Genetic polymorphisms of glutathione S-transferase A1, the major glutathione S-transferase in human liver: consequences for enzyme expression and busulfan conjugation.
BACKGROUND
High-dose busulfan is widely used as part of conditioning regimens for patients who are undergoing hematopoietic stem cell or bone marrow transplantation. High plasma concentrations of busulfan have been linked to the occurrence of hepatic venoocclusive disease (VOD), a severe complication associated with a high mortality. Because conjugation with glutathione, the major route of biotransformation of busulfan, is predominantly catalyzed by the isozyme glutathione S-transferase A1 (GSTA1), we hypothesized that low expression or function of GSTA1 in liver caused by genetic polymorphisms may be the mechanism underlying VOD.


METHODS
Immunoblot analysis of GSTA and measurement of busulfan-glutathione conjugation by liquid chromatography-mass spectrometry were performed in 48 normal human liver samples. To search for polymorphisms, the complete GSTA1 coding regions and the promoter fragment were sequenced. All results were compared by multivariate analysis.


RESULTS
Absolute levels of GSTA protein and formation rates of busulfan-glutathione conjugate displayed a 7- and 8-fold range, from 240 to 1600 pmol/mg and 25 to 205 pmol/min per milligram of total cytosolic protein, respectively, and correlate (r2 = 0.49, P <.0001). A total of 8 single nucleotide polymorphisms (SNPs) of GSTA1 were identified, 1 of which was a silent mutation in exon 5 (A375G); all others were found in the promoter region. Haplotype analysis revealed the existence of 5 defined alleles. There was no significant relationship between any of the GSTA1 SNPs or haplotypes and either hepatic glutathione S-transferase A (GSTA) expression or GSTA1 function.


CONCLUSIONS
The identified GSTA1 polymorphisms are not likely to be related to the VOD because they do not appear to be associated with changes in GSTA expression or function. Compared with other members of the GST family, GSTA1 displays surprisingly little variation.
Bredschneider
Monika
M
Dr Margarete Fischer-Bosch Institute of Clinical Pharmacology, Stuttgart, Germany.
Klein
Kathrin
K
Mürdter
Thomas E
TE
Marx
Claudia
C
Eichelbaum
Michel
M
Nüssler
Andreas K
AK
Neuhaus
Peter
P
Zanger
Ulrich M
UM
Schwab
Matthias
M
D002066:Busulfan / Q000378:metabolism*; D017931:DNA Primers; D015971:Gene Expression Regulation, Enzymologic; D005838:Genotype; D005982:Glutathione Transferase / Q000235:genetics* / Q000378:metabolism*; D006239:Haplotypes; D006801:Humans; D015151:Immunoblotting; D007166:Immunosuppressive Agents / Q000378:metabolism*; D015999:Multivariate Analysis; D010641:Phenotype; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D011401:Promoter Regions, Genetic / Q000235:genetics
D017931:DNA Primers; D007166:Immunosuppressive Agents; D005982:Glutathione Transferase; D002066:Busulfan