WO2005041662A1

WO2005041662A1 - Method for treating collagenous tissue

Info

Publication number: WO2005041662A1
Application number: PCT/US2003/034775
Authority: WO
Inventors: Mitchell Pineau; Gerald Birchem; Edwin Bon
Original assignee: Visionary Biomedical Inc
Current assignee: Visionary Biomedical Inc
Priority date: 2003-10-03
Filing date: 2003-11-03
Publication date: 2005-05-12
Anticipated expiration: 2006-04-03
Also published as: AU2003286832A1

Abstract

The present invention relates to a method for the treatment of treating collagenous tissue having suffered weakening or damage due to disease, injury or mechanical stress by the application of a proteoglycan and electromagnetic radiation. The treatment physically and visually repairs the weakened or damaged tissue in vivo or in vitro and may be used on any animal having collagenous tissue.

Description

METHOD FOR TREATING COLLAGENOUS TISSUE BACKGROUND OF THE INVENTION 1. Field of the Invention [0001] This invention relates generally to a method for treating collagenous tissue by the use of amethod, which includes the application of a proteoglycan to the collagenous tissue along with electromagnetic radiation. 2. Description of Related Art [0002] Collagen is a fibrous protein that is the major constituent of a number of critical animal tissues. These tissues include, but are not limited to bone, skin, tendons, ligaments, blood vessels, cornea, dentine, cartilage, annular discs and other connective tissue. The collagen family represents a diverse group of extracellular matrix molecules that have a triple-helix structure. A single chain of collagen is defined as an α-chain. Εach collagen molecule consists of three α-chains that are usually identical. The only known exception is Type I collagen. Type I collagen consists of two identical chains (αl) and one different chain (α2). It is the only heteropolymer among collagens. The three polypeptide chains are stabilized as a type π left handed triple helix by intramolecular bonds. The collagen triple helices are processed in the extra-cellular matrix and self-assemble into a fibrillar structure that is stabilized primarily by intermolecular bonds. The triple helix forms a rod-like structure that is important for fibril formation and structural integrity. In higher animals, at least 19 types of collagen have been identified. Type I collagen is the principal component of bone, skin, and tendon, and is the predominant type of collagen in a mature cicatrix (scar). Type II collagen is the major type found in cartilage. [0003] The length and mechanical properties of tissues comprised primarily of type I collagen, such as ligaments, tendons, intervertebral discs, and joint capsule, can become altered as a result of an injury, a disease process, aging and everyday mechanical stress. Similarly, articular cartilage, comprised primarily of type II collagen, can suffer from the same alterations as type I collagen, as well as other specific problems, such as softening in the early stages of osteoarthritis. Other injuries or diseases can result in the fraying or tearing of collagenous tissue.

These types of ailments can be painful and even crippling to the afflicted patient. Traditional treatments may include medication, physical therapy and/or surgery to treat or replace the damaged tissue, however these methods are not fully effective, include a number of side effects and or require invasive procedures. Due to the current medical practice for treating injuries, defects and disease of collagenous tissues, the recovery period required is often time-consuming. This leads to unnecessary loss of productivity and mobility for patients. [0004] Alternative treatments to types of collagenous tissue have been advanced. One area of such treatments involves administering to a patient collagen or smaller molecules that are also related to connective tissue and extracellular matrices. Such other molecules, including long chain proteoglycans, especially glycosaminoglycans, have also been used to supplement the diets of patients with collagenous tissue injury or disease. These glycosaminoglycans include hyaluronic acid, dermatan sulfate, chondroitin, heparin, heparan sulfate, and keratan sulfate. Supplements containing glycosaminoglycans may be administered orally or by injection. [0005] Collagen and glycosaminoglycans have also been used to construct biomedical devices. A number of treatment methods show the preparation of matrices of collagen and glycosaminoglycans that are transplanted into damaged tissues.. Chemical cross-linking agents and photoinitiators have been applied directly to collagen or are used in formation of collagen/glycosaminoglycan matrices that are constructed in vitro. [0006] Other treatments have focused around the application of energy to collagenous tissue. The energy can be radiant energy, electrical energy or heat energy. Application of energy may be used to shrink and smoothen tissue, promote cross-linking between cells or destroy unwanted tissue. However, energy treatments may weaken tissue or even cause cell necrosis or DNA mutation, which may ultimately lead to the creation of cancerous cells. [0007] While the above treatments provide alternatives to traditional therapy, they do not provide complete treatment. Transplantation of exogenous matrices into the damaged tissue is both invasive and may involve issues of rejection and infection common to other types of tissue transplant. Methods that use cross-linking agents and photoinitiators are often limited to in vitro use only, as the processes and chemicals used may be cytotoxic due to acidic pHs or heat produced by exothermic reactions during the cross-linking process. Energy treatments do not have the same problem with rejection or chemical cytoxicity. However, they are limited to modifying the structure of the tissue and have not been shown to actually heal damaged tissue. Additionally, as discussed above, energy treatments may damage tissue, cause cell death or cause a mutagenic reaction at certain wavelengths or doses. [0008] Therefore there is a need for an alternative treatment for disease or injury of collagenous tissue that is more effective.

SUMMARY OF THE INVENTION [0009] In light of the present need for a more effective method of treating collagenous tissue, a brief summary of the present invention is presented. Some simplifications and omission may be made in the following summary, which is intended to highlight and introduce some aspects of the present invention, but not to limit its scope. Detailed descriptions of a preferred exemplary embodiment adequate to allow those of ordinary skill in the art to make and use the invention concepts will follow in later sections. [0010] The present invention contemplates a method for treating collagenous tissue where the method comprises applying a predetermined amount of a proteoglycan directly to an area of collagenous tissue to be treated along with the application of a predetermined amount of electromagnetic radiation to said area of collagenous tissue to be treated. In one embodiment of the present invention, the proteoglycan to be applied is a glycosaminoglycan. In a further embodiment of the present invention, the glycosaminoglycan may include hyaluronic acid. In another embodiment of the present invention the electromagnetic radiation to be applied is ultraviolet radiation. In a further embodiment of the present invention, the ultraviolet radiation has a wavelength of 300-400 nm. The present invention may be used to treat any type of collagenous tissue such as, but not limited to, joint capsule, tendons, ligaments, cartilage, bone, skin, blood vessels, annular tissue and other connective tissue. The present invention may be utilized with any animal, including human, having collagenous tissue and would provide a significant advantage over current treatment methods.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS OF THE INVENTION [0011] The present invention contemplates a method for treating collagenous tissue. Specific treatments may include, but are not limited to, modifying the texture and strength of the tissue, repairing frays, lesions or tears in the tissue and generally improving the overall health of the collagenous tissue. [0012] As used herein, the term "electromagnetic radiation" includes, but is not limited to radio waves, microwaves, infrared, visible light, ultraviolet, X-rays and gamma rays. "Ultraviolet radiation" or "UV radiation" refers to the radiation with wavelengths of 420 nm - 10 nm. The UN radiation may be generated by any known methods (such as a lamp or laser) and may be within any UV wavelength range. In the preferred embodiment of the present invention, tl e wavelength of the UV radiation is 420 nm - 285 nm. In the most preferred embodiment of the present invention, the wavelength is 400 nm - 300 nm. [0013] The term "dose", as used herein, refers to the formula of intensity of UV provided over a specific time period. The dose provided to a patient or subject further depends upon the medium through which the UV photons pass. If UV photons are required to pass through, a physiological solution to reach the target tissue the dose provided to the patient may be different than when the photons are required to only pass through air. Additionally, the dose of UV radiation would also depend on the surface area and the physiological characteristics of the particular collagenous tissue to be treated. One of ordinary skill in the art would be able to weigh these factors to provide the appropriate dose of UV photons to the tissue without undue experimentation. [0014] As used herein, the term "collagenous tissue," refers to any type of vertebrate tissue that includes collagen as a major component. Collagen in the present invention is a fibrous protein that is the chief constituent of connective tissue and bones, and includes all types of collagen, I - IXX. Collagenous tissue in the present invention includes but is not limited to joint capsules, tendons, ligaments, cartilage, bone, skin, dentine, cornea, blood vessels, annular tissue and other connective tissue, hi the preferred embodiment of the present invention, collagenous tissue includes tendons, ligaments, annular tissue and other com ective tissue. [0015] Method for treating collagenous tissue: [0016] One embodiment of the present invention provides for a method of treating collagenous tissue. The method includes applying a proteoglycan to an area of collagenous tissue to be treated, then applying ultraviolet radiation to the same area of collagenous tissue. The method may further include application of a cross-linking agent to the area to be treated, before or after application of ultraviolet radiation. [0017] The method may be used in vivo or in vitro. In the preferred embodiment, the method is used in vivo, hi order to use the method in vivo, the tissue to be treated must first be located by any medically acceptable means, such as x-ray, CAT scan, MIR, arthroscope or visually, if the procedure is invasive. The proteoglycan is then applied directly to the tissue to be treated. Following application of the proteoglycan, electromagnetic radiation is applied to the tissue and may be applied through a number of physiological layers. In addition to the proteoglycan, a cross-linking agent may be applied before or during application of electromagnetic radiation. The cross-linking agent may include any physiologically acceptable cross-linking agent. In a preferred embodiment of the present invention, the cross-linking agent is methylene blue. [0018] Any proteoglycan may be used within the scope of the present invention. In the preferred embodiment,' the proteoglycan is a glycosaminoglycan, including but not limited to hyaluronic acid, keratan sulphate, heparan sulphate, heparin, chondroitin sulphate, and dermatan sulphate. In the further preferred embodiment, the glycosaminoglycan is hyaluronic acid. The amount of proteoglycan to be applied is determined by a number of factors, including the surface area of the collagenous tissue to be treated, the location of treatment, the type of tissue treated and the depth and severity of the injury or disease. [0019] In the preferred embodiment, electromagnetic radiation is applied to the collagenous tissue to be treated for a predetermined amount of time after application of the proteoglycan. h the preferred embodiment the ultraviolet radiation has a wavelength of 420 - 285 nm. In the most preferred embodiment, the ultraviolet radiation has a wavelength of 400 - 300 nm. The electromagnetic radiation may vary in intensity depending on the dose. The dose is dependent on the surface area of the collagenous tissue to be treated, the location of the treatment, the type of tissue to be treated and the depth and severity of the injury or disease. The radiation provided to the tissue further depends upon the distance and medium through which it passes. This is especially true in the case of ultraviolet radiation in the preferred wavelengths thai has a penetration of only several millimeters. In the case of layered tissue, such as annular tissue, the imier layers would have to be treated before moving on to the outer layers according to the preferred method but could vary based on the tissue, injury and desired level of tissue treatment and repair. Alternatively, in the case of cartilage, only the outer layers can be treated due to the difficulty in penetrating the tissue, hi the preferred embodiment, the dose of ultraviolet radiation may range from 400 mJ to 6000 J. hi the most preferred embodiment, the dose may range from 1500 J to 4000 J. [0020] Another variable as to the dose of the ultraviolet radiation is the proximity to which the probe generating the radiation may be brought to the tissue. In the preferred embodiment the electromagnetic radiation is applied by probe located 1-10 mrri from the tissue to be treated. In the further prefeιτed embodiment, the radiation is applied by probe located approximately 5 mm from the tissue.

EXAMPLES [0021] The present invention will be better understood by reference to the following examples, which are provided as exemplary of the invention, and not by way of limitation.

EXAMPLE 1: [0022] The method was performed on annulus fibrosus tissue from the lumbar disc (L3, L4 and L5) of a 16-year old male with scoliosis. Freshly harvested discs were sharply dissected to remove and separate the top three layers of the annulus fibrosus. The samples were visually examined for damage, such as tears, rents or other signs of degeneration. In order to standardize the samples for experimental purposes, a microtome was used to obtain uniform samples having a size of 5 mm X 15 mm. Three specimens were examined for this experiment. To maintain tension on the test specimens during the experiment, the samples were mounted on a tensioning apparatus devised for clamping the annulus with an adjustable arm and spring and tested for baseline biomechanical characteristics. The tissue samples were then placed in a saline bath with sufficient isotonic saline solution to just cover the tissue. The saline bath was circulated through an independent water bath to maintain a constant temperature of 35-38° C. A 5 mm incision was then made with a number 11 blade scalpel and the incisions was opened to approximate a 5 mm X 5 mm open tear. The specimens were photographed prior to treatment for comparison. [0023] Specimens were treated with approximately 2 ml of 25 mg/ml Hyaluronic Acid and maintained at an approximate temperature of 35-38° C. A preprogrammed Ultraviolet dose of 3600 Joules was then applied to the specimen. Photographs were then taken of the results. [0024] With all three specimens, the incision/tear was completely sealed and minor tearing and fraying resulting from the initial dissection was also sealed. Cellular pathology of the specimens was tested but no detrimental effects to cellular structure were found.

EXAMPLE 2: [0025] The method was performed on annulus fibrosus tissue from the lumbar disc (L2,

L3 and L5) of a 14-year old male with scoliosis. Each disc sample was sharply dissected to remove and separate annular layers. A visual examination was performed to ensure that the harvested tissue was relatively normal and free of tears and weak areas. The tissue samples were then placed in a saline bath. The saline bath was circulated through an independent water bath to maintain a constant temperature of 35-37° C. The samples were tested for load-deformation characteristics in an histron™ testing machine. Total deformation of each sample was 10% of original length under a pre-stress load of 0.1 N. Measured response was reported in Newtons to realize a deformation of 1.5 mm for each 15 mm sample of annular tissue. [0026] The trial involved examining load-deformation characteristics of four groups of tissue: 1) a control group receiving no Ultraviolet exposure or incision; 2) untreated incised annular tissue; 3) UV-irradiated tissue; and 4) tissue treated with combinations of UV, hyaluronic acid and methylene blue. The following table displays the results of the trial:

[0027] As shown by the examples, application of electromagnetic radiation to collagenous tissue with the addition of a proteoglycan provides unexpected benefits in the treatment of collagen containing tissue in animals. [0028] Although the present invention has been described in detail with particular reference to preferred embodiments thereof, it should be understood that the invention is capable of other different embodiments, and its details are capable of modifications in various obvious respects. As is readily apparent to those skilled in the art, -variations and modifications can be affected while remaining within the spirit and scope of the invention. Accordingly, the foregoing disclosure, description, and figures are for illustrative purpo ses only, and do not in any way limit the invention, which is defined only by the claims.

Claims

What is claimed is:

1. A method for treating collagenous tissue comprising: applying a predetermined amount of a proteoglycan directly to an area of collagenous tissue to be treated, wherein said proteoglycan is applied to the area of collagenous tissue in vivo; and applying a predetermined amount of electromagnetic radiation to said area of collagenous tissue to be treated.

2. The method of claim 1, wherein the proteoglycan is a glycosaminoglycan.

3. The method of claim 2, wherein the glycosaminoglycan is hyaluronic acid.

4. The method of claim 2, wherein the glycosaminoglycan is dermatan sulfate and pharmaceutically effective salts thereof.

5. The method of claim 2, wherein the glycosaminoglycan is chondroitin.

6. The method of claim 2, wherein the glycosaminoglycan is heparin.

7. The method of claim 2, wherein the glycosaminoglycan is heparan sulfate.

8. The method of claim 2, wherein the glycosaminoglycan is keratan sulfate and pharmaceutically effective salts thereof.

9. The method of claim 1, wherein the electromagnetic radiation is ultraviolet radiation.

10. The method of claim 9, wherein the ultraviolet radiation has a wavelength in the range of 400-300 nm.

11. The method of claim 10, wherein the proteoglycan is hyaluronic acid.

12. The method of claim 1, wherein the method further comprises applying a cross-linking agent to the area of collagenous tissue to be treated.

13. The method of claim 12, wherein the cross-linking agent is methylene blue.

14. The method of claim 1, wherein the electromagnetic radiation is applied to the area of collagenous tissue after the predetermined amount of proteoglycan has been applied.

15. The method of claim 1, wherein the area of collagenous tissue is naturally occurring in vivo and has not been transplanted from an exogenous source.

16. The method of claim 15, wherein the proteoglycan is hyaluronic acid and the electromagnetic radiation is ultraviolet radiation.

17. A method for treating collagenous tissue comprising: applying a predetermined amount of hyaluronic acid directly to an area of collagenous tissue to be treated, wherein the hyaluronic acid is applied in vivo and the collagenous tissue to be treated has not been transplanted from an exogenous source; and applying a predetermined amount of ultraviolet radiation to said area of collagenous tissue to be treated.

8. A method for treating collagenous tissue comprising: applying a predetermined amount of hyaluronic acid directly to an area of collagenous tissue to be treated, applying a predetermined amount of ultraviolet radiation in the range of 400 - 300 nm for a predetermined time period to said area of collagenous tissue to be treated after application of said hyaluronic acid.