The glycosylation of AGP and its associations with the binding to methadone

Abstract

Methadone remains the most common form of pharmacological therapy for opioid dependence; however, there is a lack of explanation for the reports of its relatively low success rate in achieving complete abstinence. One hypothesis is that in vivo binding of methadone to the plasma glycoprotein alpha-1-acid glycoprotein (AGP), to a degree dependent on the molecular structure, may render the drug inactive. This study sought to determine whether alterations present in the glycosylation pattern of AGP in patients undergoing various stages of methadone therapy (titration < two weeks, harm reduction < one year, long-term > one and a half years) could affect the affinity of the glycoprotein to bind methadone. The composition of AGP glycosylation was determined using high pH anion exchange chromatography (HPAEC) and intrinsic fluorescence analysed to determine the extent of binding to methadone. The monosaccharides galactose and N-acetyl-glucosamine were elevated in all methadone treatment groups indicating alterations in AGP glycosylation. AGP from all patients receiving methadone therapy exhibited a greater degree of binding than the normal population. This suggests that analysing the glycosylation of AGP in patients receiving methadone may aid in determining whether the therapy is likely to be effective.

Figure 1

Relative level of AGP isolated from treatment groups. Summary of the mean level of AGP isolated from individuals at different stages of methadone therapy. *Statistically different to “normal” blood (P < 0.05).

Figure 2

Mean level of monosaccharides in AGP glycans isolated from treatment groups. “Normal blood” represents heparinised blood from the Blood Transfusion Service. *Statistically significant difference compared to normal heparinised blood sample (P < 0.05) HR: harm reduction; LT: long-term treatment. Standard deviation calculated from triplicate analysis of a single sample while others were calculated from data from more than one patient (n = 4/+).

Figure 3

Oligosaccharide library trace generated during separation by HPAEC. Separation based on the charge of oligosaccharide chains as measured using nano-Coulomb (nC). Degree of sialylation highlighted in a dashed line.