Table 4 Summary of key research assays
Assay | Sample | Purpose | Time-point |
|---|---|---|---|
T cell immunology | • Pre-ART where available • On ART • Pre-bNAb • Fortnightly during ATI • Following viral suppression | ||
HIV-specific T cell intracellular cytokine staining for CD4 and CD8 phenotypic and functional responses | PBMC | Screen CD4 and CD8 T cell responses to HIV in response to bNAbs | |
HIV ELISpots: γ-interferon responses with peptides targeted according to ICS responses | PBMC | Higher resolution analyses of specific responses to peptide level and how they are impacted by intervention | |
CD8 tetramer responses targeted to specific peptides | PBMC | Further in-depth resolution of CD8 T cell functionality at cellular level | |
HLA Class I and II typing (4-digit) | PBMC | Profiling of HLA alleles to shape immune responses and confer advantage/disadvantages | |
T cell killing assays | PBMC | Functional killing assay to augment intracellular cytokine staining data | |
Natural Killer (NK) cell responses | |||
Flow cytometry for NK cell phenotype and function | PBMC | Characterisation of NK function and profile in response to bNAbs | |
ADCC and NK cell killing | PBMC | Functional assessment of bNAbs using characterised killing protocols | |
NK-like population analyses (flow cytometry) | PBMC | High resolution characterisation of cells that fall between the innate and adaptive responses | |
Virology | |||
Next-generation sequencing (NGS) single genome amplification (SGA) of HIV ENV: bNAb sensitivity | PBMC | Env sequencing to screen for bNAb sensitivity | Screening |
Full length HIV sequencing | Plasma CD4 T cells | Full length haplotype analysis of viral and proviral populations pre and post intervention to determine correlations with response | • Pre-ART (virus) where available (HEATHER cohort) • On ART, and pre-ATI (provirus) • Rebound (virus) • Post resuppression (provirus) |
HIV Integration site analysis | CD4 T cells | Understanding of clonality of viral reservoir and relationship of integration sites to reactivation potential and susceptibility to inhibition | Pre-ATI |
HIV DNA and cell-associated RNA quantitation | CD4 T cells | Standard molecular analyses of proviral and HIV transcript quantitation, both surrogate markers of persistent infection and biomarkers of remission and rebound | • Pre-ART where available (HEATHER cohort) • On ART • Pre-bNAb • Pre-ATI • Post resuppression |
Host genomics | |||
Transcriptome profiling* *Initially bulk, but with single-cell 10× resolution dependent on results | CD4 and CD8 T cells NK cells | Full host genomic analyses at bulk and single cell (likely tetramer driven for CD8 cells) to understand host determinants of outcomes | • Pre-ART where available (HEATHER cohort) • On ART • Pre- ATI |
Pharmacokinetics and anti-drug antibodies | |||
bNAb pharmacokinetics Quantitation of ‘anti-drug antibodies’ (ADA) | Plasma | To determine bNAb plasma concentrations To determine development of inhibitory antibodies against bNAbs | • Post bNAb infusion |