Antibodies targeting the HIV Envelope (Env) protein can limit viral infection and provide immunity. However, the field has yet to generate a vaccine capable of producing protective antibodies in humans. With epitope-based vaccine design we can engineer immunogens to elicit antibodies against specific regions of Env, but we are limited in part by the low throughput nature of current epitope mapping methods. Here we developed and tested a high-throughput, comprehensive approach to map the epitopes of recently identified HIV-specific monoclonal antibodies (mAbs) that mediate ADCC.
We applied a phage display method that used deep sequencing, Phage Immunoprecipitation-Sequencing (PhIP-seq), to identify the linear epitopes of four newly identified gp41-specific monoclonal antibodies (mAbs): QA255.006, QA255.016, QA255.067, and QA255.072 as well as 240-D, a gp41-specific antibody previously mapped using peptide arrays. We first generated a synthesized oligonucleotide library encoding for 39 amino acid long peptides that tile along the entire length of several Env and full-length HIV sequences from different clades. This library was cloned into a T7 bacteriophage display vector. To perform a PhIP-seq experiment, mAbs were coated on beads and incubated with the phage library, samples were sequenced in parallel, and then peptides specifically enriched by the mAb were computationally identified. Competition ELISAs were performed to compare epitope mapping results using a more traditional approach.
We mapped the linear epitope of QA255.067 and QA255.072 to Env amino acids 592-606 and 596-609 (HXB2 numbering), a region corresponding to the immunodominant C-C loop region of gp41. Competition ELISAs confirmed these results. We also more finely mapped the epitope of 240-D to amino acids 596-605, which is consistent with findings from structural studies. We were unable to see specific enrichment of any peptides in PhIP-seq with QA255.006 and QA25.016, but competition ELISA results indicated these mAbs target a discontinuous epitope on gp41.
PhIP-seq mapped overlapping but distinct epitopes of two newly identified gp41 mAbs and 240-D. This method may be useful for mapping HIV antibodies that target linear epitopes, and particularly, antibodies that recognize the gp41 protein, which is an attractive vaccine target because it is relatively conserved.