Novel microdosing device

Abstract

The present invention concerns a microdosing test device comprising: 1) a layer impermeable to aqueous liquids wherein the layer comprises a biocompatible and biodegradable adhesive for a mucosal surface of an animal, and 2) a carrier adapted for bringing a test compound in a non-therapeutic concentration in physical contact with the mucosal surface; wherein the layer when applied to the mucosal surface forms a microdosing compartment between the layer and the mucosal surface which compartment contains the carrier.

Description

[0001] NOVEL MICRODOSING DEVICE

[0002] Technical field

[0003] The present invention relates to a microdosing test device, use of such device as well as a method for assessment of PK / PD using said device.

[0004] Background Art

[0005] Development of drugs is costly with a high risk of failure.1An analysis of where the main expenses and time consumption of investigational products (IPs) development lie clearly shows that this in the clinical phases of drug development programs, i.e., from first-in-human phase one studies to large phase three randomized clinical trials; with two thirds of the cost lying in the clinical phase. Further, 7-17% of costs lies in toxicology and bioavailability experiments on lead compounds before the program goes into the clinical phase.1Termination in clinical phase is often due to poor animal models of the disease of interest and / or differences in bioavailability between animals and humans.1, 2A method to provide earlier PD / PK screening of lead NMEs in humans without large toxicology programs would decrease risk of NME development. This is the background for the microdosing device development.

[0006] The intestine has a dual function of providing the necessities for the maintenance and growth of the cells of the body as well as being the main interface between the environment and the immune system. Further, the intestine is the preferred and main route of administration of most therapeutics. The interface function is an intricate interplay between an efficient barrier function, the regulatory layers of the innate and adaptive immune system resulting in a constant processing and appropriate reaction towards environmental microorganisms and antigens. Accordingly, the intestine is by far the biggest immune organ of the body in terms of cell numbers involved, and dysfunctions in this immune balance will result in inflammation and barrier impairment as well as collateral decreased absorption capacity of the intestine, as seen in conditions like uncontrolled sustained infections, coeliac disease or inflammatory bowel disease (IBD).3

[0007] Although mice share more than 90% of genes with humans striking differences exist in the intestinal barrier function and immune system. Thus, the development and maintenance of the epithelial barrier varies substantially across species.4Even greater differences are present in immune function where humans produce a dynamic antibody repertoire of private clonotypes whereas mice maintain a stable core of public clonotypes - a difference mainly explained by the body size dependent limitation in ability to produce clonotypes.5Immune signatures from both specific immune cells and from disease models in mice compared with the corresponding human condition has revealed a number of similarities but also significant differences in the immune responses, e.g. in the involvement of interferon signaling and regeneration.6Further, the intestinal microbiota is increasingly recognized as a key player in human intestinal disease development, but is highly species dependent and restricted in modern breeding facilities, which specifically is affecting research in intestinal homeostasis processes.7Also highly specific pathways have been shown to differ significantly between mice and humans in ways, where a pathway could be manipulated with therapeutic effects in one species, but not in the other: an example is the orphan chemoattractant receptor GPR15 which is mainly expressed on regulatory T (Treg) cells and controlling their intestinal homing in mice, whereas human Treg cells do not express GPR15.8

[0008] There is inter alia an unmet need for a sealed off compartment (the microdosing device) over intact intestinal surface or an experimental confined microinjury will allow molecular manipulation in a confined area of the intestine. Such device can thus be used to screen lead IPs by determination of bioavailability and pharmaco-dynamics at very low IP-concentrations in humans.

[0009] Summary of the Disclosure

[0010] The present inventors have experienced that development of IPs that make it to become new medical treatment for diseases is costly and there is a high risk of new treatment principles failing when the development process goes from the preclinical animal model-based phase to the clinical phase in humans. This is due to substantial differences in animal and human physiology. For example, there are fundamental differences between the immune systems of rodents and humans, which can partly be explained in the number of immune cells (and thus the possibility of diversity in response) based on body weight. This is also because there are no good animal models for a number of diseases, including polygenic diseases (diseases that require multiple genetic factors to manifest) in humans. The currently available animal models are often genetically simplified and accelerated compared to the diseases they model. This is also because treatment outcomes are fundamentally different in animal models than in the disease being modelled.

[0011] The present inventors have developed a human microdosing model (“the microdosing test device”) to investigate the effect of new medicines (IPs) that allow minimal toxicological studies in animal models and accelerated test of bioavailability (BA) and mechanism of action (MO A) in humans. The microdosing model can inter alia be used for MOA / BA studies of IPs developed for gastrointestinal diseases. The microdosing model may also be used for BA studies and some MOA studies of orally administered IPs developed for non-gastrointestinal diseases.

[0012] The present invention concerns in one aspect a microdosing test device comprising:

[0013] 1) a layer impermeable to aqueous liquids wherein the layer comprises a biocompatible and biodegradable adhesive for a mucosal surface of an animal, and

[0014] 2) a carrier adapted for bringing a test compound in a non-therapeutic concentration in physical contact with the mucosal surface; wherein the layer when applied to the mucosa forms a microdosing compartment between the layer and the mucosal surface which compartment contains the carrier.

[0015] In one embodiment the animal is a mammal. Preferably, a human.

[0016] In an embodiment the layer is made of an elastic and / or flexible polymer. Typically, the polymer is selected from the group consisting of a polyurethane (PU), a polyethylene (PE), a gas permeable polydimethylsiloxane (PDMS; silicone), a polycarbonate (PC), a polymethylmethacrylate (PMMA), and a polyvinylchloride (PVC).

[0017] In a further embodiment the carrier is a liquid, gel, or a solid carrier, preferably a cellulose carrier. The cellulose carrier is typically a filter paper.

[0018] In a still further embodiment the adhesive is selected from a group consisting of a polyethylene glycol (PEG) based adhesive, a cyanoacrylates (CA) based adhesive, a thrombine / fi- brin-based adhesive, a collagen / gelatin based adhesive, a polysaccharide based adhesive, such as chitosan (CHI), poly(lactic-co-glycolic) acid (PLGA), and a polyurethane based adhesives; as well as combinations hereof.

[0019] In a further embodiment the test compound is loaded into or on the carrier.

[0020] In a still further embodiment the concentration of the test compound in physical contact with the mucosa is non-toxic to the animal.

[0021] In a further embodiment the concentration of the test compound in physical contact with the mucosa is at least 1: 1000 of a therapeutic concentration to the animal. Typically, at least 1 : 10000. A preferred embodiment is from 1 : 10'5to 1 : 10'7such as 1 : 10'6of a therapeutic concentration to the animal.

[0022] In a still further embodiment the mucosa is a gastrointestinal tract of the animal. Typically, the mucosa is an oral cavity, stomach, small, or large intestine, a stoma, an ileostomy or a colostomy.

[0023] In a further embodiment the layer has a surface area of at least 50 mm2, a circumference of from 25-63 mm and a diameter of from 8-20 mm and is made of a water impermeable polymer. Typically, the water impermeable polymer is a transparent polyethylene or a transparent PU.

[0024] In a still further embodiment the layer has a border applied to the circumference of the layer, which border is 3-5 mm wide and 0.4-0.6 mm high and the adhesive is applied to the border. Typically, the border is made of PU applied to the circumference of the layer.

[0025] In a further embodiment PK / PD of the test compound is investigated. Typically, bioavailability and / or mechanism of action of the test compound is investigated.

[0026] In a still further embodiment the test compound is investigated for a gastrointestinal disease. In a further embodiment the gastrointestinal disease is selected from gastroduodenal ulcer diseases, gastritis, celiac disease, Crohn’s disease, ulcerative colitis, microscopic colitis, infectious gastrointestinal diseases, ulcerating diseases, barrier defect diseases, vascular intestinal diseases, or neoplastic gastrointestinal diseases.

[0027] In a still further embodiment the test compound is for oral administration and is investigated for a non-gastrointestinal disease. Typically, the non-gastrointestinal disease is diabetes type I or type II. Each of the above embodiments of the microdosing test device should be seen as referring to any one of the embodiments described above in any combination unless it is specified that an embodiment relates to a certain aspect or a certain embodiment of the present invention.

[0028] In a second aspect the present invention concerns use of a microdosing test device comprising:

[0029] 1) a layer impermeable to aqueous liquids wherein the layer comprises a biocompatible and biodegradable adhesive for a mucosal surface of an animal, such as a mammal, e.g., a human, and

[0030] 2) a carrier adapted for bringing a test compound in a non-therapeutic concentration in physical contact with the mucosal surface; wherein the layer when applied to the mucosa forms a compartment between the layer and the mucosa which compartment contains the carrier; for assessment of PK / PD of the test compound. Typically, bioavailability and / or mechanism of action of the test compound is assessed.

[0031] Embodiments of the second aspect are any one of the above embodiments of the first aspect and should be seen as embodiments of the second aspect in any combination unless it is specified that an embodiment relates to a certain aspect or a certain embodiment of the present invention.

[0032] In a third aspect the present invention concerns a method for assessment of PK / PD of a test compound in an animal comprising a) applying the test device of the first aspect wherein the test compound is loaded into or on the carrier to a mucosal surface of an animal, b) allowing exposure of the mucosal surface to the test compound for sufficient time, c) obtaining a biopsy or micropipette / syringe sample from the mucosa, d) analyzing a body surface fluid from the compartment, e) analyzing the biopsy or micropipette / syringe sample to obtain data on PK / PD of the test compound.

[0033] In an embodiment the method comprising pretreatment of the exposed area to induce local inflammation or barrier breach is optional, wherein inflammation / perfusion / regeneration can be assessed by imaging. In a further embodiment the present invention concerns a method for assessment of bioavailability and / or mechanism of action of a test compound.

[0034] In a still further embodiment wherein sufficient time is from 4 hours to 4 days.

[0035] Embodiments of the third aspect are any one of the above embodiments of the first aspect and should be seen as embodiments of the third aspect in any combination unless it is specified that an embodiment relates to a certain aspect or a certain embodiment of the present invention.

[0036] Brief description of drawing

[0037] Figure 1 shows an embodiment of the microdosing devise of the present invention as separate elements of the device and assembled.

[0038] Figure 2 shows the embodiment of the microdosing devise of the present invention assembled.

[0039] Figure 3 shows the embodiment of the microdosing devise of the present invention placed on a mucosal surface.

[0040] Figure 4 illustrates regulation of enterocyte differentiation marker CDX2 at experimental microinjury compartment of the human intestine determined by PCR.

[0041] Detailed description

[0042] The present invention concerns a microdosing model based on extensive human studies of mucosal damage in micro-models that have been shown to model active Crohn’s disease and ulcerative colitis. The invention is described below in more detail referring to embodiments which by no means should be seen as limiting the scope of the claims.

[0043] The microdosing model includes a medical device that allows microdosing of a test compound (here test compound is used interchangeably with IP, that is Investigational Product) on a well-defined area of the small or large intestinal mucosa (see Figures 1-3).

[0044] Figure 1 shows an embodiment of the microdosing device as separate elements. The first element of figure 1 A is a microdosing carrier (2) having a circumference (4) adapted to be applied to a layer impermeable to aqueous liquids (6) said layer typically made of a polymer which is elastic and / or flexible. The layer (6) as shown is also circular having a circumference (8). A further element is a border (10) which is adapted to be applied to the layer (6) at the circumference (8) of the layer (6). The border (10) has an opening (16) fitted to the microdosing carrier (2) and being circular having an outer diameter (12) and an inner diameter (14), wherein the outer diameter fits the circumference (8) of the layer (6) without extending beyond said circumference (8). When being ready for a test compound adhesive is applied to the border (10) facing a mucosal surface.

[0045] Figure 2 shows the embodiment of figure 1 assembled. In figure 2A the microdosing test device is seen from the surface to be applied to the mucosal surface wherein the microdosing carrier (2) has a circumference (4) which circumference (4) fits inside the border (10) having the outer diameter (12) with the same diameter as the layer (6). This is further shown in figure 2B wherein the microdosing test device (2, 6) is seen from the surface facing away from the mucosal surface. The dashed lines (4) indicate the carrier (2) applied to the opposite site.

[0046] Figure 3 shows the embodiment of figure 2 (2, 6, 10) seen from the side and in a cross- sectional view when being applied to the mucosal surface (18). When applied to the mucosal surface (18) the adhesive is applied to the border (10) on the surface facing the mucosal surface (18).

[0047] Typically, the edge of the device having the border allows adhesion to the human intestinal mucosa by a biocompatible, biodegradable polyethylene glycol-based adhesive (Coseal, Baxter, Deerfield, IL, USA),9‘10.

[0048] The microdosing principle, as described herein is performed on a human intestine, wherein the microdosing test device is loaded, via a carrier, with the IP such as by using a filter paper in physiological concentration but microdosed in total mass of IP per bodyweight mass (typically 10'6of therapeutic concentration).

[0049] Microdosing allows the use of negligible doses of IP than will not results in significant toxic effects of the mucosal surface or the test person. If injury or other priming of the mucosa is desired [related to the disease / disease mechanism being investigated], injury can be applied to the area to be microdosed immediately before microdosing,n’12. The microdosing test device of the present invention is attached to the stoma, e.g. an ileostomy or a colostomy, of an otherwise healthy subject, or a person with the disorder that is to be investigated.

[0050] The microdosing test device is left in situ until it spontaneously falls off after 4-5 days. If considered necessary, microdosing test device may be removed during the test period.

[0051] Outcome is measured by micro-biopsy of the area across the microdosing test device [after opening the microdosing compartment with a pair of scissors] after the desired exposure time (typically 4 hours - 4 days). Biopsy is performed with standard micro-endoscopic biopsy forceps,n.

[0052] Alternatively, outcome is measured by microcannula sampling.

[0053] MOA outcome as described above could be protein chemistry (e.g. IHC / WB / ELISA) / proteomics responses, specific RNA / bulk mRNAseq responses, single cell RNAseq responses, digital spatial profiling responses, microbiota responses or responses assessed by any other molecular biochemistry method,n’12.

[0054] BA outcomes could be assessment of specific signaling pathways known to be activated by IP or measurement of mucosal IP concentration. Typically, outcome is measured by high- resolution imaging of the dosed compartment,n’13.

[0055] An advantage of using a human stoma is that it, like the rest of the human intestine, the stoma is numb to biopsy / cannulation.

[0056] Further embodiments of the process are described in the experimental section herein, and each individual process as well as each starting material constitutes embodiments that may form part of embodiments.

[0057] The above embodiments should be seen as referring to any one of the aspects (such as ‘microdosing test device, ‘Use of the microdosing test device’, or ‘method for assessment of PK / PD’) described herein as well as any one of the embodiments described herein unless it is specified that an embodiment relates to a certain aspect or aspects of the present invention.

[0058] All references, including publications, patent applications and patents, cited herein are hereby incorporated by reference to the same extent as if each reference was individually and specifically indicated to be incorporated by reference and was set forth in its entirety herein. All headings and sub-headings are used herein for convenience only and should not be construed as limiting the invention in any way.

[0059] Any combination of the above-described elements in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or otherwise clearly contradicted by context.

[0060] The terms “a” and “an” and “the” and similar referents as used in the context of describing the invention are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. Thus, the terms “a” and “an” means “one or more” or “at least one” and are used interchangeably during the text.

[0061] The term “and / or” as used herein is intended to mean both alternatives as well as each of the alternatives individually. For instance, the expression “xxx and / or yyy” means “xxx and yyy”; “xxx”; or “yyy”, all three alternatives are subject to individual embodiments.

[0062] Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein. Unless otherwise stated, all exact values provided herein are representative of corresponding approximate values (e.g., all exact exemplary values provided with respect to a particular factor or measurement can be considered to also pro-vide a corresponding approximate measurement, modified by “about,” where appropriate).

[0063] All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context.

[0064] The use of any and all examples, or exemplary language (e.g., “such as”, “typically, “preferably”) provided herein, is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention unless otherwise indicated. No language in the specification should be construed as indicating any element is essential to the practice of the invention unless as much is explicitly stated.

[0065] The citation and incorporation of patent documents herein is done for convenience only and does not reflect any view of the validity, patentability and / or enforceability of such patent documents. The description herein of any aspect or embodiment of the invention using terms such as “comprising”, “having”, “including” or “containing” with reference to an element or elements is intended to provide support for a similar aspect or embodiment of the invention that “consists of’, “consists essentially of’, or “substantially comprises” that particular element or elements, unless otherwise stated or clearly contradicted by context (e.g., a composition described herein as comprising a particular element should be understood as also describing a composition consisting of that element, unless otherwise stated or clearly contradicted by context).

[0066] This invention includes all modifications and equivalents of the subject matter recited in the aspects or claims presented herein to the maximum extent permitted by applicable law.

[0067] The present invention is further illustrated by the following examples that, however, are not to be construed as limiting the scope of protection. The features disclosed in the foregoing description and in the following examples may, both separately and in any combination thereof, be material for realizing the invention in diverse forms thereof.

[0068] METHODS

[0069] Microdosing compartment

[0070] The microdosing test device having the microdosing compartment has been tried on newly removed surgical specimens from humans undergoing intestinal resection. The newly removed piece of intestine (the specimen) was opened by surgical scissors to reveal the mucosal surface of the intestine. The surface of the exposed mucosa was at least 8 ^ 10 cm. The microdosing compartment was assembled from 18 mm diameter circular sheet of PU film (Tegaderm™ Transparent Film, 3M, St. Paul, MN, USA) mounted with a quadratic 8 x 8 mm clinical grade cellulose-based filter paper (the microdosing compartment). Outside the microdosing compartment a border of 90% butyl and 10% octyl CA adhesive border (LiquiBand, Advanced Medical Solutions, Winsford, UK) was applied to the PU film on the side facing the mucosa surface of the opened human intestine specimen by a pipette. Sterile saline (isotonic NaCl 0.9%; Safecare Nordic, Jyllinge, Denmark) was added with or without indigo carmine (8 mg / mL) before the placement of the device unto the mucosal surface of the surgical specimen. The compartment devise was left on the mucosal surface for up to 30 min. No visible leakage of the fluid was observed.

[0071] Readout methods

[0072] Sampling from intestinal mucosa of a restricted area (7.0 x 1.5 mm) by forceps (Radial Jaw 4; Boston Scientific, Marlborough, MA, USA) through the devise or after removal of device can be done at any timepoint desired. The forceps biopsy can be taken without help of other instruments if the device is placed on a stoma or via the working channel of an endoscope if placed inside the intestine. A sample obtained this way is tested by:

[0073] • 16S rRNA gene sequencing to describe the mucosa-near microbiota of the compartment.

[0074] • Bulk RNA sequencing to describe the host transcriptome of the compartment.

[0075] • Quantitative reverse-transcription polymerase chain reaction RNA (qRT-PCR) of the expression of a specific gene or part of the genome.

[0076] • Histological scoring using clinical inflammatory scoring systems (e.g. Geboes score).14

[0077] • Multiplex protein expression to assess specific proteins regulated. Immunohistochemistry to assess surface markers or protein expression.

[0078] • NMR to assess metabolome profiles.

[0079] All these methods have been applied previously to limited area manipulation of the human intestine by us using a different model developed internally, the micro-injury model.11'13

[0080] The mucosal surface can be described according to a macroscopic inflammation scoring system developed to assess inflammation of small areas using endoscopic high-definition imaging.11The mucosal surface can be primed by a micro-injury11prior to applying the microdosing compartment device.

[0081] In the normal human intestine, a micro-injury was done to measure kinetics of epithelial differentiation after an intestinal compartment with micro-injury. The transcription factor caudal type homeobox 2 (CDX2) is a homebox protein specifically expressed in differentiating epithelial cells, where it is a master regulator of differentiation. We measured expression of this gene in the micro-compartment with micro-injury over time to determine the abundance of epithelial cell differentiation after an injury and during regeneration as an example of how epithelial cell differentiation is regulated in regenerating tissue in the human.

[0082] Twelve healthy subjects without gastrointestinal symptoms were recruited for the study. Six of the twelwe subjects had a micro-compartment with micro-injury was placed approximately 15 cm from the anocutaneous junction with a Radial Jaw 4 forceps. The injury response was documented by endoscopic high-definition image recording after 5, 24 and 48 h. At the same timepoints biopsies were taken from the micro-injury compartment. The other six subjects only had a single tissue sample taken without follup-samples. The tissue samples were minced and lysed with RP1 lysis buffer (Macherey -Nagel, Duren, Germany). Total RNA from isolated from tissue samples were isolated and quantitative reverse-transcription polymerase chain reaction qRT-PCR was performed.15CDX2 mRNA expression were calculated by their ratios to the general human intestinal epithelial marker cytokeratin 20 (CK20), which was amplified in parallel reactions. Sequences of the primers have been used before16and are listed below.

[0083] Distinct phases in the tissue regeneration here defined by expression of the epithelial differentiation marker CDX2 were recognized (Figure 4). CDX2 was decreased significantly (p<0.05, Mann-Whitney test) to 57% and 62% of the initial level after 5 and 24 h, respectively, and returned to the initial level after 48 h. Figure 4 shows regulation of enterocyte differentiation marker CDX2 at experimental microinjury compartment of the human intestine determined by PCR. Normalised to expression of the general enterocyte marker CK20. Sampling was done at timepoint 0, 5h, 24h, and 48h. * p<0.05.

[0084] The experiment shows how intestinal micro-compartments can be used to follow physiological mechanisms over time, in this case regeneration after an injury, where an initial phase of epithelial de-differentiation and, later, re-differentiation was found.

[0085] REFERENCES

[0086] 1. Sun D, Gao W, Hu H, et al. Why 90% of clinical drug development fails and how to improve it? Acta Pharm Sin B 2022;12:3049-3062.

[0087] 2. Ocana A, Garcia-Alonso S, Amir E, et al. Refining Early Antitumoral Drug Development. Trends Pharmacol Sci 2018;39:922-925.

[0088] 3. de Souza HSP, Fiocchi C, Iliopoulos D. The IBD interactome: an integrated view of aetiology, pathogenesis and therapy. Nat. Rev. Gastroenterol. Hepatol 2017;14:739-749.

[0089] 4. Walton KD, Mishkind D, Riddle MR, et al. Blueprint for an intestinal villus: Speciesspecific assembly required. Wiley. Interdiscip. Rev. Dev. Biol 2018.

[0090] 5. Collins AM, Jackson KJL. On being the right size: antibody repertoire formation in the mouse and human. Immunogenetics 2018;70: 143-158.

[0091] 6. Godec J, Tan Y, Liberzon A, et al. Compendium of Immune Signatures Identifies Conserved and Species-Specific Biology in Response to Inflammation. Immunity 2016;44: 194-206.

[0092] 7. Masopust D, Sivula CP, Jameson SC. Of Mice, Dirty Mice, and Men: Using Mice To Understand Human Immunology. J. Immunol 2017;199:383-388.

[0093] 8. Nguyen LP, Pan J, Dinh TT, et al. Role and species-specific expression of colon T cell homing receptor GPR15 in colitis. Nat. Immunol 2015;16:207-213.

[0094] 9. Wu J, Yuk H, Sarrafian TL, et al. An off-the-shelf bioadhesive patch for sutureless repair of gastrointestinal defects. Sci Transl Med 2022;14:eabh2857.

[0095] 10. Dhandapani V, Ringuette V, Desrochers M, et al. Composition, host responses and clinical applications of bioadhesives. J Biomed Mater Res B Appl Biomater 2022;110:2779-2797.

[0096] 11. Seidelin JB, Bahl MI, Licht TR, et al. Acute Experimental Barrier Injury Triggers Ulcerative Colitis-Specific Innate Hyperresponsiveness and Ulcerative Colitis-Type Microbiome Changes in Humans. Cell Mol Gastroenterol Hepatol 2021;12: 1281-1296. Bjerrum JT, Wang Y, Zhang J, et al. Lipidomic Trajectories Characterize Delayed Mucosal Wound Healing in Quiescent Ulcerative Colitis and Identify Potential Novel Therapeutic Targets. Int J Biol Sci 2022;18: 1813-1828. Seidelin JB, Larsen S, Linnemann D, et al. Cellular inhibitor of apoptosis protein 2 controls human colonic epithelial restitution, migration, and Rael activation. Am J

[0097] Physiol Gastrointest Liver Physiol 2015;308:G92-9. Geboes K, Riddell R, Ost A, et al. A reproducible grading scale for histological assessment of inflammation in ulcerative colitis. Gut 2000;47:404-409. Bergenheim F, Seidelin JB, Pedersen MT, et al. Fluorescence-based tracing of transplanted intestinal epithelial cells using confocal laser endomicroscopy. Stem Cell

[0098] Res. Ther 2019;10: 148. Coskun M, Olsen AK, Holm TL, et al. TNF-alpha-induced down-regulation of CDX2 suppresses MEP1A expression in colitis. Biochim. Biophys. Acta 2012;1822:843-851.

Claims

WE CLAIM:

1. A microdosing test device comprising:1) a layer impermeable to aqueous liquids wherein the layer comprises a biocompatible and biodegradable adhesive for a mucosal surface of an animal, preferably a human, and2) a carrier adapted for bringing a test compound in a non-therapeutic concentration in physical contact with the mucosal surface; wherein the layer when applied to the mucosal surface forms a microdosing compartment between the layer and the mucosal surface which compartment contains the carrier.

2. The test device of claim 1 wherein the layer is made of an elastic and / or flexible polymer.

3. The test device of claim 2 wherein the polymer is selected from the group consisting of a polyurethane (PU), a polyethylene (PE), a gas permeable polydimethylsiloxane (PDMS; silicone), a polycarbonate (PC), a polymethylmethacrylate (PMMA), and a polyvinylchloride (PVC).

4. The test device of any one of claims 1-3 wherein the carrier is a liquid, gel, or a solid carrier, preferably a cellulose carrier.

5. The test device of any one of claims 1-4 wherein the adhesive is selected from a group consisting of a polyethylene glycol (PEG) based adhesive, a cyanoacrylates (CA) based adhesive, a thrombine / fibrin-based adhesive, a collagen / gelatin based adhesive, a polysaccharide based adhesive, such as chitosan (CHI), poly(lactic-co-glycolic) acid (PLGA), and a polyurethane based adhesives; as well as combinations hereof.

6. The test device of any one of claims 1-5 wherein the test compound is loaded into or on the carrier.

7. The test device of any one of claims 1-6 wherein the concentration of the test compound in physical contact with the mucosa is non-toxic to the animal, preferably the human.

8. The test device of claim 6 or 7 wherein the concentration of the test compound in physical contact with the mucosa is at least 1 : 1000 of a therapeutic concentration to the animal, preferably the human.

9. The test device of any one of claims 1-8 wherein the mucosa is a gastrointestinal tract of the animal, preferably the human.

10. The test device of claim 9 wherein the mucosa is an oral cavity, stomach, small, or large intestine, a stoma, an ileostomy or a colostomy.

11. The test device of any one of claims 1-10 wherein the layer has a surface area of at least 50 mm2, a circumference of from 25-63 mm and a diameter of from 8-20 mm and is made of a water impermeable polymer.

12. The test device of claim 11 wherein the layer has a border applied to the circumference of the layer, which border is 3-5 mm wide and 0.4-0.6 mm high and the adhesive is applied to the border.

13. The test device of any one of claims 1-12 wherein the test compound is investigated for a gastrointestinal disease.

14. The test device of any one of claims 1-12 wherein the test compound is for oral administration and is investigated for a non-gastrointestinal disease.

15. A method for assessment of PK / PD, such as bioavailability and / or mechanism of action of a test compound in an animal comprising a) applying the test device of any one of claims 6-15 toa mucosal surface of an animal, b) allowing exposure of the mucosal surface to the test compound for sufficient time, c) obtaining a biopsy or micropipette / syringe sample from the mucosa, d) analyzing a body surface fluid from the compartment, e) analyzing the biopsy or micropipette / syringe sample to obtain data on PK / PD of the test compound.

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