Adenovirus serotype 5 infects human dendritic cells via a coxsackievirus–adenovirus receptor-independent receptor pathway mediated by lactoferrin and DC-SIGN

Abstract

The coxsackievirus-adenovirus receptor (CAR) is the described primary receptor for adenovirus serotype 5 (Ad5), a common human pathogen that has been exploited as a viral vector for gene therapy and vaccination. This study showed that monocytes and dendritic cells (DCs), such as freshly isolated human blood myeloid DCs, plasmacytoid DCs and monocyte-derived DCs, are susceptible to recombinant Ad5 (rAd5) infection despite their lack of CAR expression. Langerhans cells and dermal DCs from skin expressed CAR, but blocking CAR only partly decreased rAd5 infection, together suggesting that other receptor pathways mediate viral entry of these cells. Lactoferrin (Lf), an abundant protein in many bodily fluids known for its antiviral and antibacterial properties, promoted rAd5 infection in all cell populations except plasmacytoid DCs using a CAR-independent process. Lf caused phenotypic differentiation of the DCs, but cell activation played only a minor role in the increase in infection frequencies. The C-type lectin receptor DC-SIGN facilitated viral entry of rAd5-Lf complexes and this was dependent on highmannose-type N-linked glycans on Lf. These results suggest that Lf present at high levels at mucosal sites can facilitate rAd5 attachment and enhance infection of DCs. A better understanding of the tropism and receptor mechanisms of Ad5 may help explain Ad5 pathogenesis and guide the engineering of improved rAd vectors. INTRODUCTIONAdenoviruses are frequent causes of acute upper respiratory tract infections and can be responsible for ocular and gastrointestinal illnesses in humans. They have also been exploited for the development of replication-incompetent viral vectors because their genomes allow large inserts of expression cassettes, thereby offering efficient gene delivery. Recombinant adenoviruses (rAds), serotype 5 (rAd5) in particular, are commonly used vectors for gene therapy and vaccination trials (Barouch & Nabel, 2005;McConnell & Imperiale, 2004;Tatsis & Ertl, 2004). The primary receptor described for infection of most Ad species (A, C, D, E and F) is the coxsackievirus and adenovirus receptor (CAR) (Bergelson et al., 1997;Roelvink et al., 1998;Tomko et al., 1997). As currently understood, infection of Ad5 (species C) involves binding of the viral fiber knob to CAR on the target cells (Roelvink et al., 1996), followed by an interaction between the viral penton base with a v integrins on the cell surface (Wickham et al., 1993). This allows virus-receptor complexes to enter clathrin-coated pits via endocytosis Wang et al., 1998;Wickham et al., 1993). Efficient infection of Ad5 via CAR has mainly been demonstrated in vitro using immortalized cell lines with well-exposed CAR. However, CAR is naturally expressed in tight junctions between cells and therefore et al., 2007). In this study, we investigated the involvement of CAR and the influence of Lf in rAd5 infection in relevant primary human APC populations from both blood and skin. METHODSIsolation of blood DCs. This study was approved by the ethical commi...

Adenovirus type-35 vectors block human CD4 + T-cell activation via CD46 ligation

“…Freshly isolated immature CD11c + myeloid DCs (MDCs), CD123 + PDCs, and CD1a + monocyte-derived DCs (MDDCs) were exposed to rAd5 or rAd35 encoding a GFP transgene over a range of infectious particles per cell (ip/cell). The frequency of GFP + cells was used as a measure of infection after 24 h. In agreement with our previous reports (4,10), rAd35 infected all DC subsets more efficiently than rAd5 (Fig. 1A).…”

“…All cells were cultured in complete media [CM; containing RPMI 1640, 10% (vol/vol) FBS, L-glutamine, penicillin, and streptomycin; Sigma-Aldrich] at 37°C. Primary human MDCs and PDCs were isolated using anti-CD1c and anti-BDCA-4 microbead kits, respectively, on an Automacs (Miltenyi) and cultured as previously described (4,10,40 Intracellular Cytokine Staining. Staining of intracellular cytokines was performed as previously described (10).…”

“…Ad5 and Ad35 are distinguished by their use of cellular receptors. Ad5 interacts with coxsackieadenovirus receptor (CAR) (3) and several soluble factors, including lactoferrin (4,5) and coagulation factor X (6, 7), which enhance binding to host cells. In contrast, Ad35 binds CD46 with high affinity and avidity (8).…”

Therapeutic Applications of Camel’s Milk and Urine against Cancer: Current Development Efforts and Future Perspectives

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Adenovirus-triggered innate signalling pathways

“…As these cells possess strong phagocytic activity, Ad entry into them is generally believed to be mediated by phagocytosis. Indeed, macrophage receptors, such as Fc receptor and DC-Sign have been shown to facilitate cellular binding requiring anti Ad antibodies and lactoferrin, respectively [67][68][69]. If modified by an Fc fusion protein, they can directly attach to the high affinity FcR CD64 [70].…”