P49.07
Background: Griffithsin (GRFT), an anti-HIV protein isolated from the red alga Griffithsia sp., is being developed as a microbicide. GRFT binds N-linked glycans on gp120 to prevent HIV entry. Earlier studies evaluating GRFT binding in cervicovaginal lavage (CVL) samples using lectin microarrays led us to investigate how GRFT interacts with glycoproteins in the vaginal microenvironment
Methods: CVL was collected in PBS from 135 healthy asymptomatic reproductive aged women and stored at −70°C until used. Vaginal smears were evaluated using the Nugent criteria for bacterial vaginosis (BV). The samples were evaluated for GRFT binding in an ELISA using anti- GRFT primary antibody and a horseradish peroxidase labeled secondary antibody. The ELISA data were analyzed using one-way analysis of variance. GRT binding proteins in the CVLs were identified by SDS-PAGE and Western blotting.
Results: GRFT bound at higher levels in the CVLs obtained from women having a Lactobacillus-predominant flora (Nugent score 0-3) or intermediate flora (Nugent 4-6) compared to BV flora (Nugent 7-10) in pg/ng protein: 10.74+5.82, 10.56+8.09, 4.03+3.22 pg/ng protein, respectively, P value = <0.001(Normal vs BV P<0.001; Intermediate vs BV P=0.001). Western blots of normal CVL revealed five GRFT binding proteins at apparent molecular weights (in kDa) of 185, 142, 120, 81 and 71, while 2/4 BV CVL demonstrated no GRFT binding proteins and two had binding to proteins with apparent MWs of 54 to>250. The CVL from women with BV had reduced levels of these GRFT binding proteins.
Conclusions: GRFT binds to at least 5 proteins found in CVL, with different protein patterns in women with normal flora vs those with BV. The lower binding of GRFT in the CVL of women with BV suggests a protective function for GRFT binding proteins. It also suggests that a GRFT based microbicide may be bound to endogenous vaginal proteins reducing availability to compete with the HIV gp120 protein in women with normal flora.
