Variability of Human Systemic Humoral Immune Responses to Adenovirus Gene Transfer Vectors Administered to Different Organs

Abstract

Administration of adenovirus (Ad) vectors to immunologically naive experimental animals almost invariably results in the induction of systemic anti-Ad neutralizing antibodies. To determine if the human systemic humoral host responses to Ad vectors follow a similar pattern, we evaluated the systemic (serum) anti-Ad serotype 5 (Ad5) neutralizing antibodies in humans after administration of first generation (E1− E3−) Ad5-based gene transfer vectors to different hosts. AdGVCFTR.10 (carrying the normal human cystic fibrosis [CF] transmembrane regulator cDNA) was sprayed (8 × 107 to 2 × 1010 particle units [PU]) repetitively (every 3 months or every 2 weeks) to the airway epithelium of 15 individuals with CF. AdGVCD.10 (carrying the Escherichia coli cytosine deaminase gene) was administered (8 × 108 to 8 × 109PU; once a week, twice) directly to liver metastasis of five individuals with colon cancer and by the intradermal route (8 × 107 to 8 × 109 PU, single administration) to six healthy individuals. AdGVVEGF121.10 (carrying the human vascular endothelial growth factor 121 cDNA) was administered (4 × 108 to 4 × 109.5 PU, single administration) directly to the myocardium of 11 individuals with ischemic heart disease. Ad vector administration to the airways of individuals with CF evoked no or minimal serum neutralizing antibodies, even with repetitive administration. In contrast, intratumor administration of an Ad vector to individuals with metastatic colon cancer resulted in a robust antibody response, with anti-Ad neutralizing antibody titers of 102 to >104. Healthy individuals responded to single intradermal Ad vector variably, from induction of no neutralizing anti-Ad antibodies to titers of 5 × 103. Likewise, individuals with ischemic heart disease had a variable response to single intramyocardial vector administration, ranging from minimal neutralizing antibody levels to titers of 104. Evaluation of the data from all trials showed no correlation between the peak serum neutralizing anti-Ad response and the dose of Ad vector administered (P > 0.1, all comparisons). In contrast, there was a striking correlation between the peak anti-Ad5 neutralizing antibody levels evoked by vector administration and the level of preexisting anti-Ad5 antibodies (P = 0.0001). Thus, unlike the case for experimental animals, administration of Ad vectors to humans does not invariably evoke a systemic anti-Ad neutralizing antibody response. In humans, the extent of the response is dictated by preexisting antibody titers and modified by route of administration but is not dose dependent. Since the extent of anti-Ad neutralizing antibodies will likely modify the efficacy of administration of Ad vectors, these observations are of fundamental importance in designing human gene therapy trials and in interpreting the efficacy of Ad vector-mediated gene transfer.

Immune reactivity after adenoviral‐mediated aquaporin‐1 cDNA transfer to human parotid glands

“…PBMCs were then plated into 96-well round bottom polystyrene plates at 10 5 cells/well in triplicate. The plated PBMCs were stimulated as described by Harvey et al, (1999aHarvey et al, ( , 2001 with 2.5 lg protein ml À1 Ad5 vector (AdMLPa1AT (Mastrangeli et al, 1994;Rosenfeld et al, 1991)) for 6 days or with 2.5 lg ml À1 phytohemagglutinin (PHA-P, Sigma, Cat#L0917) for 3 days at 37°C in 5% CO 2 . Cells cultured under similar conditions without any stimulation served as the negative control.…”

An adenoviral vector-based mucosal vaccine is effective in protection against botulism

“…11,51 Anti-adenovirus neutralizing antibody titer assay Anti-adenovirus neutralizing antibody titers were evaluated using a modification of a previously described method. 31,[52][53][54] Briefly, AD293 cells (Stratagene) were seeded in 96-well plates at a density of 10 4 cells/well in 200 ml of Eagle's minimum essential medium (EMEM) containing 10% fetal bovine serum (FBS) and incubated at 37 1C, 5% CO 2 overnight before infection. Mouse sera were serially twofold diluted in EMEM containing 2% FBS after heat-inactivated at 55 1C for 45 min.…”

Concordant activity of transgene expression cassettes inserted into E1, E3 and E4 cloning sites in the adenovirus genome

“…The clinical advancement of gene therapy will be greatly facilitated if pharmacokinetic data on the profiles of transgene expression can be obtained in human subjects [14,18]. Ideally, it should be possible to determine the number, distribution and fate of the genetically modified transduced, nor of the level at which the therapeutic gene will be expressed because vector biodistribution, gene transfer efficiency, gene expression level and the fate of the genetically modified cells have proven to be highly variable between individuals and with repeat dosing [19][20][21]. Mapping the distribution of genetically modified cells for most therapeutic transgenes requires euthanasia and thorough post-mortem analysis of all tissues using in situ hybridization, immunohistochemical analysis, or determination of enzymatic activity in tissue extracts.…”