Discovering metabolic pathways associated with immune activation in people living with HIV following ChAdOx1 nCoV-19 vaccination using mass spectrometry.

Immune response to COVID-19 vaccination in people living with HIV (PLHIV) remains unexplored, particularly from a metabolic perspective. This study used high-resolution mass spectrometry-based plasma metabolomics to investigate metabolic changes following ChAdOx1 nCoV-19 vaccination in PLHIV and HIV-negative (HN) populations. Plasma samples from 54 PLHIV and 69 HN were analyzed in a cross-sectional design and grouped by vaccination status. A validated UHPLC-MS/MS method was developed and assessed for environmental sustainability using the Analytical Eco-Scale (AES), Analytical GREEnness Metric (AGREE), and RGBfast-based whiteness assessment. The method exhibited strong analytical performance (RSD < 20 % for the internal standard, 110 % recovery) and favorable sustainability (AGREE: 0.53, AES: >81, Whiteness: 65), outperforming comparable published methods. Untargeted metabolomics revealed 213 differentially expressed metabolites (DEMs) in HN after the third dose, compared with 194 DEMs in PLHIV (log2 FC ≥ 0.25 or ≤ -0.25; p < 0.05) groups. Partial least squares-discriminant analysis (PLS-DA) revealed 53 key endogenous metabolites in HN and 70 in PLHIV (VIP >1, FDR< 0.05) groups. Pathway enrichment analysis revealed 23 key metabolites with significant immunological relevance, particularly within amino acid, butanoate, and lipid metabolism in both the HN and PLHIV groups. Arachidonic acid metabolism emerged as one of the most affected pathways, with > 2-fold upregulation (p < 0.05) of pro-inflammatory lipid mediators, including leukotriene B4 (LTB4) and prostaglandin E2 (PGE2). These results indicate that vaccination induces marked metabolic reprogramming in PLHIV, particularly within lipid-derived inflammatory pathways, providing novel insights into vaccine-induced immune regulation in the PLHIV.