Colchicine is used to treat connective tissue disorders.

Abstract

A method for treating a patient's connective tissue disorder. The method comprises oral administration of a therapeutically effective dose of colchicine. This treatment can inhibit pathological inflammation, fibrosis, or angiogenesis associated with such a connective tissue disorder. As an example, the connective tissue disorder can be tendinopathy (such as carpal tunnel syndrome). The treatment can have various beneficial effects at the target connective tissue site, such as reducing fibrosis, increasing collagen turnover, increasing extracellular matrix turnover, reducing inflammation, and reducing angiogenesis.

Description

Technical Field

[0001] This invention relates to the drug colchicine. Background Technology

[0002] Repetitive strain injury (sometimes called repetitive stress injury) is a musculoskeletal disorder caused by repetitive overuse and can occur in many different parts of the body. Two common types of repetitive strain injury are tendinitis and carpal tunnel syndrome. Tendinitis is an inflammation of the tendon, resulting in pain and swelling. In carpal tunnel syndrome, for example, repetitive use of the fingers (such as at work or during activities) causes inflammation of the transverse carpal ligament (carpal tunnel) or flexor tendons in the wrist. This leads to compression of the median nerve, which causes pain, numbness, tingling, and weakness in the hand and wrist. These injuries usually require treatment through rehabilitation, physical therapy, NSAIDs (nonsteroidal anti-inflammatory drugs), and sometimes even surgery. However, these approaches do not completely address the underlying fibrosis, inflammation, abnormal healing, or angiogenesis that contribute to the progression of connective tissue disorders. Therefore, alternative treatments are needed for carpal tunnel syndrome and other connective tissue disorders. Summary of the Invention

[0003] Treatment Method. In one aspect, the present invention is a method for treating a patient with a connective tissue disorder. This treatment can suppress pathological inflammation, fibrosis, or angiogenesis associated with such a connective tissue disorder. The treatment can be applied to any suitable type of connective tissue, such as muscle, tendon, tendon sheath, subsynovial connective tissue (SSCT), fascia, or ligament. The term "connective tissue" as used herein excludes joints and blood vessels. Therefore, the term "connective tissue disorder" excludes arthritis and conditions involving vascular inflammation (as examples). The treatment can be particularly suitable for connective tissues composed of dense collagen fibers, such as tendons or ligaments. In some cases, the target treatment site is the upper limb (e.g., arm, wrist, hand, fingers).

[0004] This method involves administering a therapeutically effective dose of colchicine to a patient to treat a connective tissue disorder. Any suitable route of administration can be used, including oral, intravenous, intramuscular, transdermal, etc. In some embodiments, the route of administration is oral, and the dose ranges from 0.2 to 1.9 mg; and in some cases, 0.5 to 1.5 mg. A single dose or multiple (two or more) doses may be given daily. The total duration of colchicine treatment may vary depending on the circumstances. For example, the total duration of treatment may be at least 7 days or at least 14 days. Depending on the circumstances, treatment may be repeated in multiple cycles; for example, a 30-day cycle may be repeated three times.

[0005] This treatment can be administered intermittently. That is, the treatment can be administered for a period of time, stopped for another period of time, and then resumed. For example, the treatment can be administered for 2-6 weeks, then stopped for 2-6 weeks, and then resumed.

[0006] Pyridoxine. Treatment may further include co-administration of pyridoxine (a form of vitamin B6) with colchicine. Pyridoxine can be administered via any suitable route of treatment, including oral, intravenous, intramuscular, transdermal, etc. In some cases, pyridoxine is administered via the same route as colchicine. Pyridoxine can be administered alone or together with colchicine (e.g., simultaneously).

[0007] Pyridoxine enhances the therapeutic efficacy of colchicine. Pyridoxine inhibits many of the same inflammatory cytokines as colchicine. Therefore, pyridoxine can synergize with colchicine to enhance the efficacy of treatments for connective tissue disorders (e.g., anti-inflammatory effects). Furthermore, pyridoxine can support neurological health in the context of connective tissue disorders and help control associated neuropathic symptoms. Pyridoxine inhibits the inflammatory response and supports neuropathy recovery, potentially contributing to nerve regeneration and normalizing metabolic processes essential for neurological health. Therefore, pyridoxine may be particularly useful for inflammatory or fibrotic conditions that lead to neuropathy. In addition, the pain reduction associated with this treatment can further enable increased physical activity, which is beneficial for proper collagen alignment and promotes extracellular matrix transformation, thereby improving tissue health and function.

[0008] As used herein, the term "pyridoxine" encompasses any pharmaceutically acceptable salt form of pyridoxine, such as pyridoxine hydrochloride. Pyridoxine is administered at a therapeutically effective dose, which enhances the effectiveness of colchicine against connective tissue disorders. In some embodiments, the route of administration is oral, and the dose range of pyridoxine is 20-300 mg; and in some cases, 30-250 mg. A single dose or multiple (two or more) doses may be given daily. The pyridoxine used in this invention may be in any suitable pharmaceutically acceptable salt form (e.g., pyridoxine hydrochloride).

[0009] Solid dosage forms. In another aspect, the present invention comprises solid oral dosage forms of colchicine and pyridoxine. As used herein, “solid oral dosage form” means any orally ingestible form for drug administration having solid components. Examples include tablets, capsules, powders, sachets, etc. This combination can be used to treat connective tissue disorders as described above, or any other suitable condition, including gout, as well as those in oncology, immunology, cardiology, or dermatology.

[0010] Both colchicine and pyridoxine possess physicochemical and pharmacokinetic properties, indicating good compatibility and the absence of significant interactions or interferences. Examples of such physicochemical properties include the lack of highly reactive groups, melting points >40°C, and stability at room temperature. Examples of such pharmacokinetic properties include robust water solubility, indicating rapid dissolution in the gastrointestinal tract, and similar time-to-volume (Tg) values ​​after oral ingestion. 最大 Time (pharmacokinetic parameter), and different non-competitive absorption pathways. Both colchicine and pyridoxine are metabolized and excreted primarily via renal clearance. We anticipate no significant interactions between the metabolites. We also anticipate no significant interactions between the drugs or their metabolites during renal clearance. There are no contraindications to prescribing the two drugs together.

[0011] Solid oral dosage forms contain amounts of colchicine and pyridoxine that are effective in synergistic combination therapy. The amount of colchicine contained in a solid dosage form may be 0.2–1.9 mg; and in some cases, 0.5–1.5 mg. The amount of pyridoxine contained in a solid dosage form may range from 20–300 mg; and in some cases, 30–250 mg. The amount of colchicine, pyridoxine, or both may be less than the amounts conventionally used individually to achieve the relevant therapeutic effect.

[0012] Solid oral dosage forms may further contain one or more excipients that function as fillers, disintegrants, wet granulation lubricants, etc. Examples of usable excipients include carnauba wax, hydroxypropyl methylcellulose, polydextrose, polyethylene glycol, triacetin, cellulose gel, croscarmellose sodium, maltodextrin, pregelatinized starch, microcrystalline cellulose, sodium glycolate starch, magnesium stearate, and lactose monohydrate.

[0013] Liquid Formulations. In another aspect, the present invention comprises liquid oral dosage forms of colchicine and pyridoxine. As used herein, “liquid oral dosage form” means any ingestible liquid pharmaceutical mixture intended for oral administration. Examples of such formulations include syrups, elixirs, solutions, suspensions, emulsions, etc. This combination can be used to treat connective tissue disorders as described above, or any other suitable condition, including gout, as well as those in oncology, immunology, cardiology, or dermatology. As explained above, both colchicine and pyridoxine possess physicochemical and pharmacokinetic properties indicating good compatibility with no significant interactions or interferences.

[0014] Liquid oral dosage forms contain amounts of colchicine and pyridoxine, which are effective in synergistic combination therapy. Expressed by weight, the liquid dosage form may contain colchicine in an amount of 0.2–1.9 mg per dose volume; and in some cases, 0.5–1.5 mg. Expressed by weight, the liquid dosage form may contain pyridoxine in an amount of 20–300 mg per dose volume; and in some cases, 30–250 mg.

[0015] Liquid dosage forms can be administered orally by any suitable means. For example, the liquid can be administered via a drinking cup, spoon, syringe, or medicine dropper. The dosage volume of the liquid dosage form may be less than 15 ml, less than 12 ml, or less than 7 ml. Expressed as a concentration, the liquid dosage form may contain colchicine in an amount of 0.02–0.65 mg / ml; and in some cases, 0.05–0.5 mg / ml. Expressed as a concentration, the liquid dosage form may contain pyridoxine in an amount of 2–100 mg / ml; and in some cases, 3–83 mg / ml. The amount of colchicine, pyridoxine, or both may be less than the amount conventionally used to achieve the relevant therapeutic effect. Detailed Implementation

[0016] This invention treats connective tissue disorders. For better understanding, the following is a classification of various treatable connective tissue disorders: tendinopathy (such as tendinosis, tendinitis, or tenosynovitis), nerve compression disorders (such as carpal tunnel syndrome or tarsal tunnel syndrome), pathological angiogenesis within connective tissue (such as angiogenesis that causes pain in tendinopathy), and repetitive use injuries (e.g., repetitive strain injuries or stress injuries).

[0017] Please note that the above classification is neither exclusionary nor exhaustive. Specific conditions may cross or extend into different categories as they are a primary cause or manifestation of other underlying conditions, their proximate (upstream) causes, or their downstream effects. All such alternative classifications should be considered as connective tissue disorders treatable by this invention.

[0018] For example, considering the background of carpal tunnel syndrome, it can have two or more classifications. Carpal tunnel syndrome can be primarily viewed as inflammation of the transverse carpal ligament or flexor tendons in the wrist, causing median nerve compression neuropathy; or it can be viewed as a manifestation of repetitive strain injury of the wrist.

[0019] Alternatively, from an alternative perspective of wrist tendinitis, repetitive strain may be a proximate (upstream) cause of carpal tunnel inflammation or pathological fibrosis (e.g., where inflammation and swelling of the wrist tendons lead to median nerve compression); while median nerve compression (pinching) (causing symptoms of carpal tunnel syndrome) can be considered a downstream effect. In either case, reducing inflammation or fibrosis at the carpal tunnel reduces median nerve compression and alleviates symptoms of carpal tunnel syndrome. Therefore, in the context of carpal tunnel syndrome, all such related conditions encompass the connective tissue disorders treated by this invention.

[0020] In some implementations, connective tissue disorders are inflammatory conditions. A specific example of a related inflammatory condition is tendinitis (inflammation in the tendon), such as Achilles tendinitis, medial epicondylitis (golfer's elbow), lateral epicondylitis (tennis elbow), patellar tendonitis, or wrist tendinitis. In some cases, connective tissue disorders are not associated with radiographically visible calcium deposits in the connective tissue. In some cases, connective tissue disorders are not fluoroquinolone-induced tendinopathy. In some cases, connective tissue disorders are not tenosynovitis.

[0021] Colchicine is a drug primarily used to treat gout. It works by disrupting the formation of microtubules within cells. Microtubules are essential components of the cytoskeleton and are crucial for various cellular processes, including cell division and intracellular material transport. By inhibiting microtubule polymerization, colchicine impairs the ability of white blood cells to migrate and perform their inflammatory functions. This reduces the inflammatory response and helps relieve pain and swelling associated with gout.

[0022] There are multiple possible mechanisms and effects of the treatment methods described in this article. As mentioned above, colchicine is known to have an inhibitory effect on inflammation. Colchicine also has inhibitory effects on fibrosis and angiogenesis. Regarding pathological fibrosis, this treatment method can regulate the collagen healing process by modulating (i.e., increasing or decreasing) collagen synthesis. Regulating the amount of collagen in healing tissue can reduce the accumulation of fibrotic tissue and control the growth of excessively proliferating tissue. Regulating the amount of collagen in healing tissue can also improve the healing process and promote type I collagen.

[0023] Modulating collagen in healing tissue can also limit the structural capacity for angiogenesis. Therefore, this treatment can have a dual anti-angiogenic effect. Through this same anti-angiogenic effect, the treatment can also inhibit the formation of new neurogenesis that leads to pathological innervation in the tissue. Therefore, this treatment can provide a multipronged approach that targets the interactions of inflammation, fibrosis, angiogenesis, swelling, or other abnormal tissue remodeling that occur in connective tissue disorders. This is a significant improvement over conventional therapies that are only effective in symptom relief.

[0024] In the context of tendinopathy, treatment can be based on the patient's stage. Specifically, tendinopathy can have three stages: first, reactive tendinopathy; then, tendon deterioration; and finally, degenerative tendinopathy. Applying treatment after the reactive tendinopathy stage may be the optimal time to reduce collagen synthesis. It should also be noted that tendinitis is characterized by tendon inflammation, while tendon degeneration is characterized by tendon deterioration. Therefore, treatment can address tendinopathy by targeting inflammation, fibrosis, angiogenesis, or a combination thereof.

[0025] Treatment can have several beneficial effects on the pathophysiology of tendinopathy. Tendinopathy is characterized by an increase in type III collagen relative to type I collagen. Therefore, treatment can improve collagen composition (e.g., by reducing the ratio of type III collagen to type I collagen). Other potential beneficial effects on the pathophysiology of tendinopathy include reducing fibrosis, increasing collagen turnover (e.g., by regulating collagen synthesis or increasing collagenase activity), increasing extracellular matrix turnover, reducing inflammation, and reducing angiogenesis.

[0026] Colchicine derivatives. Instead of colchicine, the present invention may use chemical derivatives of colchicine. Such colchicine derivatives may be used in specific medical applications, such as cancer treatment. See Krzywik et al, “NewSeries of Double-Modified Colchicine Derivatives: Synthesis, Cytotoxic Effect and Molecular Docking” (2020). Molecules 25, 3540. Many of these derivatives can have similar physicochemical properties to colchicine and are therefore suitable for the same dosage forms. Similarly, the physicochemical properties of colchicine can be altered by changing its solid form. See U.S. Patent No. 8,309,764 B2 (issued November 13, 2012), entitled "Colchicine Solid-State Forms: Methods of Making and Methods of Use Thereof". Again, many of these modified solid forms can have similar physicochemical properties to colchicine and are therefore suitable for the same dosage forms.

[0027] The selected colchicine chemical derivatives can be metabolites of colchicine. Colchicine has two major metabolites; 2-O-demethylcolchicine and 3-O-demethylcolchicine (2-DMC and 3-DMC, respectively), and one minor metabolite, 10-O-demethylcolchicine.

[0028] Experimental work (clinical) The experimental trial was conducted on a 19-year-old male who had suffered from bilateral upper extremity tendinitis for a duration of 7 months. Symptoms radiated down from his wrists to his fingers and up to his elbows. He reported his symptoms as a severity level of 5-7 on a standard discomfort scale of 1-10. The cause of his extensive tendinitis was repetitive strain injury.

[0029] The patient began treatment with 0.6 mg of colchicine orally once daily. Symptoms began to improve on the second day of treatment. After a total of two weeks of continuous daily administration of colchicine, symptoms almost completely disappeared. He was asymptomatic during his daily activities. At the end of the two-week treatment period, the patient reported a reduction in symptom severity to level 2 on a standard 1-10 discomfort scale.

[0030] To observe the effects of discontinuing colchicine after initial successful treatment, the experimental trial was extended. Following discontinuation, his tendinitis symptoms gradually recurred. After 7 days without treatment, he reported that the severity of his symptoms had worsened to a standard level 5. After another 14 days without treatment (a total of 21 days), his symptoms remained at level 5, occasionally rising to level 6 with increased arm use.

[0031] Experimental work (in vitro) In vitro tendonopathy model: The following experiments were conducted using an in vitro tendonopathy model. Cells derived from human tendons (tendon cells) can serve as in vitro tendon models. See Miller et al., “MicroRNA29a regulates IL-33-mediated tissue remodeling in tendon disease.” Nat Commun 6, 6774 (2015). Cultured tendon cells can be used to study the effects of pro-inflammatory cytokines on the most fundamental components of tendon structure. See Ellis et al., “Defining the profile: Characterizing cytokines in tendon injury to improve clinical therapy.” Journal of Immunology and Regenerative Medicine 16,100059 (2022). When tendons are injured, they trigger a local inflammatory response involving immune cell infiltration and the expression of pro-inflammatory mediators. This inflammatory environment can alter tendon cell physiology, activating them and making them pro-inflammatory. Exposing tendon cells to the pro-inflammatory cytokines TNF-α and IL-1β can mimic the effects of tendon injury. See Smith et al., “Tumour necrosis factor alpha, interleukin 1 beta and interferon gamma have detrimental effects on equine tenocytes that cannot be rescued by IL-1RA ormesenchymal stromal cell-derived factors.” Cell Tissue Res 391, 523-544 (2023).

[0032] Cell culture system. Immortalized, healthy, commercially derived human tendon cells (patellar tendon from a 58-year-old female donor) were reconstructed and seeded in T-75 culture flasks coated with bovine collagen I, and maintained in tendon cell growth medium supplemented with tendon cell growth supplements, FBS, and penicillin / streptomycin. Culture conditions were maintained at 37°C under a humidified 5% CO2 atmosphere. Cells were seeded at a density of 100,000 cells / well in commercially derived 24-well plates pre-coated with rat tail tendon collagen I.

[0033] Treatment Protocol. This study was designed to track the time course of collagen production following cytokine exposure to mimic the effects of tendon injury. The time variable was the duration of cytokine exposure (6 hours, 1 day, 3 days, and 5 days). A 24-hour drug treatment period followed each of these cytokine exposure time periods. Inflammatory symptoms were induced using a combination of IL-1β (1 nM; 17 ng / mL) and TNF-α (10 ng / mL). Two therapeutic interventions were evaluated. The treatment designated "Treatment A" was colchicine (2 nM). The treatment designated "Treatment B" was colchicine (2 nM) plus pyridoxine (100 μM).

[0034] The experimental and treatment groups were as follows: (1) exposure to inflammatory cytokines plus treatment A; (2) exposure to inflammatory cytokines plus treatment B; (3) exposure to inflammatory cytokines (negative control, no drug). The other reference control groups were as follows: (A) treatment A without inflammatory cytokine exposure (drug control only); (B) treatment B without inflammatory cytokine exposure (drug control only); (C) healthy controls (no inflammatory cytokines, no drug treatment).

[0035] Sample collection time. Samples were collected at the following time points after the start of cytokine exposure: • 6-hour sample: collected over 30 hours (6 hours of cytokine exposure + 24 hours of drug treatment) • 1-day sample: collected over 48 hours (24 hours of cytokine exposure + 24 hours of drug treatment) • 3-day samples: collected over 96 hours (72 hours of cytokine exposure + 24 hours of drug treatment) • 5-day sample: collected over 144 hours (120 hours of cytokine exposure + 24 hours of drug treatment; cell culture medium was replenished on day 3) Assays used: Collagen I (human) content in cell culture supernatant was measured using enzyme-linked immunosorbent assay (ELISA). TGF-β1 in cell culture supernatant was measured using the MSD multiplex platform. Other inflammatory cytokines in cell culture supernatant were measured using the MSD multiplex platform.

[0036] Collagen I Results Control. Healthy control tendon cells showed baseline collagen I production of 30 ng / mL. Treatment A alone, without inflammatory stimulation via cytokines, showed collagen I levels of 17 ng / mL. Negative controls (exposed to inflammatory cytokines but without drug treatment) were measured at each time point.

[0037] Six-hour collagen response. The results given below are expressed as the concentration levels of human collagen I in cell culture supernatant. Six hours after cytokine exposure, the negative control showed a moderately reduced collagen I level (24 ng / mL) compared to healthy controls. Treatments A and B produced similar collagen I levels (both 20 ng / mL).

[0038] One-day collagen response. On day 1, the negative control with cytokine exposure maintained collagen I levels reduced to 20 ng / mL. Comparable collagen I levels were shown in the two treatment groups: treatment A at 19 ng / mL and treatment B at 19 ng / mL.

[0039] Three-day collagen response. On day 3, the negative control with cytokine exposure showed partial recovery of collagen I to 23 ng / mL. Treatment A maintained collagen I levels at 21 ng / mL.

[0040] Five-day collagen response. On day 5, the negative control with cytokine exposure showed an increase in collagen I levels to 28 ng / mL. Treatment A maintained a similar level (28 ng / mL), while treatment B produced a moderately reduced level of 22 ng / mL.

[0041] Results Summary: These data indicate that both treatment A (colchicine) and treatment B (colchicine + pyridoxine) maintained stable collagen I production under inflammatory conditions. Cytokine exposure induced an initial inflammatory response, followed by a compensatory increase in collagen I production with drug treatment. Both treatments A and B demonstrated the ability to maintain collagen I levels within physiological ranges, with treatment B (the combination of colchicine and pyridoxine) showing more consistent measurements at earlier time points. These results suggest that colchicine (with and without pyridoxine) maintains collagen I production under inflammatory conditions of tendon injury.

[0042] TGF-β1 results Control. Healthy control tendon cells showed baseline TGF-β1 production of 1.73 ng / mL. Treatment A alone, without inflammatory stimulation via cytokines, showed TGF-β1 levels of 1.90 ng / mL. Negative controls (exposed to inflammatory cytokines but without drug treatment) were measured at each time point.

[0043] Six-hour response. The results given below are expressed as the concentration level of TGF-β1 in the cell culture supernatant. Six hours after cytokine exposure, the negative control showed a significantly reduced TGF-β1 level (to 0.97 ng / mL) compared to the healthy control. Treatment A produced an elevated TGF-β1 level of 1.23 ng / mL (127% of the negative control), while treatment B showed a similar improvement to 1.16 ng / mL (120% of the negative control).

[0044] One-day response. On day 1, the negative control with cytokine exposure maintained a reduced TGF-β1 level of 1.22 ng / mL (71% of healthy baseline). Both treatment groups showed improved TGF-β1 levels: treatment A at 1.28 ng / mL (104% of negative control) and treatment B at 1.43 ng / mL (117% of negative control). The drug-only control showed enhanced TGF-β1 production of 1.90 ng / mL (110% of healthy baseline).

[0045] Three-day response. On day 3, the negative control with cytokine exposure showed TGF-β1 recovery to 1.70 ng / mL (98% of healthy baseline). Treatment A maintained elevated TGF-β1 levels at 1.94 ng / mL (114% of the negative control and 112% of healthy baseline).

[0046] Five-day response. On day 5, the negative control with cytokine exposure showed that TGF-β1 was restarted to inhibition at 1.22 ng / mL (71% of healthy baseline). Treatment A maintained a higher level at 1.32 ng / mL (108% of the negative control), while treatment B showed an excellent response at 1.52 ng / mL (124% of the negative control and 88% of healthy baseline).

[0047] Results Summary. These data indicate that both treatment A (colchicine) and treatment B (colchicine + pyridoxine) effectively counteracted cytokine-induced TGF-β1 inhibition. The inflammatory response initially reduced TGF-β1 levels to approximately 56% of healthy baseline at 6 hours, with persistent inhibition through day 5, except for a temporary recovery on day 3. Both treatments showed the ability to increase TGF-β1 levels above the negative control, with treatment B demonstrating a superior sustained effect through day 5. Treatment A showed a particularly strong effect on day 3 (114% of the negative control), which then lessened by day 5. These results suggest that colchicine, particularly in combination with pyridoxine, effectively maintains TGF-β1 production in the inflammatory context of tendon injury.

[0048] IFN-γ results Controls. For healthy control samples, the IFN-γ level was 8.2 pg / mL. Treatment A alone, without inflammatory stimulation by cytokines, showed an IFN-γ level of 3.1 pg / mL. Negative controls (exposed to inflammatory cytokines but without drug treatment) were measured at each time point.

[0049] Six-hour response. The results given below are expressed as the concentration levels of IFN-γ in cell culture supernatant. Six hours after cytokine exposure, the negative control showed a significantly elevated IFN-γ level of 17.6 pg / mL (215.7% of the healthy level). Treatment A showed a significant decrease to 12.6 pg / mL (71.6% of the negative control and 154.5% of the healthy level), while treatment B resulted in a further increase to 21.4 pg / mL (121.4% of the negative control and 261.8% of the healthy level).

[0050] One-day response. On day 1, the negative control with cytokine exposure maintained elevated IFN-γ levels at 15.6 pg / mL (190.7% of healthy levels). Treatment A showed a modest decrease to 15.3 pg / mL (98.0% of negative controls, 186.9% of healthy levels), while treatment B resulted in a further increase to 19.9 pg / mL (127.6% of negative controls, 243.4% of healthy levels).

[0051] Three-day response. On day 3, the negative control with cytokine exposure showed a sustained increase in IFN-γ, reaching 12.7 pg / mL (154.8% of healthy levels). Treatment A reduced IFN-γ levels to 9.2 pg / mL (72.4% of the negative control and 112.1% of healthy levels).

[0052] Five-day response. On day 5, the negative control with cytokine exposure showed a sustained increase in IFN-γ to 16.3 pg / mL (199.0% of healthy levels). Treatment A showed a modest decrease to 13.1 pg / mL (80.7% of negative controls, 160.7% of healthy levels), while treatment B maintained the level at 15.2 pg / mL (93.2% of negative controls, 185.4% of healthy levels).

[0053] Results Summary. These data indicate that exposure to inflammatory cytokines leads to a sustained increase in IFN-γ levels to approximately 170–190% of healthy baseline. Treatment A (colchicine) showed early efficacy in counteracting this increase, particularly at the 6-hour time point, reducing IFN-γ levels by 28.4% compared to the negative control. This antagonistic effect persisted until day 5, although by orders of magnitude. Treatment B (colchicine + pyridoxine) exhibited a very different response pattern, generally maintaining or increasing the IFN-γ elevation. These results suggest that in inflammatory conditions, colchicine monotherapy can modulate the IFN-γ response more effectively than combination therapy.

[0054] IL-6 results Controls. For healthy control samples, the IL-6 level was 342.3 pg / mL. Treatment A alone, without inflammatory stimulation via cytokines, showed a reduced IL-6 level of 73.7 pg / mL. Negative controls (exposed to inflammatory cytokines but without drug treatment) were measured at each time point.

[0055] Six-hour response. The results given below are expressed as the concentration levels of IL-6 in the cell culture supernatant. Six hours after cytokine exposure, the negative control showed an elevated IL-6 level of 1614.8 pg / mL (471.7% of the healthy level). Treatment A showed a significant increase to 2989.5 pg / mL (185.1% of the negative control and 873.4% of the healthy level), while treatment B resulted in a moderate increase to 2239.1 pg / mL (138.7% of the negative control and 654.1% of the healthy level).

[0056] One-day response. On day 1, the negative control with cytokine exposure maintained elevated IL-6 levels at 1726.6 pg / mL (504.4% of healthy levels). Treatment A showed a further increase to 2114.9 pg / mL (122.5% of negative control, 617.8% of healthy levels), while treatment B showed a decrease to 1543.9 pg / mL (89.4% of negative control, 451.0% of healthy levels).

[0057] Three-day response. On day 3, the negative control with cytokine exposure showed a decrease in IL-6 to 1067.0 pg / mL (311.7% of healthy levels). Treatment A showed a significant increase to 1360.1 pg / mL (127.5% of the negative control and 397.3% of healthy levels), counteracting the cytokine-induced decrease.

[0058] Five-day response. On day 5, the negative control with cytokine exposure maintained a reduced IL-6 level of 696.6 pg / mL (203.5% of healthy levels). Treatment A showed a sustained antagonistic effect, with levels increasing to 708.5 pg / mL (101.7% of negative controls and 207.0% of healthy levels), while treatment B maintained a level of 938.0 pg / mL (134.7% of negative controls and 274.0% of healthy levels).

[0059] Results Summary. These data indicate that exposure to inflammatory cytokines produces a biphasic IL-6 response, with an initial increase followed by a sustained decrease. Treatment A (colchicine) showed a consistent modulatory effect throughout the time course, with a significant antagonistic effect, particularly during the late stage (days 3–5), in which it partially restored IL-6 levels to baseline. Treatment B (colchicine + pyridoxine) showed a more moderate effect, suggesting a potential mechanistic difference in IL-6 regulation between colchicine monotherapy and combination therapy.

[0060] TNF-α results Controls. For healthy control samples, the TNF-α level was 211.3 pg / mL. Treatment A alone, without inflammatory stimulation by cytokines, showed the lowest TNF-α level, 1.1 pg / mL. Negative controls (exposed to inflammatory cytokines but without drug treatment) were measured at each time point.

[0061] Six-hour response. The results given below are expressed as the concentration levels of TNF-α in the cell culture supernatant. Six hours after cytokine exposure, the negative control showed a significant increase in TNF-α levels to 1678.4 pg / mL (794.4% of healthy levels). Treatment A resulted in a further increase to 1958.4 pg / mL (116.7% of negative control, 926.9% of healthy levels), while treatment B showed a modest increase to 1782.0 pg / mL (106.2% of negative control, 843.4% of healthy levels).

[0062] One-day response. On day 1, the negative control with cytokine exposure maintained an elevated level of 1317.2 pg / mL (623.4% of healthy levels). Treatment A showed an increase to 1703.5 pg / mL (129.3% of negative control, 806.3% of healthy levels), while treatment B showed a modest increase to 1460.3 pg / mL (110.9% of negative control, 691.1% of healthy levels).

[0063] Three-day response. On day 3, the negative control with cytokine exposure showed a decrease in TNF-α to 960.1 pg / mL (454.4% of healthy levels). Treatment A increased TNF-α levels to 1303.6 pg / mL (135.8% of the negative control and 617.0% of healthy levels).

[0064] Five-day response. On day 5, the negative control with cytokine exposure showed a sustained decrease in TNF-α to 824.1 pg / mL (390.1% of healthy levels). Treatment A showed a significant increase to 895.9 pg / mL (108.7% of the negative control and 424.1% of healthy levels), while treatment B maintained a level of 1036.5 pg / mL (125.8% of the negative control and 490.6% of healthy levels).

[0065] Results Summary: These data suggest that exposure to inflammatory cytokines leads to a complex TNF-α response pattern, with an initial increase followed by a gradual decrease. Treatment A (colchicine) showed a consistent regulatory effect throughout the time course, with a particularly significant antagonistic effect in the late stage, where it partially restored TNF-α levels to baseline. Treatment B (colchicine + pyridoxine) exhibited a more moderate effect, indicating a different mechanism of action in TNF-α regulation compared to treatment A.

[0066] IL-33 Results Controls. For healthy control samples, the IL-33 level was 8.7 pg / mL. Treatment A alone, without inflammatory stimulation by cytokines, showed a reduced IL-33 level of 3.4 pg / mL. Negative controls (exposed to inflammatory cytokines but without drug treatment) were measured at each time point.

[0067] Six-hour response. The results given below are expressed as the concentration level of IL-33 in the cell culture supernatant. Six hours after cytokine exposure, the negative control showed a significant increase in IL-33 levels to 18.0 pg / mL (206.6% of healthy levels). Treatment A showed a significant decrease to 9.9 pg / mL (55.0% of negative control, 113.6% of healthy levels), while treatment B maintained an elevated level of 16.7 pg / mL (92.8% of negative control, 191.7% of healthy levels).

[0068] One-day response. On day 1, the negative control with cytokine exposure maintained elevated IL-33 levels at 13.1 pg / mL (150.6% of healthy levels). Treatment A showed a modest decrease to 11.2 pg / mL (85.1% of negative controls, 128.1% of healthy levels), while treatment B resulted in a further increase to 19.4 pg / mL (147.4% of negative controls, 221.9% of healthy levels).

[0069] Three-day response. On day 3, the negative control with cytokine exposure showed a sustained increase in IL-33, reaching 10.7 pg / mL (123.1% of healthy levels). Treatment A showed a sustained antagonistic effect, with a decrease to 7.6 pg / mL (70.8% of the negative control and 87.1% of healthy levels).

[0070] Five-day response. On day 5, the negative control with cytokine exposure maintained elevated IL-33 levels at 17.1 pg / mL (195.9% of healthy levels). Treatment A showed a sustained antagonistic effect, with levels decreasing to 14.6 pg / mL (85.4% of negative controls and 167.4% of healthy levels), while treatment B maintained levels at 13.0 pg / mL (76.3% of negative controls and 149.4% of healthy levels).

[0071] Results Summary. These data indicate that exposure to inflammatory cytokines led to a sustained increase in IL-33 levels throughout the experimental period. Treatment A (colchicine) showed a consistent antagonistic effect (i.e., reduction in IL-33) at all time points, with a particularly significant reduction at 6 hours, decreasing IL-33 levels by 45% compared to the negative control. This antagonistic effect persisted until day 5, indicating sustained therapeutic activity. Treatment B (colchicine + pyridoxine) showed minimal modulation of IL-33 levels, suggesting that colchicine monotherapy is more effective than combination therapy for IL-33 regulation.

[0072] VEGF results Controls. For healthy control samples, VEGF levels were 199.0 pg / mL. Treatment A alone, without inflammatory stimulation via cytokines, showed elevated VEGF levels of 621.7 pg / mL. Negative controls (exposed to inflammatory cytokines but without drug treatment) were measured at each time point.

[0073] Six-hour response. The results given below are expressed as VEGF concentration levels in cell culture supernatants. Six hours after cytokine exposure, the negative control showed an increase in VEGF levels to 333.6 pg / mL (167.6% of healthy levels). Treatment A showed a significant increase to 523.8 pg / mL (157.0% of negative controls, 263.2% of healthy levels), while treatment B showed a similar increase to 476.4 pg / mL (142.8% of negative controls, 239.4% of healthy levels).

[0074] One-day response. On day 1, the negative control with cytokine exposure showed elevated VEGF levels of 599.5 pg / mL (301.2% of healthy levels). Treatment A resulted in an increase to 799.7 pg / mL (133.4% of negative controls and 401.8% of healthy levels), while treatment B showed a decrease to 455.4 pg / mL (76.0% of negative controls and 228.8% of healthy levels).

[0075] Three-day response. On day 3, the negative control with cytokine exposure showed an increase in VEGF to 555.0 pg / mL (278.9% of healthy levels). Treatment A showed an increase to 624.9 pg / mL (112.6% of the negative control and 314.0% of healthy levels).

[0076] Five-day response. On day 5, the negative control with cytokine exposure showed a sustained increase in VEGF to 386.6 pg / mL (194.2% of healthy levels). Treatment A maintained an increase to 455.8 pg / mL (117.9% of the negative control and 229.0% of healthy levels), while treatment B showed a level of 518.8 pg / mL (134.2% of the negative control and 260.7% of healthy levels).

[0077] Results Summary. These data indicate that exposure to inflammatory cytokines led to a sustained increase in VEGF levels throughout most of the experimental time. Treatment A (colchicine) showed a consistent increase in VEGF at all time points, with a particularly significant increase at 6 hours, increasing VEGF levels by 57% compared to the negative control. This effect persisted until day 5, indicating sustained activity. Treatment B (colchicine + pyridoxine) showed a similar early increase as colchicine monotherapy, but with variable activity at later time points.

[0078] The foregoing description and embodiments are illustrative only and are not intended to be limiting. Each aspect and embodiment disclosed in this invention can be considered individually or in combination with other aspects, embodiments, and variations of the invention. Furthermore, unless otherwise stated, the steps of the method of the invention are not limited to any particular order of execution. Modifications to these embodiments will be appreciated by those skilled in the art, which incorporate the spirit and essence of the invention. Such modifications are within the scope of the invention.

[0079] Unless the context clearly specifies otherwise, any use of the word "or" in this document is intended to be inclusive and equivalent to "and / or". Thus, for example, expressing "A or B" means A, or B, or both A and B. Similarly, for example, expressing "A, B, or C" means A, or B, or C, or any combination thereof.

Claims

1. A method for treating a patient’s connective tissue disorder, comprising oral administration of colchicine in a dose range of 0.2-1.9 mg.

2. The method according to claim 1, wherein, The method treats target connective tissue, wherein the target connective tissue is tendon, ligament, tendon sheath, or subsynovial connective tissue.

3. The method according to claim 1, wherein, The connective tissue disorder mentioned is tendinopathy.

4. The method according to claim 3, wherein, The tendinopathy mentioned is carpal tunnel syndrome.

5. The method according to claim 1, wherein, The dosage is in the range of 0.5-1.5 mg.

6. The method according to claim 2, wherein, The treatment reduces fibrosis in the target connective tissue.

7. The method according to claim 2, wherein, The treatment promotes collagen conversion in the target connective tissue.

8. The method according to claim 1, wherein, Pyridoxine was not administered to the patient in combination with colchicine.

9. The method according to claim 2, wherein, The treatment increases the ratio of type I collagen to type III collagen.

10. The method according to claim 2, wherein, The treatment promotes extracellular matrix turnover in the target connective tissue.

11. The method according to claim 2, wherein, The treatment reduces inflammation in the target connective tissue.

12. The method according to claim 1, wherein, The treatment reduces angiogenesis in the target connective tissue.

13. The method according to claim 1, wherein, The colchicine is administered multiple times daily.

14. The method according to claim 1, wherein, The treatment lasts for at least 7 days.

15. The method according to claim 14, wherein, The treatment lasts for at least 14 days.

16. The method of claim 1, further comprising administering pyridoxine in combination with the colchicine.

17. The method according to claim 16, wherein, The pyridoxine is administered at doses ranging from 20 to 300 mg.

18. The method according to claim 2, wherein, The target connective tissue does not have X-ray visible calcium deposits.

19. The method according to claim 1, wherein, The connective tissue disorder is not fluoroquinolone-induced tendinopathy.

20. The method according to claim 1, wherein, The connective tissue disorder is not tenosynovitis.