Role of SLC22A1 Polymorphic Variants in Drug Disposition, Therapeutic Responses, and Drug–Drug Interactions

REVIEW Role of SLC22A1 polymorphic variants in drug disposition, therapeutic responses, and drug–drug interactions

C Arimany-Nardi1, H Koepsell2 and M Pastor-Anglada1

The SCL22A1 gene encodes the broad selectivity transporter hOCT1. hOCT1 is expressed in most epithelial barriers thereby contributing to drug pharmacokinetics. It is also expressed in different drug target cells, including immune system cells and others. Thus, this membrane protein might also contribute to drug pharmacodynamics. Up to 1000 hOCT1 polymorphisms have been identified so far, although only a small fraction of those have been mechanistically studied. A paradigm in the field of drug transporter pharmacogenetics is the impact of hOCT1 gene variability on metformin clinical parameters, affecting area under the concentration–time curve, Cmax and responsiveness. However, hOCT1 also mediates the translocation of a variety of drugs used as anticancer, antiviral, anti-inflammatory, antiemetic agents as well as drugs used in the treatment of neurological diseases among. This review focuses exclusively on those drugs for which some pharmacogenetic data are available, and aims at highlighting the need for further clinical research in this area.

The Pharmacogenomics Journal (2015) 15, 473–487; doi:10.1038/tpj.2015.78; published online 3 November 2015

SLC22 GENE FAMILY HOCT1 The SLC22 gene family encodes for more than 30 proteins, some The first organic cation transporter (rOCT) was cloned from rat of them acting as organic cation transporters (OCTs), organic kidney in 1994.4 Some years later the mouse (mOCT1)5 and rabbit cation and zwitterion transporters (OCTNs) and organic anion orthologues (rbOCT1)6 were isolated. hOCT1 was independently transporters (OATs). OCTs, OCTNs and organic anion transporters, cloned by two groups.7,8 A difference between both human show broad substrate selectivity, being able to transport multiple complementary DNA clones resulting in three amino-acid compounds different in size and structure. substitutions was observed being the human orthologue 78% identical to rOCT1.9 hOCT1 is encoded by the SLC22A1 gene, which contains 11 exons and 10 introns localised on chromo- HUMAN ORGANIC CATION TRANSPORTERS (HOCTS) some 6.q26-7. 10 Although OCT1 is highly conserved in mammals, Organic cation transporters (OCT1-3), such as many transporters in only a few amino acids are also conserved in fish, Drosophila the SLC22 family, have a predicted membrane topology of melanogaster or Caenorhabditis elegans (Figure 1). 12 α-helical transmembrane domains (TMDs), an intracellular N OCT1 protein is expressed mostly in epithelial cells. Regarding terminus, a large glycosylated extracellular loop between TMDs 1 its tissue distribution, hOCT1 is mainly expressed in the liver, and 2, a large intracellular loop with phosphorylation sites where it is located at the sinusoidal membrane of the between TMDs 6 and 7 and an intracellular C terminus. OCTs hepatocytes.11 OCT1 orthologues in rat, mouse and rabbit are are equilibrative transporters. Translocation of cations by these also expressed strongly in kidney, although SLC22A1 messenger transporters is also electrogenic. Thus, the membrane potential RNA abundance in human renal tissue appears to be low.10,12 might serve as a driving force in addition to the concentration Nevertheless, the OCT1 protein has been identified although gradient. They can translocate a broad variety of substrates with differentially localized in rat (basolateral membrane of proximal highly variable molecular structures, and similarly, they can also be tubules),10 and human kidneys (localised at the apical and inhibited by a large number of compounds, which are able to subapical domains of both proximal and distal tubules).12 interact with the transporters without being translocated. OCTs hOCT1 is also expressed in many other organs including small are potentially bidirectional transporters able to interact with intestine, at the brush border membrane; lung, at the luminal inhibitors with different affinities depending on the site of drug– membrane of bronchial epithelial cells; heart; skeletal muscle; transporter interaction.1 OCT substrates include endogenous compounds, drugs, xeno- brain, in endothelial cells of microvessels; placenta; mammary gland; adrenal gland; eye, in cornea, iris–ciliary body and blood– biotics as well as a variety of model substrates, which are routinely 10,13–20 used for OCT function assays. Most substrates and inhibitors retina barrier; adipose tissue; and immune cells (Table 1). of OCTs broadly overlap among the different members of the hOCT1 expression in tumours has also been broadly 10,21–23 family.2,3 reported, although in some cases its expression levels

1Molecular Physiology and Experimental Therapeutics, Department of Biochemistry and Molecular Biology, Institute of Biomedicine (IBUB), University of Barcelona and Oncology Program, National Biomedical Research Institute on Liver and Gastrointestinal Diseases (CIBER EHD), Instituto de Salud Carlos III, Barcelona, Spain and 2Department of Molecular Plant Physiology and Biophysics, Julius-von-Sachs-Institute, University Würzburg, Würzburg, Germany. Correspondence: Professor M Pastor-Anglada, Departament de Bioquímica i Biologia Molecular, Universitat de Barcelona, Avinguda Diagonal 643, Barcelona 08028, Spain. E-mail: [email protected] Received 5 February 2015; revised 20 August 2015; accepted 8 September 2015; published online 3 November 2015 Drug pharmacogenetics and SLC22A1 polymorphisms C Arimany-Nardi et al 474 may be downregulated when compared with surrounding normal bases and some non-charged compounds. Substrates of human tissue, as recently shown for liver cancer.24 OCT1 include model cations, endogenous compounds and drugs In summary, the anatomical distribution of hOCT1 in most among others. Some of the model compounds used to asses epithelial barriers anticipates a major role for this membrane hOCT1 activity are tetraethylammonium, 1-methyl-4-phenyl- transporter in controlling drug pharmacokinetics and, eventually, pyridinium and 4-[4-(dimthylamino)-styryl]-N-methylpyridinium. pharmacodynamics. hOCT1 was originally included in the ﬁrst set Endogenous compounds transported by human OCT1 include of human transporters relevant to drug development by the choline, acetylcholine, the neuromodulators histidyl-proline dike- 25 International Transporter Consortium. topiperazine (cyclo(His-Pro)) and salsolinol, the L-arginine meta- Human OCT1 can transport not only monovalent organic bolite agmatine, the polyamine putrescine and vitamine B1.10,26–28 cations, but also some divalent organic cations, some weak Drugs transported by hOCT1 include metformin, used for the

B. taurus ------MLTVDDVLEQVGEFGWFQKQTFLILCLLSAAFAPIYVG 38 C. elegans MSFQAMETFAEISQEILMSATKPPDFDFVLEQVGNYGTYQIVFFFIICLPTSLPSAFSAF 60 D. melanogaster ------MGYDDVITHLGEFGPYQKRIYYLLCLP-AIVCAFHKL 36 H. sapiens ------MPTVDDILEQVGESGWFQKQAFLILCLLSAAFAPICVG 38 M. musculus ------MPTVDDVLEHVGEFGWFQKQAFLLLCLISASLAPIYVG 38 R. norvegicus ------MPTVDDVLEQVGEFGWFQKQAFLLLCLISASLAPIYVG 38 S. salar ------MGFADLLNDVGGFGRFQWIHVTLLSIPGLLMASQNLL 37 S. enterica ------

B. taurus -IVFLAFTPDHRCRSPGVAELSRRCGWSLAEELNYTVPGPGPE------SQCLRYEVDWN 91 C. elegans NIPFVVGNPPHTCHIP------EGKEYLRPLTNDTQILSCK------QYNET 100 D. melanogaster AGVFLLAKPDFRCALP------YENGSIYELSPHLWNLSYP------ENERC 76 H. sapiens -IVFLGFTPDHHCQSPGVAELSQRCGWSPAEELNYTVPGLGPAGEA-FLGQCRRYEVDWN 96 M. musculus -IVFLGFTPDHHCRSPGVAELSQRCGWSPAEELNYTVPGLGSAGEASFLSQCMKYEVDWN 97 R. norvegicus -IVFLGFTPGHYCQNPGVAELSQRCGWSQAEELNYTVPGLGPSDEASFLSQCMRYEVDWN 97 S. salar -NNFTAGMPGHHCTIPNRTSIASSQNISQSEVDDRELLRAFIPMDASGTKLSKCTRYVEA 96 S. enterica ------MEDNDH 6

B. taurus QSTLGCLDPLAS-LATNGSPLPLGPCEQGWVYDTP--GSSIVTEFNLVCDDSWKVDLFQS 148 C. elegans QINVFRAFTSAP-VDTYSDRISLVPCQNGWDYDNSTYLDSLVTEFNLVCDQQAWIEISTT 159 D. melanogaster SYYDVDYTEEYL-NGSIPRSSNETKTCSSYVYDRSKYLNSAVTEWNLVCSRSLLSATSDS 135 H. sapiens QSALSCVDPLAS-LATNRSHLPLGPCQDGWVYDTP--GSSIVTEFNLVCADSWKLDLFQS 153 M. musculus QSTLDCVDPLSS-LAANRSHLPLSPCEHGWVYDTP--GSSIVTEFNLVCGDAWKVDLFQS 154 R. norvegicus QSTLDCVDPLSS-LVANRSQLPLGPCEHGWVYDTP--GSSIVTEFNLVCGDAWKVDLFQS 154 S. salar QWHLLESNVSVIGHQANFSKLETEICLDGWTYDKTEFLSTVVSEWDLVCTLRPMKQMSQT 156 S. enterica IGARLDRLPLSRFHFRIFGIISFGLLLTGFLSYSG--NVVLAKLINNGWSNNYLNAAFTS 64

B. taurus CVNLGFFLGSLGVGYIADRFGRKVCLLATTLTCASLGVLTAVAPDYTSLLIFRLLQGLVS 208 C. elegans SFYVGSFIGNCLFGYVADKFGRRRSFFVILTVLIVCGTASSFAKDIESFIILRFFTGLAF 219 D. melanogaster LFMLGVLLGSFIFGQMSDKLGRKPTFFASLVLQLIFGVLAAVAPEYFSYTISRMIVGATT 195 H. sapiens CLNAGFLFGSLGVGYFADRFGRKLCLLGTVLVNAVSGVLMAFSPNYMSMLLFRLLQGLVS 213 M. musculus CVNLGFFLGSLVVGYIADRFGRKLCLLVTTLVTSLSGVLTAVAPDYTSMLLFRLLQGMVS 214 R. norvegicus CVNLGFFLGSLVVGYIADRFGRKLCLLVTTLVTSVSGVLTAVAPDYTSMLLFRLLQGMVS 214 S. salar IYMGGVLAGAIIFGGLSDRFGRKALLIWSYFQLATLGTCTAFSPSFMTYCIFRFMTGMAV 216 S. enterica ALMLGYFIGSLTGGFIGDYLGRRKAFRINLLLVGISATAAAFVPNMYWLIFFRCLMGTGM 124

B. taurus KGSWTAGYTLITEFVGLGYRRTVAILYQMAFTVGLVLLSGLAYILPHWRWLQLAVSLP-- 266 C. elegans PALFQIPFIICMEFMGNSGRIFSGLMTSLFFGAAMALLGVVAMFIRRWRQLTFFCNAP-- 277 D. melanogaster SGVFLVAYVIALEMVGSSYRLFAGVAMQMFFSVGFMLTAGFAYFIHDWRWLQIAITLP-- 253 H. sapiens KGNWMAGYTLITEFVGSGSRRTVAIMYQMAFTVGLVALTGLAYALPHWRWLQLAVSLP-- 271 M. musculus KGSWVSGYTLITEFVGSGYRRTTAILYQVAFTVGLVGLAGVAYAIPDWRWLQLAVSLP-- 272 R. norvegicus KGSWVSGYTLITEFVGSGYRRTTAILYQMAFTVGLVGLAGVAYAIPDWRWLQLAVSLP-- 272 S. salar SGVILNTVSLKVEWIPTKSRTLVGTLSSFFFTFGQMVLAGIAHSLRDWRKLQVAVCAP-- 274 S. enterica GALIMVGYASFTEFIPPIVRGKWSARLSFVGNWSPMLSAGIGVVVIAFLSWRMMFLLGGI 184

B. taurus --IFLLLFRFWFVPESPRWLLSQKRNTEAIKIMDHIAQKN------GKLPPADLKML 315 C. elegans --FAFYIIYYFFLPESPRWSVSVGKWADAKKQLKKIAKMN--GKSNVDVDELVDSMKNHQ 333 D. melanogaster --GLLFLCYYWIIPESARWLLMKGRKDEAFVIIEKAAKENKVEVPNEIYEQLVDEVAEKK 311 H. sapiens --TFLFLLYYWCVPESPRWLLSQKRNTEAIKIMDHIAQKN------GKLPPADLKML 320 M. musculus --TFLFLLYYWFVPESPRWLLSQKRTTQAVRIMEQIAQKN------RKVPPADLKMM 321 R. norvegicus --TFLFLLYYWFVPESPRWLLSQKRTTRAVRIMEQIAQKN------GKVPPADLKML 321 S. salar --FFLFFLYSWWYSESARWLVLNRRSDKALKHIHRVARIN---RKPEMVEKITLEVLECH 329 S. enterica GMLLAWSLSGKYFIESPRWLAGKGRQREAEQQLLMIESQIE------KEKSIVLPPHQE 237

B. taurus SLEEDVTEKLSPSFIDLFRTPNLRKYTFILMYLWFTSSVVYQG---LIMHVGATGGNLYL 372 C. elegans NAAEEKETKRSHNVTDLFKTPNLRRKTLIVTYIWVMNAIIYNG---LTLNVSNLPVDDYW 390 D. melanogaster KQDEMAASQPAATVFDLLRYPNLRRKTLLIFFDWFVNSGVYYG---LSWNTNNLGGNQLV 368 H. sapiens SLEEDVTEKLSPSFADLFRTPRLRKRTFILMYLWFTDSVLYQG---LILHMGATSGNLYL 377 M. musculus CLEEDASERRSPSFADLFRTPSLRKHTLILMYLWFSCAVLYQG---LIMHVGATGANLYL 378 R. norvegicus CLEEDASEKRSPSFADLFRTPNLRKHTVILMYLWFSCAVLYQG---LIMHVGATGANLYL 378 S. salar MHKEVQSSKTTHTAYDLIRTRVMRRISLCLMVVWFSTSFAYYG---LAMDLQKFGVNIYL 386 S. enterica SNKSMESRFENGTFWLLFKGQMLRRTLVAITVLIAMNISLYTITVWIPTIFVNSGIDVTK 297

Figure 1. Alignment of the organic cation transporter of different species. In orange the conserved amino-acid residues are highlighted.

The Pharmacogenomics Journal (2015), 473 – 487 © 2015 Macmillan Publishers Limited Drug pharmacogenetics and SLC22A1 polymorphisms C Arimany-Nardi et al 475 B. taurus DFLYSALVEFPA---GFIILVTIDRFGRRYPLATSNLAAGLACFLMIFIPHDLPWLNIMV 429 C. elegans SFIINGAVELPG---YFVVWPLLQCAGRRWTLAATMIVCGIGCVSAMFMPDGYPWLVASA 447 D. melanogaster NFMISGAVEIPG---YTLLLFTLNRWGRRSILCGTMMVAGISLLATIFVPSDMNWLIVAC 425 H. sapiens DFLYSALVEIPG---AFIALITIDRVGRIYPMAMSNLLAGAACLVMIFISPDLHWLNIII 434 M. musculus DFFYSSLVEFPA---AFIILVTIDRIGRIYPIAASNLVAGAACLLMIFIPHELHWLNVTL 435 R. norvegicus DFFYSSLVEFPA---AFIILVTIDRIGRIYPIAASNLVTGAACLLMIFIPHELHWLNVTL 435 S. salar IQIIFGLVDFPA---KLVALGSLTFLGRRITQGTCLLMSALMIFTNIFVPTDMQSIRTTL 443 S. enterica SIFMTAIIMIGAPVGIFIAALIIDHFPRRLFGSLLLIVIAVVGYFYSIQTEEWAILVYGL 357

B. taurus ACVGRMGITIVFQMVCLVNAELFPTFIRNLGMMVCSSLCDLGGVLTPFLVFRLMEVWQGS 489 C. elegans SFIGKFGVGSGFAVIYIFAGELYPTVVRAIGMGMSSMVAGSGLLLAPHIVN-LGKIVKIL 506 D. melanogaster AMIGKLAITSSYGTIYIFSAEQFPTVVRNVGLGASSMVARVGGILAPYLKL-LGEIWRPL 484 H. sapiens MCVGRMGITIAIQMICLVNAELYPTFVRNLGVMVCSSLCDIGGIITPFIVFRLREVWQAL 494 M. musculus ACLGRMGATIVLQMVCLVNAELYPTFIRNLGMMVCSALCDLGGIFTPFMVFRLMEVWQAL 495 R. norvegicus ACLGRMGATIVLQMVCLVNAELYPTFIRNLGMMVCSALCDLGGIFTPFMVFRLMEVWQAL 495 S. salar ACLGKAFTSASFTCIYLFTGELYPTVIRQTGMGFTSTMARVG-SMAAPAVLILEEMLPAL 502 S. enterica VMISFLYMYVCFASAVYVP-ELWPTHLRLRGSGFVNAVGRIVAVFTPYGVAVLLTRYGSV 416

B. taurus PLILFAALGLVAGGMTLLLPETKGVTLPETIEDAENLQR-KAKPKENKIYLQVQTSELNT 548 C. elegans PLLIMGLMALSAGILTFFLPETLGAPLPMTIEDAENFGK-KPEPDSGMFTQAAKKRESQP 565 D. melanogaster PLIICGALSLTAGLLSLLLPETLNKPMPETIEDGENFGK-KPAP-----QETAEEGGTQE 538 H. sapiens PLILFAVLGLLAAGVTLLLPETKGVALPETMKDAENLGR-KAKPKENTIYLKVQTSEP-- 551 M. musculus PLILFGVLGLSAGAVTLLLPETKGVALPETIEEAENLGRRKSKAKENTIYLQVQTGKS-- 553 R. norvegicus PLILFGVLGLTAGAMTLLLPETKGVALPETIEEAENLGRRKSKAKENTIYLQVQTGKS-- 553 S. salar PSMIYGGAAVVAGIIAFFLPETLNIPLPDTIEDVEEKWASKKLDAEKQAKKEAVALREMK 562 S. enterica TVFVVLGMLLAACALILFCFGIETRRVSLEALSSLR------452

B. taurus QAAERDASQGTAQQK----- 563 C. elegans LLEPHTPMDRRRRSSRLMNI 585 D. melanogaster LSGMLNGKSG------548 H. sapiens ------SGT------554 M. musculus ------PHT------556 R. norvegicus ------SST------556 S. salar KGAVEGDGEITGLNAL---- 578 S. enterica ------

Figure 1. Continued.

accepts toxins such as the carcinogens aﬂatoxin B1 and Table 1. Expression of hOCT1 determined by mRNA or protein 32 – ethidiumbromide as substrates. expression11,12,15,19,21,84 93

mRNA Protein Subcellular localization SNPS Bladder ✓ Urothelium The SLC22A1 gene is highly polymorphic in humans. In 2002, Kerb Brain ✓✓ Endothelial cells of microvessels et al.33 described for the first time 25 polymorphic variants in Eye ✓ cornea, iris–ciliary body and blood– Caucasians for this gene. Since then, over 1000 single-nucleotide retina barrier polymorphisms (SNPs) have been identified, 22 of which have ✓✓ Heart Cardiomyocytes been related to the treatment outcome when drugs transported Immune cells ✓✓ Intestine ✓✓ Lateral membranes of the by this protein are being used. Out of these, 21 are located at the enterocytes protein-coding region, causing all of them amino-acid substitu- Kindey ✓✓ tions, but one that results in an amino-acid deletion (M420del) Liver ✓✓ Sinusoidal membrane of the (Figure 2). The one left in this list is an intronic variant (rs622342). hepatocytes For a comprehensive overview of SLC22A1 gene heterogeneity, we Lung ✓✓ Luminal membrane of bronchial refer to Tables 2–8 in this review. epithelial cells SLC22A1 genetic heterogeneity is ethnic-specific. Caucasian, Ovary ✓ African and American (Puerto Ricans, Colombian and Mexican) ✓ Placenta populations present higher variability than Asians and Pacific Skeletal ✓ Islanders. Only one of the studied polymorphic variants (M408V) is muscle fi fi Skin ✓ present in all groups and is the only variant identi ed in Paci c Spleen ✓ Islanders. Three other variants (F160L, P341L and the Testis ✓ intronic rs622342) are present in the rest of the groups with allelic frequencies higher than 1%, whereas R61C and M420del Abbreviations: hOCT1, human organic cation transporter 1; mRNA, have been only identified in Caucasians, Africans and Americans. messenger RNA. Caucasian and American populations seem to share the specificity for SLC22A1 variants being G38D-, G401S- and G465R specific for these ethnic groups. The variants S14F, R342H, V461I, R488M and V519F are specific for Africans, all of them present at treatment of type-II diabetes and cancer; the antivirals lamivudine, an allelic frequency higher than 1%. Two of the variants (P117L acyclovir and ganciclovir; quinine, pentamidine and furamidine and P283L) are expressed exclusively in Asian population, used as antiparasitic drugs; and the antineoplastic drugs although their allelic frequency is lower than 1%. The other oxaliplatin, picoplatin and cis-diammine(pyridine)chloroplatin(II) studied variants (C88R, A99T, G174S, S189L and G220V) are – and bendamustine among others.10,22,29 31 Moreover, hOCT1 also expressed in less than 0.5% of the population.

Figure 2. hOCT1 model with the revised polymorphic variants highlighted.

Interestingly, two of these polymorphic variants (C88R and characterised yielding similar Km values to those described for the G465R) have been observed only in combination with the variant wild-type transporter, although Vmax was significantly reduced. A M420del. The haplotype combining C88R and M420del is clinical study in healthy volunteers showed that plasma glucose designed allele OCT1*6, whereas the haplotype with C88R and levels after metformin treatment were significantly higher in M420del is designed allele OCT1*5. Other alleles (OCT1*1–4) volunteers carrying hOCT1 polymorphisms than in those carrying contain only one polymorphic variant and will be referred in this only reference hOCT1 alleles. Consistent with the difference in the review by the SNP amino-acid change. plasma glucose levels after metformin treatment, insulin levels Many of SLC22A1 non-synonymous polymorphisms associated were significantly higher 2 h after glucose administration in with altered function are located in conserved gene regions individuals carrying loss-of-function variants compared with those among orthologues, thereby suggesting a key role of these carrying the wild type.35 A further study showed an increase in the residues on functionality. In fact, some of these polymorphic area under the concentration–time curve, higher maximal plasma variants are major contributors to variation in drug disposition, concentration (Cmax) and lower oral volume of distribution (V/F)in therapeutic and adverse drug responses and risk for human those individuals carrying reduced function alleles (R61C, G401S, disease. Moreover, the potential pharmacokinetic consequences M420del and G465R) compared with those carrying the wild-type of hOCT1 polymorphic variants vary depending on the admini- transporter.34 strated drug, making it difficult to predict the effect of a Tzvetkov et al.12 observed in healthy volunteers that homo- polymorphism on the transport of a specific compound without zygous carriers of R61C, G401S, M420del and G465R presented directly testing that compound. increased renal clearance of metformin. This increase could not be detected in another study where the variants were expressed heterozygously. However, homozygous carriers for the minor DRUGS AND PHARMACOGENOMICS allele in the intronic variant rs622342 showed significantly lower Drugs used in the treatment diabetes and polycystic ovary renal clearance of the drug.36 syndrome In a clinical study with patients with type-II diabetes mellitus, Metformin, a drug used for the treatment of type-II diabetes and only the variant rs622342 of all studied variant was associated cancer, was one of the first drugs described to be a hOCT1 with the glucose-lowering effect of metformin. In patients carrying substrate. It was also the first drug for which polymorphic variants this variant, the reduction of glycated haemoglobin (HbA1c) was of the transporter were studied for their effect on pharmaco- lower than in those expressing the wild-type protein.37 This kinetics and pharmacodynamics.34,35 Nowadays, it can be variant was also assessed in diabetic Indian population and was considered as a sort of paradigm in drug transporter pharmaco- found to be similarly associated with low response to metformin genetics. In the seminal studies by the Giacomini laboratory, seven treatment.38 The clinical impact of this variant is somehow of the polymorphic hOCT1 variants (S14F, R61C, S189L, G220V, controversial, because in the report by Christensen et al. this type G401S, M420del and G465R) showed impaired metformin uptake. of correlation was not found, in agreement with other studies In particular, S14F, S189L, G401S and M420del were kinetically focused on the impact of this variant on metformin action.39,40

The Pharmacogenomics Journal (2015), 473 – 487 © 2015 Macmillan Publishers Limited Drug pharmacogenetics and SLC22A1 polymorphisms C Arimany-Nardi et al 477 Nevertheless, this association could be identiﬁed with the variant G401S, but only when short-term treatments were evaluated.41 43 Patients expressing the variants R61C, G401S, M420del and patients (1531) G465R showed a metformin-induced HbA1c reduction similar to

Scottish diabetic that achieved in patients bearing the wild-type transporter protein.42,43

45 Polymorphic variants in hOCT1 have not only been related to the metformin effect in type-II diabetic patients but also to (150)

with PCOS secondary effects of the drug. The minor allele in the M408V Italian women variant and a 8-bp insertion (rs36056065) in the gene have been associated with the presence of common gastrointestinal side

44 effects of metformin therapy, but not with the effectiveness of the

(235) treatment. In the same study, SNPs R61C, M420del and G465R

T2D patients showed no effect either on metformin action or in its secondary 44 Latvian population effects.

74 In women with Polycystic Ovary Syndrome, metformin effects were also related to OCT1 variants. The studied population was (113) children

Caucasian stratiﬁed based on the number of carried polymorphisms. They

or the rest of the references within the text. presented the polymorphic variants, R61C, G401S, G465R and 31 e-II diabetes. Allelic frequencies as reported in the reviewed M420del. Metformin signiﬁcantly reduced total cholesterol or

(241) triglycerides in the reference group, but had no effect in subjects population Spanish CLL carrying variants, regardless of the number of polymorphisms. 63 However, the effect of metformin on glucose-stimulated insulin ﬂ with secretion was in uenced by the number of OCT1 polymorphisms. Spanish

CML (65) Thus, insulin area under the concentration–time curve was

98 similarly decreased in the reference group or in individuals expressing one polymorphism, but did not signiﬁcantly change in Italian patients carrying two or more polymorphisms.45 CML (60) In Asian population, the role of OCT1 polymorphic variants in

97 metformin handling has also been studied. The uptake of ﬁ

(156) metformin was signi cantly reduced in cells expressing Q97K, patients British CML P117L and R206C compared with the wild-type OCT1.46 Kinetic studies indicated that P117L and R206C had a reduced Vmax, 96 whereas Q97K showed an increased Km for the drug. Never-

(336) theless, the allelic frequency for these variants was so low that no CML patients

British-German clinical studies could be performed. In a clinical study in healthy

76 subjects, the variant P341L showed a trend towards higher area under the concentration–time curve and Cmax values compared (270) German with the reference OCT1, although the difference was not 95 statistically signiﬁcant.47 Two other variants were found in Asian − 4 (100)

German population to be either positively (M408V) or negatively ( 43T G in intron 1) correlated with metformin efﬁcacy. In fact, the hepatic 11 expression of OCT1 in individuals expressing the M408V variant (150) German tends to be lower than in those were the reference variant is 48 37

gene expressed. (102) Deutsch Drugs used in antiviral treatments 94 SLC22A1 Lamivudine, an antiretroviral drug, is also a hOCT1 substrate.30 (200) American European- Importantly, hOCT1 is known to be expressed and regulated in most immune system cells. Its expression in CD4+ cells is 33

ND ND 0.374particularly ND ND ND relevant 0.359 aslong ND as ND thisis ND a major ND HIV1 ND target ND being ND ND (57)

Caucasian this transporter protein a major determinant of drug accumulation within cells.49 Polymorphic variants C88R and G465R showed complete loss of function when assayed for lamivudin uptake. Variants R61C, S189L and M420del presented a signiﬁcantly Amino-acid change — decreased Vmax, although only S189L showed a signiﬁcantly

del decreased affinity for the drug. In all polymorphic variants studied 4 A R342H ND 0 ND 0 ND ND ND ND ND ND ND ND ND ND ND A G401S 0.032 0.011 ND 0.01 ND 0.024 ND 0.029 ND ND 0.029 0.044 ND 0.02 ND – A M440I ND 0 ND ND ND ND ND ND ND ND 0.01 ND ND ND ND A V461I ND 0 ND 0 ND ND ND ND ND ND ND ND ND ND ND A G465R ND 0.04 ND 0.043 ND ND 0.031 ND ND ND 0.017 0.027 0.04 0.01 ND T R488M ND 0 ND 0 ND ND 0.002 ND ND ND ND ND ND ND ND T V519F ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND – G M408V ND 0.598 0.397 0.429 ND ND 0.368 0.397 ND 0.559 0.598 ND 0.39 ND ND T P341L ND 0 ND 0.017 0.02 ND 0.02 ND ND ND 0.017 ND ND ND ND – fi fi A G38D ND ND ND 0.003 ND ND ND ND ND ND 0.015 ND ND ND ND A A99T ND ND 0.1 ND ND ND ND ND ND ND 0.002 ND ND ND ND A G174S ND ND ND ND ND ND ND ND ND ND 0.004 ND ND ND ND T G220V ND 0 ND 0 ND ND 0.002 ND ND ND ND ND ND ND ND T R61C 0.091 0.072 ND 0.097 ND 0.101 0.067 ND 0.074 0.147 0.054 0.058 0.1 0.103 0.067 T P117L ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND G F160L 0.22 0.065 ND 0.23 ND ND 0.217 ND 0.24 ND 0.209 ND ND ND ND T S189L ND 0.005 ND 0 ND ND ND ND ND ND 0.004 ND ND ND ND T P283L ND ND ND 0 0so ND far, transport ND ef NDcacy ND wassigni NDcantly ND altered ND (Arimany-Nardi ND ND ND C C88R 0.006 ND ND 0.003 ND ND ND ND ND ND 0.002 ND ND ND ND 4 4 4 4 4 4 4 4 4 4 4 T S14F ND 0 ND 0 ND ND 0 ND ND ND ND ND ND ND ND 4 4 4 4 4 4 4 4 4 4 4 4 C et al., in preparation). Moreover, when two common Asian variants

4 (P283L and P341L) were tested for lamivudine transportability, change both were shown to be less active for lamivudine uptake. In fact, kinetic studies revealed a signiﬁcantly reduced Vmax without changes in substrate afﬁnity.50 Published Caucasian allelic frequencies of the Drugs used in anticancer therapy Bendamustine is indeed an old drug but a very new one in the Exon Ref. number Nucleotide 1 rs34447885 41C 1 rs35888596 113G 1 rs12208357 181C 1 rs55918055 262T 1 295G 1 rs200684404 350C 2 rs683369 480C 3 520G 3 rs34104736 566C 3 rs36103319 659G 5 rs4646277 848C 6 rs2282143 1022C 6 rs34205214 1025G 7 rs34130495 1201G 7 rs35191146 1260G 7 rs34305973 1259T 7 rs202220802 1258A 7 rs628031 1222A 77 rs35167514 rs72552763 1257delATG 1260 GAT M420del 0.16 0.185 ND 0.17 ND 0.148 0.154 0.176 0.181 0.184 0.228 0.164 0.18 0.183 0.198 8 rs35956182 1320G 8 rs34295611 1381G 9 rs34059508 1393G 9 rs35270274 1463G 10 rs78899680 1555G Intronic rs622342 A literature. Numbers in brackets indicate the cohort of individuals genotyped in each study. The corresponding reference (superindex) is quoted as f Table 2. Abbreviations: CML, chronic myeloid leukaemia; CLL, chronic lymphocytic leukaemia; ND, not determined; PCOS, polycystic ovary syndrome; T2D,context typ of chronic lymphocytic leukaemia treatment. Bendamustine

© 2015 Macmillan Publishers Limited The Pharmacogenomics Journal (2015), 473 – 487 478 h hraoeoisJunl(05,473 (2015), Journal Pharmacogenomics The rgpamcgntc n L2A polymorphisms SLC22A1 and pharmacogenetics Drug

Table 3. Caucasian allelic frequencies found in databases

Exon Ref. number Nucleotide Amino-acid Americans with Finnish British in Iberian Toscani in European European- Utah residents with Toscani in change change European England and Spain Italy (662) American Northern and Western Italia (TSI) ancenstry Scotland European ancestry from the (204) CEPH collection (CEU) (226) – 8 05McilnPbihr Limited Publishers Macmillan 2015 © 487 1 rs34447885 41C4T S14F 0 0 0 0 0 ND 0 ND ND 1 rs35888596 113G4A G38D 0 0 0.006 0.04 0.01 0.007 0.002 ND ND 1 rs12208357 181C4T R61C 0.09 0.05 0.05 0.04 0.06 0.06 0.08 ND ND Arimany-Nardi C 1 rs55918055 262T4C C88R 0.006 0 0.006 0 0 0.002 0.004 ND ND 1 295G4A A99T ND ND ND ND ND ND ND ND ND 1 rs200684404 350C4T P117L 0 0 0 0 0 0 0 ND ND 2 rs683369 480C4G F160L 0.25 0.19 0.23 0.25 0.16 0.18 0.78 0.252 0.176 3 520G4A G174S ND ND ND ND ND ND ND ND ND 4

3 rs34104736 566C T S189L 0 0 0 0 0.005 0.002 0.001 ND ND al et 3 rs36103319 659G4T G220V 0 0 0 0 0 0.001 0 ND ND 5 rs4646277 848C4T P283L 0 0 0 0 0 ND ND ND ND 6 rs2282143 1022C4T P341L 0 0.005 0.02 0 0 0.015 0.014 0 0.005 6 rs34205214 1025G4A R342H 0 0 0 0 0.005 ND 0 ND ND 7 rs34130495 1201G4A G401S 0.02 0.02 0.02 0.07 0.03 0.02 0.03 ND ND 7 rs628031 1222A4G M408V 0.56 0.51 0.61 0.5 0.66 0.61 0.41 0.58 0.644 7 rs202220802 1258A4– 7 rs34305973 1259T4– 7 rs35191146 1260G4– 7 rs35167514 1257delATG M420del ND ND ND ND ND ND ND ND ND 7 rs72552763 1260 GAT4del 8 rs35956182 1320G4A M440I 0.006 0.03 0.006 0 0.01 0.005 0.01 ND ND 8 rs34295611 1381G4A V461I 0 0 0 0 0 ND 0 ND ND 9 rs34059508 1393G4A G465R 0.012 0.011 0.03 0 0.005 0.03 0.03 ND ND 9 rs35270274 1463G4T R488M 0 0 0 0 0.005 ND 0 ND ND 10 rs78899680 1555G4T V519F 0 0 0 0 0 ND 0 ND ND Intronic rs622342 A4C — 0.35 0.37 0.38 0.36 0.32 ND ND 0.405 0.323 Abbreviation: ND, not determined. Allelic frequencies of the SLC22A1 gene polymorphisms as available in databases are shown. In bold italic are data derived from the 1000 Genomes project, in bold roman are data from CsAgilent ClinSeq project, in underline italic are data derived from the NHLBI GO exome-sequencing project and in underline roman are Hapmap data. Numbers in brackets show the number of genotyped individuals, when available. Drug pharmacogenetics and SLC22A1 polymorphisms C Arimany-Nardi et al 479

Table 4. Published African allelic frequencies of the SLC22A1 gene

Exon Ref. number Nucleotide change Amino-acid change Africa-American (200)94 Xhosa population South Africa (148)99 Cape admixed (100)100

1 rs34447885 41C4T S14F 0.031 0.017 0.01 1 rs35888596 113G4A G38D 0 0 ND 1 rs12208357 181C4T R61C 0.01 ND 0 1 rs55918055 262T4C C88R ND ND 0 1 295G4A A99T ND ND ND 1 rs200684404 350C4T P117L ND ND ND 2 rs683369 480C4G F160L 0.005 ND ND 3 520G4A G174S ND ND ND 3 rs34104736 566C4T S189L 0 0 0.01 3 rs36103319 659G4T G220V 0.005 0 0.01 5 rs4646277 848C4T P283L ND 0 0 6 rs2282143 1022C4T P341L 0.082 0.084 0.005 6 rs34205214 1025G4A R342H 0.031 ND ND 7 rs34130495 1201G4A G401S 0.007 0 0 7 rs628031 1222A4G M408V 0.735 ND ND 7 rs202220802 1258A4– 7 rs34305973 1259T4– 7 rs35191146 1260G4– 7 rs35167514 1257delATG M420del 0.029 ND ND 7 rs72552763 1260 GAT4del 8 rs35956182 1320G4A M440I 0.005 0 0.005 8 rs34295611 1381G4A V461I 0.01 ND ND 9 rs34059508 1393G4A G465R 0 0 0.005 9 rs35270274 1463G4T R488M 0.05 ND ND 10 rs78899680 1555G4T V519F ND 0.03 0.015 Intronic rs622342 A4C — ND 0.216 0.18 Abbreviation: ND, not determined. Allelic frequencies as reported in the reviewed literature. Numbers in brackets indicate individuals genotyped in each study. The corresponding reference (superindex) is quoted as references within the text.

Table 5. African allelic frequencies found in databases

Exon Ref. number Nucleotide Amino- Americans of Luhya Yoruba African African Luhya in Yoruba in change acid African population population American ancestry in Webuye, Ibadan, change ancestry (2) (2) Southwest Kenya Nigeria USA (ASW) (LWK) (216) (YRI) (288) (114)

1 rs34447885 41C4T S14F 0.008 0.03 0.02 0.02 ND ND ND 1 rs35888596 113G4A G38D 0 0.005 0 0 ND ND ND 1 rs12208357 181C4T R61C 0.02 0 0 0.014 ND ND ND 1 rs55918055 262T4C C88R 0 0 0 0 ND ND ND 1 295G4A A99T ND ND ND ND ND ND ND 1 rs200684404 350C4T P117L 0 0 0 0 ND ND ND 2 rs683369 480C4G F160L 0.04 0 0 0.95 0.035 0.028 0.017 3 520G4A G174S ND ND ND ND ND ND ND 3 rs34104736 566C4T S189L 0 0 0 0 ND ND ND 3 rs36103319 659G4T G220V 0.008 0 0 0 ND ND ND 5 rs4646277 848C4T P283L 0 0 0 ND ND ND ND 6 rs2282143 1022C4T P341L 0.04 0.08 0.09 0.07 0.053 0.077 0.082 6 rs34205214 1025G4A R342H 0 0.02 0.02 0.02 ND ND ND 7 rs34130495 1201G4A G401S 0.008 0 0 0.005 ND ND ND 7 rs628031 1222A4G M408V 0.73 0.75 0.76 0.27 0.711 0.75 0.741 7 rs202220802 1258A4– 7 rs34305973 1259T4– 7 rs35191146 1260G4– 7 rs35167514 1257delATG M420del ND ND ND ND ND ND ND 7 rs72552763 1260 GAT4del 8 rs35956182 1320G4A M440I 0 0 0 0.002 ND ND ND 8 rs34295611 1381G4A V461I 0.008 0.02 0.03 0.009 ND ND ND 9 rs34059508 1393G4A G465R 0 0 0 0.003 ND ND ND 9 rs35270274 1463G4T R488M 0.02 0.05 0.06 0.03 ND ND ND 10 rs78899680 1555G4T V519F 0.008 0.02 0.05 0.02 ND ND ND Intronic rs622342 A4C — 0.21 0.22 0.15 ND 0.196 0.2 0.148 Abbreviation: ND, not determined. Allelic frequencies of the SLC22A1 gene polymorphisms as available in databases are shown. In bold italic are data derived from the 1000 Genomes project, in underline italic are data derived from the NHLBI GO exome-sequencing project and in underline roman are Hapmap data. Numbers in brackets show the number of genotyped individuals, when available.

© 2015 Macmillan Publishers Limited The Pharmacogenomics Journal (2015), 473 – 487 480 h hraoeoisJunl(05,473 (2015), Journal Pharmacogenomics The rgpamcgntc n L2A polymorphisms SLC22A1 and pharmacogenetics Drug

Table 6. Published Asian allelic frequencies of the SLC22A1 gene

Exon Ref. number Nucleotide Amino- Asian- Japanese Japanese Japanese Japanese with Malay Chinese Chinese Chinese-Malay- Korean Korean Vietnamese India Indian change acid American (116)101 (194)80 with diabetes primary billiar (70)66 (100)95 (70)66 Indian with CML (150)95 (96)47 (100)95 Tamilian (70)66 change (60)94 type II (66)46 cirrhosis (275)80 (38)66 (112)102 – 8 05McilnPbihr Limited Publishers Macmillan 2015 © 487 1 rs34447885 41C4T S14F 0 0 ND ND ND ND ND ND ND ND ND ND ND ND 1 rs35888596 113G4A G38D 0 0 ND ND ND ND ND ND ND ND ND ND ND ND 1 rs12208357 181C4T R61C 0 ND ND ND ND ND ND ND ND ND ND ND ND ND Arimany-Nardi C 1 rs55918055 262T4C C88R ND ND ND ND ND ND ND ND ND ND 0 ND ND ND 1 295G4A A99T ND ND ND ND ND ND ND ND ND ND ND ND ND ND 1 rs200684404 350C4T P117L ND ND ND 0.023 ND ND ND ND ND ND 0 ND ND ND 2 rs683369 480C4G F160L 0.017 0.086 0.16 ND 0.1 0.2 ND 0.21 ND ND ND ND ND 0.21 3 520G4A G174S ND ND ND ND ND ND ND ND ND ND ND ND ND ND

3 rs34104736 566C4T S189L 0 0 ND 0.008 ND ND ND ND ND ND 0 ND ND ND al et 3 rs36103319 659G4T G220V 0 0 ND ND ND ND ND ND ND ND ND ND ND ND 5 rs4646277 848C4T P283L ND ND ND ND ND ND 0.005 ND ND 0.013 ND 0 ND ND 6 rs2282143 1022C4T P341L 0.117 0.168 0.16 ND 0.17 0.09 0.11 0.12 ND 0.167 0.135 0.055 0.089 0.11 6 rs34205214 1025G4A R342H 0 ND ND ND ND ND ND ND ND ND ND ND ND ND 7 rs34130495 1201G4A G401S 0 0 ND ND ND ND ND ND ND ND ND ND ND ND 7 rs628031 1222A4G M408V 0.762 0.81 ND 0.85 ND 0.69 ND 0.73 0.78 ND ND ND 0.803 0.58 7 rs202220802 1258A4– 7 rs34305973 1259T4– 7 rs35191146 1260G4– 7 rs35167514 1257delATG M420del 0 0 ND ND ND ND ND ND ND ND ND ND ND ND 7 rs72552763 1260 GAT4del 8 rs35956182 1320G4A M440I 0 0 ND ND ND ND ND ND ND ND ND ND ND ND 8 rs34295611 1381G4A V461I 0 0 ND ND ND ND ND ND ND ND ND ND ND ND 9 rs34059508 1393G4A G465R 0 0 ND ND ND ND ND ND ND ND ND ND ND ND 9 rs35270274 1463G4T R488M 0 0 ND ND ND ND ND ND ND ND ND ND ND ND 10 rs78899680 1555G4T V519F ND ND ND ND ND ND ND ND ND ND ND ND ND ND Intronic rs622342 A4C — ND ND 0.18 ND 0.15 0.2 ND 0.15 ND ND ND ND 0.245 0.37 Abbreviations: CML, chronic myeloid leukaemia; ND, not determined. Allelic frequencies as reported in the reviewed literature. Numbers in brackets indicate the cohort of individuals genotyped in each study. The corresponding reference (superindex) is quoted as for the rest of references within the text. Drug pharmacogenetics and SLC22A1 polymorphisms C Arimany-Nardi et al 481

Table 7. Asian allelic frequencies found in databases

Exon Ref. number Nucleotide Amino- Han- Southern Japanese Han-Chinese Chinese in Gujarati Indians Japanese change acid Chinese Han- in Beijing, Metropolitan in Houston, in Tokyo, change Bejing Chinese China (CHB) Denver, Colorado Texas (GIH) Japan (JPT) (270) (CHD) (218) (202) (224)

1 rs34447885 41C4T S14F 0 0 0 ND ND ND ND 1 rs35888596 113G4A G38D 0 0 0 ND ND ND ND 1 rs12208357 181C4T R61C 0 0 0 ND ND ND ND 1 rs55918055 262T4C C88R 0 0 0 ND ND ND ND 1 295G4A A99T ND ND ND ND ND ND ND 1 rs200684404 350C4T P117L 0.005 0 0 ND ND ND ND 2 rs683369 480C4G F160L 0.11 0.18 0.12 0.163 0.138 0.153 0.129 3 520G4A G174S ND ND ND ND ND ND ND 3 rs34104736 566C4T S189L 0 0 0 ND ND ND ND 3 rs36103319 659G4T G220V 0 0 0 ND ND ND ND 5 rs4646277 848C4T P283L 0.005 0.005 0.011 ND ND ND ND 6 rs2282143 1022C4T P341L 0.14 0.11 0.16 0.139 0.161 0.094 0.146 6 rs34205214 1025G4A R342H 0 0 0 ND ND ND ND 7 rs34130495 1201G4A G401S 0 0 0 ND ND ND ND 7 rs628031 1222A4G M408V 0.78 0.7 0.81 0.746 0.736 0.663 0.805 7 rs202220802 1258A4– 7 rs34305973 1259T4– 7 rs35191146 1260G4– 7 rs35167514 1257delATG M420del ND ND ND ND ND ND ND 7 rs72552763 1260 GAT4del 8 rs35956182 1320G4A M440I 0 0 0 ND ND ND ND 8 rs34295611 1381G4A V461I 0 0 0 ND ND ND ND 9 rs34059508 1393G4A G465R 0 0 0 ND ND ND ND 9 rs35270274 1463G4T R488M 0 0 0 ND ND ND ND 10 rs78899680 1555G4T V519F 0 0 0 ND ND ND ND Intronic rs622342 A4C — 0.12 0.15 0.19 0.157 0.144 0.194 0.179 Abbreviation: ND, not determined. Allelic frequencies of the SLC22A1 gene polymorphisms as available in databases are shown. In bold italic are data derived from the 1000 Genomes project and in underline italic are Hapmap data. Numbers in brackets show the number of genotyped individuals, when available.

Table 8. American allelic frequencies

Exon Ref. number Nucleotide change Amino-acid change Mexican-American (20)94 Colombian Mexican Puerto Ricans

1 rs34447885 41C4T S14F 0 0 0.008 0 1 rs35888596 113G4A G38D 0.056 0.008 0 0.02 1 rs12208357 181C4T R61C 0 0.03 0.05 0.009 1 rs55918055 262T4C C88R ND 0 0 0 1 295G4A A99T ND ND ND ND 1 rs200684404 350C4T P117L ND 0 0 0 2 rs683369 480C4G F160L 0.05 0.15 0.07 0.12 3 520G4A G174S ND ND ND ND 3 rs34104736 566C4T S189L 0 0 0 0.009 3 rs36103319 659G4T G220V 0 0 0 0 5 rs4646277 848C4T P283L 0 0 0 6 rs2282143 1022C4T P341L 0 0.03 0.02 0.03 6 rs34205214 1025G4A R342H 0 0 0 0 7 rs34130495 1201G4A G401S 0 0.08 0 0 7 rs628031 1222A4G M408V 0.786 0.69 0.86 0.75 7 rs202220802 1258A4– 7 rs34305973 1259T4– 7 rs35191146 1260G4– 7 rs35167514 1257delATG M420del 0.214 ND ND ND 7 rs72552763 1260 GAT4del 8 rs35956182 1320G4A M440I 0 0 0 0.09 8 rs34295611 1381G4A V461I 0 0 0 0 9 rs34059508 1393G4A G465R 0 0.008 0.03 0.02 9 rs35270274 1463G4T R488M 0 0 0 0 10 rs78899680 1555G4T V519F ND 0.008 0 0 Intronic rs622342 A4C — ND 0.47 0.37 0.35 Abbreviation: ND, not determined. Allelic frequencies of the SLC22A1 gene polymorphisms as reported in the reviewed literature and available in databases are shown. In bold italic are data from literature and in underline italic are data derived from the 1000 Genomes project.

© 2015 Macmillan Publishers Limited The Pharmacogenomics Journal (2015), 473 – 487 Drug pharmacogenetics and SLC22A1 polymorphisms C Arimany-Nardi et al 482 has recently been described to be a hOCT1 substrate, being overall response to the treatment in two independent studies,62,63 bendamustine-induced cytotoxicity dependent on hOCT1 function. whereas a recent clinical study in newly diagnosed CML patients Two of the studied polymorphic variants (S189L and G465R) were showed that this polymorphic variant could modulate the not associated with hOCT1-mediated cytotoxicity, whereas R61C outcome during imatinib treatment, showing significant correla- and M420del variants showed a decreased hOCT1-induced tion with a higher risk of treatment failure.71 cytotoxicity when compared with the wild-type transporter. In addition, the SNPS combination, R61C, L160F, P341L and When these loss-of-function variants were evaluated in primary M420del were shown to be significantly associated with imatinib cells from chronic lymphocytic leukaemia patients, it was observed response.70 that a subset of chronic lymphocytic leukaemia cases showed a Although imatinib does not seem to be translocated by significant correlation between those cells expressing disfunctional hOCT1,55 the presence of the transporter as well as some of its variants of the transporter and bendamustine ex vivo cytotoxicity.31 polymorphic variants have widely reported to affect outcome. Sorafenib is a novel tyrosine kinase inhibitor that has been Thus, it is tempting to speculate that hOCT1 expression is either reported to be beneficial in the treatment of hepatocellular linked to the expression of other transporters and/or metabolic carcinoma and cholangiocarcinoma. A recent study identified proteins relevant to imatinib efficacy, or hOCT1 polymorphic hOCT1 as a sorafenib transporter, although previous studies ruled variants show genetic linkage to other genes that can indeed out the possibility of hOCT1 mediating sorafenib uptake.51,52 Two contribute to imatinib therapeutic response. novel variants, hOCT1-R61S and hOCT1-C88A, which were fi identi ed either in hepatocellular carcinoma, cholangiocarcinoma Other drugs or both but not in the adjacent healthy tissue, showed impaired sorafenib transportability that may affect the ability of this drug to O-desmethyltramadol, the active form of tramadol, is internalised reach active intracellular concentrations in tumours retaining through hOCT1. Polymorphic variants R61C, G401S, M420del and hOCT1 expression.51 transporters bearing those combinations of amino-acid substitu- Interestingly, Zebularine, an epigenetic drug that is being tions more commonly found as polymorphisms in Caucasian developed for the treatment of the myelodysplastic syndrome, population (M420del/C88R and M420del/G465R) alleles OCT1*5 has been shown to be extruded by the wild-type form of hOCT1. and *6 showed impaired O-desmethyltramadol uptake compared Accordingly, when the polymorphic variants C88R, M408V, with the wild-type transporter. Clinical studies demonstrated that M420del and G465R were tested for their efflux capacity, drug those individuals carrying loss-of-function variants presented efflux was significantly lower than that observed in the wild-type higher plasma concentrations of the drug associated with high form in all cases.53 Although this would require further clinical concentrations in the central nervous system and increased μ 72 assessment, it is tempting to speculate that, in case hOCT1 is stimulation of the -opioid receptor. fi behaving as an efflux instead of an influx drug transporter, loss-of- Morphine, is a high-af nity hOCT1 substrate. However, the function variants would be associated with increased drug polymorphic variant G401S as well as the haplotypes OCT1*5 and sensitivity. OCT1*6 were not able to transport morphine. R61C and M420del Imatinib, is a tyrosine kinase inhibitor currently used in the showed low morphine transport capacity (Vmax) compared with fi treatment of chronic myeloid leukaemia (CML). Although some the wild-type protein without signi cant changes in substrate in vitro studies indicated that imatinib is a weak or even is not a affinity, despite a tendency to increased Km reported for the hOCT1 substrate,54,55 some studies have reported that hOCT1 M420del variant. In a clinical study involving healthy volunteers, all expression and its associated activity may be the most important these hOCT1 variants were considered to be non-functional. determinants for imatinib treatment outcome in CML patients.56–58 Individuals carrying a loss-of-function allele had significantly The role of the transporter polymorphic variants on the pharma- higher plasma concentrations of morphine than those found in cokinetics of this drug and its clinical response is also controversial. the non-carriers. The maximal plasma concentration showed also Kim et al.59 determined that the polymorphic variant L160F was a strong trend to increase along with the number of loss-of- 73 responsible for a higher rate of loss of response to imatinib in a function hOCT1 alleles. In a parallel clinical study in children, cohort of 229 patients. However, a further study in a smaller cohort mean individual morphine clearence in subjects homozygous for of 33 patients showed no correlation between this polymorphic defective hOCT1 alleles was significantly lower than clearence 74 variant and response to treatment.60 All studies involving the R61C found in carriers of the active OCT1 allele. variant showed no correlation between its occurrence in patients The antihypertensive drug debrisoquine has also been and response to imatinib.61–64 Otherwise, patients carrying the shown to be translocated by hOCT1. The analysis of the funcional G401S variant showed high probability of achieving major impact of its polymorphic variants on drug uptake showed that molecular response to the drug.64 Takahashi et al.65 determined the R61C, G401S and M420del variants presented a decreased that Japanese patients, carrying the polymorphic variant M408V, Vmax compared with the wild type, whereas the Km value was showed significantly major molecular response compared with not significantly affected. The haplotypes OCT1*5 and OCT1*6 those carrying the wild type. In addition, the haplotype implicating resulted in a complete loss of function for debriosquine this polymorphic variant with the two intronic variants rs3798168 transport.75 and rs80301372 was shown by others to be significantly associated The antiemetic drug topisetron, is also a hOCT1 substrate. This with diminished imatinib clearance.66 Another study in Caucasians drug is primarily metabolised by the hepatic enzyme CYP2D6. could not demonstrate any significant correlation between this Thus, in order to be metabolised, it has to be internalised into variant and overall inadequate response to imatinib.63 Another hepatocytes by hOCT1. In this context, transporter polymorphic polymorphic variant with high allelic frequency in Asians, P341L, has variants have been postulated to alter drug response. A recent been repeatedly reported not to be associated with altered imatinib study by Tzvetkov et al. showed an increase in drug plasma pharmacokinetics and drug response.67–69 However, this was not concentrations and response to treatment in patients carrying the case in a study with mixed Caucasian, Asian and African reduced functioning hOCT1 variants (R61C, G401S, M420del and patients, in which an association between this polymorphic variant the haplotypes OCT1*5 and OCT1*6) independently of CYP2D6 and cytogenic response to imatinib was observed.70 genetic variability. Although ondansetron inhibited the uptake of This apparent controversy involving hOCT1 polymorphic 1-methyl-4-phenylpyridinium mediated by hOCT1, direct uptake variants and imatinib, has also been found when addressing the of the drug could not be measured. Nevertheless, similar to what pharmacogenetic impact of the Caucasian and American high- was observed with topisetron, ondansetron plasma concentra- allelic-frequent variant M420del. No correlation was observed with tions were higher in patient lacking any fully active hOCT1 allele

Table 9. SLC22A1 SNPs role in drug functional effects and its clinical impact

Drugs SNPs Functional effects Clinical Impact Lack of clinical impact

35 Diabetes Metformin S14F ↓Vmax; = Km 36 R61C ↑AUC; ↑Cmax; No altered CLrenal ↓V/F, = HbA1c34,42,43 Homozygous ↑CLrenal12 No effect44 35 34 S189L ↓Vmax; = Km ↑AUC; ↑Cmax; ↓V/F 35 36 G401S ↓Vmax; = Km ↑AUC; ↑Cmax; ↓V/F; No altered CLrenal ↑HbA1ca34,41–43 Homozygous ↑CLrenal12 No effect44 35 36 M420del ↓Vmax; = Km ↑AUC; ↑Cmax; No altered CLrenal ↓V/F, = HbA1c34,42,43 Homozygous ↑CLrenal12 G465R Homozygous No altered CLrenal36 ↑CLrenal, = HbA1c12,42,43 No effect44 rs622342 ↑HbA1c37 No effect39,40 Homozygous ↑CLrenal36 ↓Metformin response37 46 Q97K ↑Km; = Vmax 46 P117L ↓Vmax; = Km 46 R206C ↓Vmax; = Km 47 P341L ↑AUC; ↑Cmax M408V Positive marker metformin efficiency48 -43T4G intron 1 Negative marker metformin efficiency48 PCOS Metformin45 R61C/G401S/ ↓Metformin efficiency M420del/G465R (regardless number of variants) ↑AUC (in patients carrying two or more polymorphisms) 50 Antiviral Lamivudine R61C ↓Vmax; = Km C88R Complete loss of function S189L ↓Vmax; ↑Km M420del ↓Vmax; = Km G465R Complete loss of function P283L ↓Vmax; = Km P341L ↓Vmax; = Km Anticancer Bendamustine31 R61C ↓ Cytotoxicity S189L ↓↓Cytotoxicity M420del ↓ Cytotoxicity G465R ↓↓Cytotoxicity Sorafenib51 R61S ↓ Transportability C88A ↓ Transportability Zebularine56 C88R ↓ Efflux M408V ↓ Efflux M420del ↓ Efflux G465R ↓ Efflux Imatinib L160F ↓Imatinib efficiency59 No effect60 R61C No correlation61–64 G401S ↑MMR64 M408V ↑MMR65 No effect63 M408V+rs3798168 ↓Imatinib CL66 +rs80301372 P341L ↓Cytogenic response70 No effect67–69 M420del ↑Risk treatment failure71 No effect62,63 R61C/L160F/ ↓Imatinib efficiency70 P341L/M420del Others O-desmethyltramadol72 R61C ↓ Uptake ↑Plasma concentrations G401S ↓ Uptake ↑Plasma concentrations M420del ↓ Uptake ↑Plasma concentrations OCT1*5 ↓ Uptake ↑Plasma concentrations OCT1*6 ↓ Uptake ↑Plasma concentrations 73,74 Morphine R61C ↓ Vmax ↑Plasma concentrations Homozygous ↓CL G401S No uptake ↑Plasma concentrations Homozygous ↓CL M420del ↓ Vmax ↑Plasma concentrations Homozygous ↓CL

Table 9. (Continued)

Drugs SNPs Functional effects Clinical Impact Lack of clinical impact

OCT1*5 No uptake ↑Plasma concentrations Homozygous ↓CL OCT1*6 No uptake ↑Plasma concentrations Homozygous ↓CL 75 Debrisoquine R61C ↓Vmax; = Km G401S ↓Vmax; = Km M420del ↓Vmax; = Km OCT1*5 No uptake OCT1*6 No uptake Topisetron76 R61C ↑Plasma concentrations G401S ↑Plasma concentrations M420del ↑Plasma concentrations OCT1*5 ↑Plasma concentrations OCT1*6 ↑Plasma concentrations Ondansetron76 R61C ↑Plasma concentrations G401S ↑Plasma concentrations M420del ↑Plasma concentrations OCT1*5 ↑Plasma concentrations OCT1*6 ↑Plasma concentrations Anti-parkinson drugs77 rs622342 ↑Prescribed doses of Anti- Parkinson drugs ↓Survival after treatment with levodopa Amisulpride and R61C ↓ Uptake sulpride78 G401S ↓ Uptake M420del ↓ Uptake OCT1*5 ↓ Uptake OCT1*6 ↓ Uptake Vitamin B128 R61C ↓ Uptake S189L ↓ Uptake G220V ↓ Uptake G401S ↓ Uptake M420del ↓ Uptake G465R ↓ Uptake The colour code reﬂects the population in which the polymorphic variant is more present; african (purple); caucasian and american (green); asian (yellow); rare variant (pink). All cited variants in the table present a reduced tranport activity. aonly in short-term treatments.

and decreased with the increasing number of fully active hOCT1 M420del, as well as the haplotypes OCT1*5 and OCT1*6 resulted alleles. When drug effect was assessed by number of vomiting in decreased drug uptake.78 episodes in the first 24 h after chemotherapy, those patients Vitamine B1 has recently been identified as a hOCT1 substrate. carrying lacking-of-function alleles vomited less frequently than In vitro studies showed that hOCT1 polymorphic variants R61C, those with one or two functional alleles in agreement with the S189L, G220V, G401S, M420del and G465R affected vitamine B1 observed increased plasma concentration. The effect was uptake.28 significant in the subgroup of patients that received topisetron, but not in the subgroup treated with ondansetron although a DRUG–DRUG INTERACTIONS similar trend could be observed. When the group of patients treated with ondansetron was enlarged, the significance for this Polymorphic hOCT1 proteins have shown to be more susceptible drug could be also determined.76 to inhibition by drugs compared with a hOCT1 reference. For Finally, the minor C allele at rs622342, the intronic variant that metformin, variants R61C and M420del, were more sensitive to was previously shown to be associated with metformin response, drug inhibition, with half maximal inhibitory concentration up to was also associated with higher prescribed doses of anti- 23 times lower than those of OCT1 reference. The concomitant Parkinsonian drugs and shorter survival after levodopa therapy. administered drugs, verapamil and amitriptyline, revealed poten- The exact drug or drugs that contributed to the difference in tial drug–drug interactions at clinical plasma concentrations prescribed daily dose of anti-Parkinsonian drugs could not exactly of metformin for M420del.79 A study with lamivudine and be identified. The only drug for which the prescribed daily doses co-administered antivirals showed that, at Cmax concentrations, were significantly associated with rs622342 was amantadine. abacavir, zidovudine, efavirenz and raltegravir could significantly However, it could not be excluded that the polymorphic variant inhibit lamivudine uptake mediated by M420del. Abacavir was was associated with lower response to levodopa, and that this had also able to inhibit the uptake mediated by R61C, whereas it to be compensated by prescribing higher doses of amantadine.77 increased significantly the uptake of lamivudine when raltegravir The psychotropic drugs amisulpride and sulpride have recently was co-administered. Efavirenz, otherwise, did not affect lamivu- been reported to be hOCT1 substrates also. In vitro studies dine uptake mediated by this polymorphic variant when showed that the hOCT1 polymorphic variants R61C, G401S and compared with the OCT1-reference transporter. Moreover, the

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