Gender Differences in the Renal Excretion of Organic Cations in Rats: Correlation with Kidney Expression of OCT-2 and MATE-2K

Typically, clinical pharmacokinetic studies are performed in young healthy male volunteers. As a result, a gender-specific analysis is usually not included in the evaluation of results. Several publications have reported gender differences (GDs) in pharmacokinetics (PK) and pharmacodynamics (PD) of drugs. In addition, based on the analysis of FDA's Adverse Event Reporting System (AERS) data and other resources, it is evident that women encounter adverse drug reactions more than men [1, 2]. With an increase in awareness of GDs in drug response and a trend towards development of personalized medicine, it has become critical to identify factors that contribute to these GDs. FDA's Office of Women's Health (OWH) was established by a congressional mandate to advocate for the participation of women in clinical trials and the analyses of data by sex. OWH supports studies to track the participation of women in clinical trials and collaborates with FDA centers and external partners to understand sex and gender differences in disease prevalence and response to therapy through science and research [3]. Today, understanding the role of PK and PD in GDs in pharmacological response is an active area of research. Given the fact that gender pharmacology and human clinical trials are quite complex, investigators are using various non-clinical models to understand specific differences that may impact the PK and PD of a given drug.;Drug elimination from the body is one of the major determinants of a drug's pharmacokinetic profile. The kidney is primarily responsible for the excretion of drugs and it's metabolites from the body. Differences in renal excretion of drugs between males and females therefore may impact the safety and efficacy of drugs and can have important clinical implications.;The isolated perfused rat kidney (IPK) is a well-established ex-vivo model and has been used to study numerous aspects of drug disposition. The IPK model was used in this research to evaluate the effect of gender on the renal elimination of the organic cations, trimethoprim (TMP) and metformin (MET). Both TMP and MET are primarily excreted unchanged in the urine. The organic cation transporters (OCTs) and multi-drug and toxin extrusion transporters (MATEs) have been implicated in the renal transport of TMP and MET. From the OCT and MATE family of transporters, the transporters OCT-2 and MATE-2K are primarily expressed in the kidneys and play a critical role in the renal secretion of several cationic drugs. Several drug-drug interactions (DDIs) involving both TMP and MET have been documented. GDs in the expression of these transporters may impact the PK of the cationic drugs that are substrates of these transporters and are primarily excreted in the urine.;The fundamental objective of this research was to evaluate the effect of gender on the renal disposition of the TMP and MET in the IPK model and to correlate the findings with the expression of OCT-2 and MATE-2K transporters in the male and female rat kidneys.;A gender difference in the renal excretion was observed for both TMP and MET in the IPK. For TMP, the renal clearance was more than two-fold higher in the IPK experiments using kidneys from female donors. This difference can be primarily attributed to the higher tubular secretion observed in females. A significant difference in the kidney weight-corrected GFR was found among control groups, and was higher in the female kidneys compared to males. This correlates with the significantly higher filtration clearance noted in females in the TMP treated group. However, the difference in secretion clearance between the groups was much higher compared to the difference in the filtration clearance (nearly two fold higher secretion clearance in females compared to males vs. 1.4 folds higher filtration clearance in females). The study design also included a castrated group to elucidate the effect of the sex hormone testosterone on the renal transport of TMP. No significant differences were observed in any of the parameters between the male and the castrated groups suggesting that testosterone has minimal effect on TMP's urinary disposition. Similar to the male group, the renal excretion parameters (renal clearance, filtration clearance, secretion clearance and XR) were significantly lower in the castrated group as compared to the female group, which confirms the findings of the male group in an indirect manner. An XR ratio greater than 1 was observed in all study groups suggesting a mechanism of net secretion, and it was nearly two fold higher in the female groups. The disappearance of TMP from the perfusate was not found to be significantly different among the three groups. It is worth noting however that the average initial concentration in the perfusate in the female experiments was higher (although not significantly higher), and despite the higher initial concentration, the amount of TMP remaining in the perfusate was the lowest in the female group. The lack of significant difference in the perfusate disposition of TMP suggests a greater role of transport systems on the luminal membrane contributing to the observed GDs in the renal clearance of TMP.;Based on the previously reported higher expression levels of OCT2 in male rats [25] and assuming a role of OCT2 in the renal transport of TMP, the findings above were surprising. Thus, the data suggests a possible involvement of multiple transport systems at both the basolateral and luminal membrane and further studies are needed to identify the source of these GDs in TMP excretion. For a very widely used antibiotic, especially among women, these differences could have clinically significant consequences.;For MET, a well-documented OCT-2 substrate, the renal clearance was nearly three-fold higher in males as compared to females. Nearly the entire amount of drug dosed in the male groups was recovered in the urine by the end of the study duration. The excretion ratio was also three-fold greater in the male study groups. This finding correlates with the previously reported greater expression levels of OCT-2 in males compared to the females. Upon comparing the amount of MET remaining in the perfusate between the male and female groups, no significant differences were observed in the initial perfusate concentrations between the two groups. However, nearly all of the MET was cleared from the perfusate by the end of the study duration in the male IPK groups but approximately 23% of the initially dosed amount remained in the perfusate from the female IPK groups.;The observed differences in the drug excretion in the IPK were attributed to differences in the expression levels of transporters in males and females. To establish the correlation of the IPK results with the expression of transporters, protein levels for OCT-2 and MATE-2K (cation transporting transporters found primarily in the kidney) transporters were measured using the western blot technique. Analysis of the western blot data revealed that the expression of both OCT-2 (present on the basolateral membrane) and MATE-2K (present on the luminal membrane) was significantly greater in males as compared to females. The expression of OCT-2 was approximately 20% greater in male rat kidneys, and the expression of MATE-2K was nearly three-fold higher in the male rat kidneys. Although the expression levels of OCT2 are not as high as reported previously (1.2 fold greater in the current study vs. nearly two fold higher in a previous study), the findings of this investigation still correlate with the previous finding of higher levels of OCT2 proteins in males compared to females. The previous study evaluated the expression levels at both the mRNA level and the protein level. The differences in the current findings and the previously reported data could be due to several variables such as test system (kidney tissue homogenate vs. kidney slices and basolateral membrane vesicles), methodology and instrumentation and investigator.;MATE' s have been recently identified as proton/cation anti-porters mediating the final step in the excretion of various cationic drugs. It transports a wide range of cationic substrates, and certain drugs such as lamivudine known to be substrates of the OCTs, are also substrates of the MATE transporters [120]. In such cases, the combined effect of the higher expression levels of both transporters can lead to a significantly higher renal clearance of the particular drug in males and inversely, a significantly lower clearance (and therefore accumulation) in females. This could be a potential reason for the higher ADRs and/or DDIs in females and may require a different dosing regimen of both the victim and the perpetrator drug to avoid toxic effects in the female population.;The higher expression levels of the MATE-2K transporter in the males also presents an anomaly for the TMP clearance, found to be greater in the females while confirming that it could be a potential factor that causes the GDs in the renal excretion of MET. MATE-2K was chosen as the transporter of interest for this particular study, as it is primarily expressed in the kidneys and therefore assumed to play a greater role in the excretion of cationic compounds as compared to the other MATE transporters. MATE1 is also reported to be present in the luminal membrane of the kidney and could potentially be one the transporter responsible for the GDs observed in the TMP excretion [120]. Additional studies to further characterize the renal excretion of TMP should be performed to determine the underlying cause of the GDs in TMP excretion.;Taken together, these results suggest that gender-related differences exist in the renal excretion of drugs and the expression of OCT-2 and MATE-2K transporter expression levels is males and females. This could be a factor that contributes to the differences observed in males and females in the renal drug excretion and thus have an impact on the overall PK profile of drugs in the female population.