Hepatitis C computer virus broadly neutralizing monoclonal antibodies isolated 25 years after spontaneous clearance

Hepatitis C computer virus broadly neutralizing monoclonal antibodies isolated 25 years after spontaneous clearance. of two impartial experiments, each performed in duplicate. HCVcc values are from a single experiment performed in duplicate. MAb names are color coded according to hierarchical clustering in Fig. 2. In many cases, the neutralizing breadth of C18 MAbs was consistent with the previously described neutralizing breadth of Ldb2 closely related reference MAbs. Two of the three most broadly neutralizing C18 MAbs (HEPC153 and HEPC151-1) bound at AR3, the target of many previously described bNAbs (19). In addition to these AR3-site MAbs, HEPC111 was also broadly neutralizing (17 of 24 strains [4 of 6 genotypes] neutralized), which was similar to the previously described neutralizing breadth of closely related reference MAb AR4A (12 of 19 genotype 1 HCVpps were neutralized by AR4A in reference 31). HEPC167, which clustered with the weakly neutralizing reference MAb AR1A in the binding analysis, also exhibited poor neutralizing breadth against the HCVpp panel (2 of 24 strains [1 of 6 genotypes] neutralized). The neutralizing breadth of AS108 MAbs varied widely. HEPC108 was broadly neutralizing (19 of 24 strains [5 of 6 genotypes] neutralized) despite sharing probable binding residues with weakly neutralizing reference MAb AR1A and weakly neutralizing C18 MAb HEPC167. Furthermore, HEPC132, which also bound at AS108 and shared 10 of 15 HEPC108 probable binding residues, neutralized 0 of 24 strains, further demonstrating that this neutralizing breadth of MAbs is not determined solely by KY02111 the antigenic site targeted. HEPC112, which binds a novel site in E1 (AS112), neutralized 7 of 24 strains (1 of 6 genotypes), which did not meet our threshold of broad neutralization. Taken together, these results demonstrate that C18 MAbs targeting known antigenic sites (AR3 and AR4-5) as well as non-AR1C5 antigenic sites (AS108 and AS146) were broadly neutralizing. bNAbs targeting multiple antigenic sites were encoded by IgHV1-69. We sequenced the heavy and light chain variable gene sequences of each of the MAbs (Table 3). As we and others have previously observed (19, 31, 32, 35), multiple AR3-site MAbs (HEPC122, HEPC151-1, and HEPC153) were encoded by the same antibody heavy chain variable gene segment, VH1-69. Of note, one AR4-5-site MAb (HEPC111) and one AS108-site MAb (HEPC108) also used VH1-69. Collectively, these data indicate that VH1-69 usage favors broad neutralization and binding of HCV across multiple distinct antigenic sites. Of note, we also found that HEPC151-2 and HEPC158, which were biologically cloned from different B cells using limiting dilution and flow sorting, displayed identical heavy chain and light chain-variable gene sequences, indicating that this clonotype was relatively frequent among HCV-specific B cells in this subject. As we have previously observed, all MAbs, including bNAbs, were encoded by antibody genes with relatively sparse somatic mutations, ranging from 87% to 94% identity to their germ line heavy chain variable heavy (VH) gene sequences and 89% to 98% identity to their germ line light chain variable light (VL) gene sequences, indicating that extensive somatic hypermutation was not necessary for acquisition of broad neutralizing activity. TABLE 3 Germ line origin genes and variable region analysis of subject C18 MAbs axis and another MAb around the axis. (B) Pearson values of pairwise correlations between neutralization profiles of each C18 MAb (axis) and each reference MAb (axis). Only values that are statistically significant (< 0.05) are shown, and darker green color indicates a stronger positive correlation. The highest KY02111 KY02111 value for each C18 MAb is usually boxed and in strong type. C18 MAbs clustered into three functional groups, namely, AR3-like, AR1-like, and AR5-like. In many cases, neutralization profiles of C18 MAbs correlated best with neutralization profiles of reference MAbs that bound to the same antigenic site. As shown in Fig. 8B, neutralization profiles of C18 MAbs HEPC153, HEPC122, and HEPC154, which each target the AR3 antigenic site, showed the greatest correlation with reference MAbs AR3B, HEPC43, and AR3A, respectively, which are also AR3-site MAbs. Neutralization profiles of HEPC111 and HEPC130, which bind at the AR4-5 antigenic site, each showed the greatest KY02111 correlation with reference MAb AR5A, which also binds at this site. Similarly, the neutralization profile of HEPC167, which binds at the AR1 antigenic site, correlated best with the neutralization profile of the AR1A reference MAb. In other cases, neutralization profiles of C18 MAbs could not be predicted from the antigenic site targeted. Unexpectedly, the neutralization profile of HEPC151-1, which bound at the AR3 site, correlated best with reference MAb AR1A. The neutralization profile of HEPC108, which bound the AS108 site, correlated best with the neutralization profile of AR3A..