Abstract
Human immunodeficiency virus (HIV) remains a significant public health issue. In recent years, passive immunization with broadly neutralizing antibodies (bNabs) is being considered as a potentially efficacious approach for fighting HIV. One candidate that holds great promise is represented by the CD4-binding site targeted bNab capable of neutralizing over 90% of circulating HIV strains, VRC01. VRC01 along with its variants and clonal relatives - VRC01-LS and VRC07-523LS are currently being evaluated as vaccines in a number of clinical trials for HIV treatment and prevention. While mucosal areas of the body serve as major ports of HIV entry, reliable quantification of bNabs for pharmacokinetic and bioavailability analyses has been challenging due to low antibody concentrations in these samples. We developed an immunoassay on the Singulex platform which enables ultra-sensitive quantification of VRC01, VRC07, VRC01-LS and VRC07-523LS with a greater than 4-log linear dynamic range (LDR) and less than 120 pg/mL lower limit of quantitation (LLOQ). We implemented this assay to quantify VRC01 levels in rectal, cervical and oral mucosal samples in two passive immunization studies conducted with VRC01 - VRC 601 and VRC 602. Our assay was able to successfully quantify VRC01 levels in mucosal samples from all dosage groups (5 - -40 mg/kg) in these trials. VRC01 levels in a significant proportion of these samples (37% in oral and 25% in rectal mucosa) were below the lower limits of quantitation of other traditional immunoassays used for VRC01 quantification. We also measured VRC01 levels in sera from these trials and found that VRC01 measurements made using our assay exhibited excellent correlation (r2 = 0.9509) with measurements made previously using Enzyme-linked immunosorbent assay (ELISA). Our assay provides a reliable, sensitive and accurate method for quantification of clinically relevant bNabs and will help delineate antibody infiltration and bioavailability characteristics in complex biological matrices (CBM) such as mucosal tissues. This will in turn help determine clinical antibody threshold concentrations required to mediate protection against HIV acquisition and serve to inform dosing regimens and clinical trial design for future efficacy trials with these bNabs.