Antibody Responses Are Generated to Immunodominant ELK/KLE-Type Motifs on the Nonstructural-1 Glycoprotein during Live Dengue Virus Infections in Mice and Humans: Implications for Diagnosis, Pathogenesis, and Vaccine Design

Abstract

Antibodies generated to the purified dengue type 2 virus (D-2V) nonstructural-1 (NS1) protein in mice and rabbits were compared with those generated to this protein in congeneic (H-2 class II) mouse strains and humans after D-2V infections. Unlike the profiles observed with the rabbits, similar antibody reaction profiles were generated by mice and humans with severe D-2V disease (dengue hemorrhagic fever [DHF]/dengue shock syndrome [DSS]). Many of these epitopes contained the core acidic-hydrophobic-basic (tri-amino-acid; ELKtype) motifs present in the positive or negative orientations. Antibody responses generated to these ELK/KLEtype motifs and the epitope LX1 on this protein were influenced by class II molecules in mice during D-2V infections; but these antibodies cross-reacted with human fibrinogen and platelets, as implicated in DHF/DSS pathogenesis. The core LX1 epitope ( 113 YSWKTWG 119 ), identified by the dengue virus complex-specific monoclonal antibody (MAb) 3D1.4, was prepared so that it contained natural I-A d -binding and ELK-type motifs. This AFLX1 peptide, which appropriately displayed the ELK-type and LX1 epitopes in solid-phase immunoassays, generated a similar, but lower, immunodominant anti-ELK-motif antibody reaction in I-A dpositive mice, as generated in mice and humans during D-2V infections. These antibody responses were much stronger in the high-responding mouse strains and each of the DHF/DSS patients tested and may therefore account for the association of DHF/DSS resistance or susceptibility with particular class II molecules and autoantibodies, antibody-stimulating cytokines (e.g., interleukin-6), and complement product C3a being implicated in DHF/DSS pathogenesis. These results are likely to be important for the design of a safe vaccine against this viral disease and showed the AFLX1 peptide and MAb 3D1.4 to be valuable diagnostic reagents.

Determination of Viremia and Concentration of Circulating Nonstructural Protein 1 in Patients Infected with Dengue Virus in Mexico

“…Statistical significance was P 0.05. infection may be related to inefficient detection of NS1 during secondary infections. [35][36][37][38] In summary, these results suggest that during primary infections, levels of circulating NS1 are higher in patients with DHF than in patients with DF. Nonetheless, our results and other results 18,20 also suggest that concentrations of circulating NS1 are affected by the infecting serotype and the immunologic status (i.e., primary or secondary infection) of the patient.…”

Monoclonal Antibodies That Bind to Common Epitopes on the Dengue Virus Type 2 Nonstructural-1 and Envelope Glycoproteins Display Weak Neutralizing Activity and Differentiated Responses to Virulent Strains: Implications for Pathogenesis and Vaccines

“…Indirect ELISAs and immunoblot assays using purified DENV-2 particles and the DENV-2 NS1 glycoprotein were described previously (7,10). For ELISA, optimal concentrations of purified DENV-2 particles (1/250 dilution; 0.58 g/ml) and DENV-2 eNS1 glycoprotein (2 to 3 g/ml) were diluted in 0.16%/0.29% (wt/vol) sodium carbonate/bicarbonate (pH 9.8), loaded at 50 l/well into ELISA plates (Immulon 2; Dynatech), and adsorbed overnight at 4°C.…”

“…Three other MAbs generated to the DENV-2 NS1 protein also recognized short sequential amino acid sequences. MAb 1C6.3 reacted more specifically with multiple KELK-type motifs present in either orientation (KELK/KLEK-type motifs), MAb 3D1.4 recognized the LX1 (113-YSWKTWG-119) epitope, and MAb 1G5.3 recognized the 24C (301-TTASGKLIT-309) epitope (7,9,12).…”

“…During DENV infections, human PAb responses were generated to multiple acidic (E or D)-aliphatic/aromatic (G, A, I, L or V/F, W, or Y)-basic (K or R) (ELK-type triamino acid) motifs present in either orientation (ELK/KLEtype motifs) on the DENV NS1 glycoproteins, and these responses were higher in DSS patients than in patients with mild disease (dengue fever [DF]) (7). Monoclonal antibody (MAb) 1G5.4-A1-C3 displayed the same reaction pattern as that for human DSS patient PAbs against multiple ELK/KLE-type epitopes on the DENV-2 NS1 glycoprotein, and therefore the cross-reaction of this MAb with other DENV proteins and human proteins is likely to be highly relevant in studies of DHF/DSS pathogenesis (7,9). Three other MAbs generated to the DENV-2 NS1 protein also recognized short sequential amino acid sequences.…”

Dengue Virus NS1 Protein Modulates Cellular Energy Metabolism by Increasing Glyceraldehyde-3-Phosphate Dehydrogenase Activity

“…Furthermore, the anti-NS1 antibodies elicited at high titers during infection may form immune complexes with NS1, which trigger the inflammatory response and bind some coagulation elements to deregulate vascular permeability (13,14). Conversely, active or passive immunization with NS1 promotes the survival of DENVchallenged mice, making it an attractive target for vaccine development (15-17).…”