Bifunctional CD4-DC-SIGN fusion proteins demonstrate enhanced avidity to gp120 and inhibit HIV-1 infection and dissemination.

Early stages of mucosal infection are potential targets for HIV-1 prevention. CD4 is the primary receptor in HIV-1 infection whereas DC-SIGN likely plays an important role in HIV-1 dissemination, particularly during sexual transmission. To test the hypothesis that an inhibitor simultaneously targeting both CD4 and DC-SIGN binding sites on gp120 may provide a potent anti-HIV strategy, we designed constructs by fusing the extracellular CD4 and DC-SIGN domains together with varied arrangements of the lengths of CD4, DC-SIGN and the linker. We expressed, purified and characterized a series of soluble CD4-linker–DC-SIGN (CLD) fusion proteins. Several CLDs, composed of a longer linker and an extra neck domain of DC-SIGN, had enhanced affinity for gp120 as evidenced by molecular-interaction analysis. Furthermore, such CLDs exhibited significantly enhanced neutralization activity against both laboratory-adapted and primary HIV-1 isolates. Moreover, CLDs efficiently inhibited HIV-1 infection in trans via a DC-SIGN-expressing cell line and primary human dendritic cells. This was further strengthened by the results from the human cervical explant model, showing that CLDs potently prevented both localized and disseminated infections. This is the first time that soluble DC-SIGN-based bifunctional proteins have demonstrated anti-HIV potency. Our study provides proof of the concept that targeting both CD4 and DC-SIGN binding sites on gp120 represents a novel antiviral strategy. Given that DC-SIGN binding to gp120 increases exposure of the CD4 binding site and that the soluble forms of CD4 and DC-SIGN occur in vivo, further improvement of CLDs may render them potentially useful in prophylaxis or therapeutics

The HIV-1 transmission bottleneck

It is well established that most new systemic infections of HIV-1 can be traced back to one or a limited number of founder viruses. Usually, these founders are more closely related to minor HIV-1 populations in the blood of the presumed donor than to more abundant lineages. This has led to the widely accepted idea that transmission selects for viral characteristics that facilitate crossing the mucosal barrier of the recipient’s genital tract, although the specific selective forces or advantages are not completely defined. However, there are other steps along the way to becoming a founder virus at which selection may occur. These steps include the transition from the donor’s general circulation to the genital tract compartment, survival within the transmission fluid, and establishment of a nascent stable local infection in the recipient’s genital tract. Finally, there is the possibility that important narrowing events may also occur during establishment of systemic infection. This is suggested by the surprising observation that the number of founder viruses detected after transmission in intravenous drug users is also limited. Although some of these steps may be heavily selective, others may result mostly in a stochastic narrowing of the available founder pool. Collectively, they shape the initial infection in each recipient

Transferrin conjugation confers mucosal molecular targeting to a model HIV-1 trimeric gp140 vaccine antigen☆

The generation of effective immune responses by mucosal vaccination without the use of inflammatory adjuvants, that compromise the epithelial barrier and recruit new cellular targets, is a key goal of vaccines designed to protect against sexually acquired pathogens. In the present study we use a model HIV antigen (CN54gp140) conjugated to transferrin (Tf) and evaluate the ability of the natural transferrin receptor CD71 to modulate immunity. We show that the conjugated transferrin retained high affinity for its receptor and that the conjugate was specifically transported across an epithelial barrier, co-localizing with MHC Class II+ cells in the sub-mucosal stroma. Vaccination studies in mice revealed that the Tf-gp140 conjugate elicited high titres of CN54gp140-specific serum antibodies, equivalent to a systemic vaccination, when conjugate was applied topically to the nasal mucosae whereas gp140 alone was poorly immunogenic. Moreover, the Tf-gp140 conjugate elicited both IgG and IgA responses and significantly higher gp140-specific IgA titre in the female genital tract than unconjugated antigen. These responses were achieved after mucosal application of the conjugated protein alone, in the absence of any pro-inflammatory adjuvant and suggest a potentially useful and novel molecular targeting approach, delivering a vaccine cargo to directly elicit or enhance pathogen-specific mucosal immunity