New chimeric TLR7/NOD2 agonist is a potent adjuvant to induce mucosal immune responses

International audienceBackground: PRR (Pattern Recognition Receptor) agonists have been widely tested as potent vaccine adjuvants. TLR7 (Toll-Like Receptor 7) and NOD2 (nucleotide-binding oligomerization domain 2) are key innate receptors widely expressed at mucosal levels.Methods: Here, we evaluated the immunostimulatory properties of a novel hybrid chemical compound designed to stimulate both TLR7 and NOD2 receptors.Finding: The combined TLR7/NOD2 agonist showed increase efficacy than TLR7L or NOD2L agonists alone or combined in different in vitro models. Dual TLR7/NOD2 agonist efficiently stimulates TLR7 and NOD2, and promotes the maturation and reprogramming of human dendritic cells, as well as the secretion of pro-inflammatory or adaptive cytokines. This molecule also strongly induces autophagy in human cells which is a major intracellular degradation system that delivers cytoplasmic constituents to lysosomes in both MHC class I and II-restricted antigen presentation. In vivo, TLR7/NOD2L agonist is a potent adjuvant after intranasal administration with NP-p24 HIV vaccine, inducing high-quality humoral and adaptive responses both in systemic and mucosal compartments. Use of TLR7/NOD2L adjuvant improves very significantly the protection of mice against an intranasal challenge with a vaccinia virus expressing the p24.Interpretation: Dual TLR7/NOD2L agonist is a very potent and versatile vaccine adjuvant and promote very efficiently both systemic and mucosal immunity

STING agonists enable antiviral cross-talk between human cells and confer protection against genital herpes in mice

<div><p>In recent years, there has been an increasing interest in immunomodulatory therapy as a means to treat various conditions, including infectious diseases. For instance, Toll-like receptor (TLR) agonists have been evaluated for treatment of genital herpes. However, although the TLR7 agonist imiquimod was shown to have antiviral activity in individual patients, no significant effects were observed in clinical trials, and the compound also exhibited significant side effects, including local inflammation. Cytosolic DNA is detected by the enzyme cyclic GMP-AMP (2’3’-cGAMP) synthase (cGAS) to stimulate antiviral pathways, mainly through induction of type I interferon (IFN)s. cGAS is activated upon DNA binding to produce the cyclic dinucleotide (CDN) 2’3’-cGAMP, which in turn binds and activates the adaptor protein Stimulator of interferon genes (STING), thus triggering type I IFN expression. In contrast to TLRs, STING is expressed broadly, including in epithelial cells. Here we report that natural and non-natural STING agonists strongly induce type I IFNs in human cells and in mice <i>in vivo</i>, without stimulating significant inflammatory gene expression. Systemic treatment with 2’3’-cGAMP reduced genital herpes simplex virus (HSV) 2 replication and improved the clinical outcome of infection. More importantly, local application of CDNs at the genital epithelial surface gave rise to local IFN activity, but only limited systemic responses, and this treatment conferred total protection against disease in both immunocompetent and immunocompromised mice. In direct comparison between CDNs and TLR agonists, only CDNs acted directly on epithelial cells, hence allowing a more rapid and IFN-focused immune response in the vaginal epithelium. Thus, specific activation of the STING pathway in the vagina evokes induction of the IFN system but limited inflammatory responses to allow control of HSV2 infections <i>in vivo</i>.</p></div

IgG1 Allotypes Influence the Pharmacokinetics of Therapeutic Monoclonal Antibodies through FcRn Binding

International audienceBecause IgG1 allotypes might have different half-lives, their influence on infliximab (G1m17,1 allotype) pharmacokinetics was investigated in a group of spondyloarthritis patients. Infliximab was found to have a shorter half-life in patients homozygous for the G1m17,1 allotypes than in those carrying the G1m3 with no G1m1 (G1m3,-1) allotype. Because the neonatal FcR (FcRn) is involved in the pharmacokinetics of mAbs, the interaction of different IgG1 allotypes with FcRn was examined using cellular assays and surface plasmon resonance. G1m17,1 mAbs, such as infliximab and rituximab, were shown to bind more efficiently to FcRn and to be transcytosed better than the G1m3,-1 mAb cetuximab, which explains why infliximab is a better competitor for endogenous IgG1 in G1m3,-1 allotype-bearing patients. A set of four allotype variants of adalimumab (G1m17,1; G1m17,-1; G1m3,1; and G1m3,-1) was also tested for its binding to FcRn, revealing that the G1m3,1 variant, not present in commercial mAbs, binds more efficiently to FcRn and is transcytosed better than the other three variants, all of which are found in therapeutic mAbs

STING agonists induce IFN responses in the vaginal epithelium faster and more efficiently than TLR agonists.

<p>(<b>A</b>) HaCaT cells were treated with imiquimod (1μg/ ml), ODN2216 (1μg/ ml), and 3’3’-cAIMP (100μg/ ml) for 24 h. Levels of ISG15 and viperin were determined in the cell lysate by Western blotting. (<b>B</b>) Mice were treated intravaginally with imiquimod or ODN1826 (25 μg per mouse) 12 h prior to infection with HSV2. Vaginal washes were collected 48 h p.i. and viral load was determined. n = 5 per group. (<b>C-D</b>) Mice were anesthetized for 30 min and imiquimod, ODN1826, or 3’3’-cAIMP was applied to the vagina. Tissues were isolated (<b>C</b>) 6 h or (<b>D</b>) 36 h after treatment. Paraffin sections of the vaginal tissues were stained for viperin (red). DAPI (blue) marks the nuclei and the dotted white lines mark basal membrane between the epithelium and stroma. White arrows highlight examples of viperin positive cells. L = lumen, E = epithelium, S = stroma. n = 4. One representative picture is shown for each staining. (<b>E-G</b>) RNA was isolated from vaginal tissue treated as indicated for 6 h, and levels of <i>Ifnb</i>, <i>Mx1</i>, and <i>Tnfa</i> mRNA were determined by RT-qPCR. n = 4–5. mRNA levels were normalized to <i>Gapdh</i> and shown as relative levels of expression compared to mock-treated mice. (<b>B, E-G</b>) Statistics, Kruskal-Wallis test with Dunn’s multiple comparisons test. * = p<0,05 compared to mock treated.</p

HSV2 and STING agonists induce antiviral genes in vaginal epithelium.

<p>Mice were injected i.p. with 2’3’-cGAM(PS)₂ (125 μg/mouse) or infected with HSV2 (6.7×10⁴ p.f.u.). Tissues were isolated from mice 6 h and 24 h after CDN stimulation and HSV2 infection, respectively. Paraffin sections of the vaginal tissues were stained for viperin (red) and HSV2 (green). DAPI (blue) marks the nuclei and the dotted white lines mark basal membrane between the epithelium and stroma. White arrows highlight examples of viperin positive cells, and arrowheads mark examples of HSV2-infected cells. L = lumen, E = epithelium, S = stroma. n = 3. One representative picture is shown for each staining and treatment group.</p