ObjectiveBrain infection with Theiler's murine encephalomyelitis virus (TMEV) in C57BL/6J mice produces an etiologically relevant model of acquired seizures. Dietary changes can modify seizure presentation following TMEV brain infection and influence intestinal microbiome diversity and composition. Intestinal dysbiosis may thus similarly affect seizure burden and antiseizure medicine (ASM) activity in this model, independent of pharmacokinetic effects. We thus sought to define the influence of antibiotic (ABX)-induced gut dysbiosis on acute seizure presentation, anticonvulsant activity of carbamazepine (CBZ), and CBZ pharmacokinetics with TMEV infection.MethodsMale C57BL/6J mice (4-5 weeks old) received oral ABX or saline (SAL) once daily beginning on arrival through day 7 after TMEV infection (postinfection [p.i.]). Mice were infected with TMEV or phosphate-buffered saline on day 0. Mice received intraperitoneal (20 mg/kg) CBZ or vehicle (VEH) twice daily on days 3-7 p.i. and were assessed for handling-induced seizures 30 min after treatment. Plasma was collected on day 7 p.i. at 15 and 60 min after CBZ administration for bioanalysis.ResultsTMEV infection induced acute seizures, but ABX-induced gut dysbiosis altered seizure presentation. There were 75% SAL-VEH, 35% SAL-CBZ, 35% ABX-VEH, and 72% ABX-CBZ mice with seizures during the 7-day monitoring period. There was a significant pretreatment x ASM interaction (p = .0001), with differences in seizure burden in SAL- versus ABX-pretreated mice (p = .004). CBZ significantly increased latency to seizure presentation, an effect absent in ABX-CBZ mice. Plasma CBZ concentrations did not differ between SAL and ABX pretreatment groups, suggesting that ABX did not influence CBZ pharmacokinetics.SignificanceABX-induced gut dysbiosis markedly altered acute disease trajectory with TMEV-induced encephalitis, reflecting a novel contribution of the gut microbiome to seizure presentation. ABX-induced gut dysbiosis also significantly changed acute seizure control by CBZ, but did not influence plasma CBZ concentrations. The gut-brain axis is thus an underrecognized contributor to TMEV infection-induced seizures, ASM activity, and disease burden.

Continued growth in global sales of natural products has led to an increased risk of natural product-drug interactions that can compromise drug efficacy and safety. One such natural product, goldenseal, was shown to decrease systemic exposure to a subtherapeutic dose of oral metformin in healthy adults. A follow-up study involving therapeutic metformin doses and adults with type II diabetes demonstrated a metformin dose-dependent pharmacokinetic interaction with goldenseal. These results, along with no change in metformin half-life or renal clearance in both studies, suggested that the goldenseal-metformin interaction occurred in the gut via inhibition of an unidentified saturable intestinal transport process. We used enteroid monolayers derived from the duodenum of 4 healthy human adult donors to recapitulate the goldenseal-metformin interaction in vitro and identify the transporters involved in the observed in vivo interaction. Our results implicate thiamine transporter (ThTr) 2 as the predominant transporter involved in metformin uptake through the apical membrane, accounting for approximately 45% of total metformin intracellular accumulation. Additionally, goldenseal inhibited ThTr-2, but only under subsaturating metformin dosing concentrations. The goldenseal-metformin interaction mediated under therapeutic metformin dose conditions involves a low-affinity basolateral transporter, ThTr-1, which accounts for approximately 50% of inhibitable metformin apical to basolateral flux. However, a substantial fraction of metformin flux appears to involve paracellular transport. These results further elucidate the mechanism underlying the goldenseal-metformin interaction and suggest that enteroid monolayers are a promising model to study intestinal natural product-drug interactions. SIGNIFICANCE STATEMENT: The research presented in this article demonstrates the utility of enteroid monolayers to predict and ascertain the mechanisms of drug-drug and natural product-drug interactions. Using this model, the study was able to identify the transporters (thiamine transporter-1 and thiamine transporter-2) involved in metformin absorption that are inhibited by the natural product, goldenseal, which were previously unidentified.

CYP2A6 is the hepatic enzyme responsible for the metabolic inactivation of nicotine. Variation in CYP2A6 alters nicotine clearance, affecting numerous smoking behaviors and tobacco-related diseases, making investigating sources of variation important. A published molecular study of microRNA-126-5p, the microRNA-126 functional arm, showed it decreased CYP2A6 expression post-transcriptionally; it also showed that higher CYP2A7 mRNA competed for microRNA-126-5p binding, mitigating the CYP2A6 reduction. To extend these observations, we investigated relationships between microRNA-126-5p and CYP2A6 protein and activity using a large human liver bank (n = 282). MicroRNA-126-5p was not inversely correlated with CYP2A6 protein (rs = 0.04, p > 0.05), nor was it significant in an unadjusted regression model (p > 0.05) or in an adjusted model (with genotype, age, and sex) (p > 0.05). Although CYP2A7 mRNA was positively correlated with CYP2A6 protein (rs = 0.48, p < 0.001), adding CYP2A7 mRNA to the adjusted model did not alter the relationship between microRNA-126-5p and CYP2A6 protein (p > 0.05), nor did CYP2A7 mRNA interact with microRNA-126-5p on CYP2A6 (p > 0.05). Similar results were found in modeling CYP2A6 activity. MicroRNA-21 was used as a positive control (inversely correlated with CYP2A6 protein, rs = -0.33, p < 0.001) and microRNA-152 as a negative control (not correlated with CYP2A6 protein, rs = -0.06, p > 0.05). These data do not support a role for microRNA-126-5p in downregulating CYP2A6 protein or activity, or for CYP2A7 mRNA in playing a decoy role, even when other predictors (genotype, age, and sex) were included in the model.

We evaluated the impact of spaceflight on a microphysiologic model of calcium oxalate (CaOx) kidney stone disease. Proximal tubule epithelial cells cultured as confluent microtubules were exposed to CaOx crystals with or without potassium citrate (a potential countermeasure) to determine the impact on gene expression. Nine genes were differentially expressed in response to CaOx crystal exposure during spaceflight. This project presents the use of microgravity as a unique environment to study kidney pathophysiology.

Altered drug pharmacokinetics during inflammation or infection have been linked to elevated plasma concentrations of proinflammatory cytokines. Data on how these cytokines affect the expression and activity of intestinal drug transporters and, therefore, bioavailability of transported drugs, remain limited. Here, we used a novel human enteroid in vitro model to investigate the effects of key proinflammatory cytokines (ie, interleukin [IL]-1β, IL-6, tumor necrosis factor-α, and interferon-gamma) on the mRNA expression of major intestinal transporters and activity of intestinal breast cancer resistance protein (BCRP) and P-glycoprotein (P-gp). Differentiated enteroid monolayers (in 96-well plates) were treated for 48 hours with each cytokine individually or in combination (cocktail) at 0.1, 1, or 10 ng/mL, encompassing their pathophysiological plasma concentrations in various inflammatory conditions. In a concentration-dependent manner, the cytokine cocktail significantly reduced the mRNA expression of BCRP, P-gp, multidrug resistance proteins 2/3, organic solute transporter α/β, serotonin transporter, and organic anion transporter polypeptide 2B1, while increasing multidrug resistance protein4 mRNA expression. Among individual cytokines, IL-1β elicited the most pronounced effects. To quantify the effect of cytokines on mRNA expression and activity of BCRP and P-gp, these treatments, at 1 ng/mL of individual cytokines or the cocktail, were repeated in the Transwell format. The efflux ratio of nitrofurantoin (a selective BCRP substrate), after exposure to 1 ng/mL of each cytokine or the cytokine cocktail for 48 hours, was significantly reduced, whereas the efflux ratio of digoxin (a P-gp substrate) remained unchanged. SIGNIFICANCE STATEMENT: Proinflammatory cytokines significantly downregulate major intestinal drug transporter expression and breast cancer resistance protein activity in human enteroid monolayers, highlighting the potential impact of inflammation on oral drug bioavailability. These results can be used to populate physiologically-based pharmacokinetic models to predict transporter-mediated drug absorption under inflammatory conditions, guiding safer and more effective dosing regimens.

Accelerating development of complex in vitro models (CIVMs) drives a need for additional methods to characterize these systems for use in toxicology and drug development. Relative to traditional cell culture, CIVMs can use a lower number of cells, are cultured for longer periods of time, and typically involve engineered 3-D cell organization. Standard single-cell assessment tools are not conducive to these types of complex models due to cell isolation stress, cell loss, and high cost. In this report, we benchmark RNA-seq deconvolution, which utilizes publicly available scRNA-seq datasets to predict cell proportions from bulk RNA-seq data derived from two CIVMs: a stem-cell-based human intestinal organoid CIVM and a neonatal rodent testis CIVM. We consider the impact of multiple imputation methods for scRNA-seq to restore the gene distribution of the original tissue and benchmark multiple deconvolution methods. The accuracy of deconvolution methods varied significantly in our analyses but provided valuable information on the emergence of an enterocyte cell population from the LGR5+ crypt stem cells following differentiation in the intestinal organoid CIVM. In the testis CIVM, deconvolution indicated that a small population of germ cells were retained over time, peritubular myoid cells proliferated over time, and that Leydig cell estimates remained stable with physiologically relevant hormone stimulation. In our analysis, using imputed single-cell references improved deconvolution accuracy. Deconvolution can be a useful tool for novel CIVM characterization, especially with rapidly growing libraries of single-cell data across tissues and developmental time.

The application of human enteroid systems presents a significant opportunity within the drug development pipeline, highlighting considerable potential for advancements in the characterization and evaluation of new molecular entities. Derived from LGR5+ crypt-based columnar cells, enteroid systems more accurately recapitulate the microanatomy and physiological processes of the human intestinal mucosa compared to traditionally used systems. They contain the complement of major mucosal epithelial cell types, maintain the genetic identity of the donor and intestinal segment they were derived from, and exhibit biological functions and specific activities that are seen in vivo. In this review, we examine the applications of human enteroid systems in nonclinical drug development and compare findings to existing and emerging in vitro models of the small intestine. Specifically, we explore enteroid systems in the context of predicting oral drug disposition, focusing on apparent permeability, intestinal first-pass metabolism, and drug interactions, as well as their utility in assessing drug-induced gastrointestinal toxicity and screening therapeutic efficacy against enteric diseases. Additionally, we highlight aspects of enteroid systems that warrant further study.

ABSTRACT Pharmacokinetic drug interactions can lead to unexpected changes in plasma concentrations of the object drug, potentially increasing the risk for adverse effects and/or decreasing therapeutic efficacy. The botanical product goldenseal was previously shown to decrease metformin systemic exposure in healthy adults. This three‐arm, open‐label, crossover clinical study assessed the pharmacokinetic goldenseal–metformin interaction in adults with type 2 diabetes stabilized on therapeutic doses of metformin (500–2550 mg daily). The aggregate pharmacokinetic data indicated no clinically meaningful interaction as determined by the metformin area under the plasma concentration‐time curve (AUC) geometric mean ratio [90% confidence interval] of 0.93 [0.86–1.01] laying within the predefined no‐effect range (0.80–1.25). However, metformin AUC decreased by ~20%, 14%, and 0% after goldenseal coadministration at low (500–750 mg), moderate (1000–1500 mg), and high (2000–2550 mg) metformin doses, respectively; renal clearance and half‐life remained unchanged throughout. The exploratory pharmacodynamic endpoint, HbA1c, decreased on average from 6.8% to 6.5%, regardless of the effects of goldenseal on metformin pharmacokinetics. The decreasing effect of goldenseal on metformin systemic exposure with increasing metformin dose, coupled with no changes in renal excretion and elimination half‐life, indicated that both the pharmacokinetic goldenseal–metformin interaction and the nonlinear absorption of metformin are governed by saturable, intestinal transport mechanism(s). The disconnect between changes in metformin systemic exposure and therapeutic effects emphasizes the need to evaluate clinical biomarkers to comprehensively assess drug interaction risks, particularly those involving natural products. Healthcare providers may consider cautioning patients about supplementing metformin pharmacotherapy with goldenseal to avoid risks for undesired changes in glycemic control. Trial Registration: ClinicalTrials.gov identifier: NCT05081583

This study evaluated the underlying mechanistic links between genetic variability in vitamin K metabolic pathway genes (CYP4F2 and CYP4F11) and phylloquinone hydroxylation activity using genotype- and haplotype-based approaches. Specifically, we characterized genetic variability in the CYP4F2/CYP4F11 locus and compared common single allele genotypes and common haplotypes as predictors of hepatic gene expression, enzyme abundance, and phylloquinone (VK1) ω-hydroxylation kinetics. We measured CYP4F2 and CYP4F11 mRNA levels, CYP4F2 and CYP4F11 protein abundances, and the VK1 concentration-dependent ω-hydroxylation rate in matched human liver nucleic acid and microsome samples, utilizing a novel in vitro population modeling approach. Results indicate that accounting for the CYP4F2*3 allele alone is sufficient to capture most of the genetic-derived variability in the observed phenotypes. Additionally, our findings highlight the important contribution that CYP4F11 makes toward vitamin K metabolism in the human liver.

The microgravity environment aboard the International Space Station (ISS) provides a unique stressor that can help understand underlying cellular and molecular drivers of pathological changes observed in astronauts with the ultimate goals of developing strategies to enable long- term spaceflight and better treatment of diseases on Earth. We used this unique environment to evaluate the effects of microgravity on kidney proximal tubule epithelial cell (PTEC) response to serum exposure and vitamin D biotransformation capacity. To test if microgravity alters the pathologic response of the proximal tubule to serum exposure, we treated PTECs cultured in a microphysiological system (PT-MPS) with human serum and measured biomarkers of toxicity and inflammation (KIM-1 and IL-6) and conducted global transcriptomics via RNAseq on cells undergoing flight (microgravity) and respective controls (ground). Given the profound bone loss observed in microgravity and PTECs produce the active form of vitamin D, we treated 3D cultured PTECs with 25(OH)D3 (vitamin D) and monitored vitamin D metabolite formation, conducted global transcriptomics via RNAseq, and evaluated transcript expression of CYP27B1, CYP24A1, or CYP3A5 in PTECs undergoing flight (microgravity) and respective ground controls. We demonstrated that microgravity neither altered PTEC metabolism of vitamin D nor did it induce a unique response of PTECs to human serum, suggesting that these fundamental biochemical pathways in the kidney proximal tubule are not significantly altered by short-term exposure to microgravity. Given the prospect of extended spaceflight, more study is needed to determine if these responses are consistent with extended (>6 months) exposure to microgravity.

To further the development of an in vitro model that faithfully recapitulates drug disposition of orally administered drugs, we investigated the utility of human enteroid monolayers to simultaneously assess intestinal drug absorption and first-pass metabolism processes. We cultured human enteroid monolayers from 3 donors, derived via biopsies containing duodenal stem cells that were propagated and then differentiated atop permeable Transwell inserts, and confirmed transformation into a largely enterocyte population via RNA sequencing analysis and immunocytochemistry (ICC) assays. Proper cell morphology was assessed and confirmed via bright field microscopy and ICC imaging of tight junction proteins and other apically and basolaterally localized proteins. Enteroid monolayer barrier integrity was demonstrated by elevated transepithelial electrical resistance that stabilized after 10 days in culture and persisted for 42 days. These results were corroborated by low paracellular transport probe permeability at 7 and 21 days in culture. The activity of a prominent drug metabolizing enzyme, CYP3A, was confirmed at 7, 21, and 42 days culture under basal, 1α,25(OH)2 vitamin D3-induced, and 6',7'-dihydroxybergamottin-inhibited conditions. The duration of these experiments is particularly noteworthy, because, to our knowledge, this is the first study to assess drug metabolizing enzymes and transporters expression/function for enteroids cultured for greater than 12 days. The sum of these results suggests enteroid monolayers are a promising ex vivo model to investigate and quantitatively predict an orally administered drug's intestinal absorption and/or metabolism. SIGNIFICANCE STATEMENT: This study presents a novel ex vivo model of the human intestine, human intestinal organoid (enteroid) monolayers that maintain barrier function and metabolic functionality for up to 42 days in culture. The incorporation of both barrier integrity and metabolic function over an extended period within the same model is an advancement over historically used in vitro systems, which either lack one or both of these attributes or have limited viability.

Abstract ID 96848Poster Board 203Rationale: Brain infection with Theiler’s murine encephalomyelitis virus (TMEV) in C57BL/6J mice models infection-induced acute seizures and epileptogenesis. Diet manipulation can modify the presentation of acute seizures in TMEV-infected mice (Zierath et al, BioRxiV 2023); however, no study has yet assessed whether antibiotic-induced intestinal dysbiosis influences the activity of antiseizure medicines (ASM) in the TMEV model. Whether the gut microbiome influences the phenotype of symptomatic seizures after TMEV infection is also unclear. We thus sought to define the extent to which antibiotic administration influences acute seizure presentation, the activity of ASMs, and the pharmacokinetic profile of ASMs in this mouse seizure model.Methods: Male C57BL/6J mice (4-5 weeks-old) received a broad-spectrum antibiotic cocktail (ABX) containing ampicillin, metronidazole, neomycin sulfate, and vancomycin (n=55) or vehicle (n=60) by oral gavage once daily beginning at arrival (Day -2) to Day 7 post-TMEV infection. Mice were infected with either intracerebral TMEV or PBS on Day 0. Mice received carbamazepine (CBZ; 20 mg/kg, i.p.) or vehicle (0.5% MC) twice daily Days 3-7 p.i. and were assessed for handling-induced seizures 30 min after CBZ treatment. Plasma samples were collected on Day 7 p.i. at 15 and 60 min post-CBZ treatment to quantify the extent to which ABX-induced gut dysbiosis influences ASM pharmacokinetics.Results: TMEV infection induced acute symptomatic seizures, regardless of pretreatment and ASM history as 18/25 (72%) ABX-CBZ mice, 7/20 (35%) ABX-VEH mice, 7/20 (35%) SAL-CBZ mice, and 15/20 (75%) SAL-VEH mice presented with seizures during the 7-day monitoring period. Average seizure burden was: 12.5 in ABX-CBZ, 4.7 in ABX-VEH, 5.7 in SAL-CBZ, and 16.1 in SAL-VEH mice. There was a significant pretreatment x ASM interaction (F (1, 81) = 16.0, p=0.0001), with post-hoc tests revealing marked differences in seizure burden in SAL- versus ABX-pretreated mice (p=0.004). Further, the latency to Stage 5 seizure was substantially increased by CBZ during days 3-7 post-infection; an effect absent in ABX-treated mice similarly administered CBZ. In TMEV-infected mice, spleens were 0.32% of body weight in ABX-CBZ mice, 0.34% in ABX-VEH mice, 0.36% in SAL-CBZ mice, and 0.38% in SAL-VEH mice. In sham-infected mice, spleens were 0.45% of body weight in ABX-CBZ mice, 0.65% in SAL-CBZ mice, and 0.36% in SAL-VEH mice. Plasma CBZ concentrations in TMEV-infected mice receiving ABX were 9.8±2.0 mg/mL 15 min post-dosing and 4.0±0.5 at 60 min post-dose, consistent with published CBZ plasma concentrations in mice (Bialer et al, 2004) suggesting no pharmacokinetic differences because of ABX history.Conclusions: Gut dysbiosis markedly alters the presentation of symptomatic seizures and acute disease burden in the TMEV mouse model, reflecting a novel therapeutic target for seizure control. The gut-brain axis is an understudied target in epilepsy that may benefit from greater investigation.Acknowledgements: This work was supported by the University of Washington Department of Pharmacy and the University of Washington Plein Center for Geriatric Pharmacy.

Abstract ID 95893Poster Board 034Background: The widely used botanical dietary supplement goldenseal is an established clinical inhibitor of cytochrome P450 (CYP) 3A activity, as evidenced by a 40-50% increase in the area under the plasma concentration vs. time curve (AUC) of the CYP3A probe drug midazolam after oral administration. Our recently observed lack of change in midazolam half-life further suggested that the interaction occurs primarily in the intestine. We next developed and verified a physiologically based pharmacokinetic (PBPK) model of the goldenseal-midazolam interaction to elucidate the mechanism of CYP3A inhibition (reversible vs. time-dependent) and anatomical site of the interaction (intestine vs. liver). Simulations indicated that the primary mechanism involves time-dependent inhibition of CYP3A in the intestine. Clinical verification that the intestine is the primary site of the interaction would enhance the robustness of this PBPK model. The objective of this study was to compare the pharmacokinetics of intravenously administered midazolam (to bypass first-pass metabolism) between the absence and presence of goldenseal.Methods: Adult volunteers (n = 22) participated in this 3-arm crossover study. Midazolam (0.5 mg) was administered intravenously alone (baseline), after a single oral dose (3 g) of goldenseal (acute), and after self-administration of goldenseal (1 g 3x daily) for 27 days (chronic). Blood (5 mL) was collected before and from 0.083-24 hours post-midazolam administration. Plasma was harvested and analyzed for midazolam and the primary metabolite, 1’-hydroxymidazolam, using a validated LC/MS/MS method. The pharmacokinetics of each analyte were determined via noncompartmental analysis methods using Phoenix WinNonlin™ (v8.1).Results: The geometric mean plasma concentration vs. time profile for both midazolam and 1’-hydroxymidazolam after intravenous administration of midazolam were essentially superimposable, regardless of goldenseal exposure (Figure 1). The geometric mean ratio (GMR) [90% confidence interval] of midazolam AUC in the presence to absence of goldenseal was 1.05 [0.87-1.23] and 0.97 [0.81-1.13] after acute and chronic goldenseal, respectively. Corresponding values for half-life were 0.98 [0.84-1.12] and 1.02 [0.88-1.16], respectively and for 1'-hydroxymidazolam AUC were 1.03 [0.91-1.15] and 1.04 [0.70-1.38], respectively. The GMR of the 1’-hydroxymidazolam/midazolam AUC ratio was 1.16 [0.92-1.40] and 1.09 [0.85-1.33] after acute and chronic goldenseal, respectively.Conclusions: The lack of change in the pharmacokinetics of midazolam and 1’-hydroxymidazolam between baseline and goldenseal exposure, whether goldenseal was administered acutely or chronically for 27 consecutive days, confirmed that the goldenseal-midazolam interaction occurs primarily in the intestine. These novel observations further verify the PBPK model developed for this natural product-drug combination. This more robust model could be applied to other oral CYP3A substrates that may be co-consumed with goldenseal (e.g., calcium channel blockers, HCV and HIV protease inhibitors, statins), as well as various specific populations (e.g., hepatically impaired, renally impaired, pediatrics, pregnancy), to better predict interaction risks and manage pharmacotherapeutic regimens.Supported by the National Institutes of Health (U54 AT008909).

Abstract ID 92080Poster Board 032In vivo studies suggest that inflammation or infections can markedly alter drug clearance, thereby affecting drug effectiveness and safety (Morgan E. T. et al., 2009). This alteration in drug clearance is caused by the elevation in plasma pro-inflammatory cytokines, which can modulate the in vitro expression and activity of human hepatic CYP enzymes and transporters as well as renal transporters. However, the impact of these cytokines on the human intestine, a critical site for drug absorption, remains underexplored, presenting challenges in applying these findings in vivo. This study aimed to systematically investigate the influence of pro-inflammatory cytokines on the mRNA abundance of major intestinal drug transporters using an in vitro human enteroid model. These cells, chosen for their robust drug transport and metabolizing activities, closely mimic the functions of the intestinal mucosal epithelial barrier. Enteroid monolayers derived from three duodenal crypt stem cell donors were cultured in 96-well format, differentiated for 14 days, and then exposed for 48 hours, in triplicate, to interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ), either individually or in combination, at concentrations of 0.1, 1, and 10 ng/mL. These concentrations span from basal to supra-pathophysiological levels, corresponding to those observed in various infectious/inflammatory diseases. Following treatment, total RNA was isolated and subjected to quantitative PCR (qPCR) measurement of key intestinal transporter mRNAs: P-Glycoprotein (P-gp), Breast Cancer Resistance Protein (BCRP), Multidrug Resistance-Associated Protein 1, 2, 3, and 4 (MRP1/2/3/4), Organic Solute Transporter Alpha and Beta (OST-α/β), Organic Cation Transporter 1 and 3 (OCT1/3), Plasma Membrane Monoamine Transporter (PMAT), and Organic Anion Transporting Polypeptide 2B1 (OATP2B1). Exposure to a high concentration of pro-inflammatory cytokines down-regulated intestinal P-gp, PCRP, MRP2/3, OST α/ β, PMAT, and OATP2B1 mRNA levels, and these changes are expected to be concentration-dependent based on prior investigations of DME perturbations by cytokine exposure. To our knowledge, this is the first systematic investigation of the impact of cytokines on human intestinal drug transporters. Future work will focus on determining whether these changes in enteroid mRNA abundance correspond to alterations in intestinal transporter activity and/or protein abundance. These data are pivotal for developing physiologically based pharmacokinetic models to predict transporter-mediated drug pharmacokinetics during inflammation or infection.This work was supported by the NIH Grant R01HD102786.