Antiviral immunity, Biotherapy and Vaccines   


  HEADDr. Marie-Lise GOUGEON / marie-lise.gougeon@pasteur.fr
  MEMBERSDr. Maxime BATTISTELLA (MD, graduate student), Dr. Marlène BRAS (PhD), Maryse BRANDT (Secretary), Pauline FORMAGLIO (Graduate student), Emily LOISON (PhD student), Marie-Thérèse MELKI (PhD student), Dr. Béatrice POIRIER-BEAUDOUIN (Engineer), Valérie SEFFER (Technician), Evelyne ROSSIGNOL (Laboratory assistant), Dr. Héla SAIDI (PhD)


  Annual Report

Our research aims to understand the interplay between innate and adaptive immunity in chronic viral infections, in order to identifymechanisms responsible for the lack of immune control of persistent viruses, such as HIV. In particular, we dissect the modalities of the cross-talk between innate effectors, i.e. dendritic cells (DCs) and natural killer (NK) cells, and their impact on adaptive immunity. Moreover, we are deeply involved in immunomonitoring of vaccine immunity in phase I and II vaccine trials, evaluating new vaccine candidates specific for influenza, malaria, shigellosis, anthrax or cancer.

Interplay between innate and adaptive immunity against persistent viruses.

NK-DC cross-talk, through HMGB1, promotes HIV replication in infected DCs and contributes to the establishment of viral reservoirs

HIV-1 has evolved ways to exploit DCs, thereby facilitating viral dissemination and allowing evasion of antiviral immunity. The fate of DCs is extremely dependent on the interaction with autologous NK cells, but the mechanisms by which NK-DC interaction controls viral infections remain unclear. We have investigated the impact of NK-DC cross-talk on maturation and functions of HIV-infected immature DCs, and found that activated NK cells are required for the induction of DC maturation, whether HIV-1-infectedor not, and this process involves the alarmin HMGB1. However, derived mature infected DCs show a strong impairment in their ability to induce Th1 polarization of naïve CD4 T cells. This is associated with the defective production of IL-12 and IL-18 by infected DCs. Moreover, NK-DC cross-talk results in a dramatic increase in HIV viral replication and proviral DNA expression in DCs, and HMGB1 has a pivotal role in this process. These observations provide evidence for the crucial role of NK-DC cross-talk in promoting HIV dissemination, and they challenge the question of the in vivoinvolvement of HMGB1 in the replenishment of viral reservoirs.

HIV-infected DCs are resistant to NK-mediated killing. HMGB1-dependent inhibition of DC apoptosis

NK cells are able to kill virus-infected cells without previous sensitization, and during viral infections they are rapidly recruited to inflamed tissues where they should destroy infected DCs. We addressed the question of the susceptibility of HIV-infected DCs to NK-mediated cytotoxicity, and found that, whereas NK cells are able to kill non infected immature DCs (i.e. “editing process” to keep in check the quality of DCs)(Figure),they fail to eliminate infected DCs. The resistance of infected DCs to TRAIL-mediated NK killing is associated with the upregulation of two anti-apoptotic molecules, c-FLIP and c-IAP2, induced in DCs by HMGB1 released at NK-DC synapse. Blocking HMGB1 activity by specific antibodies or glycyrrhizin (a specific inhibitor of HMGB1)restoresthe susceptibility of infected DCs to NK killing. These observations provide evidence for the pivotal role of HMGB1 in the survival of infected DCs and their escape from cytotoxicity, thus contributing to viral persistence and dissemination.

Immunomonitoring Platform for the ex-vivoanalysis of vaccine-induced immunity and in vivoevaluation of new vaccine candidates

The objectives are to follow-up innate and adaptive immunity in healthy volunteers or patients involved in the evaluation of new vaccine candidates or in new therapies. The monitoring of specific immune effectors is performed using complementary methods, including multicolor flow cytometry, intracellular cytokine staining, ELISPOT, and cytokine multiplex analyses, with the aim to quantify the frequency of vaccine-specific cells and determine the cytokine/chemokine signatures of vaccine responses. Monitoring the antibody response induced by a vaccine candidates with adapted ELISA tests and quantifying the frequency of Ig-producing B cells by ELISPOT is also performed by our laboratory in the context of phase I or II trials. We also characterize innate effectors, such as NK cells and DC, and analyze their stage of differentiation and their effectors/regulatory functions. This platform has recently evaluated, in a phase II trial, the effectiveness of an oral vaccine (developed by P. Sansonetti’s team at Institut Pasteur) against Shigella dysenteriae serotype 1, the bacterium responsible for the epidemic form of Shigellosis or bacillary dysentery (Launay O et al. Vaccine, 2009, 27:1184-91). We arecurrently involved in the in vivoanalysis of Flu-specific cellular immune response directed against seasonal vaccine and H1N1 pandemic vaccine.

Keywords: HIV, innate and adaptive immunity, NK cells, dendritic cells, apoptosis, AIDS, Immunomonitoring, Clinical trials

gougeon.jpg

(A) NK-DC cross-talk (NK and DCs were stained with red and green mitotracker, respectively), leading to apoptosis of DCs (B). (C) NK-DC cross-talk involves the alarmin HMGB1 (DCs were costained with CD40 and HMGB1 specific Abs).



  Publications

1. Launay O, Sadorge C, Jolly N, Poirier B, Béchet S, van der Vliet D, Seffer V, Fenner N, Dowling K, Giemza R, Johnson J, Ndiaye A, Vray M, Sansonetti P, Morand P, Poyart C, Lewis D, Gougeon M-L (2009). Safety and immunogenicity of SC599, an oral live attenuated Shigella dysenteriae type-1 vaccine in healthy volunteers: Results of a Phase 2, randomized, double-blind placebo-controlled trial. Vaccine,27(8):1184-111 (PMID: 19135496).

2. Saïdi H, Melki MT, Gougeon M-L. (2008) HMGB1-dependent triggering of HIV-1 replication and persistence in dendritic cells as a consequence of NK-DC cross-talk. PLoS One,3(10):e3601 (PMID: 8974890).

3. Bristeau-Leprince A, Mateo V, Lim A, Magerus-Chatinet A, Solary E, Fischer A, Rieux-Laucat F, Gougeon M-L (2008). Human TCR alpha/beta+CD4-CD8-double-negative T cells in patients with autoimmune lymphoproliferative syndrome express restricted Vbeta TCR diversity and are clonally related to CD8+T cells. Journal of Immunology181(1):440-448 (PMID: 18566410).

4. Lecoeur H, Melki M-T, Saïdi H, Gougeon M-L (2008). Analysis of apoptotic pathways by multiparametric flow cytometry: application to HIV infection. Methods in Enzymology442:51-82 (PMID:18662564).

5- Fazilleau N, Bachelez H, Gougeon M-L, Viguier M (2007). Cutting edge: size and diversity of CD4+CD25highFoxp3+ regulatory T cell repertoire in humans: evidence for similarities and partial overlapping with CD4+CD25- T cells. Journal of Immunology179(6):3412-3416 (PMID:17785774).





Activity Reports 2009 - Institut Pasteur
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