic neurons exhibited reduced receptor tropomyosin-related kinase B (TRKB) levels and decreased survival. HAP1 stabilized the association of TRKB with the intracellular sorting protein sortilin, prevented TRKB degradation, and promoted its anterograde transport. Our findings indicate that intracellular sorting of neurotrophin receptors is critical for postnatal neurogenesis and could provide a therapeutic target for defective postnatal neurogenesis.
Authors
Jianxing Xiang, Hao Yang, Ting Zhao, Miao Sun, Xingshun Xu, Xin-Fu Zhou, Shi-Hua Li, Xiao-Jiang Li
×Abstract
High-dose (HD) IL-2 therapy in patients with cancer increases the general population of Tregs, which are positive for CD4, CD25, and the Treg-specific marker Foxp3. It is unknown whether specific subsets of Tregs are activated and expanded during HD IL-2 therapy or whether activation of any particular Treg subset correlates with clinical outcome. Here, we evaluated Treg population subsets that were induced in patients with melanoma following HD IL-2 therapy. We identified a Treg population that was positive for CD4, CD25, Foxp3, and the inducible T cell costimulator (ICOS). This Treg population increased more than any other lymphocyte subset during HD IL-2 therapy and had an activated Treg phenotype, as indicated by high levels of CD39, CD73, and TGF-β. ICOS+ Tregs were the most proliferative lymphocyte population in the blood after IL-2 therapy. Patients with melanoma with enhanced expansion of ICOS+ Tregs in blood following the first cycle of HD IL-2 therapy had worse clinical outcomes than patients with fewer ICOS+ Tregs. However, there was no difference in total Treg expansion between HD IL-2 responders and nonresponders. These data suggest that increased expansion of the ICOS+ Treg population following the first cycle of HD IL-2 therapy may be predictive of clinical outcome.
Authors
Geok Choo Sim, Natalia Martin-Orozco, Lei Jin, Yan Yang, Sheng Wu, Edwina Washington, Deborah Sanders, Carol Lacey, Yijun Wang, Luis Vence, Patrick Hwu, Laszlo Radvanyi
×Abstract
Aniridia is a congenital and progressive panocular condition with poor visual prognosis that is associated with brain, olfactory, and pancreatic abnormalities. Development of aniridia is linked with nonsense mutations that result in paired box 6 (
PAX6 ) haploinsufficiency. Here, we used a mouse model of aniridia to test the hypothesis that manipulation ofPax6 dosage through a mutation-independent nonsense mutation suppression strategy would limit progressive, postnatal damage in the eye. We focused on the nonsense suppression drugs 3-[5-(2-fluorophenyl)-1,2,4-oxadiazol-3-yl]benzoic acid (ataluren) and gentamicin. Remarkably, we demonstrated that nonsense suppression not only inhibited disease progression but also stably reversed corneal, lens, and retinal malformation defects and restored electrical and behavioral responses of the retina. The most successful results were achieved through topical application of the drug formulation START (0.9%s odium chloride, 1%T ween 80, 1% powdereda taluren, 1% car boxymet hylcellulose), which was designed to enhance particle dispersion and to increase suspension viscosity. These observations suggest that the eye retains marked developmental plasticity into the postnatal period and remains sensitive to molecular remodeling. Furthermore, these data indicate that other neurological developmental anomalies associated with dosage-sensitive genetic mutations may be reversible through nonsense suppression therapeutics.Authors
Cheryl Y. Gregory-Evans, Xia Wang, Kishor M. Wasan, Jinying Zhao, Andrew L. Metcalfe, Kevin Gregory-Evans
×Abstract
Approximately 85% of lung cancers are non–small-cell lung cancers (NSCLCs), which are often diagnosed at an advanced stage and associated with poor prognosis. Currently, there are very few therapies available for NSCLCs due to the recalcitrant nature of this cancer. Mutations that activate the small GTPase KRAS are found in 20% to 30% of NSCLCs. Here, we report that inhibition of superoxide dismutase 1 (SOD1) by the small molecule ATN-224 induced cell death in various NSCLC cells, including those harboring
KRAS mutations. ATN-224–dependent SOD1 inhibition increased superoxide, which diminished enzyme activity of the antioxidant glutathione peroxidase, leading to an increase in intracellular hydrogen peroxide (H2O2) levels. We found that ATN-224–induced cell death was mediated through H2O2-dependent activation of P38 MAPK and that P38 activation led to a decrease in the antiapoptotic factor MCL1, which is often upregulated in NSCLC. Treatment with both ATN-224 and ABT-263, an inhibitor of the apoptosis regulators BCL2/BCLXL, augmented cell death. Furthermore, we demonstrate that ATN-224 reduced tumor burden in a mouse model of NSCLC. Our results indicate that antioxidant inhibition by ATN-224 has potential clinical applications as a single agent, or in combination with other drugs, for the treatment of patients with various forms of NSCLC, includingKRAS -driven cancers.Authors
Andrea Glasauer, Laura A. Sena, Lauren P. Diebold, Andrew P. Mazar, Navdeep S. Chandel
×Abstract
Cardiac resynchronization therapy (CRT), the application of biventricular stimulation to correct discoordinate contraction, is the only heart failure treatment that enhances acute and chronic systolic function, increases cardiac work, and reduces mortality. Resting myocyte function also increases after CRT despite only modest improvement in calcium transients, suggesting that CRT may enhance myofilament calcium responsiveness. To test this hypothesis, we examined adult dogs subjected to tachypacing-induced heart failure for 6 weeks, concurrent with ventricular dyssynchrony (HFdys) or CRT. Myofilament force-calcium relationships were measured in skinned trabeculae and/or myocytes. Compared with control, maximal calcium-activated force and calcium sensitivity declined globally in HFdys; however, CRT restored both. Phosphatase PP1 induced calcium desensitization in control and CRT-treated cells, while HFdys cells were unaffected, implying that CRT enhances myofilament phosphorylation. Proteomics revealed phosphorylation sites on Z-disk and M-band proteins, which were predicted to be targets of glycogen synthase kinase-3β (GSK-3β). We found that GSK-3β was deactivated in HFdys and reactivated by CRT. Mass spectrometry of myofilament proteins from HFdys animals incubated with GSK-3β confirmed GSK-3β–dependent phosphorylation at many of the same sites observed with CRT. GSK-3β restored calcium sensitivity in HFdys, but did not affect control or CRT cells. These data indicate that CRT improves calcium responsiveness of myofilaments following HFdys through GSK-3β reactivation, identifying a therapeutic approach to enhancing contractile function.
Authors
Jonathan A. Kirk, Ronald J. Holewinski, Viola Kooij, Giulio Agnetti, Richard S. Tunin, Namthip Witayavanitkul, Pieter P. de Tombe, Wei Dong Gao, Jennifer Van Eyk, David A. Kass
×Abstract
Malaria, which is the result of
Plasmodium falciparum infection, is a global health threat that resulted in 655,000 deaths and 216 million clinical cases in 2010 alone. Recent phase 3 trials with malaria vaccine candidate RTS,S/AS01 (RTS,S) in children has demonstrated modest efficacy against clinical and severe malaria. RTS,S targets the pre-erythrocytic phase of the disease and induces high antibody titers against theP. falciparum circumsporozoite protein (CSP) and a moderate CD4+ T cell response. The individual contribution of these adaptive immune responses to protection from infection remains unknown. Here, we found that prophylactic administration of anti-CSP mAbs derived from an RTS,S-vaccinated recipient fully protected mice with humanized livers from i.v.- and mosquito bite–deliveredP. falciparum sporozoite challenge. Titers of anti-CSP that conveyed full protection were within the range observed in human RTS,S vaccine recipients. Increasing anti-CSP titers resulted in a dose-dependent reduction of the liver parasite burden. These data indicate that RTS,S-induced antibodies are protective and provide sterilizing immunity againstP. falciparum infection when reaching or exceeding a critical plasma concentration.Authors
Lander Foquet, Cornelus C. Hermsen, Geert-Jan van Gemert, Eva Van Braeckel, Karin E. Weening, Robert Sauerwein, Philip Meuleman, Geert Leroux-Roels
×Abstract
The renal disorder C3 glomerulopathy with dense deposit disease (C3G-DDD) pattern results from complement dysfunction and primarily affects children and young adults. There is no effective treatment, and patients often progress to end-stage renal failure. A small fraction of C3G-DDD cases linked to factor H or C3 gene mutations as well as autoantibodies have been reported. Here, we examined an index family with 2 patients with C3G-DDD and identified a chromosomal deletion in the complement factor H–related (
CFHR ) gene cluster. This deletion resulted in expression of a hybrid CFHR2-CFHR5 plasma protein. The recombinant hybrid protein stabilized the C3 convertase and reduced factor H–mediated convertase decay. One patient was refractory to plasma replacement and exchange therapy, as evidenced by the hybrid protein quickly returning to pretreatment plasma levels. Subsequently, complement inhibitors were tested on serum from the patient for their ability to block activity of CFHR2-CFHR5. Soluble CR1 restored defective C3 convertase regulation; however, neither eculizumab nor tagged compstatin had any effect. Our findings provide insight into the importance of CFHR proteins for C3 convertase regulation and identify a genetic variation in theCFHR gene cluster that promotes C3G-DDD. Monitoring copy number and sequence variations in theCFHR gene cluster in C3G-DDD and kidney patients with C3G-DDD variations will help guide treatment strategies.Authors
Qian Chen, Michael Wiesener, Hannes U. Eberhardt, Andrea Hartmann, Barbara Uzonyi, Michael Kirschfink, Kerstin Amann, Maike Buettner, Tim Goodship, Christian Hugo, Christine Skerka, Peter F. Zipfel
×Abstract
Breast cancer (BC) can recur as metastatic disease many years after primary tumor removal, suggesting that disseminated tumor cells survive for extended periods in a dormant state that is refractory to conventional therapies. We have previously shown that altering the tumor microenvironment through fibrosis with collagen and fibronectin deposition can trigger tumor cells to switch from a dormant to a proliferative state. Here, we used an in vivo preclinical model and a 3D in vitro model of dormancy to evaluate the role of Src family kinase (SFK) in regulating this dormant-to-proliferative switch. We found that pharmacological inhibition of SFK signaling or
Src knockdown results in the nuclear localization of cyclin-dependent kinase inhibitor p27 and prevents the proliferative outbreak of dormant BC cells and metastatic lesion formation; however, SFK inhibition did not kill dormant cells. Dormant cell proliferation also required ERK1/2 activation. Combination treatment of cells undergoing the dormant-to-proliferative switch with the Src inhibitor (AZD0530) and MEK1/2 inhibitor (AZD6244) induced apoptosis in a large fraction of the dormant cells and delayed metastatic outgrowth, neither of which was observed with either inhibitor alone. Thus, targeting Src prevents the proliferative response of dormant cells to external stimuli, but requires MEK1/2 inhibition to suppress their survival. These data indicate that treatments targeting Src in combination with MEK1/2 may prevent BC recurrence.Authors
Lara H. El Touny, Anthony Vieira, Arnulfo Mendoza, Chand Khanna, Mark J. Hoenerhoff, Jeffrey E. Green
×Abstract
While murine-based systems to identify cancer-promoting agents (carcinogens) are established, models to identify compounds that promote aging (gerontogens) have not been described. For this purpose, we exploited the transcription of
p16INK4a , which rises dynamically with aging and correlates with age-associated disease. Activation ofp16INK4a was visualized in vivo using a murine strain that harbors a knockin of the luciferase gene into theCdkn2a locus (p16LUC mice). We exposedp16LUC mice to candidate gerontogens, including arsenic, high-fat diet, UV light, and cigarette smoke and serially imaged animals to monitor senescence induction. We show that exposure to a high-fat diet did not acceleratep16INK4a expression, whereas arsenic modestly augmented, and cigarette smoke and UV light potently augmented, activation ofp16INK4a -mediated senescence. This work provides a toxicological platform to study mammalian aging and suggests agents that directly damage DNA promote molecular aging.Authors
Jessica A. Sorrentino, Janakiraman Krishnamurthy, Stephen Tilley, James G. Alb Jr., Christin E. Burd, Norman E. Sharpless
×Abstract
Naive T helper cells differentiate into functionally distinct effector subsets that drive specialized immune responses. Recent studies indicate that some of the effector subsets have plasticity. Here, we used an EAE model and found that Th17 cells deficient in the transcription factor BCL11B upregulated the Th2-associated proteins GATA3 and IL-4 without decreasing RAR-related orphan receptor γ (RORγt), IL-17, and GM-CSF levels. Surprisingly, abnormal IL-4 production affected Th17 cell trafficking, diverting migration from the draining lymph nodes/CNS route to the mesenteric lymph nodes/gut route, which ameliorated EAE without overt colitis. T helper cell rerouting in EAE was dependent on IL-4, which enhanced retinoic acid (RA) production by dendritic cells, which further induced expression of gut-homing receptors CCR9 and α4β7 on
Bcl11b -deficient CD4+ T cells. Furthermore, IL-4 treatment or Th2 immunization of wild-type mice with EAE caused no alteration in Th17 cytokines or RORγt, but diverted T helper cell trafficking to the gut, which improved EAE outcome without overt colitis. Our data demonstrate that Th17 cells are permissive to Th2 gene expression without affecting Th17 gene expression. This Th17 plasticity has an impact on trafficking, which is a critical component of the immune response and may represent a possible avenue for treating multiple sclerosis.Authors
Danielle Califano, Keith J. Sweeney, Hung Le, Jeffrey VanValkenburgh, Eric Yager, William O’Connor Jr., Jeffrey S. Kennedy, David M. Jones, Dorina Avram
×Abstract
Recent studies have underscored the importance of memory T cells in mediating protective immunity against pathogens and cancer. Pharmacological inhibition of regulators that mediate T cell differentiation promotes the differentiation of activated CD8+ T cells into memory cells. Nonetheless, pharmacological agents have broad targets and can induce undesirable immunosuppressive effects. Here, we tested the hypothesis that aptamer-targeted siRNA inhibition of mTOR complex 1 (mTORC1) function in CD8+ T cells can enhance their differentiation into memory T cells and potentiate antitumor immunity more effectively than the pharmacologic inhibitor rapamycin. To specifically target activated cells, we conjugated an siRNA targeting the mTORC1 component raptor to an aptamer that binds 4-1BB, a costimulatory molecule that is expressed on CD8+ T cells following TCR stimulation. We found that systemic administration of the 4-1BB aptamer-raptor siRNA to mice downregulated mTORC1 activity in the majority of CD8+ T cells, leading to the generation of a potent memory response that exhibited cytotoxic effector functions and enhanced vaccine-induced protective immunity in tumor-bearing mice. In contrast, while treatment with the general mTORC1 inhibitor rapamycin also enhanced antigen-activated CD8+ T cell persistence, the cytotoxic effector functions of the reactivated memory cells were reduced and the alloreactivity of DCs was diminished. Consistent with the immunological findings, mice treated with rapamycin, but not with 4-1BB aptamer-raptor siRNA, failed to reject a subsequent tumor challenge.
Authors
Alexey Berezhnoy, Iris Castro, Agata Levay, Thomas R. Malek, Eli Gilboa
×Abstract
The ability of individual T cells to perform multiple effector functions is crucial for protective immunity against viruses and cancer. This polyfunctionality is frequently lost during chronic infections; however, the molecular mechanisms driving T cell polyfunctionality are poorly understood. We found that human T cells stimulated by a high concentration of antigen lacked polyfunctionality and expressed a transcription profile similar to that of exhausted T cells. One specific pathway implicated by the transcription profile in control of T cell polyfunctionality was the MAPK/ERK pathway. This pathway was altered in response to different antigen concentrations, and polyfunctionality correlated with upregulation of phosphorylated ERK. T cells that were stimulated with a high concentration of antigen upregulated sprouty-2 (
SPRY2 ), a negative regulator of the MAPK/ERK pathway. The clinical relevance of SPRY2 was confirmed by examiningSPRY2 expression in HIV-specific T cells, where high levels of SPRY2 were seen in HIV-specific T cells and inhibition ofSPRY2 expression enhanced the HIV-specific polyfunctional response independently of the PD-1 pathway. Our findings indicate that increasedSPRY2 expression during chronic viral infection reduces T cell polyfunctionality and identify SPRY2 as a potential target for immunotherapy.Authors
Yen-Ling Chiu, Liang Shan, Hailiang Huang, Carl Haupt, Catherine Bessell, David H. Canaday, Hao Zhang, Ya-Chi Ho, Jonathan D. Powell, Mathias Oelke, Joseph B. Margolick, Joel N. Blankson, Diane E. Griffin, Jonathan P. Schneck
×Abstract
Congenital diaphragmatic hernia (CDH) is a common birth malformation with a heterogeneous etiology. In this study, we report that ablation of the heparan sulfate biosynthetic enzyme NDST1 in murine endothelium (
Ndst1ECKO mice) disrupted vascular development in the diaphragm, which led to hypoxia as well as subsequent diaphragm hypoplasia and CDH. Intriguingly, the phenotypes displayed inNdst1ECKO mice resembled the developmental defects observed in slit homolog 3 (Slit3 ) knockout mice. Furthermore, introduction of a heterozygous mutation in roundabout homolog 4 (Robo4 ), the gene encoding the cognate receptor of SLIT3, aggravated the defect in vascular development in the diaphragm and CDH. NDST1 deficiency diminished SLIT3, but not ROBO4, binding to endothelial heparan sulfate and attenuated EC migration and in vivo neovascularization normally elicited by SLIT3-ROBO4 signaling. Together, these data suggest that heparan sulfate presentation of SLIT3 to ROBO4 facilitates initiation of this signaling cascade. Thus, our results demonstrate that loss of NDST1 causes defective diaphragm vascular development and CDH and that heparan sulfate facilitates angiogenic SLIT3-ROBO4 signaling during vascular development.Authors
Bing Zhang, Wenyuan Xiao, Hong Qiu, Fuming Zhang, Heather A. Moniz, Alexander Jaworski, Eduard Condac, Gerardo Gutierrez-Sanchez, Christian Heiss, Robin D. Clugston, Parastoo Azadi, John J. Greer, Carl Bergmann, Kelley W. Moremen, Dean Li, Robert J. Linhardt, Jeffrey D. Esko, Lianchun Wang
×Abstract
Acute myelogenous leukemia (AML) subtypes that result from oncogenic activation of homeobox (HOX) transcription factors are associated with poor prognosis. The HOXA9 transcription activator and growth factor independent 1 (GFI1) transcriptional repressor compete for occupancy at DNA-binding sites for the regulation of common target genes. We exploited this HOXA9 versus GFI1 antagonism to identify the genes encoding microRNA-21 and microRNA-196b as transcriptional targets of HOX-based leukemia oncoproteins. Therapeutic inhibition of microRNA-21 and microRNA-196b inhibited in vitro leukemic colony forming activity and depleted in vivo leukemia-initiating cell activity of HOX-based leukemias, which led to leukemia-free survival in a murine AML model and delayed disease onset in xenograft models. These data establish microRNA as functional effectors of endogenous HOXA9 and HOX-based leukemia oncoproteins, provide a concise in vivo platform to test RNA therapeutics, and suggest therapeutic value for microRNA antagonists in AML.
Authors
Chinavenmeni S. Velu, Aditya Chaubey, James D. Phelan, Shane R. Horman, Mark Wunderlich, Monica L. Guzman, Anil G. Jegga, Nancy J. Zeleznik-Le, Jianjun Chen, James C. Mulloy, Jose A. Cancelas, Craig T. Jordan, Bruce J. Aronow, Guido Marcucci, Balkrishen Bhat, Brian Gebelein, H. Leighton Grimes
×Abstract
Angioinvasion is a hallmark of mucormycosis. Previously, we identified endothelial cell glucose-regulated protein 78 (GRP78) as a receptor for Mucorales that mediates host cell invasion. Here we determined that spore coat protein homologs (CotH) of Mucorales act as fungal ligands for GRP78. CotH proteins were widely present in Mucorales and absent from noninvasive pathogens. Heterologous expression of
CotH3 andCotH2 inSaccharomyces cerevisiae conferred the ability to invade host cells via binding to GRP78. Homology modeling and computational docking studies indicated structurally compatible interactions between GRP78 and both CotH3 and CotH2. A mutant ofRhizopus oryzae , the most common cause of mucormycosis, with reduced CotH expression was impaired for invading and damaging endothelial cells and CHO cells overexpressing GRP78. This strain also exhibited reduced virulence in a diabetic ketoacidotic (DKA) mouse model of mucormycosis. Treatment with anti-CotH Abs abolished the ability ofR. oryzae to invade host cells and protected DKA mice from mucormycosis. The presence of CotH in Mucorales explained the specific susceptibility of DKA patients, who have increased GRP78 levels, to mucormycosis. Together, these data indicate that CotH3 and CotH2 function as invasins that interact with host cell GRP78 to mediate pathogenic host-cell interactions and identify CotH as a promising therapeutic target for mucormycosis.Authors
Teclegiorgis Gebremariam, Mingfu Liu, Guanpingsheng Luo, Vincent Bruno, Quynh T. Phan, Alan J. Waring, John E. Edwards Jr., Scott G. Filler, Michael R. Yeaman, Ashraf S. Ibrahim
×Abstract
Alpha-1 antitrypsin deficiency (AATD) is a rare genetic disease that results from mutations in the alpha-1 antitrypsin (
AAT ) gene. The mutant AAT protein aggregates and accumulates in the liver leading to AATD liver disease, which is only treatable by liver transplant. The PiZ transgenic mouse strain expresses a humanAAT (hAAT ) transgene that contains the AATD-associated Glu342Lys mutation. PiZ mice exhibit many AATD symptoms, including AAT protein aggregates, increased hepatocyte death, and liver fibrosis. In the present study, we systemically treated PiZ mice with an antisense oligonucleotide targeted againsthAAT (AAT-ASO) and found reductions in circulating levels of AAT and both soluble and aggregated AAT protein in the liver. Furthermore, AAT-ASO administration in these animals stopped liver disease progression after short-term treatment, reversed liver disease after long-term treatment, and prevented liver disease in young animals. Additionally, antisense oligonucleotide treatment markedly decreased liver fibrosis in this mouse model. Administration of AAT-ASO in nonhuman primates led to an approximately 80% reduction in levels of circulating normal AAT, demonstrating potential for this approach in higher species. Antisense oligonucleotides thus represent a promising therapy for AATD liver disease.Authors
Shuling Guo, Sheri L. Booten, Mariam Aghajan, Gene Hung, Chenguang Zhao, Keith Blomenkamp, Danielle Gattis, Andrew Watt, Susan M. Freier, Jeffery H. Teckman, Michael L. McCaleb, Brett P. Monia
×Abstract
Background. The majority of patients receiving the platinum-based chemotherapy drug oxaliplatin develop peripheral neurotoxicity. Because this neurotoxicity involves ROS production, we investigated the efficacy of mangafodipir, a molecule that has antioxidant properties and is approved for use as an MRI contrast enhancer.Methods. The effects of mangafodipir were examined in mice following treatment with oxaliplatin. Neurotoxicity, axon myelination, and advanced oxidized protein products (AOPPs) were monitored. In addition, we enrolled 23 cancer patients with grade ≥2 oxaliplatin-induced neuropathy in a phase II study, with 22 patients receiving i.v. mangafodipir following oxaliplatin. Neuropathic effects were monitored for up to 8 cycles of oxaliplatin and mangafodipir.Results. Mangafodipir prevented motor and sensory dysfunction and demyelinating lesion formation. In mice, serum AOPPs decreased after 4 weeks of mangafodipir treatment. In 77% of patients treated with oxaliplatin and mangafodipir, neuropathy improved or stabilized after 4 cycles. After 8 cycles, neurotoxicity was downgraded to grade ≥2 in 6 of 7 patients. Prior to enrollment, patients received an average of 880 ± 239 mg/m2 oxaliplatin. Patients treated with mangafodipir tolerated an additional dose of 458 ± 207 mg/m2 oxaliplatin despite preexisting neuropathy. Mangafodipir responders managed a cumulative dose of 1,426 ± 204 mg/m2 oxaliplatin. Serum AOPPs were lower in responders compared with those in nonresponders.Conclusion. Our study suggests that mangafodipir can prevent and/or relieve oxaliplatin-induced neuropathy in cancer patients.Trial registration. Clinicaltrials.gov NCT00727922.Funding. Université Paris Descartes, Ministère de la Recherche et de l’Enseignement Supérieur, and Assistance Publique-Hôpitaux de Paris.Authors
Romain Coriat, Jérôme Alexandre, Carole Nicco, Laurent Quinquis, Evelyne Benoit, Christiane Chéreau, Hervé Lemaréchal, Olivier Mir, Didier Borderie, Jean-Marc Tréluyer, Bernard Weill, Joel Coste, François Goldwasser, Frédéric Batteux
×Abstract
Mammals transport blood through a high-pressure, closed vascular network and lymph through a low-pressure, open vascular network. These vascular networks connect at the lymphovenous (LV) junction, where lymph drains into blood and an LV valve (LVV) prevents backflow of blood into lymphatic vessels. Here we describe an essential role for platelets in preventing blood from entering the lymphatic system at the LV junction. Loss of CLEC2, a receptor that activates platelets in response to lymphatic endothelial cells, resulted in backfilling of the lymphatic network with blood from the thoracic duct (TD) in both neonatal and mature mice. Fibrin-containing platelet thrombi were observed at the LVV and in the terminal TD in wild-type mice, but not
Clec2 -deficient mice. Analysis of mice lacking LVVs or lymphatic valves revealed that platelet-mediated thrombus formation limits LV backflow under conditions of impaired valve function. Examination of mice lacking integrin-mediated platelet aggregation indicated that platelet aggregation stabilizes thrombi that form in the lymphatic vascular environment to prevent retrograde blood flow. Collectively, these studies unveil a newly recognized form of hemostasis that functions with the LVV to safeguard the lymphatic vascular network throughout life.Authors
Paul R. Hess, David R. Rawnsley, Zoltán Jakus, Yiqing Yang, Daniel T. Sweet, Jianxin Fu, Brett Herzog, MinMin Lu, Bernhard Nieswandt, Guillermo Oliver, Taija Makinen, Lijun Xia, Mark L. Kahn
×Alveolar rhabdomyosarcoma–associated PAX3-FOXO1 promotes tumorigenesis via Hippo pathway suppression
Abstract
Alveolar rhabdomyosarcoma (aRMS) is an aggressive sarcoma of skeletal muscle characterized by expression of the paired box 3-forkhead box protein O1 (
PAX3-FOXO1 ) fusion oncogene. Despite its discovery nearly two decades ago, the mechanisms by which PAX3-FOXO1 drives tumor development are not well characterized. Previously, we reported that PAX3-FOXO1 supports aRMS initiation by enabling bypass of cellular senescence checkpoints. We have now found that this bypass occurs in part through PAX3-FOXO1–mediated upregulation ofRASSF4 , a Ras-association domain family (RASSF) member. RASSF4 expression was upregulated inPAX3-FOXO1 –positive aRMS cell lines and tumors. Enhanced RASSF4 expression promoted cell cycle progression, senescence evasion, and tumorigenesis through inhibition of the Hippo pathway tumor suppressor MST1. We also found that the downstream Hippo pathway target Yes-associated protein 1 (YAP), which is ordinarily restrained by Hippo signaling, was upregulated in RMS tumors. These data suggest that Hippo pathway dysfunction promotes RMS. This work provides evidence for Hippo pathway suppression in aRMS and demonstrates a progrowth role for RASSF4. Additionally, we identify a mechanism used by PAX3-FOXO1 to inhibit MST1 signaling and promote tumorigenesis in aRMS.Authors
Lisa E.S. Crose, Kathleen A. Galindo, Julie Grondin Kephart, Candy Chen, Julien Fitamant, Nabeel Bardeesy, Rex C. Bentley, Rene L. Galindo, Jen-Tsan Ashley Chi, Corinne M. Linardic
×Abstract
The cytokines RANKL and TNF activate NF-κB signaling in osteoclast precursors (OCPs) to induce osteoclast (OC) formation. Conversely, TNF can limit OC formation through NF-κB p100, which acts as an inhibitor, and TNF receptor–associated receptor 3 (TRAF3); however, a role for TRAF3 in RANKL-mediated OC formation is unknown. We found that TRAF3 limits RANKL-induced osteoclastogenesis by suppressing canonical and noncanonical NF-κB signaling. Conditional OC-specific
Traf3 -KO (cKO) mice had mild osteoporosis and increased OC formation. RANKL induced TRAF3 degradation via the lysosome/autophagy system. The autophagy/lysosome inhibitor chloroquine reduced RANKL-induced OC formation and function by increasing TRAF3 expression in OCPs in vitro and in vivo. Although chloroquine had no effect on basal bone resorption, it inhibited parathyroid hormone– and ovariectomy-induced OC activation in WT, but not cKO, mice. Deletion of the transcription factor geneRelb resulted in increased TRAF3 expression in OCPs, which was associated with decreased RANKL-induced TRAF3 degradation. RelB directly increased expression of BECN1, a key autophagy regulator, by binding to its promoter. These data indicate that autophagic/lysosomal degradation of TRAF3 is an important step in RANKL-induced NF-κB activation in OCPs. Furthermore, treatments that increase TRAF3 levels in OCPs, including pharmacological inhibition of its degradation with compounds such as chloroquine, may limit bone destruction in common bone diseases.Authors
Yan Xiu, Hao Xu, Chen Zhao, Jinbo Li, Yoshikazu Morita, Zhenqiang Yao, Lianping Xing, Brendan F. Boyce
×Abstract
Lyme disease, caused by the spirochete
Borrelia burgdorferi , is the most prevalent arthropod-borne illness in the United States and remains a clinical and social challenge. The spectrum of disease severity among infected patients suggests that host genetics contribute to pathogenic outcomes, particularly in patients who develop arthritis. Using a forward genetics approach, we identified the lysosomal enzyme β-glucuronidase (GUSB), a member of a large family of coregulated lysosomal enzymes, as a key regulator of Lyme-associated arthritis severity. Severely arthritic C3H mice possessed a naturally occurring hypomorphic allele,Gusbh . C57BL/6 mice congenic for the C3HGusb allele were prone to increased Lyme-associated arthritis severity. Radiation chimera experiments revealed that resident joint cells drive arthritis susceptibility. C3H mice expressing WTGusb as a transgene were protected from severe Lyme arthritis. Importantly, theGusbh allele also exacerbated disease in a serum transfer model of rheumatoid arthritis. A known GUSB function is the prevention of lysosomal accumulation of glycosaminoglycans (GAGs). Development of Lyme and rheumatoid arthritis inGusbh -expressing mice was associated with heightened accumulation of GAGs in joint tissue. We propose that GUSB modulates arthritis pathogenesis by preventing accumulation of proinflammatory GAGs within inflamed joint tissue, a trait that may be shared by other lysosomal exoglycosidases.Authors
Kenneth K.C. Bramwell, Ying Ma, John H. Weis, Xinjian Chen, James F. Zachary, Cory Teuscher, Janis J. Weis
×Abstract
Central congenital hypothyroidism (CCH) is more prevalent in children born to women with hyperthyroidism during pregnancy, suggesting a role for thyroid hormone (TH) in the development of central thyroid regulation. Using the zebrafish embryo as a model for thyroid axis development, we have characterized the ontogeny of negative feedback regulation of thyrotrope function and examined the effect of excess TH on thyrotrope development. We found that thyroid-stimulating hormone β subunit (
tshb ) and type 2 deiodinase (dio2 ) are coexpressed in zebrafish thyrotropes by 48 hours after fertilization and that TH-driven negative feedback regulation oftshb transcription appears in the thyroid axis by 96 hours after fertilization. Negative feedback regulation correlated with increased systemic TH levels from the developing thyroid follicles. We used a transgenic zebrafish that expresses GFP under the control of thetshb promoter to follow thyrotrope fates in vivo. Time-lapse imaging revealed that early exposure to elevated TH leads to thyrotrope cell death. Thyrotrope numbers slowly recovered following the removal of excess TH. These data demonstrate that transient TH exposure profoundly impacts the thyrotrope population during a critical period of pituitary development and may have long-term implications for the functional reserve of thyroid-stimulating hormone (TSH) production and the TSH set point later in life.Authors
Ksenia N. Tonyushkina, Meng-Chieh Shen, Theresa Ortiz-Toro, Rolf O. Karlstrom
×Abstract
Multiple intestinal atresia (MIA) is a rare cause of bowel obstruction that is sometimes associated with a combined immunodeficiency (CID), leading to increased susceptibility to infections. The factors underlying this rare disease are poorly understood. We characterized the immunological and intestinal features of 6 unrelated MIA-CID patients. All patients displayed a profound, generalized lymphocytopenia, with few lymphocytes present in the lymph nodes. The thymus was hypoplastic and exhibited an abnormal distribution of epithelial cells. Patients also had profound disruption of the epithelial barrier along the entire gastrointestinal tract. Using linkage analysis and whole-exome sequencing, we identified 10 mutations in tetratricopeptide repeat domain–7A (
TTC7A ), all of which potentially abrogateTTC7A expression. Intestinal organoid cultures from patient biopsies displayed an inversion of apicobasal polarity of the epithelial cells that was normalized by pharmacological inhibition of Rho kinase. Our data indicate that TTC7A deficiency results in increased Rho kinase activity, which disrupts polarity, growth, and differentiation of intestinal epithelial cells, and which impairs immune cell homeostasis, thereby promoting MIA-CID development.Authors
Amélie E. Bigorgne, Henner F. Farin, Roxane Lemoine, Nizar Mahlaoui, Nathalie Lambert, Marine Gil, Ansgar Schulz, Pierre Philippet, Patrick Schlesser, Tore G. Abrahamsen, Knut Oymar, E. Graham Davies, Christian Lycke Ellingsen, Emmanuelle Leteurtre, Brigitte Moreau-Massart, Dominique Berrebi, Christine Bole-Feysot, Patrick Nischke, Nicole Brousse, Alain Fischer, Hans Clevers, Geneviève de Saint Basile
×Abstract
Pancreatic ductal adenocarcinoma (PDAC) is often associated with overexpression of TGF-β. Given its tumor suppressor functions, it is unclear whether TGF-β is a valid therapeutic target for PDAC. Here, we found that proliferating pancreatic cancer cells (PCCs) from human PDAC patients and multiple murine models of PDAC (mPDAC) often exhibit abundant levels of phosphorylated retinoblastoma 1 (RB) and Smad2. TGF-β1 treatment enhanced proliferation of PCCs isolated from
KrasG12D -driven mPDAC that lacked RB (KRC cells). This mitogenic effect was abrogated by pharmacological inhibition of type I TGF-β receptor kinase, combined inhibition of MEK/Src or MEK/PI3K, and restoration of RB expression. TGF-β1 promoted epithelial-to-mesenchymal transition (EMT), invasion, Smad2/3 phosphorylation, Src activation, Wnt reporter activity, and Smad-dependent upregulation of Wnt7b in KRC cells. Importantly, TGF-β1–induced mitogenesis was markedly attenuated by inhibition of Wnt secretion. In an in vivo syngeneic orthotopic model, inhibition of TGF-β signaling suppressed KRC cell proliferation, tumor growth, stroma formation, EMT, metastasis, ascites formation, and Wnt7b expression, and markedly prolonged survival. Together, these data indicate that RB dysfunction converts TGF-β to a mitogen that activates known oncogenic signaling pathways and upregulates Wnt7b, which synergize to promote PCC invasion, survival, and mitogenesis. Furthermore, this study suggests that concomitantly targeting TGF-β and Wnt7b signaling in PDAC may disrupt these aberrant pathways, which warrants further evaluation in preclinical models.Authors
A. Jesse Gore, Samantha L. Deitz, Lakshmi Reddy Palam, Kelly E. Craven, Murray Korc
×Abstract
The shelterin complex plays dual functions in telomere homeostasis by recruiting telomerase and preventing the activation of a DNA damage response at telomeric ends. Somatic stem cells require telomerase activity, as evidenced by progressive stem cell loss leading to bone marrow failure in hereditary dyskeratosis congenita. Recent work demonstrates that dyskeratosis congenita can also arise from mutations in specific shelterin genes, although little is known about shelterin functions in somatic stem cells. We found that mouse hematopoietic stem cells (HSCs) are acutely sensitive to inactivation of the shelterin gene
Acd , encoding TPP1. Homozygosity for a hypomorphicacd allele preserved the emergence and expansion of fetal HSCs but led to profoundly defective function in transplantation assays. Upon completeAcd inactivation, HSCs expressed p53 target genes, underwent cell cycle arrest, and were severely depleted within days, leading to hematopoietic failure. TPP1 loss induced increased telomeric fusion events in bone marrow progenitors. However, unlike in epidermal stem cells,p53 deficiency did not rescue TPP1-deficient HSCs, indicating that shelterin dysfunction has unique effects in different stem cell populations. Because the consequences of telomere shortening are progressive and unsynchronized, acute loss of shelterin function represents an attractive alternative for studying telomere crisis in hematopoietic progenitors.Authors
Morgan Jones, Gail Osawa, Joshua A. Regal, Daniel N. Weinberg, James Taggart, Hande Kocak, Ann Friedman, David O. Ferguson, Catherine E. Keegan, Ivan Maillard
×Abstract
There is a considerable resurgence of interest in the role of aerobic glycolysis in cancer; however, increased glycolysis is frequently viewed as a consequence of oncogenic events that drive malignant cell growth and survival. Here we provide evidence that increased glycolytic activation itself can be an oncogenic event in a physiologically relevant 3D culture model. Overexpression of glucose transporter type 3 (GLUT3) in nonmalignant human breast cells activated known oncogenic signaling pathways, including EGFR, β1 integrin, MEK, and AKT, leading to loss of tissue polarity and increased growth. Conversely, reduction of glucose uptake in malignant cells promoted the formation of organized and growth-arrested structures with basal polarity, and suppressed oncogenic pathways. Unexpectedly and importantly, we found that unlike reported literature, in 3D the differences between “normal” and malignant phenotypes could not be explained by HIF-1α/2α, AMPK, or mTOR pathways. Loss of epithelial integrity involved activation of RAP1 via exchange protein directly activated by cAMP (EPAC), involving also O-linked N-acetylglucosamine modification downstream of the hexosamine biosynthetic pathway. The former, in turn, was mediated by pyruvate kinase M2 (PKM2) interaction with soluble adenylyl cyclase. Our findings show that increased glucose uptake activates known oncogenic pathways to induce malignant phenotype, and provide possible targets for diagnosis and therapeutics.
Authors
Yasuhito Onodera, Jin-Min Nam, Mina J. Bissell
×Abstract
Corneal integrity and transparency are indispensable for good vision. Cornea homeostasis is entirely dependent upon corneal stem cells, which are required for complex wound-healing processes that restore corneal integrity following epithelial damage. Here, we found that leucine-rich repeats and immunoglobulin-like domains 1 (
LRIG1 ) is highly expressed in the human holoclone-type corneal epithelial stem cell population and sporadically expressed in the basal cells of ocular-surface epithelium. In murine models, LRIG1 regulated corneal epithelial cell fate during wound repair. Deletion ofLrig1 resulted in impaired stem cell recruitment following injury and promoted a cell-fate switch from transparent epithelium to keratinized skin-like epidermis, which led to corneal blindness. In addition, we determined that LRIG1 is a negative regulator of the STAT3-dependent inflammatory pathway. Inhibition of STAT3 in corneas ofLrig1–/– mice rescued pathological phenotypes and prevented corneal opacity. Additionally, transgenic mice that expressed a constitutively active form of STAT3 in the corneal epithelium had abnormal features, including corneal plaques and neovascularization similar to that found inLrig1–/– mice. Bone marrow chimera experiments indicated that LRIG1 also coordinates the function of bone marrow–derived inflammatory cells. Together, our data indicate that LRIG1 orchestrates corneal-tissue transparency and cell fate during repair, and identify LRIG1 as a key regulator of tissue homeostasis.Authors
Takahiro Nakamura, Junji Hamuro, Mikiro Takaishi, Szandor Simmons, Kazuichi Maruyama, Andrea Zaffalon, Adam J. Bentley, Satoshi Kawasaki, Maho Nagata-Takaoka, Nigel J. Fullwood, Satoshi Itami, Shigetoshi Sano, Masaru Ishii, Yann Barrandon, Shigeru Kinoshita
×Abstract
Metabolic profiling of cancer cells has recently been established as a promising tool for the development of therapies and identification of cancer biomarkers. Here we characterized the metabolomic profile of human breast tumors and uncovered intrinsic metabolite signatures in these tumors using an untargeted discovery approach and validation of key metabolites. The oncometabolite 2-hydroxyglutarate (2HG) accumulated at high levels in a subset of tumors and human breast cancer cell lines. We discovered an association between increased 2HG levels and MYC pathway activation in breast cancer, and further corroborated this relationship using MYC overexpression and knockdown in human mammary epithelial and breast cancer cells. Further analyses revealed globally increased DNA methylation in 2HG-high tumors and identified a tumor subtype with high tissue 2HG and a distinct DNA methylation pattern that was associated with poor prognosis and occurred with higher frequency in African-American patients. Tumors of this subtype had a stem cell–like transcriptional signature and tended to overexpress glutaminase, suggestive of a functional relationship between glutamine and 2HG metabolism in breast cancer. Accordingly, 13C-labeled glutamine was incorporated into 2HG in cells with aberrant 2HG accumulation, whereas pharmacologic and siRNA-mediated glutaminase inhibition reduced 2HG levels. Our findings implicate 2HG as a candidate breast cancer oncometabolite associated with MYC activation and poor prognosis.
Authors
Atsushi Terunuma, Nagireddy Putluri, Prachi Mishra, Ewy A. Mathé, Tiffany H. Dorsey, Ming Yi, Tiffany A. Wallace, Haleem J. Issaq, Ming Zhou, J. Keith Killian, Holly S. Stevenson, Edward D. Karoly, King Chan, Susmita Samanta, DaRue Prieto, Tiffany Y.T. Hsu, Sarah J. Kurley, Vasanta Putluri, Rajni Sonavane, Daniel C. Edelman, Jacob Wulff, Adrienne M. Starks, Yinmeng Yang, Rick A. Kittles, Harry G. Yfantis, Dong H. Lee, Olga B. Ioffe, Rachel Schiff, Robert M. Stephens, Paul S. Meltzer, Timothy D. Veenstra, Thomas F. Westbrook, Arun Sreekumar, Stefan Ambs
×Abstract
How glucose sensing by the nervous system impacts the regulation of β cell mass and function during postnatal development and throughout adulthood is incompletely understood. Here, we studied mice with inactivation of glucose transporter 2 (
Glut2 ) in the nervous system (NG2KO mice). These mice displayed normal energy homeostasis but developed late-onset glucose intolerance due to reduced insulin secretion, which was precipitated by high-fat diet feeding. The β cell mass of adult NG2KO mice was reduced compared with that of WT mice due to lower β cell proliferation rates in NG2KO mice during the early postnatal period. The difference in proliferation between NG2KO and control islets was abolished by ganglionic blockade or by weaning the mice on a carbohydrate-free diet. In adult NG2KO mice, first-phase insulin secretion was lost, and these glucose-intolerant mice developed impaired glucagon secretion when fed a high-fat diet. Electrophysiological recordings showed reduced parasympathetic nerve activity in the basal state and no stimulation by glucose. Furthermore, sympathetic activity was also insensitive to glucose. Collectively, our data show that GLUT2-dependent control of parasympathetic activity defines a nervous system/endocrine pancreas axis that is critical for β cell mass establishment in the postnatal period and for long-term maintenance of β cell function.Authors
David Tarussio, Salima Metref, Pascal Seyer, Lourdes Mounien, David Vallois, Christophe Magnan, Marc Foretz, Bernard Thorens
×Abstract
Studies have established that pigmentation can provide strong, protective effects against certain human diseases. For example, angiogenesis-dependent diseases such as wet age-related macular degeneration and infantile hemangioma are more common in light-skinned individuals of mixed European descent than in African-Americans. Here we found that melanocytes from light-skinned humans and albino mice secrete high levels of fibromodulin (FMOD), which we determined to be a potent angiogenic factor. FMOD treatment stimulated angiogenesis in numerous in vivo systems, including laser-induced choroidal neovascularization, growth factor–induced corneal neovascularization, wound healing, and Matrigel plug assays. Additionally, FMOD enhanced vascular sprouting during normal retinal development. Deletion of
Fmod in albino mice resulted in a marked reduction in the amount of neovascularization induced by retinal vein occlusion, corneal growth factor pellets, and Matrigel plugs. Our data implicate the melanocyte-secreted factor FMOD as a key regulator of angiogenesis and suggest an underlying mechanism for epidemiological differences between light-skinned individuals of mixed European descent and African-Americans. Furthermore, inhibition of FMOD in humans has potential as a therapeutic strategy for treating angiogenesis-dependent diseases.Authors
Irit Adini, Kaustabh Ghosh, Avner Adini, Zai-Long Chi, Takeru Yoshimura, Ofra Benny, Kip M. Connor, Michael S. Rogers, Lauren Bazinet, Amy E. Birsner, Diane R. Bielenberg, Robert J. D’Amato
×Abstract
The protective role of hemeoxygenase-1 (HO-1) in various inflammatory conditions is mediated in part by its products, carbon monoxide (CO) and biliverdin. Here we investigated a therapeutic role for CO and CO-primed cells in acute pancreatitis (AP). In a mouse model of AP, treatment with CO-releasing molecule–2 (CORM-2) decreased mortality, pancreatic damage, and lung injury. CORM-2 decreased systemic inflammatory cytokines, suppressed systemic and pancreatic macrophage TNF-α secretion, and inhibited macrophage TLR4 receptor complex expression. In both human and mouse cells, CORM-2 inhibited endogenous and exogenous ligand-dependent TLR4 activation, which indicates that CORM-2 could be therapeutic for both early and late stages of AP, which involve sterile- and endotoxin-mediated inflammation, respectively. Mice engrafted with TLR4-deficient hematopoietic cells were protected against caerulein-induced AP. In the absence of leukocyte TLR4 expression, CORM-2 did not confer additional protection, which indicates that CORM-2–dependent effects are mediated via suppression of macrophage TLR4 activation. We determined that CO was directly responsible for the protective effects of CORM-2 in AP, as inactive forms of CORM-2 were ineffective. Importantly, adoptive transfer of CORM-2–primed cells reduced AP. Such a therapeutic approach would translate the beneficial effects of CO-based therapies, avoiding CO- or CO-RM–mediated toxicities in AP and a wide range of diseases.
Authors
Jing Xue, Aida Habtezion
×Abstract
Loeys-Dietz syndrome (LDS) is a connective tissue disorder that is characterized by a high risk for aneurysm and dissection throughout the arterial tree and phenotypically resembles Marfan syndrome. LDS is caused by heterozygous missense mutations in either TGF-β receptor gene (
TGFBR1 orTGFBR2 ), which are predicted to result in diminished TGF-β signaling; however, aortic surgical samples from patients show evidence of paradoxically increased TGF-β signaling. We generated 2 knockin mouse strains with LDS mutations in eitherTgfbr1 orTgfbr2 and a transgenic mouse overexpressing mutantTgfbr2 . Knockin and transgenic mice, but not haploinsufficient animals, recapitulated the LDS phenotype. While heterozygous mutant cells had diminished signaling in response to exogenous TGF-β in vitro, they maintained normal levels of Smad2 phosphorylation under steady-state culture conditions, suggesting a chronic compensation. Analysis of TGF-β signaling in the aortic wall in vivo revealed progressive upregulation of Smad2 phosphorylation and TGF-β target gene output, which paralleled worsening of aneurysm pathology and coincided with upregulation of TGF-β1 ligand expression. Importantly, suppression of Smad2 phosphorylation and TGF-β1 expression correlated with the therapeutic efficacy of the angiotensin II type 1 receptor antagonist losartan. Together, these data suggest that increased TGF-β signaling contributes to postnatal aneurysm progression in LDS.Authors
Elena M. Gallo, David C. Loch, Jennifer P. Habashi, Juan F. Calderon, Yichun Chen, Djahida Bedja, Christel van Erp, Elizabeth E. Gerber, Sarah J. Parker, Kimberly Sauls, Daniel P. Judge, Sara K. Cooke, Mark E. Lindsay, Rosanne Rouf, Loretha Myers, Colette M. ap Rhys, Kathleen C. Kent, Russell A. Norris, David L. Huso, Harry C. Dietz
Copyright © 2025 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)