Treatment of pain with polysulfated polysaccharides

Abstract

To provide a novel and improved analgesic.SOLUTION: The present disclosure relates to the medical use of polysulfated polysaccharides and compositions thereof for the treatment of pain or pain conditions. In particular, the disclosure relates to the use of polysulfated polysaccharides and compositions thereof for the treatment of pain or pain conditions mediated by mature Nerve Growth Factor (NGF) or its precursor pro-Nerve Growth Factor (pro-NGF).SELECTED DRAWING: Figure 23

Description

[Technical field]

[0001] The present disclosure relates to pharmaceutical compositions and methods for treating pain or painful conditions, including the use of polysulfated polysaccharides and compositions thereof. In particular, the present disclosure relates to the therapeutic use of mature nerve growth factor (NGF) or its precursor pro- A method for treating pain or painful conditions mediated by pro-NGF The present invention relates to uses of resulfated polysaccharides and compositions thereof. [Background technology]

[0002] Safe and effective analgesics continue to represent an unmet medical need. It is a symptom of a bone disease and a disease in itself. Examples of bone diseases are osteoarthritis and osteoporosis. The disease results in significant disability, pain and reduced quality of life. Metastatic bone cancer causes excruciating bone pain. Chronic pain is estimated to cost $560 billion to $635 billion. It is reported to affect approximately 100 million Americans, with an estimated annual cost of $1. The public health and economic burden of pain is enormous [1].

[0003] The mainstay of pain treatment has typically been opioids and nonsteroidal anti-inflammatory drugs (NSAIDs). Despite accounting for the majority of clinical situations, these agents are used in limited are generally effective, have a significant side effect profile, and in the case of opioids, are There are problems with morbidity. [2] NSAIDs have been found to worsen disease. For example, in diseases such as bone cancer, osteoporosis and osteoarthritis, NSAIDs have been shown to It has a detrimental effect on healing and cartilage metabolism ([3]-[6]). Subjects in need of therapy are at increased risk of both traumatic and idiopathic fractures.

[0004] Within this context, the neurotrophins nerve growth factor (NGF) and the NGF precursor pro- Related molecular targets, such as NGF, have been implicated in malignant and non-malignant pain [7] as well as in arthritis and osteoarthritis. osteoporosis, interstitial cystitis, prostatitis, pancreatitis, chronic headache, cancer pain, non-cancer pain, back pain in the treatment of nociceptive and neuropathic pain conditions [1], including diabetic neuropathy Provides an alternative to NSAIDs and opioids. Other relevant molecular targets have also been implicated in other pain conditions, such as fibromyalgia [1A] and endometriosis. It also has potential in the treatment of inflammatory pain, such as that associated with NG. F has been found to play a role in the pathophysiology of central neuropathic pain. In this regard, high levels of NGF have been observed in the cerebrospinal fluid of patients with multiple sclerosis. [1C]

[0005] Drugs targeting NGF are being developed to treat pain conditions. Among these, Anti-NGF antibodies showed the most promise, and NGF sequestering antibodies Currently under development: TANEZUMAB® (Pfizer and Eli Lilly), FASINUMAB® (TEVA and Regen) eron) and FULRANUMAB® (Janssen and Amgen) gen). Summary of the Invention [Problem to be solved by the invention]

[0006] However, anti-NGF antibodies have been associated with joint destruction, leading to joint replacement and neurotoxicity. There are some reports that anti-NGF antibodies may be effective in treating chronic conditions [1]. The efficacy and safety profile of anticancer drugs have not yet been established. There is a need to expand the repertoire of medicines available to provide pain relief. [Means for solving the problem]

[0007] Structurally-based peptides such as heparin, pentosan polysulfate, chitosan polysulfate, and fucan Related polysulfated polysaccharides have been used as anticoagulants for many years [8–13] and are safe. Pentosan polysulfate (PPS) is used postoperatively and prophylactically as a thrombolytic agent.

[13] , and PPS is the same drug as ELMIRON, which is currently prescribed for interstitial cystitis. (registered trademark). The use of PPS in related pathologies has been described, as well as in asthma, allergic rhinitis and and / or the potential of PPS in the treatment of inflammatory conditions such as chronic obstructive pulmonary disease (COPD) Indirect uses have also been described.

[14]

[0008] The present disclosure provides the surprising finding that PPS can act to inhibit or reduce NGF activity. Thus, PPS is a novel stimulator of NGF that acts as a blocker of NGF. It acts via a different mechanism of action than the currently available anti-NGF antibodies, tanezumab, fasinumab, and fulranumab. Surprisingly, PPS acts as a pro-NGF inhibitor. It has been found that the experimental data can act to inhibit or reduce the activity of For example, PPS inhibits the expression of NGF at both the mRNA and protein levels. PPS inhibits or reduces the expression of pro-NGF. It has been demonstrated that this can affect the levels of NGF and its activity. Furthermore, experimental data suggest that PPS may be a mediator of pain and pain-related conditions. The pharmacological action of NGF is exerted by targeting NGF-producing bone cells as a target. Experimental data have demonstrated that it can mediate pain and pain-related conditions. Although bone cells are used as an example of cells capable of treating pain and pain-related conditions, the present disclosure is directed to Contemplates using other target cells that secrete NGF or pro-NGF as mediators Examples of such target cells are immune cells and primary and secondary cancer cells. These are cancer cells that contain cytoplasmic cells.

[0009] According to one embodiment, a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal is provided. The method comprises the steps of administering to a mammal a polysulfated polysaccharide or an acceptable salt thereof. A method is provided, comprising:

[0010] According to another aspect, a method for inhibiting pro-nerve growth factor (pro-NGF) activity in a mammal is provided. The method comprises administering to a mammal a polysulfated polysaccharide or an acceptable salt thereof. A method is provided that includes the step of providing a

[0011] According to another aspect, a method for treating pain mediated by nerve growth factor (NGF) comprises administering to the subject a and administering a polysulfated polysaccharide or an acceptable salt thereof to a mammal in need of such treatment. A method is provided comprising administering to an object.

[0012] According to another aspect, a method for treating pain mediated by pro-nerve growth factor (NGF) is provided. The method comprises administering a polysulfated polysaccharide or an acceptable salt thereof to a pet in need of such treatment. A method is provided comprising administering to a mammal.

[0013] According to another aspect, a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal is provided. A composition for administering a pharmaceutical composition to a subject, comprising a polysulfated polysaccharide or an acceptable salt thereof and an acceptable excipient. Things are provided.

[0014] According to another aspect, a method for inhibiting pro-nerve growth factor (pro-NGF) activity in a mammal is provided. or reducing the amount of a polysulfated polysaccharide or an acceptable salt thereof and an acceptable excipient. A composition comprising:

[0015] According to another aspect, a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal is provided. A polysulfated polysaccharide or an acceptable salt thereof and an acceptable excipient for use in preparing a pharmaceutical composition comprising the .... A composition comprising:

[0016] According to another aspect, a method for inhibiting pro-nerve growth factor (pro-NGF) activity in a mammal is provided. and a polysulfated polysaccharide or an acceptable salt thereof for use in reducing or inhibiting and an excipient adapted to said composition.

[0017] According to another aspect, the present invention relates to a method for treating pain mediated by nerve growth factor (NGF) in a mammal. The present invention relates to a method for treating atopic dermatitis, comprising administering to a patient a composition comprising a polysulfated polysaccharide or an acceptable salt thereof and an acceptable excipient. A composition comprising:

[0018] According to another aspect, the present invention provides a method for the treatment of neurodegenerative diseases mediated by pro-nerve growth factor (pro-NGF) in mammals. A method for treating pain mediated by a polysulfated polysaccharide or an acceptable salt thereof and an acceptable salt thereof and an excipient.

[0019] According to another aspect, the present invention relates to a method for treating pain mediated by nerve growth factor (NGF) in a mammal. The present invention relates to a polysulfated polysaccharide or an acceptable salt thereof and an acceptable and an excipient.

[0020] According to another aspect, the present invention provides a method for the treatment of neurodegenerative diseases mediated by pro-nerve growth factor (pro-NGF) in mammals. and a polysulfated polysaccharide or an acceptable salt thereof for use in treating pain mediated by and an acceptable excipient.

[0021] According to another aspect, a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal is provided. The present invention provides the use of a polysulfated polysaccharide or an acceptable salt thereof in

[0022] According to another aspect, a method for inhibiting or reducing pro-nerve growth factor (NGF) activity in a mammal is provided. The present invention provides the use of a polysulfated polysaccharide, or an acceptable salt thereof, in reducing

[0023] According to another aspect, a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal is provided. The present invention provides a use of a polysulfated polysaccharide or an acceptable salt thereof in the manufacture of a medicament for the treatment of a disease caused by a pulmonary artery disease. will be done.

[0024] According to another aspect, a method for inhibiting pro-nerve growth factor (pro-NGF) activity in a mammal is provided. or reducing the amount of a polysulfated polysaccharide or an acceptable salt thereof in the manufacture of a medicament for Use is provided.

[0025] According to another aspect, the present invention relates to a method for treating pain mediated by nerve growth factor (NGF) in a mammal. The present invention provides the use of a polysulfated polysaccharide, or an acceptable salt thereof, in treating

[0026] According to another aspect, the present invention provides a method for the treatment of neurodegenerative diseases mediated by pro-nerve growth factor (pro-NGF) in mammals. The present invention provides a use of a polysulfated polysaccharide or an acceptable salt thereof in treating pain mediated by do.

[0027] According to another aspect, the present invention relates to a method for treating pain mediated by nerve growth factor (NGF) in a mammal. The use of a polysulfated polysaccharide or an acceptable salt thereof in the manufacture of a medicament for treating Provided.

[0028] According to another aspect, the present invention provides a method for the treatment of neurodegenerative diseases mediated by pro-nerve growth factor (pro-NGF) in mammals. The use of a polysulfated polysaccharide or an acceptable salt thereof in the manufacture of a medicament for treating mediated pain. The use of such salts is provided.

[0029] definition Unless otherwise defined herein, scientific and technical terms used in connection with this application Terms used herein shall have the meanings commonly understood by those of ordinary skill in the art. Unless otherwise required by paragraph, singular terms shall include the plural and plural terms shall include the singular. Therefore, as used in this specification and the appended claims, the singular The forms "a," "an," and "the" are clear from the context. Unless otherwise indicated, "a cell" includes plural referents. Reference to includes a population of cells.

[0030] With respect to the definitions provided herein, unless otherwise stated or implied from the context, Unless otherwise specified, defined terms and phrases have the meanings given. Unless otherwise clear from context, the following terms and phrases are understood to be of the same meaning as those used herein by those of ordinary skill in the relevant art. The scope of the invention extends to the claims contained therein. The definitions are provided to aid in describing particular embodiments, since the invention is limited only by , and are not intended to limit the invention as defined by the claims.

[0031] Throughout this specification, various aspects and components of this invention may be presented in a range format. The range format is included for convenience and should not be construed as an inflexible limitation on the scope of the invention. Therefore, ranges should be stated to cover all possible parts unless otherwise indicated. In addition to the range, each individual numerical value within the range should be deemed to be specifically disclosed. For example, a range such as 1-5 can be expressed as 1-2, 1-3, 1-4, 2-3, 2-4, 2 In addition to subranges such as ~5, 3-4, etc., individual numbers and subranges within the listed ranges are also included. The numbers, for example, 1, 2, 3, 4 and 5 should be considered to be specifically disclosed. This applies regardless of the breadth of a range disclosed. These will be presented herein.

[0032] The term "acceptable excipient" refers to a vehicle that is physiologically compatible and capable of carrying the compounds described herein. or any solvent, diluent, dispersion medium, coating agent, antimicrobial agent, or any other additive that is not harmful to the use thereof. and excipients or agents such as antifungal agents, isotonic and absorption delaying agents. The use of such carriers and agents to prepare compositions of active substances is well known in the art. It is well known (e.g., Remington: The Science and Practice of Pharmacy, 21st Edition, n; Lippincott Williams & Wilkins: Philadelphia, PA, 2005).

[0033] The term "acceptable salt" includes, but is not limited to, hydrochloride, hydrobromide, sulfate, lysine, phosphate, phosphate, phosphate-containing salts, etc. Inorganic acid salts such as phosphates; formates, acetates, trifluoroacetates, maleates, alcohols, etc. Organic acid salts such as tartrates, methanesulfonates, benzenesulfonates, p-toluenesulfonates, etc. sulfonates, such as arginates, aspartates, glutamines, etc. Amino acid salts such as amine salts; sodium salts, potassium salts, cesium salts, etc. Metal salts; alkaline earth metals such as calcium salts, magnesium salts, etc.; and triethylene salts. amidoamine salt, pyridine salt, picoline salt, ethanolamine salt, triethanolamine salt, Organic salts such as dicyclohexylamine salts, N,N'-dibenzylethylenediamine salts, etc. Includes amine salts.

[0034] Base salts include, but are not limited to, sodium, potassium, lithium, calcium, magnesium Alkylamines such as salts formed from naphthalene, zinc, ammonium, and triethylamine ammonium, alkoxyammonium such as those formed with ethanolamine, and and ethylenediamine, choline or amines such as arginine, lysine or histidine. These include salts formed with pharma- ceutically acceptable cations, such as salts formed with acids. The types of acceptable salts and general information regarding their formation are known to those skilled in the art. “Handbook of Pharmaceutical salts” PHStahl, CGWermuth, 1 st edition, 2002, This is as described in general textbooks such as Wiley-VCH.

[0035] The terms "administration of" and / or "administering" a compound refer to the administration of a compound of the invention to a It should be understood to mean providing treatment to an individual in need thereof.

[0036] As used herein, the term "composition" refers to a preparation containing specific ingredients in specific amounts. In addition to the composition, any product derived directly or indirectly from the combination of specified ingredients in specified amounts. The term "product" is intended to encompass any product.

[0037] Throughout this specification the words "comprise" or "comprise" Variations such as "comprising" or "comprising" , the inclusion of the stated elements, integers or steps, or groups of elements, integers or steps It is meant to include, but is not limited to, any other element, integer or step, or It will be understood that no exclusion of groups of steps is implied.

[0038] The present disclosure relates to the treatment of mammalian subjects. Treatment of a "mammalian" subject may also refer to a "patient" or Treatment of an individual is sometimes referred to as treatment of an "individual." A "mammalian" subject is a person who is receiving treatment for a particular condition or treatment. whether the patient exhibits clinical signs of a condition suggesting the need for medical treatment, is treated for a condition, or or has been diagnosed with a condition that is being treated.

[0039] As used herein, "malignant pain" refers to pain associated with cancer; "Pain" refers to pain that is not related to cancer.

[0040] Thus, unless otherwise specifically indicated, the present invention relates to methods for treating and / or preventing the development of inflammatory bowel disorders in humans and other non-human mammals. It should be understood that the term "human" refers to a human subject that is a human being. Human mammals include primates, domestic animals and farm animals (e.g., sheep, horses, etc.). animals such as cats and dogs; performance animals; ) (e.g., race horses, camels, greyhounds), clinical laboratory animals (e.g., mice, rabbits In addition to the reptiles (dogs, rats, and guinea pigs), which are usually found in the wild, they are also found in zoos, wildlife parks, etc. Even though they are mammals that may be susceptible to treatment because they are found in many places, good.

[0041] As used herein, the terms "treating," "treat," or "treatment" The term and variations thereof refer to a polysulfated polysaccharide or an acceptable salt thereof, or a polysulfated polysaccharide. or an acceptable salt thereof during the course of clinical pathology, compared to not administering a composition comprising the Refers to a clinical intervention intended to alter the natural course of a subject's condition. Therapeutic options include slowing the progression of the disease, reversing or alleviating the disease state, and improving prognosis. For example, if one or more of the above therapeutic outcomes are achieved, the subject may: "Treatment" refers to being successfully "treated." As used herein, "treating" and "treat" refer to The term "treatment" and its variations are intended to avoid the occurrence of clinical pathological processes. "Preventing," "preventing," or "preventing" will be understood to refer to a clinical intervention that has been "Prevention."

[0042] An "effective amount" is the amount needed to produce a measurable improvement in a particular disease (e.g., bone marrow edema). The term "therapeutically effective" refers to at least the minimum concentration or amount that is achieved by administering the compound to a subject. The effective amounts herein will depend on the disease state, age, sex, and weight of the patient, as well as the desired individual This may vary depending on factors such as the ability of the PPS to produce a new response. An effective dose is an amount in which the therapeutically beneficial effects outweigh any toxic or adverse effects of the PPS. "An effective amount" refers to the amount of drug or agent required to obtain the desired prophylactic result. It also encompasses a "prophylactically effective" amount, which refers to the rate of administration of an agent.

[0043] Description of the embodiments The present disclosure relates to a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal. comprising the step of administering to a mammal a polysulfated polysaccharide or an acceptable salt thereof. , relates to a method.

[0044] The present disclosure also provides a method for inhibiting or preventing pro-nerve growth factor (pro-NGF) activity in a mammal. The present invention relates to a method for reducing the level of inflammatory bowel disease, comprising administering to a mammal a polysulfated polysaccharide or an acceptable salt thereof. The present invention relates to a method comprising the steps of:

[0045] In the above methods, inhibition or reduction of NGF activity or pro-NGF activity is preferred. Or, to treat pain in a mammal.

[0046] Experimental data show that PPS upregulates NGF at both the mRNA and protein levels. It was found that PPS inhibits or reduces the expression of pro-NGF. It has been shown that it can reduce the levels of NGF and its activity. Furthermore, experimental data suggest that PPS may be a mediator of pain and pain-related conditions. The pharmacological action of NGF is exerted by targeting NGF-producing bone cells as a target. Experimental data demonstrate that serotonin receptor agonists can mediate pain and pain-related conditions. Although bone cells are used as an example of cells that can be used to treat pain and pain-related conditions, the present disclosure is not limited to the use of The present invention contemplates the use of other target cells that secrete NGF or pro-NGF as a mediator. Examples of such target cells include immune cells and primary and secondary cancer cells. These are cancer cells that contain

[0047] Thus, in the examples of the methods, compositions or uses described herein, a mammal Inhibition or reduction of nerve growth factor (NGF) activity in the brain inhibits or reduces NGF expression In one example, the inhibition or reduction of NGF expression is achieved by suppressing the transcription of mRNA. In one embodiment, the inhibition or reduction of expression of NGF is By inhibiting or reducing the translation of RNA into protein.

[0048] Thus, in the examples of the methods, compositions or uses described herein, a mammal Inhibition or reduction of pro-nerve growth factor (pro-NGF) activity in In one example, by inhibiting or reducing expression of pro-NGF. In one example, the inhibition or reduction is by inhibiting or reducing transcription of mRNA. -Inhibition or reduction of NGF expression inhibits or reduces translation of mRNA into protein. This is by making it possible.

[0049] The present disclosure relates to the inhibition and reduction of pro-NGF activity, as a result of which NGF activity is inhibited and reduced. Further reductions are planned.

[0050] The present disclosure is also directed to a method of treating pain mediated by nerve growth factor (NGF). and administering the polysulfated polysaccharide or an acceptable salt thereof to a mammal in need of such treatment. The present invention relates to a method comprising the step of administering

[0051] The present disclosure also provides a method for treating pain mediated by pro-nerve growth factor (pro-NGF). The method comprises administering a polysulfated polysaccharide, or an acceptable salt thereof, to a patient in need of such treatment. The present invention relates to a method comprising administering to a mammal comprising the steps of:

[0052] In the above methods, the pain is non-malignant pain or malignant pain.

[0053] In the above method, the pain is preferably musculoskeletal pain. Preferably, the musculoskeletal pain is non-malignant. Preferably, the musculoskeletal pain is malignant musculoskeletal pain. Preferably, the skeletal pain is non-malignant skeletal pain. Preferably, the skeletal pain is malignant It's skeletal pain.

[0054] Preferably, the skeletal pain is back pain. Preferably, the back pain is lower back pain. The pain is bone and / or joint pain. Preferably, the skeletal pain is joint pain. Preferably, The skeletal pain is bone pain. Preferably, the bone pain is non-malignant bone pain. Preferably, the non-malignant bone pain is The pain is associated with osteoporosis.

[0055] Preferably, the non-malignant bone pain is pain associated with an arthritic condition. The arthritic condition is selected from rheumatoid arthritis or osteoarthritis. Preferably, the arthritic condition is osteoarthritis. Preferably, the arthritic condition is osteoarthritis. is a joint selected from the group consisting of ankle, hip, knee, shoulder, spine and wrist. Preferably, the osteoarthritis is in the hip joint. Preferably, the osteoarthritis is in the knee joint. Preferably, the bone pain is malignant bone pain. Preferably, malignant bone pain is pain associated with a primary bone cancer. Pain associated with bone cancer.

[0056] Preferably, the pain is acute pain; cancer pain; chronic pain; generalized pain; non-cancer pain and persistent pain. Preferably, the pain is selected from the group consisting of a nociceptive pain condition or a neuropathic pain. Preferably, the pain is an acute pain condition; a chronic pain condition; a generalized pain condition, and and persistent pain conditions. Preferably, the pain condition is a non-malignant pain condition. The pain condition is a malignant pain condition.

[0057] Preferably, the pain is due to arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; interstitial cystitis. ;Osteoporosis;Abdominal pain;Arthralgia;Endometriosis;Pancreatitis;Chronic headaches;Primary cancer;Secondary (metastatic) )Cancer;Back pain;Bone injuries or fractures;Diabetic neuropathy;Fibromyalgia;Migraines;Multiple myeloma Bone disease; multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; post-herpetic neuralgia; neoplasia The present invention is associated with a condition selected from the group consisting of: sciatica; sciatica; and visceral pain.

[0058] Preferably, the pain is due to arthritis; osteoarthritis; rheumatoid arthritis; prostatitis and interstitial bladder pain. The condition is associated with a condition selected from the group consisting of cystitis.

[0059] Preferably, the pain is abdominal pain; joint pain; endometriosis; pancreatitis; chronic headache; primary cancer; secondary Metastatic cancer; back pain; bone injury or fracture; diabetic neuropathy; fibromyalgia; migraine; Myeloma bone disease; multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; postherpetic It is associated with a condition selected from the group consisting of menstrual pain; neoplasms; sciatica; and visceral pain.

[0060] The present disclosure also provides a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal. The present invention relates to a composition comprising a polysulfated polysaccharide or an acceptable salt thereof for use in a pharmaceutical composition comprising a polysulfated polysaccharide or an acceptable salt thereof and an acceptable excipient. do.

[0061] The present disclosure also provides a method for inhibiting or preventing pro-nerve growth factor (pro-NGF) activity in a mammal. The present invention relates to a method for the preparation of a polysulfated polysaccharide or an acceptable salt thereof, and an acceptable excipient, for reducing the The present invention relates to a composition comprising:

[0062] The present disclosure also provides a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal. The present invention relates to a method for producing a polysulfated polysaccharide or an acceptable salt thereof, and an acceptable excipient, for use in the production of a pharmaceutical composition comprising the steps of: The present invention relates to a composition comprising:

[0063] The present disclosure also provides a method for inhibiting or preventing pro-nerve growth factor (pro-NGF) activity in a mammal. and a polysulfated polysaccharide or an acceptable salt thereof and an acceptable salt thereof for use in reducing and an excipient.

[0064] In the above compositions, the inhibition or reduction treats pain in a mammal.

[0065] The present disclosure also provides a method for treating nerve growth factor (NGF)-mediated pain in a mammal. A composition for treating a patient suffering from atopic dermatitis, comprising a polysulfated polysaccharide or an acceptable salt thereof and an acceptable excipient. Related to the composition.

[0066] The present disclosure also relates to a method for treating neuroprotection mediated by pro-nerve growth factor (pro-NGF) in mammals. A polysulfated polysaccharide or an acceptable salt thereof and an acceptable excipient for treating pain in a patient suffering from a chronic condition. The present invention relates to a composition comprising:

[0067] The present disclosure also provides a method for treating nerve growth factor (NGF)-mediated pain in a mammal. A polysulfated polysaccharide or an acceptable salt thereof and an acceptable excipient for use in treating The present invention relates to a composition comprising:

[0068] The present disclosure also relates to a method for treating neuroprotection mediated by pro-nerve growth factor (pro-NGF) in mammals. The present invention relates to a polysulfated polysaccharide or an acceptable salt thereof and an acceptable salt thereof for use in treating pain associated with and an acceptable excipient.

[0069] In the above composition, the pain is non-malignant pain or malignant pain.

[0070] In the above composition, the pain is preferably musculoskeletal pain. Preferably, the musculoskeletal pain is non- Preferably, the musculoskeletal pain is malignant musculoskeletal pain. The pain is skeletal pain. Preferably, the skeletal pain is non-malignant skeletal pain. It is skeletal pain.

[0071] Preferably, the skeletal pain is back pain. Preferably, the back pain is lower back pain. The pain is bone and / or joint pain. Preferably, the skeletal pain is joint pain. Preferably, The skeletal pain is bone pain. Preferably, the bone pain is non-malignant bone pain. Preferably, the non-malignant bone pain is The pain is associated with osteoporosis.

[0072] Preferably, the non-malignant bone pain is pain associated with an arthritic condition. The arthritic condition is selected from rheumatoid arthritis or osteoarthritis. Preferably, the arthritic condition is osteoarthritis. Preferably, the arthritic condition is osteoarthritis. is a joint selected from the group consisting of ankle, hip, knee, shoulder, spine and wrist. Preferably, the osteoarthritis is in the hip joint. Preferably, the osteoarthritis is in the knee joint. Preferably, the bone pain is malignant bone pain. Preferably, malignant bone pain is pain associated with a primary bone cancer. Pain associated with bone cancer.

[0073] Preferably, the pain is acute pain; cancer pain; chronic pain; generalized pain; non-cancer pain and persistent pain. Preferably, the pain is selected from the group consisting of a nociceptive pain condition or a neuropathic pain. Preferably, the pain is an acute pain condition; a chronic pain condition; a generalized pain condition, and and persistent pain conditions. Preferably, the pain condition is a non-malignant pain condition. The pain condition is a malignant pain condition.

[0074] Preferably, the pain is due to arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; interstitial cystitis. ;Osteoporosis;Abdominal pain;Arthralgia;Endometriosis;Pancreatitis;Chronic headaches;Primary cancer;Secondary (metastatic) )Cancer;Back pain;Bone injuries or fractures;Diabetic neuropathy;Fibromyalgia;Migraines;Multiple myeloma Bone disease; multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; post-herpetic neuralgia; neoplasia sciatica; and visceral pain. The fracture is a femur fracture. The femur fracture is preferably a femoral neck fracture.

[0075] Preferably, the pain is due to arthritis; osteoarthritis; rheumatoid arthritis; prostatitis and interstitial bladder pain. The condition is associated with a condition selected from the group consisting of cystitis.

[0076] Preferably, the pain is abdominal pain; joint pain; endometriosis; pancreatitis; chronic headache; primary cancer; secondary Metastatic cancer; back pain; bone injury or fracture; diabetic neuropathy; fibromyalgia; migraine; Myeloma bone disease; multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; postherpetic The present invention is associated with a condition selected from the group consisting of menstrual pain; neoplasms; sciatica; and visceral pain. Preferably, the fracture is a femur fracture. The femur fracture is preferably a femoral neck fracture.

[0077] The present disclosure also provides a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal. The present invention relates to the use of a polysulfated polysaccharide or an acceptable salt thereof in the manufacture of a pharmaceutical composition comprising the steps of:

[0078] The present disclosure also provides a method for inhibiting or preventing pro-nerve growth factor (pro-NGF) activity in a mammal. relates to the use of polysulfated polysaccharides or acceptable salts thereof in reducing

[0079] The present disclosure also provides a method for inhibiting or reducing nerve growth factor (NGF) activity in a mammal. The present invention relates to the use of a polysulfated polysaccharide or an acceptable salt thereof in the manufacture of a medicament for treating a patient suffering from atopic dermatitis.

[0080] The present disclosure also provides a method for inhibiting or preventing pro-nerve growth factor (pro-NGF) activity in a mammal. Use of polysulfated polysaccharides or acceptable salts thereof in the manufacture of a medicament for reducing Regarding.

[0081] In the above uses, the inhibition or reduction treats pain in a mammal.

[0082] The present disclosure also provides a method for treating nerve growth factor (NGF)-mediated pain in a mammal. The present invention relates to the use of a polysulfated polysaccharide or an acceptable salt thereof in treating a patient suffering from atopic dermatitis.

[0083] The present disclosure also relates to a method for treating neuroprotection mediated by pro-nerve growth factor (pro-NGF) in mammals. The present invention relates to the use of a polysulfated polysaccharide or an acceptable salt thereof in treating pain associated with chronic pain.

[0084] The present disclosure also provides a method for treating nerve growth factor (NGF)-mediated pain in a mammal. Use of a polysulfated polysaccharide or an acceptable salt thereof in the manufacture of a medicament for treating do.

[0085] The present disclosure also relates to a method for treating neuroprotection mediated by pro-nerve growth factor (pro-NGF) in mammals. A polysulfated polysaccharide or an acceptable salt thereof in the manufacture of a medicament for treating pain Regarding the use of.

[0086] In the above uses, the pain is non-malignant pain or malignant pain.

[0087] In the above uses, the pain is preferably musculoskeletal pain. Preferably, the musculoskeletal pain is non-malignant. Preferably, the musculoskeletal pain is malignant musculoskeletal pain. Preferably, the skeletal pain is non-malignant skeletal pain. Preferably, the skeletal pain is malignant It's skeletal pain.

[0088] Preferably, the skeletal pain is back pain. Preferably, the back pain is lower back pain. The pain is bone and / or joint pain. Preferably, the skeletal pain is joint pain. Preferably, The skeletal pain is bone pain. Preferably, the bone pain is non-malignant bone pain. Preferably, the non-malignant bone pain is The pain is associated with osteoporosis.

[0089] Preferably, the non-malignant bone pain is pain associated with an arthritic condition. The arthritic condition is selected from rheumatoid arthritis or osteoarthritis. Preferably, the arthritic condition is osteoarthritis. Preferably, the arthritic condition is osteoarthritis. is a joint selected from the group consisting of ankle, hip, knee, shoulder, spine and wrist. Preferably, the osteoarthritis is in the hip joint. Preferably, the osteoarthritis is in the knee joint. Preferably, the bone pain is malignant bone pain. Preferably, malignant bone pain is pain associated with a primary bone cancer. Pain associated with bone cancer.

[0090] Preferably, the pain is acute pain; cancer pain; chronic pain; generalized pain; non-cancer pain and persistent pain. Preferably, the pain is selected from the group consisting of a nociceptive pain condition or a neuropathic pain. Preferably, the pain is an acute pain condition; a chronic pain condition; a generalized pain condition, and and persistent pain conditions. Preferably, the pain condition is a non-malignant pain condition. The pain condition is a malignant pain condition.

[0091] Preferably, the pain is caused by arthritis, osteoarthritis, rheumatoid arthritis, interstitial cystitis, abdominal pain; Joint pain; endometriosis, osteoporosis, prostatitis, pancreatitis, chronic headaches, primary cancer, secondary (metastatic) )Cancer, back pain, bone injury or fracture, diabetic neuropathy, fibromyalgia; migraine headaches; multiple myeloma Bone disease, multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; post-herpetic neuralgia; neoplasia sciatica; and visceral pain. The fracture is a femur fracture. The femur fracture is preferably a femoral neck fracture.

[0092] Preferably, the pain is due to arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; and interstitial The condition is associated with a condition selected from the group consisting of cystitis.

[0093] Preferably, the pain is abdominal pain; joint pain; endometriosis; pancreatitis; chronic headache; primary cancer; secondary Metastatic cancer; back pain; bone injury or fracture; diabetic neuropathy; fibromyalgia; migraine; Myeloma bone disease; multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; postherpetic The present invention is associated with a condition selected from the group consisting of menstrual pain; neoplasms; sciatica; and visceral pain. Preferably, the fracture is a femur fracture. The femur fracture is preferably a femoral neck fracture.

[0094] The present disclosure also contemplates the methods, compositions and uses described above, wherein the polysulfated polysaccharides are preferably These include high molecular weight heparin, low molecular weight heparin, heparan sulfate, pentosan polysulfate, poly Chondroitin sulfate, chitosan polysulfate, dermatan polysulfate sulodexide, dex sulfate Tolan, polysulfated inulin, sulfated lactobionic acid amide, sulfated bis-aldonic acid amide Mido, sucrose octasulfate, fucoidan-1, fucoidan-2, sulfated beta-cyclodextrin cyclodextrin, sulfated gamma-cyclodextrin and inositol hexasulfate, The compound is selected from the group consisting of small sulfated compounds including benzoyl sulfate.

[0095] Preferably, the polysulfated polysaccharide is a high molecular weight heparin, a low molecular weight heparin, a polysulfate pentasaccharide, or a combination thereof. Selected from the group consisting of chitosan (PPS), chondroitin polysulfate and chitosan polysulfate will be done.

[0096] The polysulfate pentosan (PPS) is preferably a sodium salt of polysulfate pentosan ( NaPPS), magnesium salt of pentosan polysulfate (MgPPS), pentosan polysulfate The calcium salt of pentosan polysulfate (CaPPS) and the zinc salt of pentosan polysulfate (ZnPPS) were The compound is selected from the group consisting of:

[0097] Preferably, the polysulfate pentosan (PPS) is sodium polysulfate pentosan (Na PPS).

[0098] In one preferred embodiment, NaPPS is available from Bene-PharmaChem Gm US FDA and EU FDA approved by bH & Co KG, Geretsried, Germany Manufactured according to specifications submitted to the ropean Community EMEA.

[0099] PPS and PPS compositions suitable for administration by various routes are described in Remington: The Science and Practice of Pharmacy, 21st Edition; Lippincott Williams & Wilkins: Philadel The formulations are reviewed by referring to standard textbooks in this field, such as Fischer, Phia, PA, 2005. It will be appreciated by those skilled in the art that these compositions can be formulated for use in injection, oral administration, or in combination with other suitable formulations. (including tablets and capsules containing gastrointestinal drug absorption enhancers and enhancers), intravenous, etc. This includes those caused by:

[0100] Preferably, the inhibition or reduction, or treatment, is administered intramuscularly (IM) or subcutaneously (SC). by injection via the intraventricular, intravesical or intrathecal route; by intravenous (IV) route , intra-articular (IA), peri-articular, topical, via suppository, or orally. by.

[0101] The present disclosure relates to a method for the production of nerve growth factor (NGF) or pro-nerve growth factor (proGF) in mammals. -NGF) activity, or to inhibit or reduce polysulfated polysaccharides or their tolerance The salt is administered to a mammal in an effective amount ranging from about 10 ng to about 1000 ng as a fixed dose. By administering NGF or pro-NGF-mediated In certain embodiments, the effective amount is about 100 mg / kg / day, as a fixed dose. 50ng~approx. 950ng;approx. 100ng~approx. 900ng;approx. 150ng~approx. 850ng; Approx. 200ng~Approx. 800ng; Approx. 250ng~Approx. 750ng; Approx. 300ng~Approx. 700n g; about 350ng to about 650ng; about 400ng to about 600ng; about 450ng to about 50 In certain embodiments, the effective amount is in the range of about 10 ng, about 50 ng, about 100ng, about 150ng, about 200ng, about 250ng, about 300ng, about 350ng , about 400ng, about 450ng, about 500ng, about 550ng, about 600ng, about 650 ng, about 700ng, about 750ng, about 800ng, about 850ng, about 900ng, about 9 It is a fixed dose of 50ng or about 1000ng.

[0102] At the doses mentioned above (i.e., the doses range from about 10 ng to about 1000 ng), In this regard, the inhibition or reduction, or treatment, is preferably performed intraventricularly, intravesically or intrathecally. By injection of polysulfated polysaccharides or acceptable salts thereof via the route.

[0103] The present disclosure relates to a method for the production of nerve growth factor (NGF) or pro-nerve growth factor (prGGF) in mammals. Inhibiting or reducing o-NGF activity, or inhibiting or reducing polysulfated polysaccharides or their receptors The tolerable salt is administered to a mammal in an effective amount ranging from about 10 μg to about 1000 μg as a fixed dose. mediated by NGF or pro-NGF in mammals by administration In certain embodiments, the effective amount is about 100 mg / kg / day, as a fixed dose. 50μg~about 950μg;about 100μg~about 900μg;about 150μg~about 850μg; Approx. 200μg~Approx. 800μg; Approx. 250μg~Approx. 750μg; Approx. 300μg~Approx. 700μ g; about 350 μg to about 650 μg; about 400 μg to about 600 μg; about 450 μg to about 50 In certain embodiments, the effective amount is about 10 μg, about 50 μg, about 100 μg, about 200 μg, about 300 μg, about 400 μg, about 500 μg, about 600 μg, about 700 μg, about 800 μg, about 900 μg, about 1000 μg, about 1500 μg, about 2000 μg, about 3000 μg, approx. 150 μg, approx. 200 μg, approx. 250 μg, approx. 300 μg, approx. 350 μg, approx. 4 00μg, about 450μg, about 500μg, about 550μg, about 600μg, about 650μg, Approx. 700 μg, approx. 750 μg, approx. 800 μg, approx. 850 μg, approx. 900 μg, approx. 950 μg g or a fixed dose of approximately 1000 μg.

[0104] Regarding the doses mentioned above (i.e., they range from about 10 μg to about 1000 μg), The inhibition, reduction or treatment is preferably achieved by intraventricular, intravesicular or intrathecal routes. by injection of polysulfated polysaccharides or acceptable salts thereof.

[0105] Preferably, the inhibition or reduction, or treatment, is by the administration of polysulfated polysaccharides or their acceptable derivatives. The salt is administered to the mammal in an effective amount of about 1 mg / kg to about 2 mg / kg of the mammal per dose. By doing so.

[0106] Preferably, the inhibition or reduction, or treatment, is by the administration of polysulfated polysaccharides or their acceptable derivatives. By administering the salt to the mammal in an effective amount of about 2 mg / kg of the mammal per dose. The effective amount is preferably about 1.0 to 2.0 mg / kg of subject per dose. In embodiments, the effective amount is about 1.0-1.5 mg / kg; 1.5-2.0 mg / kg; 0.5mg / kg; 1.0mg / kg; 1.5mg / kg; or 2.0mg / kg do.

[0107] The present disclosure relates to a method for the production of nerve growth factor (NGF) or pro-nerve growth factor (prGGF) in mammals. Inhibiting or reducing o-NGF activity, or inhibiting or reducing polysulfated polysaccharides or their receptors The tolerable salt is administered to a mammal in an effective amount ranging from about 1 mg to about 25 mg as a fixed dose. By this, pain mediated by NGF or pro-NGF in mammals can be prevented. In certain embodiments, the effective amount is from about 2 mg to about 24 mg. between about 3 mg and about 23 mg; between about 4 mg and about 22 mg; between about 5 mg and about 2 Between 1 mg; Between about 6 mg and about 20 mg; Between about 7 mg and about 19 mg; Between about 8 mg and Between about 18 mg; Between about 9 mg and about 17 mg; Between about 10 mg and about 16 mg; About 11 between about 12 mg and about 14 mg. In certain embodiments, the effective amount is about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, About 6mg, about 7mg, about 8mg, about 9mg, about 10mg, about 11mg, about 12mg, about 1 3mg, about 14mg, about 15mg, about 16mg, about 17mg, about 18mg, about 19mg, Fixation of about 20 mg, about 21 mg, about 22 mg, about 23 mg, about 24 mg or about 25 mg The dosage.

[0108] In certain embodiments, the effective amount is a fixed dose between about 25 mg and 4000 mg. In certain embodiments, the effective amount is about 25 mg, 50 mg, 75 mg, 100 mg, , 125mg, 150g, 175mg, 200mg, 225mg, 250mg, 275m In certain embodiments, the effective amount is a fixed dose of about 350 mg or 300 mg. g, 400mg, 450mg, 500mg, 550mg, 600mg, 650mg, 70 0mg, 750mg, 800mg, 850mg, 900mg, 950mg, 1000mg , fixed doses of 2000mg, 3000mg or 4000mg.

[0109] Preferably, administration is by injection. Preferably, administration is by subcutaneous (SC) injection. Alternatively, the SC injection is a slow SC injection. Administration is preferably intramuscular (IM) ) by injection.

[0110] Preferably, administration to a human is by dosing in a once-daily, twice-weekly or three-times-weekly treatment regimen. Administration to humans is preferably by dosing in a twice weekly treatment regimen. The drug is administered to humans in a twice-weekly treatment regimen with a minimum dosing interval of 3 days and a maximum dosing interval of 4 days. Administration to humans is preferably by a twice weekly treatment regimen for six weeks. Preferably, the total dose of polysulfated polysaccharide administered in the treatment regimen is about 200-300 mg / kg. It is 4000mg.

[0111] Thus, dosing is adjusted for heavier or lighter weight individuals. The regimen may be adapted depending on the severity of pain experienced by the subject. In some cases, patients may experience high levels of pain. If you experience pain in the bell, it is advisable to reach the therapeutic load of the PPS as quickly as possible. This may be, for example, about 1.0 mg / kg or more per day until pain is relieved. Administration of the above PPS may be required.

[0112] If administration is by injection, this is usually performed in a clinical setting by a nurse / physician. Those skilled in the art will appreciate that the key to successful treatment is to administer sufficient PPS to achieve an optimal therapeutic dose. It will be understood that the administration of PPS to a subject is It is known that the loading can be done over time, e.g., 1 mg PPS / day for 4-5 days. For severe chronic cases, subjects should be administered the drug more frequently than once a year. It is thought that multiple treatment courses, perhaps two or three courses per year, may be required. can be done.

[0113] From a safety perspective, a lower dose range (1 ~2 mg PPS / kg or a fixed dose of about 25-50 mg PPS) is preferred. If S is a known anticoagulant and the basal APT is increased by a higher dose (>3 mg PPS / kg or The blood pressure can be increased by 150-200 mg (fixed dose) and bleeding from any open wound can be prevented. This is because it may encourage

[0114] Although administration by injection is preferred, oral or topical formulations of PPS may be used initially IM or SC. It may also be used as a follow-up (maintenance dose) for the treatment of PPS. It can also be administered orally. For administration by IV infusion, the dose is 0.5 to 1 ml per day. The lower dose of g PPS / kg is preferred.

[0115] The present disclosure also contemplates co-administration of polysulfated polysaccharides with other therapeutic agents, such as NGF sequestrants. When other therapeutic agents are utilized in combination with the compounds of the invention, they may be, for example, Is listed in the Physician Desk Reference (PDR) or may otherwise be used in amounts determined by one of skill in the art.

[0116] The specific dose level and dosing frequency for any particular patient may vary. The activity of the particular compound utilized, the metabolic stability and the length of action of that compound, age, body weight , general health, sex, diet, mode and time of administration, excretion rate, drug combinations, specific conditions The results are expected to depend on a variety of factors, including the severity of the condition, and the host receiving the therapy. It will be understood that, for example, larger animals will require larger doses. By way of example, a large animal such as a horse would require a fixed dose of approximately 4000 mg. It may be necessary.

[0117] Determination of suitability of the disclosed treatments may be assessed using patient-reported outcome measurement instruments. Patient-reported pain and / or functional outcomes may be used. the Numerical Rating Scale (NRS) for pain

[17] ; Lysholm Knee Score for Function

[18] ; pain, symptoms, function and quality of life. the Oarthritis Outcome Score (KOOS)

[19] ; ​​and the Ossow for low back pain function. Oswestry Disability Index (Oswestry Back Pain Disability Questionnaire) (also known as the Oswestry Low Back Pain Disability Questionnaire)

[0020] .

[0118] Preferably, pain in the mammal is reduced following administration. Preferably, the decrease is determined by a rating scale. is a numeric rating scale (NRS).

[0119] Preferably, administration of the polysulfated polysaccharide or an acceptable salt thereof further improves the health of the mammal. Preferably, the administration of the polysulfated polysaccharide or an acceptable salt thereof is The weight of the dairy animal is further maintained. Preferably, the polysulfated polysaccharide or an acceptable salt thereof is Administration further maintains or improves musculoskeletal integrity and / or function. Administration of polysulfated polysaccharides or acceptable salts thereof may improve skeletal integrity and / or function. Preferably, the administration of the polysulfated polysaccharide or an acceptable salt thereof is , further maintaining or improving joint or bone integrity and / or function. Administration of polysulfated polysaccharides or acceptable salts thereof may further improve bone integrity and / or function. maintain or improve

[0120] Preferably, the administration of the polysulfated polysaccharide or an acceptable salt thereof is to treat cancer-induced The present invention relates to a method for preventing bone fractures in a mammal suffering from bone cancer. The administration of an acceptable salt of the compound of the present invention delays the time to cancer-induced bone fractures and The patient is suffering from bone cancer. Preferably, the administration of the polysulfated polysaccharide or an acceptable salt thereof is Prevent or reverse bone loss. Preferably, the polysulfated polysaccharide or its tolerable derivative is The administration of the salt prevents or reverses the dysregulation of mineral homeostasis. Preferably, the mineral homeostasis is calcium and / or phosphate homeostasis. In addition, administration of polysulfated polysaccharides or acceptable salts thereof may prevent nerve compression syndrome or Or restore.

[0121] Preferably, function in the mammal improves following treatment. Preferably, there is improvement as determined by the Lysholm Knee Assessment. [Brief description of the drawings]

[0122] [Figure 1] FIG. 1 is an example of an NGF / proNGF ELISA plate layout. [Diagram 2]Representative Alizarin Red staining of differentiated KOA and NOF osteoblast / osteocyte-like cultures. [Diagram 3] Gene expression of osteoblast / osteocyte differentiation markers in KOA cultures from three donors (KOA1-3): DMP1. A) KOA1-DMP1: 18S mRNA expression; B) KOA2-DMP1: 18S mRNA expression; and C) KOA3-DMP1 mRNA expression. Data are triplicate real-time RT-PCR reactions (mean + standard deviation (SD)) for expression of 18S rRNA. [Figure 4] Gene expression of osteoblast / osteocyte differentiation markers in KOA cultures from three donors (KOA1-3:SOST). A) KOA1-SOST: 18S mRNA expression; B) KOA2-SOST: 18S mRNA expression; and C) KOA3-SOST: 18S mRNA expression. Data are triplicate real-time RT-PCR reactions (mean + standard deviation (SD)) for expression of 18S rRNA. [Diagram 5] Gene expression of osteoblast / osteocyte differentiation markers in KOA cultures from three donors (KOA1-3:OCN). A) KOA1-OCN: 18S mRNA expression; B) KOA2-OCN: 18S mRNA expression; and C) KOA3-OCN: 18S mRNA expression. Data are triplicate real-time RT-PCR reactions (mean + standard deviation (SD)) for expression of 18S rRNA. [Figure 6] Gene expression of osteoblast / osteocyte differentiation markers in KOA cultures from three donors (KOA1-3):RANKL. A) KOA1-RANKL: 18S mRNA expression; B) KOA2-RANKL: 18S mRNA expression; and C) KOA3-RANKL: 18S mRNA expression. Data are triplicate real-time RT-PCR reactions (mean + standard deviation (SD)) for expression of 18S rRNA. [Figure 7]Gene expression of osteoblast / osteocyte differentiation markers in KOA cultures from OPGs of three donors (KOA1-3). A) KOA1-OPG: 18S mRNA expression; B) KOA2-OPG: 18S mRNA expression; and C) KOA3-OPG: 18S mRNA expression. Data are triplicate real-time RT-PCR reactions (mean + standard deviation (SD)) for expression of 18S rRNA. [Figure 8] FIG. 1. NGF expression by human bone cells. Relative NGF mRNA expression over differentiation of KOA cultures for three donors was analyzed. Data are triplicate real-time RT-PCR reactions (mean + standard deviation (SD)) for expression of 18S rRNA. A) KOA1-NGF: 18S mRNA expression; B) KOA2-NGF: 18S mRNA expression; and C) KOA3-NGF: 18S mRNA expression. [Figure 9] Confocal microscopy images showing NGF protein expression (red) by freshly harvested human bone cells (rhs) compared to isotype control IgG immunostaining (lhs) Φ. Nuclei (blue) are stained with DAPI. [Figure 10] Figure 1 shows secretion levels of NGF protein as determined by human NGF ELISA in cultures of 28-day differentiated NOF osteocyte-like cells that were either untreated (UT) for an additional 72 hours or pretreated with 1, 5 and 50 μg / mL PPS for the same time periods. Data are the mean + SD of triplicate reactions. A) -NOF1; B) -NOF2; C) -NOF3; and D) NOF: all donors. [Figure 11] Figure 1. Secretion levels of NGF protein as determined by human NGF ELISA in cultures of 28-day differentiated KOA osteocyte-like cells untreated (UT) for an additional 72 hours or pretreated with 1, 5 and 50 μg / mL PPS for the same time periods. Data are the mean + SD of triplicate reactions. A) -KOA1; B) -KOA2; C) -KOA3; and D) KOA: all donors. [Figure 12]Apparent levels of NGF secreted by NOF osteocyte-like cultures in response to a combination of rhTNF and PPS. Data are the mean + SD of supernatants collected from triplicate wells. A) -NOF1; B) -NOF2; C) -NOF3; and D) NOF: all donors. [Figure 13] Apparent levels of NGF secreted by KOA osteocyte-like cultures in response to a combination of rhTNF and PPS. Data are the mean + SD of supernatants collected from triplicate wells. A) -KOA1; B) -KOA2; C) -KOA3; and D) KOA: all donors. [Figure 14] Secretion of proNGF from cultures of NOF osteocyte-like cells treated with a combination of rhTNF and PPS. Data are the mean + SD of supernatants collected from triplicate wells. Significant differences relative to untreated control (UT) are indicated by * (p<0.05); significant differences relative to cultures treated with rhTNF are indicated by # (p<0.05). A) -NOF1; B) -NOF2; C) -NOF3; and D) NOF: all donors. [Figure 15] Secretion of proNGF from cultures of NOF and KOA osteocyte-like cells treated with a combination of rhTNF and PPS. Data are the mean + SD of supernatants collected from triplicate wells. Significant differences relative to untreated control (UT) are indicated by * (p<0.05); significant differences relative to cultures treated with rhTNF are indicated by # (p<0.05). A) -KOA1; B) -KOA2; C) -KOA3; and D) KOA: all donors. [Figure 16] Effect of the combination of rhTNF and PPS on gene expression in KOA osteocyte-like cultures. Real-time RT-PCR was performed for NGF mRNA. Data are the mean + SD of triplicate real-time RT-PCR reactions for expression of 18S rRNA. A) KOA1-NGFR: 18S mRNA expression; B) KOA2-NGFR: 18S mRNA expression. [Figure 17]Effect of the combination of rhTNF and PPS on gene expression in KOA osteocyte-like cultures. Real-time RT-PCR was performed for IL-6 mRNA. Data are the mean + SD of triplicate real-time RT-PCR reactions for expression of 18S rRNA. A) KOA1-IL6: 18S mRNA expression; B) KOA2-IL6: 18S mRNA expression. [Figure 18] FIG. 1. Effect of the combination of rhTNF and PPS on gene expression in KOA osteocyte-like cultures. Real-time RT-PCR was performed for COX2 mRNA. Data are the mean + SD of triplicate real-time RT-PCR reactions for expression of 18S rRNA. A) KOA1-COX2: 18S mRNA expression; B) KOA2-COX2: 18S mRNA expression. [Figure 19] Gene expression of NGF receptor in KOA1 and KOA2 osteocyte-like cultures. Real-time RT-PCR was performed for NGFR (P75NTR) and NTRKA (TrkA) mRNA. Data are the mean + SD of triplicate real-time RT-PCR reactions for expression of 18S rRNA. Note that in many cases, gene expression with reliable melting curves was undetectable or PCR products were derived from only one or two of the triplicate reactions. A) KOA1-NGFR:18S mRNA expression; B) KOA1-NTRKA:18S mRNA expression; C) KOA2-NGFR:18S mRNA expression; and D) KOA2-NTRKA:18S mRNA expression. [Figure 20] FIG. 1. Effect of PPS treatment on basal and TNFα-induced MMP13 expression in human KOA osteocyte-like cultures. Real-time RT-PCR analysis of MMP13 mRNA relative to 18S rRNA expression. Data are the mean + SD of triplicate real-time RT-PCR reactions. A) KOA1-MMP13:18S mRNA expression; and B) KOA2-MMP13:18S mRNA expression. [Figure 21]FIG. 1. Effect of PPS treatment on basal and TNFα-induced MMP13 expression in human KOA bone cell-like cultures. Confocal microscopy imaging of immunostaining of untreated KOA2 cells stained with isotype control primary antibody or mouse anti-human anti-MMP13 antibody (top row), or KOA2 cells treated with rhTNFα alone or rhTNFα+PPS (50 μg / mL) and stained for MMP13 (green staining). All nuclei are stained with DAPI (blue). A) isotype IgG control; B) untreated; anti-MMP13; C) rhTNF-α-treated; anti-MMP13; and D) rhTNFα+PPS50; anti-MMP13. [Figure 22] Figure 1. NGF expression in isolated human bone cells. Osteocyte-enriched fractions of serial human trabecular bone digests were cultured for 24 h untreated (A), treated with rhTNFα (B), PPS (0.5 μg / mL) (C) or a combination of both (D) and then examined by confocal microscopy for NGF immunoreactivity. Control cells were also immunostained using an isotype-matched negative control primary antibody (E). Images are representative data from two individual donors. Scale bars in each image represent 100 μm. [Figure 23] FIG. 1. Effect of combination of rhTNF and PPS on NGF gene expression in osteocyte-like cultures. Human differentiated osteocyte-like cultures were untreated or pretreated with PPS (0.1 or 1.0 μg / mL) for 24 hours, then pretreated with or without rhTNFα (1 ng / mL) for an additional 48 hours, and then real-time RT-PCR for NGF mRNA was performed. Data are the mean + SD of triplicate real-time RT-PCR reactions normalized to mRNA expression of the housekeeping gene ACTB. [Figure 24] Immunostaining of KOA-derived bone cells for TrkA and P-75. At day 28, differentiated human primary bone cell-like cultures were immunostained for (A) TrkA, (B) P-75 and examined by confocal microscopy as described in Materials and Methods, or stained with an isotype control monoclonal antibody (C). Scale bars represent 50 μm in each case. [Diagram 25] Figure 1: Human differentiated KOA osteoblast / osteocyte cultures express NGF: Cells from three KOA patients were cultured under osteogenic conditions for up to 28 days as described in Materials and Methods. Gene expression was measured by real-time RT-PCR at various time points for A) DMP1; B) SOST; C) OCN; D) NGF. Data (mean + standard error (SEM)) were normalized to expression of 18S rRNA using the 2-(ΔCT) method and are shown for the expression of each gene at the end of the time course pooled from cells of three donors. The mean ΔCT for each time point is shown in parentheses above each histogram. [Figure 26] Diagram of NGF expression in isolated human bone cells and KOA bone. Directly conjugated α-NGF MAb was tested against NCI-H266 cells and compared against directly conjugated negative control antibody, X-63 (top row). Fractions IV-VI of KOA bone digests were similarly stained for NGF and staining was compared against expression of the osteocyte marker SOST / sclerostin (middle row). Double staining revealed intracellular staining for both NGF and SOST in these cells, but no colocalized staining. A negative control IgG1 MAb was used to confirm the specificity of the staining. Finally, as described in Materials and Methods, here, using unconjugated α-NGF MAb, NGF positivity was evident in osteocytes (white arrows) in situ in demineralized KOA bone (bottom row). Bone morphology is revealed by digital interference contrast (DIC). In all cases, nuclei were visualized by DAPI staining (blue). Scale bar represents 50 μm. [Figure 27] Human osteocyte-like cells secrete proNGF. Secretion of proNGF was examined from cultures of KOA osteocyte-like cells treated with a combination of rhTNF and PPS. Data are the mean + SD of supernatants collected from triplicate wells. Significant differences relative to untreated control (UT) are indicated by * (p<0.05); significant differences relative to cultures treated with rhTNF are indicated by # (p<0.05). [Figure 28]FIG. 1. Effect of combination of rhTNF and PPS on NGF gene expression in osteocyte-like cultures. Human differentiated osteocyte-like cultures were untreated or pretreated with PPS (0.1 or 1.0 μg / mL) for 24 hours, then pretreated with or without rhTNFα (1 ng / mL) for an additional 48 hours, and then real-time RT-PCR for NGF mRNA was performed. Data are the mean + SD of triplicate real-time RT-PCR reactions normalized to mRNA expression of the housekeeping gene ACTB. [Figure 29] Human NOF osteocyte-like cells secrete proNGF. Secretion of proNGF was examined from cultures of NOF osteocyte-like cells treated with a combination of rhTNF and PPS. Data are the mean + SD of supernatants collected from triplicate wells. Significant differences relative to untreated control (UT) are indicated by * (p<0.05); significant differences relative to cultures treated with rhTNF are indicated by # (p<0.05). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0123] Example embodiment 1. Nerve Growth Factor (NGF) or Pro-Nerve Growth Factor (pro-NGF) in Mammals F) A method for inhibiting or reducing activity, comprising administering to a subject a polysulfated polysaccharide or an acceptable salt thereof. to a mammal.

[0124] 2. The method of embodiment 1, in which the inhibition or reduction of NGF activity treats pain in a mammal. The method described.

[0125] 3. Mediated by nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) The present invention relates to a method for treating pain caused by a disease, the method comprising administering a polysulfated polysaccharide or an acceptable salt thereof to a patient having such a disease. The method comprises administering to a mammal in need of such treatment.

[0126] 4. The method of any one of embodiments 1 to 3, wherein the pain is musculoskeletal pain.

[0127] 5. The method of embodiment 4, wherein the musculoskeletal pain is non-malignant musculoskeletal pain.

[0128] 6. The method of embodiment 4, wherein the musculoskeletal pain is malignant musculoskeletal pain.

[0129] 7. The method of embodiment 4, wherein the musculoskeletal pain is skeletal pain.

[0130] 8. The method of embodiment 7, wherein the skeletal pain is non-malignant skeletal pain.

[0131] 9. The method of embodiment 8, wherein the skeletal pain is malignant skeletal pain.

[0132] 10. The method of any one of embodiments 7 to 9, wherein the skeletal pain is back pain.

[0133] 11. The method of embodiment 10, wherein the back pain is lower back pain.

[0134] 12. The method of embodiment 7, wherein the skeletal pain is bone and / or joint pain.

[0135] 13. The method of embodiment 12, wherein the skeletal pain is arthralgia.

[0136] 14. The method of embodiment 12, wherein the skeletal pain is bone pain.

[0137] 15. The method of embodiment 14, wherein the bone pain is non-malignant bone pain.

[0138] 16. The method of embodiment 15, wherein the non-malignant bone pain is pain associated with osteoporosis.

[0139] 17. The method of embodiment 15, wherein the non-malignant bone pain is pain associated with an arthritic condition.

[0140] 18. Embodiment 17, in which the arthritic condition is selected from rheumatoid arthritis or osteoarthritis. The method according to

[0141] 19. The method of embodiment 18, wherein the arthritic condition is rheumatoid arthritis.

[0142] 20. The method of embodiment 18, wherein the arthritic condition is osteoarthritis.

[0143] 21. Osteoarthritis of the ankle, hip, knee, shoulder, spine, and wrist 21. The method of embodiment 20, wherein the in an articulating joint is selected from the group:

[0144] 22. The method of embodiment 21, wherein the osteoarthritis is in the hip joint.

[0145] 23. The method of embodiment 21, wherein the osteoarthritis is in the knee joint.

[0146] 24. The method of embodiment 21, wherein the osteoarthritis is in the spine (spondylosis). .

[0147] 25. The method of embodiment 14, wherein the bone pain is malignant bone pain.

[0148] 26. The method of embodiment 25, wherein the malignant bone pain is pain associated with primary bone cancer.

[0149] 27. The method of embodiment 25, wherein the malignant bone pain is pain associated with secondary (metastatic) bone cancer. Method of posting.

[0150] 28. Pain, arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; interstitial cystitis; abdominal pain ;joint pain;endometriosis;pancreatitis;chronic headaches;primary cancer;secondary (metastatic) cancer;back pain;bone Injuries or fractures; diabetic neuropathy; fibromyalgia; migraines; multiple myeloma bone disease; multiple sclerosis osteoporosis, neuralgia, herpetic neuralgia, postherpetic neuralgia, neoplasms, sciatica, and visceral pain. The method described in the example.

[0151] 29. Pain in arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; and interstitial cystitis 29. The method of embodiment 28, wherein the condition is associated with a condition selected from the group consisting of:

[0152] 30. Pain is abdominal pain; joint pain; endometriosis; pancreatitis; chronic headache; primary cancer; secondary (transplant) (metastatic) cancer; back pain; bone injury or fracture; diabetic neuropathy; fibromyalgia; migraine; multiple bone Myeloma bone disease; multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; post-herpetic neuralgia; In embodiments associated with a condition selected from the group consisting of: neoplasm; sciatica; and visceral pain. 28. The method according to claim 28.

[0153] 31. The pain is selected from the group consisting of acute pain; chronic pain; generalized pain and persistent pain. The method according to any one of embodiments 1 to 30.

[0154] 32. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals A polysulfated polysaccharide or an acceptable salt thereof for inhibiting or reducing GF activity, and an acceptable excipient.

[0155] 33. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals Polysulfated polysaccharides or their corresponding receptors for use in inhibiting or reducing GF activity. A composition comprising the salt and an acceptable excipient.

[0156] 34. The method of embodiment 32 or embodiment 33, wherein the inhibition or reduction treats pain in a mammal. 34. The composition of claim 33.

[0157] 35. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals Polysulfated polysaccharides or their tolerable derivatives for treating pain mediated by GFs. A composition comprising a salt and an acceptable excipient.

[0158] 36. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals Polysulfated polysaccharides or their derivatives for use in treating pain mediated by fibroblast growth factor (GF) and an acceptable excipient.

[0159] 37. The composition of any one of embodiments 32 to 36, wherein the pain is musculoskeletal pain.

[0160] 38. The composition of embodiment 37, wherein the musculoskeletal pain is non-malignant musculoskeletal pain.

[0161] 39. The composition of embodiment 37, wherein the musculoskeletal pain is malignant musculoskeletal pain.

[0162] 40. The composition of embodiment 37, wherein the musculoskeletal pain is skeletal pain.

[0163] 41. The composition of embodiment 40, wherein the skeletal pain is non-malignant skeletal pain.

[0164] 42. The composition of embodiment 40, wherein the skeletal pain is malignant skeletal pain.

[0165] 43. The composition of any one of embodiments 40 to 42, wherein the skeletal pain is back pain.

[0166] 44. The composition of embodiment 43, wherein the back pain is lower back pain.

[0167] 45. The composition of embodiment 40, wherein the skeletal pain is bone and / or joint pain.

[0168] 46. ​​The composition of embodiment 45, wherein the skeletal pain is arthralgia.

[0169] 47. The composition of embodiment 45, wherein the skeletal pain is bone pain.

[0170] 48. The composition of embodiment 47, wherein the bone pain is non-malignant bone pain.

[0171] 49. The composition of embodiment 48, wherein the non-malignant bone pain is pain associated with osteoporosis.

[0172] 50. The composition of embodiment 48, wherein the non-malignant bone pain is pain associated with an arthritic condition.

[0173] 51. The method of embodiment 50, wherein the arthritic condition is selected from rheumatoid arthritis or osteoarthritis. The composition described in

[0174] 52. The composition of embodiment 51, wherein the arthritic condition is rheumatoid arthritis.

[0175] 53. The composition of embodiment 51, wherein the arthritic condition is osteoarthritis.

[0176] 54. Osteoarthritis of the ankle, hip, knee, shoulder, spine, and wrist The composition of embodiment 53, wherein the composition is in an articulating joint selected from the group consisting of:

[0177] 55. The composition of embodiment 54, wherein the osteoarthritis is in the hip joint.

[0178] 56. The composition of embodiment 54, wherein the osteoarthritis is in the knee joint.

[0179] 57. The composition of embodiment 54, wherein the osteoarthritis is in the spine (spondylosis). thing.

[0180] 58. The composition of embodiment 47, wherein the bone pain is malignant bone pain.

[0181] 59. The composition of embodiment 58, wherein the malignant bone pain is pain associated with primary bone cancer. .

[0182] 60. The method according to embodiment 58, wherein the malignant bone pain is pain associated with secondary (metastatic) bone cancer. The composition described above.

[0183] 61. Pain, arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; interstitial cystitis; abdominal pain ;joint pain;endometriosis;pancreatitis;chronic headaches;primary cancer;secondary (metastatic) cancer;back pain;bone Injuries or fractures; diabetic neuropathy; fibromyalgia; migraines; multiple myeloma bone disease; multiple sclerosis osteoporosis, neuralgia, herpetic neuralgia, postherpetic neuralgia, neoplasms, sciatica, and visceral pain. or a composition as described in one example.

[0184] 62. Pain in arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; and interstitial cystitis The composition of embodiment 61, wherein the composition is associated with a condition selected from the group consisting of:

[0185] 63. Pain: abdominal pain; joint pain; endometriosis; pancreatitis; chronic headache; primary cancer; secondary (transplant) (metastatic) cancer; back pain; bone injury or fracture; diabetic neuropathy; fibromyalgia; migraine; multiple bone Myeloma bone disease; multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; post-herpetic neuralgia; In embodiments associated with a condition selected from the group consisting of: neoplasm; sciatica; and visceral pain. 62. The composition described in 61.

[0186] 64. The pain is selected from the group consisting of acute pain; chronic pain; generalized pain and persistent pain. The composition of any one of embodiments 32 to 63.

[0187] 65. Pain is classified as a nociceptive pain state; a neuropathic pain state; an acute pain state; or a chronic pain state. condition;generalized pain condition;persistent pain condition;non-malignant pain condition;malignant pain condition;chronic pain condition;generalized pain condition Any one of embodiments 32 to 36, wherein the patient is selected from the group consisting of chronic pain conditions and persistent pain conditions. The composition described.

[0188] 66. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals Use of polysulfated polysaccharides or acceptable salts thereof in inhibiting or reducing GF activity - Patents.com .

[0189] 67. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals A polysulfated polysaccharide or a polysaccharide thereof in the manufacture of a medicament for inhibiting or reducing GF activity. Use of acceptable salts.

[0190] 68. The method of embodiment 66 or embodiment 67, wherein the inhibition or reduction treats pain in a mammal. 68. The use according to form 67.

[0191] 69. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals Polysulfated polysaccharides or acceptable salts thereof in treating pain mediated by GF Use of.

[0192] 70. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals The use of polysulfated polysaccharides or or its acceptable salts.

[0193] 71. The use according to any one of embodiments 66 to 70, wherein the pain is musculoskeletal pain.

[0194] 72. The use according to embodiment 71, wherein the musculoskeletal pain is non-malignant musculoskeletal pain.

[0195] 73. The use according to embodiment 71, wherein the musculoskeletal pain is malignant musculoskeletal pain.

[0196] 74. The use according to embodiment 71, wherein the musculoskeletal pain is skeletal pain.

[0197] 75. The use according to embodiment 74, wherein the skeletal pain is non-malignant skeletal pain.

[0198] 76. The use according to embodiment 74, wherein the skeletal pain is malignant skeletal pain.

[0199] 77. The use according to any one of embodiments 74 to 76, wherein the skeletal pain is back pain.

[0200] 78. The use according to embodiment 77, wherein the back pain is lower back pain.

[0201] 79. The use according to embodiment 74, wherein the skeletal pain is bone and / or joint pain.

[0202] 80. The use according to embodiment 79, wherein the skeletal pain is arthralgia.

[0203] 81. The use according to embodiment 79, wherein the skeletal pain is bone pain.

[0204] 82. The use according to embodiment 81, wherein the bone pain is non-malignant bone pain.

[0205] 83. The use according to embodiment 82, wherein the non-malignant bone pain is pain associated with osteoporosis.

[0206] 84. The use according to embodiment 82, wherein the non-malignant bone pain is pain associated with an arthritic condition.

[0207] 85. Embodiment 84, in which the arthritic condition is selected from rheumatoid arthritis or osteoarthritis. The use described in.

[0208] 86. The use according to embodiment 85, wherein the arthritic condition is rheumatoid arthritis.

[0209] 87. The use according to embodiment 85, wherein the arthritic condition is osteoarthritis.

[0210] 88. Osteoarthritis of the ankle, hip, knee, shoulder, spine, and wrist The use according to embodiment 87, which is in an articulating joint selected from the group:

[0211] 89. The use according to embodiment 88, wherein the osteoarthritis is in the hip joint.

[0212] 90. The use according to embodiment 88, wherein the osteoarthritis is in the knee joint.

[0213] 91. The use according to embodiment 88, wherein the osteoarthritis is in the spine (spondylosis). .

[0214] 92. The use according to embodiment 81, wherein the bone pain is malignant bone pain.

[0215] 93. The use according to embodiment 92, wherein the malignant bone pain is pain associated with primary bone cancer.

[0216] 94. The method according to embodiment 92, wherein the malignant bone pain is pain associated with secondary (metastatic) bone cancer. Use of the above.

[0217] 95. Pain, arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; interstitial cystitis; abdominal pain ;joint pain;endometriosis;pancreatitis;chronic headaches;primary cancer;secondary (metastatic) cancer;back pain;bone Injuries or fractures; diabetic neuropathy; fibromyalgia; migraines; multiple myeloma bone disease; multiple sclerosis osteoporosis, neuralgia, herpetic neuralgia, postherpetic neuralgia, neoplasms, sciatica, Any of embodiments 66 to 70, wherein the condition is associated with a condition selected from the group consisting of: or use as described in one example.

[0218] 96. Pain in arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; and interstitial cystitis The use according to embodiment 95, in association with a condition selected from the group consisting of:

[0219] 97. Pain: abdominal pain; joint pain; endometriosis; pancreatitis; chronic headache; primary cancer; secondary (transplant) (metastatic) cancer; back pain; bone injury or fracture; diabetic neuropathy; fibromyalgia; migraine; multiple bone Myeloma bone disease; multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; post-herpetic neuralgia; In embodiments associated with a condition selected from the group consisting of: neoplasm; sciatica; and visceral pain. 95. Use as described in claim 95.

[0220] 98. The pain is selected from the group consisting of acute pain; chronic pain; generalized pain and persistent pain. The use according to any one of embodiments 66 to 97.

[0221] 99. Pain is classified as: nociceptive pain state; neuropathic pain state; acute pain state; chronic pain state condition;generalized pain condition;persistent pain condition;non-malignant pain condition;malignant pain condition;chronic pain condition;generalized pain condition Any one of embodiments 66 to 70, wherein the patient is selected from the group consisting of chronic pain conditions and persistent pain conditions. Use as described.

[0222] 100. Polysulfated polysaccharides are high molecular weight heparin, low molecular weight heparin, heparan sulfate, poly Pentosan polysulfate, chondroitin polysulfate, chitosan polysulfate, dermatan polysulfate Rodexide, dextran sulfate, polysulfated inulin, sulfated lactobionic acid amide, sulfate Oxidized bis-aldonic acid amide, sucrose octasulfate, fucoidan-1, fucoidan-2, sulfur Oxidized beta-cyclodextrin, sulfated gamma-cyclodextrin and other However, inositol hexasulfate is preferably selected from the group consisting of small sulfated compounds. 99. The method, composition or use according to any one of claims 1 to 99.

[0223] 101. Polysulfated polysaccharides are high molecular weight heparin, low molecular weight heparin, and polysulfated pentose. Polysulfate (PPS), chondroitin polysulfate and chitosan polysulfate. 101. The method, composition or use according to embodiment 100,

[0224] 102. Polysulfate pentosan (PPS) is a sodium salt of polysulfate pentosan (Na PPS), magnesium salt of polysulfate pentosan (MgPPS), It consists of calcium salt (CaPPS) and zinc salt of pentosan polysulfate (ZnPPS). 102. The method, composition or use of embodiment 101, selected from the group consisting of:

[0225] 103. Polysulfate pentosan (PPS) is a sodium polysulfate pentosan (NaPP 103. The method, composition or use of embodiment 102, wherein

[0226] 104. Inhibition, reduction, or treatment of intraventricular, intravesicular, or intrathecal Administration via the muscular route, injection via the intramuscular (IM) or subcutaneous (SC) routes, intravenous (IV) V) administration, intra-articular (IA), peri-articular, topical, via suppository, or The method, composition or use according to any one of the preceding embodiments, by oral administration. .

[0227] 105. Inhibition or reduction, or treatment using polysulfated polysaccharides or acceptable salts thereof by administering to a mammal an effective amount of about 1-2 mg / kg of mammal per dose. 105. The method, composition or use according to any one of embodiments 1 to 104.

[0228] 106. Inhibition or reduction, or treatment using polysulfated polysaccharides or acceptable salts thereof and administering to a mammal an effective amount of about 2 mg / kg of mammal per dose. 106. The method, composition or use according to any one of embodiments 1 to 105.

[0229] 107. The method according to any one of embodiments 104 to 106, wherein administration is by injection. Composition or Use.

[0230] 108. The method, composition or method according to embodiment 107, wherein administration is by subcutaneous (SC) injection. use.

[0231] 109. The method according to embodiment 108, wherein the SC injection is a slow SC injection. Composition or Use.

[0232] 110. Administration to humans is by dosing in a once-daily, twice-weekly, or three-weekly treatment regimen. 110. The method, composition or use according to any one of embodiments 104 to 109.

[0233] 111. The method according to embodiment 110, wherein the administration to a human is by dosing in a twice-weekly treatment regimen. The method, composition or use described herein.

[0234] 112. The therapeutic regimen for human administration is twice weekly with a minimum dosing interval of 3 days and a maximum dosing interval of 4 days. The method, composition or use according to embodiment 111, by administration in men.

[0235] 113. Administration to humans is by twice weekly treatment regimen for 6 weeks, The method, composition or use according to embodiment 111 or embodiment 112.

[0236] 114. The total dose of polysulfated polysaccharide administered in the treatment regimen is approximately 200-4000 mg. 114. The method, composition or use of embodiment 113, wherein

[0237] 115. Inhibition or reduction, or treatment using polysulfated polysaccharides or acceptable salts thereof The compound is administered to a mammal in an effective amount ranging from about 10 ng to about 1000 ng as a fixed dose. The method, composition or use according to any one of the preceding embodiments, wherein

[0238] 116. Inhibition, reduction, or treatment of a disease caused by the use of polysulfated polysaccharides or acceptable salts thereof The compound is administered to a mammal in an effective amount ranging from about 10 μg to about 1000 μg as a fixed dose. The method, composition or use according to any one of the preceding claims, wherein

[0239] 117. Inhibition, reduction, or treatment is by intraventricular, intravesical, or intrathecal route 115 or 116, by administering a polysulfated polysaccharide or an acceptable salt thereof to 117. The method, composition or use according to embodiment 116.

[0240] 118. Inhibition or reduction, or treatment using polysulfated polysaccharides or acceptable salts thereof By administering to a mammal an effective amount in the range of about 1 mg to about 25 mg as a fixed dose, The method, composition or use according to any one of embodiments 104 to 113,

[0241] 119. Any of embodiments 1 to 118, in which pain in a mammal is reduced after administration. In one example, the method, composition or use.

[0242] 120. Pain in a mammal is reduced as determined by a rating scale. 120. The method, composition or use according to embodiment 119.

[0243] 121. The method according to embodiment 120, wherein the rating scale is a numerical rating scale (NRS). Methods, compositions or uses.

[0244] 122. The administration of a polysulfated polysaccharide or an acceptable salt thereof further maintains the health of a mammal. 122. The method, composition or use according to any one of embodiments 1 to 121, For.

[0245] 123. The administration of a polysulfated polysaccharide or an acceptable salt thereof helps a mammal to further maintain its body weight. 123. The method, composition or use according to any one of the preceding embodiments, wherein

[0246] 124. The administration of polysulfated polysaccharides or their acceptable salts improves musculoskeletal integrity and / or 124. The method, composition or method according to any one of the preceding embodiments 1 to 123, further maintaining or improving the function or activity of the thing or use.

[0247] 125. The administration of polysulfated polysaccharides or acceptable salts thereof has been shown to improve skeletal integrity and / or 125. The method, composition or use of embodiment 124, wherein said method, composition or use further maintains or improves function.

[0248] 126. The administration of polysulfated polysaccharides or their acceptable salts has been shown to improve joint or bone integrity and 126. The method, composition or method of embodiment 125, further maintaining or improving the function and / or function of is used.

[0249] 127. The administration of polysulfated polysaccharides or acceptable salts thereof has been shown to improve bone integrity and / or 127. The method, composition or use according to embodiment 126, further maintaining or improving function.

[0250] 128. Administration of polysulfated polysaccharides or their acceptable salts improves bone function in cancer-induced bone marrow tumors. 128. The method of claim 1, wherein the mammal is suffering from bone cancer. The method, composition or use described herein.

[0251] 129. Administration of polysulfated polysaccharides or their acceptable salts improves bone function in cancer-induced bone marrow tumors. 129. The method of embodiment 128, wherein the time to fracture is delayed and the mammal is suffering from bone cancer. , composition or use.

[0252] 130. Administration of polysulfated polysaccharides or acceptable salts thereof prevents or reverses bone loss. 128. The method, composition or use according to any one of the preceding embodiments, wherein

[0253] 131. The administration of polysulfated polysaccharides or their acceptable salts is effective in preventing abnormal mineral homeostasis. 131. The method or composition according to any one of the preceding claims, which prevents or restores joint function. Or use.

[0254] 132. Mineral homeostasis is calcium and / or phosphate homeostasis. 132. The method, composition or use according to embodiment 131.

[0255] 133. The administration of polysulfated polysaccharides or their acceptable salts is effective in preventing or treating nerve compression syndrome. 133. The method, composition or use according to any one of the preceding embodiments, wherein the

[0256] 134. Any of embodiments 1 to 133, wherein function in a mammal is improved after administration. In one embodiment, the method, composition or use.

[0257] 135. Function in mammals is determined by the Lysholm knee assessment method described herein. 135. The method, composition or use according to embodiment 134, which, as determined, improves. EXAMPLES

[0258] Study Example 1: Effect of pentose polysulfate on human bone cell response to inflammatory mediators Effects of Sansodium (NaPPS) A pilot study was conducted to investigate the potential effect of PPS on pain mediators. This study also demonstrated that PPS mediates pain mediation via bone cells. We are also paying attention to the potential of this.

[0259] Introduction to research Knee osteoarthritis (KOA) is a common and painful condition, and to control pain It is primarily managed by prescription painkillers. The etiology of KOA is not fully understood, , tumor necrosis factor alpha (TNFα) and interleukin 1-beta (IL-1β) It is known to be associated with increased expression of proinflammatory mediators, including These include matrix metallopeptidase (MMP) family member-13 (MM P-13) and other cartilage-degrading enzymes. Total knee arthroplasty (TKA) is performed to replace the affected joint with a prosthetic total knee replacement. Patient-reported pain was the primary outcome measure for TKA.

[22]

[0260] Bones are innervated with bone sensory neurons originating exclusively from the dorsal root ganglia of the spinal cord.

[23] The secreted neurotropic protein beta-nerve growth factor (NGF) It is a major causative factor for pain in many chronic conditions, including KOA [24–26]. Furthermore, NGF expression was associated with increased TNFα and IL-1 expression in experimental mouse models of osteoarthritis. NGF is known to be induced by both β and β-lactamase

[27] . At least two receptors are expressed in the tropomyosin receptor kinase A (TrkA) and and the pan-neurotropin receptor p75 / NTR, which Neuronal signaling depends on whether the neuron also expresses the co-receptor sortilin. Various biological mechanisms are involved in the regulation of neuronal proliferation and proliferation, and in the regulation of neuronal apoptosis. Tanezumab, a neutralizing antibody treatment targeting NGF, has been shown to be effective in treating NGF-related disorders. Bruxism has shown promising results in the treatment of pain associated with KOA [21, 24-26], and NGF is a key readout and mediator of pain in this condition. NGF is initially expressed in a proprotein form (proNGF). ), which is the enzyme that activates furin or furin-like proprotein convertases. It is post-translationally processed (proteolytically cleaved) to its mature form by the use of phospholipase A (GPA)

[0028] . Currently, soluble NGF levels can be measured using commercially available enzyme-linked immunosorbent assays (ELISAs). Accurate detection of the protein is problematic due to its effect on the readout of proNGF; In this case, proNGF was predictive of both the magnitude and direction of effect (increase or decrease). This prevents the readout of many commercially available ELISA kits in an unpredictable manner

[29] . Therefore, it is necessary to extend the quantitative evaluation to the level of proNGF.

[0261] Pentosan polysulfate (PPS) sodium for the treatment of interstitial cystitis and bladder pain syndrome It is a drug approved by the FDA for the treatment of KOA and has an excellent safety profile

[0030] . It is currently being tested for its efficacy as a treatment for KOA, with encouraging results. The mode of action of PPS appears to be multifactorial, with interstitial bladder involvement. Replenishing the glycosaminoglycan (GAG) layer in the case of its effect on inflammation, as well as cell signaling, particularly nuclear factor kappa-B (NFκB) in chondrocytes

[32] , and IL-1β-iNOS

[33] pathway. Importantly, KOA has been shown to reduce inflammation in the joints. It is a systemic disease involving changes to the subchondral bone, as well as to the synovium and cartilage.

[21] The contribution of each tissue to disease progression and associated pain is not fully understood. In KOA, there is almost complete degradation of the cartilage, with few healthy chondrocytes remaining. This suggests that pain mediators may, to a large extent, originate from the underlying subchondral bone. The most abundant cell type in hard bone tissue is the osteocyte, and these cells They are increasingly recognized as a key regulatory cell type in many local and systemic physiological processes. In osteoarthritis-related conditions, bone cells It is involved in inflammatory and osteolytic reactions to wear particles from implants

[36] and in periprosthetic joints. In conditions of peripheral infection, it also induces a strong pro-inflammatory response to bacteria

[37] .

[0262] Hypothesis Research The present inventors have demonstrated that bone cells produce NGF in the inflammatory environment of the subchondral bone in KOA. We hypothesized that PPS may act by inhibiting this production.

[0263] To test this hypothesis, we performed a randomized controlled trial in patients with advanced KOA who underwent TKA. The effect of PPS on the response of human primary osteocyte-like cultures differentiated from the proximal tibia of patients with osteoporosis Treatment with recombinant TNFα was used as a proinflammatory stimulus, and NGF was used as a proinflammatory stimulus. The effect on the relative expression of a number of genes was examined. Bell was also measured by the respective ELISA.

[0264] The present inventors have demonstrated for the first time that human bone cells can produce NGF. These results suggest that PP may be responsible for the local pain response. S inhibited NGF mRNA transcription and proNGF secretion by bone cells, and these pro We also show that the stimulatory effect of TNFα on the IL-1 deficiency process is reversed. The findings highlight a previously unknown role for bone cells in pain responses and PP These results suggest a mechanism for beneficial pain in KOA patients taking S.

[0265] material and method ethical statement All studies using material from human patients were supported by the Royal Adelaide Hospital. Ital's Human Research Ethics Committee The study was approved by the local ethical committee of the University of Tokyo (approval number 130114). was obtained along with the patients' written informed consent.

[0266] research design Donors and cells To maximize the clinical relevance of findings from this study, the effects of PPS were evaluated in patients with prosthetic knee joints. Three patients with advanced knee osteoarthritis (KOA) who underwent total knee arthroplasty (TKA) were randomly selected for inclusion. 1) Cells isolated from the subchondral bone of the proximal tibia were cultured in vitro for a period of 28 days. The effect of endothelial cell differentiation on osteocyte-like cultures [36, 38-42] was examined. The group consisted of three patients who underwent total hip arthroplasty (THA) for femoral neck fractures ( Cells isolated from the proximal femur of NOF (group 2) were differentiated and tested similarly. All The donors were female, and the groups were age-matched. The mean age of KOA was 77.0 ± 8.5. The mean age of the NOF group was 77.7 ± 5.5 years (p = 0.91).

[0267] Thaw cryopreserved cells from each donor (all passages 0 or 1) and culture them in T75cm 2 tissue culture The cells were cultured in flasks for 10 days. When the cells became confluent, collagenase / dispersal was performed. Cells were removed by enzyme digestion, washed by centrifugation, counted, and diluted to 5 x 10. 5 cells / mL The cells were then diluted to 1 × 10 5 and 2 × 10 4 12 cells / well each The cells were seeded into either 8-well tissue culture trays or 8-well chamber slides for 24 h. After that, the medium was changed to αMEM, 5% v / v fetal calf serum (FCS), 1.8 mM calcium dihydrogen phosphate, and potassium (KH2PO4), 100 μM ascorbyl 2-phosphate (As2P), 10 mM M HEPES, 10 -8 M dexamethasone and 0.2 mM L-glutamine The medium was replaced with osteogenic differentiation medium. During the differentiation process, samples were analyzed for total RNA preparation and gene expression. For the present analysis, cells were collected in Trizol reagent at 3, 14, and 28 days (see below). Cultures seeded in chamber slides were incubated with 100% CO2 as previously described

[41] . , tested for mineralization in vitro using Alizarin Red staining technique did.

[0268] PPS and TNFα treatment PPS is a wholly owned subsidiary of bene pharmaChem GmbH & Co.KG, Geret The stock solution was manufactured by Striead, Germany at 1.0 mg / mL. The differentiated cells were either left untreated or incubated in culture medium for 72 h. The mice were pretreated with PPS at final concentrations of 1, 5 or 50 μg / mL for 1 h. S was based on the effective and maximal activity levels published in a previous study

[43] . The medium was supplemented with or without recombinant human (rh) TNFα (1 ng / mL). The cells were then replaced with the same concentration of PPS without any additional incubation for 48 hours. As described above, culture supernatants were collected and total RNA and cDNA were prepared. From the nurses, eight conditions (untreated, TNF, PPS1, PPS5, PPS50, TNF+PP S1, TNF+PPS5, TNF+PPS50) were obtained.

[0269] Total RNA production and real-time RT-PCR analysis Due to evidence of PPS interference in the generation of assayable cDNA, residual PPS was removed. In an attempt to remove the RNA, the RNA precipitate was washed with 75% ethanol instead of the usual single wash step. Total lysate was extracted from Trizol lysate according to the manufacturer's instructions, except for washing three times in RNA was prepared. The RNA preparation was analyzed using a Nanodrop microvolume spectrophotometer (Therm The RNA from each sample was tested for yield and purity using the Fisher Scientific PCR kit. Micrograms were analyzed using Superscript™ II kit according to the manufacturer's instructions. The housekeeping gene ( 18S, ACTB) expression, nerve growth factor (NGF) and its receptor NTRK1 (T RKA) and NGFR (P75NTR), MMP13, cyclooxygenase-2 (C OX-2), interleukin-6 (IL-6), RANKL, OPG, OCN, DMP1 Real-time RT-PCR was performed for genes including SOST. The oligonucleotide primer sequences for each are shown in Table 1.

[0270] [Table 1]

[0271] ELISA analysis Culture supernatants stored frozen (-80°C) until use were thawed at 4°C and immediately diluted with the manufacturer's Human NGF (catalog number: EHNGF; Thermo Fisher Scientific) was prepared according to the manufacturer's instructions. r Scientific) or proNGF (Cat. No.: BEK-2226-2 P; Biosensis) protein levels were assayed by ELISA. The overall plate layout is shown generally in Figure 1.

[0272] immunostaining were seeded in 8-well chamber slides and allowed to differentiate for 28 days or freshly digested from bone The cells were left untreated or treated with rhTNFα (1 ng / mL), PPS (50 μg / mL; "PPS50") or PPS50 + rhTNFα. The medium was removed and all wells were rinsed a total of three times with PBS (pH 7.4, Gibco). The cells were then incubated for 1 hour at room temperature with 100 μL of Histochoice (Sigma-Aldrich). After fixation, the wells were rinsed twice with distilled H2O and stained. The cells were incubated in 50 μL of bleach for 20 min at room temperature in a humidified chamber. Blocked with rocking buffer (5% v / v normal rabbit serum in 1x PBS) The cells were then rinsed three times with wash buffer (0.1% v / v normal rabbit serum in PBS). The cells were incubated with mouse monoclonal IgG1 anti-human NGF (25623; Thermo Scientific). o Fisher Scientific), anti-human NGFR (2F1C2; Thermo Scientific) o Fisher Scientific), or anti-human TrkA (6B2; mo Fisher Scientific), or anti-human SOST (MAb 220 902.11;R&D Systems, Minneapolis, MN, USA) Primary antibodies and their respective IgG1 isotype control (MAb 1B5) Primary antibodies and IgG controls were diluted between 1:50 and 1:400. The cells were incubated with the primary antibody for 40 min at 4° C. For unconjugated MAbs The chamber slides were then rinsed three times with washing buffer and stained with nuclear DAPI (1:20 00 dilution; diamidino-2-phenylindole; Thermo Fisher) 50 μL of rabbit α-mouse Alexa-fluor secondary antibody (1:2000 dilution) After 1 hour of incubation at room temperature, the secondary antibody was removed and the wells were The cells were washed three times with washing buffer. Finally, the cells were resuspended in FluoroBrite DMEM (Li fe Technologies) was added to each well and analyzed by confocal microscopy (FV300 0 Confocal Microscope, Olympus Lifescience;Adelaide Mic The images were captured using a fluoroscopy.

[0273] For double labeling purposes, anti-NGF was coupled with fluorescein isothiocyanate (FITC; Sigma Chemical Co., St. Louis, MO, USA) For this, MAb25623 was first incubated overnight at 4°C in carbonate / bicarbonate The solution was dialyzed against salt buffer (1 L; pH 9.6). FITC was diluted to 1 mM in anhydrous DMSO. 15 μL of FITC solution was added to 100 μg of anti-NGF, and the tube was then filled with 10 μg of FITC. The mixture was mixed on a rotator at room temperature for 2 hours. Size exclusion chemistry at Macia Biotech, Piscataway, NJ, USA Unbound FITC was removed using chromatography. NanoDrop One Spectroscopy A spectrophotometer (Thermo Fisher Scientific) was used to measure the fluorescence intensity at 280 nm and The absorbance of 0.5 mL fractions at 492 nm was determined and calculated using the formula: Concentration (mg / mL) = A 28 0-(A 492Determine the concentration of FITC-conjugated antibody by (x0.35) / 1.4 To remove aggregates, the antibody solution was centrifuged at 16,400 RCF for 15 minutes before use. As a positive control for immunostaining, the Harmonizome database [ Rouillard AD, Gundersen GW, Fernandez NF, W ang Z, Monteiro CD, McDermott MG, Ma'ayan A ] was used to culture the small cell lung cancer cell line NCI-H266 (ATCC, Massachusetts, VA). For these assays, FITC conjugates were used. DUGATEX-63 MAb (Biosensis, Thebarton, SA, Australia) laria) was used as a negative control; direct conjugate was incubated for 40 min, Aspirated and wells washed 3 times as above.

[0274] Bones isolated from KOA patients were fixed, decalcified, embedded, and sectioned as described. First, bone sections (5 μm) were heated at 60°C for 15 min to remove excess paraffin. The slides were then melted and dewaxed. For antigen retrieval, the slides were then soaked in 10% distilled water. Incubate in formic acid for 10 min, rinse in PBS, and then immunostain as above did.

[0275] Isolation of human primary bone cells from fresh bone For some experiments, freshly obtained human patient bones were used for immunostaining purposes as described. As previously described

[46] , primary bone cells were used as a source of bone marrow cells.

[0276] Data and statistical analysis: ELISA data was analyzed using GraphPad Prism software (GraphPad Prism, La Jolla, CA, USA) and Holm-Sidak Analysis was performed by two-way analysis of variance (ANOVA) with multiple comparison post-hoc test. p<0.0 A value of 5 was considered statistically significant.

[0277] Results and Discussion NGF expression by cultured bone cells TKA for Knee Osteoarthritis (KOA) or Neck of Femoral Fracture (NOF) Human primary osteoblasts isolated from the subchondral bone of patients undergoing THA were cultured for a period of 28 days. The NOF cultures were cultured under differentiation conditions [35-40, 47]. Overall, the NOF cultures significantly greater than KOA donors, as assayed by rin Red staining The results are consistent with previous unpublished observations (Figure 2). As previously reported by our colleagues for cells isolated from the proximal femur

[41] , It may be related to the proximal femur in NOF and the subchondral proximal tibia in KOA, or may be more The likely explanation is the apparent lack of differentiation of osteoblasts from patients with osteoarthritis. Cells from all donors were able to form prebone-specific fibroblasts by day 14. The tumor showed strong osteocyte / osteocyte characteristics, and in two cases, by day 3, DMP1, SOST and In 2 / 3 cases, DM had progressed by day 28. P1, SOST and OCN mRNA expression declined from day 14 levels, and organelle This coincides with the acquisition of a mature osteocyte-like phenotype associated with the loss of Cultures from all donors also expressed the osteoclastogenesis-related gene RANK1. They expressed mRNAs encoding KL and OPG (Figs. 3 to 7) and showed osteocyte-like phenotypes. The expression of NGF mRNA was consistent with that of the donors tested

[35] . These results suggest that the expression of differentiation markers is relatively abundant in the cultures of The expression peaked on the 2nd day and declined by the 28th day (KOA2 ​​in Fig. 8). - cells secreted significant full-length NGF protein that was detected in the supernatant:KOA11 1.9±48.8pg / ml; NOF85.0±60.3pg / ml.

[0278] In further studies, cells from all KOA donors were transformed into pre-osteocytes / osteocytes by day 3. showed strong signatures of β-lactamase inhibitors and expressed significant mRNA for DMP1, SOST, and OCN. DMP1 and SOST mRNAs were expressed at the delta cycle threshold (ΔC Although expressed to similar levels overall based on T values, OCN mRNA was more abundant Expression of DMP1 and OCN increased by day 14, then decreased by day 28. and acquisition of a mature osteocyte-like phenotype associated with loss of organelles and overall metabolism The expression of NGF mRNA was significantly decreased in all the organelles tested [35, 48]. Similar to the expression of OCN from the cells of the donor, it was relatively abundant in these cultures and This reflects the expression of differentiation markers listed above, which peaked at day 14 and declined by day 28. Furthermore, cells from all KOA donors showed significant total IL-1 expression detected in the supernatants. They secreted long NGF protein (111.9±48.8pg / ml).

[0279] Importantly, cells from all three donors expressed abundant mRNA encoding NGF. We have demonstrated that human bone cells are a source of NGF protein and are involved in the treatment of knee osteoarthritis (Fig. 8). This is consistent with the possibility that this may be a cause of pain responses. We explanted osteocytes from additional donor bones according to our published protocol

[46] . Immunostaining revealed strong positivity for cytoplasmic NGF protein ( Figure 9). This is believed to be the first report of bone cell expression of NGF.

[0280] Effect of PPS pretreatment on the secretion of NGF Human osteocyte-like cultures derived from KOA or NOF osteoblasts were left untreated (control). or pretreated with different concentrations of PPS (1, 5 or 50 μg / mL) for 72 h. The supernatants were tested for mature NGF protein secretion by ELISA. Cells from all donors expressed detectable secreted NGF. Interestingly, the secretion levels were It increased in response to PPS treatment (FIGS. 10 and 11).

[0281] Effect of PPS on TNF-induced NGF and proNGF secretion To further examine the effect of PPS on NGF expression, three KOA and three N Osteocyte-like cultures derived from osteoblasts of OF donors were pretreated with different concentrations of PPS. Three days later, the cells were treated with rhTNFα in the absence or presence of a pretreatment concentration of PPS. The cells were harvested 48 hours after treatment and subjected to ELISA analysis as described in Materials and Methods. A study by Malerba and colleagues

[29] found that maturation in experimental samples Along with NGF, the presence of an immature proprotein form of NGF, proNGF, These effects are not predictable and give false readings in many commercial ELISAs for It was therefore demonstrated that the magnitude and direction of the sintering were inconceivable in this study. ProNGF levels were also measured in treated supernatants as described in and Methods.

[0282] As observed for the mature protein, basal proNGF was expressed in all donor cell cultures. It was detectable in the culture supernatant (Figures 14, 15, and 27). Recombinant human TNFα treatment increased the levels of proNGF in both cultured and NOF cells. Significantly increased NGF expression in response to this proinflammatory stimulus in the setting of osteoarthritis This was consistent with the induction of PPS in cells from all donors assayed. and potently suppressed basal proNGF secretion at all PPS concentrations tested. Importantly from a therapeutic perspective, PPS also inhibited the proliferation of IL-1 in KOA medium at all concentrations tested. In both nutrient and NOF cultures and in Fig. 27, rhT on proNGF secretion It potently reversed the effect of NF (Figures 14 and 15).

[0283] The concentrations of PPS selected were based on those previously reported

[43] . A study by Wes et al. showed that PPS administered subcutaneously to volunteers had a mean dose of approximately 1-3 μg / mL. The plasma concentration of PPS in ml is reported and the effective doses used herein have clinical relevance. However, the lack of a dose response in our assay supports the finding that Remarkably similar findings have been obtained for NOF cells. (Figure 29) NGF expression and its regulation by PPS are involved in the association of bone cells between skeletal sites and pathology. It was suggested that this may be a common feature of

[0284] Effect of PPS on gene expression induced by TNF After 3 days of pretreatment of osteocyte-like cultures with PPS and an additional 48 h treatment time, Isolate total RNA for the purpose of analyzing gene expression using real-time RT-PCR The ratio of absorbance at 260 nm / 280 nm is generally between 1.9 and 2.0. All cultures were cultured at 100°C for 10 min as determined by Nanodrop microspectrophotometer, which showed The quality of RNA isolated from the samples appeared to be high, but with the exception of one or two samples, five or produced complementary DNA (cDNA) from all samples exposed to 50 μg / mL PPS. It was consistently not possible to achieve this with assayable housekeeping genes. (18S, BACT) expression, but in the general presence of PPS, housekeeping All of the 6 donors with severely reduced or absent gene expression This was demonstrated by untreated and TNF-treated samples. This was despite increased attempts to remove potentially contaminating PPS. The present inventors have demonstrated that PPS contamination of precipitated RNA occurs due to the coprecipitation of pentose-like structures with the RNA. They concluded that the ribosomal protein inhibits reverse transcription of RNA into cDNA, possibly due to its similarity to the ribosomal protein. Such inhibitory activity of PPS on reverse transcriptase has been demonstrated previously [30-32 However, after repeated washing of the RNA pellet prior to reverse transcription, as described above, untreated, TNFα-treated, PPS (1 μg / mL) and TNFα+PPS (1 μg / mL)-treated media cDNA was produced from two of the three KOA donor cultures from the culture and these were then ligated. The results were analyzed by real-time RT-PCR, and are shown in Figures 16 to 18.

[0285] Most interesting was the effect of PPS on NGF mRNA expression in human bone cells. The cultures expressed measurable NGF mRNA (Figure 16). In the presence of TNFα, The expression level in one donor (KOA1) was unchanged, whereas in the second donor (KOA2) It should be noted that 48 hours after TNFα exposure is a relatively late time point. This is because many genes are stimulated within minutes to hours of exposure and then Ideally, the time course of rhTNFα-treated cells would be It will be interesting to examine whether PPS alone or in combination with rhTNFα Together, NGF mRNA levels were potently suppressed (FIG. 16).

[0286] We further investigated whether the effect of PPS on NGF / proNGF expression is at the transcriptional level. To investigate this, we used real-time RT-PCR to probe for NGF mRNA expression. Further studies were performed to determine whether exposure to TNFα increased NGF mRNA levels. However, PPS at 1 mg / mL increased the relative expression of N GF expression was significantly decreased, suggesting

[0287] This finding is consistent with the protein expression data, suggesting that PPS may be involved in the expression of pain mediators. This supports the hypothesis that IFN-γ reverses the effects of proinflammatory mediators in KOA. Expression by bone cells of two of the known receptors for NGF was also observed in the immunohistochemical study. The high affinity receptor tropomyosin receptor kinase (PKK) was also investigated by RT-PCR. The expression of A (TrkA) was confirmed by RT-PCR in both KOA1 and KOA2. 19), consistent with the complete lack of signal by immunostaining / confocal microscopy. The lack of expression of TrkA by human bone cells indicates that only neurons express TrkA in the mouse Castaneda reported that bone expressed detectable levels of this protein. -Consistent with the findings of Corral et al.

[53] . However, very low levels of NGF Receptor P75NTR mRNA was detected, but with sporadic detection across samples tested. As for TRKA (Fig. 19), no detectable immunostaining for this molecule was observed. These observations suggest that NGF expression by osteocytes is paracrine in bone. This work supports the notion that the most likely target cells are bone sensory neurons. I support it.

[0288] Interestingly, PPS also suppressed the effect of rhTNFα on IL-6 mRNA expression. IL-6 transcription and secretion were also inhibited by TNF-induced NFκB This is a key readout for signal transduction.

[54] Furthermore, rhTNFα inhibits COX2 COX2 mRNA expression was induced by IL-1 (Figure 18). COX2 is an important proinflammatory protein involved in bone destruction. It is an enzyme responsible for the expression of prostaglandin E2 (PGE2), a mediator of It is a target for some nonsteroidal anti-inflammatory drugs (NSAIDs) and is used to relieve OA-related pain. It has been claimed that PPS alone and in combination with rhTNFα are effective. , completely inhibited COX2 mRNA expression.

[0289] These findings suggest that PPS inhibits multiple downstream proinflammatory effects of TNFα. PPS is a cellular response induced by TNFα / TNF receptor signaling. This is consistent with previous reports that it acts as a transcriptional inhibitor of endogenous signaling pathways

[32] .

[0290] Regarding the mechanism of cartilage destruction associated with KOA, the role of phospholipids produced by osteocytes in regulating bone quality was investigated. MMP13 is a key enzyme involved in the destruction of cartilage in OA

[56] , and MMP14 is associated with cartilage destruction in OA. It was also of interest to examine the expression of p13

[57] . PPS is a IL-1β-responsive M It has been implicated as an inhibitor of MP13 expression, but this has not been clearly demonstrated

[33] . In a study, rhTNFα suppressed MMP13 mRNA expression in KOA donor osteocyte-like cultures. This effect was strongly inhibited by PPS treatment (Figure 20). This was confirmed at the protein level by (Figure 21). This supports the notion that its effect on the endothelium protects against cartilage destruction in KOA. Preliminary mechanistic data are provided.

[0291] conclusion This study demonstrated that PPS can be used to inhibit or reduce NGF or pro-NGF activity. Furthermore, this study demonstrated that cartilage repair in patients with knee osteoarthritis can be used For the first time, we present the effect of PPS on human primary bone cells isolated from subcutaneous bone. Osteocytes are involved in pain responses in pathological conditions, including, but not exclusively, KOA. This is the first demonstration of NGF / proNGF production and secretion by this cell type. induced levels of proNGF secretion and TNFα-induced levels of proNGF secretion and TNFα-induced PPS also inhibited TNFα-induced NGF mRNA expression. In conclusion, this suggests that PPS inhibits the expression of collagenase MMP-13 at levels that are not expected to be altered. NGF in subchondral bone and its potential to alleviate pain associated with osteoarthritis of the knee may act at multiple levels to inhibit the release of other pain mediators, if present. Provide evidence that:

[0292] Study Example 2: Effect of rhTNF and NGF gene expression in human bone cell-like cultures Effect of PPS combination We investigated whether PPS exerts its effect on NGF / proNGF expression at the transcriptional level. We describe a follow-up study to determine whether

[0293] material and method The methodology for this study was the same as for Example 1, with the following additions.

[0294] Isolation of human bone cells In some experiments, osteocytes were cultured in human bone marrow following published protocols

[46] . Briefly, bone from TKA was vigorously phosphorylated in sterile PBS. without intervention, followed by recovery of the released cells by centrifugation and washing in PBS. The osteocyte-enriched fraction was then subjected to six consecutive collagenase / dispase / EDTA digestions. Cells from digests 4–6, corresponding to minutes

[46] , were pooled and further analyzed by centrifugation. Wash twice, resuspend in PBS, and then plate into 8-well glass-bottomed chamber slides. After cells were allowed to recover for 72 h, they were immunostained or pretreated with PPS. were treated with PPS and then with a combination of rhTNFα as indicated.

[0295] Results and Discussion NGF expression by freshly isolated human bone cells NGF expression was also examined in cells directly digested from human KOA bone. As previously reported, a large number of cells in the “osteocyte-enriched” fraction of serial bone digests

[46] express NGF. This is our first study on NGF expression by human bone cells. This is the first report of findings from the study. It was qualitatively upregulated in the cells and co-incubated with PPS to upregulate the DWP. It was regulated.

[0296] Effect of PPS on NGF mRNA expression induced by TNF.ALPHA. To examine whether the effect of PPS on NGF / proNGF expression is at the transcriptional level To investigate the effect of NGF on the expression of NGF mRNA in mice, we also used real-time RT-PCR. The relative expression of NGF mRNA was increased by exposure to TNFα (Figure 23). Both 1 and 1 μg / mL PPS significantly increased basal NGF mRNA levels. However, 1 μg / mL PPS inhibited NG in the presence of TNFα. F expression, suggesting that at least part of the effect of PPS on bone cells is at the transcriptional level. This suggests that PPS is a tumour-like cell that is mediated by TNFα / TNF receptor signalling. Previous reports have shown that it acts as a transcriptional inhibitor of intracellular signaling pathways triggered by This is consistent with the findings of

[32] . This suggests that PPS may have a KOA effect on the expression of pain mediators. This supports the hypothesis that IL-16 reverses the effects of proinflammatory mediators in the immune system.

[0297] Conclusions for Example 2 This study examined the proximal tibia of patients with advanced KOA undergoing TKA. The effect of PPS on the response of human primary bone cell-like cultures to recombinant TNFα. Treatment with β-lactam α-lactam was used as a proinflammatory stimulus to examine its effect on the relative expression of NGF mRNA. ProNGF protein levels were also determined.

[0298] The present inventors have again shown that human bone cells are capable of producing NGF and that they These results suggest that PPS may be a cause of localized pain responses. inhibited NGF mRNA transcription and proNGF secretion by bone cells, and these pro We also show that the stimulatory effect of TNFα on the inflammatory process is reversed. The findings highlight the previously unknown role of bone cells in pain responses and PPS These results suggest a mechanism for the beneficial effects of pain in patients with KOA taking

[0299] Study Example 3: NGF expression by freshly isolated human bone cells and in human bone. This study continues from Study Example 2 and presents further studies characterizing NGF expression.

[0300] material and method The methodology for this study was the same as for Example 2.

[0301] Results and Discussion To examine NGF protein expression, we directly co-transfected NCI-H266 cells with The staining of the conjugated anti-NGF antibody was optimized (Figure 26, top). was tested in cells obtained by sequential digestion from human KOA bone. Fractions IV to VI obtained by the method have been previously published to be enriched in mature osteocytes

[0046] . As shown in FIG. 26 (middle), many cells in the osteocyte-enriched fraction express NGF. stained brightly for sclerostin and co-stained for the osteocyte marker SOST / sclerostin. Furthermore, osteocyte expression of NGF was evident in stained sections of human KOA subchondral bone. Combined with the above observations in the differentiation cultures, this suggests that human bone cells This is the first report of our findings regarding NGF expression in mouse bone. Previous studies using a fluorescent reporter system to identify NGF expression in the ulnar mechanoreceptor Expression of NGF in osteoblasts in response to mechanical loading has been reported, but expression in osteocytes has not been reported.

[23] The reporter system used lacked the sensitivity to detect low levels of NGF. Potential or observed differences may be due to differences in species, relative age, skeletal site, stimuli (inflammatory processes, This may be due to the effects of osteoarthritis on osteocyte expression, as well as the effect of osteoarthritis on osteocyte expression. be.

[0302] As described above in Study Example 2, in freshly isolated KOA osteocyte-like cells In an attempt to examine the regulation of NGF protein expression, they or a combination of these. In all cases, NGF immunostaining was However, cells exposed to rhTNF-α alone and PPS alone were qualitatively more resistant than controls. Cells with high NGF expression and treated with a combination of rhTNF-α and PPS showed qualitatively The IgG4-dependent markers showed significantly less staining than the IgG4-dependent markers (Figure 22).

[0303] Due to the unpredictable yield of bone cells from individual patient bones, for quantitative evaluation of treatment However, it is technically difficult to achieve uniform and sufficient numbers of viable, adherent cells across wells. Furthermore, since NGF is a secreted protein, it is difficult to determine its intracellular level. Therefore, quantitative regulation of NGF secretion into the supernatant of osteocyte-like cell differentiation cultures is difficult. was studied.

[0304] In conclusion, PPS can be used to inhibit or reduce NGF or pro-NGF activity. It is demonstrated herein that the present invention can be used in patients with osteoarthritis of the knee. The effect of PPS on human primary bone cells isolated from subchondral bone in humans is reported herein for the first time. It has also been demonstrated to be a potent, but not exclusive, inhibitor of pain response in pathological conditions, including KOA. The bone cells are involved in the response to NGF / proNGF production and secretion by this cell type. Disclosed herein is the first demonstration that PPS is a potent anti-inflammatory drug. Induced levels of proNGF secretion and NGF mRNA induced by TNFα. PPS also inhibited the expression of collagenase MMP-13 induced by TNFα. Taken together, this suggests that PPS may be able to alleviate pain associated with knee osteoarthritis. To restore normal function, NGF and possibly other pain mediators in the subchondral bone were Our results provide evidence that phosphodiesterase inhibitors can act at multiple levels to inhibit catabolite release.

[0305] Numerous variations and / or modifications may be made to the above without departing from the broad overall scope of the present disclosure. It will be understood by those skilled in the art that various modifications may be made to the embodiments. The embodiments are to be considered in all respects as illustrative and not restrictive.

[0306] References

[0307] [Table 2-1]

[0308] [Table 2-2]

[0309]

Table 2-3

[0310]

Table 2-4

[0311]

Table 2-5

[0312]

Table 2-6

[0313]

Table 2-7

[0314]

Table 2-8

[0315]

Table 2-9

Claims

1. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals The method for inhibiting or reducing the activity of a polysulfated polysaccharide or an acceptable salt thereof is A method comprising the step of administering to the mammal.

2. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals Use of a polysulfated polysaccharide or an acceptable salt thereof in inhibiting or reducing activity.

3. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals The use of a polysulfated polysaccharide or its tolerable compound in the manufacture of a medicament for inhibiting or reducing the activity of said polysulfated polysaccharide or its tolerable compound. Use of salt as prescribed.

4. The inhibition or reduction of NGF or pro-NGF activity is a method for treating pain in a mammal.

4. The method according to claim 1 or the use according to claim 2 or claim 3 for treating

5. A method for treating neuronal cell proliferation mediated by nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) The present invention relates to a method for treating pain caused by a rheumatoid arthritis, comprising administering a polysulfated polysaccharide or an acceptable salt thereof to such a patient. A method comprising the step of administering to a mammal in need of treatment.

6. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals Use of polysulfated polysaccharides or acceptable salts thereof in treating pain mediated by 。

7. Nerve growth factor (NGF) or pro-nerve growth factor (pro-NGF) in mammals The use of polysulfated polysaccharides or polysaccharides thereof in the manufacture of a medicament for treating pain mediated by Use of acceptable salts.

8. The method or method according to any one of claims 1, 4 or 5, wherein the pain is musculoskeletal pain. The use according to any one of claims 2, 3, 6 or 7.

9. The method or use according to claim 8, wherein the musculoskeletal pain is skeletal pain.

10. The method or use of claim 9, wherein the skeletal pain is non-malignant skeletal pain.

11. The method or use of claim 10, wherein the skeletal pain is malignant skeletal pain.

12. The skeletal pain is selected from the group consisting of back pain; lower back pain; bone and joint pain; bone pain; and joint pain.

12. The method or use according to any one of claims 9 to 11, which is selected.

13. The method or use of claim 12, wherein the bone pain is non-malignant bone pain.

14. The non-malignant bone pain is selected from the group consisting of osteoporosis; rheumatoid arthritis; and osteoarthritis.

14. The method or use of claim 13, wherein the treatment is selected from pain associated with a condition to be treated.

15. The method or use of claim 12, wherein the bone pain is malignant bone pain.

16. The malignant bone pain is pain associated with primary bone cancer or pain associated with secondary (metastatic) bone cancer.

16. The method or use of claim 15, wherein the pain is

17. The pain is caused by any one of: arthritis; osteoarthritis; rheumatoid arthritis; prostatitis; and interstitial cystitis. The method according to any one of claims 1, 4 or 5, which is associated with a condition selected from the group consisting of: or the use according to any one of claims 2, 3, 6 or 7.

18. The pain is selected from the group consisting of abdominal pain, joint pain, endometriosis, pancreatitis, chronic headache, primary cancer, secondary (metastatic) (sexual) cancer; back pain; bone injury or fracture; diabetic neuropathy; fibromyalgia; migraine; multiple myelopathy Bone disease; multiple sclerosis; osteoporosis; neuralgia; herpetic neuralgia; post-herpetic neuralgia; new 10. The method of claim 1, wherein the inflammatory bowel disease is associated with a condition selected from the group consisting of: chronic inflammation; sciatica; and visceral pain. 4 or 5 or any of claims 2, 3, 6 or 7.

2. The use according to claim 1.

19. The pain is selected from the group consisting of acute pain; chronic pain; generalized pain and persistent pain.

19. The method or use according to any one of claims 1 to 18.

20. The pain is selected from the group consisting of a nociceptive pain state, a neuropathic pain state, an acute pain state, and a chronic pain state. ; Generalized pain state; Persistent pain state; Non-malignant pain state; Malignant pain state; Chronic pain state; Generalized pain state and persistent pain conditions.

8. The method or use according to any one of claims 2, 3, 6 or 7.

21. The polysulfated polysaccharide is selected from the group consisting of high molecular weight heparin, low molecular weight heparin, heparan sulfate, polysulfate, and the like. Acid pentosan, chondroitin polysulfate, chitosan polysulfate, dermatan polysulfate oxide, dextran sulfate, polysulfated inulin, sulfated lactobionic acid amide, sulfated Bis-aldonic acid amide, sucrose octasulfate, fucoidan-1, fucoidan-2, sulfated Beta-cyclodextrin, sulfated gamma-cyclodextrin and other , and small sulfated compounds including inositol hexasulfate.

21. The method or use according to any one of claims 20 to 29.

22. The polysulfated polysaccharide is selected from the group consisting of high molecular weight heparin, low molecular weight heparin, and polysulfated pentosan ( PPS), chondroitin polysulfate and chitosan polysulfate; 22. The method or use of claim 21.

23. The polysulfate pentosan (PPS) is a sodium salt of polysulfate pentosan (NaPP S), magnesium salt of polysulfate pentosan (MgPPS), calcium salt of polysulfate pentosan the group consisting of the zinc salt of pentosan polysulfate (ZnPPS) and the zinc salt of pentosan polysulfate (ZnPPS) 23. The method or use of claim 22, selected from:

24. The pentosan polysulfate (PPS) is sodium pentosan polysulfate (NaPPS).

24. The method or use of claim 23, wherein 25. The method of claim 25, wherein the inhibition, reduction, or treatment is administered by intramuscular (IM) or subcutaneous (SC) routes, cardiac Intravenous (IV) administration; intraarticular administration; (IA) administration, periarticular administration, topical administration, administration via suppository, or oral administration; 25. The method or use according to any one of claims 1 to 24.

26. The inhibition, reduction, or treatment comprises administering to a subject a polysulfated polysaccharide or an acceptable salt thereof. and administering to the mammal an effective amount of about 1 mg / kg to about 2 mg / kg of the mammal per dose.

26. The method or use according to any one of claims 1 to 25, by administering

27. ​​The administration to the mammal is by dosing in a once-daily, twice-weekly, or three-times-weekly treatment regimen.

27. The method or use of claim 25 or claim 26,

28. The total dose of polysulfated polysaccharide administered in the treatment regimen is about 200 to 4000 mg.

28. The method or use of claim 27, wherein

29. The inhibition, reduction, or treatment comprises administering to a subject a polysulfated polysaccharide or an acceptable salt thereof. and administering to said mammal an effective amount ranging from about 10 ng to about 1000 ng as a fixed dose.

26. The method according to any one of claims 1 to 25, by

30. The inhibition, reduction, or treatment comprises administering to a subject a polysulfated polysaccharide or an acceptable salt thereof. and administering to said mammal an effective amount ranging from about 10 μg to about 1000 μg as a fixed dose.

26. The method according to any one of claims 1 to 25, by

31. The inhibition or reduction, or treatment, is achieved by intraventricular, intravesical, or intrathecal routes. 29 or claim 29 by administering the polysulfated polysaccharide or an acceptable salt thereof.

31. The method according to claim 30.

32. The inhibition, reduction, or treatment comprises administering to a subject a polysulfated polysaccharide or an acceptable salt thereof. administering to said mammal an effective amount in the range of about 1 mg to about 25 mg as a fixed dose.

26. The method according to claim 1 , 33. The administration of the polysulfated polysaccharide or an acceptable salt thereof, (i) the health and / or function of the mammal; (ii) musculoskeletal integrity and / or function; (iii) skeletal integrity and / or function; or (iv) joint or bone integrity and / or function 33. The method or use according to any one of claims 1 to 32, further maintaining or improving 。

34. The invention relates to a method for treating cancer-induced bone fractures, comprising administering the polysulfated polysaccharide or an acceptable salt thereof to a patient having a cancer-induced bone fracture.

34. The method of claim 1, wherein the mammal is suffering from bone cancer. Method or use.

35. The administration of the polysulfated polysaccharide or an acceptable salt thereof prevents or reduces bone fractures induced by cancer.

35. The method of claim 34, wherein the mammal is afflicted with bone cancer. is used.

36. The administration of the polysulfated polysaccharide or an acceptable salt thereof, (i) bone loss; (ii) dysregulation of mineral homeostasis; or (iii) Nerve compression syndrome 36. The method or use according to any one of claims 1 to 35, for preventing or ameliorating

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