Gerard Zurawski Ph.D.

Ceglia, V., S. Zurawski, M. Montes, A. Bouteau, Z. Wang, J. Ellis, B. Z. Igyártó, Y. Lévy and G. Zurawski (2021). “Anti-CD40 Antibodies Fused to CD40 Ligand Have Superagonist Properties.” J Immunol 207(8): 2060-2076.

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CD40 is a potent activating receptor within the TNFR family expressed on APCs of the immune system, and it regulates many aspects of B and T cell immunity via interaction with CD40 ligand (CD40L; CD154) expressed on the surface of activated T cells. Soluble CD40L and agonistic mAbs directed to CD40 are being explored as adjuvants in therapeutic or vaccination settings. Some anti-CD40 Abs can synergize with soluble monomeric CD40L. We show that direct fusion of CD40L to certain agonistic anti-CD40 Abs confers superagonist properties, reducing the dose required for efficacy, notably greatly increasing total cytokine secretion by human dendritic cells. The tetravalent configuration of anti-CD40-CD40L Abs promotes CD40 cell surface clustering and internalization and is the likely mechanism of increased receptor activation. CD40L fused to either the L or H chain C termini, with or without flexible linkers, were all superagonists with greater potency than CD40L trimer. The increased anti-CD40-CD40L Ab potency was independent of higher order aggregation. Moreover, the anti-CD40-CD40L Ab showed higher potency in vivo in human CD40 transgenic mice compared with the parental anti-CD40 Ab. To broaden the concept of fusing agonistic Ab to natural ligand, we fused OX40L to an agonistic OX40 Ab, and this resulted in dramatically increased efficacy for proliferation and cytokine production of activated human CD4(+) T cells as well as releasing the Ab from dependency on cross-linking. This work shows that directly fusing antireceptor Abs to ligand is a useful strategy to dramatically increase agonist potency.

Ceglia, V., S. Zurawski, M. Montes, A. Bouteau, Z. Wang, J. Ellis, B. Z. Igyártó, Y. Lévy and G. Zurawski (2021). “Anti-CD40 Antibodies Fused to CD40 Ligand Have Superagonist Properties.” J Immunol 207(8): 2060-2076.

Full text of this article.

CD40 is a potent activating receptor within the TNFR family expressed on APCs of the immune system, and it regulates many aspects of B and T cell immunity via interaction with CD40 ligand (CD40L; CD154) expressed on the surface of activated T cells. Soluble CD40L and agonistic mAbs directed to CD40 are being explored as adjuvants in therapeutic or vaccination settings. Some anti-CD40 Abs can synergize with soluble monomeric CD40L. We show that direct fusion of CD40L to certain agonistic anti-CD40 Abs confers superagonist properties, reducing the dose required for efficacy, notably greatly increasing total cytokine secretion by human dendritic cells. The tetravalent configuration of anti-CD40-CD40L Abs promotes CD40 cell surface clustering and internalization and is the likely mechanism of increased receptor activation. CD40L fused to either the L or H chain C termini, with or without flexible linkers, were all superagonists with greater potency than CD40L trimer. The increased anti-CD40-CD40L Ab potency was independent of higher order aggregation. Moreover, the anti-CD40-CD40L Ab showed higher potency in vivo in human CD40 transgenic mice compared with the parental anti-CD40 Ab. To broaden the concept of fusing agonistic Ab to natural ligand, we fused OX40L to an agonistic OX40 Ab, and this resulted in dramatically increased efficacy for proliferation and cytokine production of activated human CD4(+) T cells as well as releasing the Ab from dependency on cross-linking. This work shows that directly fusing antireceptor Abs to ligand is a useful strategy to dramatically increase agonist potency.

Godot, V., Tcherakian, C., Gil, L., Cervera-Marzal, I., Li, G., Cheng, L., Ortonne, N., Lelièvre, J.D., Pantaleo, G., Fenwick, C., Centlivre, M., Mouquet, H., Cardinaud, S., Zurawski, S.M., Zurawski, G., Milpied, P., Su, L. and Lévy, Y. (2020). “TLR-9 agonist and CD40-targeting vaccination induces HIV-1 envelope-specific B cells with a diversified immunoglobulin repertoire in humanized mice.” PLoS Pathog 16(11): e1009025.

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The development of HIV-1 vaccines is challenged by the lack of relevant models to accurately induce human B- and T-cell responses in lymphoid organs. In humanized mice reconstituted with human hematopoietic stem cells (hu-mice), human B cell-development and function are impaired and cells fail to efficiently transition from IgM B cells to IgG B cells. Here, we found that CD40-targeted vaccination combined with CpG-B adjuvant overcomes the usual defect of human B-cell switch and maturation in hu-mice. We further dissected hu-B cell responses directed against the HIV-1 Env protein elicited by targeting Env gp140 clade C to the CD40 receptor of antigen-presenting cells. The anti-CD40.Env gp140 vaccine was injected with CpG-B in a homologous prime/boost regimen or as a boost of a NYVAC-KC pox vector encoding Env gp140 clade C. Both regimens elicited Env-specific IgG-switched memory hu-B cells at a greater magnitude in hu-mice primed with NYVAC-KC. Single-cell RNA-seq analysis showed gp140-specific hu-B cells to express polyclonal IgG1 and IgG3 isotypes and a broad Ig VH/VL repertoire, with predominant VH3 family gene usage. These cells exhibited a higher rate of somatic hypermutation than the non-specific IgG+ hu-B-cell counterpart. Both vaccine regimens induced splenic GC-like structures containing hu-B and hu-Tfh-like cells expressing PD-1 and BCL-6. We confirmed in this model that circulating ICOS+ memory hu-Tfh cells correlated with the magnitude of gp140-specific B-cell responses. Finally, the NYVAC-KC heterologous prime led to a more diverse clonal expansion of specific hu-B cells. Thus, this study shows that CD40-targeted vaccination induces human IgG production in hu-mice and provides insights for the development of a CD40-targeting vaccine to prevent HIV-1 infection in humans.

Fovet, C. M., L. Stimmer, V. Contreras, P. Horellou, A. Hubert, N. Seddiki, C. Chapon, S. Tricot, C. Leroy, J. Flament, J. Massonneau, N. Tchitchek, B. A. t Hart, S. Zurawski, P. Klucar, P. Hantraye, K. Deiva, G. Zurawski, S. Oh, R. Le Grand and C. Serguera (2019). “Intradermal vaccination prevents anti-MOG autoimmune encephalomyelitis in macaques.” EBioMedicine Sep 3. [Epub ahead of print].

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BACKGROUND: Autoimmune demyelinating diseases (ADD) are a major cause of neurological disability due to autoreactive cellular and humoral immune responses against brain antigens. A cure for chronic ADD could be obtained by appropriate immunomodulation. METHODS: We implemented a preclinical scheme to foster immune tolerance to myelin oligodendrocyte glycoprotein (MOG), in a cynomolgus-macaque model of experimental autoimmune encephalomyelitis (EAE), in which administration of recombinant human MOG (rhMOG) elicits brain inflammation mediated by MOG-autoreactive CD4(+) lymphocytes and anti-MOG IgG. For immunotherapy, we used a recombinant antibody (Ab) directed against the dendritic cell-asialoglycoprotein receptor (DC-ASGPR) fused either to MOG or a control antigen PSA (prostate-specific antigen). FINDINGS: rhMOG and the anti-DC-ASGPR-MOG were respectively detected in CD1a(+) DCs or CD163(+) cells in the skin of macaques. Intradermal administration of anti-DC-ASGPR-MOG, but not control anti-DC-ASGPR-PSA, was protective against EAE. The treatment prevented the CD4(+) T cell activation and proinflammatory cytokine production observed in controls. Moreover, the administration of anti-DC-ASGPR-MOG induced MOG-specific CD4(+)CD25(+)FOXP3(+)CD39(+) regulatory lymphocytes and favoured an upsurge in systemic TGFbeta and IL-8 upon rhMOG re-administration in vivo. INTERPRETATION: We show that the delivery of an anti-DC-ASGPR-MOG allows antigen-specific adaptive immune modulation to prevent the breach of immune tolerance to MOG. Our findings pave the way for therapeutic vaccines for long-lasting remission to grave encephalomyelitis with identified autoantigens, such as ADD associated with anti-MOG autoantibodies. FUND: Work supported by the French ANR (ANR-11-INBS-0008 and ANR-10-EQPX-02-01), NIH (NIH 1 R01 AI 105066), the Baylor Scott and White Healthcare System funding and Roche Research Collaborative grants.

Flamar, A. L., H. Bonnabau, S. Zurawski, C. Lacabaratz, M. Montes, L. Richert, A. Wiedemann, L. Galmin, D. Weiss, A. Cristillo, L. Hudacik, A. Salazar, C. Peltekian, R. Thiebaut, G. Zurawski and Y. Levy (2018). “HIV-1 T cell epitopes targeted to Rhesus macaque CD40 and DCIR: A comparative study of prototype dendritic cell targeting therapeutic vaccine candidates.” PLoS One 13(11): e0207794.

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HIV-1 infection can be controlled by anti-retroviral drug therapy, but this is a lifetime treatment and the virus remains latent and rapidly rebounds if therapy is stopped. HIV-1-infected individuals under this drug regimen have increased rates of cancers, cardiovascular diseases, and autoimmunity due to compromised immunity. A therapeutic vaccine boosting cellular immunity against HIV-1 is therefore desirable and, possibly combined with other immune modulating agents, could obviate the need for long-term drug therapies. An approach to elicit strong T cell-based immunity is to direct virus protein antigens specifically to dendritic cells (DCs), which are the key cell type for controlling immune responses. For eliciting therapeutic cellular immunity in HIV-1-infected individuals, we developed vaccines comprised of five T cell epitope-rich regions of HIV-1 Gag, Nef, and Pol (HIV5pep) fused to monoclonal antibodies that bind either, the antigen presenting cell activating receptor CD40, or the endocytic dendritic cell immunoreceptor DCIR. The study aimed to demonstrate vaccine safety, establish efficacy for broad T cell responses in both primed and naive settings, and identify one candidate vaccine for human therapeutic development. The vaccines were administered to Rhesus macaques by intradermal injection with poly-ICLC adjuvant. The animals were either i) naive or, ii) previously primed with modified vaccinia Ankara vector (MVA) encoding HIV-1 Gag, Pol, and Nef (MVA GagPolNef). In the MVA-primed groups, both DC-targeting vaccinations boosted HIV5pep-specific blood CD4+ T cells producing multiple cytokines, but did not affect the MVA-elicited CD8+ T cell responses. In the naive groups, both DC-targeting vaccines elicited antigen-specific polyfunctional CD4+ and CD8+ T cell responses to multiple epitopes and these responses were unchanged by a subsequent MVA GagPolNef boost. In both settings, the T cell responses elicited via the CD40-targeting vaccine were more robust and were detectable in all the animals, favoring further development of the CD40-targeting vaccine for therapeutic vaccination of HIV-1-infected individuals.