Join MatTek at SOT 2017

Posted on March 9, 2017 |
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Assessment of the Phototoxicity of Three Different TiO2 Nano-Forms Using Reconstructed Human Tissue Model Epiderm. Abstract:1526 Poster: P614

Monday Afternoon, March 13 1:15 PM to 4:30 PM CC Exhibit Hall Poster Session: Nanotoxicology: In Vitro Author Attended: 3:00 PM-4:30 PM

Liskova1 , T. Milasova1 , S. Jantova2 , V. Brezova2 , and H. Kandarova1 . 1 Mattek In Vitro Life Science Laboratories, Bratislava, Slovakia; and 2 FCHPT, Bratislava, Slovakia.

Absorption of the solar light by photo-sensitive substances and consequent formation of reactive oxygen species (ROS) and other photo-products may lead to the cellular damage as well as to responses of the immune system. Taken that into the consideration, the determination of phototoxicity of substances absorbing UV and visible spectra of the solar light (VIS) belongs therefore to the basic toxicology tests. One of the methods used for the determination of phototoxicity is a test based on the use of 3D in vitro reconstructed human skin tissue model-EpiDermTM, the EpiDerm H3D-PT. This test was developed and pre-validated by organization ZEBET already in 1997. The main objective of this work was to verify and determine the phototoxicity and phototoxic potential of the selected reference substances and three different types of TiO2 nanoparticles using the EpiDermTM H3D-PT. We firstly evaluated and standardized the measurement conditions of the sunlight simulator SOL-500 and verified the sensitivity of the EpiDermTM tissues towards UV/VIS light. Next, we evaluated correct prediction of phototoxicity of the EpiDermTM H3D-PT using six reference substances, of which four were known phototoxins (chlorpromazine hydrochloride, two types of bergamot oil and anthracene) and two compounds were UV-absorbing, but without phototoxic potential (cinnamaldehyde, p-aminobenzoic acid). Finally, we have used this method to predict the phototoxicity of three different types of titanium dioxide (P25 AEROXID, Eusolex T-2000, TIG-115). Based on the results obtained in this work, we conclude that the EpiDermTM H3D-PT is a reliable test for the detection of phototoxicity and prediction of the phototoxic potential of selected substances. This conclusion is supported by the fact that during the measurements we obtained the same or better results as published by Liebsch et al. (1997). Phototoxicity of TiO2 has not been demonstrated in any of the three samples tested. This is because TiO2 nanoparticles do not penetrate deep enough into the epidermis to cause cytotoxicity by irradiation with UVA/VIS.

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CON4EI: EpiOcular Eye Irritation Tests (OECD TG 492 and ET-50 Protocols). Abstract: 1767 Poster: P303

Tuesday Morning, March 14 9:30 AM to 12:45 PM CC Exhibit Hall Poster Session: Ocular Toxicology Author Attended: 11:15 AM-12:45 PM

H. Kandarova1 , S. Letasiova1 , E. Adriaens2 , N. Alépée3 , A. Drzewiecka4 , P.M. Fochtman4 , K.D. Gruszka4 , R. Guest5 , G. Maglennon5 , J. Schofield6 , J.A. Willoughby7 , S. Verstraelen8 , and A.R. Van Rompay8 . 1 MatTek In Vitro Life Science Laboratories, Bratislava, Slovakia; 2 Adriaens Consulting BVBA, Aalter, Belgium; 3 L’Oréal Research & Innovation, Aulnay-sous-Bois, France; 4 Institute of Industrial Organic Chemistry Branch Pszczyna, Pszczyna, Poland; 5 ENVIGO, Derby, United Kingdom; 6 ENVIGO, Cambridgeshire, United Kingdom; 7 Cyprotex US, Kalamazoo, MI; and 8 VITO NV, Applied Bio & Molecular Systems, Mol, Belgium

Assessment of the acute eye irritation potential is a part of the international regulatory requirements for testing of chemicals. The objective of the CON4EI (CONsortium for in vitro Eye Irritation Testing Strategy) project is to develop tiered testing strategies for eye irritation assessment for all drivers of classification. For this, a set of 80 reference chemicals (38 liquids and 42 solids) was tested with eight different alternative methods. Here, the results obtained with reconstructed human cornea-like epithelium (RhCE) EpiOcular and the two EpiOcular Eye IrritationTests (EITand ET-50) are shown. The primary aim of this study was an evaluation of the performance of the test methods to discriminate chemicals not requiring classification for serious eye damage/eye irritancy (No Category) from chemicals requiring classification and labeling (Category 1 and 2). In addition, the predictive capacity in terms of in vivo driver of classification was investigated. In a second step, it was investigated whether the EpiOcular EIT and ET-50 can be used also for prediction of sub-classes of ocular irritation.

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Hyper-Osmolal Vaginal Lubricants Markedly Increase Epithelial Damage in a ThreeDimensional Vaginal Epithelium Model. Abstract: 1871 Poster: P447

Tuesday Morning, March 14 9:30 AM to 12:45 PM CC Exhibit Hall Poster Session: Non-Pharmaceutical Safety Assessment Author Attended: 11:15 AM-12:45 PM

S. Ayehunie1 , T. Landry1 , S. Bogojevic1 , P. Hayden1 , R. Cone2 , and Y. Wang2 . 1 MatTek Corporation, Ashland, MA; and 2 Johns Hopkins University, Baltimore, MD.

Widely used vaginal lubricants in the U.S. and Europe are strongly hyperosmolal, formulated with high concentrations of glycerol, propylene glycol, poly-quaternary compounds, or other ingredients that make these lubricants 4 to 30 times the osmolality of healthy vaginal fluid.  Hyperosmolal formulations have been shown to cause marked toxicity to human colorectal epithelia in vivo and significantly increase vaginal transmission of genital herpes infections in the mouse/HSV model.  They also cause toxicity to explants of vaginal epithelia or cultured vaginal epithelial cells and increase susceptibility to pathogenic organisms such as HIV in target cells in a cell culture system. Here, we report that the average osmolality of healthy vaginal fluid collected from eight donors with an average of five samples per donor is 370±40 mOsm/Kg.  Using a well-characterized three-dimensional human vaginal epithelium tissue model (EpiVaginal™), we demonstrated that hyperosmolal vaginal lubricants with osmolality greater than 4 times that of vaginal fluid (>1500 mOsm/Kg) cause disruption of epithelial barrier properties and structural damage.  Four out of four such lubricants caused tissue disruption in the parabasal and basal cell layers as observed histologically.  These lubricants also reduced/compromised barrier integrity as measured by the trans-epithelial electrical resistance (TEER).  No epithelial damage or reduction in MTT viability was noted for hyposmolal lubricants (N=3) with osmolality of <370 mOsm/Kg. The results showed the utility of the organotypic EpiVaginal tissue model for lubricant screening and this in vitro test method confirm the extensive reports of safety concerns of hyperosmolal lubricants, which strongly suggest that the FDA review the toxic effects of currently marketed vaginal lubricants.

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Determination of Contact Sensitization Potential of Chemicals Using In Vitro Reconstructed Normal Human Epidermal Model Epiderm: Impact of the Modality of Application. Abstract: 2746 Poster: P114

Wednesday Afternoon, March 15 1:15 PM to 4:30 PM CC Exhibit Hall Poster Session: Alternatives to Mammalian Models I: Liver, Ocular, and Skin Alternatives Author Attended: 1:15 PM-2:45 PM

S. Letasiova1 , E. Corsini2 , V. Galbiati2 , H. Kandarova1 , D. Lehmeier3 , and H. Gehrke3 . 1 Mattek In Vitro Life Science Laboratories, Bratislava, Slovakia; 2 Università degli Studi di Milano, Milan, Italy; and 3 Eurofins BioPharma Product Testing Munich GmbH, Munich, Germany.

Assessment of skin sensitization potential has traditionally been conducted in animal models, such as the Mouse Local Lymph Node Assay (LLNA) and the Guinea Pig Maximisation Test (GPMT). However, a growing focus and consensus for minimizing animal use have stimulated the development of in vitro methods to assess skin sensitization. Interleukin-18 (IL-18) release in reconstructed human epidermal models has been identified as a potentially useful endpoint for the identification and classification of skin sensitizing chemicals, including chemicals of low water solubility or stability (1). The purpose of this study was to investigate the impact of the modality of chemical exposure on the predictive capacity of the assay. EpiDerm tissue viability assessed by MTT assay and IL-18 release assessed by ELISA were evaluated after 24 h topical exposure to test chemicals either impregnated in 8 mm diameter paper filters or directly applied to the surface of EpiDerm. Acetone: olive oil (4:1) was used as vehicle in all cases. A total of five chemicals from 3 different sources were tested. The testing set included 3 sensitizers, namely 2,4-dinitrochlorobenzene, cinnamaldehyde and isoeugenol/eugenol, and 2 non sensitizers, lactic acid and salicylic acid. Four independent dose – response experiments were conducted in 3 laboratories, resulting in correct prediction of the sensitizing potency of test chemicals. The assessment of IL-18 release using in vitro reconstructed normal human epidermal model EpiDerm appears to be a promising tool for in vitro determination of contact sensitization potential.

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Molecular Mechanisms of Corneal Oxidative Stress: In Vitro-Reconstructed Human Corneal Tissue Model. Abstract: 2756 Poster: P124

Wednesday Afternoon, March 15 1:15 PM to 4:30 PM CC Exhibit Hall Poster Session: Alternatives to Mammalian Models I: Liver, Ocular, and Skin Alternatives Author Attended: 1:15 PM-2:45 PM

Y. Kaluzhny, M.W. Kinuthia, T. Truong, A. Plotkin, P. Hayden, and M. Klausner. MatTek Corporation, Ashland, MA.

The cornea is directly exposed to UV radiation which is an environmental stress factor. Oxidative damage plays an important role in the pathogenesis of many ocular diseases, including dry eye disease (DED). Current methods used to investigate the mechanisms of corneal injuries utilize monolayer cell cultures or animals.  As a result, there is a need for more physiologically relevant, human-based in vitro models for ocular research. This study explores the relationship between oxidative stress and DED using an in vitro reconstructed corneal tissue model comprised of normal human corneal epithelial cells cultured at the air-liquid interface.  The tissues express corneal-specific genes essential for metabolism and detoxification: aldehyde dehydrogenases (ALDH3A1 and ALDH1A1), thioredoxin reductase (TXNRD1), UDP glucoronosyltransferase 1A1 (UGTIA1), and cytochrome P450 enzymes.  Oxidative stress was generated by exposing the tissues to simulated solar UV radiation  (60 mJ and  120 mJ UVB), H2O2 (20 mM and 50 mM) or to desiccating stress conditions (DSC, 40% RH, 40°C, and 5% CO2) to stimulate morphological, cellular, and molecular changes relevant to DED. The effect of oxidative stress on the corneal tissues was analyzed at 0, 2, 24, and 48h after UV exposure or DSC. Reactive oxygen species (ROS), cytokine release, transepithelial electrical resistance, tissue viability, histology, and gene expression were evaluated.  UV exposure resulted in 38.5 to 60.2 fold increases in cellular ROS accumulation, which decreased by 73% 2h post-exposure.  24h incubation at DSC induced 3 to 5.5 fold increases in intracellular ROS as detected by the DCFH-DA assay (intracellular H2O2) and a 41.5 fold as detected by 8-Isoprostane assay (phospholipid oxidation).  Both conditions caused considerable release of IL-8 and gene upregulation for pro-inflammatory cytokines and enzymes (IL-1, IL-6, PTGS2, SOD1, and MMP13), but didn’t affect tissue viability.  Following exposure to UV or DSC, topical application of lubricant gel drops (25 µl/tissue) improved tissue morphology and barrier function, but didn’t have an effect on cytokine release or gene expression. The results demonstrate that the in vitro organotypic human corneal tissue structurally and functionally reproduces oxidative stress and DED markers.  The model will avoid species extrapolation, be more cost effective and more reproducible than animal methods, and will facilitate drug discovery by allowing screening and optimization of pharmaceutical formulations.

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In Vitro Reconstructed Human Corneal Tissue Model for Ophthalmic Drug Delivery. Abstract: 2757 Poster: P125

Wednesday Afternoon, March 15 1:15 PM to 4:30 PM CC Exhibit Hall Poster Session: Alternatives to Mammalian Models I: Liver, Ocular, and Skin Alternatives Author Attended: 1:15 PM-2:45 PM

Y. Kaluzhny1 , M.W. Kinuthia1 , A.M. Gremilogianni2 , C.G. Tsoli2 , P. Hayden1 , and M. Klausner1 . 1 MatTek Corporation, Ashland, MA; and 2 University of Athens, Athens, Greece.

The corneal barrier is vitally important for eye protection, but it also presents a major challenge for delivery of ophthalmic drugs.  Another complication is that the cornea expresses drug-metabolizing enzymes and transport systems that can affect drug delivery. Most current studies use excised rabbit corneas that have many shortcomings including poor species extrapolation, difficulty to standardize, high cost, and ethical concerns. To address the need for physiologically relevant, human-based in vitro models for ophthalmic drug delivery studies, we developed an in vitro reconstructed corneal model. The multilayered cultures contain normal human corneal epithelial cells that grow at the air-liquid interface and develop barrier properties comparable to the in vivo human cornea. Reconstructed tissues were characterized by histology, confocal microscopy, transepithelial electrical resistance, esterase activity, and expression of metabolic enzymes and drug transporters.  Corneal permeability was evaluated using model compounds with a wide range of hydrophobicity, molecular weights, and excipients; ophthalmic related antibiotics, and seven formulations of Latanoprost eye drops. The tissues express mucins 4 and 16, as well as corneal-specific genes essential for metabolism and detoxification: aldehyde dehydrogenases (ALDH3A1 and ALDH1A1), thioredoxin reductase (TXNRD1), UDP glucoronosyltransferase 1A1 (UGTIA1), carboxyl esterase 1 (CES1), and low, but detectable levels of cytochrome P450’s (CYP1A1, CYP2C19, CYP2C8, CYP2C9, CYP2D6, CYP3A5). mRNA expression of efflux transporters (ABCB1, ABCC1, ABCC3, ABCC5, ABCG2), peptide transporters (SLC15A1, SLC15A2), and uptake transporters (SLC22A1, SLC22A3) could be detected in the tissues. The correlation of permeation coefficients (r2) to excised animal corneas for model drugs was 0.84. Esterase activity was evident as Latanoprost acid was readily detected in the receiver compartment. Of seven formulations of Latanoprost eye drops tested, Xalatan (containing 0.02% BAC) had the highest permeation (Papp = 8.81cm×s-1) and Monoprost (preservative-free) had the lowest (Papp =1.15 cm×s-1) (1). The reconstructed in vitro corneal tissue morphology, barrier properties, gene expression, and permeability resemble that of the in vivo human cornea. This model is anticipated to be a useful tool to evaluate new ophthalmic formulations.

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Development of an Alternative In Vitro Method for Acute Inhalation Toxicity Testing. Abstract: 2777 Poster: P145

Wednesday Afternoon, March 15 1:15 PM to 4:30 PM CC Exhibit Hall Poster Session: Alternatives to Mammalian Models II Author Attended: 3:00 PM-4:30 PM

G.R. Jackson, A. Hunter, A. Maione, M. Klausner, and P.J. Hayden. MatTek Corporation, Ashland, MA.

Knowledge of acute inhalation toxicity and irritation potential is important for establishing safe handling, labeling and emergency response procedures for chemicals. The US EPA High Production Volume Chemical Challenge, and the EU REACH programs have further increased the need for inhalation toxicity information. The Globally Harmonized System (GHS) of Classification and Labeling of Chemicals utilized by government regulators worldwide specifies 5 inhalation toxicity categories. The EPA has established a separate system that uses 4 toxicity categories. Acute inhalation toxicity tests currently accepted within the GHS and EPA systems involve in vivo 4 hr rat inhalation LC50 tests (OECD TG 403/436). In the current work, a newly developed in vitro toxicity test was evaluated in comparison to the established in vivo tests.

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Use of In Vitro Human 3D Airway Tissue Models to Study Influenza Virus Infection of the Upper and Lower Respiratory Tract. Abstract: 2778 Poster: P146

Wednesday Afternoon, March 15 1:15 PM to 4:30 PM CC Exhibit Hall Poster Session: Alternatives to Mammalian Models II Author Attended: 3:00 PM-4:30 PM

A. Martyna1 , A.G. Maione2 , G.R. Jackson2 , O. O’Connell2 , B. Bahsoun1 , D. Cantoni1 , P. Hayden2 , and J. Rossman1 . 1 University of Kent, Canterbury, United Kingdom; and 2 MatTek Corporation, Ashland, MA.

Although influenza infection can be prevented through vaccination, continual mutations require rapid and ongoing production of new vaccines.  Our ability to design effective vaccines depends on better understanding the range of virus-host interactions.  Here, we evaluate how cellular differentiation in both the upper and lower airways affects viral replication and host response. To more accurately recapitulate the complexity of tissue structure and differentiation within the respiratory tract, two commercially available human 3D tissue models of tracheal/bronchial tissue, EpiAirwayTM, and alveolar tissue, EpiAlveolarTM, were utilized.  Tissues were infected with H3N2 Influenza A Virus (IAV) in parallel with the immortalized alveolar epithelial cell line, A549.  Cultures were assessed for viral replication, viral protein production and trafficking, viral assembly and morphology, transepithelial electrical resistance (TEER) and inflammatory cytokine secretion. The A549 cell line is one of the most widely used cell lines for modeling the host-pathogen responses to IAV infection.  However, our results show that both viral and host responses differ significantly between differentiated primary airway tissue models and immortalized 2D monolayer cells.  While all cell systems tested were capable of being productively infected with IAV strains, the profile of viral replication and host response differed greatly between systems.  In all cases, the differentiated primary airway tissues showed lower viral replication and altered protein trafficking and viral morphology, when compared to A549 cell monolayers, despite showing significant reductions in TEER upon infection.  EpiAlveolarTM and EpiAirwayTM tissues showed aberrant protein localization and altered host response, including cytokine secretion. In addition, viral protein trafficking appears to be significantly different in upper vs. lower airway tissues, though the impact of this on the overall host-pathogen interaction remains unclear. Overall our results show that differentiated primary airway tissue models offer important insight into IAV replication when compared to standard tissue culture model systems.

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Use of an Organotypic Small Intestinal Tissue Model for Drug Toxicity, Permeability, Drug-Drug Interaction, and Metabolism Studies. Abstract: 3146 Poster: P105

Thursday Morning, March 16 8:30 AM to 11:45 AM CC Hall A Poster Session: Alternatives to Mammalian Models III Author Attended: 8:30 AM-10:00 AM

S. Ayehunie1 , Z. Stevens1 , T. Landry1 , A. Armento1 , B. Press2 , and P. Hayden1 , and M. Klausner1 . 1 MatTek Corporation, Ashland, MA; and 2 Cyprotex, Waltham, MA.

The goal of this study was to validate the biological relevance of an in vitro organotypic small intestinal (SMI) tissue model for predicting intestinal drug toxicity and permeation. A primary human cell based small intestinal (SMI) 3D tissue model that recapitulates the in vivo counterpart was used to evaluate the apparent permeability coefficient of N = 16 drugs for which in vivo data are available (Biopharmaceutics Classification System).  Toxicity was assessed using transepithelial electrical resistance (TEER), MTT viability, and Lucifer Yellow (LY) leakage assays.  Bioavailability or efflux transport of drugs was analyzed using LC-MS/MS.  The metabolic activity of the SMI tissue model was assessed using specific substrates and metabolites were analyzed using LC-MS/MS.  Test drugs with human absorption of >80% (high permeability drugs) had in vitro apparent permeability coefficients (Papp) of >2 x106 cm sec-1 and drugs with <80% human absorption (low permeability drugs) had  in vitro Papp values of <2 x106 cm sec-1.  Using these criteria, the SMI tissue model categorized the test drugs as high permeability and low permeability with a sensitivity of 100%, specificity of 89%, and accuracy of 94% compared to historical human absorption data.  Drug-drug interactions were also examined using efflux transporter inhibitors.  The inhibitors increased drug bioavailability while decreasing the efflux ratio.  Efflux ratios for substrates talinolol, digoxin, and loperamide were reduced by 45%, 40%, and 60%, respectively, in the presence of the P-gp inhibitor, verapamil.  The efflux ratio of nitrofurantoin (BCRP substrate) was reduced by 63% in the presence of the BCRP inhibitor, novobiocin.  Results from drug metabolism studies also showed midazolam (CPY3A substrate) was metabolized (6.5% conversion) by the intestinal tissue model. In conclusion, the SMI tissue model appear to be promising new tool for evaluation of drug safety, permeability, and metabolism.

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Inter-Laboratory Reproducibility of a Three Dimensional Model of Human Buccal Tissue. Abstract: 3147 Poster: P106

Thursday Morning, March 16 8:30 AM to 11:45 AM CC Hall A Poster Session: Alternatives to Mammalian Models III Author Attended: 8:30 AM-10:00 AM

M. Bachelor1 , F. Zanetti2 , H. Behrsing3 , H. Raabe3 , L. Ortega-Torres2 , C. Merg2 , R. Duliz2 , N. Sadowski3 , B. Breyfogle1 , J. Molignano1 , G. Booker1 , E. Guedj2 , S. Frentzel2 , A. Elamin2 , C. Mathis2 , M. Peitsch1 , and J. Hoeng2 . 1 MatTek Corporation, Ashland, MA; 2 Philip Morris International R&D, Neuchâtel, Switzerland; and 3 Institute for In Vitro Sciences, Gaithersburg, MD. Sponsor: P. Hayden.

Human three-dimensional (3D) tissue models of the human oral epithelia have been developed as an alternative to animal testing.  3D tissue models are powerful tools for investigating basic biological function in response to various challenges including oral care products and environmental insult.  To evaluate intra-laboratory reproducibility of the oral epithelial model for broader utility, tissues were exposed to three different inducers in three phases using independently produced lots of EpiOral, a commercially available 3D model of human buccal tissue .  Tissues were exposed to TNF-a/IL-1b to evaluate inflammatory response, 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) to evaluate xenobiotic metabolism,or TritonX-100 to evaluate cytotoxicity using the EpiOral model.  Adenylate kinase release, MTT activity, pro-inflammatory marker release and xenobiotic phase I enzyme expression were analyzed following exposure of the tissues to TCDD or TNF-a/IL-1b.  A total of three experimental phases were conducted to evaluate inter-phase reproducibility.  Following exposure of oral tissues to inflammatory inducers TNF-a/IL-1b, significant increases in secreted IL-1b, IL-8, IP-10 and MMP-1 were observed.  Exposure of oral tissues to TCDD resulted in significant increases in the gene expression of Phase I metabolism enzymes including CYP1A1 and CYP1B1.  A concomitant increase in CYP1A1/1B1 activity, albeit varied, was also observed in response to TCDD treatment in all phases.  A significant induction in cytokine release (IL-8 and IL-1b, and IP-10) was also observed in all phases.  In summary, we demonstrate utility of the EpiOral model for evaluation of cytotoxicity, xenobiotic metabolism, gene activation and cytokine profiling in response to various stimuli.

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Prediction of Drug-Induced Gastrointestinal (GI) Toxicity and Wound Healing Using 3D-Human Small Intestinal Tissue Model System. Abstract: 3148 Poster: P107

Thursday Morning, March 16 8:30 AM to 11:45 AM CC Hall A Poster Session: Alternatives to Mammalian Models III Author Attended: 8:30 AM-10:00 AM

S. Ayehunie1 , T. Landry1 , Z. Stevens1 , A. Armento1 , M. Wagoner2 , M. Klausner1 , and P. Hayden1 . 1 MatTek Corporation, Ashland, MA; and 2 AstraZeneca, Waltham, MA.

The mechanisms of off-target, dose-limiting gastrointestinal toxicity for therapeutic compounds are often poorly understood. Lack of physiologically-relevant in vitro models and species differences in intestinal physiology is a challenge for gastrointestinal (GI) toxicity studies. Here, we evaluated the utility of a primary human cell based 3D small intestinal tissue (SMI) as an investigational tool for drug induced GI toxicity and wound healing studies. To achieve this goal, a blinded study was performed using N=8 drugs (5 problematic and 3 well-tolerated in humans) for which dog and rat toxicity studies were not predictive of human toxicity. The compounds were tested at five concentrations. Using the in vitro SMI model, we examined cytotoxicity by MTT and LDH assays and tissue barrier integrity using transepithelial electrical resistance (TEER) measurements, following two exposures over 96 hours. The results showed that the SMI tissue detected drug induced reduction in TEER in 5/5 of the problematic drugs at concentrations at or below 30x clinical Cmax, while the 3 well-tolerated drugs showed no effect on TEER at up to 1,000x clinical Cmax.  The utility of the tissue model for wound healing studies was examined by inducing a 2 mm wound area. Tissues were cultured for up to 10 days and wound healing was monitored by confocal imaging for epithelial cell migration and fibroblast spreading. Tissues cultured in human serum completely healed at day 6 of the culture period compared to >10 days for control tissues. In summary, TEER measurements on the SMI tissue is a sensitive, predictive tool to assess clinically-relevant intestinal toxicity that appears to give improved predictions versus animal preclinical toxicology models.  Furthermore, the small intestinal tissue model can be used to evaluate therapeutics for their wound healing potential.

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Development of an In Vitro Model of Mucociliary Rat Airway Epithelium for Inhalation Toxicity Testing. Abstract: 3241 Poster: P182

Thursday Morning, March 16 8:30 AM to 11:45 AM CC Hall A Poster Session: Emerging Technologies Author Attended: 10:00 AM-11:45 AM

G.R. Jackson, A. Maione, M. Klausner, and P.J. Hayden. MatTek Corporation, Ashland, MA.

Knowledge of inhalation toxicity and irritation potential is important for development of new inhalable therapeutics, assessment of environmental air pollutants, and for establishing safe handling, labeling, and emergency response procedures for chemicals. Animal models have been traditionally used to assess the respiratory effect of new drugs or environmental agents.  However, there is growing awareness and concern that animal models do not always accurately predict human outcomes. Therefore, development of in vitro model systems based on primary human cells has been a high priority in the scientific community. In vitro organotypic airway models based on primary human airway cells are well established and are commercially available.  However, validation of these systems for prediction of in vivo human outcomes is problematic because most available in vivo toxicity data was produced from animal models. Thus, discrepancies between human in vitro data and animal in vivo data may either be model differences or species differences. The missing link for validation is a corresponding in vitro animal airway model. In vitro organotypic airway models of various species including mice, rats and dogs are also well established in the literature, but have been challenging to produce on a large scale. The goal of the current project is to develop a scalable in vitro organotypic model of rat mucociliary airway epithelium to allow for comparison with human in vitro airway models and thus close the loop in the current validation scheme for human derived in vitro organotypic mucociliary airway epithelium. Airway tissues were obtained from 8-week old, male, CD rats (Charles River).  Conducting airways were dissected and epithelial cells were isolated following enzymatic digestion. Isolated cells were seeded onto microporous membrane inserts and cultured at the air-liquid interface for up to 27 days. H&E stained histological evaluation demonstrated a high level of organotypic mucociliary differentiation including a pseudostratified epithelial structure with copious development of functional cilia. Robust barrier function was demonstrated by development of transepithelial electrical resistance of >800 Ω x cm2.  These results suggest that in vitro rat airway models may be a promising tool to allow rodent to human translation of in vitro inhalation toxicology data, and ultimately a full transition to human based in vitro models.

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