Adeno-associated virus 9-mediated airway expression of antibody protects old and immunodeficient mice against influenza virus

Abstract

Influenza causes serious and sometimes fatal disease in individuals at risk due to advanced age or immunodeficiencies. Despite progress in the development of seasonal influenza vaccines, vaccine efficacy in elderly and immunocompromised individuals remains low. We recently developed a passive immunization strategy using an adeno-associated virus (AAV) vector to deliver a neutralizing anti-influenza antibody at the site of infection, the nasal airways. Here we show that young, old, and immunodeficient (severe combined immunodeficient [SCID]) mice that were treated intranasally with AAV9 vector expressing a modified version of the broadly neutralizing anti-influenza antibody FI6 were protected and exhibited no signs of disease following an intranasal challenge with the mouse-adapted H1N1 influenza strain A/Puerto Rico/8/1934(H1N1) (PR8) (Mt. Sinai strain). Nonvaccinated mice succumbed to the PR8 challenge due to severe weight loss. We propose that airway-directed AAV9 passive immunization against airborne infectious agents may be beneficial in elderly and immunocompromised patients, for whom there still exists an unmet need for effective vaccination against influenza.

FIG 1

AAV9-mediated ffLuc expression in mouse airways. (A) Imaging of ffLuc expression. Young, old, and SCID BALB/c mice were dosed i.n. with 10¹¹ GC of AAV9.ffLuc vector and underwent imaging of ffLuc expression 14 days later. Luminescence was observed in both the nasal and lung airways. (B) Quantification of levels of luminescence in the airways (nose and lung) of young, old, and SCID mice (n = 3/group). **, P < 0.01; ***, P < 0.001, Tukey's test. (C and D) Assessment of FI6 expression in lavage fluid samples. At day 14, FI6 expression was assessed in BALF (C) and NLF (D) samples from young and old mice (n = 3/group) treated with 10¹¹ GC of AAV9.FI6-IA. As controls for nonspecific expression, BALF and NLF samples from mice injected i.n. with 10¹¹ GC of AAV.201Ig-IA (a vector expressing an antibody irrelevant to influenza; n = 3/group) were analyzed.

FIG 2

AAV9.FI6-IA protection of old BALB/c mice from influenza challenge. (A and D) Young vaccinated (○) and nonvaccinated (●) mice (n = 8/group) (A) and old vaccinated (○) and nonvaccinated (●) mice (n = 8/group) (D) were challenged i.n. with PR8 (30 PFU) and weighed daily. Percent changes in weight were calculated based on the weight on the day of the challenge. (B and E) Young (B) and old (E) mice were euthanized when they appeared in distress or their body weight declined >30%, as depicted in the Kaplan-Meier plots. Solid lines, vaccinated mice; dashed lines, nonvaccinated mice. (C and F) Young (C) and old (F) mice (n = 3) from the vaccinated and nonvaccinated groups were necropsied at day 6 (d6) to quantify viral loads in the lung. (G and I) Young (G) and old (I) mice (n = 5/group) were vaccinated with either AAV9.FI6 (○) or an irrelevant vaccine (AAV9.201Ig-IA) (●), challenged i.n. with PR8 (30 PFU), and weighed daily. Percent changes in weight were calculated based on the weight on the day of the challenge. (H and J) Young (H) and old (J) mice were euthanized when they appeared in distress or their body weight declined >30%. Solid lines, mice vaccinated with AAV9.FI6; dashed lines, mice vaccinated with the irrelevant vector. ****, P ≤ 0.0001, Mann-Whitney test; ***, P ≤ 0.001, Mantel-Cox test.

FIG 3

Sustained protection against challenge with PR8. BALB/c mice were dosed i.n. with 10¹¹ GC of AAV9.FI6-IA vector in 50 μl (n = 5 mice/group). Mice were challenged with PR8 (30 PFU) 7 days (A), 14 days (B), 28 days (C), 3 months (D), or 6 months (E) after the single instillation of vector. The weights of the animals were assessed daily, and mice were euthanized when they appeared in distress or they lost >30% of their initial body weight. Percent changes in weight were calculated based on the weight on the day of the challenge. ○, AAV9.FI6-IA vector-treated mice; ●, naïve mice. ****, P ≤ 0.0001, Mann-Whitney test.

FIG 4

AAV9.FI6-IA protection of SCID mice from influenza challenge. (A) Vaccinated (○) and nonvaccinated (●) BALB/c mice were challenged i.n. with 30 PFU of PR8 and weighed daily. Percent changes in weight were calculated based on the weight on the day of the challenge. (B) BALB/c mice were euthanized when they appeared in distress or their body weight declined >30%, as depicted in the Kaplan-Meier plot. Solid lines, vaccinated mice; dashed lines, nonvaccinated mice. (C) Vaccinated (○) and nonvaccinated (●) SCID mice were challenged i.n. with 30 PFU of PR8 and weighed daily. Percent changes in weight were calculated based on the weight on the day of the challenge. (D) SCID mice were euthanized when they appeared in distress or their body weight declined >30%, as depicted in the Kaplan-Meier plot. Solid lines, vaccinated mice; dashed lines, nonvaccinated mice. ****, P ≤ 0.0001, Mann-Whitney test.