Metabolomics reveals metabolite profiles associated with clinical cure of chronic HBV infection under different drug interventions

Objectives: To identify differential metabolites associated with complete cure in chronic hepatitis B patients treated with various regimens of entecavir and pegylated interferon-alpha. Methods: The retrospective, case-control study was conducted at the Mudan People's Hospital of Heze, Shandong, China, and comprised samples from the institutional laboratory department of hepatitis B virus patients enrolled between June 2, 2021, and December 30, 2023. The patients were grouped based on the treatment protocol and the evidence of complete cure. Groups A and B comprised patients who were and were not clinically cured after receiving 48 weeks of sequential entecavir and pegylated interferon-alpha. Group E patients received entecavir monotherapy for over 2 years without a clinical cure, and group W included untreated patients. Ultra-performance liquid chromatography-mass spectrometry and metabolomics databases were utilised to identify differential metabolites and networks indicative of clinical cure. Data was analysed using the R language Ropls package. Results: Of the 50 patients, 13(26%) each were in Group A and Group B, while 12(24%) each were in Group E and Group W. The median age was similar across groups (Group A: 42 [40-47] years; Group B: 43 [40-49] years; Group E: 43.5 [39-48] years; Group W: 43 [40-48] years), and male patients predominated in each group (Group A: 10(77%); Group B: 10(77%); Group E: 9(75%); Group W: 9(75%)). A total of 52 differential metabolites showing altered abundance in Group A compared with the other groups were identified. Notably, 9,10-dihydroxy-12Z-octadecenoic acid, 12,13-dihydroxy-9Z-octadecenoic acid and kaempferide were common differential metabolites across all intergroup comparisons. Beyond these common metabolites, the top-ranked differential metabolites unique to each comparison included guanidoacetic acid, caffeine and uracil 5-carboxylate in Group A versus Group B; 3-ketosphingosine in Group A versus Group E; and L-2-hydroxyglutaric acid and 3,4-dihydroxybenzeneacetic acid in Group A versus Group W. The fold-change values were 1.28 for 9,10-DHOME, 1.12 for 12,13-DHOME and 1.33 for kaempferide, and guanidoacetic acid ranked first among the differential metabolites in Group A versus Group B (fold-change: 1.27, p=2.56×10-8). These metabolites were enriched in 42 metabolic pathways, including linoleic acid, caffeine and amino acid metabolism. Conclusions: Kaempferide, 9,10-dihydroxy-12Z-octadecenoic acid and 12,13-dihydroxy-9Z-octadecenoic acid were common differential metabolites, indicating potential significance for achieving clinical cure in chronic hepatitis B virus infection. Guanidoacetic acid, caffeine, uracil 5-carboxylate, 3-ketosphingosine, L-2-hydroxyglutaric acid, and 3,4-dihydroxybenzeneacetic acid were the top three differential metabolites in each group beyond the common ones.