Modification of the cyclopropyl moiety of abacavir provides insight into the structure activity relationship between HLA‐B*57:01 binding and T‐cell activation

Abstract

Background Abacavir is associated with hypersensitivity reactions in individuals positive for the HLA‐B*57:01 allele. The drug binds within the peptide binding groove of HLA‐B*57:01 altering peptides displayed on the cell surface. Presentation of these HLA‐abacavir‐peptide complexes to T‐cells is hypothesized to trigger a CD8+ T‐cell response underpinning the hypersensitivity. Thus, the aim of this study was to explore the relationship between the structure of abacavir with HLA‐B*57:01 binding and the CD8+ T‐cell activation. Methods Seventeen abacavir analogues were synthesized and cytokine secretion from abacavir/abacavir analogue‐responsive CD8+ T‐cell clones was measured using IFN‐γ ELIspot. In silico docking studies were undertaken to assess the predicted binding poses of the abacavir analogues within the HLA‐B*57:01 peptide binding groove. In parallel, the effect of selected abacavir analogues on the repertoire of self‐peptides presented by cellular HLA‐B*57:01 was characterized using mass spectrometry. Results Abacavir and ten analogues stimulated CD8+ T‐cell IFN‐γ release. Molecular docking of analogues that retained antiviral activity demonstrated a relationship between predicted HLA‐B*57:01 binding orientations and the ability to induce a T‐cell response. Analogues that stimulated T‐cells displayed a perturbation of the natural peptides displayed by HLA‐B*57:01. The antigen‐specific CD8+ T‐cell response was dependent on the enantiomeric form of abacavir at both cyclopropyl and cyclopentyl regions. Conclusion Alteration of the chemical constitution of abacavir generates analogues that retain a degree of pharmacological activity, but have variable ability to activate T‐cells. Modelling and immunopeptidome analysis delineate how drug HLA‐B*57:01 binding and peptide display by antigen presenting cells relate to the activation of CD8+ T‐cells.

CD8+T-cell landscape in Indigenous and non-Indigenous people restricted by influenza mortality-associated HLA-A*24:02 allomorph

“…Peptide/MHC complexes were dissociated, and fractionated by reversed phase high performance liquid chromatography (RP-HPLC) as described 27,38,74 . 500µL fractions were collected throughout the gradient, and the peptide containing fractions combined into 9 pools, vacuum-concentrated and reconstituted in 15µL 0.1% formic acid (Honeywell) in Optima™ LC-MS water.…”

“…500µL fractions were collected throughout the gradient, and the peptide containing fractions combined into 9 pools, vacuum-concentrated and reconstituted in 15µL 0.1% formic acid (Honeywell) in Optima™ LC-MS water. Reconstituted fraction pools were analysed by LC-MS/MS using a SCIEX 5600+ TripleTOF mass spectrometer equipped with a Nanospray III ion source as previously described 74 .…”

In-Vitro Approaches to Predict and Study T-Cell Mediated Hypersensitivity to Drugs

“…This was achieved via the prolonged incubation of C1R-B*57:01 cells with abacavir followed by peptide elutions from the class I MHC and analysis by mass spectrometry (m/s). This was further reinforced in 2019 when abacavir analogues with a similar T-cell liability were found to perturb the HLA-B*57:01 peptide repertoire in a similar convention to abacavir, while those with no T-cell liability did not ( 180 ). This concept was further explored via the use of the β-lactam antibiotic flucloxacillin which was identified to covalently haptenate HLA-B*57:01 native peptides which were subsequently processed and presented on the MHC for T-cell recognition.…”

“…Proof-of-concept iterative medicinal chemistry studies in pursuit of an analogue of abacavir devoid of hypersensitivity liabilities with preserved pharmacological action have embodied a promising prototypical approach to disconnect pharma-and immunocophores. Cross-disciplinary laboratories operated using in-silico docking models alongside functional studies to simultaneously decipher pharmacological (anti-viral) and immunological (T-cell activation) structure-activity-relationships of compound series (152,153). Digressing from such idealistic goals, many of the following experimental assays have essentially been devised to address various aspects that lie within the void of knowledge between such conventional theoretical chemistry-based wisdom, and pragmatic transition of a compound to clinical use.…”

Advances and novel developments in drug hypersensitivity diagnosis

“…Abacavir DHRs have been related with the allele HLA‐B*57:01. Recently, abacavir analogues retaining their antiviral capacity but not altering the HLA‐B*57:01 peptide binding repertoire, and thus avoiding CD8 + T‐cell activation, were synthetized 98 . Furthermore, this allele has been associated with SCARs induced by other drugs like carbamazepine (CBZ), increasing the risk of SJS/TEN but not DRESS in the European population 99 .…”