ABSTRACT
Soluble forms of the human immunodeficiency virus type 1 (HIV-1) primary receptor CD4 (soluble CD4 [sCD4]) have been extensively characterized for a quarter of a century as promising HIV-1 inhibitors, but they have not been clinically successful. By combining a protein cavity-filling strategy and the power of library technology, we identified an engineered cavity-altered single-domain sCD4 (mD1.22) with a unique combination of excellent properties, including broad and potent neutralizing activity, high specificity, stability, solubility, and affinity for the HIV-1 envelope glycoprotein gp120, and small molecular size. To further improve its neutralizing potency and breadth, we generated bispecific multivalent fusion proteins of mD1.22 with another potent HIV-1 inhibitor, an antibody domain (m36.4) that targets the coreceptor-binding site on gp120. The fusion proteins neutralized all HIV-1 isolates tested, with potencies about 10-, 50-, and 200-fold higher than those of the broadly neutralizing antibody VRC01, the U.S. FDA-approved peptide inhibitor T20, and the clinically tested sCD4-Fc fusion protein CD4-Ig, respectively. In addition, they exhibited higher stability and specificity and a lower aggregation propensity than CD4-Ig. Therefore, mD1.22 and related fusion proteins could be useful for HIV-1 prevention and therapy, including eradication of the virus.
Subject(s)
Anti-HIV Agents/immunology , CD4 Antigens/immunology , HIV Antibodies/immunology , HIV Infections/virology , HIV-1/immunology , Single-Domain Antibodies/immunology , Anti-HIV Agents/chemistry , Anti-HIV Agents/therapeutic use , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/genetics , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/therapeutic use , CD4 Antigens/genetics , Cross Reactions , HIV Antibodies/chemistry , HIV Antibodies/genetics , HIV Antibodies/therapeutic use , HIV Envelope Protein gp120/immunology , HIV Infections/drug therapy , HIV Infections/immunology , HIV-1/drug effects , HIV-1/genetics , Humans , Neutralization Tests , Single-Domain Antibodies/chemistry , Single-Domain Antibodies/genetics , Single-Domain Antibodies/therapeutic useABSTRACT
Sera from patients with Chagas' disease were used to screen a Trypanosoma cruzi amastigote cDNA library. Characterization of 50 positive clones showed that 21 (42%) encode previously identified T. cruzi ribosomal and flagellar proteins, heat-shock proteins or proteins with repetitive motifs. Twenty-nine clones (58%) correspond to nine genes not previously described in T. cruzi. Three cDNAs, encoding novel repetitive antigens with homology to ribosomal proteins and to other RNA binding proteins, were further characterized. Patient humoral responses against the recombinant proteins encoded by these cDNAs were evaluated in anticipation that they may constitute potential new targets for serodiagnostic assays.
