Intratracheal Gene Delivery with Adenoviral Vector Induces Elevated Systemic IgG and Mucosal IgA Antibodies to Adenovirus and β -Galactosidase

Abstract

One major concern about using adenoviral vectors for repetitive gene delivery to lung epithelial cells is the induction of an immune response to the vector, thus, impeding effective gene transduction. To assess the immune response to the adenoviral vector, repetitive intratracheal (i.t.) gene dosing was performed in CD-1 mice using the replication-deficient adenovirus 5 (Ade5) vector carrying the lacZ gene, and compared to the antibody responses induced by conventional intranasal (i.n.) and intraperitoneal (i.p.) routes of immunization. Kinetics of serum IgG, IgA, and IgM antibody responses to the adenoviral vector and to β-galactosidase (β-Gal) were evaluated. Two or three adenoviral vector doses given by i.t., i.n., or i.p. routes resulted in serum IgG titers in excess of 1:200,000, whereas serum IgM and IgA were moderately induced. Analysis of the predominant murine IgG subclass was determined to be IgG_2b and IgG_2a. To determine the localization of this antibody response, the ELISPOT assay was employed. Lymphocytes were isolated from the lung, the lower respiratory lymph nodes (LRLN), the nasal passages (NP), and the spleen. For i.t- and i.n.-administered mice, the highest IgA spot-forming cell (SFC) response to Ade5 and β-Gal was located in the NP and in the lung. Both the lung and the LRLN showed elevated numbers of IgG SFCs (4- to 12-fold greater than splenic IgG SFC response) for Ade5 and β-Gal. This evidence suggests that the lung and associated lymphoid tissues were the source for serum antibodies. Further analysis of serum antibodies showed that the i.p.- and i.t.-administered groups yielded the greatest neutralization titers to Ade5, suggesting that the reduced effectiveness of repetitive gene transfer is in part due to circulating neutralizing antibodies. Thus, repetitive i.t. instillation will stimulate a localized and systemic antibody response to the vector.

Overview summary

I.t. delivery of an adenoviral vector into CD-1 mice induces both a localized and systemic antibody response to the adenoviral vector. Elevated IgG anti-Ade5 antibodies were detected in serum following two or three i.t. doses, and were of equal magnitude to the antibody response elicited by i.n. or i.p. adenoviral vector administration. Serum IgA anti-Ade5 antibodies were also detected. Antibodies to β-galactosidase were induced and were of equal magnitude with anti-Ade5 antibody titers. The immune sera from i.t.-dosed group did exhibit neutralization activity, and inhibited adenoviral vector infection of 293 cells. The elicited immune antibodies were derived from antibody-producing cells in the lung, LRLN, NP, and spleen. Thus, repetitive adenoviral vector gene transfer will induce a host antibody response capable of diminishing effective gene transfer.