A Single Dose, Thermostable, Trivalent Human Papillomavirus Vaccine Formulated Using Atomic Layer Deposition

Abstract

Formulations of human papillomavirus (HPV) 16, 18, and 31 L1 capsomere protein antigens were spray dried to obtain glassy microspheres that were then coated by atomic layer deposition (ALD) with nanometer-thin protective layers of alumina. Spray-drying was used to formulate human papillomavirus (HPV) 16, 18, and 31 L1 capsomere protein antigens within glassy microspheres to which nanoscopic protective layers of alumina were applied using ALD. Suspensions of alumina-coated, capsomere-containing microparticles were administered in a single dose to mice. ALD-deposited alumina coatings provided thermostability and a delayed in vivo release of capsomere antigens, incorporating both a prime and a boost dose in one injection. Total serotype-specific antibody titers as well as neutralizing titers determined from pseudovirus infectivity assays were unaffected by incubation of the ALD-coated vaccines for at 4, 50, or 70 °C for three months prior to administration. In addition, even after incubation for three months at 70 °C, single doses of ALD-coated vaccines produced both higher total antibody responses and higher neutralizing responses than control immunizations that used two doses of conventional liquid formulations stored at 4 °C.

Keywords: Atomic layer deposition; Controlled release; Human papilloma virus vaccine; Thermostability.

Figure 1.. Titers of antibodies against serotypes HPV 16, 18, and 31 in vaccinated BALB/C mice.

Anti-HPV 16 titers are shown in the first row, anti-HPV 18 titers in the second row, and anti-HPV 31 titers in the bottom row. Each point is the geometric mean of titers for all animals in the group. Titers for groups of mice that received 7.5 μg doses of HPV capsomeres in liquid or spray-dried and reconstituted (SD) formulations on days 0 and 21 are shown in black closed squares. Titers for groups of mice that received single, 7.5 μg doses of HPV capsomeres formulated in spray-dried microspheres coated with nanoscopic alumina layers deposited in 250 ALD cycles that had been incubated for three months at 4, 50 or 70 °C are shown as open orange circles, open blue triangles, and inverted open green triangles, respectively.

Figure 2.. Maximum anti-HPV 16, 18, and 31 ELISA titers.

Maximum anti-HPV16 (A), anti-HPV18 (B), or anti-HPV31 (C) ELISA titers measured in sera from each group of mice. All mice were administered a 7.5 μg dose of HPV capsomeres on day 0; groups of mice that received liquid and SD formulations were administered an additional 7.5 μg booster dose on day 21. ALD-coated microspheres were incubated for 3 months at 4, 50 or 70 °C prior to administration. Maximum titers were reached on the day noted on each of the bars of the graph.

Figure 3.. Day 72 neutralization titers for HPV16 (A), 18 (B), and 31(C).

Groups received liquid formulations (black squares), spray-dried formulations (red circles), ALD-coated microspheres incubated at 4 °C (blue triangles), 50 °C (green inverted triangles) or 70 °C (gold diamonds). Mice were administered 7.5 μg doses on day 0. Groups receiving liquid or spray-dried formulations were administered additional booster doses on day 21 whereas only a single dose was administered to groups receiving ALD-coated microsphere formulations.