Functional analysis of novel polymorphisms in the human SLCO1A2 gene that encodes the transporter OATP1A2

Fanfan Zhou, Jian Zheng, Ling Zhu, Andreas Jodal, Pei H. Cui, Mark Wong, Howard Gurney, W. Bret Church, Michael Murray

Research output: Contribution to journalArticleResearchpeer-review

Abstract

The solute carrier organic anion transporting polypeptide 1A2 (OATP1A2, SLCO1A2) is implicated in the cellular influx of a number of drugs. We identified five novel single nucleotide polymorphisms (SNPs) in coding exons of the SLCO1A2 gene in a cohort of subjects: G550A, G553A, G673A, A775C, and G862A, that encoded the OATP1A2 variants E184K, D185N, V255I, T259P, and D288N, respectively. The function and expression of these variant transporters were assessed in HEK-293 cells. We found that the novel variants, E184K, D185N, T259P, and D288N, were associated with impaired estrone-3-sulfate, imatinib, and methotrexate transport (∼20-50% of wild-type control); function was retained by OATP1A2-V255I. From biotinylation assays, the decreased function of these variants was due, at least in part, to impaired plasma membrane expression. The four loss-of-function variants were studied further using mutagenesis to produce variants that encode residues with different charges or steric properties. From immunoblotting, the replacement of negatively charged residues at amino acid positions 184 and 185 impaired membrane expression, while either a positive or negative charge at residue 288 supported the correct membrane targeting of OATP1A2. Replacement of T259 with bulky residues disrupted transporter stability. From molecular models, E184, D185, and D288 were located near several charged residues such that intramolecular ionic interactions may stabilize the transporter structure. Individuals who carry these novel SNPs in the SLCO1A2 gene may be at risk from impaired efficacy or enhanced toxicity during treatment with drugs that are substrates for OATP1A2.

LanguageEnglish
Pages1099-1108
Number of pages10
JournalAAPS Journal
Volume15
Issue number4
DOIs
Publication statusPublished - 2013
Externally publishedYes

Fingerprint

Single Nucleotide Polymorphism
Biotinylation
Molecular Models
Membranes
HEK293 Cells
Immunoblotting
Methotrexate
Mutagenesis
Pharmaceutical Preparations
Genes
Anions
Exons
Cell Membrane
Amino Acids
Peptides
Imatinib Mesylate
estrone sulfate

Cite this

Zhou, F., Zheng, J., Zhu, L., Jodal, A., Cui, P. H., Wong, M., ... Murray, M. (2013). Functional analysis of novel polymorphisms in the human SLCO1A2 gene that encodes the transporter OATP1A2. AAPS Journal, 15(4), 1099-1108. https://doi.org/10.1208/s12248-013-9515-1
Zhou, Fanfan ; Zheng, Jian ; Zhu, Ling ; Jodal, Andreas ; Cui, Pei H. ; Wong, Mark ; Gurney, Howard ; Church, W. Bret ; Murray, Michael. / Functional analysis of novel polymorphisms in the human SLCO1A2 gene that encodes the transporter OATP1A2. In: AAPS Journal. 2013 ; Vol. 15, No. 4. pp. 1099-1108.
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abstract = "The solute carrier organic anion transporting polypeptide 1A2 (OATP1A2, SLCO1A2) is implicated in the cellular influx of a number of drugs. We identified five novel single nucleotide polymorphisms (SNPs) in coding exons of the SLCO1A2 gene in a cohort of subjects: G550A, G553A, G673A, A775C, and G862A, that encoded the OATP1A2 variants E184K, D185N, V255I, T259P, and D288N, respectively. The function and expression of these variant transporters were assessed in HEK-293 cells. We found that the novel variants, E184K, D185N, T259P, and D288N, were associated with impaired estrone-3-sulfate, imatinib, and methotrexate transport (∼20-50{\%} of wild-type control); function was retained by OATP1A2-V255I. From biotinylation assays, the decreased function of these variants was due, at least in part, to impaired plasma membrane expression. The four loss-of-function variants were studied further using mutagenesis to produce variants that encode residues with different charges or steric properties. From immunoblotting, the replacement of negatively charged residues at amino acid positions 184 and 185 impaired membrane expression, while either a positive or negative charge at residue 288 supported the correct membrane targeting of OATP1A2. Replacement of T259 with bulky residues disrupted transporter stability. From molecular models, E184, D185, and D288 were located near several charged residues such that intramolecular ionic interactions may stabilize the transporter structure. Individuals who carry these novel SNPs in the SLCO1A2 gene may be at risk from impaired efficacy or enhanced toxicity during treatment with drugs that are substrates for OATP1A2.",
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Zhou, F, Zheng, J, Zhu, L, Jodal, A, Cui, PH, Wong, M, Gurney, H, Church, WB & Murray, M 2013, 'Functional analysis of novel polymorphisms in the human SLCO1A2 gene that encodes the transporter OATP1A2', AAPS Journal, vol. 15, no. 4, pp. 1099-1108. https://doi.org/10.1208/s12248-013-9515-1

Functional analysis of novel polymorphisms in the human SLCO1A2 gene that encodes the transporter OATP1A2. / Zhou, Fanfan; Zheng, Jian; Zhu, Ling; Jodal, Andreas; Cui, Pei H.; Wong, Mark; Gurney, Howard; Church, W. Bret; Murray, Michael.

In: AAPS Journal, Vol. 15, No. 4, 2013, p. 1099-1108.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Zhou, Fanfan

AU - Zheng, Jian

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