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WO2023218095A1 - Agents de stimulation de régéneration tissulaire - Google Patents

Agents de stimulation de régéneration tissulaire Download PDF

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Publication number
WO2023218095A1
WO2023218095A1 PCT/EP2023/062965 EP2023062965W WO2023218095A1 WO 2023218095 A1 WO2023218095 A1 WO 2023218095A1 EP 2023062965 W EP2023062965 W EP 2023062965W WO 2023218095 A1 WO2023218095 A1 WO 2023218095A1
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WIPO (PCT)
Prior art keywords
seq
agent
use according
skin
wound
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English (en)
Inventor
Anna HULTGÅRDH NILSSON
Jan Alenfall
Jan Nilsson
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Coegin Pharma AB
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Coegin Pharma AB
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism

Definitions

  • the present invention relates to agents for the regeneration of tissue.
  • the regenerated tissue is preferably skin, i.e. the regeneration of tissue is preferably the healing of wounds.
  • Said wounds can be acute or pathological wounds.
  • Advanced wound care is a fast-growing market mainly composed of 4 main categories: dressings, wound cleansers, negative pressure wound therapy devices and biologies. It is estimated that 1 % to 2 % of the population will experience a chronic wound during their lifetime in developed countries (Gottrup) and this rate is increasing year after year. Stalled wounds may persist for months, even years (Kirker et al., Frykberg et al.) before healing. Hence, non-healing/hard-to-heal wounds represent an economic burden for health services and a large medical problem for patients, representing a global cost of $143 bn.
  • Treatment for acute wounds include debridement, decontamination, moist wound healing, and early closure.
  • Treatment for chronic wounds include debridement, anti-inflammation therapy, antibiotic therapy, synthetic substitutes, modulating cytokines and/or matrix metalloproteinases (MMPs), or recruiting stem cells.
  • MMPs matrix metalloproteinases
  • Wounds may arise from a variety of aetiologies such as peripheral arterial disease, venous hypertension, neuropathy (e.g., diabetes mellitus), pressure (in patients with neuropathy or decreased mobility), vasculitis and burns, etc and are very often hard to heal.
  • a large proportion of acute wounds heal with appropriate management, but some wounds fail to heal and proceed to becoming chronic, non-healing wounds.
  • a ‘completely healed wound’ can be defined as one that has returned to its normal anatomic structure, function, and appearance within a reasonable period of time (normally in about four to six weeks). Any wound that has not reached the above state would be considered a chronic, non-healing wound.
  • Wound care has become increasingly important given the rise of chronic wounds and the morbidity associated with them.
  • An important concept in wound care is the role of debridement, or the removal of non-viable tissue material. Disruption at one or more points in the distinct phases of wound healing (haemostasis, inflammation, proliferation, or remodelling) can cause a wound not to heal.
  • An alternation in the balance between various cytokines, growth factors, protease activity, matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) is observed in chronic wounds.
  • alteration in the morphology and proliferation rate of fibroblasts, keratinocyte activity, and accumulation of oxygen derived free radicals, necrotic tissue and underlying disease all play a role in inhibiting wound healing.
  • the present inventors have identified agents with use in healing wounds.
  • One aspect of the disclosure provides for an agent comprising: a) a peptide selected from the group consisting of:
  • X 5 is D or G
  • X14 is E or G
  • X15 is S or T
  • VDTYDGDISVVYGL SEQ ID NO: 34
  • VDTYDGDISVVYG SEQ ID NO: 35
  • VDTYDGDISVVY SEQ ID NO: 36
  • VDTYDGDISVV SEQ ID NO: 37
  • VDTYDGDISV SEQ ID NO: 38
  • VDTYDGDIS SEQ ID NO: 39
  • VDTYDGRGDSVVYGLR SEQ ID NO: 40
  • VDVPNGDISLAYGL SEQ ID NO: 41
  • VDVPNGDISLAYG SEQ ID NO: 42
  • VDVPNGDISLA SEQ ID NO: 43
  • VDVPNGDIS SEQ ID NO: 44
  • GDPNDGRGDSVVYGLR SEQ ID NO: 45
  • LDGLVRAYDNISPVG SEQ ID NO: 46
  • GDPNGDISVYGLR SEQ ID NO: 47
  • GDPNGDISVYGLR SEQ ID NO
  • the present invention relates to a method for treatment and/or regeneration of a tissue defect in a subject, the method comprising administering a therapeutically effective amount of an agent as defined herein to an subject in need thereof.
  • the present invention relates to a method for treating and/or promoting healing of a tissue defect in a subject, the method comprising administering a therapeutically effective amount of an agent as defined herein to an subject in need thereof.
  • the present invention relates to a method for regeneration of skin at the site of a skin defect and/or for reducing scar formation resulting from healing of a skin defect in a subject, said method comprising administering an agent as defined herein.
  • the present invention relates to use of an agent as defined herein for the manufacture of a medicament for the treatment and/or regeneration of a skin defect in a subject.
  • the present invention relates to use of an agent defined herein for the manufacture of a medicament for the treating and/or promoting healing of a tissue defect in a subject.
  • the present invention relates to use of an agent defined herein for the manufacture of a medicament for the treatment of a skin defect in a subject.
  • the present invention relates to use of an agent defined herein for reducing scar formation resulting from healing of a skin defect in a subject.
  • the present invention relates to use of an agent defined herein for promoting the healing of a skin defect in a subject.
  • the present invention relates to use of an agent defined in herein for promoting vascularisation in a healing tissue, such as skin healing from a skin defect, such as a wound.
  • the present invention relates to use of an agent defined herein in combination with a tissue meshing technique, for promoting healing of a skin defect, such as a wound.
  • the present invention relates to use of an agent as defined herein for promoting grafting of skin.
  • the present invention relates to a method for inserting a stent in a subject, said method comprising administering an agent as defined herein.
  • the present invention relates to a medical device comprising the agent as defined herein.
  • FIG. 1 FOL26 stimulates endothelial and vascular smooth muscle cell proliferation.
  • Human umbilical cord endothelial cells (A) and human coronary artery smooth muscle cells (B) were treated with FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM, and DNA synthesis analyzed by measuring the uptake of BrdU.
  • FIG. 2 FOL26 enhances endothelial cell tube formation and expression of PECAM-1.
  • Human umbilical cord endothelial cells were treated with FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM, and endothelial tube formation and gene expression of PECAM-1 (CD31) analyzed.
  • FOL26 increased tube formation compared to control.
  • the expression of PECAM-1 was increased for 100 nM and 1000 nM FOL26.
  • FIG. 3 FOL26 increases endothelial cell migration in a cell culture scratch wound assay. Confluent cultures of human umbilical cord endothelial cells were treated with FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM, and ability of the cells to migrate into an in vitro scratch injury was analyzed with image J software. Migration of cells increased with increasing concentration of FOL26.
  • FIG. 4 FOL26 increases arterial smooth muscle cell migration in a cell culture scratch wound assay.
  • PCR polymerase chain reaction
  • FOL26 increased expression of both COL1 A1 and COL1 A2.
  • the wound closure at 5h was increased for FOL26 at 10 nM and 100 nM, while the 1000 nM also trended towards increased closure compared to control.
  • FIG. 5 FOL26 inhibits endothelial and arterial smooth muscle cell apoptosis.
  • Cultured human umbilical cord endothelial and coronary artery smooth cells were exposed to soluble Fas ligand (sFasL) to induce apoptosis that subsequently was assessed by determining activation of caspase 3 and 7.
  • Cells were also treated with FOL 26 in concentrations of 10 nM, 100 nM, and 1000 nM to determine the effect on apoptosis activated by sFasL.
  • FOL26 was capable of reversing the sFasL-induced apoptosis, to the highest extent at the 100 nM concentration.
  • FOL26 affects the gene expression of VEGF and the VEGF receptor family in endothelial cells.
  • Human umbilical cord endothelial cells were treated with FOL 26 in concentrations of 10 nM, 100 nM, and 1000 nM and the expression of the genes encoding c-Met, neuropilin-1 (NRP-1), VEGF-A and VEGF receptor-2 (VEGFR-2) determined by polymerase chain reaction (PCR).
  • FOL026 was reducing the c-Met expression, in the lowest dose reducing however in the 100nM and 1000nM doses were inducing NRP-1 expression.
  • FOL026 had no effect on VEGF-A expression, however in 10nM it reduced and in 1000nM it induced the expression of VEGFR-2 levels.
  • FOL26 affects the gene expression of cytokines and matrix metalloproteinases in endothelial cells.
  • Human umbilical cord endothelial cells were treated with FOL 26 in concentrations of 10 nM, 100 nM, and 1000 nM and the expression of the genes encoding TNF-a, IL-6, MMP2 and MMP3 determined by polymerase chain reaction (PCR).
  • TNF-a is strong pro-inflammatory cytokine while MMP2 is required for activation of smooth muscle cell proliferation and migration.
  • FOL026 reduced the TNF-a expression in 10nM and 100nM doses, had no effect on IL-6 or MMP3 expression, however, MMP2 expression was induced with 200nM and 1000nM doses.
  • FOL26 affects the gene expression of VEGF and the VEGF receptor family in arterial smooth muscle cells.
  • Human coronary artery smooth cells were treated with FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM and the expression of the genes encoding c-Met, neuropilin-1 (NRP-1), VEGF-A and VEGF receptor-2 (VEGFR- 2) determined by polymerase chain reaction (PCR).
  • FOL026 reduced the c-MET expression in 10nM and 1000nM doses, as well as the NRP-1 expression in 10, 100, and 1000nM doses.
  • FOL26 affects the gene expression of cytokines and matrix metalloproteinases in arterial smooth muscle cells.
  • Human coronary artery smooth cells were treated with FOL 26 in concentrations of 10 nM, 100 nM, and 1000 nM and the expression of the genes encoding TNF-a, IL-6, MMP2 and MMP3 determined by polymerase chain reaction (PCR).
  • TNF-a is strong pro-inflammatory cytokine while MMP2 is required for activation of smooth muscle cell proliferation and migration.
  • FOL026 100 and 1000nM induced the TNF-a, IL-6 and MMP3 expression, furthermore, FOL026 induced in 10, 100, and 1000nM doses the MMP2 expression.
  • FIG. 10 FOL26 stimulation of endothelial cell proliferation is NRP-1 dependent.
  • Human umbilical cord endothelial cells were first treated with a scramble non-coding siRNA (NC) or siRNA for NRP-1, subsequently they were treated with FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM, and DNA synthesis analyzed by measuring the uptake of BrdU.
  • NC scramble non-coding siRNA
  • FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM
  • DNA synthesis analyzed by measuring the uptake of BrdU.
  • An increase in O.D. (450 nm) for the FOL 26 treated cell lines compared to the control showed that FOL26 induced angiogenesis may be NRP-1 dependent as a reduction was observed for FOL26 10 and 100nM.
  • Figure 11 Human umbilical cord endothelial cells were treated with a scramble noncoding siRNA (NC) or siRNA for NRP-1 in concentrations of 1nM, and 5nM, and the NRP-1 expression values and Glyceraldehyde-phosphate dehydrogenase (GAPDH) mRNA expression values was evaluated, the latter was used as an internal control to normalize the data.
  • NC scramble noncoding siRNA
  • GPDH Glyceraldehyde-phosphate dehydrogenase
  • Figure 12 FOL26 enhances endothelial cell tube in an NRP-1 specific manner.
  • Human umbilical cord endothelial cells were first treated with a scramble non-coding siRNA (NC) or siRNA for NRP-1, subsequently treated with FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM, and endothelial tube formation and the total master segemnts were measured using Imaged, gene expression of PECAM-1 (CD31) analyzed.
  • FOL26 increased tube formation compared to control in a NRP-1 dependent way. Scale bar: 150 pm.
  • Figure 13 Overview of acute wound healing and pathological wound healing models, including experimental parameters.
  • A) Acute wound healing - one way ANOVA and Dunnett’s multiple comparison test vs. vehicle n.s. Unpaired t test * ⁇ 0.05.
  • C) Pathological wound healing - One way ANOVA p 0.143, Dunnett’s multiple comparison test vs vehicle #p ⁇ 0.05. Unpaired t test *p ⁇ 0.05, **p ⁇ 0.01.
  • Figure 15 Representative images of wound tongue at day 3, acute model (donor 1). PAS stained skin sections. Dashed lines show the newly formed epithelial tongues. Scale bar represents 100 pm.
  • Figure 16 Representative images of wound tongue at day 3, pathological model (donor 2). PAS stained skin sections. Dashed lines show the newly formed epithelial tongues. Scale bar represents 100 pm.
  • Figure 17 Increased proliferation of epidermal keratinocytes adjacent to the wound bed and decreased keratinocyte apoptosis at 300 nM FOL-005 peptide in the ex vivo pathological wound healing model (6 day protocol, 3 topical applications of FOL peptide).
  • FIG. 18 FOL56 stimulates endothelial and vascular smooth muscle cell proliferation.
  • Human umbilical cord endothelial cells (A) and human coronary artery smooth muscle cells (B) were treated with FOL 56 in concentrations of 10 nM, 100 nM, and 1000 nM and DNA synthesis analyzed by measuring the uptake of BrdU.
  • FOL56 increases proliferation at all concentrations in both HUVECs and HCASMCs compared to the control.
  • FIG. 19 FOL56 enhances endothelial cell tube formation and expression of PECAM- 1.
  • Human umbilical cord endothelial cells were treated with FOL 56 in concentrations of 10 nM, 100 nM, and 1000 nM and endothelial tube formation and gene expression of PECAM-1 (CD31) analyzed.
  • FOL56 increased tube formation at all concentrations compared to control, and also increased expression of PECAM-1.
  • FIG. 20 FOL56 increases endothelial cell migration in a cell culture wound model. Confluent cultures of human umbilical cord endothelial cells were treated FOL 56 in concentrations of 10 nM, 100 nM, and 1000 nM and ability of the cells to migrate into an in vitro scratch injury was analyzed with image J software. FOL56 increased cell migration at all concentrations compared to the control.
  • FIG. 21 FOL56 increases arterial smooth muscle cell migration in a cell culture wound model.
  • Confluent cultures of human coronary artery smooth cells were treated with FOL 56 in concentrations of 10 nM, 100 nM, and 1000 nM and ability of the cells to migrate into an in vitro scratch injury was analyzed with image J software. Expression of the collagen gene a1 and a2 chains were determined by polymerase chain reaction (PCR).
  • FOL56 increased expression of both COL1A1 and COL1A2. The wound closure at 5h was increased for FOL56 at all concentrations compared to the control.
  • FIG. 22 FOL56 inhibits endothelial and arterial smooth muscle cell apoptosis.
  • Cultured human umbilical cord endothelial and coronary artery smooth cells were exposed to soluble Fas ligand (sFasL) to induce apoptosis that subsequently was assessed by determining activation of caspase 3 and 7.
  • Cells were also treated with FOL 56 in concentrations of 10 nM, 100 nM, and 1000 nM to determine the effect on apoptosis activated by sFasL.
  • FOL56 was capable of inhibiting, or trended towards inhibiting apoptosis in both HUVECs and HCASMCs.
  • FOL56 affects the gene expression of VEGF and the VEGF receptor family in endothelial cells.
  • Human umbilical cord endothelial cells were treated with FOL 56 in concentrations of 10 nM, 100 nM, and 1000 nM and the expression of the genes encoding c-Met, neuropilin-1 (NRP-1), VEGF-A and VEGF receptor-2 (VEGFR-2) determined by polymerase chain reaction (PCR).
  • FOL56 affects the gene expression of cytokines and matrix metalloproteinases in endothelial cells.
  • Human umbilical cord endothelial cells were treated with FOL 56 in concentrations of 10 nM, 100 nM, and 1000 nM and the expression of the genes encoding TNF-a, IL-6, MMP2 and MMP3 determined by polymerase chain reaction (PCR).
  • TNF-a is strong pro-inflammatory cytokine while MMP2 is required for activation of smooth muscle cell proliferation and migration.
  • FOL56 affects the gene expression of VEGF and the VEGF receptor family in arterial smooth muscle cells.
  • Human coronary artery smooth cells were treated with FOL 56 in concentrations of 10 nM, 100 nM, and 1000 nM and the expression of the genes encoding c-Met, neuropilin-1 (NRP-1), VEGF-A and VEGF receptor-2 (VEGFR- 2) determined by polymerase chain reaction (PCR).
  • FOL56 affects the gene expression of cytokines and matrix metalloproteinases in arterial smooth muscle cells.
  • Human coronary artery smooth cells were treated with FOL 56 in concentrations of 10 nM, 100 nM, and 1000 nM and the expression of the genes encoding TNF-a, IL-6, MMP2 and MMP3 determined by polymerase chain reaction (PCR).
  • TNF-a is strong pro-inflammatory cytokine while MMP2 is required for activation of smooth muscle cell proliferation and migration.
  • FIG. 27 Effect of FOL-026 peptide on endothelial cell function and gene expression.
  • the ROS level (C) activated by 50ug/ml oxidized Low-density Lipoprotein (oxLDL) for 2 hours in cells pre-incubated with FOL-026 peptides was assessed using H2O2 measurement (n 4-8 per group).
  • oxLDL oxidized Low-density Lipoprotein
  • the effect of FOL-026 peptides on gene expression of VEGF-A (F) and its receptors (G and H), as well as HGF receptor c-Met (I) in HUVECs (n 4 for F-l).
  • Scale bar 150um.
  • FIG. 29 Effect of FOL-026 peptide on angiogenesis in vitro and vivo.
  • A Western blot analysis of phosphorylated AKT (pAKT-T308), AKT, phosphorylated ERK1/2 (p-ERK1/2) and ERK1/2 in endothelial cells stimulated with FOL-26 peptides for 48 hours (B).
  • Figure 31 Effect of NRP-1 knock-down on endothelial cell function induced with FOL- 026 peptide.
  • FIG 32 Representative pictures of endothelial cells at 0 and 5 hours after scratch wounding are shown in (A).
  • FIG 33 Effect of NRP-1 knock-down on tube formation induced with FOL-026 peptide.
  • Figure 34 Effect of FOL-026 peptide on smooth muscle cell function.
  • Figure 36 Effect of NRP-1 knock-down on smooth muscle cell function induced with FOL-026 peptide.
  • qRT-PCR analysis of NRP-1 mRNA expression in HCASMCs with small interfering RNA transfection for 48 hours (n 4) (A).
  • FIG 37 Representative images of smooth muscle cells at 0 and 5 hours after scratch wounding are shown in (A).
  • FIG 38 Effect of FOL-005 peptide on endothelial and smooth muscle cell proliferation and wound healing ability.
  • FIG 40 Effect of FOL-005 peptide on tube formation and the role of NRP-1 in that. Representative images of tube formation in human umbilical vascular endothelial cells (HUVECs) treated with FOL-005 peptides are shown in (A and B).
  • HUVECs human umbilical vascular endothelial cells
  • Preventing includes delaying, stopping, reducing the risk of the onset, of disease, disorder, or condition.
  • the term "skin defect" refers to, but is not limited to: injuries or trauma to internal or external tissue, preferably injury or trauma to the epidermis and/or dermis of the skin.
  • the wound may be an acute wound or a chronic wound.
  • an acute wound may be an incision, laceration, abrasion graze or burn, a puncture wound, a penetration wound or a wound due to dermatologic diseases such as psoriasis, acne and eczema.
  • a chronic wound may be a venous ulcer, a diabetic ulcer, a pressure ulcer, corneal ulcer, digestive ulcer or wounds as a result of ischemia and radiation poisoning.
  • treatment and/or regeneration of a skin defect relates to the promotion, the acceleration, and/or the improvement of healing at the wounded site, i.e. the formation of a functional skin at the wounded site.
  • treatment and/or regeneration of a skin defect ideally result in formation or regeneration of dermis and epidermis, including a basal layer, which characterize functional skin.
  • hard-to-heal wound refers to a wound that has not healed. Wounds that do not heal within approximately 4 to 6 weeks, for example, are considered chronic.
  • a "chronic wound” may be a wound that fails to progress through an orderly and timely sequence of repair or a wound that does not respond to treatment and/or the demands of treatment are beyond the patient's physical health, tolerance or stamina. Many wounds that are first considered to be acute wounds ultimately become chronic wounds due to factors still not well understood. One significant factor is the transition of planktonic bacteria within the wound to form a biofilm. For example, a chronic wound may have an epithelial layer that fails to cover the entire surface of the wound and is subject to bacterial colonization, which can result in biofilm formation, which is resistant to treatment with anti-bacterial agents.
  • venous insufficiency venous insufficiency
  • arterial insufficiency venous insufficiency
  • diabetic complications or is a pressure-related.
  • Hard-to-heal wounds due to venous insufficiency account for 70% to 90% of all hard-to-heal wounds and commonly affect the elderly.
  • Venous insufficiency results in venous hypertension, in which blood flow is abrogated resulting in subsequent ischaemia. Venous insufficiency can occur as a result of obstructions to venous outflow or reflux due to valve damage. Following a period of ischaemia, tissue reperfusion can result in reperfusion injury, causing the tissue damage that leads to wound formation.
  • Exemplary chronic wounds can include "burn ulcers", including first degree burn, which may be a superficial, reddened area of skin; second degree burn, which may be a blistered injury site which may heal spontaneously after the blister fluid has been removed; third degree burn, which may be a burn through the entire skin and usually require surgical intervention for wound healing; scalding burns, which may occur from scalding hot water, grease or radiator fluid; thermal burns, which may occur after contact with flames, usually deep burns; chemical burns, which may come from acid and alkali, usually deep burns; electrical burns; and contact burns, which are usually deep and may occur from muffler tail pipes, hot irons and stoves, or other materials.
  • first degree burn which may be a superficial, reddened area of skin
  • second degree burn which may be a blistered injury site which may heal spontaneously after the blister fluid has been removed
  • third degree burn which may be a burn through the entire skin and usually require surgical intervention for wound healing
  • vascular regeneration Angiogenesis and neovascularisation are understood to be important processes of vascular regeneration and may collectively or alternatively be referred to herein as vascular regeneration.
  • vascular regeneration is to be understood in its broadest sense to also include other repair processes including endothelial cell migration, proliferation and mobilisation.
  • wound dressing particularly refers to any material applied to a wound for protection, absorbance, drainage, etc.
  • films e.g., polyurethane films
  • hydrocolloids hydrophilic colloidal particles bound to polyurethane foam
  • hydrogels cross-linked polymers containing about at least 60% water
  • foams hydrophilic or hydrophobic
  • calcium alginates nonwoven composites of fibers from calcium alginate
  • cellophane cellulose with a plasticizer
  • tissue such as in the context of “tissue defect”, is meant epithelial surfaces of epithelial origin.
  • the agent of the present invention may be a peptide; a polynucleotide encoding said peptide; a vector comprising said polynucleotide; or a cell comprising said polynucleotide or vector.
  • the agent is a peptide or a pharmaceutically acceptable salt thereof.
  • amino acid as used herein includes the standard twenty genetically-encoded amino acids and their corresponding stereoisomers in the ‘D’ form (as compared to the natural ‘L’ form), omega-amino acids and other naturally-occurring amino acids, unconventional amino acids (e.g., a,a-disubstituted amino acids, N-alkyl amino acids, etc.) and chemically derivatized amino acids (see below).
  • amino acid when an amino acid is being specifically enumerated, such as ‘alanine’ or ‘Ala’ or ‘A’, the term refers to both L-alanine and D-alanine unless explicitly stated otherwise.
  • Other unconventional amino acids may also be suitable components for peptides of the present disclosure, as long as the desired functional property is retained by the peptide.
  • each encoded amino acid residue where appropriate, is represented by a single letter designation, corresponding to the trivial name of the conventional amino acid.
  • the peptide is non-naturally occurring, such as a peptide comprising non-proteinogenic amino acid residues.
  • the agent comprises or consists of a tandem repeat comprising two or more repeat units.
  • the repeat unit comprises or consists of the amino acid sequence of any one or more of the sequences as described herein.
  • the peptide is cyclic.
  • the cyclic structure may be achieved by any suitable method of synthesis.
  • heterodetic linkages may include, but are not limited to formation via disulphide, cysteine, alkylene or sulphide bridges.
  • the peptide is capable of forming at least one intramolecular cysteine bridge.
  • the agent comprises or consists of a peptide comprising or consisting of an amino acid sequence of the general formula:
  • Xs is D or G
  • Xe is I or G
  • X7 is V or L; and Xa is V or A.
  • the peptide comprises or consists of an amino acid sequence of the general formula:
  • VDX2X3X4GX5X6SX7X8YGLR (SEQ ID NO: 2) wherein:
  • X 2 is T or V
  • X 3 is Y or P
  • X4 is D or N
  • Xs is D or G
  • Xe is I or G
  • X 7 is V or L
  • X 8 is V or A.
  • the peptide comprises or consists of an amino acid sequence of the general formula:
  • VDTYX 4 GX5X6SX 7 X 8 YGLR (SEQ ID NO: 3) wherein:
  • X 4 is D or N
  • X 5 is D or G
  • X 6 is I or G
  • X 7 is V or L
  • X 8 is V or A.
  • the peptide comprises or consists of an amino acid sequence of the general formula:
  • VDTYDGZ 7 Z 8 SZIOZIIYGLR (SEQ ID NO: 4) wherein:
  • X 5 is D or G
  • X 8 is I or G
  • X 7 is V or L
  • X 8 is V or A.
  • the peptide comprises or consists of an amino acid sequence of the general formula:
  • VDTYDGZ 7 Z 8 SVVYGLR (SEQ ID NO: 5) wherein:
  • Xs is D or G
  • X 8 is I or G;
  • the agent comprises or consists of a peptide comprising or consisting of an amino acid sequence of the general formula:
  • X14 is E or G
  • X15 is S or T.
  • the peptide comprises or consists of an amino acid sequence of the general formula:
  • X 9 is C, P or G
  • X is E or G
  • Xu is C, D or I
  • X12 is D, I, S or G
  • X13 is S, D or G
  • X14 is E or G
  • X15 is S or T.
  • the peptide comprises or consists of an amino acid sequence of the general formula:
  • X 9 is C, P or G
  • X10 is E or G
  • Xu is C, I or absent
  • X12 is D, G or absent
  • X13 is S, G or absent
  • X14 is E or G.
  • the peptide comprises or consists of an amino acid sequence of the general formula:
  • X 9 is C, P or G; Xio is E or G; and
  • X14 is E or G.
  • the peptide comprises or consisted of the amino acid sequence IELSYGIK (SEQ ID NO: 109).
  • the peptide comprises or consists of VDTYDGGISVVYGLR (SEQ ID NO: 6). In one embodiment, the peptide comprises or consists of AEIDSIELSYGIK (SEQ ID NO: 110). In one embodiment, the peptide comprises or consists of VDTYDGDISVVYGLR (SEQ ID NO: 7). In one embodiment, the peptide comprises or consists of DTYDGDISVVYGLR (SEQ ID NO: 8). In one embodiment, the peptide comprises or consists of TYDGDISVVYGLRS (SEQ ID NO: 9). In one embodiment, the peptide comprises or consists of TYDGDISVVYGLR (SEQ ID NO: 10).
  • the peptide comprises or consists of YDGDISWYGLRS (SEQ ID NO: 11). In one embodiment, the peptide comprises or consists of YDGDISVVYGLR (SEQ ID NO: 12). In one embodiment, the peptide comprises or consists of DGDISVVYGLRS (SEQ ID NO: 13). In one embodiment, the peptide comprises or consists of DGDISVVYGLR (SEQ ID NO: 14). In one embodiment, the peptide comprises or consists of GDISVVYGLRS (SEQ ID NO: 15). In one embodiment, the peptide comprises or consists of GDISVVYGLR (SEQ ID NO: 16).
  • the peptide comprises or consists of DISVVYGLRS (SEQ ID NO: 17). In one embodiment, the peptide comprises or consists of DISVVYGLR (SEQ ID NO: 18). In one embodiment, the peptide comprises or consists of VDVPNGDISLAYGLR (SEQ ID NO: 19). In one embodiment, the peptide comprises or consists of DVPNGDISLAYGLRS (SEQ ID NO: 20). In one embodiment, the peptide comprises or consists of DVPNGDISLAYGLR (SEQ ID NO: 21). In one embodiment, the peptide comprises or consists of VPNGDISLAYGLRS (SEQ ID NO: 22).
  • the peptide comprises or consists of VPNGDISLAYGLR (SEQ ID NO: 23). In one embodiment, the peptide comprises or consists of PNGDISLAYGLRS (SEQ ID NO: 24). In one embodiment, the peptide comprises or consists of PNGDISLAYGLR (SEQ ID NO: 25). In one embodiment, the peptide comprises or consists of NGDISLAYGLRS (SEQ ID NO: 26). In one embodiment, the peptide comprises or consists of NGDISLAYGLR (SEQ ID NO: 27). In one embodiment, the peptide comprises or consists of GDISLAYGLRS (SEQ ID NO: 28). In one embodiment, the peptide comprises or consists of GDISLAYGLR (SEQ ID NO: 29).
  • the peptide comprises or consists of DISLAYGLRS (SEQ ID NO: 30). In one embodiment, the peptide comprises or consists of DISLAYGLR (SEQ ID NO: 31). In one embodiment, the peptide comprises or consists of VDTYDGDGSVVYGLR (SEQ ID NO: 32). In one embodiment, the peptide comprises or consists of VDVPEGDISLAYGLR (SEQ ID NO: 33). In one embodiment, the peptide comprises or consists of KPLAEIDSIELSYGIK (SEQ ID NO: 111). In one embodiment, the peptide comprises or consists of KCLAECDSIELSYGIK (Cyclic) (SEQ ID NO: 112).
  • the peptide comprises or consists of KPLAEDISIELSYGIK (SEQ ID NO: 113). In one embodiment, the peptide comprises or consists of KPLAEISDIELSYGIK (SEQ ID NO: 114). In one embodiment, the peptide comprises or consists of KPLAEIGDIELSYGIK (SEQ ID NO: 115). In one embodiment, the peptide comprises or consists of KPLAEGDIELSYGIK (SEQ ID NO: 116). In one embodiment, the peptide comprises or consists of KPLAEIELSYGIK (SEQ ID NO: 117). In one embodiment, the peptide comprises or consists of KPLAEIDSIELTYGIK (SEQ ID NO: 118).
  • the peptide comprises or consists of KPLAEIDGIELSYGIK (SEQ ID NO: 119). In one embodiment, the peptide comprises or consists of KPLAEIDGIELTYGIK (SEQ ID NO: 120). In one embodiment, the peptide comprises or consists of KPLAEIGSIELSYGIK (SEQ ID NO: 121). In one embodiment, the peptide comprises or consists of KGLAEIDSIELSYGIK (SEQ ID NO: 122). In one embodiment, the peptide comprises or consists of KPLAGIDSIGLSYGIK (SEQ ID NO: 123). In one embodiment, the peptide comprises or consists of Cyclic KCLAEIDSCELSYGIK (SEQ ID NO: 124).
  • the peptide comprises or consists of LAEIDSIELSYGIK (SEQ ID NO: 125). In one embodiment, the peptide comprises or consists of EIDSIELSYGIK (SEQ ID NO: 126). In one embodiment, the peptide comprises or consists of IDSIELSYGIK (SEQ ID NO: 127). In one embodiment, the peptide comprises or consists of DSIELSYGIK (SEQ ID NO: 128). In one embodiment, the peptide comprises or consists of SIELSYGIK (SEQ ID NO: 129). In one embodiment, the peptide comprises or consists of IELSYGIK (SEQ ID NO: 109).
  • the peptide comprises or consists of VDTYDGDISVVYGL (SEQ ID NO: 34). In one embodiment, the peptide comprises or consists of VDTYDGDISVVYG (SEQ ID NO: 35). In one embodiment, the peptide comprises or consists of VDTYDGDISVVY (SEQ ID NO: 36). In one embodiment, the peptide comprises or consists of VDTYDGDISVV (SEQ ID NO: 37). In one embodiment, the peptide comprises or consists of VDTYDGDISV (SEQ ID NO: 38). In one embodiment, the peptide comprises or consists of VDTYDGDIS (SEQ ID NO: 39).
  • the peptide comprises or consists of VDTYDGRGDSVVYGLR (SEQ ID NO: 40). In one embodiment, the peptide comprises or consists of VDVPNGDISLAYGL (SEQ ID NO: 41). In one embodiment, the peptide comprises or consists of VDVPNGDISLAYG (SEQ ID NO: 42). In one embodiment, the peptide comprises or consists of VDVPNGDISLA (SEQ ID NO: 43). In one embodiment, the peptide comprises or consists of VDVPNGDIS (SEQ ID NO: 44).
  • the peptide comprises or consists of GDPNDGRGDSVVYGLR (SEQ ID NO: 45 In one embodiment, the peptide comprises or consists of LDGLVRAYDNISPVG (SEQ ID NO: 46). In one embodiment, the peptide comprises or consists of GDPNGDISVVYGLR (SEQ ID NO: 47). In one embodiment, the peptide comprises or consists of VDVPNGDISLAYRLR (SEQ ID NO: 48). In one embodiment, the peptide comprises or consists of VDVPEGDISLAYRLR (SEQ ID NO: 49). In one embodiment, the peptide comprises or consists of V(beta-D)TYDGDISVVYGLR (SEQ ID NO: 50).
  • the peptide comprises or consists of VDTY(beta-D)GDISVVYGLR (SEQ ID NO: 51 In one embodiment, the peptide comprises or consists of VDTYDG(beta- D)ISVVYGLR (SEQ ID NO: 52). In one embodiment, the peptide comprises or consists of CLAEIDSC (Cyclic) (SEQ ID NO: 130). In one embodiment, the peptide comprises or consists of CFKPLAEIDSIECSYGIK (Cyclic) (SEQ ID NO: 131). In one embodiment, the peptide comprises or consists of Cyclic CFKPL EIDSIEC (SEQ ID NO: 132).
  • the peptide comprises or consists of KPLAEIDSIELSYGI (SEQ ID NO: 133). In one embodiment, the peptide comprises or consists of KPLAEIDSIELSYG (SEQ ID NO: 134). In one embodiment, the peptide comprises or consists of KPLAEIDSIELSY (SEQ ID NO: 135), KPLAEIDSIELS (SEQ ID NO: 136). In one embodiment, the peptide comprises or consists of KPLAEIDSIEL (SEQ ID NO: 137). In one embodiment, the peptide comprises or consists of KPLAEIDSIE (SEQ ID NO: 138).
  • the agent as defined herein can be in the form of a pharmaceutically acceptable salt or prodrug of said agent.
  • the agent as defined herein can be formulated as a pharmaceutically acceptable addition salt or hydrate of said agent, such as but not limited to K + , Na + , as well as non-salt e.g. H + .
  • the agent is a chemical derivative of a peptide.
  • Chemical derivatives of one or more amino acids may be achieved by reaction with a functional side group.
  • Such derivatives include, for example, those molecules in which free amino groups have been derivatized to form amine hydrochlorides, p-toluene sulphonyl groups, carboxybenzoxy groups, f-butyloxycarbonyl groups, chloroacetyl groups or formyl groups.
  • Free carboxyl groups may be derivatized to form salts, methyl and ethyl esters or other types of esters and hydrazides.
  • Free hydroxyl groups may be derivatized to form O-acyl or O-alkyl derivatives.
  • Also included as chemical derivatives are those peptides which contain naturally occurring amino acid derivatives of the twenty standard amino acids.
  • 4-hydroxyproline may be substituted for proline
  • 5- hydroxylysine may be substituted for lysine
  • 3-methylhistidine may be substituted for histidine
  • homoserine may be substituted for serine and ornithine for lysine.
  • Derivatives also include peptides containing one or more additions or deletions as long as the requisite activity is maintained.
  • Other included modifications are amidation, amino terminal acylation (e.g. acetylation or thioglycolic acid amidation), terminal carboxylamidation (e.g. with ammonia or methylamine), and the like terminal modifications.
  • the agent is further modified such by glycosylation, PEGylation, amidation, esterification, acylation, acetylation and/or alkylation.
  • the agent is further conjugated to a moiety, which may be selected from the group consisting of polyethylene glycol (PEG), monosaccharides, fluorophores, chromophores, radioactive compounds, and cell-penetrating peptides.
  • PEG polyethylene glycol
  • monosaccharides include fluorophores, chromophores, radioactive compounds, and cell-penetrating peptides.
  • the fluorophore is selected from the group consisting of Lucifer yellow, biotin, 5,6-carboxyltetramethylrhodamine (TAMRA), indodicarbocyanine (C5) Alexa Fluor® 488, Alexa Fluor® 532, Alexa Fluor® 647, ATTO 488, ATTO 532, 6- carboxyfluorescein (6-FAM), Alexa Fluor® 350, DY-415, ATTO 425, ATTO 465, Bodipy® FL, fluorescein isothiocyanate, Oregon Green® 488, Oregon Green® 514, Rhodamine GreenTM, 5’-Tetrachloro-Fluorescein, ATTO 520, 6-carboxy-4',5'-dichloro- 2',7'-dimethoxyfluoresceine, Yakima YellowTM dyes, Bodipy® 530/550, hexachlorofluorescein, Alexa Fluor® 555, DY-549
  • the agent further comprises a detectable moiety, such as a moiety that is detectable by an imaging technique such as SPECT, PET, MRI, optical or ultrasound imaging.
  • the detectable moiety comprises or consists of a radioisotope, such as selected from the group consisting of 99m Tc, 111 In, 67 Ga, 58 Ga, 72 As, 89 Zr, 123 l and 201 TI.
  • the peptide comprises or consists of a fusion.
  • the peptide may comprise a fusion of two amino acid sequences as disclosed herein.
  • fusion of a peptide relates to an amino acid sequence fused to another peptide.
  • the said peptide may be fused to a polypeptide such as glutathione-S-transferase (GST) or protein A in order to facilitate purification of said peptide. Examples of such fusions are well known to those skilled in the art.
  • the said peptide may be fused to an oligo-histidine tag such as His6 or to an epitope recognised by an antibody such as the well-known Myc tag epitope. Fusions to any variant or derivative of said peptide are also included in the scope of the disclosure.
  • the fused portion may be a lipophilic molecule or peptide domain that is capable of promoting cellular uptake of the polypeptide, as known to those skilled in the art.
  • said peptide is of no more than no more than 85, such as no more than 80, such as no more than 75, such as no more than 70, such as no more than 65, such as no more than 60, such as nor more than 55, such as no more than 50, such as no more than 55, such as no more than 40 amino acids, such as no more than 35, such as no more than 30, such as no more than 28, such as no more than 26, such as no more than 24, such as no more than 22, such as no more than 20, such as no more than 19, such as no more than 18, such as no more than 17, such as no more than 16, such as no more than 15, such as no more than 14, such as no more than 13, such as no more than 12, such as no more than 11, such as no more than 10 amino acids in length.
  • no more than 80 such as no more than 75, such as no more than 70, such as no more than 65, such as no more than 60, such as nor more than 55, such as no more than 50, such as no more than 55, such as no more than 40 amino acids, such as no
  • the peptide comprises at least 2 additional amino acids, such as at least 3, such as at least 4, such as at least 5, such as at least 6, such as at least 7, such as at least 8, such as at least 9, such as at least 10, such as at least 15 or such as at least 20 amino acids conjugated to the N- or C-terminus of the peptide.
  • said peptide is between 5 and 30 amino acids in length, such as between 5 and 20, such as between 8 and 20, such as between 8 and 18, such as between 10 and 16 amino acids in length.
  • the peptide when is T, then the peptide comprises no more than 25 amino acid residues.
  • the agent is a fragment of a peptide described herein, and the fragment comprises 15 or fewer amino acids in length, such as fewer than 14 amino acids, such as fewer than 13 amino acids, such as fewer than 12 amino acids, such as fewer than 11 amino acids, such as fewer than 10 amino acids, such as fewer than 9 amino acids, such as fewer than 8 amino acids, such as fewer than 7 amino acids, such as fewer than 6 amino acids, such as fewer than 5 amino acids in length.
  • fragment at least 5 contiguous amino acids of the amino acid sequence are included, for example at least 6, 7, 8, 9, 10, 11 , 12, 13, 14 or 15 contiguous amino acids of the amino acid sequence.
  • the fragment may be 15 or fewer amino acids in length, for example 14, 13, 12, 11 , 10, 9, 8, 7, 6 or 5 amino acids in length.
  • the agent is a variant of a peptide as described herein, wherein the variant comprises or consists of a sequence wherein any one amino acid has been altered for another proteinogenic or non-proteinogenic amino acid, with the proviso that no more than five amino acids are so altered.
  • variant refers to a peptide that does not share 100% amino acid sequence identity with the parent peptide, i.e. one or more amino acids must be mutated.
  • “Mutated” refers to altering an amino acid at a specified position in the parent peptide. For example, an amino acid at a specified position may be deleted, altered, substituted or may be the site of an insertion/addition of one or more amino acids. It will be appreciated by persons skilled in the art that the substitutions may be conservative or non-conservative.
  • said peptide variant comprises or consists of a sequence wherein no more than five amino acids are altered for another proteinogenic or non- proteinogenic amino acid, such as no more than 4 amino acids, such as no more than 3 amino acids, such as no more than 2 amino acids, such as no more than 1 amino acid is altered.
  • one or more amino acids are conservatively substituted. “Conservatively substituted” refers to a substitution of one amino acid with another with similar properties (size, hydrophobicity, etc.), such that the function of the peptide is not significantly altered. Thus, by “conservative substitutions” is intended combinations such as Gly, Ala; Vai, He, Leu; Asp, Glu; Asn, Gin; Ser, Thr; Lys, Arg; and Phe, Tyr.
  • the peptide has one additional amino acid.
  • said peptide comprises or consists of one or more additional amino acids, inserted at the N- and/or C-terminus and/or internally within the sequence.
  • at least 2 additional amino acids such as at least 3, such as at least 4, such as at least 5, such as at least 6, such as at least 7, such as at least 8, such as at least 9, such as at least 10, such as at least 15 or such as at least 20 additional amino acids are inserted.
  • the agent is a polynucleotide encoding upon expression a peptide as described herein.
  • the agent is a vector comprising a polynucleotide as described herein.
  • the agent is a cell comprising a polynucleotide or a vector as described herein.
  • the agent is in a composition.
  • the composition is a pharmaceutical composition.
  • the composition is a cosmetic composition.
  • the composition is a coating.
  • Coatings on various implants are known in the art. Applications in humans include central venous catheters, coronary stents, ventricular assist devices, extracorporeal blood circuits, blood sampling devices, and vascular grafts. Such coatings can be in a gel or non-gel form.
  • a "coating comprising the agent” includes that the agent adsorbed to the surface, bonded to the surface, and imbedded in the polymer surface.
  • the present invention relates to an implant comprising the agent described herein.
  • the implant is coated with a composition comprising the agent.
  • the agent may for example be adsorbed to the surface, bonded to the surface, and imbedded in the polymer surface of the implant.
  • the implant is of a biomaterial, such as bone.
  • the implant is a medical device, such as a stent.
  • the present invention relates to a medical device comprising the agent as defined herein.
  • the medical device is a stent.
  • the medical device is a wound dressing.
  • the medical device is a mesh.
  • a normal wound healing process typically includes the stages of 1) haemostasis via platelets, fibrin, and proteoglycans; 2) inflammation comprising recruitment of neutrophils, macrophages, and lymphocytes; 3) proliferation comprising formation of fibroblasts, epithelial cells, collagen, and endothelial cells; and 4) remodelling including collagen fibril cross-linking and scar maturation.
  • the mode of action of the disclosed polypeptides as outlined herein is likely to play a favourable role in the inflammation, proliferation, and remodelling stages of wound healing through the interaction with the NRP-1 co-receptors such as VEGFR and several others. An interplay of the agents of the disclosure with several important growth factors can therefore improve the wound healing process.
  • the composition of the disclosure is a pharmaceutical composition.
  • the composition of the disclosure is a cosmetic composition.
  • the composition is a coating composition.
  • Tissue meshes have found use in facilitating wound healing.
  • One embodiment of the present disclosure provides for a mesh comprising an agent of the disclosure.
  • One embodiment of the disclosure provides for a stent comprising an agent of the disclosure.
  • One embodiment of the present disclosure provides for a wound dressing comprising the agent of the disclosure.
  • the agents of the present disclosure were found to increase vascularisation/induce angiogenesis, and to induce healing of wounds.
  • One embodiment of the disclosure provides for the agent of the disclosure for use in the treatment and/or regeneration of a tissue defect in a subject.
  • One embodiment of the disclosure provides for an agent of the disclosure for use in the treatment and/or promoting healing of a tissue defect.
  • the agents of the disclosure induce healing of epithelial tissue.
  • the tissue defect is an epithelial tissue defect.
  • the tissue defect is a skin defect.
  • the tissue is epithelium.
  • the tissue is epithelial cells.
  • the tissue is skin.
  • the skin defect is a wound, such as an open wound.
  • the wound is a hard-to-heal wound, such as a chronic wound.
  • the wound is a surgical wound.
  • the wound is associated with venous insufficiency, arterial insufficiency, diabetic complications, sickle cell anaemia, or is a pressure-related wound.
  • the diabetic complication is due to diabetes mellitus, such as Type 1 diabetes mellitus, such as Type 1 diabetes mellitus with foot ulcer and/or Type 1 diabetes mellitus with other skin ulcer, or Type 2 diabetes mellitus, such as Type 2 diabetes mellitus with foot ulcer and/or Type 2 diabetes mellitus with other skin ulcer.
  • the diabetes mellitus presents with gangrene.
  • the diabetes mellitus is associated with a disease of the connective tissue, such as fibrosis. In one embodiment, the diabetes mellitus presents with fibrosis. In one embodiment, the diabetes mellitus is associated with fibrosis. On embodiment of the disclosure provides for an agent of the disclosure for prevention or treatment of fibrosis. In one embodiment of the disclosure, the fibrosis is associated with diabetes mellitus.
  • the wound is a dermal ulcer, or a skin defect due to vascular insufficiency or diabetic affection of the blood vessels.
  • the tissue defect is a diabetic complication.
  • the diabetic complication is a diabetes mellitus related eye condition.
  • the diabetic complication is retinopathy.
  • the tissue defect is damage to the eye and/or orbit, such as injury (trauma) of the eye and/or orbit.
  • the skin defect is a skin disorder provoked by external factors.
  • the skin disorder provoked by external factors is pressure ulceration, dermatoses provoked by friction or mechanical stress, dermatoses due to foreign bodies, dermatoses provoked or exacerbated by exposure to cold, dermatoses provoked by heat or electricity, dermatoses provoked by light or UV radiation, dermatoses due to ionizing radiation, allergic contact dermatitis, photo-allergic contact dermatitis, irritant contact dermatitis, allergic contact urticarial, protein contact dermatitis, allergic contact sensitisation, phototoxic reactions to skin contact with photoactive agents, cutaneous reactions to venomous or noxious animals.
  • the skin defect is an inflammatory dermatosis.
  • the inflammatory dermatosis is dermatitis, eczema, atopic dermatitis, papulosquamous dermatoses, urticaria, angioedema or other urticarial disorders, inflammatory erythemas and other reactive inflammatory dermatoses, immunobullous diseases of the skin, cutaneous lupus erythematosus, scarring or sclerosing inflammatory dermatoses.
  • the skin defect is sclerosis.
  • the sclerosis is systemic sclerosis (scleroderma), such as circumscribed scleroderma.
  • the skin defect is a postprocedural disorder of the skin.
  • the postprocedural disorder of the skin is an unsatisfactory surgical scar of skin, a cutaneous flap necrosis, a myocutaneous flap necrosis, a skin graft failure, a composite graft failure.
  • the postprocedural disorder of the skin is a surgical incision.
  • the skin defect is a genetic and/or developmental disorder affecting the skin.
  • the genetic and/or developmental disorder affecting the skin is epidermolysis bullosa, or genodermatosis, such as pemphigus genodermatosis.
  • the epidermolysis bullosa is selected from the group consisting of epidermolysis bullosa simplex, junctional epidermolysis bullosa, dystrophic epidermolysis bullosa, recessive dystrophic epidermolysis bullosa, syndromic epidermolysis bullosa and epidermolysis bullosa.
  • the skin defect is an external defect. In one embodiment, the skin defect is an open defect. In one embodiment, the skin defect is an inflamed wound. In one embodiment, the skin defect is infected. Infected wounds can be difficult to heal due to formation of biofilm, e.g. from bacteria.
  • the skin defect is an injury or trauma to internal or external tissue, such as injury or trauma to the epidermis and/or dermis of the skin.
  • the skin defect is an acute defect or a chronic defect.
  • the skin defect is full thickness skin defect.
  • the wound is an acute wound or a chronic wound.
  • the acute wound is an incision, laceration, abrasion graze or burn, a puncture wound, a penetration wound or a wound due to dermatologic diseases such as psoriasis, acne and eczema.
  • the chronic wound is a venous ulcer, a diabetic ulcer, a pressure ulcer, corneal ulcer, digestive ulcer or wounds as a result of ischemia and radiation poisoning.
  • the wound is a non-pressure chronic ulcer, such as a nonpressure chronic ulcer of lower limb.
  • the agent is administered topically, orally, subcutaneously, or systemically. In one embodiment, the administration is by instillation.
  • One embodiment of the disclosure provides for an agent of the disclosure for use in the treatment of a disease of the skin, such as injury of external cause.
  • the skin defect is a disease of the skin, such as injury of external cause.
  • the disease of the skin is selected from the group consisting of: a. a skin disorder involving specific cutaneous structures, such as disorders of cutaneous blood and lymphatic vessels, such as dermatoses resulting from vascular insufficiency, such as ischaemic ulceration of the skin, and b. a skin disorder associated with pregnancy, the neonatal period, and infancy, such as dermatoses of infancy, such as erythrodermas of infancy (Leiner’s disease).
  • the injury of external cause is selected from the group consisting of: a. injury to the head, such as intracranial injury, such as diffuse brain injury, such as cerebral contusion, and b. injury to unspecified part of trunk, limb, or body region, such as other injuries of spine or trunk, such as injury of the spinal cord.
  • the subject is suffering from diabetes mellitus.
  • the subject is suffering from stoma.
  • the wound is generated as part of internal acidic fluids.
  • the subject is a mammal. In one embodiment, the mammal is a human.
  • One embodiment of the disclosure provides for a method for treatment and/or regeneration of a tissue defect in a subject, the method comprising administering a therapeutically effective amount of an agent of the disclosure to an subject in need thereof.
  • One embodiment of the disclosure provides for a method for treating and/or promoting healing of a tissue defect in a subject, the method comprising administering a therapeutically effective amount of an agent of the disclosure to an subject in need thereof.
  • One embodiment of the disclosure provides for a method for regeneration of skin at the site of a skin defect and/or for reducing scar formation resulting from healing of a skin defect in a subject, said method comprising administering an agent of the disclosure.
  • One embodiment of the disclosure provides for a use of an agent of the disclosure for the manufacture of a medicament for the treatment and/or regeneration of a skin defect in a subject.
  • One embodiment of the disclosure provides for a use of an agent of the disclosure for the manufacture of a medicament for the treating and/or promoting healing of a tissue defect in a subject.
  • One embodiment of the disclosure provides for a use of an agent of the disclosure for the manufacture of a medicament for the treatment of a skin defect in a subject.
  • One embodiment provides for a use of an agent of the disclosure for reducing scar formation resulting from healing of a skin defect in a subject.
  • One embodiment provides for a use of an agent of the disclosure for promoting the healing of a skin defect in a subject.
  • One embodiment of the disclosure provides a use of an agent of the disclosure for promoting vascularisation in a healing tissue, such as skin healing from a skin defect, such as a wound.
  • One embodiment provides for a use of an agent of the disclosure in combination with a tissue meshing technique, for promoting healing of a skin defect, such as a wound.
  • One embodiment provides for a use of an agent of the disclosure for promoting grafting of skin.
  • One embodiment of the disclosure provides for a method for inserting a stent in a subject, said method comprising administering an agent of the disclosure.
  • the stent is coated with the agent of the disclosure.
  • the agent of the disclosure is part of a cosmetic composition.
  • the peptide comprises or consists of the amino acid sequence VDTYDGDISVVYGLR (SEQ ID NO: 7), and the tissue defect is a wound.
  • the peptide comprises or consists of the amino acid sequence VDTYDGGISVVYGLR (SEQ ID NO: 6), and the tissue defect is a wound. In one embodiment, the peptide comprises or consists of the amino acid sequence AEIDSIELSYGIK (SEQ ID NO: 110), and the tissue defect is a wound.
  • the peptide comprises or consists of the amino acid sequence KPLAEIDSIELSYGIK (SEQ ID NO: 111), and the tissue defect is a wound.
  • One aspect of the disclosure provides for the agent of the disclosure for use in the prevention of fibrosis of the skin in a subject.
  • One aspect of the disclosure provides for an agent of the disclosure for use in the treatment of a disease of the skin, such as injury of external cause.
  • One aspect of the disclosure provides for a method for treatment and/or regeneration of a tissue defect in a subject, the method comprising administering a therapeutically effective amount of an agent of the disclosure to an subject in need thereof.
  • One aspect of the disclosure provides for a method for treating and/or promoting healing of a tissue defect in a subject, the method comprising administering a therapeutically effective amount of an agent of the disclosure to an subject in need thereof.
  • One aspect provides for a use of an agent of the disclosure for reducing scar formation resulting from healing of a skin defect in a subject.
  • One aspect of the disclosure provides a use of an agent of the disclosure for promoting vascularisation in a healing tissue, such as skin healing from a skin defect, such as a wound.
  • An agent comprising: a) a peptide selected from the group consisting of:
  • X 5 is D or G; Xe is I or G;
  • X? is V or L
  • Xs is V or A
  • X14 is E or G
  • X15 is S or T
  • VDTYDGDISVVYGL (SEQ ID NO: 34),
  • VDTYDGDISVVYG (SEQ ID NO: 35),
  • VDTYDGDISVVY (SEQ ID NO: 36),
  • VDTYDGDISVV (SEQ ID NO: 37),
  • VDTYDGDISV (SEQ ID NO: 38),
  • VDTYDGDIS (SEQ ID NO: 39),
  • VDTYDGRGDSVVYGLR (SEQ ID NO: 40),
  • VDVPNGDISLAYGL (SEQ ID NO: 41),
  • VDVPNGDISLAYG (SEQ ID NO: 42),
  • VDVPNGDISLA (SEQ ID NO: 43),
  • VDVPNGDIS (SEQ ID NO: 44),
  • VDVPNGDISLAYRLR (SEQ ID NO: 48)
  • VDVPEGDISLAYRLR (SEQ ID NO: 49),
  • V(beta-D)TYDGDISVVYGLR (SEQ ID NO: 50)
  • VDTYDG(beta-D)ISVVYGLR (SEQ ID NO: 52);
  • KPLAEIDSIELSYG (SEQ ID NO: 134), KPLAEIDSIELSY (SEQ ID NO: 135), KPLAEIDSIELS (SEQ ID NO: 136), KPLAEIDSIEL (SEQ ID NO: 137), and KPLAEIDSIE (SEQ ID NO: 138); b) a polynucleotide encoding upon expression, the peptide of a); c) a vector comprising the polynucleotide of b); or d) a cell comprising the polynucleotide of b), or the vector of c).
  • the agent according to item 1 wherein the peptide comprises an amino acid sequence of the general formula:
  • VDX 2 X3X4GX5X 6 SX7X 8 YGLR (SEQ ID NO: 2) wherein: X 2 is T or V;
  • X 3 is Y or P
  • X 4 is D or N
  • X 5 is D or G
  • X s is I or G
  • X 7 is V or L; and X 8 is V or A.
  • X 4 is D or N
  • Xs is D or G
  • X 8 is I or G
  • X7 is V or L; and Xs is V or A.
  • Xs is D or G
  • Xs is I or G
  • X7 is V or L
  • Xs is V or A.
  • the agent according to any one of the preceding items, wherein the peptide comprises an amino acid sequence of the general formula: VDTYDGZ 7 Z 8 SVVYGLR (SEQ ID NO: 5) wherein:
  • X 5 is D or G; and X 5 is I or G;
  • the agent according to item 1 wherein the peptide comprises an amino acid sequence of the general formula:
  • X 9 is C, P or G
  • X is E or G
  • Xn is C, D or I
  • X12 is D, I, S or G
  • X13 is S, D or G
  • X14 is E or G; and X15 is S or T.
  • the agent according to item 1 wherein the peptide comprises an amino acid sequence of the general formula:
  • Xg is C, P or G
  • X is E or G
  • X11 is C, I or absent
  • X12 is D, G or absent
  • X13 is S, G or absent
  • X14 is E or G. 8. The agent according to item 1 , wherein the peptide comprises an amino acid sequence of the general formula:
  • X9 is C, P or G
  • X is E or G; and X14 is E or G.
  • the agent according to item 1 wherein the peptide comprises or consists of an amino acid sequence selected from the group consisting of: VDTYDGGISVVYGLR (SEQ ID NO: 6),
  • VDTYDGDISVVYGLR (SEQ ID NO: 7), DTYDGDISVVYGLR (SEQ ID NO: 8), TYDGDISVVYGLRS (SEQ ID NO: 9), TYDGDISVVYGLR (SEQ ID NO: 10), YDGDISVVYGLRS (SEQ ID NO: 11), YDGDISVVYGLR (SEQ ID NO: 12), DGDISVVYGLRS (SEQ ID NO: 13), DGDISVVYGLR (SEQ ID NO: 14), GDISVVYGLRS (SEQ ID NO: 15), GDISVVYGLR (SEQ ID NO: 16), DISVVYGLRS (SEQ ID NO: 17), DISVVYGLR (SEQ ID NO: 18), VDVPNGDISLAYGLR (SEQ ID NO: 19), DVPNGDISLAYGLRS (SEQ ID NO: 20), DVPNGDISLAYGLR (SEQ ID NO: 21), VPNGDISLAYGLRS (SEQ ID NO: 22
  • DISLAYGLRS (SEQ ID NO: 30), DISLAYGLR (SEQ ID NO: 31), VDTYDGDGSVVYGLR (SEQ ID NO: 32), VDVPEGDISLAYGLR (SEQ ID NO: 33).
  • KPLAEGDIELSYGIK (SEQ ID NO: 116), KPLAEIELSYGIK (SEQ ID NO: 117), KPLAEIDSIELTYGIK (SEQ ID NO: 118), KPLAEIDGIELSYGIK (SEQ ID NO: 119), KPLAEIDGIELTYGIK (SEQ ID NO: 120), KPLAEIGSIELSYGIK (SEQ ID NO: 121), KGLAEIDSIELSYGIK (SEQ ID NO: 122), KPLAGIDSIGLSYGIK (SEQ ID NO: 123), Cyclic KCLAEIDSCELSYGIK (SEQ ID NO: 124), LAEIDSIELSYGIK (SEQ ID NO: 125), EIDSIELSYGIK (SEQ ID NO: 126), IDSIELSYGIK (SEQ ID NO: 127), DSIELSYGIK (SEQ ID NO: 128), SIELSYGIK (SEQ ID NO: 129), and IEL
  • the peptide comprises or consists of an amino acid sequence VDTYDGGISVVYGLR (SEQ ID NO: 6). 13. The agent according to item 1 , wherein the peptide comprises or consists of an amino acid sequence VDTYDGDISVVYGLR (SEQ ID NO: 7).
  • the peptide comprises no more than 85, such as no more than 80, such as no more than 75, such as no more than 70, such as no more than 65, such as no more than 60, such as nor more than 55, such as no more than 50, such as no more than 55, such as no more than 40 amino acids, such as no more than 35, such as no more than 30, such as no more than 28, such as no more than 26, such as no more than 24, such as no more than 22, such as no more than 20, such as no more than 19, such as no more than 18, such as no more than 17, such as no more than 16, such as no more than 15, such as no more than 14, such as no more than 13, such as no more than 12, such as no more than 11 , such as no more than 10 amino acids.
  • no more than 80 such as no more than 75, such as no more than 70, such as no more than 65, such as no more than 60, such as nor more than 55, such as no more than 50, such as no more than 55, such as no more than 40 amino acids, such as no more than 35,
  • the peptide comprises at least 2 additional amino acids, such as at least 3, such as at least 4, such as at least 5, such as at least 6, such as at least 7, such as at least 8, such as at least 9, such as at least 10, such as at least 15 or such as at least 20 amino acids conjugated to the N- or C-terminus of the peptide.
  • agent according to any one of the preceding items, wherein the agent comprises or consists of a tandem repeat comprising two or more repeat units.
  • agent is a variant of the peptide, wherein the variant comprises or consists of a sequence wherein any one amino acid has been altered for another proteinogenic or non- proteinogenic amino acid, with the proviso that no more than five amino acids are so altered.
  • the variant comprises or consists of a sequence wherein no more than five amino acids are altered for another proteinogenic or non-proteinogenic amino acid, such as no more than 4 amino acids, such as no more than 3 amino acids, such as no more than 2 amino acids, such as no more than 1 amino acid is altered.
  • agent according to any of the preceding items, wherein the agent further comprises a detectable moiety.
  • radioisotope is selected from the group consisting of 99m Tc, 111 ln, 67 Ga, 68 Ga, 72 As, 89 Zr, 123 l and 201 TI.
  • composition comprising the agent according to any one of the preceding items.
  • composition according to item 38 wherein the composition is a pharmaceutical composition.
  • composition according to item 38 wherein the composition is a cosmetic composition.
  • 41 The composition according to any one of the preceding items, wherein the composition is a coating composition.
  • a medical device comprising an agent according to any one of the preceding items.
  • a mesh comprising an agent according to any one of the preceding items.
  • a stent comprising an agent according to any one of the preceding items.
  • a wound dressing comprising the agent according to any one of the preceding items.
  • tissue defect is a skin defect.
  • the skin defect is a wound, such as an open wound.
  • the wound is a hard-to-heal wound, such as a chronic wound.
  • the wound is associated with venous insufficiency, arterial insufficiency, diabetic complications, sickle cell anaemia, or is a pressure-related wound.
  • diabetes mellitus such as Type 1 diabetes mellitus, such as Type 1 diabetes mellitus with foot ulcer and/or Type 1 diabetes mellitus with other skin ulcer, or Type 2 diabetes mellitus, such as Type 2 diabetes mellitus with foot ulcer and/or Type 2 diabetes mellitus with other skin ulcer.
  • diabetes mellitus is associated with a disease of the connective tissue, such as fibrosis.
  • the wound is a dermal ulcer, or a skin defect due to vascular insufficiency or diabetic affection of the blood vessels.
  • tissue defect is a diabetic complication.
  • the agent for use according to any one of the preceding items, wherein the diabetic complication is retinopathy.
  • the agent for use according to any one of the preceding items, wherein the tissue defect is damage to the eye and/or orbit, such as injury (trauma) of the eye and/or orbit.
  • the agent for use according to any one of the preceding items, wherein the skin defect is a skin disorder provoked by external factors.
  • the skin disorder provoked by external factors is pressure ulceration, dermatoses provoked by friction or mechanical stress, dermatoses due to foreign bodies, dermatoses provoked or exacerbated by exposure to cold, dermatoses provoked by heat or electricity, dermatoses provoked by light or UV radiation, dermatoses due to ionizing radiation, allergic contact dermatitis, photo-allergic contact dermatitis, irritant contact dermatitis, allergic contact urticarial, protein contact dermatitis, allergic contact sensitisation, phototoxic reactions to skin contact with photoactive agents, cutaneous reactions to venomous or noxious animals.
  • the agent for use according to any one of the preceding items, wherein the skin defect is an inflammatory dermatosis.
  • the agent for use according to any one of the preceding items, wherein the inflammatory dermatosis is dermatitis, eczema, atopic dermatitis, papulosquamous dermatoses, urticaria, angioedema or other urticarial disorders, inflammatory erythemas and other reactive inflammatory dermatoses, immunobullous diseases of the skin, cutaneous lupus erythematosus, scarring or sclerosing inflammatory dermatoses.
  • the agent for use according to any one of the preceding items, wherein the skin defect is sclerosis.
  • sclerosis is systemic sclerosis (scleroderma), such as circumscribed scleroderma.
  • skin defect is a postprocedural disorder of the skin.
  • the postprocedural disorder of the skin is an unsatisfactory surgical scar of skin, a cutaneous flap necrosis, a myocutaneous flap necrosis, a skin graft failure, a composite graft failure.
  • agent for use according to any one of the preceding items, wherein the genetic and/or developmental disorder affecting the skin is epidermolysis bullosa, or genodermatosis, such as pemphigus genodermatosis.
  • the epidermolysis bullosa is selected from the group consisting of epidermolysis bullosa simplex, junctional epidermolysis bullosa, dystrophic epidermolysis bullosa, recessive dystrophic epidermolysis bullosa, syndromic epidermolysis bullosa and epidermolysis bullosa.
  • the skin defect is infected.
  • the skin defect is an injury or trauma to internal or external tissue, such as injury or trauma to the epidermis and/or dermis of the skin.
  • the acute wound is an incision, laceration, abrasion graze or burn, a puncture wound, a penetration wound or a wound due to dermatologic diseases such as psoriasis, acne and eczema.
  • the chronic wound is a venous ulcer, a diabetic ulcer, a pressure ulcer, corneal ulcer, digestive ulcer or wounds as a result of ischemia and radiation poisoning.
  • agent for use according to any one of the preceding items wherein the agent is administered topically, orally, subcutaneously, or systemically.
  • the disease of the skin is selected from the group consisting of: a. a skin disorder involving specific cutaneous structures, such as disorders of cutaneous blood and lymphatic vessels, such as dermatoses resulting from vascular insufficiency, such as ischaemic ulceration of the skin, and b. a skin disorder associated with pregnancy, the neonatal period, and infancy, such as dermatoses of infancy, such as erythrodermas of infancy (Leiner’s disease).
  • the injury of external cause is selected from the group consisting of: a. injury to the head, such as intracranial injury, such as diffuse brain injury, such as cerebral contusion, and b. injury to unspecified part of trunk, limb, or body region, such as other injuries of spine or trunk, such as injury of the spinal cord.
  • agent for use according to item 94 wherein the mammal is a human.
  • a method for treatment and/or regeneration of a tissue defect in a subject comprising administering a therapeutically effective amount of an agent according to any one of the preceding items to an subject in need thereof.
  • a method for treating and/or promoting healing of a tissue defect in a subject comprising administering a therapeutically effective amount of an agent according to any one of the preceding items to an subject in need thereof.
  • a method for regeneration of skin at the site of a skin defect and/or for reducing scar formation resulting from healing of a skin defect in a subject comprising administering an agent according to any one of the preceding items.
  • a method for inserting a stent in a subject comprising administering an agent according to any one of the preceding items.
  • 108. The method according to item 107, wherein the stent is coated with the agent according to any one of the preceding items.
  • Example 1 Polypeptides of the disclosure stimulates endothelial and vascular smooth muscle cell proliferation Methods
  • human umbilical cord endothelial cells were first treated with a scramble non-coding siRNA (NC) or siRNA for NRP-1 , subsequently they were treated with FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM, and DNA synthesis analyzed by measuring the uptake of BrdU.
  • NC scramble non-coding siRNA
  • FOL26 FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM
  • polypeptides of the disclosure stimulates endothelial and vascular smooth muscle cell proliferation.
  • Cells were transfected for 48 hours or performed with 0.5% supplement starvation for 24 hours on 6-well plates as described above and dissociated by Accutase (Gibco, USA) to obtain cell suspension.
  • Geltrex reduced growth factor basement membrane matrix ((Invitrogen, CA, USA) was thawed at 4°C overnight and added into a 96-well plate (50 pl/well) next. The coated 96-well plate was placed in the incubator more than 30 mins for further use and the cell suspension was seeded into it at a density of 1.5 x 104/well.
  • NRP-1 silencing experiments cells were cultured with complete medium.
  • polypeptide treatment experiments cells were stimulated with complete medium plus the different concentration of polypeptide.
  • NRP-1 rescued experiments cells transfected with siRNA were cultured with complete medium and NRP-1 stimulations. After a 24-hour incubation, images were recorded by an inverted microscope (Nikon, Eclipse TE2000-U, Japan) and the sum length of master segments of vessels in each image were quantified using Angiogenesis Analyzer plug-in ImageJ software program (pixel/unit). Data were analyzed with photographs from 8-14 wells per condition.
  • Human umbilical cord endothelial cells were first treated with a scramble non-coding siRNA (NC) or siRNA for NRP-1 , subsequently treated with FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM, and endothelial tube formation and the total master segemnts were measured using ImageJ. gene expression of PECAM-1 (CD31) analyzed.
  • NC scramble non-coding siRNA
  • FOL26 FOL26 in concentrations of 10 nM, 100 nM, and 1000 nM
  • test polypeptides significantly increased tube formation in the HUVECs.
  • the polypeptides also increased expression of PECAM-1.
  • Figure 12 shows that FOL26 increased tube formation compared to control in a NRP-1 dependent way.
  • Polypeptides of the disclosure induces tube formation in HUVECs.
  • Example 3 Polypeptides of the disclosure induces wound healing in a scratch wound assay on HUVECs and HCASMCs
  • Ibidi Culture-Insert 2 well (Ibidi, Nanterre, France) was used to assess the ability of wound healing in vitro. Briefly, one silicone culture-insert (cut 2 well inlet) was placed in the one well of 24-well plate and 3.5x104 cells were seeded into each inlet with 70 pl complete medium. For NRP-1 silencing and NRP-1 rescued experiments, cells were transfected with siRNA/siRNA-NC for 48 hours before seeding. For test polypeptide treatment experiments, cells were starved with respective low-serum medium for 24 hours and 48 hours after adhering in the inlet. The cell-free gap between two inlets was created by removing the culture-insert, followed by washing with DPBS gently to remove suspended cells.
  • wound closure % 1 - (wound area at T x I wound area at To)] * 100 (To is the wound area at 0 h, and T x is the wound area at corresponding hours after creating the gap (pixel/unit). Data were quantified with pictures from 8-20 wells per group.
  • Polypeptides of the disclosure induces wound healing in a scratch wound assay on HUVECs and HCASMCs.
  • Example 4 Polypeptides of the disclosure prevents apoptosis in HUVECs and HCASMCs
  • HAVECs Human Umbilical Vein Endothelial Cells
  • HCASMCs Human Coronary Artery Smooth Muscle Cells
  • Test polypeptide and sFasL were dissolved in the respective complete medium.
  • Caspase-Gio 3/7 Assay was performed to evaluate cell apoptosis. HUVECs and HCASMCs were seeded into 96-well plates at a density of 3-5 x 103 per well. For additional NRP-1 on cells transfected siRNA, NRP-1 was added 24 hours after transfection and the mixture of Caspase-Gio 3/7 was added 48 hours after polypeptide stimulations. Subsequent steps were performed as manufacturer’s instructions using the FOL26 and FOL56 as test polypeptides. Data were read by Wallac 1420 Victor 2 (Perkin Elmer, USA).
  • Polypeptides of the disclosure prevents apoptosis in HUVECs and HCASMCs.
  • Example 5 Effect on gene expression of polypeptides of the disclosure in HUVECs and HCASMCs
  • real-time qPCR was performed using by KAPA SYBR FAST One-Step qRT- PCR Master Mix Kit (KAPA Biosystems, USA) on LightCycler 480 system (Roche, Mannheim, Germany).
  • Table 1 Growth Factors and Cytokines Affecting Various Steps in Wound Healing.
  • Polypeptides of the disclosure induces a milieu inducing wound healing by TNF and IL- 6 expression likely to promote fibroblast migration and proliferation, collagen secretion, angiogenesis, and induce correct wound healing structure by inducing cell remodelling through MMP expression.
  • Example 6 Healing of wounds in combination with tissue meshing.
  • tissue meshes having a coating of a polypeptide of the disclosure, and/or having a polypeptide dispersed within it will effect faster and/or more complete healing of a wound, compared to tissue meshes without the polypeptide of the disclosure.
  • Example 7 Effect of test polypeptides in acute and pathological wound healing Methods
  • test polypeptides The pharmacological profile of the test polypeptides has been tested using two models, with full-thickness human abdominal skin cultured ex-vivo under normal conditions (WCM medium only, 37 °C, 5% CO2,) representing “acute wound healing” and under extreme culture conditions (VCM, high glucose, H2O2, 37 °C, 5% CO2, 5% O2) representing “pathological wound healing” (Figure 13) (Monasterium Labs, Prof. Ralf Paus, Munster, Germany).
  • Table 2 Number of wound closure observed in acute wound healing-pooled data at 72 hours timer point*. *No full wound closure was observed in the pathological punches. The wound closure in day 6 time point from the first experiment were excluded from this table.
  • Polypeptides of the disclosure induce full wound closure.
  • the ROS level (figure27b) activated by 50ug/ml oxidized Low-density Lipoprotein (oxLDL) for 2 hours in cells pre-incubated with FOL- 026 peptides was assessed using H2O2 measurement (n 4-8 per group).
  • the FOL-026 peptide has a stimulatory effect on the proliferation of endothelial cells.
  • Figures 30 a-d shows the positive effect FOL-026 has on tube formation. It demonstrates the increased vessel density effect of FOL-026 and how FOL-026 induces upregulation of angiogenesis related genes.
  • the FOL-026 peptide affects angiogenesis both in vitro and in vivo.
  • Example 10 Effect of NR P-1 knock-down on endothelial cell function induced with
  • Figures 31a-c shows that reducing the NRP-1 expression blocks the effect of FOL-026
  • Figures 32a-b show that FOL-026 has a positive effect on wound closure in siRNA-NC/ siNRP-1-transfected HUVECs.
  • FOL-026 stimulates wound closure in HUVECs by binding to NRP-1
  • the FOL-026 peptide induce tube formation in siRNA-NC and siNRP-1.
  • Example 12 Effect of FOL-026 peptide on smooth muscle cell function
  • FOL-026 induces smooth muscle cell proliferation, inhibits apoptosis, reduces ROS formation and accelerates wound closure
  • Example 13 Effect of NRP-1 knock-down on smooth muscle cell function induced with FOL-026 peptide
  • Method qRT-PCR analysis of NRP-1 mRNA expression in HCASMCs with small interfering RNA transfection for 48 hours (n 4) (figure 36a).
  • Representative images of smooth muscle cells at 0 and 5 hours after scratch wounding were shown (figure 37a).
  • FOL-026 has a positive effect on wound closure in siRNA-NC/ si NRP-1 -transfected HUVECs.
  • Example 14 The FOL-005 peptide stimulates endothelial and smooth muscle cell proliferation and wound healing ability
  • the results shown in figure 38a indicates that the proliferation of human umbilical vascular endothelial cells (HUVECs) increases with the addition of the FOL-005 peptide.
  • the scratch wound assay in figure 38b and 38c indicates that the FOL-005 peptide accelerate HUVEC wound closure.
  • the FOL-005 peptide has a positive effect on endothelial and smooth muscle cell proliferation and wound healing ability.
  • Gottrup F. A specialized wound-healing center concept: importance of a multidisciplinary department structure and surgical treatment facilities in the treatment of chronic wounds.

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Abstract

La présente divulgation concerne des agents comprenant des polypeptides pour traiter des défauts tissulaires, en particulier pour favoriser la cicatrisation de plaies.
PCT/EP2023/062965 2022-05-13 2023-05-15 Agents de stimulation de régéneration tissulaire Ceased WO2023218095A1 (fr)

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Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6482410B1 (en) * 1994-09-16 2002-11-19 The Scripps Research Institute Cytotactin derivatives that stimulate attachment and neurite outgrowth, and methods of making same
WO2013021212A2 (fr) * 2011-08-10 2013-02-14 Follicum Ab Nouvelles compositions et leurs utilisations
WO2018202870A1 (fr) * 2017-05-04 2018-11-08 Follicum Ab Peptides pour le traitement du diabète
WO2020094797A1 (fr) * 2018-11-07 2020-05-14 Follicum Ab Fragments peptidiques pour le traitement du diabète
WO2020216896A1 (fr) * 2019-04-24 2020-10-29 Follicum Ab Formulation topique
WO2022090413A1 (fr) * 2020-10-28 2022-05-05 Follicum Ab Peptides destinés à être utilisés dans la pigmentation de la peau et des cheveux

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6482410B1 (en) * 1994-09-16 2002-11-19 The Scripps Research Institute Cytotactin derivatives that stimulate attachment and neurite outgrowth, and methods of making same
WO2013021212A2 (fr) * 2011-08-10 2013-02-14 Follicum Ab Nouvelles compositions et leurs utilisations
WO2018202870A1 (fr) * 2017-05-04 2018-11-08 Follicum Ab Peptides pour le traitement du diabète
WO2020094797A1 (fr) * 2018-11-07 2020-05-14 Follicum Ab Fragments peptidiques pour le traitement du diabète
WO2020216896A1 (fr) * 2019-04-24 2020-10-29 Follicum Ab Formulation topique
WO2022090413A1 (fr) * 2020-10-28 2022-05-05 Follicum Ab Peptides destinés à être utilisés dans la pigmentation de la peau et des cheveux

Non-Patent Citations (3)

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Title
FRYKBERG R. G.BANKS J.: "Challenges in the Treatment of Chronic Wounds", ADV WOUND CARE (NEW ROCHELLE, vol. 4, no. 9, 2015, pages 560 - 82
GOTTRUP F.: "A specialized wound-healing center concept: importance of a multidisciplinary department structure and surgical treatment facilities in the treatment of chronic wounds", THE AMERICAN JOURNAL OF SURGERY, vol. 187, no. 5, 2004, pages S38 - S43
KIRKER K. R.JAMES G. A.: "In vitro studies evaluating the effects of biofilms on wound-healing cells: a review", APMIS, vol. 125, no. 4, 2017, pages 344 - 52

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