Although most drugs are considered safe and compatible with breastfeeding, cases of toxic drug exposure have been reported. Active efflux transporters have been implicated as a mechanism in the transfer of drugs from mother to baby via breastmilk. Using breastmilk as a source of human mammary epithelial cells, this novel longitudinal study investigated the expression of four active transporters, namely, MDR1, MRP1, MRP2 and BCRP in the lactating human breast. BCRP gene was found to be strongly overexpressed with levels peaking at 5 months postpartum, potentially indicating a time where a breastfed infant may be at risk of inadvertent exposure to BCRP substrates. Serum albumin, a major component of human breastmilk was increasingly downregulated as lactation progresses. Xanthine oxidase/dehydrogenase, an enzyme in breastmilk attributed to a reduced risk of gastroenteritis caused by Escherichia coli and Salmonella enteritides, was downregulated. Lysozyme and fatty acid synthase are progressively upregulated. This study also shows that breastmilk-derived epithelial cells, when propagated in culture, exhibit characteristics significantly different to those derived directly from breastmilk. This serves to warn that in vitro studies are not a true representation of in vivo processes in the lactating breast; hence, application of in vitro data should be conducted with caution.

Assessing the interactions of a new drug candidate with transporters, either as a substrate, inhibitor, or inducer, is no simple matter. There are many clinically relevant transporters, as many as nine to be evaluated for an FDA submission and up to 11 for the EMA as of 2020. Additionally, it is likely that if a compound is a substrate or inhibitor of one transporter, it will be so for other transporters as well. There are practically no specific substrates or inhibitors, presumably because the specificities of drug transporters are so broad and overlapping, and even fewer clinically relevant probes that can be used to evaluate transporter function in humans. In the case of some transporters, it is advisable to evaluate an NCE with more than one test system and/or more than one probe substrate in order to convince oneself (and regulatory authorities) that a clinical drug interaction study is not warranted. Finally, each test system has its own unique set of advantages and disadvantages. One has to appreciate the nuances of the available tools (test systems, probe substrates, etc.) to select the most relevant tools for the study and design the optimal in vitro experiment. In this chapter, several examples are used to illustrate the successful interpretation of in vitro data for both efflux and uptake transporters. Some data presented in this chapter are unpublished at the time of the compilation of this book. It has been included in this chapter to provide a sense of the complexities in transporter kinetics to the reader.

Platinum-based doublets are the standard chemotherapy for lung cancer. The identification of markers associated with drug toxicity may improve the success of the treatment. Single nucleotide polymorphisms (SNPs) mapping into the genes involved in platinum transport or detoxification may explain the occurrence of toxicities. In this study, we evaluated the role of three SNPs in predicting the onset of adverse events for lung cancer patients receiving cisplatin or carboplatin in adjuvant, neo-adjuvant and metastatic settings.
Eighty-two patients affected by non-small-cell and small-cell lung cancer treated with cisplatin- or carboplatin-based chemotherapy (stage II-IV) were enrolled. Before genetic analysis, patients signed a written informed consent. DNA was extracted from peripheral blood samples and genotypes were determined by real-time PCR. We selected and analyzed three SNPs: ABCB1 c.3435C>T/rs1045642, ABCC2 -24C>T/rs717620 and GSTP1 c.313A>G/rs1695. Patient characteristics and genotypes were correlated with hematological, gastrointestinal and renal toxicity as recorded by Common Terminology Criteria for Adverse Event (CTCAE) v4.03. No neurological toxicity was observed in our patients.
Variant alleles were present in 53% of patients for ABCB1 c.3435C >T, 18.3% for ABCC2 -24C> T, and 34.8% for GSTP1 c.313A>G. Heterozygous CT at ABCB1 c.3435 was associated to a lower risk of hematological toxicity compared to homozygous CC (OR = 0.20; 95% CI 0.05, 0.69; p = 0.01). Similar results were observed by genetic dominant model (CT + TT vs CC) and hematological toxicity (OR = 0.26; 95% CI 0.09, 0.79; p = 0.02). No other significant associations were found between toxicity and SNPs. Multivariate analysis confirmed an independent value for the ABCB1 c.3435 C >T polymorphism.
The present study reveals that ABCB1 c.3435C>T polymorphism influences platinum toxicity. The T allele seems to exert a protective effect on the development of toxicities. Further studies, such as epigenetic regulation ones, are needed to validate and shed more light on this association.

Glucuronidation is a major process of drug metabolism and elimination that generally governs drug efficacy and toxicity. Publications have demonstrated that efflux transporters control intracellular glucuronidation metabolism. However, it is still unclear whether and how efflux transporters interact with UDP-glucuronosyltransferases (UGTs) in subcellular organelles. In this study, kaempferol, a model fluorescent flavonoid, was used to investigate the interplay of glucuronidation with transport at the subcellular level. Human recombinant UGTs and microsomes were utilized to characterize the in vitro glucuronidation kinetics of kaempferol. The inhibition of UGTs and efflux transporters on the subcellular disposition of kaempferol were determined visually and quantitatively in Caco-2/TC7 cells. The knockout of transporters on the subcellular accumulation of kaempferol in liver and intestine were evaluated visually. ROS and Nrf2 were assayed to evaluate the pharmacological activities of kaempferol. The results showed that UGT1A9 is the primary enzyme responsible for kaempferol glucuronidation. Visual and quantitative data showed that the UGT1A9 inhibitor carvacrol caused a significant rise in subcellular aglycone and reduction in subcellular glucuronides of kaempferol. The inhibition and knockout of transporters, such as P-glycoprotein (P-gp), breast cancer resistance protein (BCRP), and multidrug resistance-associated proteins (MRPs), exhibited a marked increase in subcellular kaempferol and decrease in its subcellular glucuronides. Correspondingly, inhibition of UGT1A9 and transporters led to increased kaempferol and, consequently, a significantly enhanced ROS scavenging efficiency and nuclear translocation of Nrf2. In conclusion, the interplay of efflux transporters (P-gp, BCRP, and MRPs) and UGTs govern the subcellular exposure and corresponding pharmacological activity of kaempferol.

Associations between polymorphisms of UDP-glucuronosyltransferases (UGTs) or efflux transporters (e.g., P-glycoprotein and MRP2) and different types of cancer have been described, whereas the role of influx transporters (e.g. OATP1B1 and OATP2B1) has been seldom explored. The GenColon study investigated potential associations between variant alleles of UGTs, efflux and influx transporters and CRC.
Three hundred CRC cases were matched with 300 controls for age, sex and enrolment site. Fifteen SNPs in UGT1A6-9, UGT2B7, ABCB1, ABCC2, SLCO1B1 and SLCO2B1 genes were characterized using Taqman® PCR. Using multivariate conditional logistic regression, we investigated the relationships between CRC and \"environmental\" risk factors (physical activity, housing and working areas, consumption of red meat, tobacco, alcohol); genetic polymorphisms, in the study population and in the subgroups with \"environmental\" risk factors.
No significant association was observed for the analyzed SNPs (or haplotypes). However, an increased CRC risk was found in carriers of the UGT1A8 rs1042597-G variant allele (additive risk OR = 3.39[1.29-8.89], p = 0.02951) in the subgroup of meat-consumers (n = 84), and in carriers of the ABCB1 rs1045642-T (exon26) variant allele (additive risk; OR = 1.89[1.10-3.39], p = 0.0257) in the \"never alcohol consumption subgroup\" (n = 125). In addition, as previously reported, the following CRC risk factors were identified: absence of physical activity (OR = 6.35[3.70-10.9], p < 0.0001), living or working in rural or mix area (OR = 2.50[1.48-4.23], p = 0.0006 and OR = 2.99[1.63-5.48], p = 0.004, respectively) and tobacco exposure >30 years (3.37[1.63-6.96], p = 0.0010).
Variant genotypes of influx transporters (OATP1B1 and 2B1) were not associated with CRC. This study confirmed the influence of lifestyle factors, but not the previously reported detrimental effect of SNPs in intestinal UGTs or efflux transporters, except for a UGT1A8 variant in subjects consuming meat and the exon 26 SNP of ABCB1 in the never alcohol consumption subgroup.
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