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WO2008125864A1 - Sfrp3 utilisée pour accélérer la guérison de plaies - Google Patents

Sfrp3 utilisée pour accélérer la guérison de plaies Download PDF

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Publication number
WO2008125864A1
WO2008125864A1 PCT/GB2008/001338 GB2008001338W WO2008125864A1 WO 2008125864 A1 WO2008125864 A1 WO 2008125864A1 GB 2008001338 W GB2008001338 W GB 2008001338W WO 2008125864 A1 WO2008125864 A1 WO 2008125864A1
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WO
WIPO (PCT)
Prior art keywords
sfrp3
wound
therapeutically effective
healing
wounds
Prior art date
Application number
PCT/GB2008/001338
Other languages
English (en)
Inventor
Mark William James Ferguson
Hugh Laverthy
Nicholas Occleston
Sharon O'kane
Kerry Nield
Original Assignee
Renovo Limited
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Filing date
Publication date
Application filed by Renovo Limited filed Critical Renovo Limited
Publication of WO2008125864A1 publication Critical patent/WO2008125864A1/fr

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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
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/1703—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • A61K38/1709—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • 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

Definitions

  • the present invention relates to medicaments for accelerating wound healing, and to methods of accelerating wound healing.
  • Wounds are a source of discomfort to those afflicted, and may be associated with, or give rise to, a number of clinical difficulties or complications.
  • the wound healing response proceeds through a number of overlapping processes, beginning with the inflammatory response, and proceeding via the production of granulation tissue, wound contraction, and reconstitution of a functional epithelial covering. Perturbation of any of these processes may retard or otherwise disrupt the normal healing response.
  • Clinical approaches to wound management will generally depend on the desired outcome. This outcome may, for example, be considered with reference to the speed at which a wound heals, or with reference to the degree of scarring occurring. In management of some wounds increasing the speed of wound healing is of primary importance, while control of the degree of scarring that occurs is of much lesser importance, hi management of other wounds controlling the degree of scarring occurring is of primary importance, while increasing the speed of wound healing is of much lesser importance.
  • the present invention is preferably applicable to the management of wounds in which the primary clinical concern relates to the speed of healing of the wound.
  • Wounds are painful, even aside from the events associated with their formation, and delays in the healing of wounds may be associated with extended incidences of pain to the sufferer. Wounds can also decrease the mechanical function of the injured area.
  • the secreted Frizzled-related proteins are a group of secreted glycoproteins which structurally resemble the Frizzled family of proteins (which are receptors for WNT signalling proteins). sFRPs possess a cysteine-rich domain (CRD) homologous to the CRD of Frizzled proteins but lack the transmembrane and cytosolic domains of the full length Frizzled proteins.
  • CRDs of sFRPs which share 30-50% sequence similarity with those of Frizzled proteins, include ten conserved cysteine residues which are shared with the Frizzled CRD domain, in addition to further conserved residues such as a proline located four residues C-terminal to cysteine-9.
  • sFRPs The domain spans 120-125 amino acids near the N-terminus of the protein (Melkonyan et al., 1997; Jones and Jomary 2002; Dann et al., 2001).
  • the mechanism of this antagonism has not been fully elucidated, but it has been suggested that it is brought about by a direct interaction between WNT and either the CRD, or the C-terminal domain lying outside the CRD. It is believed that sFRPs may block WNT signalling either by interacting with WNT proteins, and thus preventing binding of WNTs to Frizzled receptors, or by forming non-functional complexes with the Frizzled receptors themselves.
  • sFRP3 Secreted Frizzled Related Protein 3
  • Frizzled-related protein 1 FrzB-1, Frezzled, and Fritz.
  • the amino acid sequence of the human sFRP3 protein is shown in Sequence ID No. 1 (this sequence had previously been deposited as Accession Number: Q92765).
  • the N-terminal CRD domain of sFRP-3 has been shown to be able to bind and inhibit the activity of WNTl and WNT8.
  • sFRP3 has also been shown to bind WNT5A in immuno-precipitation experiments, although this interaction is not sufficient to block WNT5A activity.
  • DNA encoding human sFRP3 is shown in Sequence ID No. 3 (this sequence has previously been deposited as Accession Number NM OO 1463).
  • the present invention provides the use of sFRP3, or a therapeutically effective fragment or derivative thereof, in the manufacture of a medicament for accelerating wound healing.
  • This first aspect of the invention also provides sFRP3, or a therapeutically effective fragment or derivative thereof, for use as a medicament for accelerating wound healing
  • the invention provides a method of accelerating wound healing, the method comprising providing a therapeutically effective amount of sFRP3, or a therapeutically effective fragment or derivative thereof, to a patient in need of such accelerated wound healing.
  • the sFRP3, or therapeutically effective fragment or derivative thereof may be provided to a site where it is desired to accelerate wound healing by any suitable means. It may be preferred that sFRP3, or a therapeutically effective fragment or derivative thereof, is administered to the site where wound healing is to be accelerated.
  • the site may be a site where a wound is to be formed, or preferably may be a wound.
  • the present invention is based on the inventors' new and surprising finding that sFRP3, as well as suitable fragments or derivatives thereof, may be used to accelerate wound healing. Without wishing to be bound by any hypothesis, the inventors believe that the acceleration of wound healing observed occurs as a result of increased wound contraction associated with the provision of sFRP3. There is nothing in the prior art that would previously have led the skilled person to believe that sFRP3, or its fragments or derivatives, may be used to accelerate wound healing or to promote wound contraction.
  • Wound contraction is a normal part of the wound healing process, in which the margins of a wound are drawn together by contractile elements within and surrounding the wound, generally believed to be myofibroblasts.
  • excessive contraction can cause deleterious effects, such as pathological contraction associated with hypertrophic scars which normally occurs after scar formation, the ability to promote contraction during the healing process provides a useful and beneficial manner by which wound healing may be accelerated.
  • the recognition that sFRP3, or therapeutically effective fragments or derivatives thereof, may be used to accelerate wound healing provides a new mode by which wound healing may be therapeutically accelerated. This finding also gives rise to the prospect of improved medicaments and methods by which wound healing may be accelerated.
  • medicaments or methods of the invention are used to accelerate healing of skin wounds.
  • the medicaments or methods of the invention utilise sFRP3 itself.
  • the sFRP3 to be used may preferably be human sFRP3, as set out in Sequence ID No. 1.
  • sFRP3 therapeutically effective fragments or derivatives of sFRP3
  • Preferred means by which such acceleration may be assessed (and quantified if required) are considered elsewhere in the specification.
  • therapeutically effective derivatives suitable for use in the medicaments or methods of the invention may be derived either from sFRP3 itself, or from therapeutically effective fragments of sFRP3.
  • Preferred fragments or derivatives of sFRP3 for use in the medicaments and methods of the invention may be those based on human sFRP3, the amino acid sequence of which is shown in Sequence ID No. 1.
  • the biological activity of sFRP3 has frequently been ascribed to the CRD of sFRP3 and/or to areas of the C- terminal domain of sFRP3.
  • preferred therapeutically effective fragments of sFRP3 for use in medicaments or methods of the invention may be fragments comprising all, or part, of the CRD (as shown in Sequence ID No. 2), or of the C-terminal domain of sFRP3.
  • a therapeutically effective fragment or derivative of sFRP3 may be a fragment or derivative that is able to accelerate the rate of healing of a treated wound by at least 10% compared to the rate of healing of a comparable untreated or control wound.
  • a therapeutically effective fragment or derivative of sFRP3 may be capable of accelerating healing by at least 20%, more preferably at least 50%, even more preferably at least 75% and yet more preferably of accelerating healing by at least 90% compared to the rate of healing of an untreated or control wound.
  • a most preferred therapeutically effective fragment or derivative of sFRP3 may be capable of accelerating the healing of a wound by 100% or more, compared to the rate of healing of an untreated or control wound.
  • a therapeutically effective fragment or derivative of sFRP3 may be a fragment or derivative that is able to accelerate the rate of contraction of a treated wound by at least 10% compared to the rate of contraction of a comparable untreated or control wound.
  • a therapeutically effective fragment or derivative of sFRP3 may be capable of accelerating contraction by at least 20%, more preferably at least 50%, even more preferably at least 75% and yet more preferably of accelerating contraction by at least 90% compared to the rate of contraction of an untreated or control wound.
  • a most preferred therapeutically effective fragment or derivative of sFRP3 may be capable of accelerating the rate of contraction of a treated wound by 100% or more, compared to the rate of contraction of an untreated or control wound.
  • therapeutically effective fragments or derivatives of sFRP3 suitable for use in the medicaments or methods of the invention may be those able to decrease the area or width of a wound.
  • Suitable therapeutically effective fragments or derivatives of sFRP3 may be those capable of increasing the rate of granulation tissue formation.
  • a therapeutically effective fragment or derivative of sFRP3 may be one that is capable of accelerating the healing of a wound to which the fragment or derivative of sFRP3 is added.
  • Suitable therapeutically effective amounts of sFRP3, as well as suitable therapeutically effective fragments or derivatives of sFRP3, are considered elsewhere in the specification.
  • Therapeutically effective fragments of sFRP3 suitable for use in accordance with the present invention may comprise 10 or more amino acid residues from Sequence ID No. 1, preferably up to 100 amino acid residues, more preferably up to 200 amino acid residues, and even more preferably up to 300 amino acid residues. Fragments suitable for use in the medicaments and methods of the present invention include those comprising up to 324 amino acids residues of Sequence ID No. 1.
  • Preferred therapeutically effective fragments of sFRP3 suitable for use in the medicaments or methods of the invention include those comprising some, or all, of the CRD domain of sFRP3 (shown in Sequence ID No. 2), and/or those comprising some, or all, of the C-terminal domain of sFRP3.
  • Therapeutically effective fragments of sFRP3 suitable for use in accordance with the present invention may comprise up to 10 contiguous amino acid residues from Sequence ID No. 2, preferably up to 50 contiguous amino acid residues, more preferably up to 100 contiguous amino acid residues, and even more preferably up to 124 contiguous amino acid residues from Sequence ID No. 2.
  • Preferred fragments may include amino acid residues involved in binding of sFRP3 to its biological targets.
  • Preferred therapeutically effective fragments or derivatives of sFRP3 will be those that incorporate a WNT-binding region of sFRP3 (either in whole or in part), such as the CRD of sFRP3, or C-terminal domain of sFRP3.
  • Preferred fragments of sFRP3 for use in the medicaments or methods of the invention may include those comprising at least five of the conserved cysteine residues in the CRD of sFRP3, and preferably may comprise at least six, seven, eight, nine or all ten of these residues. It will be appreciated that it is the three dimensional structure of sFRP3 that is important in considering its binding to biological targets, and that accordingly suitable fragments may be selected based upon their ability to assume the requisite three dimensional conformation necessary for target binding.
  • the invention provides the use of an agent comprising a therapeutically effective portion of the CRD of sFRP3 in the manufacture of a medicament for the acceleration of healing.
  • the therapeutically effective portion of the CRD of sFRP3 may preferably be the entire CRD, as shown in Sequence ID No. 2.
  • Such medicaments may be used in the treatment of wounds of the types described elsewhere in the specification.
  • Medicaments according to this embodiment of the invention may be formulated and/or administered in accordance with information provided throughout the present disclosure.
  • peptides comprising all or part of sFRP3 (as defined by Sequence ID No. 1) represent preferred agents for use in accordance with the present invention, it will be recognised that there are contexts in which the sensitivity of peptides to degradation may be disadvantageous. There are many known techniques by which peptide derivatives may be produced that have greater resistance to degradation than do the original peptides from which they are derived. Such derivatives may represent preferred active agents suitable for use in accordance with the invention.
  • Preferred therapeutically effective derivatives of sFRP3 for use in the medicaments or methods of the invention may include derivatives corresponding to all or part of the CRD of sFRP3, or to the C-terminal domain of sFRP3, as considered with reference to therapeutically effective fragments above. Therapeutically effective derivatives of such fragments represent preferred derivatives in accordance with the present invention.
  • the medicaments or methods of the invention may make use of therapeutically effective peptides derivable from the CRD of sFRP3.
  • Suitable therapeutically effective derivatives of sFRP3 for use in the medicaments or methods of the invention may be selected from the group consisting of: therapeutically effective derivatives based on the WNT-binding domain of sFRP3; therapeutically effective derivatives based on the pharmacophore of sFRP3; therapeutically effective derivatives based on glycosylated residues of sFRP3; therapeutically effective peptoid derivatives of sFRP3 or its fragments; therapeutically effective D-amino acid derivatives of sFRP3 or its fragments; therapeutically effective peptidomimetics based on sFRP3 or its fragments; therapeutically effective peptide analogues of sFRP3 or its fragments; therapeutically effective pseudopeptides based on sFRP3 or its fragments; therapeutically effective retro-inverso peptides based on sFRP3 or its fragments; therapeutically effective depsipeptide derivatives based on sFRP3 or its fragments;
  • Peptoid derivatives may be expected to have greater resistance to degradation than do peptide agents of the invention, whilst retaining the same ability to accelerate wound healing. Suitable peptoid derivatives may be readily designed from knowledge of sFRP3's sequence and structure (and in particular the sequence and structure of the WNT- binding domain). Commercially available software and well-established protocols may be used to develop peptoid derivatives suitable for use in accordance with the invention. It will be appreciated that the therapeutic effectiveness of peptoid and other derivatives may be investigated using the same techniques that allow assessment of therapeutic effectiveness of peptide fragments.
  • Retropeptoids (based on sFRP3 or its therapeutically effective fragments) in which all amino acids are replaced by peptoid residues in reversed order may also be used in the medicaments or methods of the invention to accelerate wound healing.
  • a retropeptoid may be expected to bind to its binding partner in the opposite direction to the naturally occurring peptide or a peptoid-peptide hybrid containing one peptoid residue.
  • D-amino acid forms of sFRP3 or its therapeutically effective fragments also confer the requisite ability to accelerate wound healing.
  • the order of the amino acid residues comprising the derivative is reversed as compared to those in the original peptide.
  • the preparation of derivatives using D-amino acids rather than L-amino acids greatly decreases any unwanted breakdown of such an agent by normal metabolic processes, decreasing the amounts of agent which need to be administered, along with the frequency of its administration.
  • derivatives suitable for use in the medicaments and methods of the invention clearly include both those derived from full length sFRP3 and those derived from therapeutically effective fragments of sFRP3.
  • Derivatives of sFRP3, suitable for use in the medicaments or methods of the invention also include peptide derivatives capable of increasing the rate of the healing of wounds.
  • Such peptide derivatives may be based on sFRP3, or on fragments thereof, but may comprise alterations or substitutions of the naturally occurring amino acid sequence. It will be appreciated that amino acid residues involved in the binding of sFRP3 to WNTs may be retained in peptide derivatives for use in accordance with the invention, but that amino acid residues that are not involved in binding to WNTs may be substituted without adversely effecting the activity of such derivatives.
  • the group of therapeutically effective derivatives of sFRP3 also encompasses therapeutically effective small molecule mimics of sFRP3, or its fragments.
  • small molecule mimics of sFRP3 encompass any small molecule chemical entities that are able to mimic sFRP3 activity, and thereby accelerate wound healing, for example by inhibiting WNT signalling. It may be preferred that small molecule mimics of sFRP3 are organic molecules with a weight less than 33OkDa, and preferably with a weight less than IkDa.
  • Suitable small molecule mimics of sFRP3 that may be used in the medicaments or methods of the invention include those that are able to increase the rate of contraction of a wound to which they are provided.
  • a therapeutically effective derivative of sFRP3 suitable for use in accordance with the present invention may share at least 10% homology with Sequence ID No. 1, preferably at least 25% homology, more preferably at least 50% homology, and even more preferably at least 75% homology. Particularly preferred derivatives may share at least 80%, 85%, 90%, 95% or greater homology with Sequence ID No. 1.
  • Therapeutically effective derivatives of sFRP3 suitable for use in accordance with the present invention may share at least 10% identity with Sequence ID No. 1, preferably at least 25% identity, more preferably at least 50% identity, and even more preferably at least 75% identity. Particularly preferred derivatives may share at least 80%, 85%, 90%, 95% or greater identity with Sequence ID No. 1.
  • a therapeutically effective amount of sFRP3, or a fragment or derivative thereof is any amount of sFRP3, or a therapeutically effective fragment or derivative thereof, which is able to accelerate the healing of a wound.
  • a therapeutically effective amount of sFRP3, or a fragment or derivative thereof is preferably an amount of sFRP3, or a fragment or derivative thereof, which is able to accelerate the healing of a wound to which the sFRP3, or fragment or derivative, is administered.
  • a therapeutically effective amount of a medicament of the invention is any amount of a medicament of the invention that is able to accelerate the healing of a wound. This acceleration of healing may preferably be achieved in a wound to which the medicament of the invention is administered.
  • a therapeutically effective amount of sFRP3, or a fragment or derivative thereof, or a therapeutically effective amount of a medicament of the invention may preferably be an amount that is effective to accelerate healing of a treated wound by at least 10% compared to the rate of healing of a comparable untreated or control wound.
  • a therapeutically effective amount of sFRP3, or a fragment or derivative thereof, or of a medicament of the invention may be an amount capable of accelerating healing by at least 20%, more preferably at least 50%, even more preferably at least 75% and yet more preferably of accelerating healing by at least 90% compared to the rate of healing of an untreated or control wound.
  • a most preferred therapeutically effective amount of sFRP3, or a fragment or derivative thereof, or of a medicament of the invention may be capable of accelerating the healing of a wound by 100% or more, compared to the rate of healing of an untreated or control wound.
  • sFRP3, or its therapeutically effective fragments or derivatives is also able to inhibit scar formation.
  • a preferred therapeutically effective amount of sFRP3, or its fragments or derivatives may be an amount that is able to accelerate wound healing, and also to inhibit scarring.
  • a therapeutically effective amount of sFRP3 to be used in a single incidence of treatment may preferably be between O.lng and 1500ng per wound centimetre, more preferably between Ing and lOOOng per wound centimetre, even more preferably between Ing and 500ng per wound centimetre, and still more preferably between Ing and IOng per wound centimetre.
  • a most preferred therapeutically effective amount of sFRP3 that may be used in a single incidence of treatment to accelerate healing in accordance with the present invention may be determined with reference to the type of wound healing of which is to be accelerated.
  • a most preferred therapeutically effective amount to be administered to a centimetre of an incisional wound in a single incidence of treatment may be approximately Ing of sFRP3 per wound centimetre, whilst a most preferred therapeutically effective amount to be administered to a centimetre of an excisional wound in a single incidence of treatment may be approximately IOng.
  • therapeutically effective fragments or derivatives of sFRP3 may have weights that differ significantly from that of sFRP3.
  • guidance as to suitable therapeutically effective amounts of the fragments or variants to be employed in the medicaments or methods of the invention may be derived from the numbers of moles of sFRP3 provided by the therapeutically effective amounts of sFRP3 considered above.
  • a therapeutically effective amount of sFRP3, or a fragment or derivative thereof, to be used in a single incidence of treatment may preferably be between 2.6fml and 40pmol per wound centimetre, more preferably between 0.02pmol and 26pmol per wound centimetre, even more preferably between 0.02pmol and 13pmol per wound centimetre, and still more preferably between 0.02pmol and 0.26pmol per wound centimetre.
  • a most preferred therapeutically effective amount of a fragment or derivative of sFRP3 to be administered to a centimetre of an incisional wound in a single incidence of treatment may be approximately 0.02pmol, whilst a most preferred therapeutically effective amount to be administered to a centimetre of an excisional wound in a single incidence of treatment may be approximately 0.26pmol.
  • the inventors believe that the total amount of sFRP3 to be provided to a centimetre of wound in a 24 hour period may preferably be between 0.2ng and 3000ng per wound centimetre, more preferably between 2ng and 2000ng per wound centimetre, even more preferably between 2ng and lOOOng per wound centimetre, and still more preferably between 2ng and 20ng per wound centimetre.
  • the total amount of sFRP3, or a fragment or derivative thereof, to be provided per centimetre of wound in a 24 hour period may preferably be between 5.2fml and 80pmol per wound centimetre, more preferably between 0.04pmol and 52pmol per wound centimetre, even more preferably between 0.04pmol and 26pmol per wound centimetre, and still more preferably between 0.04pmol and 0.52pmol per wound centimetre.
  • These suggested amounts may represent therapeutically effective amounts for administration to a centimetre of wound over an entire course of treatment.
  • a fragment or derivative of sFRP3 comprises a different number of receptor binding sites to the number of receptor binding sites found in native sFRP3, this may alter the number of moles of such a fragment or derivative required in order to provide a therapeutically effective amount.
  • the amount of the derivative that will be needed to provide a therapeutically effective amount will generally be half of the amount(s) suggested above.
  • Other such variations will be readily apparent to the skilled person.
  • the amount of an active compound or agent of the invention to be provided via topical administration may be altered depending on permeability of the tissue or organ to which the topical composition is administered.
  • Such an increased amount of sFRP3, or a fragment or derivative thereof may still represent a therapeutically effective amount, if the amount of the agent taken up into the tissue or organ where wound healing is to be accelerated is therapeutically effective (i.e. if a therapeutically effective amount permeates the tissue or organ where wound healing is to be accelerated, irrespective of the fact that a larger amount of the agent may remain on the surface of, and unable to penetrate, the tissue or organ being treated).
  • this volume may be administered to either one or both of the margins of a wound to be treated (i.e. in the case of a reference to lOO ⁇ l of a medicament, this may be administered as lOO ⁇ l along one of two wound margins to be joined together, or as 50 ⁇ l to each of the wound margins to be joined together).
  • medicaments of the invention should be taken as encompassing any medicament manufactured in accordance with any aspect or embodiment of the invention.
  • a medicament of the invention will generally constitute a preferred means for putting into practice any method of treatment in accordance with the present invention.
  • Suitable compositions, formulations and routes of delivery that may be used for medicaments of the invention are considered elsewhere in the specification.
  • medicaments of the invention may comprise sFRP3, or a therapeutically effective fragment of sFRP3, or a therapeutically effective derivative of sFRP3 (or of a fragment of sFRP3), or any combination of such agents.
  • a medicament of the invention may comprise a compound able to increase the expression of sFRP3 (or a therapeutically effective fragment or derivative thereof), such as a nucleic acid construct encoding a suitable agent.
  • a medicament of the invention may also comprise a cell comprising such a nucleic acid construct.
  • An “active compound”, for the purposes of the present disclosure, should be taken to be sFRP3, or any therapeutically effective fragment or derivative thereof.
  • An active agent may also encompass a nucleic acid construct encoding sFRP3, or a therapeutically effective fragment or derivative thereof, or a cell comprising such a nucleic acid construct.
  • a “topical medicament”, for the purposes of the present disclosure, is to be construed as a medicament that is applied at a site where it is intended to have its effect. This site may be a wound, or a site where a wound is to be formed.
  • Topical medicaments suitable for use in accordance with the present invention include, but are not limited to, ointments; creams; lotions; gels; sprays; wound dressings capable of releasing active agents to the body; and injectable solutions administered by local injections (e.g. intradermal injections).
  • wounds within the present disclosure should also be taken to encompass sites where wounds are to be formed, since the inventors have found that such sites may benefit from prophylactic treatment using the medicaments or methods of the invention.
  • wounds will primarily be described with reference to skin wounds, which comprise preferred wounds the contraction of which may be promoted in accordance with the present invention.
  • skin wounds which comprise preferred wounds the contraction of which may be promoted in accordance with the present invention.
  • the promotion of wound contraction in accordance with the invention should is not limited to skin wounds. The inventors believe that contraction of wounds may be promoted in wounds of all tissues.
  • Skin wounds include both chronic wounds and acute wounds. Examples of suitable chronic or acute wounds, the healing of which may be accelerated in accordance with the invention, are set out elsewhere in the specification.
  • the medicaments and methods of the invention may be used to accelerate healing of wounds selected from the group consisting of: abrasions; avulsions; crush wounds; incisional wounds; lacerations; punctures; ulcers, abscesses, and missile wounds, all of which may be suffered by the skin (among other tissues or organs).
  • suitable wounds include surgical wounds; pre-tibial lacerations; graft recipient sites; gastrointestinal ulcers, and lung abscesses.
  • Preferred chronic wounds that may be treated with the medicaments or methods of the invention in order to accelerate their healing include ulcers such as, diabetic ulcers, decubitus ulcers, and venous ulcers.
  • a wound the contraction of which is to be accelerated using the medicaments of the invention may be selected from the group consisting of: abrasions, cuts, pressure ulcers stages i) to iv); venous stasis ulcers, ulcers caused by mixed etiologies; lower extremity ulcers; diabetic ulcers; radiation ulcers; arterial ulcers; partial thickness excisions; full thickness excisions; wounds in the immunocompromised, elderly or paraplegics; pre-tibial lacerations; wounds that have been debrided; and surgical wounds (including surgical incisions or excisions).
  • promotion of contraction of wounds may be of particular benefit in cases in which the wound healing response is impaired, inhibited, retarded or otherwise defective as compared to the normal rate of healing.
  • the methods and medicaments of the invention may also be used to promote contraction of wounds in patients that are not subject to an impaired healing response. Illustrative examples of both contexts are set out below. It is well known that dermal injuries in the aged heal more slowly than do those of younger individuals. The aged may therefore particularly benefit from promotion of wound contraction that may be brought about using the medicaments and methods of the invention. There are also many other conditions or disorders that are associated with a delayed or otherwise impaired wound healing response.
  • patients with diabetes For example patients with diabetes, patients with polypharmacy (for example as a result of old age), postmenopausal women, patients susceptible to pressure injuries (for example paraplegics), patients with venous disease, clinically obese patients, patients receiving chemotherapy, patients receiving radiotherapy, patients receiving steroid treatment or immunocompromised patients may all suffer from impaired healing.
  • the slower healing response exhibited by such patients may contribute to the development of infections at the site of wounds.
  • the slow wound healing response may also be associated with the formation of chronic wounds, as considered below. Accordingly, it will be appreciated that such patients represent a preferred group that may benefit from increased wound contraction using the methods or medicaments of the invention.
  • Aged or senescent patients may be defined as comprising patients aged 65 years or older, more preferably aged 75 years or older. In the case of female patients, it may be preferred that the medicaments or methods of the invention be provided to postmenopausal patients, to whom they may be of marked benefit.
  • the medicaments or methods of the invention may be utilised to promote contraction of wounds of patients not subject to delayed wound healing. Promoting contraction in this way will give rise to a faster wound healing response than would normally be achieved by such patients in the absence of promoted wound contraction (i.e. will give rise to faster healing than in control wounds). Accordingly the wounds of patients treated in this manner may be induced to heal more rapidly.
  • promotion of wound contraction in this manner can help reduce time spent in convalescence, and can thus benefit productivity. Accordingly, promotion of the contraction of wounds of healthy patients is a preferred embodiment of all aspects of the present invention.
  • a chronic wound may be defined as any wound that does not show any healing tendency within eight weeks of formation when subject to appropriate (conventional) therapeutic treatment.
  • Acute wounds may be any wound other than a chronic wound.
  • Chronic wounds that may benefit from promoted contraction provided by the medicaments or methods of the invention may be selected from the group comprising: leg ulcers; venous ulcers; diabetic ulcers; bed sores; decubitus ulcers; foot ulcers; and pressure ulcers.
  • leg ulcers venous ulcers
  • diabetic ulcers diabetic ulcers
  • bed sores decubitus ulcers
  • foot ulcers foot ulcers
  • pressure ulcers a group comprising: leg ulcers; venous ulcers; diabetic ulcers; bed sores; decubitus ulcers; foot ulcers; and pressure ulcers.
  • Incidences of wound infection are also much increased in chronic, as opposed to acute, wounds.
  • the ability of wound contraction promoted using the medicaments or methods of the invention to decrease the "open area" of treated wounds may be of benefit in reducing the possible ingress of pathogens, and also reducing fluid loss from the damaged tissue.
  • a preferred use of the medicaments or methods of the invention lies in their use to promote contraction of wounds prior to grafting or suturing to close the treated wound. This use is suitable for both chronic and acute wounds, hi general the treatment of wounds using the medicaments or methods of the invention may be stopped prior to grafting or suturing, so that the grafted material, or sutured tissue, is not subject to further contraction once in situ.
  • Chronic wounds are also subject to many disadvantages that are not generally associated with acute wounds. For example, chronic wounds frequently expand beyond the limits of the original wounded area. This may arise as a result of infection (which may increase the damage around the margins of the wound, thereby leading to expansion) or through maceration of the tissue surrounding the wound (typically as a consequence of increased liquid loss through the chronic wound).
  • the propensity for chronic wounds to expand beyond the boundary of the original injury means that such wounds are frequently of great surface area.
  • wound contraction promoted using the medicaments or methods of the invention may be useful in reducing the area of chronic wounds, and may help to prevent the expansion of such wounds.
  • Pretibial lacerations are acute wounds of the leg that are very frequently slow to heal, and which frequently give rise to the development of leg ulcers.
  • Existing treatments used for pretibial lacerations include the use of surgical procedures (such as the use of skin grafts and flaps) in an attempt to heal the wound before chronic wound development.
  • Pretibial lacerations constitute acute wounds that may particularly benefit from treatment with the medicaments and methods of the invention, in order to promote contraction of the wound, thereby hastening healing and reducing incidences of chronic wound formation.
  • Examples of specific wounds, other than those of the skin, which may benefit from accelerated healing of wounds in accordance with the present invention include, but are not limited to, those selected from the group consisting of: gastrointestinal ulcers, and lung abscesses.
  • medicaments or methods of the invention may be used to effectively promote the contraction of various gastrointestinal ulcers, such as peptic, gastric, duodenal and esophageal ulcers. These ulcers may be considered for the purposes of the present invention to constitute chronic wounds.
  • Lung abscesses may arise as a result of inflamed pleura, which lead to the formation of a pus-filled cavity and the subsequent loss of lung parenchyma.
  • treatment of lung abscesses tends to involve the use of antibiotics to address the infection, but such treatment may still leave a hole in the lung that is eventually repaired to leave a dense scar. The presence of such a scar can significantly decrease lung function.
  • promotion of wound contraction using the medicaments and methods of the invention would be beneficial to speed up the repair process, and to decrease the size of the scar formed by decreasing wound size.
  • the medicaments and methods of the invention be used to accelerate healing of skin wounds. This may be through treatment of such wounds themselves and/or the sites where such wounds are to be formed.
  • a "treated wound” in the context of the present disclosure is any wound that has been provided with a therapeutically effective amount of sFRP3 (or a therapeutically effective fragment or derivative thereof) whether by a medicament of the invention, or in accordance with a method of treatment of the invention.
  • Control-treated wounds and “untreated wounds” in the present context are respectively wounds treated with a relevant control, and wounds that have not been treated before, or during, healing.
  • Control wounds will not be treated with a medicament of the invention, and preferably will not be treated with a therapeutically effective amount of an active compound. That said, wounds treated with medicaments known from the prior art may constitute suitable control wounds for comparative purposes (for example to illustrate increased efficiency or effectiveness of medicaments of the invention as compared to those already known).
  • a "diluent control-treated wound” will be an untreated wound to which a control diluent has been administered, and a “na ⁇ ve control” will be an untreated wound made without administration of an active compound or a suitable control diluent, and left to heal without therapeutic intervention.
  • wound centimetre constitutes a unit by which the size of a wound to be treated may be measured.
  • a wound centimetre may be taken to comprise any square centimetre of a body surface that is wounded in whole or in part.
  • a wound of two centimetres length and one centimetre width i.e. with a total surface area of two centimetres 2
  • a wound having a length of two centimetres and a width of two centimetres i.e. a total surface area of four centimetres 2
  • a linear wound of two centimetres length, but of negligible width i.e. with negligible surface area
  • the size of a wound in wound centimetres should generally be assessed when the wound is in its relaxed state (i.e. when the body site bearing the wounded area is in the position adopted when the body is at rest). In the case of skin wounds, the size of the wound should be assessed when the skin is not subject to external tension.
  • An inch of wound may be similarly defined, save that the relevant units of length or area are measured in inches rather than centimetres.
  • a centimetre or inch of wounding may thus provide a unit by which the size of a wound to be treated may be measured, and the required amount of a medicament of the invention (or of an active compound administered in accordance with a method of treatment of the invention) may be determined.
  • Acceleration of wound healing achieved using the medicaments or methods of the invention may preferably lead to a treated wound healing at a rate at least 5% faster than an untreated or control wound, preferably at a rate at least 10% faster, more preferably at least 15%, 20% or 25% faster; yet more preferably at least 50% faster, still more preferably at least 75% faster, and most preferably 100% (or more) faster.
  • the acceleration of healing may be brought about through promotion of wound contraction. Suitable methods by which promotion of contraction of wounds may be quantified to assess improvements in the rate of healing are described elsewhere in the specification.
  • Measurements that may be of use in assessing the rate of contraction of a wound include the rate at which the area or width of a wound decreases. Accelerated healing achieved using the medicaments or methods of the invention may preferably lead to a treated wound in which the wound's area or width decreases at a rate at least 5% faster than an untreated or control treated wound, preferably at a rate at least 10% faster, more preferably at least 15%, 20% or 25% faster; yet more preferably at least 50% faster, still more preferably at least 75% faster, and most preferably 100% (or more) faster. Such accelerated healing may be brought about by promotion of wound contraction. Suitable methods by which wound width may be measured in order to assess promotion of contraction of wounds are described elsewhere in the specification.
  • Acceleration of healing using the medicaments or methods of the invention may also give rise to a treated wound having an increased "healing age" when compared with an untreated or control treated wound. Such an increase in healing age may be assessed macroscopically, visually or clinically to determine maturity of the treated wound compared to a suitable untreated or control wound.
  • a wound treated with the medicaments or methods of the invention may preferably have a healing age that is 1, 2, 3, 4, 5 or more days greater than that of an untreated, or control treated wound of the same chronological age.
  • a preferred measurement that may be used in assessing acceleration of wound healing is the rate at which the area of a wound decreases. Acceleration of wound healing using the medicaments or methods of the invention may preferably lead to a treated wound in which wound area decreases at a rate at least 5% faster than a control or untreated wound, preferably at a rate at least 10% faster, more preferably at least 15%, 20% or 25% faster; yet more preferably at least 50% faster, still more preferably at least 75% faster, and most preferably 100% (or more) faster.
  • the rate at which the area of a treated wound decreases may be compared with control or untreated wounds, or with reference data regarding the rate at which areas of control or untreated wounds decrease in order to assess any differences in the rates observed.
  • a therapeutically effective amount of sFRP3, or a fragment or derivative thereof, in the context of the present invention may preferably be an amount of sFRP3, or a fragment or derivative thereof, that is able to give rise to a treated wound in which wound area decreases at a rate at least 5% faster than a control or untreated wound, preferably at a rate at least 10% faster, more preferably at least 15%, 20% or 25% faster; yet more preferably at least 50% faster, still more preferably at least 75% faster, and most preferably 100% (or more) faster.
  • Such a therapeutically effective amount of sFRP3, or its fragments or derivatives may be provided in a therapeutically effective amount of a medicament of the invention.
  • the area of a wound may be assessed macroscopically or microscopically in order to determine the rate of wound healing. Suitable assessments of wound area may, for example, utilise photographs or tracings of the wound margins. These may be considered over time, or with reference to standard comparison data, to assess whether or not wound area is being therapeutically decreased.
  • the area of a wound assessed in this way should be distinguished from the degree of re- epithelialisation observed in the wound. Since the re-epithelialisation response will lead to overgrowth of the wound margins his may render assessment of wound area difficult. In general it may be preferred to assess wound area microscopically.
  • a preferred measurement that may be used in assessing acceleration of wound healing is the rate at which the width of a wound decreases.
  • Acceleration of wound healing using the medicaments or methods of the invention may preferably lead to a treated wound in ⁇ which wound width decreases at a rate at least 5% faster than a control or untreated wound, preferably at a rate at least 10% faster, more preferably at least 15%, 20% or 25% faster; yet more preferably at least 50% faster, still more preferably at least 75% faster, and most preferably 100% (or more) faster.
  • wound width may be measured in order to assess promotion of wound contraction. It may generally be preferred that wound width be assessed microscopically, using histological slides.
  • a preferred protocol for the assessment of wound width in full thickness wounds involves assessing the width of the wound at its mid point (i.e. a point mid way into the depth of the wound). The mid point is preferably in the dermis of the wound, well below the level at which re-epithelialisation occurs. Measurement of wound width at this point may avoid any inaccuracies that may otherwise be arise if wound width is not clearly distinguished from the portion of a wound that has not undergone re-epithelialisation..
  • sFRP3 or a fragment or derivative thereof capable of increasing wound contraction and/or granulation tissue formation represent preferred therapeutically effective amounts suitable for use in accordance with the present invention. It will also be appreciated that acceleration of wound healing may thus also be assessed with reference to the amount of wound contraction taking place, and alternatively or additionally, with reference to the amount of granulation tissue being formed.
  • Wound contraction is generally accepted to be dependent on the action of fibroblasts located both at the periphery of the wound and within the wound. Contraction is linked to fibroblast proliferation rate and connection of these cells to extracellular matrix components.
  • the increase in the rate of wound contraction that may be achieved using the medicaments or methods of the invention should be distinguished from the rate at which the wound is covered with a new epithelial layer (in the skin a new epidermis), which is related to the rate of re-epithelialisation.
  • Granulation tissue formation arises primarily as a result of the influx of cells, such as fibroblasts, from the unwounded tissue surrounding the wound. The formation of granulation tissue plays an important role in filling the wound defect.
  • the width of a wound (and hence the rate at which the width of a wound decreases, and so the acceleration, or otherwise, of wound healing) may be assessed macroscopically, or microscopically.
  • wound width may be measured at a standardised "reference" point within the wound.
  • the inventors have found that measurements taken midway through the depth of the wound allow for an accurate and reproducible assessment of wound width.
  • Suitable image analysis software may aid the assessment of wound width in this manner.
  • Macroscopic assessment of wound width may either be performed directly (i.e. with measurements taken directly from a wound), or indirectly, in which case measurements may be taken using representations of the wound, such as photographs, traced outlines, mouldings, or the like.
  • Image analysis software may be useful in the macroscopic assessment of wound width, particularly as assessed from photographs. In assessing wound width microscopically it is important that the width of a wound is differentiated from the degree of re-epithelialisation of the wound.
  • Preferred routes of administration by which therapeutically effective amounts of sFRP3, or a fragment or derivative thereof, may be provided to a wound the healing of which it is desired to accelerate, are discussed more fully elsewhere in the specification. However, it may generally be preferred that sFRP3, or its fragments or derivatives, are provided by local administration to a wound, healing of which is to be accelerated. Suitable methods by which such local administration may be achieved will depend on the identity of the tissue in question, and may also be influenced by the size or location of the wound. Preferred routes of administration may include local injection (for example intradermal injection in the case where it is wished to accelerate healing of skin wounds). Other suitable means of administration include the use of topical medicaments such as sprays; powders; drops (e.g. for the ear or eye); ointments or creams; or release from local devices e.g. stents, implants, polymers, wound dressings, sutures or the like.
  • topical medicaments such as sprays; powders; drops (e.g. for
  • medicaments of the invention may be formulated and manufactured in any form that allows for the medicament to be administered to a patient such that a therapeutically effective amount of the active compound is provided to a wound the healing of which is to be accelerated.
  • certain routes of administration normally associated with systemic administration may also be suitable for topical administration of active compounds to an otherwise "inaccessible" wound in which it is desired to accelerate healing (for example, inhalation or intranasal administration of active compounds may be of use in accelerating healing of wounds of the respiratory system).
  • Medicaments of the invention may preferably be provided in the form of one of more dosage units providing a therapeutically effective amount (or a known fraction or multiple of a therapeutically effective amount) of an active compound.
  • Methods of preparing such dosage units will be well known to the skilled person; for example see Remington's Pharmaceutical Sciences 18 th Ed. (1990).
  • the medicaments of the invention should be taken to encompass any composition, material or device from which sFRP3, or a fragment or derivative thereof, may be provided to a wound in a therapeutically effective quantity (as defined elsewhere in the specification).
  • medicaments of the invention that comprise a solid material
  • the medicament be formulated such that a predetermined area of the medicament provides a therapeutically effective amount of an active compound to a wound to which the solid material is applied. This will be particularly beneficial in the case of solid medicaments, such as wound dressings, that are to be placed over a site where wound healing is to be accelerated.
  • Medicaments of the invention may be provided in the form of discrete dosage units capable of providing a therapeutically effective amount of an active compound.
  • a suitable dosage unit in accordance with this embodiment of the invention may comprise a sufficient amount of a medicament of the invention to accelerate the healing of a given length or area of a wound.
  • a suitable dosage unit may comprise sufficient of a medicament of the invention to accelerate the healing of one centimetre of a wound.
  • a suitable dosage unit may comprise sufficient of a medicament of the invention to accelerate the healing of one inch of a wound.
  • medicaments of the invention may be formulated to provide single dosage units or to provide multiple dosage units, as required.
  • a medicament of the invention may be packaged to provide one or more dosage units.
  • Each dosage unit may provide a known fraction or multiple of a therapeutically effective amount of sFRP3, or a fragment or derivative thereof.
  • Suitable forms in which such discrete dosage units may be provided can be selected with reference to the nature of the medicament to be administered.
  • medicaments of the invention comprising injectable solutions may be provided in the form of vials or pre-filled syringes comprising one or more dosage units.
  • Other liquid medicaments in accordance with the invention such as gels, creams, ointments, irrigation fluids or the like, may be provided in the form of tubes, sachets, cartons or blister packs comprising one or more dosage units.
  • Solid medicaments of the invention may readily be formulated such that a given area of the solid medicament is capable of providing sufficient sFRP3, or a fragment or derivative thereof, to accelerate the healing of a matching-sized area of wound.
  • a solid medicament of the invention may be cut to the required size and/or shape to cover a wound and the medicament will release a therapeutically effective amount of an active compound sufficient to accelerate the healing of the wound.
  • an active compound released in this manner may be taken up via the wound, where the barrier function of the skin is impaired, but that the therapeutic effects of such a compound may actually occur as a result of its activity in the surrounding skin.
  • a solid medicament in accordance with the invention may alternatively be cut to a size that covers both a wound to be treated, and an area of normal skin surrounding the wound.
  • Solid formulations for use in medicaments of the invention may, or may not, be contained in a containment membrane or coating, microspheres, microgranules or microcapsules.
  • the materials for such containment membranes or coatings may be selected from any of a variety of biodegradable natural or synthetic materials. Suitable materials may provide resistance to diffusion of the active compound.
  • suitable materials for use in containment membranes or coatings be selected such that they allow sustained release of the active compound to the wound healing of which is to be accelerated. Examples of techniques by which this may be achieved will be well known to those skilled in the art, and will include the use of alternating layers of a suitable containment membrane or coating with layers of a carrier incorporating the active compound.
  • Suitable materials for use in containment membranes or coatings will generally degrade or be broken down over a period of time, thereby exposing the carrier, and allowing therapeutic release of the active compound from the carrier to the wound.
  • the degradation or breakdown of suitable containment membranes or coatings may be caused by prolonged exposure to a wound. Factors that may mediate the degradation of such containment membranes or coatings will generally be the same as those that may cause the release of the active compound from the carrier.
  • Such degradation or breakdown may typically be caused by moisture associated with the wound, or by the activity of enzymes active during wound healing.
  • Suitable solid formulations that may be used in medicaments of the invention may, for example, be selected from: powders; sprays, crystals; microneedles; solid compositions comprising microparticles, microspheres, microgranules, microcapsules, nanoparticles or liposomes; sutures and wound dressings.
  • Suitable liquid formulations that may be used in medicaments of the invention may, for example, be selected from: gels: thermosetting gels; creams; ointments; sprays; injectable solutions; irrigation solutions; other solutions of active compounds; and liquid compositions comprising microparticles, nanoparticles or liposomes.
  • suitable solid formulations may be applied throughout the dressing, and particularly to the surface of the dressing that is to be placed into contact with the wound.
  • Solid formulations may be applied as a coating that may be applied to all the material of the dressing, or may be applied to discrete portions of the dressing (for instance the surface of the dressing that is to be placed in contact with the wound).
  • Solid formulations may also be provided as granules, microgranules, microparticles, nanoparticles or liposomes adhered to the material of the dressing.
  • acceleration of wound healing using the medicaments or method of the invention may give rise to a healing time 1 day, 2 days, or 3 days faster than that occurring in a control-treated or untreated wound.
  • Healing time may be calculated as the time elapsing between formation of a wound and complete closure of the wound (i.e. the point at which wound width becomes zero, or imperceptible).
  • the acceleration of wound healing in accordance with the invention may give rise to a healing time that is at least 4 days, 5 days or 6 days faster than that occurring in a control- treated or untreated wound.
  • acceleration of wound healing may give rise to a healing time that is at least 7 days, 8 days or 9 days faster than that occurring in a control-treated or untreated wound, and most preferably acceleration of wound healing may give rise to a time to wound closure that is at least 10 days or greater than that occurring in a control-treated or untreated wound.
  • the inventors have found that the medicaments and methods of the invention are able to promote accelerated wound healing when used either prior to the formation of a wound, or when used after a wound has already been formed.
  • prophylactic use for the purposes of the present disclosure.
  • the prophylactic use of agents in accordance with the invention to accelerate wound healing is a preferred mode of use in accordance with the invention. It will be appreciated that such use is most suitable in the case where the time and location of prospective wound formation is known, and may be particularly suitable for accelerating healing of wounds associated with surgical procedures. However, prophylactic use of the medicaments or methods of the invention may also be of use in situations where there is an increased likelihood of wounding occurring.
  • the inventors have found that administration of agents in accordance with the invention immediately prior to formation of a wound (e.g. in the hour preceding wounding, or preferably in the forty minutes or thirty minutes preceding wounding, and more preferably in the ten minutes preceding wounding) is highly effective, though administration at earlier times (e.g. up to 24 or 48 hours before wounding) may also beneficially accelerate wound healing.
  • the prophylactic use of methods and medicaments of the invention is a preferred embodiment of the invention, and is particularly preferred in the event that it is wished to accelerate healing of surgical wounds.
  • Injection and particularly intradermal injection, constitutes a preferred manner in which the medicaments of the invention may be administered (or the methods of the invention effected), as considered elsewhere in the specification.
  • a medicament of the invention be administered by intradermal injection to a site where wounding will take place. If the medicament is administered only a short time prior to wound, then intradermal injection of this type will typically lead to the formation of a raised bleb which will remain at the time of wounding. A wound may then be formed through the bleb. Wounds formed in this way will benefit from accelerated healing in accordance with the present invention.
  • blebs formed by intradermal injection of medicaments of the invention may be allowed to resolve before a wound is formed.
  • the medicaments and methods of the invention may also be used to accelerate wound healing after a wound has been formed. This use will be the use generally adopted in respect of accidental wounds (and indeed most wounds formed other than in association with a surgical procedure).
  • medicaments in accordance with the invention When used to treat existing wounds medicaments in accordance with the invention may be applied along the margins of a wound to be treated. Preferably, medicaments of the invention may be injected along the margins of wounds to be treated. Injection in this manner also constitutes a preferred route of administration in accordance with the methods of treatment of the invention. In the case of skin wounds it is preferred that the route of injection selected is intradermal injection.
  • topical medicaments may also be applied to the margins of wounds the healing of which it is wished to accelerate.
  • suitable medicaments may be administered to the wound defect itself, from where they may permeate the surrounding tissue, and thereby achieve their effect.
  • the medicaments or methods of the invention may help to accelerate healing of a wound if used at any time up until the healing process is complete (for example even if administered to a partially healed wound the medicaments of the invention may usefully accelerate healing of the un-healed portion of the wound in a manner that will therapeutically decrease the time until the whole wound is fully healed).
  • a number of factors may be considered in determining the "window" in which medicaments or methods of the invention may be beneficially employed to accelerate healing of wounds.
  • wound in question may include: i) the nature of the wound in question (for example: is the wound at a site that is generally subject to "fast” or “slow” healing?); ii) the severity of the wound (what is the extent of the damage that has occurred?); and iii) the size of the damaged area.
  • the methods or medicaments of the invention may still be effective to accelerate healing of the wound even if administered relatively late in the healing response.
  • the medicaments or methods of the invention may preferably be administered within the first one to 24 hours after formation of an acute wound, beneficial acceleration of healing may also be achieved if administered up to ten, or more, days after the wound is formed.
  • Therapeutic acceleration of wound healing may be achieved using only a single administration of the medicaments or methods of the invention. Due to the simplicity of this therapeutic regime it may constitute a preferred use of the medicaments and methods of the invention.
  • medicaments or methods of the invention be used in repeated incidences of therapy.
  • treatment to accelerate wound healing may involve administration of medicaments of the invention on more than once occasion. Use in this manner may be preferred in the case of large wounds, or of wounds that are resistant to treatment, or subject to retarded healing (such as chronic wounds).
  • medicaments of the invention may be administered to a wound as required until therapeutically effective acceleration of healing has been achieved (for example, until the wound has closed, or no further acceleration of healing may be achieved).
  • medicaments of the invention may be administered daily (or on multiple occasions within a given day), or may be administered after a delay of multiple days.
  • sFRP3, or a therapeutically effective fragment or derivative thereof may be administered prior to formation of a wound, and then again after a wound has been formed.
  • the inventors have found that this regime of treatment is particularly effective for the acceleration of wound healing, as demonstrated in the Experimental Results section.
  • Both the first and second administrations of sFRP3, or a therapeutically effective fragment or derivative thereof may be achieved utilising the medicaments of the invention. It may be preferred that the second administration of sFRP3 takes place a day after formation of a wound the healing of which is to be accelerated.
  • medicaments or methods of the invention are utilised both before and after wounding.
  • the medicaments of the invention are administered to a site where they are to have their effect around the time of wounding, or immediately prior to the forming of a wound (for example in the period up to six hours before wounding, and particularly in the period of 10, 20, 40 or 60 minutes prior to wound formation) or the medicaments may be administered at an earlier time before wounding (for example up to 48 hours before a wound is formed).
  • the most preferred times of administration prior to formation of a wound will be determined with reference to a number of factors, including the formulation and route of administration of the selected medicament, the dosage of the medicament to be administered, the size and nature of the wound to be formed, and the biological status of the patient (which may be determined with reference to factors such as the patient's age, health, and predisposition to healing complications) and the half-life of the active compound in the body, (where non-peptide derivatives of sFRP3 may generally be expected to have longer half-lives than sFRP3 or its fragments).
  • the methods or medicaments of the invention may be administered both before and after formation of a wound.
  • administration of an active compound prior to wound formation (and preferably immediately prior to the formation of a wound) followed by administration of an active compound on one or more days following wounding is particularly effective in accelerating wound healing, through promotion of wound contraction
  • an “agent” or “agent of the invention” will be sFRP3, or a therapeutically effective fragment or derivative thereof, or a compound able to promote expression of such an active compound. It will be appreciated that all such suitable agents may be incorporated in medicaments in accordance with the invention, and all may be used in the methods or uses of the invention.
  • the medicaments of the invention represent preferred compositions by which a therapeutically effective amount of an active compound may be administered in order to put the methods of the invention into practice.
  • sFRP3 (as shown in Sequence ID No 1) is a preferred example of an agent, or agent of the invention, as considered elsewhere in the specification.
  • the amount of a medicament of the invention (and hence an agent of the invention) that should be provided to a wound in order that a therapeutically effective amount of an active compound may be administered depends on a number of factors. A number of these are discussed elsewhere in the specification, and include the biological activity and bioavailability of the agent present in the medicament, which in turn depends, among other factors, on the nature of the agent and the mode of administration of ⁇ the medicament. Other factors in determining a suitable therapeutic amount of a medicament may include:
  • the specific wound to be treated e.g. accelerating healing of an acute wound or a chronic wound.
  • the frequency of administration will also be influenced by the above-mentioned factors and particularly the half-life of the chosen agent within the subject being treated.
  • Frequency of administration will depend upon the biological half-life of the agent used.
  • a cream or ointment containing an agent of the invention should be administered to a target tissue such that the concentration of the agent at a wound is maintained at a level suitable to accelerate wound healing. This may be achieved by a single administration of a composition incorporating an agent of the invention or may require administration of such a composition daily or even several times daily.
  • Medicaments of the invention may be administered by any suitable route capable of achieving the desired effect of accelerating wound healing, but it is preferred that the medicaments be administered such that an agent of the invention is provided locally at a wound site or site where a wound is to be formed.
  • Administration and particularly topical administration of the medicaments of the invention may be effected as part of the initial and/or follow up care for the wounded area.
  • the agents of the invention may be provided on a dressing or patch, which may be used to cover a wound, the healing of which is to be accelerated. It will be appreciated that such a dressing or patch used to administer an agent of the invention may preferably be provided in a sterile form.
  • the agents of the invention may be released from a device or implant, or may be used to coat such a device e.g. sutures, or a stent or controlled release device.
  • the vehicle of a composition comprising agents of the invention should be one that is well tolerated by the patient and allows release of the agent to a wound to which the composition is provided.
  • a vehicle is preferably biodegradable, biocompatible, bioresolveable, bioresorbable and/or non-inflammatory. If the composition is to be applied to an existing wound then the pharmaceutically acceptable vehicle will be one that is relatively "mild”.
  • An agent of the invention may be incorporated within a slow or delayed release device. Such devices may, for example, be placed on or inserted under the skin and the agent may be released over days, weeks or even months. Such use may be of particular benefit when the selected agent is a nucleic acid encoding an active compound, or in the case of active compounds prone to degradation.
  • Delayed release devices may be particularly useful for patients requiring protracted treatment with the medicaments or methods of the invention, such as those requiring accelerated healing of chronic wounds. Delayed release devices may be particularly advantageous when used for the administration of an agent or nucleic acid that would otherwise normally require frequent administration (e.g. at least daily administration by other routes).
  • Daily doses of an agent of the invention may be given as a single administration (e.g. a daily application of a topical formulation or a daily injection).
  • the agent of the invention may require administration twice or more times during a day.
  • Each such administration may provide a therapeutically effective amount of the agent, or a known fraction of such a therapeutically effective amount.
  • a slow release device may be used to provide optimal doses of an agent of the invention to a patient without the need to administer repeated doses.
  • a dose of a composition comprising agents of the invention may preferably be sufficient to provide a therapeutically effective amount of an active compound in a single administration.
  • each dose need not in itself provide a therapeutically effective amount of an agent or an active compound, but that a therapeutically effective amount of an agent or active compound may instead be built up through repeated administration of suitable doses.
  • compositions comprising agents of the invention.
  • a pharmaceutical vehicle for administration of an agent of the invention may be a liquid and a suitable pharmaceutical composition would be in the form of a solution.
  • the pharmaceutically acceptable vehicle is a solid and a suitable composition is in the form of a powder or tablet.
  • the agent of the invention may be formulated as a part of a pharmaceutically acceptable patch or other transdermal delivery device capable of providing an agent of the invention to a wound or a site where a wound is to be formed.
  • a solid vehicle can include one or more substances that may also act as flavouring agents, lubricants, solubilizers, suspending agents, fillers, glidants, compression aids, binders or tablet-disintegrating agents; it can also comprise an encapsulating material.
  • the vehicle is a finely divided solid that is in admixture with the finely divided agent of the invention.
  • the agent of the invention is mixed with a vehicle having the necessary compression properties in suitable proportions and compacted in the shape and size desired.
  • the powders and tablets preferably contain up to 99% of the agent of the invention.
  • Suitable solid vehicles include, for example, calcium phosphate, magnesium stearate, talc, sugars, lactose, dextrin, starch, gelatin, cellulose, polyvinylpyrrolidine, low melting waxes and ion exchange resins.
  • Liquid vehicles may be used in preparing solutions, suspensions, emulsions, syrups, elixirs and pressurized compositions.
  • the agent of the invention can be dissolved or suspended in a pharmaceutically acceptable liquid vehicle such as water, an organic solvent, a mixture of both, or pharmaceutically acceptable oils or fats.
  • the liquid vehicle can contain other suitable pharmaceutical additives such as solubilizers, emulsifiers, buffers, preservatives, sweeteners, flavouring agents, suspending agents, thickening agents, colours, viscosity regulators, stabilizers or osmo-regulators.
  • suitable examples of liquid vehicles for oral and parenteral administration include water (partially containing additives as above, e.g.
  • cellulose derivatives preferably sodium carboxymethyl cellulose solution
  • alcohols including monohydric alcohols and polyhydric alcohols, e.g. glycols) and their derivatives, and oils (e.g. fractionated coconut oil and arachis oil).
  • Phosphate buffered saline PBS
  • the vehicle can be an oily ester such as ethyl oleate and isopropyl myristate.
  • Sterile liquid vehicles are useful in sterile liquid form compositions for parenteral administration.
  • the liquid vehicle for pressurized compositions can be halogenated hydrocarbon or other pharmaceutically acceptable propellant.
  • Liquid pharmaceutical compositions which are sterile solutions or suspensions can be utilized by, for example, intramuscular, intrathecal, epidural, intraperitoneal, intradermal, intraadventitial (blood vessels) or subcutaneous injection. Sterile solutions can also be administered intravenously.
  • the agent of the invention may be prepared as a sterile solid composition that maybe dissolved or suspended at the time of administration using sterile water, saline, or other appropriate sterile injectable medium, such as PBS. Vehicles are intended to include necessary and inert binders, suspending agents, lubricants and preservatives.
  • Agents of the invention may be used to accelerate healing of "internal” wounds (i.e. wounds occurring within the body, rather than on an external surface of the body).
  • internal wounds include penetrative wounds that pass through the skin into underlying tissues, and wounds associated with surgical procedures conducted within the body.
  • medicaments or methods of the invention to accelerate healing of internal wounds will necessitate the use of suitable routes of administration, thereby requiring the formulation of the agents of the invention in a manner that allows their delivery to the wound in question.
  • medicaments in accordance with the invention for accelerating healing of wounds in the lungs or other respiratory tissues may be formulated for inhalation.
  • medicaments in accordance with the invention for accelerating healing of wounds in the body cavities may be formulated as a lavage, gel or instillate.
  • compositions comprising agents of the invention and precise therapeutic regimes for administration of such compositions (such as daily doses of the active agent and the frequency of administration).
  • Medicaments or methods of the invention may be used to accelerate wound healing as a monotherapy (e.g. through use of medicaments or methods of the invention alone).
  • the methods or medicaments of the invention may be used in combination with other compounds or treatments for the acceleration of wound healing.
  • Suitable compounds that may be used as parts of such combination therapies will be well known to those skilled in the art.
  • sFRP3, or its fragments or derivatives may be administered at the sites of wounds (or sites where wounds are to be formed) where it is wished to accelerate healing by virtue of cellular expression (commonly referred to as gene therapy).
  • the invention provides a method of accelerating wound healing, the method comprising inducing cellular expression of a therapeutically effective amount of sFRP3, or a fragment or derivative thereof, at a site where healing is to be accelerated.
  • a site may be a wound, or a site where a wound is to be formed, or where there is a heightened likelihood of wound formation occurring.
  • sFRP3 cellular expression of therapeutically effective amounts of sFRP3 may be achieved by manipulating naturally occurring expression of this molecule by cells in the region of the wound to be treated.
  • cells in the region of wound (or site where a wound is to be formed) that is to be treated may be induced to express sFRP3, or therapeutically effective fragments or derivatives thereof, by means of the introduction of materials encoding such agents.
  • Suitable materials may typically comprise nucleic acids such as DNA or RNA, and these may be devised based upon the sequences referred to in this specification.
  • Nucleic acids for use in this embodiment of the invention may be administered "as is", for example by means of ballistic transfection, or as parts of a larger construct, which may be able to incorporate stably into cells so transfected.
  • Suitable constructs may also contain regulatory elements, by which expression of a therapeutically effective amount of sFRP3, or a fragment or derivative thereof, may be achieved. Such constructs give rise to further aspects of the present invention.
  • the invention also provides a construct encoding sFRP3, or a therapeutically effective fragment or derivative thereof, said construct being capable of expression at a site where wound healing is to be accelerated to give rise to a therapeutically effective amount of the sFRP3, or therapeutically effective fragment or derivative.
  • the invention also provides a method of accelerating wound healing, the method comprising administering a construct (as described above) to a site where wound healing is to be accelerated such that a therapeutically effective amount of sFRP3, or a therapeutically effective fragment or derivative thereof, is expressed.
  • the invention also provides the use of such a construct in the manufacture of a medicament for the acceleration of wound healing.
  • Figure 1 is a bar chart comparing the widths (at day 3 post-wounding) of treated incisional wounds and "no drug control incisional wounds" (untreated na ⁇ ve control incisional wounds). "*" indicates p ⁇ 0.05 versus na ⁇ ve no drug control wounds.
  • Figure 2 is a bar chart comparing the widths (at day 3 post-wounding) of treated excisional wounds and "no drug" control excisional wounds (untreated na ⁇ ve control excisional wounds). "*" indicates p ⁇ 0.05 versus na ⁇ ve no drug control wounds.
  • the inventors investigated the ability of sFRP3 to accelerate wound healing using in an in vivo model of healing. This assessed the ability of sFRP3, provided at different concentrations, to accelerate the healing of full thickness incisional or excisional wounds in rats.
  • Carrier free human sFRP3 (Catalogue number 192-SF/CF) was purchased from R&D Systems.
  • the sFRP3 was diluted in phosphate buffered saline (PBS) to produce three solutions having concentrations as follows:
  • lng/lOO ⁇ L a concentration of 0.26 nM, in which each lOO ⁇ l of the medicament provides 0.026 pmol of sFRP3)
  • lOng/lOO ⁇ L a concentration of 2.6 nM, in which each lOO ⁇ l of the medicament provides 0.26 pmol of sFRP3)
  • lOOOng/lOO ⁇ L a concentration of 263.0 nM, in which each lOO ⁇ l of the medicament provides 26.3 pmol of sFRP3).
  • sFRP3 incorporated in the solutions described above (Ing, IOng or lOOOng of sFRP3 in lOO ⁇ l of PBS) were injected intradermally at the sites where wounds were to be formed, an amount sufficient to prophylactically treat lcm of wound (either lcm length or lcm 2 ).
  • the intradermal injections caused the formation of a raised bleb, which was then immediately incised or excised to form full thickness experimental wounds.
  • each injection of the lng/lOO ⁇ l solution provided 0.026pmoles of sFRP3, whilst each injection of the lOng/lOO ⁇ l solution provided 0.26pmoles of sFRP3, and each injection of the lOOOng/lOO ⁇ l solution provided 26.3pmoles of sFRP3.
  • All treated wounds were re-injected again 1 day post- wounding with the appropriate solution via injection of 50 ⁇ l to each of the two margins of the incisional wounds, or by injection of 25 ⁇ l of the relevant solution to each of the quadrants of the excisional wounds. Wounds were harvested at day 3 post-wounding.
  • the wounds were photographed after wounding, prior to re-injection on day 1 and on day of harvest.
  • the wounds were analysed microscopically, by image analysis, to measure wound width in order assess the ability of the medicaments of the invention to accelerate wound healing.
  • the widths of experimental wounds were assessed in order to investigate the ability of the medicaments of the invention to promote wound contraction, and thereby accelerate wound healing.
  • Wounds were excised from the experimental animals (incorporating a small amount of surrounding normal tissue) and fixed in 10% (v/v) buffered formal saline. Both the anterior and posterior halves of the wound were processed to wax but only the anterior half was cut to produce tissue sections. Five slides were prepared from each wound, each slide comprising four, 5 ⁇ m thick, serial sections. One slide from each wound was stained with Haematoxylin and Eosin (H & E) and from this wound width measurements were taken. The width of each wound was measured using image analysis equipment and software. Measurements were taken at the mid-wound point (midway throught the depth of the wound, at the level of the dermis) in the case of incisional wounds, and at the top of the wound, in the case of excisional wounds.
  • medicaments of the invention (incorporating sFRP3) was able to accelerate the healing of wounds, as assessed by measurement of width of incisional or excisional wounds three days after wound formation, at all concentrations investigated.
  • sFRP3 at a concentration of lng/lOO ⁇ l, lOng/lOO ⁇ l or lOOOng/lOO ⁇ l (respectively equivalent to a total dose of 0.052 pmoles, 0.52 pmoles, or 52.6 pmol of sFRP3 per cm of wound) significantly decreased wound width, as compared to na ⁇ ve controls (p ⁇ 0.05), when incisional wounds were assessed at three days after wounding.
  • sFRP3 at a concentration of either lng/lOO ⁇ l or lOng/lOO ⁇ l (respectively equivalent to a total dose of 0.052 pmoles or 0.52 pmoles of sFRP3 per cm 2 of wound) significantly decreased wound width, as compared to na ⁇ ve controls (p ⁇ 0.05), when excisional wounds were assessed at three days after wounding.

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Abstract

La présente invention concerne une protéine 3 apparentée à Frizzled sécrétée (sFRP3) ou un fragment ou dérivé de celle-ci thérapeutiquement efficace, utilisé comme médicament pour accélérer la guérison de plaies. La plaie peut être une plaie cutanée ou une plaie chirurgicale. Le médicament peut être un médicament topique ou peut être conçu pour une injection locale. Le médicament peut augmenter le taux de contraction d'une plaie traitée. Cette invention concerne également un procédé permettant d'accélérer la guérison de plaies, lequel procédé consiste à administrer une quantité thérapeutiquement efficace de sFRP3 ou d'un fragment ou dérivé de celle-ci thérapeutiquement efficace, au niveau d'un site où il faut accélérer la guérison d'une plaie.
PCT/GB2008/001338 2007-04-17 2008-04-17 Sfrp3 utilisée pour accélérer la guérison de plaies WO2008125864A1 (fr)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20030009023A1 (en) * 1996-10-11 2003-01-09 Luyten Frank P. Isolation and method of using tissue growth-inducing Frzb protein
WO2004113513A2 (fr) * 2003-06-25 2004-12-29 Ottawa Health Research Institute Procedes et compositions pour moduler la croissance et la differentiation des cellules souches

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20030009023A1 (en) * 1996-10-11 2003-01-09 Luyten Frank P. Isolation and method of using tissue growth-inducing Frzb protein
WO2004113513A2 (fr) * 2003-06-25 2004-12-29 Ottawa Health Research Institute Procedes et compositions pour moduler la croissance et la differentiation des cellules souches

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