ORGANIC CATION TRANSPORTERS - Drug Transporters: Molecular Characterization and Role in Drug Disposition - page 25

2.7. GENETIC VARIATION

Much attention has recently been focused on the pharmacogenetics of drug trans-
porters (i.e., the effect of natural human genetic variation in drug transporter genes
on drug disposition and drug response). Research in this area has taken primarily a
sequence-based approach, beginning with identification of genetic variants likely to
produce a functional effect (e.g., amino acid substitutions, or insertions or deletions
in the coding region), followed in some cases by functional studies of the variant
proteins in heterologous expression systems.

2.7.1. OCT1

Several functionally significant polymorphisms in OCT1 have been described. In a
sample of 57 healthy Caucasians, 25 genetic variants were discovered, of which eight
resulted in a change in protein sequence. 69 Of these, five (Arg61Cys, Cys88Arg,
Phe160Leu, Gly401Ser, and Met420del) were tested for function by measurement
of transport activity in X. laevis oocytes. Cys88Arg and Gly401Ser showed virtually
complete loss of activity toward MPP + , while Arg61Cys retained approximately 30%
of wild-type activity. In contrast to MPP + , uptake of [ 3 H]serotonin by Cys88Arg
and Gly401Ser was detectable ( 10% of wild-type), suggesting that these variants

influence the substrate selectivity of OCT1.
Shu et al. discovered 15 amino acid sequence-altering variants of OCT1 that were
identified in a large sample of ethnically diverse healthy subjects, and tested these
for function by uptake of MPP + in X. laevis oocytes. 70 Variants with significantly re-
duced or complete loss of function included the Arg61Cys and Gly401Ser described
by Kerb et al. 69 , as well as Pro341Leu, Gly220Val, and Gly465Arg. An additional
variant, Ser14Phe, showed a significant increase in activity; interestingly, this vari-
ant corresponds to the likely ancestral allele, as the consensus mammalian OCT1
sequence includes phenylalanine at this position. It was noted that all of the variants
with reduced function occurred at evolutionarily conserved amino acid residues, and
that variants with reduced function tended to be amino acid substitutions that result
in a large chemical change. Of the variants with significant functional differences
from the reference OCT1, five (Ser14Phe, Arg61Cys, Pro341Leu, Gly401Ser, and
Gly465Arg) occurred at

∼

1% allele frequency in at least one ethnic group and are at-
tractive candidates for association with drug response phenotypes for OCT1 substrate
drugs.
Functional studies of several additional OCT1 variants have been performed by
two independent groups using microinjected X. laevis oocytes 71 or transiently trans-
fected HEK293 cells. 72 Three amino acid sequence variants, Pro283Leu, Arg287Gly,
and the previously described Pro341Leu, showed significantly reduced transport ac-
tivity, with Pro283Leu and Arg287Gly having no appreciable activity toward the
model substrates TEA and MPP + . The reduction in activity of these variants was not
explained by reduction in protein expression, as immunofluorescence studies showed
all three variants to have membrane expression levels similar to that of the wild-type
OCT1. 72

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