1. ¹Center for Cell and Gene Therapy, Baylor College of Medicine, Texas Children's Hospital, The Methodist Hospital, Houston, TX, USA
2. ²Department of Bioengineering, Rice University, Houston, TX 77005, USA
3. ³Department of Molecular and Human Genetics, Baylor College of Medicine, 1 Baylor Plaza, Houston, TX 77030, USA
4. ⁴Department of Microbiology and Immunology, Baylor College of Medicine, 1 Baylor Plaza, Houston, TX 77030, USA

Correspondence: Michael A. Barry, Work Departments of Immunology and Molecular and Human Genetics, Baylor College of Medicine, Center for Cell and Gene Therapy, 1 Baylor Plaza, Houston, TX 77030, USA. Fax: +1 713 798 1481. E-mail: mab@bcm.tmc.edu

Received 13 May 2004; Accepted 6 September 2004.

Abstract

Adenoviruses are robust gene delivery vectors in vivo, but are limited by their propensity to provoke strong innate and adaptive responses. Previous work has demonstrated that polyethylene glycol (PEG) modification of adenovirus can protect the vectors from preexisting and adaptive immune responses by reducing protein–protein interactions. To test whether PEGylation can reduce innate immune responses to adenovirus by reducing their interactions with immune cells, first-generation (FG-Ad) and helper-dependent (HD-Ad) Ad5 vectors were PEGylated with SPA-PEG and tested in vitro and in vivo. We demonstrate that increasing PEGylation ablated in vitro transduction, but surprisingly had no negative effect on the level or distribution of in vivo gene delivery. This poor in vitro transduction could be rescued in part by physically forcing the PEGylated vectors onto cells, suggesting that physiological forces in vivo may enable transduction via heparin sulfate proteoglycan and integrin interactions. While transduction remained the same as for unmodified vectors, the PEGylated vectors reduced innate IL-6 responses by 70 and 50% in vivo for FG-Ad and HD-Ad. These reduced innate responses paralleled similar reductions in vector uptake by macrophages in vitro and Kupffer cells in vivo. These data suggest that PEGylation of Ad vectors can reduce innate immune responses without reducing transduction in vivo. These data also suggest that nonspecific vector uptake by macrophages and Kupffer cells may be critically involved in the initial activation of innate immune responses.