Ex Vivo and In Vivo CD46 Receptor Utilization by Species D Human Adenovirus Serotype 26 (HAdV26)

Human adenovirus serotype 26 (Ad26) is used as a gene-based vaccine against SARS-CoV-2 and HIV-1. Yet, its primary receptor portfolio remains controversial, potentially including sialic acid, CAR, integrins, and CD46. We and others have shown that Ad26 can use CD46, but these observations were questioned by the inability to co-crystallize Ad26 fiber with CD46. Recent work demonstrated that Ad26 binds CD46 with its hexon protein rather than its fiber. We examined the functional consequences of Ad26 for infection in vitro and in vivo. Ectopic expression of human CD46 on Chinese hamster ovary cells increased Ad26 infection significantly. Deletion of the complement control protein domains CCP1 or CCP2 or the serine-threonine-proline (STP) region of CD46 reduced infection. Comparing wild type and sialic acid-deficient CHO cells, we show that the usage of CD46 is independent of its sialylation status. Ad26 transduction was increased in CD46 transgenic mice after intramuscular (IM) injection, but not after intranasal (IN) administration. Ad26 transduction was 10-fold lower than Ad5 after intratumoral (IT) injection of CD46-expressing tumors. Ad26 transduction of liver was 1000-fold lower than Ad5 after intravenous (IV) injection. These data demonstrate the use of CD46 by Ad26 under certain situations, but also show that the receptor has little consequence by other routes of administration. Finally, IV injection of high doses of Ad26 into CD46 mice induced release of liver enzymes in the bloodstream and reduced white blood cell counts, but did not induce thrombocytopenia. This suggests that Ad26 virions do not induce direct clotting side effects seen during COVID-19 vaccination with this serotype of adenovirus. IMPORTANCEHuman species D Ad26 is being pursued as a low seroprevalence vector for oncolytic virotherapy and gene-based vaccination against HIV-1 and SARS-CoV-2. However, there is debate in the literature about its tropism and receptor utilization, which directly influence its efficiency for certain applications. This work was aimed at determining which receptor(s) this virus uses for infection, and its role in virus biology, vaccine efficacy, and importantly, in vaccine safety.