Miltefosine as a novel influenza vaccine adjuvant

Abstract

Vaccine plays a major role in the prevention of influenza virus infection. However, the effectiveness of currently available influenza vaccines is unsatisfactory. According to the United States Centers for Disease Control and Prevention, the effectiveness of the influenza vaccine fell below 50% for H3N2 in most seasons in the past 15 years. Furthermore, the influenza vaccine generally induces a protective immune response at least two weeks after vaccination. Therefore there is a window period without protection shortly after vaccination. In this study, miltefosine, an FDA-approved anti-parasitic drug, was evaluated as a novel influenza vaccine adjuvant. I found that the miltefosine-adjuvanted vaccine improved the vaccine-induced protection in mice upon lethal virus challenge at 3 or 7 days after vaccination. The significantly higher survival rate, lower body weight loss, and milder lung damage were observed in infected mice receiving the miltefosine-adjuvanted vaccine (VCM) compared to those receiving vaccine alone (Vac). To decipher the beneficial effect of miltefosine, the immune responses were studied. The antibody titers were significantly higher in mice receiving VCM than Vac before and after the virus challenge. Besides, post-infection sera from the VCM group were found to be cross-reactive against an antigenically-drifted virus strain, suggesting the induction of cross-protection by VCM. Antigen-specific splenocyte cytokine responses were also significantly increased in mice receiving VCM. The frequencies of TFH cells elevated significantly in mice receiving VCM compared to those receiving Vac, before and after the virus challenge. Histological examination showed that the formation of germinal centers was also promoted by miltefosine-adjuvanted vaccination. Superior immunological memory was induced by VCM compared to that of Vac. Quicker and greater secondary antibody responses were observed in mice given VCM upon recall of vaccine antigens. To further elucidate the mechanisms of adjuvant effects of miltefosine, whole blood transcriptomic analysis was conducted. After miltefosine-adjuvanted vaccination, increased oxidative stress responses and erythrocytes and hemoglobin activity were observed. These were associated with improved vaccine effectiveness directly or indirectly through inflammatory responses. After virus challenge, reduced inflammatory responses, increased platelet activation, and upregulated actin cytoskeleton activity were found in the VCM group. These were linked to reduced lung tissue damage and promoted activation of monocytes and adaptive immunity. The vaccine formula based on a cocktail of three adjuvants with influenza vaccine, including miltefosine, imiquimod, and paromomycin, were found to increase the vaccine effectiveness, but the effect was not better than miltefosine adjuvanted influenza vaccine. Additionally, miltefosine was demonstrated not to be an antiviral drug against the influenza virus. Collectively, the miltefosine-adjuvanted influenza vaccine showed significantly improved protection in mice with only one dose and in a much shorter time. The improved protective response is related to augmented antibody response and TFH cell-associated germinal center response. Comparing to other vaccine adjuvants, miltefosine has the advantage of being a clinically-approved drug with known safety data in human. Repurposing existing drugs is a feasible option for identifying novel vaccine adjuvants.