Anti-LPS antibodies protect against Klebsiella pneumoniae by empowering neutrophil-mediated clearance without neutralizing TLR4

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Abstract

Initial promising results with immune sera guided early human mAb approaches against Gram-negative sepsis to an LPS neutralization mechanism, but these efforts failed in human clinical trials. Emergence of multidrug resistance has renewed interest in pathogen-specific mAbs. We utilized a pair of antibodies targeting Klebsiella pneumoniae LPS, one that both neutralizes LPS/TLR4 signaling and mediates opsonophagocytic killing (OPK) (54H7) and one that only promotes OPK (KPE33), to better understand the contribution of each mechanism to mAb protection in an acutely lethal pneumonia model. Passive immunization 24 hours prior to infection with KPE33 protected against lethal infection significantly better than 54H7, while delivery of either mAb 1 hour after infection resulted in similar levels of protection. These data suggest that early neutralization of LPS-induced signaling limits protection afforded by these mAbs. LPS neutralization prevented increases in the numbers of γδT cells, a major producer of the antimicrobial cytokine IL-17A, the contribution of which was confirmed using il17a-knockout mice. We conclude that targeting LPS for OPK without LPS signaling neutralization has potential to combat Gram-negative infection by engaging host immune defenses, rather than inhibiting beneficial innate immune pathways.

Authors

Taylor S. Cohen, Mark Pelletier, Lily Cheng, Meghan E. Pennini, Jessica Bonnell, Romana Cvitkovic, Chew-shun Chang, Xiaodong Xiao, Elisabetta Cameroni, Davide Corti, Elena Semenova, Paul Warrener, Bret R. Sellman, JoAnn Suzich, Qun Wang, C. Kendall Stover

Broadly neutralizing antibodies with few somatic mutations and hepatitis C virus clearance

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Abstract

Here, we report the isolation of broadly neutralizing mAbs (bNAbs) from persons with broadly neutralizing serum who spontaneously cleared hepatitis C virus (HCV) infection. We found that bNAbs from two donors bound the same epitope and were encoded by the same germline heavy chain variable gene segment. Remarkably, these bNAbs were encoded by antibody variable genes with sparse somatic mutations. For one of the most potent bNAbs, these somatic mutations were critical for antibody neutralizing breadth and for binding to autologous envelope variants circulating late in infection. However, somatic mutations were not necessary for binding of the bNAb unmutated ancestor to envelope proteins of early autologous transmitted/founder viruses. This study identifies a public B cell clonotype favoring early recognition of a conserved HCV epitope, proving that anti-HCV bNAbs can achieve substantial neutralizing breadth with relatively few somatic mutations, and identifies HCV envelope variants that favored selection and maturation of an anti-HCV bNAb in vivo. These data provide insight into the molecular mechanisms of immune-mediated clearance of HCV infection and present a roadmap to guide development of a vaccine capable of stimulating anti-HCV bNAbs with a physiologic number of somatic mutations characteristic of vaccine responses.

Authors

Justin R. Bailey, Andrew I. Flyak, Valerie J. Cohen, Hui Li, Lisa N. Wasilewski, Anna E. Snider, Shuyi Wang, Gerald H. Learn, Nurgun Kose, Leah Loerinc, Rebecca Lampley, Andrea L. Cox, Jennifer M. Pfaff, Benjamin J. Doranz, George M. Shaw, Stuart C. Ray, James E. Crowe Jr.

¹⁸F-FDG as an inflammation biomarker for imaging dengue virus infection and treatment response

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Abstract

Development of antiviral therapy against acute viral diseases, such as dengue virus (DENV), suffers from the narrow window of viral load detection in serum during onset and clearance of infection and fever. We explored a biomarker approach using ¹⁸F-fluorodeoxyglucose (¹⁸F-FDG) PET in established mouse models for primary and antibody-dependent enhancement infection with DENV. ¹⁸F-FDG uptake was most prominent in the intestines and correlated with increased virus load and proinflammatory cytokines. Furthermore, a significant temporal trend in ¹⁸F-FDG uptake was seen in intestines and selected tissues over the time course of infection. Notably, ¹⁸F-FDG uptake and visualization by PET robustly differentiated treatment-naive groups from drug-treated groups as well as nonlethal from lethal infections with a clinical strain of DENV2. Thus, ¹⁸F-FDG may serve as a novel DENV infection–associated inflammation biomarker for assessing treatment response during therapeutic intervention trials.

Authors

Ann-Marie Chacko, Satoru Watanabe, Keira J. Herr, Shirin Kalimuddin, Jing Yang Tham, Joanne Ong, Marie Reolo, Raymond M.F. Serrano, Yin Bun Cheung, Jenny G.H. Low, Subhash G. Vasudevan

MPEG1/perforin-2 mutations in human pulmonary nontuberculous mycobacterial infections

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Abstract

Perforin-2 is a highly conserved pore-forming protein encoded by macrophage expressed gene 1 (MPEG1). A number of studies have shown that Perforin-2–deficient mice are unable to survive following a bacterial challenge that is nonlethal in WT mice. There is also recent evidence that Mpeg1^+/– heterozygous mice display an intermediate killing ability compared with Mpeg1 WT and Mpeg1^–/– mice. Despite these in vivo findings, to date, no perforin-2 deficiencies have been associated with human disease. Here, we report four patients with persistent nontuberculous mycobacterial infection who had heterozygous MPEG1 mutations. In vitro, neutrophils, macrophages, and B cells from these patients were unable to kill Mycobacterium avium as efficiently as normal controls. CRISPR mutagenesis validated the deleterious antibacterial activity of these mutations. These data suggest that perforin-2 haploinsufficiency may contribute to human susceptibility to infections with intracellular bacteria.

Authors

Ryan M. McCormack, Eva P. Szymanski, Amy P. Hsu, Elena Perez, Kenneth N. Olivier, Eva Fisher, E. Brook Goodhew, Eckhard R. Podack, Steven M. Holland

Cross-reactive dengue human monoclonal antibody prevents severe pathologies and death from Zika virus infections

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Abstract

Zika virus (ZIKV) infections have been linked with neurological complications and congenital Zika syndrome. Given the high level of homology between ZIKV and the related flavivirus dengue virus (DENV), we investigated the level of cross-reactivity with ZIKV using a panel of DENV human mAbs. A majority of the mAbs showed binding to ZIKV virions, with several exhibiting neutralizing capacities against ZIKV in vitro. Three of the best ZIKV-neutralizing mAbs were found to recognize diverse epitopes on the envelope (E) glycoprotein: the highly conserved fusion-loop peptide, a conformation-specific epitope on the E monomer, and a quaternary epitope on the virion surface. The most potent ZIKV-neutralizing mAb (SIgN-3C) was assessed in 2 type I interferon receptor–deficient (IFNAR^–/–) mouse models of ZIKV infection. Treatment of adult nonpregnant mice with SIgN-3C rescued mice from virus-induced weight loss and mortality. The SIgN-3C variant with Leu-to-Ala mutations in the Fc region (SIgN-3C-LALA) did not induce antibody-dependent enhancement (ADE) in vitro but provided similar levels of protection in vivo. In pregnant ZIKV-infected IFNAR^–/– mice, treatment with SIgN-3C or SIgN-3C-LALA significantly reduced viral load in the fetal organs and placenta and abrogated virus-induced fetal growth retardation. Therefore, SIgN-3C-LALA holds promise as a ZIKV prophylactic and therapeutic agent.

Authors

Yiu-Wing Kam, Cheryl Yi-Pin Lee, Teck-Hui Teo, Shanshan W. Howland, Siti Naqiah Amrun, Fok-Moon Lum, Peter See, Nicholas Qing-Rong Kng, Roland G. Huber, Mei-Hui Xu, Heng-Liang Tan, Andre Choo, Sebastian Maurer-Stroh, Florent Ginhoux, Katja Fink, Cheng-I Wang, Lisa F.P. Ng, Laurent Rénia

IL-21 is required for CD4 memory formation in response to viral infection

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Abstract

IL-21 has been shown to play an important role in the CD8 T cell response during acute and chronic viral infections. However, the role of IL-21 signaling in the CD4 T cell response to viral infection remains incompletely defined. In a model of infection with vaccinia virus, we show that intrinsic IL-21 signaling on CD4 T cells was critical for the formation of memory CD4 T cells in vivo. We further reveal that IL-21 promoted CD4 T cell survival in a mechanism dependent on activation of the STAT1 and STAT3 signaling pathways. In addition, the activation of Akt is also required for IL-21–dependent survival of CD4 T cells in vivo. These results identify a critical role for intrinsic IL-21 signaling in CD4 T cell survival and memory formation in response to viral infection in vivo and may provide insights into the design of effective vaccine strategies.

Authors

Yuqing Yuan, Yiping Yang, Xiaopei Huang

BCG vaccination induces HIV target cell activation in HIV-exposed infants in a randomized trial

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Abstract

BACKGROUND. Bacillus Calmette-Guérin (BCG) vaccine is administered at birth to protect infants against tuberculosis throughout Africa, where most perinatal HIV-1 transmission occurs. We examined whether BCG vaccination alters the levels of activated HIV target T cells in HIV-exposed South African infants.

METHODS. HIV-exposed infants were randomized to receive routine (at birth) or delayed (at 8 weeks) BCG vaccination. Activated and CCR5-expressing peripheral blood CD4⁺ T cell, monocyte, and NK cell frequencies were evaluated by flow cytometry and immune gene expression via PCR using Biomark (Fluidigm).

RESULTS. Of 149 infants randomized, 92% (n = 137) were retained at 6 weeks: 71 in the routine BCG arm and 66 in the delayed arm. Routine BCG vaccination led to a 3-fold increase in systemic activation of HIV target CD4⁺CCR5⁺ T cells (HLA-DR⁺CD38⁺) at 6 weeks (0.25% at birth versus 0.08% in delayed vaccination groups; P = 0.029), which persisted until 8 weeks of age when the delayed arm was vaccinated. Vaccination of the infants in the delayed arm at 8 weeks resulted in a similar increase in activated CD4⁺CCR5⁺ T cells. The increase in activated T cells was associated with increased levels of MHC class II transactivator (CIITA), IL12RB1, and IFN-α1 transcripts within peripheral blood mononuclear cells but minimal changes in innate cells.

CONCLUSION. BCG vaccination induces immune changes in HIV-exposed infants, including an increase in the proportion of activated CCR5⁺CD4⁺ HIV target cells. These findings provide insight into optimal BCG vaccine timing to minimize the risks of HIV transmissions to exposed infants while preserving potential benefits conferred by BCG vaccination.

TRIAL REGISTRATION. ClinicalTrials.gov NCT02062580.

FUNDING. This trial was sponsored by the Elizabeth Glaser Pediatric AIDS Foundation (MV-00-9-900-01871-0-00) and the Thrasher Foundation (NR-0095); for details, see Acknowledgments.

Authors

Melanie A. Gasper, Anneke C. Hesseling, Isaac Mohar, Landon Myer, Tali Azenkot, Jo-Ann S. Passmore, Willem Hanekom, Mark F. Cotton, I. Nicholas Crispe, Donald L. Sodora, Heather B. Jaspan

M1-like monocytes are a major immunological determinant of severity in previously healthy adults with life-threatening influenza

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Abstract

In each influenza season, a distinct group of young, otherwise healthy individuals with no risk factors succumbs to life-threatening infection. To better understand the cause for this, we analyzed a broad range of immune responses in blood from a unique cohort of patients, comprising previously healthy individuals hospitalized with and without respiratory failure during one influenza season, and infected with one specific influenza A strain. This analysis was compared with similarly hospitalized influenza patients with known risk factors (total of n = 60 patients recruited). We found a sustained increase in a specific subset of proinflammatory monocytes, with high TNF-α expression and an M1-like phenotype (independent of viral titers), in these previously healthy patients with severe disease. The relationship between M1-like monocytes and immunopathology was strengthened using murine models of influenza, in which severe infection generated using different models (including the high-pathogenicity H5N1 strain) was also accompanied by high levels of circulating M1-like monocytes. Additionally, a raised M1/M2 macrophage ratio in the lungs was observed. These studies identify a specific subtype of monocytes as a modifiable immunological determinant of disease severity in this subgroup of severely ill, previously healthy patients, offering potential novel therapeutic avenues.

Authors

Suzanne L. Cole, Jake Dunning, Wai Ling Kok, Kambez Hajipouran Benam, Adel Benlahrech, Emmanouela Repapi, Fernando O. Martinez, Lydia Drumright, Timothy J. Powell, Michael Bennett, Ruth Elderfield, Catherine Thomas, MOSAIC investigators, Tao Dong, John McCauley, Foo Y. Liew, Stephen Taylor, Maria Zambon, Wendy Barclay, Vincenzo Cerundolo, Peter J. Openshaw, Andrew J. McMichael, Ling-Pei Ho

Attenuated and vectored vaccines protect nonhuman primates against Chikungunya virus

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Abstract

Chikungunya virus (CHIKV) is rapidly spreading across the globe, and millions are infected. Morbidity due to this virus is a serious threat to public health, but at present, there is no vaccine against this debilitating disease. We have recently developed a number of vaccine candidates, and here we have evaluated 3 of them in a nonhuman primate model. A single immunization with an attenuated strain of CHIKV (Δ5nsP3), a homologous prime-boost immunization with a DNA-launched RNA replicon encoding CHIKV envelope proteins (DREP-E), and a DREP-E prime followed by a recombinant modified vaccinia virus Ankara encoding CHIKV capsid and envelope (MVA-CE) boost all induced protection against WT CHIKV infection. The attenuated Δ5nsP3 virus proved to be safe and did not show any clinical signs typically associated with WT CHIKV infections such as fever, skin rash, lymphopenia, or joint swelling. These vaccines are based on an East/Central/South African strain of Indian Ocean lineage, but they also generated neutralizing antibodies against an isolate of the Asian genotype that now is rapidly spreading across the Americas. These results form the basis for clinical development of an efficacious CHIKV vaccine that generates both humoral and cellular immunity with long-term immunological memory.

Authors

Pierre Roques, Karl Ljungberg, Beate M. Kümmerer, Leslie Gosse, Nathalie Dereuddre-Bosquet, Nicolas Tchitchek, David Hallengärd, Juan García-Arriaza, Andreas Meinke, Mariano Esteban, Andres Merits, Roger Le Grand, Peter Liljeström

Clinical, virological, and biological parameters associated with outcomes of Ebola virus infection in Macenta, Guinea

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Abstract

BACKGROUND. The pathogenesis of Ebola virus (EBOV) disease (EVD) is poorly characterized. The establishment of well-equipped diagnostic laboratories close to Ebola treatment centers (ETCs) has made it possible to obtain relevant virological and biological data during the course of EVD and to assess their association with the clinical course and different outcomes of the disease.

METHODS. We were responsible for diagnosing EBOV infection in patients admitted to two ETCs in forested areas of Guinea. The pattern of clinical signs was recorded, and an etiological diagnosis was established by RT-PCR for EBOV infection or a rapid test for malaria and typhoid fever. Biochemical analyses were also performed.

RESULTS. We handled samples from 168 patients between November 29, 2014, and January 31, 2015; 97 patients were found to be infected with EBOV, with Plasmodium falciparum coinfection in 18%. Overall mortality for EVD cases was 58%, rising to 86% if P. falciparum was also present. Viral load was higher in fatal cases of EVD than in survivors, and fatal cases were associated with higher aspartate aminotransferase (AST) and alanine aminotransferase (ALT), C-reactive protein (CRP), and IL-6 levels. Furthermore, regardless of outcome, EVD was characterized by higher creatine kinase (CPK), amylase, and creatinine levels than in febrile patients without EVD, with higher blood urea nitrogen (BUN) levels in fatal cases of EVD only.

CONCLUSION. These findings suggest that a high viral load at admission is a marker of poor EVD prognosis. In addition, high AST, ALT, CRP, and IL-6 levels are associated with a fatal outcome of EVD. Damage to the liver and other tissues, with massive rhabdomyolysis and, probably, acute pancreatitis, is associated with EVD and correlated with disease severity. Finally, biochemical analyses provide substantial added value at ETCs, making it possible to improve supportive rehydration and symptomatic care for patients.

FUNDING. The French Ministry of Foreign Affairs, the Agence Française de Développement, and Institut Pasteur.

Authors

Marie-Astrid Vernet, Stéphanie Reynard, Alexandra Fizet, Justine Schaeffer, Delphine Pannetier, Jeremie Guedj, Max Rives, Nadia Georges, Nathalie Garcia-Bonnet, Aboubacar I. Sylla, Péma Grovogui, Jean-Yves Kerherve, Christophe Savio, Sylvie Savio-Coste, Marie-Laure de Séverac, Philippe Zloczewski, Sandrine Linares, Souley Harouna, Bing M’Lebing Abdoul, Frederic Petitjean, Nenefing Samake, Susan Shepherd, Moumouni Kinda, Fara Roger Koundouno, Ludovic Joxe, Mathieu Mateo, Patrick Lecine, Audrey Page, Tang Maleki Tchamdja, Matthieu Schoenhals, Solenne Barbe, Bernard Simon, Tuan Tran-Minh, Christophe Longuet, François L’Hériteau, Sylvain Baize