Coadministration of cidofovir and smallpox vaccine reduced vaccination side effects but interfered with vaccine-elicited immune responses and immunity to monkeypox

Abstract

While the smallpox vaccine, Dryvax or Dryvax-derived ACAM2000, holds potential for public immunization against the spread of smallpox by bioterror, there is serious concern about Dryvax-mediated side effects. Here, we report that a single-dose vaccination regimen comprised of Dryvax and an antiviral agent, cidofovir, could reduce vaccinia viral loads after vaccination and significantly control Dryvax vaccination side effects. However, coadministration of cidofovir and Dryvax also reduced vaccine-elicited immune responses of antibody and T effector cells despite the fact that the reduced priming could be boosted as a recall response after monkeypox virus challenge. Evaluations of four different aspects of vaccine efficacy showed that coadministration of cidofovir and Dryvax compromised the Dryvax-induced immunity against monkeypox, although the covaccinated monkeys exhibited measurable protection against monkeypox compared to that of naïve controls. Thus, the single-dose coadministration of cidofovir and Dryvax effectively controlled vaccination side effects but significantly compromised vaccine-elicited immune responses and vaccine-induced immunity to monkeypox.

FIG. 1.

Cidofovir+Dryvax vaccination regimen significantly controlled Dryvax-induced skin lesions and reduced vaccinia (Dryvax) viral loads in PBMC. (a) Representative photos show the skin rashes formed at the vaccination sites of individual monkeys in the Dryvax-alone and cidofovir+Dryvax groups on day 10 after vaccination. All animals vaccinated with Dryvax alone developed large and severe skin blisters, whereas the monkeys vaccinated with the cidofovir+Dryvax regimen developed no or very small skin rashes. (b) Comparison of sizes of skin rashes at the vaccination sites between Dryvax alone and cidofovir+Dryvax groups at different time points postvaccination. The horizontal bars indicate mean skin rash areas (mm²) for the groups; the cidofovir+Dryvax group had a much smaller skin rash than that of the Dryvax-alone group on days 3, 6, 10, 13, and 20 postvaccination (**, P < 0.01). (c) Comparison of numbers of A33R copies in the PMBC between the Dryvax alone and cidofovir+Dryvax groups using real-time quantitative PCR. Viral A33R mRNA was detected at days 4 and 7 in PBMC from all monkeys vaccinated with Dryvax alone but was low or undetectable in the monkeys covaccinated with cidofovir+Dryvax. The mean number of copies of the A33R gene in PBMC of the Dryvax-alone group (horizontal bars) is much higher than that of the cidofovir+Dryvax group at days 4 and 7 postvaccination (*, P < 0.05). The dashed line indicates the detection limit for the real-time quantitative PCR.

FIG. 2.

Cidofovir+Dryvax coadministration resulted in marked decreases in Dryvax-elicited antibody and T-cell immune responses despite the fact that the reduced priming could be boosted as a recall response after monkeypox challenge. (a to c) Mean GMT levels of anti-B5R, anti-L1R antibodies (Ab), and neutralizing antibodies (GMT ND₅₀), respectively, before and after vaccination as well as after monkeypox virus challenge, as indicated. The virus-specific antibody titers in the cidofovir+Dryvax group were lower than those in the Dryvax-group but higher than those in the mock control group (×, P < 0.05 for days 28, 35, and 55 when results for the cidofovir+Dryvax group are compared to those for the mock control group). Note that up to 2 log increases in mean GMT within 7 days after monkeypox virus challenge were seen for the cidofovir+Dryvax group, with a P value of <0.05 for anti-L1R and ND₅₀ antibodies on day 7 after challenge compared to the mock control group. (d to f) Mean absolute numbers of viral H3L-specific IFN-γ⁺ cells detected by ELISPOT (d) and mean percentages of H3L-specific IFN-γ⁺ CD4⁺ (e) and CD8⁺ T cells (f) by ICS for each group before and after vaccination as well as after monkeypox virus challenge. The numbers of H3L-specific IFN-γ⁺ CD8⁺ T cells in the cidofovir+Dryvax group were greater than those in the mock control group after vaccination (×, P < 0.05). At day 35, two naïve monkeys showed a high background of IFN-γ⁺ CD8⁺ T cells. H3L-specific IFN-γ⁺ lymphocytes and CD4⁺ and CD8⁺ T effector cells in the cidofovir+Dryvax group demonstrated some extent of recall expansion after monkeypox virus challenge. * and ** denote that P values for mean GMT levels of anti-B5R, anti-L1R antibodies, and neutralizing antibodies or mean percentage numbers of H3L-specific IFN-γ⁺ CD4⁺ and CD8⁺ T effector cells in the Dryvax-alone group were <0.05 and <0.01, respectively, compared with the mock control group.

FIG. 3.

The cidofovir+Dryvax vaccination regimen impaired Dryvax-induced immunity to monkeypox and conferred poor, although measurable, protection against monkeypox. (a) All the monkeys vaccinated with Dryvax alone showed no or a few small skin lesions after virus challenge. It seemed that the cidofovir+Dryvax-vaccinated group developed fewer skin lesions than did mock control animals on days 7, 11, and 21 after monkeypox virus challenge (*, P < 0.05), but this group did not have significantly fewer lesions than the mock control group at other time points. (b) All the monkeys vaccinated with Dryvax alone survived the lethal monkeypox virus challenge. There was no significant difference in survival rate between the cidofovir+Dryvax group and the mock control group (P > 0.05), although the cidofovir+Dryvax-vaccinated group had a longer mean survival time than the mock control group during the 28-day follow-up after the monkeypox virus challenge (P < 0.032).

FIG. 4.

The Dryvax-alone group did not have any notable pathological findings, and the cidofovir+Dryvax group appeared to exhibit less-severe pathology than the mock control group (Table 3). (a) Representative gross pathology photographs demonstrating that the Dryvax-alone group was “lesion-free,” whereas the cidofovir+Dryvax group had noticeable monkeypox lesions that were less severe than those seen with the mock group. An early necropsied monkey, 7329, and a survivor, 7336, in the mock control group exhibited apparent lung congestion and edema with many large grayish-white lesions on the surface. No apparent lung lesions were seen for monkey 7331 and others vaccinated with Dryvax alone. Monkey 7325 and other survivors in the cidofovir+Dryvax group showed no lung congestion/edema and had fewer and smaller grayish-white lesions in the lungs than the two surviving monkeys in the mock group. The monkeys, including 7322, in the cidofovir+Dryvax group exhibited less-inflamed and less-congested spleens (splenomegaly) and other organs than those of animals in the mock group (P < 0.05, see Table 3 for organ weights). (b) Representative histological photographs demonstrating that the cidofovir+Dryvax group had less-severe lung and skin monkeypox lesions than the mock control. An early necropsied monkey, 7329, developed acutely exudative inflammation with central necrosis and destruction of the involved bronchiolar wall; monkey 7336 (mock group) surviving the 28-day follow-up still showed evident histopathologic changes in which some alveoli were filled with edema fluid, macrophages, degenerative neutrophils, and necrotic cellular debris, whereas hyperplastic fibroblasts and fibrous proliferation were seen in thickened alveolar walls. The monkey in the Dryvax-alone group exhibited “normal” histology in the lungs and skin. Monkey 7320 in the cidofovir+Dryvax group had less severe histopathology, as mild fibrous proliferation and thickening were seen only in a few alveoli in the lungs, and less-severe residual lesions in the skin were seen on day 28 after challenge. The representative data shown from gross pathology and histology seem to suggest that the pathology in animals from the cidofovir+Dryvax group looks similar to that found in survivors of the mock group, while net differences in pathology are visible in comparison to the Dryvax-only animals. HE, hematoxylin and eosin.