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Although adenoviruses (AdVs) possess advantageous features as vectors, several challenges remain. These include a high prevalence of neutralizing antibodies against certain AdV types and the inability to efficiently transduce CAR-deficient cells and tissues. We showed previously that lactoferricin (Lfcin) enhances CAR-independent HAdV-C5 infection of epithelial and T-cells. Here, we assessed the ability of Lfcin to enable HAdV-C5 infection and transduction of human skeletal muscle cells. Lfcin increases HAdV-C5 infection and transduction of muscle myoblasts and myotubes by 10- to 30-fold. Enhanced infection correlates with increased cell binding, which differs mechanistically from that of coagulation factor X-mediated binding, as it remains unaffected by the removal of heparan sulfate. Additionally, Lfcin reduces the neutralizing effects of serum against HAdV-C5, suggesting it may shield key epitopes. By enabling viral binding to muscle cells and mitigating serum neutralization, Lfcin offers a novel strategy to improve the efficiency and durability of HAdV-C5-based gene delivery systems.

Most adenoviruses attach to host cells by means of the protruding fiber protein that binds to host cells via the coxsackievirus and adenovirus receptor (CAR) protein. Human adenovirus type 52 (HAdV-52) is one of only three gastroenteritis-causing HAdVs that are equipped with two different fiber proteins, one long and one short. Here we show, by means of virion-cell binding and infection experiments, that HAdV-52 can also attach to host cells via CAR, but most of the binding depends on sialylated glycoproteins. Glycan microarray, flow cytometry, surface plasmon resonance and ELISA analyses reveal that the terminal knob domain of the long fiber (52LFK) binds to CAR, and the knob domain of the short fiber (52SFK) binds to sialylated glycoproteins. X-ray crystallographic analysis of 52SFK in complex with 2-O-methylated sialic acid combined with functional studies of knob mutants revealed a new sialic acid binding site compared to other, known adenovirus: glycan interactions. Our findings shed light on adenovirus biology and may help to improve targeting of adenovirus-based vectors for gene therapy.