Mucosal delivery of microbicides by recombinant commensal bacteria: expression of the HIV-inactivating protein cyanovirin-N in gram-positive bacteria

Abstract

To investigate feasibility of using recombinant commensal bacteria for vaginal delivery of microbicides, we constructed strains of Streptococcus gordonii expressing the potent HIV-inactivating protein, cyanovirin-N (CV-N). The Siena group have been developing methods for engineering nonpathogenic commensals for stable expression of heterologous proteins as an approach to mucosal vaccine delivery (see selected references below), and now we are exploring this approach for in situ delivery of microbicidal proteins or peptides. Recombinant S. gordonii stably colonize mouse and rat vaginal mucosa, and persist in the vagina of cynomolgous monkeys, and serve as a useful model expression system. CV-N was produced in S. gordonii as a fusion protein with M6, a streptococcal surface protein. Four different CV-N/M6 fusions were expressed. Two included the C-terminal portion of M6 which allowed surface anchoring, while the others were secreted into the culture medium. The N-terminal portion of M6 in the fusions was either 5- or 121-amino acids long. The in vitro HIV-inactivating capabilities of whole recombinant bacteria expressing CV-N on the surface, and of the S. gordonii-produced soluble CV-N, are currently being assayed. Future plans include expression of CV-N in human vaginal strains of Lactobacillus, and microbicidal evaluations of CV-N-producing commensals in animal models.