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A composite MyD88/CD40 switch synergistically activates mouse and human dendritic cells for enhanced antitumor efficacy
Priyadharshini Narayanan, … , Kevin M. Slawin, David M. Spencer
Priyadharshini Narayanan, … , Kevin M. Slawin, David M. Spencer
Published March 7, 2011
Citation Information: J Clin Invest. 2011;121(4):1524-1534. https://doi.org/10.1172/JCI44327.
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Research Article Article has an altmetric score of 9

A composite MyD88/CD40 switch synergistically activates mouse and human dendritic cells for enhanced antitumor efficacy

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Abstract

The in vivo therapeutic efficacy of DC-based cancer vaccines is limited by suboptimal DC maturation protocols. Although delivery of TLR adjuvants systemically boosts DC-based cancer vaccine efficacy, it could also increase toxicity. Here, we have engineered a drug-inducible, composite activation receptor for DCs (referred to herein as DC-CAR) comprising the TLR adaptor MyD88, the CD40 cytoplasmic region, and 2 ligand-binding FKBP12 domains. Administration of a lipid-permeant dimerizing ligand (AP1903) induced oligomerization and activation of this fusion protein, which we termed iMyD88/CD40. AP1903 administration to vaccinated mice enabled prolonged and targeted activation of iMyD88/CD40-modified DCs. Compared with conventionally matured DCs, AP1903-activated iMyD88/CD40-DCs had increased activation of proinflammatory MAPKs. AP1903-activated iMyD88/CD40-transduced human or mouse DCs also produced higher levels of Th1 cytokines, showed improved migration in vivo, and enhanced both antigen-specific CD8+ T cell responses and innate NK cell responses. Furthermore, treatment with AP1903 in vaccinated mice led to robust antitumor immunity against preestablished E.G7-OVA lymphomas and aggressive B16.F10 tumors. Thus, the iMyD88/CD40 unified “switch” effectively and safely replaced exogenous adjuvant cocktails, allowing remote and sustained DC activation in vivo. DC “licensing” through iMyD88/CD40 may represent a mechanism by which to exploit the natural synergy between the TLR and CD40 signaling pathways in DCs using a single small molecule drug and could augment the efficacy of antitumor DC-based vaccines.

Authors

Priyadharshini Narayanan, Natalia Lapteva, Mamatha Seethammagari, Jonathan M. Levitt, Kevin M. Slawin, David M. Spencer

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Figure 2

iMyD88/CD40 induces potent activation of primary BMDCs.

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iMyD88/CD40 induces potent activation of primary BMDCs.
(A) Upon CID tre...
(A) Upon CID treatment, iMyD88/CD40 upregulates immunostimulatory molecules on BMDCs. Immature BMDCs were transduced with 100 MOI Ad and stimulated with CID (100 nM), LPS (1 μg/ml), or soluble CD40L (1 μg/ml) for 48 hours, followed by flow cytometry to measure surface expression of CD40, CD86, and MHC II. Histogram numbers represent percentages of CD40+, CD86+, or MHC II+ cells within the CD11c+ gate. (B) CID-induced iMyD88/CD40 activation induces robust proinflammatory cytokine production by BMDCs. Supernatants from Ad-transduced BMDCs, stimulated with CID, LPS, or recombinant mouse CD40 ligand (5 μg/ml) for 48 hours, were analyzed in duplicate in a multiplex cytokine bead assay. *P < 0.05 (1-tailed); ***P < 0.0005 (2-tailed). Similar results were obtained for mouse soluble (PeproTech Inc.) and multimeric (Axxora, LLC) CD40L. (C) iMyD88/CD40 activates proinflammatory signaling molecules in BMDCs. 48 hours after Ad treatment, BMDCs were stimulated with CID, LPS, or recombinant CD40L for 30 minutes. Cell lysates were harvested and analyzed using antibodies to phosphorylated and unphosphorylated IKKα/β, p38, JNK, ERK, and Akt. (D) iMyD88/CD40 induces synergistic activation of signaling molecules in human THP-1 cells. THP-1 cells were treated with Ad as listed in part C and activated and analyzed as above. Data are representative of at least 3 (A, B, and D) or 2 (C) independent experiments.

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