Figure 3. Analysis of RhCMV vectors with supertope-focused SIV inserts.

(A,B) Comparison of overall SIV Gag, Rev/Tat/Nef, and 5’-Pol CD8⁺ T cell responses elicited by a set of three miR-142-restricted (A) or miR-126-restricted (B) 68–1 RhCMV/SIV vectors individually expressing full length Gag, Retanef and 5’-Pol inserts versus the same responses elicited by a single miR-142-restricted or miR-126-restricted 68–1 RhCMV/SIV vector expressing either the MHC-II supertope- or the MHC-E supertope-focused insert (see fig. S7). Responses were determined by flow cytometric ICS assay using overlapping 15mer peptide mixes comprising the overall Gag, Rev/Tat/Nef, and 5’-Pol sequences, as described in Fig. 1. Two-sided Wilcoxon P-values are shown, comparing the area-under-the-curve (AUC) of response to full-length vs. supertope inserts adjusted for multiple comparisons. (C,D) Analysis of the same samples shown in panels A and B at the single 15mer epitope level, comparing responses to supertope peptides (red), adjacent non-supertope peptides in the supertope-focused insert (pink), and selected non-contiguous commonly recognized peptides (black) that are not present in the supertope-focused inserts for the both the miR-142-restricted (MHC-II-only) and miR-126-restricted (MHC-E-only) 68–1 RhCMV/SIV vectors. (E,F) Full analysis of CD8⁺ T cell responses to overlapping SIV Gag 15mers following vaccination with either the miR-142-restricted (E) or miR-126-restricted (F) 68–1 RhCMV/SIV vectors expressing the MHC-II- and MHC-E-supertope-focused inserts, respectively, in 5 representative RMs per vector, as described in Fig. 1. The blue and green shaded rectangles represent the 15mer peptides that are completely (15/15 amino acids) or mostly (11/15 amino acids) encoded in the MHC-II and MHC-E supertope-focused inserts, with the core supertope 15mers shown by arrowheads. Additional analyses are shown in fig. S8. ST – supertope; FL – full length.