Single-Domain Antibody-SH3 Fusions for Efficient Neutralization of HIV-1 Nef Functions

HIV-1 reprograms the migration of macrophages

“…Actually, this is not surprising because vesicular trafficking is essential for podosome function. 61,79,80 Thus, multiple partners could mediate Nef effects on podosomes, but strikingly, results obtained with Neffin antibodies 64 and with siRNA-mediated gene silencing demonstrated that Hck is a main effector. Consequently, we propose that pharmacological disruption of the Nef/Hck interaction 70 may specifically reduce mesenchymal migration of infected macrophages and reduce HIV-1 dissemination and pathogenesis.…”

“…All other Nef SF2 variants, including 2 Nef variants (AxxA and VGF→AAA) that impact the interaction of Nef with SH3-binding domains, were not found around podosomes and were unable to modify their size (supplemental Figure 5A-B). When wt-Nef SF2 was coexpressed with antiNef single-domain antibodies (sdAbs) and Neffins C1 and B6, 63,64 we observed that Nef SF2 was delocalized from podosomes (data not shown), and F-actin content in podosomes was significantly decreased (supplemental Figure 5C). Neffins C1 and B6 are anti-Nef sdAbs linked to the SH3 domain of the Src family member Hck, which inhibit most Nef biological activities.…”

“…Neffins C1 and B6 are anti-Nef sdAbs linked to the SH3 domain of the Src family member Hck, which inhibit most Nef biological activities. 63,64 The strong inhibitory effect of Neffins on podosome size, together with the effects of Nef mutants AxxA and VGF→AAA, suggested a pivotal role of the Nef-SH3 interaction in podosome modulation. In addition, Nef SF2 AxxA failed to increase the life span of podosomes compared with wt-Nef SF2 (supplemental Figure 5D).…”

Small molecule inhibitors of the HIV-1 virulence factor, Nef

“…These studies provide a strong rationale for the discovery and development of small molecule antagonists of Nef function as a new approach to antiretroviral therapy. Furthermore, recent studies show that engineered Nef-binding proteins block its functions in cell-based studies, including CD4 and MHC-I downregulation, viral infectivity, and kinase activation [16]. These experiments provide an important proof-of-concept that Nef antagonists may be valuable weapons in the fight against AIDS.…”

HIV-1 Nef: a master manipulator of the membrane trafficking machinery mediating immune evasion

“…A single domain antibody was also designed to target Nef [66], and based on this antibody a small peptide, termed Neffin, was produced containing a portion of the anti-Nef antibody linked to a recombinant form of the Hck SH3 domain [67]. This peptide binds Nef with high affinity and inhibits a number of Nef functions including MHC-I downregulation and Nef-mediated increases in infectivity [67,68]. The development of Nef inhibitors is currently an area of interest, as Nef inhibition represents a novel HIV-1 targeting strategy.…”