Single nucleotide polymorphisms result in impaired membrane localization and reduced atpase activity in multidrug transporter ABCG2

Abstract

ABCG2/MXR/ABCP1/BCRP is a member of the ATP-binding cassette membrane transporter, which consists of six transmembrane regions and one ATP-binding cassette. The transporter is known to be involved in the efflux of various anticancer compounds such as mitoxantrone, doxorubicin and topoisomerase I inhibitor. In this study, we analyzed the effects of polymorphisms in ABCG2, V12M and Q141K on transporter function. When polarized LLC-PK1 cells were transfected with variant ABCG2, drug-resistance to topoisomerase I inhibitor of cells expressing V12M or Q141K was less than 1/10 that of wild-type ABCG2 transfected cells, and was accompanied by increased drug accumulation and decreased drug efflux in the variant ABCG2-expressing cells. We further elucidated the molecular mechanisms of the transport dysfunction by investigating membrane localization and ATPase activity. Confocal microscopic analysis revealed that apical plasma membrane localization of V12M was disturbed, while the localization of wild-type transporters occurred specifically in the apical plasma membrane of polarized LLC-PK1 cells. Also, ATPase activities measured in the membrane of SF9 cells infected with variant ABCG2 showed that Q141K decreased activity by 1.3 below that of wild-type ABCG2. In addition, kinetic analysis of ATPase activity showed that the K m value in Q141K was 1.4-fold higher than that of wild-type ABCG2. These results indicated that naturally occurring SNPs alter transport functions of ABCG2 transporter and analysis of SNPs in ABCG2 may hold great importance in understanding the response/metabolism of chemotherapy compounds that act as substrates for ABCG2.

Synergistic interaction of ABCB1 and ABCG2 polymorphisms predicts the prevalence of toxic encephalopathy during anticancer chemotherapy

“…9,[12][13][14][15] The results are also concordant with the findings of Wang et al 11 who suggested that among linked ABCB1 SNPs, the 3435T4C silent variation is responsible for the differences in gene expression.…”

“…The twelfth amino acid is placed within the first transmembrane domain and the 12M allele may disturb the protein's proper membrane localization resulting in decreased cellular transport. 12,13 Data on the functional relevance of these SNPs and the controversies have been reviewed recently by many authors. 14,15 In order to assess the possible value of these SNPs in the prediction of neurotoxic side effects of chemotherapy, we have studied a large cohort of patients treated for childhood acute lymphoblastic leukaemia (ALL).…”

Subcellular localization of the ABCG2 transporter in normal and malignant human gallbladder epithelium

“…42 Also, impaired membrane localization caused by single-nucleotide polymorphisms results in a decline of ABCG2 ATPase activity. 43 Whether this applies to ABCG2 function in the in vivo situation must be established.…”

Determination of the Single Nucleotide Polymorphisms C3435 and G2677T in MDR1 and C421A in BCRP in Blood Samples of Patients with Inflammatory Bowel Disease and Healthy Controls in the Swiss Population

“…In vitro and in vivo studies, on the other hand, have shown this SNP to be disruptive of BCRP activity. Clinical trials with diflomotecan, a novel anticancer agent inhibiting topoisomerase I, showed decreased plasma concentration in patients heterozygous for this SNP compared to wild type [33,34]. Recently, the increased placental crossing of the antidiabetic agent glyburide (glibenclamide) has been demonstrated in stably transfected human embryonic kidney (HEK)-293 cells expressing the polymorphic ABCG2 (Q141K) [35].…”