DNA Vaccine Combinations Expressing Either Tissue Plasminogen Activator Signal Sequence Fusion Proteins or Ubiquitin-Conjugated Antigens Induce Sustained Protective Immunity in a Mouse Model of Pulmonary Tuberculosis

Abstract

DNA vaccination has emerged as a powerful approach in the search for a more efficacious vaccine against tuberculosis. In this study, we evaluated the effectiveness of immunizing with combinations of 10 different tuberculosis DNA vaccines that expressed mycobacterial proteins fused at the N terminus to eukaryotic intracellular targeting sequences. In one vaccine combination, the genes were fused to the tissue plasminogen activator signal sequence (TPA), while in a second combination the same 10 genes were expressed as ubiquitin (Ub)-conjugated proteins. In ex vivo studies in which the secretion of gamma interferon was measured, cellular immune responses were detected in mice vaccinated with either the TPA DNA vaccine combination or the Ub DNA vaccine combination at 7 and 14 days following a low-dose Mycobacterium tuberculosis challenge. Moreover, mice vaccinated with the TPA combination, the Ub combination, and Mycobacterium bovis BCG were able to limit the growth of tubercle bacilli in the lung and spleen after a virulent tuberculous aerosol challenge. Histopathological analyses also showed that mice immunized with the DNA vaccine combinations had substantially improved postinfection lung pathology relative to the naïve controls. Finally, in three different long-term experiments, the survival periods following aerogenic challenge were extended as much as sevenfold for vaccinated mice compared to naïve controls. Interestingly, in all three experiments, no significant differences were detected in the mean times to death for mice immunized with the TPA combination or the Ub combination relative to the BCG controls. In conclusion, these studies demonstrate the effectiveness of immunization with DNA vaccine combinations against tuberculosis and suggest that further testing of these plasmid cocktails is warranted.Tuberculosis (TB) is a major global threat to human health, with more than 2 million people dying each year from Mycobacterium tuberculosis infections (39). Although the current TB vaccine (Mycobacterium bovis BCG) has been widely used throughout the world for many decades, its efficacy has been shown to be highly variable in several well-controlled clinical trials (5). Moreover, since BCG is a live attenuated vaccine, its use is contraindicated in human immunodeficiency-infected and other immunocompromised patients, who ironically have the highest risk of developing TB. Clearly, the development of a new, more effective vaccine would greatly facilitate worldwide control of this ancient plague. As a consequence, the search for a new vaccine against TB has greatly intensified over the last decade, with several experimental vaccines already shown to have promise in preclinical testing (30).Since DNA vaccines can induce substantial cellular immunity and evoke both CD4 and CD8 T-cell responses, genetic immunization has become a viable strategy for developing new vaccines against intracellular parasites (18). Several recent studies carried out with animal models of TB have suggested that DNA vaccines against this d...

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