In this issue of the JCI, Edelmann et al. reveal that a JAK2-V617F mutation that is common in patients with polycythemia vera and essential thrombocytosis enhances risk of developing life-threatening thromboses by increasing β1 and β2 integrin affinity for the endothelial adhesion molecules VCAM1 and ICAM2. This aberrant interaction underlies also altered myeloid trafficking to the spleen. The cover image shows the expression of endothelial VCAM1 (white) and ICAM1 (purple) together with myeloid-derived cells (yellow) and other immune cell populations in a mouse spleen. Image credit: Lars Philipsen.
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Corinne L. Williams
Serpil C. Erzurum
Benjamin L. Ebert
Helen H. Hobbs
At implantation, the embryo expresses paternally derived alloantigens and evokes inflammation that can threaten reproductive success. To ensure a robust placenta and sustainable pregnancy, an active state of maternal immune tolerance mediated by CD4+ regulatory T cells (Tregs) is essential. Tregs operate to inhibit effector immunity, contain inflammation, and support maternal vascular adaptations, thereby facilitating trophoblast invasion and placental access to the maternal blood supply. Insufficient Treg numbers or inadequate functional competence are implicated in idiopathic infertility and recurrent miscarriage as well as later-onset pregnancy complications stemming from placental insufficiency, including preeclampsia and fetal growth restriction. In this Review, we summarize the mechanisms acting in the conception environment to drive the Treg response and discuss prospects for targeting the T cell compartment to alleviate immune-based reproductive disorders.
Sarah A. Robertson, Alison S. Care, Lachlan M. Moldenhauer
A complex DNA repair machinery has evolved to protect genomic integrity in the face of a myriad of DNA damage sources. When DNA repair fails, this damage can lead to carcinogenesis and tumor genomic instability. Indeed, many heritable cancer predisposition syndromes are attributable to germline defects in DNA repair pathways. On the other hand, these defects may also portend particular vulnerabilities of the cancer and may be exploited therapeutically. Most recently this has been demonstrated in the case of mismatch repair-deficient cancers, in which the immune checkpoint inhibitors have been demonstrated to be highly active. This observation has paved the way for further research investigating other sources of genomic instability that may serve as biomarkers to select patients for immunotherapy.
Katherine M. Bever, Dung T. Le
About one-third of the US population will develop herpes zoster (HZ, commonly known as shingles) over a lifetime, while two-thirds will not. It is not clear exactly why certain people are susceptible to HZ; however, we may be coming closer to an answer. In this issue of the JCI, a study by Levin et al. provides important details concerning pathogenesis of and protection from HZ. The authors characterized differences in the immunologic responses induced by two HZ vaccines, the live attenuated zoster vaccine (ZV) and the more recently developed adjuvanted varicella-zoster virus (VZV) glycoprotein E (gE) subunit herpes zoster vaccine (HZ/su), in vaccine-naive subjects and those previously vaccinated with HZ. The observed differences in responses paralleled the observed clinical protection of the two zoster vaccines, with HZ/su being superior to HZ. Together, these results seem to explain immunologically why the new subunit vaccine outperforms the live vaccine. These differences may also provide clues as to why HZ develops in the first place.
Anne A. Gershon
Stephen T. Oh
In spite of a very robust body of literature and definitive data demonstrating the importance of the programmed cell death receptor-1 (PD-1) pathway in T cells and their function, the data on NK cell PD-1 expression have been highly variable and, particularly in the case of mouse NK cells, scarce. In this issue of the JCI, Hsu et al. present data demonstrating PD-1 expression on mouse NK cells only within tumors and show that PD-1 blockade elicits an antitumor NK cell–mediated response. This study indicates that, given the complexity of both the biology and study of NK cells, further work is needed to more clearly determine the role of the PD-1/PD-1 ligand (PD-L1) on NK cells.
Cordelia Dunai, William J. Murphy
The polyamine metabolic pathway has been considered a rational target for antineoplastic therapy since it was discovered that polyamines are absolute requirements for tumor initiation, growth, and, in some instances, survival. Although several promising preclinical studies have demonstrated the critical nature of polyamines for tumor growth, the clinical success of agents targeting polyamine metabolism have been lacking. In the accompanying article, Bianchi-Smiraglia et al. identify both a new target and new drug that inhibits polyamine biosynthesis, reduces intracellular polyamines, and inhibits the growth of several models of human multiple myeloma. These results are both intriguing and provide promise for moving such a strategy to the clinic.
Robert A. Casero Jr.
Hepatitis B virus–specific (HBV-specific) T cells have been identified as main effector cells in HBV clearance. In contrast, B cells producing neutralizing antibodies against the HBV surface antigen (HBsAg) have been studied in little detail, mainly due to methodical limitations. In this issue of the JCI, two reports use a new technique to specifically detect and characterize HBsAg-specific B cells ex vivo. Indeed, these cells are present, but show phenotypic alterations and impaired function during acute and chronic HBV infection. Thus, HBsAg-specific B cells are a novel attractive target for antiviral strategies toward functional cure of chronic HBV infection.
Christoph Neumann-Haefelin, Robert Thimme
Nucleophosmin (NPM1) is among the most frequently mutated genes in acute myeloid leukemia (AML). It is not known, however, how the resulting oncoprotein mutant NPM1 is leukemogenic. To reveal the cellular machinery in which NPM1 participates in myeloid cells, we analyzed the endogenous NPM1 protein interactome by mass spectrometry and discovered abundant amounts of the master transcription factor driver of monocyte lineage differentiation PU.1 (also known as SPI1). Mutant NPM1, which aberrantly accumulates in cytoplasm, dislocated PU.1 into cytoplasm with it. CEBPA and RUNX1, the master transcription factors that collaborate with PU.1 to activate granulomonocytic lineage fates, remained nuclear; but without PU.1, their coregulator interactions were toggled from coactivators to corepressors, repressing instead of activating more than 500 granulocyte and monocyte terminal differentiation genes. An inhibitor of nuclear export, selinexor, by locking mutant NPM1/PU.1 in the nucleus, activated terminal monocytic fates. Direct depletion of the corepressor DNA methyltransferase 1 (DNMT1) from the CEBPA/RUNX1 protein interactome using the clinical drug decitabine activated terminal granulocytic fates. Together, these noncytotoxic treatments extended survival by more than 160 days versus vehicle in a patient-derived xenotransplant model of NPM1/FLT3-mutated AML. In sum, mutant NPM1 represses monocyte and granulocyte terminal differentiation by disrupting PU.1/CEBPA/RUNX1 collaboration, a transforming action that can be reversed by pharmacodynamically directed dosing of clinical small molecules.
Xiaorong Gu, Quteba Ebrahem, Reda Z. Mahfouz, Metis Hasipek, Francis Enane, Tomas Radivoyevitch, Nicolas Rapin, Bartlomiej Przychodzen, Zhenbo Hu, Ramesh Balusu, Claudiu V. Cotta, David Wald, Christian Argueta, Yosef Landesman, Maria Paola Martelli, Brunangelo Falini, Hetty Carraway, Bo T. Porse, Jaroslaw Maciejewski, Babal K. Jha, Yogen Saunthararajah
Central to the recognition, signaling, and repair of DNA double-strand breaks (DSBs) are the MRE11-RAD50-NBS1 (MRN) complex and mediator of DNA damage checkpoint protein 1 (MDC1), the interplay of which is essential for initiation and amplification of the DNA damage response (DDR). The intrinsic rule governing the regulation of the function of this molecular machinery remains to be investigated. We report here that the ubiquitin-specific protease USP7 was physically associated with the MRN-MDC1 complex and that the MRN-MDC1 complex acted as a platform for USP7 to efficiently deubiquitinate and stabilize MDC1, thereby sustaining the DDR. Accordingly, depletion of USP7 impaired the engagement of the MRN-MDC1 complex and the consequent recruitment of the downstream factors p53-binding protein 1 (53BP1) and breast cancer protein 1 (BRCA1) at DNA lesions. Significantly, USP7 was overexpressed in cervical cancer, and the level of its expression positively correlated with that of MDC1 and worse survival rates for patients with cervical cancer. We demonstrate that USP7-mediated MDC1 stabilization promoted cervical cancer cell survival and conferred cellular resistance to genotoxic insults. Together, our study reveals a role for USP7 in regulating the function of the MRN-MDC1 complex and activity of the DDR, supporting the pursuit of USP7 as a potential therapeutic target for MDC1-proficient cancers.
Dongxue Su, Shuai Ma, Lin Shan, Yue Wang, Yuejiao Wang, Cheng Cao, Beibei Liu, Chao Yang, Liyong Wang, Shanshan Tian, Xiang Ding, Xinhua Liu, Na Yu, Nan Song, Ling Liu, Shangda Yang, Qi Zhang, Fuquan Yang, Kai Zhang, Lei Shi
Induction of TLR2 activation depends on its association with the adapter protein MyD88. We have found that TLR2 and MyD88 levels are elevated in the hippocampus and cortex of patients with Alzheimer’s disease (AD) and in a 5XFAD mouse model of AD. Since there is no specific inhibitor of TLR2, to target induced TLR2 from a therapeutic angle, we engineered a peptide corresponding to the TLR2-interacting domain of MyD88 (TIDM) that binds to the BB loop of only TLR2, and not other TLRs. Interestingly, WT TIDM peptide inhibited microglial activation induced by fibrillar Aβ1-42 and lipoteichoic acid, but not 1-methyl-4-phenylpyridinium, dsRNA, bacterial lipopolysaccharide, flagellin, or CpG DNA. After intranasal administration, WT TIDM peptide reached the hippocampus, reduced hippocampal glial activation, lowered Aβ burden, attenuated neuronal apoptosis, and improved memory and learning in 5XFAD mice. However, WT TIDM peptide was not effective in 5XFAD mice lacking TLR2. In addition to its effects in 5XFAD mice, WT TIDM peptide also suppressed the disease process in mice with experimental allergic encephalomyelitis and collagen-induced arthritis. Therefore, selective targeting of the activated status of 1 component of the innate immune system by WT TIDM peptide may be beneficial in AD as well as other disorders in which TLR2/MyD88 signaling plays a role in disease pathogenesis.
Suresh B. Rangasamy, Malabendu Jana, Avik Roy, Grant T. Corbett, Madhuchhanda Kundu, Sujyoti Chandra, Susanta Mondal, Sridevi Dasarathi, Elliott J. Mufson, Rama K. Mishra, Chi-Hao Luan, David A. Bennett, Kalipada Pahan
Steroid-resistant nephrotic syndrome (SRNS) almost invariably progresses to end-stage renal disease. Although more than 50 monogenic causes of SRNS have been described, a large proportion of SRNS remains unexplained. Recently, it was discovered that mutations of NUP93 and NUP205, encoding 2 proteins of the inner ring subunit of the nuclear pore complex (NPC), cause SRNS. Here, we describe mutations in genes encoding 4 components of the outer rings of the NPC, namely NUP107, NUP85, NUP133, and NUP160, in 13 families with SRNS. Using coimmunoprecipitation experiments, we showed that certain pathogenic alleles weakened the interaction between neighboring NPC subunits. We demonstrated that morpholino knockdown of nup107, nup85, or nup133 in Xenopus disrupted glomerulogenesis. Re-expression of WT mRNA, but not of mRNA reflecting mutations from SRNS patients, mitigated this phenotype. We furthermore found that CRISPR/Cas9 knockout of NUP107, NUP85, or NUP133 in podocytes activated Cdc42, an important effector of SRNS pathogenesis. CRISPR/Cas9 knockout of nup107 or nup85 in zebrafish caused developmental anomalies and early lethality. In contrast, an in-frame mutation of nup107 did not affect survival, thus mimicking the allelic effects seen in humans. In conclusion, we discovered here that mutations in 4 genes encoding components of the outer ring subunits of the NPC cause SRNS and thereby provide further evidence that specific hypomorphic mutations in these essential genes cause a distinct, organ-specific phenotype.
Daniela A. Braun, Svjetlana Lovric, David Schapiro, Ronen Schneider, Jonathan Marquez, Maria Asif, Muhammad Sajid Hussain, Ankana Daga, Eugen Widmeier, Jia Rao, Shazia Ashraf, Weizhen Tan, C. Patrick Lusk, Amy Kolb, Tilman Jobst-Schwan, Johanna Magdalena Schmidt, Charlotte A. Hoogstraten, Kaitlyn Eddy, Thomas M. Kitzler, Shirlee Shril, Abubakar Moawia, Kathrin Schrage, Arwa Ishaq A. Khayyat, Jennifer A. Lawson, Heon Yung Gee, Jillian K. Warejko, Tobias Hermle, Amar J. Majmundar, Hannah Hugo, Birgit Budde, Susanne Motameny, Janine Altmüller, Angelika Anna Noegel, Hanan M. Fathy, Daniel P. Gale, Syeda Seema Waseem, Ayaz Khan, Larissa Kerecuk, Seema Hashmi, Nilufar Mohebbi, Robert Ettenger, Erkin Serdaroğlu, Khalid A. Alhasan, Mais Hashem, Sara Goncalves, Gema Ariceta, Mercedes Ubetagoyena, Wolfram Antonin, Shahid Mahmood Baig, Fowzan S. Alkuraya, Qian Shen, Hong Xu, Corinne Antignac, Richard P. Lifton, Shrikant Mane, Peter Nürnberg, Mustafa K. Khokha, Friedhelm Hildebrandt
Movement of circulating fatty acids (FAs) to parenchymal cells requires their transfer across the endothelial cell (EC) barrier. The multiligand receptor cluster of differentiation 36 (CD36) facilitates tissue FA uptake and is expressed in ECs and parenchymal cells such as myocytes and adipocytes. Whether tissue uptake of FAs is dependent on EC or parenchymal cell CD36, or both, is unknown. Using a cell-specific deletion approach, we show that EC, but not parenchymal cell, CD36 deletion increased fasting plasma FAs and postprandial triglycerides. EC-Cd36–KO mice had reduced uptake of radiolabeled long-chain FAs into heart, skeletal muscle, and brown adipose tissue; these uptake studies were replicated using [11C]palmitate PET scans. High-fat diet–fed EC-CD36–deficient mice had improved glucose tolerance and insulin sensitivity. Both EC and cardiomyocyte (CM) deletion of CD36 reduced heart lipid droplet accumulation after fasting, but CM deletion did not affect heart glucose or FA uptake. Expression in the heart of several genes modulating glucose metabolism and insulin action increased with EC-CD36 deletion but decreased with CM deletion. In conclusion, EC CD36 acts as a gatekeeper for parenchymal cell FA uptake, with important downstream effects on glucose utilization and insulin action.
Ni-Huiping Son, Debapriya Basu, Dmitri Samovski, Terri A. Pietka, Vivek S. Peche, Florian Willecke, Xiang Fang, Shui-Qing Yu, Diego Scerbo, Hye Rim Chang, Fei Sun, Svetlana Bagdasarov, Konstantinos Drosatos, Steve T. Yeh, Adam E. Mullick, Kooresh I. Shoghi, Namrata Gumaste, KyeongJin Kim, Lesley-Ann Huggins, Tenzin Lhakhang, Nada A. Abumrad, Ira J. Goldberg
GWAS have repeatedly mapped susceptibility loci for emphysema to genes that modify hedgehog signaling, but the functional relevance of hedgehog signaling to this morbid disease remains unclear. In the current study, we identified a broad population of mesenchymal cells in the adult murine lung receptive to hedgehog signaling, characterized by higher activation of hedgehog surrounding the proximal airway relative to the distal alveoli. Single-cell RNA-sequencing showed that the hedgehog-receptive mesenchyme is composed of mostly fibroblasts with distinct proximal and distal subsets with discrete identities. Ectopic hedgehog activation in the distal fibroblasts promoted expression of proximal fibroblast markers and loss of distal alveoli and airspace enlargement of over 20% compared with controls. We found that hedgehog suppressed mesenchymal-derived mitogens enriched in distal fibroblasts that regulate alveolar stem cell regeneration and airspace size. Finally, single-cell analysis of the human lung mesenchyme showed that segregated proximal-distal identity with preferential hedgehog activation in the proximal fibroblasts was conserved between mice and humans. In conclusion, we showed that differential hedgehog activation segregates mesenchymal identities of distinct fibroblast subsets and that disruption of fibroblast identity can alter the alveolar stem cell niche, leading to emphysematous changes in the murine lung.
Chaoqun Wang, Nabora S. Reyes de Mochel, Stephanie A. Christenson, Monica Cassandras, Rebecca Moon, Alexis N. Brumwell, Lauren E. Byrnes, Alfred Li, Yasuyuki Yokosaki, Peiying Shan, Julie B. Sneddon, David Jablons, Patty J. Lee, Michael A. Matthay, Harold A. Chapman, Tien Peng
JAK2-V617F–positive chronic myeloproliferative neoplasia (CMN) commonly displays dysfunction of integrins and adhesion molecules expressed on platelets, erythrocytes, and leukocytes. However, the mechanism by which the 2 major leukocyte integrin chains, β1 and β2, may contribute to CMN pathophysiology remained unclear. β1 (α4β1; VLA-4) and β2 (αLβ2; LFA-1) integrins are essential regulators for attachment of leukocytes to endothelial cells. We here showed enhanced adhesion of granulocytes from mice with JAK2-V617F knockin (JAK2+/VF mice) to vascular cell adhesion molecule 1– (VCAM1-) and intercellular adhesion molecule 1–coated (ICAM1-coated) surfaces. Soluble VCAM1 and ICAM1 ligand binding assays revealed increased affinity of β1 and β2 integrins for their respective ligands. For β1 integrins, this correlated with a structural change from the low- to the high-affinity conformation induced by JAK2-V617F. JAK2-V617F triggered constitutive activation of the integrin inside-out signaling molecule Rap1, resulting in translocation toward the cell membrane. Employing a venous thrombosis model, we demonstrated that neutralizing anti–VLA-4 and anti–β2 integrin antibodies suppress pathologic thrombosis as observed in JAK2+/VF mice. In addition, aberrant homing of JAK2+/VF leukocytes to the spleen was inhibited by neutralizing anti-β2 antibodies and by pharmacologic inhibition of Rap1. Thus, our findings identified cross-talk between JAK2-V617F and integrin activation promoting pathologic thrombosis and abnormal trafficking of leukocytes to the spleen.
Bärbel Edelmann, Nibedita Gupta, Tina M. Schnoeder, Anja M. Oelschlegel, Khurrum Shahzad, Jürgen Goldschmidt, Lars Philipsen, Soenke Weinert, Aniket Ghosh, Felix C. Saalfeld, Subbaiah Chary Nimmagadda, Peter Müller, Rüdiger Braun-Dullaeus, Juliane Mohr, Denise Wolleschak, Stefanie Kliche, Holger Amthauer, Florian H. Heidel, Burkhart Schraven, Berend Isermann, Andreas J. Müller, Thomas Fischer
BACKGROUND. Intravenous Ig (IVIg), plasma exchange, and immunoadsorption are frequently used in the management of severe autoimmune diseases mediated by pathogenic IgG autoantibodies. These approaches modulating IgG levels can, however, be associated with some severe adverse reactions and a substantial burden to patients. Targeting the neonatal Fc receptor (FcRn) presents an innovative and potentially more effective, safer, and more convenient alternative for clearing pathogenic IgGs. METHODS. A randomized, double-blind, placebo-controlled first-in-human study was conducted in 62 healthy volunteers to explore single and multiple ascending intravenous doses of the FcRn antagonist efgartigimod. The study objectives were to assess safety, tolerability, pharmacokinetics, pharmacodynamics, and immunogenicity. The findings of this study were compared with the pharmacodynamics profile elicited by efgartigimod in cynomolgus monkeys. RESULTS. Efgartigimod treatment resulted in a rapid and specific clearance of serum IgG levels in both cynomolgus monkeys and healthy volunteers. In humans, single administration of efgartigimod reduced IgG levels up to 50%, while multiple dosing further lowered IgGs on average by 75% of baseline levels. Approximately 8 weeks following the last administration, IgG levels returned to baseline. Efgartigimod did not alter the homeostasis of albumin or Igs other than IgG, and no serious adverse events related to efgartigimod infusion were observed. CONCLUSION. Antagonizing FcRn using efgartigimod is safe and results in a specific, profound, and sustained reduction of serum IgG levels. These results warrant further evaluation of this therapeutic approach in IgG-driven autoimmune diseases. TRIAL REGISTRATION. Clinicaltrials.gov NCT03457649. FUNDING. argenx BVBA.
Peter Ulrichts, Antonio Guglietta, Torsten Dreier, Tonke van Bragt, Valérie Hanssens, Erik Hofman, Bernhardt Vankerckhoven, Peter Verheesen, Nicolas Ongenae, Valentina Lykhopiy, F. Javier Enriquez, JunHaeng Cho, Raimund J. Ober, E. Sally Ward, Hans de Haard, Nicolas Leupin
Activation of HIV-1 reservoirs and induction of anti–HIV-1 T cells are critical to control HIV-1 rebound after combined antiretroviral therapy (cART). Here we evaluated in humanized mice (hu-mice) with persistent HIV-1 infection the therapeutic effect of TLR3 agonist and a CD40-targeting HIV-1 vaccine, which consists of a string of 5 highly conserved CD4+ and CD8+ T cell epitope-rich regions of HIV-1 Gag, Nef, and Pol fused to the C-terminus of a recombinant anti-human CD40 antibody (αCD40.HIV5pep). We show that αCD40.HIV5pep vaccination coadministered with poly(I:C) adjuvant induced HIV-1–specific human CD8+ and CD4+ T cell responses in hu-mice. Interestingly, poly(I:C) treatment also reactivated HIV-1 reservoirs. When administrated in therapeutic settings in HIV-1–infected hu-mice under effective cART, αCD40.HIV5pep with poly(I:C) vaccination induced HIV-1–specific CD8+ T cells and reduced the level of cell-associated HIV-1 DNA (or HIV-1 reservoirs) in lymphoid tissues. Most strikingly, the vaccination significantly delayed HIV-1 rebound after cART cessation. In summary, the αCD40.HIV5pep with poly(I:C) vaccination approach both activates replication of HIV-1 reservoirs and enhances the anti–HIV-1 T cell response, leading to a reduced level of cell-associated HIV-1 DNA or reservoirs. Our proof-of-concept study has significant implication for the development of CD40-targeting HIV-1 vaccine to enhance anti–HIV-1 immunity and reduce HIV-1 reservoirs in patients with suppressive cART.
Liang Cheng, Qi Wang, Guangming Li, Riddhima Banga, Jianping Ma, Haisheng Yu, Fumihiko Yasui, Zheng Zhang, Giuseppe Pantaleo, Matthieu Perreau, Sandra Zurawski, Gerard Zurawski, Yves Levy, Lishan Su
The paracaspase MALT1 plays an essential role in activated B cell–like diffuse large B cell lymphoma (ABC DLBCL) downstream of B cell and TLR pathway genes mutated in these tumors. Although MALT1 is considered a compelling therapeutic target, the development of tractable and specific MALT1 protease inhibitors has thus far been elusive. Here, we developed a target engagement assay that provides a quantitative readout for specific MALT1-inhibitory effects in living cells. This enabled a structure-guided medicinal chemistry effort culminating in the discovery of pharmacologically tractable, irreversible substrate-mimetic compounds that bind the MALT1 active site. We confirmed that MALT1 targeting with compound 3 is effective at suppressing ABC DLBCL cells in vitro and in vivo. We show that a reduction in serum IL-10 levels exquisitely correlates with the drug pharmacokinetics and degree of MALT1 inhibition in vitro and in vivo and could constitute a useful pharmacodynamic biomarker to evaluate these compounds in clinical trials. Compound 3 revealed insights into the biology of MALT1 in ABC DLBCL, such as the role of MALT1 in driving JAK/STAT signaling and suppressing the type I IFN response and MHC class II expression, suggesting that MALT1 inhibition could prime lymphomas for immune recognition by cytotoxic immune cells.
Lorena Fontán, Qi Qiao, John M. Hatcher, Gabriella Casalena, Ilkay Us, Matt Teater, Matt Durant, Guangyan Du, Min Xia, Natalia Bilchuk, Spandan Chennamadhavuni, Giuseppe Palladino, Giorgio Inghirami, Ulrike Philippar, Hao Wu, David A. Scott, Nathanael S. Gray, Ari Melnick
Tumor relapse is the leading cause of death in breast cancer, largely due to the fact that recurrent tumors are frequently resistant to chemotherapy. We previously reported that downregulation of the proapoptotic protein Par-4 promotes tumor recurrence in genetically engineered mouse models of breast cancer recurrence. In the present study, we examined the mechanism and functional significance of Par-4 downregulation in recurrent tumors. We found that epithelial-to-mesenchymal transition (EMT) promotes epigenetic silencing of Par-4 in recurrent tumors. Par-4 silencing proceeded through binding of the EMT transcription factor Twist to the Par-4 promoter, where Twist induced a unique bivalent chromatin domain. This bivalent configuration conferred plasticity at the Par-4 promoter, and Par-4 silencing could be reversed with pharmacologic inhibitors of Ezh2 and HDAC1/2. Using an epigenome editing approach to reexpress Par-4 by specifically reversing the histone modifications found in recurrent tumors, we found that Par-4 reexpression sensitized recurrent tumors to chemotherapy in vitro and in vivo. Upon reexpression, Par-4 bound to the protein phosphatase PP1, caused widespread changes in phosphorylation of cytoskeletal proteins, and cooperated with microtubule-targeting drugs to induce mitotic defects. These results identify Twist-induced epigenetic silencing of Par-4 as a targetable axis that promotes chemoresistance in recurrent breast cancer.
Nathaniel W. Mabe, Douglas B. Fox, Ryan Lupo, Amy E. Decker, Stephanie N. Phelps, J. Will Thompson, James V. Alvarez
The adjuvanted varicella-zoster virus (VZV) glycoprotein E (gE) subunit herpes zoster vaccine (HZ/su) confers higher protection against HZ than the live attenuated zoster vaccine (ZV). To understand the immunologic basis for the different efficacies of the vaccines, we compared immune responses to the vaccines in adults 50 to 85 years old. gE-specific T cells were very low/undetectable before vaccination when analyzed by FluoroSpot and flow cytometry. Both ZV and HZ/su increased gE-specific responses, but at peak memory response (PMR) after vaccination (30 days after ZV or after the second dose of HZ/su), gE-specific CD4+ and CD8+ T cell responses were 10-fold or more higher in HZ/su compared with ZV recipients. Comparing the vaccines, T cell memory responses, including gE–IL-2+ and VZV–IL-2+ spot-forming cells (SFCs), were higher in HZ/su recipients and cytotoxic and effector responses were lower. At 1 year after vaccination, all gE-Th1 and VZV–IL-2+ SFCs remained higher in HZ/su compared with ZV recipients. Mediation analyses showed that IL-2+ PMR were necessary for the persistence of Th1 responses to either vaccine and VZV–IL-2+ PMR explained 73% of the total effect of HZ/su on persistence. This emphasizes the biological importance of the memory responses, which were clearly superior in HZ/su compared with ZV participants.
Myron J. Levin, Miranda E. Kroehl, Michael J. Johnson, Andrew Hammes, Dominik Reinhold, Nancy Lang, Adriana Weinberg
BACKGROUND. Understanding the integrated immunogenomic landscape of advanced prostate cancer (APC) could impact stratified treatment selection. METHODS. Defective mismatch repair (dMMR) status was determined by either loss of mismatch repair protein expression on IHC or microsatellite instability (MSI) by PCR in 127 APC biopsies from 124 patients (Royal Marsden [RMH] cohort); MSI by targeted panel next-generation sequencing (MSINGS) was then evaluated in the same cohort and in 254 APC samples from the Stand Up To Cancer/Prostate Cancer Foundation (SU2C/PCF). Whole exome sequencing (WES) data from this latter cohort were analyzed for pathogenic MMR gene variants, mutational load, and mutational signatures. Transcriptomic data, available for 168 samples, was also performed. RESULTS. Overall, 8.1% of patients in the RMH cohort had some evidence of dMMR, which associated with decreased overall survival. Higher MSINGS scores associated with dMMR, and these APCs were enriched for higher T cell infiltration and PD-L1 protein expression. Exome MSINGS scores strongly correlated with targeted panel MSINGS scores (r = 0.73, P < 0.0001), and higher MSINGS scores associated with dMMR mutational signatures in APC exomes. dMMR mutational signatures also associated with MMR gene mutations and increased immune cell, immune checkpoint, and T cell–associated transcripts. APC with dMMR mutational signatures overexpressed a variety of immune transcripts, including CD200R1, BTLA, PD-L1, PD-L2, ADORA2A, PIK3CG, and TIGIT. CONCLUSION. These data could impact immune target selection, combination therapeutic strategy selection, and selection of predictive biomarkers for immunotherapy in APC. FUNDING. We acknowledge funding support from Movember, Prostate Cancer UK, The Prostate Cancer Foundation, SU2C, and Cancer Research UK.
Daniel Nava Rodrigues, Pasquale Rescigno, David Liu, Wei Yuan, Suzanne Carreira, Maryou B. Lambros, George Seed, Joaquin Mateo, Ruth Riisnaes, Stephanie Mullane, Claire Margolis, Diana Miao, Susana Miranda, David Dolling, Matthew Clarke, Claudia Bertan, Mateus Crespo, Gunther Boysen, Ana Ferreira, Adam Sharp, Ines Figueiredo, Daniel Keliher, Saud Aldubayan, Kelly P. Burke, Semini Sumanasuriya, Mariane Sousa Fontes, Diletta Bianchini, Zafeiris Zafeiriou, Larissa Sena Teixeira Mendes, Kent Mouw, Michael T. Schweizer, Colin C. Pritchard, Stephen Salipante, Mary-Ellen Taplin, Himisha Beltran, Mark A. Rubin, Marcin Cieslik, Dan Robinson, Elizabeth Heath, Nikolaus Schultz, Joshua Armenia, Wassim Abida, Howard Scher, Christopher Lord, Alan D’Andrea, Charles L. Sawyers, Arul M. Chinnaiyan, Andrea Alimonti, Peter S. Nelson, Charles G. Drake, Eliezer M. Van Allen, Johann S. de Bono
The glucocorticoid receptor (GR) is a major drug target in inflammatory disease. However, chronic glucocorticoid (GC) treatment leads to disordered energy metabolism, including increased weight gain, adiposity, and hepatosteatosis — all programs modulated by the circadian clock. We demonstrated that while antiinflammatory GC actions were maintained irrespective of dosing time, the liver was significantly more GC sensitive during the day. Temporal segregation of GC action was underpinned by a physical interaction of GR with the circadian transcription factor REVERBa and co-binding with liver-specific hepatocyte nuclear transcription factors (HNFs) on chromatin. REVERBa promoted efficient GR recruitment to chromatin during the day, acting in part by maintaining histone acetylation, with REVERBa-dependent GC responses providing segregation of carbohydrate and lipid metabolism. Importantly, deletion of Reverba inverted circadian liver GC sensitivity and protected mice from hepatosteatosis induced by chronic GC administration. Our results reveal a mechanism by which the circadian clock acts through REVERBa in liver on elements bound by HNF4A/HNF6 to direct GR action on energy metabolism.
Giorgio Caratti, Mudassar Iqbal, Louise Hunter, Donghwan Kim, Ping Wang, Ryan M. Vonslow, Nicola Begley, Abigail J. Tetley, Joanna L. Woodburn, Marie Pariollaud, Robert Maidstone, Ian J. Donaldson, Zhenguang Zhang, Louise M. Ince, Gareth Kitchen, Matthew Baxter, Toryn M. Poolman, Dion A. Daniels, David R. Stirling, Chad Brocker, Frank Gonzalez, Andrew S.I. Loudon, David A. Bechtold, Magnus Rattray, Laura C. Matthews, David W. Ray
Prostate cancer is an androgen-dependent disease subject to interactions between the tumor epithelium and its microenvironment. Here, we found that epigenetic changes in prostatic cancer-associated fibroblasts (CAF) initiated a cascade of stromal-epithelial interactions. This facilitated lethal prostate cancer growth and development of resistance to androgen signaling deprivation therapy (ADT). We identified a Ras inhibitor, RASAL3, as epigenetically silenced in human prostatic CAF, leading to oncogenic Ras activity driving macropinocytosis-mediated glutamine synthesis. Interestingly, ADT further promoted RASAL3 epigenetic silencing and glutamine secretion by prostatic fibroblasts. In an orthotopic xenograft model, subsequent inhibition of macropinocytosis and glutamine transport resulted in antitumor effects. Stromal glutamine served as a source of energy through anaplerosis and as a mediator of neuroendocrine differentiation for prostate adenocarcinoma. Antagonizing the uptake of glutamine restored sensitivity to ADT in a castration-resistant xenograft model. In validating these findings, we found that prostate cancer patients on ADT with therapeutic resistance had elevated blood glutamine levels compared with those with therapeutically responsive disease (odds ratio = 7.451, P = 0.02). Identification of epigenetic regulation of Ras activity in prostatic CAF revealed RASAL3 as a sensor for metabolic and neuroendocrine reprogramming in prostate cancer patients failing ADT.
Rajeev Mishra, Subhash Haldar, Veronica Placencio, Anisha Madhav, Krizia Rohena-Rivera, Priyanka Agarwal, Frank Duong, Bryan Angara, Manisha Tripathi, Zhenqiu Liu, Roberta A. Gottlieb, Shawn Wagner, Edwin M. Posadas, Neil A. Bhowmick
Oxidative stress is an underlying component of acute and chronic kidney disease. Apoptosis signal–regulating kinase 1 (ASK1) is a widely expressed redox-sensitive serine threonine kinase that activates p38 and c-Jun N-terminal kinase (JNK) mitogen-activated protein kinase kinases, and induces apoptotic, inflammatory, and fibrotic signaling in settings of oxidative stress. We describe the discovery and characterization of a potent and selective small-molecule inhibitor of ASK1, GS-444217, and demonstrate the therapeutic potential of ASK1 inhibition to reduce kidney injury and fibrosis. Activation of the ASK1 pathway in glomerular and tubular compartments was confirmed in renal biopsies from patients with diabetic kidney disease (DKD) and was decreased by GS-444217 in several rodent models of kidney injury and fibrosis that collectively represented the hallmarks of DKD pathology. Treatment with GS-444217 reduced progressive inflammation and fibrosis in the kidney and halted glomerular filtration rate decline. Combination of GS-444217 with enalapril, an angiotensin-converting enzyme inhibitor, led to a greater reduction in proteinuria and regression of glomerulosclerosis. These results identify ASK1 as an important target for renal disease and support the clinical development of an ASK1 inhibitor for the treatment of DKD.
John T. Liles, Britton K. Corkey, Gregory T. Notte, Grant R. Budas, Eric B. Lansdon, Ford Hinojosa-Kirschenbaum, Shawn S. Badal, Michael Lee, Brian E. Schultz, Sarah Wise, Swetha Pendem, Michael Graupe, Laurie Castonguay, Keith A. Koch, Melanie H. Wong, Giuseppe A. Papalia, Dorothy M. French, Theodore Sullivan, Erik G. Huntzicker, Frank Y. Ma, David J. Nikolic-Paterson, Tareq Altuhaifi, Haichun Yang, Agnes B. Fogo, David G. Breckenridge
Long-lived HIV-1 reservoirs that persist despite antiretroviral therapy (ART) are a major impediment to a cure for HIV-1. We examined whether human liver macrophages (LMs), the largest tissue macrophage population, comprise an HIV-1 reservoir. We purified LMs from liver explants and included treatment with a T cell immunotoxin to reduce T cells to 1% or less. LMs were purified from 9 HIV-1–infected persons, 8 of whom were on ART (range 8–140 months). Purified LMs were stimulated ex vivo and supernatants from 6 of 8 LMs from persons on ART transmitted infection. However, HIV-1 propagation from LMs was not sustained except in LMs from 1 person taking ART for less than 1 year. Bulk liver sequences matched LM-derived HIV-1 in 5 individuals. Additional in vitro experiments undertaken to quantify the decay of HIV-1–infected LMs from 3 healthy controls showed evidence of infection and viral release for prolonged durations (>170 days). Released HIV-1 propagated robustly in target cells, demonstrating that viral outgrowth was observable using our methods. The t1/2 of HIV-1–infected LMs ranged from 3.8–55 days. These findings suggest that while HIV-1 persists in LMs during ART, it does so in forms that are inert, suggesting that they are defective or restricted with regard to propagation.
Abraham J. Kandathil, Sho Sugawara, Ashish Goyal, Christine M. Durand, Jeffrey Quinn, Jaiprasath Sachithanandham, Andrew M. Cameron, Justin R. Bailey, Alan S. Perelson, Ashwin Balagopal
Fibroblast-like synoviocytes (FLSs) are critical to synovial aggression and joint destruction in rheumatoid arthritis (RA). The role of long noncoding RNAs (lncRNAs) in RA is largely unknown. Here, we identified a lncRNA, LERFS (lowly expressed in rheumatoid fibroblast-like synoviocytes), that negatively regulates the migration, invasion, and proliferation of FLSs through interaction with heterogeneous nuclear ribonucleoprotein Q (hnRNP Q). Under healthy conditions, by binding to the mRNA of RhoA, Rac1, and CDC42 — the small GTPase proteins that control the motility and proliferation of FLSs — the LERFS–hnRNP Q complex decreased the stability or translation of target mRNAs and downregulated their protein levels. But in RA FLSs, decreased LERFS levels induced a reduction of the LERFS–hnRNP Q complex, which reduced the binding of hnRNP Q to target mRNA and therefore increased the stability or translation of target mRNA. These findings suggest that a decrease in synovial LERFS may contribute to synovial aggression and joint destruction in RA and that targeting the lncRNA LERFS may have therapeutic potential in patients with RA.
Yaoyao Zou, Siqi Xu, Youjun Xiao, Qian Qiu, Maohua Shi, Jingnan Wang, Liuqin Liang, Zhongping Zhan, Xiuyan Yang, Nancy Olsen, Song Guo Zheng, Hanshi Xu
The E3 ubiquitin ligase RNF8 plays critical roles in maintaining genomic stability by promoting the repair of DNA double-strand breaks (DSBs) through ubiquitin signaling. Abnormal activation of Notch signaling and defective repair of DSBs promote breast cancer risk. Here, we found that low expression of the full-length RNF8 correlated with poor prognosis for breast cancer patients. Our data revealed that in addition to its role in the repair of DSBs, RNF8 regulated Notch1 signaling and cell-fate determination of mammary luminal progenitors. Mechanistically, RNF8 acted as a negative regulator of Notch signaling by ubiquitylating the active NOTCH1 protein (N1ICD), leading to its degradation. Consistent with abnormal activation of Notch signaling and impaired repair of DSBs in Rnf8-mutant mammary epithelial cells, we observed increased risk of mammary tumorigenesis in mouse models for RNF8 deficiency. Notably, deficiency of RNF8 sensitized breast cancer cells to combination of pharmacological inhibitors of Notch signaling and poly(ADP-ribose) polymerase (PARP), suggesting implications for treatment of breast cancer associated with impaired RNF8 expression or function.
Li Li, Kiran Kumar Naidu Guturi, Brandon Gautreau, Parasvi S. Patel, Amine Saad, Mayako Morii, Francesca Mateo, Luis Palomero, Haithem Barbour, Antonio Gomez, Deborah Ng, Max Kotlyar, Chiara Pastrello, Hartland W. Jackson, Rama Khokha, Igor Jurisica, El Bachir Affar, Brian Raught, Otto Sanchez, Moulay Alaoui-Jamali, Miguel A. Pujana, Anne Hakem, Razq Hakem
The M2 isoform of pyruvate kinase (PKM2) is highly expressed in most cancer cells, and has been studied extensively as a driver of oncogenic metabolism. In contrast, the role of PKM2 in nontransformed cells is little studied, and nearly nothing is known of its role, if any, in quiescent cells. We show here that endothelial cells express PKM2 almost exclusively over PKM1. In proliferating endothelial cells, PKM2 is required to suppress p53 and maintain cell cycle progression. In sharp contrast, PKM2 has a strikingly different role in quiescent endothelial cells, where inhibition of PKM2 leads to degeneration of tight junctions and barrier function. Mechanistically, PKM2 regulates barrier function independently of its canonical activity as a pyruvate kinase. Instead, PKM2 suppresses NF-kB and its downstream target, the vascular permeability factor angiopoietin 2. As a consequence, loss of endothelial cell PKM2 in vivo predisposes mice to VEGF-induced vascular leak, and to severe bacteremia and death in response to sepsis. Together, these data demonstrate new roles of PKM2 in quiescent cells, and highlight the need for caution in developing cancer therapies that target PKM2.
Boa Kim, Cholsoon Jang, Harita Dharaneeswaran, Jian Li, Mohit Bhide, Steven Yang, Kristina Li, Zolt Arany
Interrupting T cell costimulatory signals as a strategy to control undesired immune responses, such as occur in autoimmunity or transplantation, has the potential to alleviate many of the unwanted side effects associated with current immunosuppressive therapies. Belatacept, a high-affinity version of CTLA4-Ig that blocks ligand ligation to CD28, has been approved for use in kidney transplant recipients. Despite the long-term benefits associated with its use, such as improved renal function and lower cardiovascular risk, a subset of patients treated with belatacept experience elevated rates of acute T cell–mediated rejection, tempering enthusiasm for its use. Here we demonstrate that costimulation-independent T cell alloreactivity relies on signaling through CD122, the shared IL-2 and IL-15 receptor β-chain. Combined costimulatory and CD122 blockade improved survival of transplanted tissue in mice and nonhuman primates by controlling proliferation and effector function of CD8+ T cells. The high-affinity IL-2 receptor was dispensable for memory CD8+ T cell responses, whereas signaling through CD122 as a component of the high-affinity IL-15 receptor was critical for costimulation-independent memory CD8+ T cell recall, distinguishing specific roles for IL-2 and IL-15 in T cell activation. These studies outline a novel approach for clinical optimization of costimulatory blockade strategies in transplantation by targeting CD122.
David V. Mathews, Ying Dong, Laura B. Higginbotham, Steven C. Kim, Cynthia P. Breeden, Elizabeth A. Stobert, Joseph Jenkins, J. Yun Tso, Christian P. Larsen, Andrew B. Adams
Chronic HBV (CHB) infection suppresses virus-specific T cells, but its impact on humoral immunity has been poorly analyzed. Here, we developed a dual-staining method that utilizes hepatitis B virus (HBV) surface antigens (HBsAg) labeled with fluorochromes as “baits” for specific ex vivo detection of HBsAg-specific B cells and analysis of their quantity, function, and phenotype. We studied healthy vaccinated subjects (n = 18) and patients with resolved (n = 21), acute (n = 11), or chronic (n = 96) HBV infection and observed that frequencies of circulating HBsAg-specific B cells were independent of HBV infection status. In contrast, the presence of serum HBsAg affected function and phenotype of HBsAg-specific B cells that were unable to mature in vitro into Ab-secreting cells and displayed an increased expression of markers linked to hyperactivation (CD21lo) and exhaustion (PD-1). Importantly, B cell alterations were not limited to HBsAg-specific B cells, but affected the global B cell population. HBsAg-specific B cell maturation could be partially restored by a method involving the combination of the cytokines IL-2 and IL-21 and CD40L-expressing feeder cells and was further boosted by the addition of anti–PD-1 Abs. In conclusion, HBV infection has a marked impact on global and HBV-specific humoral immunity, yet HBsAg-specific B cells are amenable to a partial rescue by B cell–maturing cytokines and PD-1 blockade.
Loghman Salimzadeh, Nina Le Bert, Charles-A. Dutertre, Upkar S. Gill, Evan W. Newell, Christian Frey, Magdeleine Hung, Nikolai Novikov, Simon Fletcher, Patrick T.F. Kennedy, Antonio Bertoletti
B cells are increasingly recognized as playing an important role in the ongoing control of hepatitis B virus (HBV). The development of antibodies against the viral surface antigen (HBV surface antigen [HBsAgs]) constitutes the hallmark of resolution of acute infection and is a therapeutic goal for functional cure of chronic HBV (CHB). We characterized B cells directly ex vivo from the blood and liver of patients with CHB to investigate constraints on their antiviral potential. Unexpectedly, we found that HBsAg-specific B cells persisted in the blood and liver of many patients with CHB and were enriched for T-bet, a signature of antiviral potential in B cells. However, purified, differentiated HBsAg-specific B cells from patients with CHB had defective antibody production, consistent with undetectable anti-HBs antibodies in vivo. HBsAg-specific and global B cells had an accumulation of CD21–CD27– atypical memory B cells (atMBC) with high expression of inhibitory receptors, including PD-1. These atMBC demonstrated altered signaling, homing, differentiation into antibody-producing cells, survival, and antiviral/proinflammatory cytokine production that could be partially rescued by PD-1 blockade. Analysis of B cells within healthy and HBV-infected livers implicated the combination of this tolerogenic niche and HBV infection in driving PD-1hiatMBC and impairing B cell immunity.
Alice R. Burton, Laura J. Pallett, Laura E. McCoy, Kornelija Suveizdyte, Oliver E. Amin, Leo Swadling, Elena Alberts, Brian R. Davidson, Patrick T.F. Kennedy, Upkar S. Gill, Claudia Mauri, Paul A. Blair, Nadege Pelletier, Mala K. Maini
Regulatory T cells (Tregs) are critical for maintaining immune homeostasis. However, current Treg immunotherapies do not optimally treat inflammatory diseases in patients. Understanding the cellular processes that control Treg function may allow for the augmentation of therapeutic efficacy. In contrast to activated conventional T cells, in which protein kinase C-θ (PKC-θ) localizes to the contact point between T cells and antigen-presenting cells, in human and mouse Tregs, PKC-θ localizes to the opposite end of the cell in the distal pole complex (DPC). Here, using a phosphoproteomic screen, we identified the intermediate filament vimentin as a PKC-θ phospho target and show that vimentin forms a DPC superstructure on which PKC-θ accumulates. Treatment of mouse Tregs with either a clinically relevant PKC-θ inhibitor or vimentin siRNA disrupted vimentin and enhanced Treg metabolic and suppressive activity. Moreover, vimentin-disrupted mouse Tregs were significantly better than controls at suppressing alloreactive T cell priming in graft-versus-host disease (GVHD) and GVHD lethality, using a complete MHC-mismatch mouse model of acute GVHD (C57BL/6 donor into BALB/c host). Interestingly, vimentin disruption augmented the suppressor function of PKC-θ–deficient mouse Tregs. This suggests that enhanced Treg activity after PKC-θ inhibition is secondary to effects on vimentin, not just PKC-θ kinase activity inhibition. Our data demonstrate that vimentin is a key metabolic and functional controller of Treg activity and provide proof of principle that disruption of vimentin is a feasible, translationally relevant method to enhance Treg potency.
Cameron McDonald-Hyman, James T. Muller, Michael Loschi, Govindarajan Thangavelu, Asim Saha, Sudha Kumari, Dawn K. Reichenbach, Michelle J. Smith, Guoan Zhang, Brent H. Koehn, Jiqiang Lin, Jason S. Mitchell, Brian T. Fife, Angela Panoskaltsis-Mortari, Colby J. Feser, Andrew Kemal Kirchmeier, Mark J. Osborn, Keli L. Hippen, Ameeta Kelekar, Jonathan S. Serody, Laurence A. Turka, David H. Munn, Hongbo Chi, Thomas A. Neubert, Michael L. Dustin, Bruce R. Blazar
BACKGROUND. Injectable depot medroxyprogesterone acetate (DMPA) is one of the most popular contraception methods in areas of high HIV seroprevalence. Evidence is accumulating that use of DMPA might be associated with an increased risk of HIV-1 acquisition by women; however, mechanisms of this association are not completely understood. The goal of this study was to gain insight into mechanisms underlying the possible link between use of DMPA and risk of HIV-1 acquisition, exploring transcription profiling of ectocervical tissues. METHODS. Healthy women received either DMPA (n = 31) or combined oral contraceptive (COC), which has not been linked to an increased risk of HIV acquisition (n = 32). We conducted a comparative microarray-based whole-genome transcriptome profiling of human ectocervical tissues before and after 6 weeks of hormonal contraception use. RESULTS. The analysis identified that expression of 235 and 76 genes was significantly altered after DMPA and COC use, respectively. The most striking effect of DMPA, but not COC, was significantly altered expression (mostly downregulation) of many genes strategically involved in the maintenance of mucosal barrier function; the alterations, as indicated by Ingenuity Pathway Analysis (IPA), were most likely due to the DMPA-induced estrogen deficiency. Furthermore, IPA predicted that transcriptome alterations related to ectocervical immune responses were in general compatible with an immunosuppressive effect of DMPA, but, in some women, also with an inflammatory-like response. CONCLUSION. Our results suggest that impairment of cervicovaginal mucosal integrity in response to DMPA administration is an important mechanism contributing to the potential increased risk of HIV-1 acquisition in DMPA users. TRIAL REGISTRATION. ClinicalTrials.gov NCT01421368. FUNDING. This study was supported by the United States Agency for International Development (USAID) under Cooperative Agreement GPO-A-00-08-00005-00.
Irina A. Zalenskaya, Neelima Chandra, Nazita Yousefieh, Xi Fang, Oluwatosin E. Adedipe, Suzanne S. Jackson, Sharon M. Anderson, Christine K. Mauck, Jill L. Schwartz, Andrea R. Thurman, Gustavo F. Doncel
Ferroptosis is a death program executed via selective oxidation of arachidonic acid–phosphatidylethanolamines (AA-PE) by 15-lipoxygenases. In mammalian cells and tissues, ferroptosis has been pathogenically associated with brain, kidney, and liver injury/diseases. We discovered that a prokaryotic bacterium, Pseudomonas aeruginosa, that does not contain AA-PE can express lipoxygenase (pLoxA), oxidize host AA-PE to 15-hydroperoxy-AA-PE (15-HOO-AA-PE), and trigger ferroptosis in human bronchial epithelial cells. Induction of ferroptosis by clinical P. aeruginosa isolates from patients with persistent lower respiratory tract infections was dependent on the level and enzymatic activity of pLoxA. Redox phospholipidomics revealed elevated levels of oxidized AA-PE in airway tissues from patients with cystic fibrosis (CF) but not with emphysema or CF without P. aeruginosa. We believe that the evolutionarily conserved mechanism of pLoxA-driven ferroptosis may represent a potential therapeutic target against P. aeruginosa–associated diseases such as CF and persistent lower respiratory tract infections.
Haider H. Dar, Yulia Y. Tyurina, Karolina Mikulska-Ruminska, Indira Shrivastava, Hsiu-Chi Ting, Vladimir A. Tyurin, James Krieger, Claudette M. St. Croix, Simon Watkins, Erkan Bayir, Gaowei Mao, Catherine R. Armbruster, Alexandr Kapralov, Hong Wang, Matthew R. Parsek, Tamil S. Anthonymuthu, Abiola F. Ogunsola, Becca A. Flitter, Cody J. Freedman, Jordan R. Gaston, Theodore R. Holman, Joseph M. Pilewski, Joel S. Greenberger, Rama K. Mallampalli, Yohei Doi, Janet S. Lee, Ivet Bahar, Jennifer M. Bomberger, Hülya Bayır, Valerian E. Kagan
Checkpoint blockade immunotherapy targeting the PD-1/PD-L1 inhibitory axis has produced remarkable results in the treatment of several types of cancer. Whereas cytotoxic T cells are known to provide important antitumor effects during checkpoint blockade, certain cancers with low MHC expression are responsive to therapy, suggesting that other immune cell types may also play a role. Here, we employed several mouse models of cancer to investigate the effect of PD-1/PD-L1 blockade on NK cells, a population of cytotoxic innate lymphocytes that also mediate antitumor immunity. We discovered that PD-1 and PD-L1 blockade elicited a strong NK cell response that was indispensable for the full therapeutic effect of immunotherapy. PD-1 was expressed on NK cells within transplantable, spontaneous, and genetically induced mouse tumor models, and PD-L1 expression in cancer cells resulted in reduced NK cell responses and generation of more aggressive tumors in vivo. PD-1 expression was more abundant on NK cells with an activated and more responsive phenotype and did not mark NK cells with an exhausted phenotype. These results demonstrate the importance of the PD-1/PD-L1 axis in inhibiting NK cell responses in vivo and reveal that NK cells, in addition to T cells, mediate the effect of PD-1/PD-L1 blockade immunotherapy.
Joy Hsu, Jonathan J. Hodgins, Malvika Marathe, Chris J. Nicolai, Marie-Claude Bourgeois-Daigneault, Troy N. Trevino, Camillia S. Azimi, Amit K. Scheer, Haley E. Randolph, Thornton W. Thompson, Lily Zhang, Alexandre Iannello, Nikhita Mathur, Karen E. Jardine, Georgia A. Kirn, John C. Bell, Michael W. McBurney, David H. Raulet, Michele Ardolino
Chronic inflammatory diseases are characterized by recurrent inflammatory attacks in the tissues mediated by autoreactive T cells. Identity and functional programming of CD8+ T cells at the target site of inflammation still remain elusive. One key question is whether, in these antigen-rich environments, chronic stimulation leads to CD8+ T cell exhaustion comparable to what is observed in infectious disease contexts. In the synovial fluid (SF) of juvenile idiopathic arthritis (JIA) patients, a model of chronic inflammation, an overrepresentation of PD-1+CD8+ T cells was found. Gene expression profiling, gene set enrichment analysis, functional studies, and extracellular flux analysis identified PD-1+CD8+ T cells as metabolically active effectors, with no sign of exhaustion. Furthermore, PD-1+CD8+ T cells were enriched for a tissue-resident memory (Trm) cell transcriptional profile and demonstrated increased clonal expansion compared with the PD-1– counterpart, suggesting antigen-driven expansion of locally adapted cells. Interestingly, this subset was also found increased in target tissues in other human chronic inflammatory diseases. These data indicate that local chronic inflammation drives the induction and expansion of CD8+ T cells endowed with potential detrimental properties. Together, these findings lay the basis for investigation of PD-1–expressing CD8+ T cell targeting strategies in human chronic inflammatory diseases.
Alessandra Petrelli, Gerdien Mijnheer, David P. Hoytema van Konijnenburg, Maria M. van der Wal, Barbara Giovannone, Enric Mocholi, Nadia Vazirpanah, Jasper C. Broen, Dirkjan Hijnen, Bas Oldenburg, Paul J. Coffer, Sebastian J. Vastert, Berent J. Prakken, Eric Spierings, Aridaman Pandit, Michal Mokry, Femke van Wijk
Polyamine inhibition for cancer therapy is, conceptually, an attractive approach but has yet to meet success in the clinical setting. The aryl hydrocarbon receptor (AHR) is the central transcriptional regulator of the xenobiotic response. Our study revealed that AHR also positively regulates intracellular polyamine production via direct transcriptional activation of 2 genes, ODC1 and AZIN1, which are involved in polyamine biosynthesis and control, respectively. In patients with multiple myeloma (MM), AHR levels were inversely correlated with survival, suggesting that AHR inhibition may be beneficial for the treatment of this disease. We identified clofazimine (CLF), an FDA-approved anti-leprosy drug, as a potent AHR antagonist and a suppressor of polyamine biosynthesis. Experiments in a transgenic model of MM (Vk*Myc mice) and in immunocompromised mice bearing MM cell xenografts revealed high efficacy of CLF comparable to that of bortezomib, a first-in-class proteasome inhibitor used for the treatment of MM. This study identifies a previously unrecognized regulatory axis between AHR and polyamine metabolism and reveals CLF as an inhibitor of AHR and a potentially clinically relevant anti-MM agent.
Anna Bianchi-Smiraglia, Archis Bagati, Emily E. Fink, Hayley C. Affronti, Brittany C. Lipchick, Sudha Moparthy, Mark D. Long, Spencer R. Rosario, Shivana M. Lightman, Kalyana Moparthy, David W. Wolff, Dong Hyun Yun, Zhannan Han, Anthony Polechetti, Matthew V. Roll, Ilya I. Gitlin, Katerina I. Leonova, Aryn M. Rowsam, Eugene S. Kandel, Andrei V. Gudkov, P. Leif Bergsagel, Kelvin P. Lee, Dominic J. Smiraglia, Mikhail A. Nikiforov
Zika virus (ZIKV) is a teratogenic mosquito-borne flavivirus that can be sexually transmitted from man to woman. The finding of high viral loads and prolonged viral shedding in semen suggests that ZIKV replicates within the human male genital tract, but its target organs are unknown. Using ex vivo infection of organotypic cultures, we demonstrated here that ZIKV replicates in human testicular tissue and infects a broad range of cell types, including germ cells, which we also identified as infected in semen from ZIKV-infected donors. ZIKV had no major deleterious effect on the morphology and hormonal production of the human testis explants. Infection induced a broad antiviral response but no IFN upregulation and minimal proinflammatory response in testis explants, with no cytopathic effect. Finally, we studied ZIKV infection in mouse testis and compared it to human infection. This study provides key insights into how ZIKV may persist in semen and alter semen parameters, as well as a valuable tool for testing antiviral agents.
Giulia Matusali, Laurent Houzet, Anne-Pascale Satie, Dominique Mahé, Florence Aubry, Thérèse Couderc, Julie Frouard, Salomé Bourgeau, Karim Bensalah, Sylvain Lavoué, Guillaume Joguet, Louis Bujan, André Cabié, Gleide Avelar, Marc Lecuit, Anna Le Tortorec, Nathalie Dejucq-Rainsford
The resolution of inflammation is an active process that is coordinated by endogenous mediators. Previous studies have demonstrated the immunomodulatory properties of the axonal guidance proteins in the initial phase of acute inflammation. We hypothesized that the neuronal guidance protein neogenin (Neo1) modulates mechanisms of inflammation resolution. In murine peritonitis, Neo1 deficiency (Neo1–/–) resulted in higher efficacies in reducing neutrophil migration into injury sites, increasing neutrophil apoptosis, actuating PMN phagocytosis, and increasing the endogenous biosynthesis of specialized proresolving mediators, such as lipoxin A4, maresin-1, and protectin DX. Neo1 expression was limited to Neo1-expressing Ly6Chi monocytes, and Neo1 deficiency induced monocyte polarization toward an antiinflammatory and proresolving phenotype. Signaling network analysis revealed that Neo1–/– monocytes mediate their immunomodulatory effects specifically by activating the PI3K/AKT pathway and suppressing the TGF-β pathway. In a cohort of 59 critically ill, intensive care unit (ICU) pediatric patients, we found a strong correlation between Neo1 blood plasma levels and abdominal compartment syndrome, Pediatric Risk of Mortality III (PRISM-III) score, and ICU length of stay and mortality. Together, these findings identify a crucial role for Neo1 in regulating tissue regeneration and resolution of inflammation, and determined Neo1 to be a predictor of morbidity and mortality in critically ill children affected by clinical inflammation.
Martin Schlegel, Andreas Körner, Torsten Kaussen, Urs Knausberg, Carmen Gerber, Georg Hansmann, Hulda Soffia Jónasdóttir, Martin Giera, Valbona Mirakaj
Chronic inflammatory demyelinating polyneuropathy (CIDP) and Guillain-Barre syndrome (GBS) are inflammatory neuropathies that affect humans and are characterized by peripheral nerve myelin destruction and macrophage-containing immune infiltrates. In contrast to the traditional view that the peripheral nerve is simply the target of autoimmunity, we report here that peripheral nerve Schwann cells exacerbate the autoimmune process through extracellular matrix (ECM) protein induction. In a spontaneous autoimmune peripheral polyneuropathy (SAPP) mouse model of inflammatory neuropathy and CIDP nerve biopsies, the ECM protein periostin (POSTN) was upregulated in affected sciatic nerves and was primarily expressed by Schwann cells. Postn deficiency delayed the onset and reduced the extent of neuropathy, as well as decreased the number of macrophages infiltrating the sciatic nerve. In an in vitro assay, POSTN promoted macrophage chemotaxis in an integrin-AM (ITGAM) and ITGAV-dependent manner. The PNS-infiltrating macrophages in SAPP-affected nerves were pathogenic, since depletion of macrophages protected against the development of neuropathy. Our findings show that Schwann cells promote macrophage infiltration by upregulating Postn and suggest that POSTN is a novel target for the treatment of macrophage-associated inflammatory neuropathies.
Denise E. Allard, Yan Wang, Jian Joel Li, Bridget Conley, Erin W. Xu, David Sailer, Caellaigh Kimpston, Rebecca Notini, Collin-Jamal Smith, Emel Koseoglu, Joshua Starmer, Xiaopei L. Zeng, James F. Howard Jr., Ahmet Hoke, Steven S. Scherer, Maureen A. Su
Cong Jin, Christopher P. Shelburne, Guojie Li, Erin N. Potts, Kristina J. Riebe, Gregory D. Sempowski, W. Michael Foster, Soman N. Abraham
Anna Sophia McKenney, Allison N. Lau, Amritha Varshini Hanasoge Somasundara, Barbara Spitzer, Andrew M. Intlekofer, Jihae Ahn, Kaitlyn Shank, Franck T. Rapaport, Minal A. Patel, Efthymia Papalexi, Alan H. Shih, April Chiu, Elizaveta Freinkman, Esra A. Akbay, Mya Steadman, Raj Nagaraja, Katharine Yen, Julie Teruya-Feldstein, Kwok-Kin Wong, Raajit Rampal, Matthew G. Vander Heiden, Craig B. Thompson, Ross L. Levine