EP-4114172-B1 - RODENT MODEL FOR HYPERKERATOSIS AND SEPSIS

Inventors

• Becker-Pauly, Christoph
• PETERS, FLORIAN

Dates

Publication Date

20260506

Application Date

20210226

Claims (10)

1. Rodent that is genetically manipulated to develop the phenotype of sepsis and hyperkeratosis upon activation by an external stimulus, which rodent is genetically manipulated to contain an additional expression cassette encoding meprin α under the control of a promoter which upon presence of the external stimulus is activated at least or only in keratinocytes to express meprin α.
2. Rodent according to claim 1, characterized in that the expression cassette encoding meprin α contains a promoter that is cell-type specific for keratinocytes.
3. Rodent according to one of the preceding claims, characterized in that the coding sequence for meprin α is separated from its promoter by an intermediate stop element which is flanked by recognition sites for a recombinase and in that the mammal contains an expression cassette for the cognate recombinase under the control of a promoter which is induced by the presence of the external stimulus.
4. Rodent according to one of the preceding claims, characterized in that it is in addition genetically manipulated to contain an expression cassette encoding a peptidase.
5. Rodent according to one of the preceding claims, characterized by the phenotype being without administration of toxins or other pro-inflammatory stimuli and without mechanically destroying epithelia or tissues of the rodent.
6. Process for testing chemical compounds, comprising providing a rodent according to one of the preceding claims in which meprin α is expressed in keratinocytes, administering the external stimulus to the rodent for generating a phenotype of sepsis and of hyperkeratosis, and administering at least one chemical compound to the rodent.
7. Process according to claim 6, characterized in that no additional toxin or proinflammatory compound is administered to the rodent and no mechanical damage is inflicted upon the rodent.
8. Process for generating a rodent which develops a phenotype of sepsis and of hyperkeratosis , characterized by introducing into the rodent a genetic manipulation that results in the presence of a promoter which is inducible by presence of an external stimulus, which promoter upon presence of the external stimulus is activated at least or only in keratinocytes and which stimulus induces overexpression of meprin α.
9. Process for generating a phenotype of sepsis and/or of hyperkeratosis and/or of systemic inflammation in a rodent, comprising the steps of providing a rodent according to one of claims 1 to 5, and administering the external stimulus to the rodent.
10. Process according to claim 9, characterized in that it is without administration of any additional toxin or other pro-inflammatory stimuli, and without mechanically destroying epithelia or tissues of the rodent.

Description

The present invention relates to a rodent model for hyperkeratosis and/or for sepsis and/or for systemic inflammation, which rodent has the advantage of developing hyperkeratosis and sepsis upon administration of an external stimulus at an age of the rodent that can be chosen freely. The rodent is e.g. for use in research, especially for use in processes for testing pharmaceutical compounds in respect of an effect on hyperkeratosis and/or on sepsis and/or on systemic inflammation. The animal is a rodent, e.g. a mouse, rat, rabbit, guinea pig. Generally herein, the non-human mammal is also simply referred to as animal. In the context of the invention, the animal is a rodent. The animal has the advantage that the sepsis and/or systemic inflammation is not elicited against an antigen, and hence the phenotype of sepsis and/or of systemic inflammation is not antigen-specific. Accordingly, the invention also relates to processes for testing pharmaceutical compounds, e.g. for use in screening test compounds, for their effect on hyperkeratosis and/or on sepsis and/or on systemic inflammation as induced in an animal model as provided herein. The rodent of the invention is genetically manipulated to develop the phenotype of hyperkeratosis and sepsis upon activation by an external stimulus, which preferably is a synthetic compound. Accordingly, the invention also relates to a process for generating the animal model. The rodent of the invention has the advantage of reproducibly generating a phenotype of hyperkeratosis and of sepsis without administration of toxins or other pro-inflammatory stimuli and without mechanically destroying epithelia or tissues of the rodent. State of the art Stortz et al., ILAR Journal 90-105 (2017), review the advantages of using mice as animal models and describe murine sepsis models which are generated by exogenous administration of toxin or of a viable pathogen, or by puncture of intestines. Holly et al., Kidney International 496-506 (2006), describe that upon induction of sepsis by puncture of intestines in rats, which resulted in acute kidney failure, changes of the concentration of a number of proteins in urine were found, the proteins including albumin, brush-border enzymes (e.g. meprin-1-alpha), and serine protease inhibitors. Meprin-1-alpha is mentioned as a potential biomarker and drug target. Wang et al., SHOCK 141-147 (2011), describe that actinonin, a meprin-1-A inhibitor, should be evaluated as a therapeutic agent in targeting early and later organ-damaging effects of sepsis that was induced by puncturing the intestine subsequent to administration of actinonin. Herzog et al. in the review article Cytokine 18-25 (2019) quote that meprin α-KO mice upon sepsis induced by LPS showed reduction in pro-inflammatory cytokines IL-1β and TNF-α and exhibited less bladder edema, leukocyte infiltration and bladder permeability than wt mice. Herzog et al. conclude that meprin A contributes to renal and urogenital pathogenesis in LPS model of sepsis. Herzog et al. further quote that meprin α was identified as a susceptibility gene for inflammatory bowel disease in ulcerative colitis patients, and quote that meprin α-KO mice were more susceptible to DSS-induced experimental colitis and displayed more colon damage and inflammation than wt mice. Herzog et al. only quote detrimental effects of knock-out of meprin α, and Herzog et al. do not relate to possible effects of inducing expression of meprin α. Lai et al., Arthritis & Rheumatism 1184-1197 (2006) describe a mouse model that was genetically manipulated to allow conditional overexpression of IL-1β. Becker-Pauly et al., Journal of Investigative Dermatology 1115-1125 (2007) in 5 patients diagnosed for Ichthyosis vulgaris, a disease model of retention hyperkeratosis, mentions high levels of meprin α. However, cultivated keratinocytes (HaCaT) were not influenced by meprin α. It is mentioned that meprin α may play a role in colorectal cancer and in melanoma formation or in wound healing. Broder and Becker-Pauly, Biochem. Journal, 253-264 (2014) relates to meprin α-KO mice in relation to kidney damage and describes release of meprin α upon activation of TGF-α and EGF, leading to inflammatory effects, and describes an increase of melanoma metastasis. Meprin α is said to be overexpressed in skin fibroblasts of fibrotic tumours. An effect of induced expression of meprin α in a genetically manipulated experimental animal cannot be inferred from the prior publications. Object of the invention It is an object of the invention to provide an animal model, which generates hyperkeratosis and sepsis upon induction of an external stimulus, which stimulus does not require the administration of a toxin nor of a pro-inflammatory molecule to the animal, nor mechanically damaging the animal model, e.g. no puncturing or other damaging of intestines. Preferably, the animal model is genetically manipulated such that administration of an external stimulus, which is a