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 x 10(7) to 2 x 10(10) 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 x 10(8) to 8 x 10(9) PU; once a week, twice) directly to liver metastasis of five individuals with colon cancer and by the intradermal route (8 x 10(7) to 8 x 10(9) PU, single administration) to six healthy individuals. AdGVVEGF121.10 (carrying the human vascular endothelial growth factor 121 cDNA) was administered (4 x 10(8) to 4 x 10(9.5) 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 10(2) to >10(4). Healthy individuals responded to single intradermal Ad vector variably, from induction of no neutralizing anti-Ad antibodies to titers of 5 x 10(3). Likewise, individuals with ischemic heart disease had a variable response to single intramyocardial vector administration, ranging from minimal neutralizing antibody levels to titers of 10(4). 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.

FIG. 1

Serum anti-Ad5 neutralizing antibody levels as a function of time after administration of E1⁻ Ad gene transfer vectors by various routes to different groups of individuals. Serum levels of anti-Ad5 neutralizing antibodies were quantified pretherapy (Pre) and at various time points as indicated after vector administration. Note that the scale for the titers varies among the panels. Each symbol represents a different individual; each arrow represents a single vector administration. The dose (each administration) of vector (PU) is indicated for each individual. For convenience in plotting the data, only the log dose is given; for the actual doses (e.g., 8 × 10⁷ instead of 10⁷ in panel A), see Table 2. (A) Single intradermal administration (day 1) of the Ad_GVCD.10 vector to normal individuals; (B) repeat administration (q3m × 3; days 1, 81, and 181) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF; (C) repeat administration (q15d × 4; days 1, 15, 30, and 45) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF; (D) repeat administration (q7d × 2; days 1 and 7) of the Ad_GVCD.10 vector directly to liver metastasis of individuals with colon carcinoma; (E) single direct myocardial administration (day 1) of Ad_GVVEGF.121.10 to individuals with diffuse coronary artery disease. In all panels, the dashed horizontal line represents the lower limit of detection of the assay.

FIG. 1

Serum anti-Ad5 neutralizing antibody levels as a function of time after administration of E1⁻ Ad gene transfer vectors by various routes to different groups of individuals. Serum levels of anti-Ad5 neutralizing antibodies were quantified pretherapy (Pre) and at various time points as indicated after vector administration. Note that the scale for the titers varies among the panels. Each symbol represents a different individual; each arrow represents a single vector administration. The dose (each administration) of vector (PU) is indicated for each individual. For convenience in plotting the data, only the log dose is given; for the actual doses (e.g., 8 × 10⁷ instead of 10⁷ in panel A), see Table 2. (A) Single intradermal administration (day 1) of the Ad_GVCD.10 vector to normal individuals; (B) repeat administration (q3m × 3; days 1, 81, and 181) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF; (C) repeat administration (q15d × 4; days 1, 15, 30, and 45) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF; (D) repeat administration (q7d × 2; days 1 and 7) of the Ad_GVCD.10 vector directly to liver metastasis of individuals with colon carcinoma; (E) single direct myocardial administration (day 1) of Ad_GVVEGF.121.10 to individuals with diffuse coronary artery disease. In all panels, the dashed horizontal line represents the lower limit of detection of the assay.

FIG. 1

Serum anti-Ad5 neutralizing antibody levels as a function of time after administration of E1⁻ Ad gene transfer vectors by various routes to different groups of individuals. Serum levels of anti-Ad5 neutralizing antibodies were quantified pretherapy (Pre) and at various time points as indicated after vector administration. Note that the scale for the titers varies among the panels. Each symbol represents a different individual; each arrow represents a single vector administration. The dose (each administration) of vector (PU) is indicated for each individual. For convenience in plotting the data, only the log dose is given; for the actual doses (e.g., 8 × 10⁷ instead of 10⁷ in panel A), see Table 2. (A) Single intradermal administration (day 1) of the Ad_GVCD.10 vector to normal individuals; (B) repeat administration (q3m × 3; days 1, 81, and 181) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF; (C) repeat administration (q15d × 4; days 1, 15, 30, and 45) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF; (D) repeat administration (q7d × 2; days 1 and 7) of the Ad_GVCD.10 vector directly to liver metastasis of individuals with colon carcinoma; (E) single direct myocardial administration (day 1) of Ad_GVVEGF.121.10 to individuals with diffuse coronary artery disease. In all panels, the dashed horizontal line represents the lower limit of detection of the assay.

FIG. 2

Peak systemic anti-Ad5 neutralizing antibody levels as a function of dose and frequency of administration. E1⁻ Ad vectors were administered by different routes to different groups of individuals as indicated. Each symbol represents a different individual. For convenience in plotting the data, only the log dose is given; for the actual doses (e.g., 8 × 10⁷⁻ instead of 10⁷ in panel A), see Table 2. (A) Single intradermal administration of the Ad_GVCD.10 vector to normal individuals; (B) repeat administration (q3mo × 3) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with cystic fibrosis; (C) repeat administration (q15d × 4) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF; (D) repeat administrations (q7d × 2) of the Ad_GVCD.10 vector directly to liver metastasis of individuals with colon carcinoma; (E) single direct myocardial administration of the Ad_GVVEGF.121.10 vector to individuals with diffuse coronary artery disease. In all panels, the dashed horizontal line represents the lower limit of detection of the assay. For panels B and C, the timing of the repetitive doses are indicated on the abscissa.

FIG. 2

Peak systemic anti-Ad5 neutralizing antibody levels as a function of dose and frequency of administration. E1⁻ Ad vectors were administered by different routes to different groups of individuals as indicated. Each symbol represents a different individual. For convenience in plotting the data, only the log dose is given; for the actual doses (e.g., 8 × 10⁷⁻ instead of 10⁷ in panel A), see Table 2. (A) Single intradermal administration of the Ad_GVCD.10 vector to normal individuals; (B) repeat administration (q3mo × 3) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with cystic fibrosis; (C) repeat administration (q15d × 4) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF; (D) repeat administrations (q7d × 2) of the Ad_GVCD.10 vector directly to liver metastasis of individuals with colon carcinoma; (E) single direct myocardial administration of the Ad_GVVEGF.121.10 vector to individuals with diffuse coronary artery disease. In all panels, the dashed horizontal line represents the lower limit of detection of the assay. For panels B and C, the timing of the repetitive doses are indicated on the abscissa.

FIG. 2

Peak systemic anti-Ad5 neutralizing antibody levels as a function of dose and frequency of administration. E1⁻ Ad vectors were administered by different routes to different groups of individuals as indicated. Each symbol represents a different individual. For convenience in plotting the data, only the log dose is given; for the actual doses (e.g., 8 × 10⁷⁻ instead of 10⁷ in panel A), see Table 2. (A) Single intradermal administration of the Ad_GVCD.10 vector to normal individuals; (B) repeat administration (q3mo × 3) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with cystic fibrosis; (C) repeat administration (q15d × 4) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF; (D) repeat administrations (q7d × 2) of the Ad_GVCD.10 vector directly to liver metastasis of individuals with colon carcinoma; (E) single direct myocardial administration of the Ad_GVVEGF.121.10 vector to individuals with diffuse coronary artery disease. In all panels, the dashed horizontal line represents the lower limit of detection of the assay. For panels B and C, the timing of the repetitive doses are indicated on the abscissa.

FIG. 3

Peak systemic anti-Ad5 neutralizing antibody levels as a function of anti-Ad5 neutralizing antibody titer before vector administration. E1⁻ Ad vectors were administered by different routes to different groups of individuals. Each symbol represents a different individual. (A) Single intradermal administration of the Ad_GVCD.10 vector to normal individuals; (B) repeat administration (q3mo × 3) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF. Open symbols, first administration; light gray symbols, second administration; dark gray symbols, third administration. (C) Repeat administration (q15d × 4) of the Ad_GVCFTR.10 vector to the airway epithelium of individuals with CF. Open symbols, first administration; light gray symbols, second administration; dark gray symbols, third administration; black symbols, fourth administration. (D) Repeat administration (q7d × 2) of the Ad_GVCD.10 vector directly to liver metastasis of individuals with colon carcinoma. (E) Single direct myocardial administration of the Ad_GVVEGF.121.10 vector to individuals with diffuse coronary artery disease. (F) Data combined from panels A to E. ●, intradermal, normal, from panel A; ▴, airway, CF, from panel B; ▵, airway, CF, from panel C; ■, liver, colon carcinoma, from panel D; □, heart, coronary artery disease, from panel E. In all panels, the dashed horizontal line represents the lower limit of detection of the assay. For convenience in plotting the data, only the log dose is given; for the actual doses (e.g., 8 × 10⁷ instead of 10⁷ in panel A), see Table 2.