Intestine related DMPK analysis using RepliGut® Planar Models

Replace Caco-2 cells and reduce late-stage failures

Importance of intestinal contribution to DMPK

In vitro modeling of intestinal absorption plays a pivotal role in advancing preclinical drug development by bridging the gap between laboratory research and clinical application. Predictions of bioavailability derived from cell culture models serve to refine drug design, optimize formulations, and flag potential safety issues. The RepliGut® Planar model is the first intestinal epithelial model that accurately reflects human intestinal contribution to metabolism and transport in a format similar to Caco-2 cells, enabling scientists to further close the gap between in vitro and in vivo bioavailability prediction and reduce late-stage clinical failures.

How RepliGut® Planar compares to Caco-2 models

Ensuring accuracy in intestinal absorption modeling necessitates the presence and functionality of pertinent influx and efflux transporters along with metabolic enzymes. While Caco-2 cells have conventionally served as the benchmark cell culture model for in vitro absorption investigations, their fidelity to native human intestinal tissues is compromised by unregulated proliferation, physiologically inaccurate differentiation processes, and altered expression of drug-related transporters and enzymes, thereby impeding their reliability in mimicking in vivo drug absorption and metabolism.

Caco-2 Cells
RepliGut® Planar
In Vivo
Human origin
Polarized epithelial monolayer
Multiple cell lineages
Regional specificity maintained
Interindividual variability
Mucus excretion
Phase I and II metabolism
96-well format

Unlike Caco-2 cells, RepliGut® models are comprised of multiple cell lineages found in the in vivo gut, which better reflect in vivo DMPK processes. The cell populations in RepliGut® models are confirmed to be ALP+, MUC2+ and CHGA+ positive and form a polarized monolayer with intact tight junctions.1 Furthermore, RepliGut® Jejunum models express more physiologically relevant levels of several genes encoding phase I and phase II metabolic enzymes reducing the chance of overpredicting bioavailability compared to Caco-2 cells

RepliGut® Models enable you to assess region-specific DMPK endpoints, from more than one donor, for a more thorough vetting of drug candidate bioavailability.

Schematic of RepliGut® Planar. Intestinal crypt-derived stem cells plated onto a semi-permeable membrane differentiate into multiple cell lineages including enterocytes, goblet cells, and enteroendocrine cells. DMPK relevant mechanisms include paracellular and transcellular passive uptake as well as active uptake and efflux. Metabolic processes could include P450, UGT, and CES enzymes.

Capabilities Summary

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Apparent permeability (Papp) and efflux ratios

Human jejunum epithelial stem cells are cultured and differentiated on semi-permeable membrane inserts in a 96-well plate to facilitate access to both the apical and basal sides of the cell monolayer. Test compounds are introduced into either compartment to assess their transport in both directions. Accurate mass spectrometry analysis of compound concentrations in each compartment allows for the calculation of permeability (Papp) and efflux ratios. Controls such as atenolol and propranolol, representing low and high permeability respectively, can be included. Additionally, transporter inhibitors can be applied to assess compound interactions with specific transporters. These assays are adaptable to specific research needs.

Typical study design
Cell Culture Timeline
10-11 days
Number of Replicates
Typically 3 wells per treatment
Incubation Time
Up to 120 min
Incubation Buffer
HBSS + 10mM HEPES + 10mM Glucose, pH 7.4
Barrier Integrity Assessment
Automated TEER using EVOM™ Auto (WPI) measured before and after compound incubation
Control Drugs
Atenolol and propranolol
Analysis Method
Accurate mass spectrometry measurement using LC/MS
Data Readout
Papp (apparent permeability coefficient), Efflux ratio, TEER
RepliGut® Planar- Jejunum accurately models passive and active drug transport. (A) Passive and (B) active transport drugs were applied to either the apical or basal compartment and drug concentration was measured in the receiver compartment at 2 hours post exposure. Digoxin and E3S transport were conducted in the presence and absence of P-gp inhibitor, tariquidar, and BCRP inhibitor, Ko143, respectively. Efflux ratios (green) were calculated as the ratio of B→A (pink) to A→B (blue).

Permeability markers tested in RepliGut® Planar-Jejunum

Absorption Group
RepliGut® A→B Papp
(10-6 cm/s)
Human Fraction Absorbed (Fa)
High absorption
(Fa ≥ 85%)
14.7 ±2.3
10.5 ±5.3
22.1 ±1.0
12.8 ±3.2
Moderate absorption
(Fa = 50 – 84%)
0.98 ±0.2
0.97 ±0.9
Low absorption
(Fa < 50%)
0.47 ±0.29
0.17 ±0.14
Absorption Group
Efflux Ratio
Efflux Substrates
12.5 ±5.8
30.7 ±14.4

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