Use of trans-ferulic acid in the preparation of a medicament for the treatment of acute and chronic contact dermatitis

Abstract

The application provides application of tetrahydropalmatine in preparation of a medicine for treating acute and chronic contact dermatitis. Experiments show that tetrahydropalmatine can significantly inhibit redness, desquamation and ear swelling of acute and chronic contact dermatitis, reduce itching, inhibit epidermal thickening and inflammatory infiltration. The mechanism of action includes double inhibition of TSLP and TNF-alpha protein levels, and recovery of EDEM3 expression by directly combining EDEM3 protein (KD=3.69±0.21 muM), inhibition of endoplasmic reticulum stress, and thus down-regulation of epithelial alarm substances IL25, IL33 and TSLP. The effect of tetrahydropalmatine disappears after EDEM3 is silenced, confirming that EDEM3 is a functional target protein. In addition, tetrahydropalmatine is also effective for non-histamine-dependent allergic contact dermatitis, and has no side effects such as skin atrophy and weight loss, and the safety is better than that of dexamethasone. The application also relates to similar application of tetrahydropalmatine and cortex phellodendri extract. The application provides a safe, effective and mechanism-confirmed new type of therapeutic drug for contact dermatitis.

Description

Technical Field

[0001] This invention relates to the field of drugs for treating acute and chronic contact dermatitis, and particularly to the use of corydaline in the preparation of drugs for treating acute and chronic contact dermatitis. Background Technology

[0002] Contact dermatitis is an inflammation of the skin caused by direct contact with irritants (irritant contact dermatitis) or allergens (allergic contact dermatitis). Symptoms include itching, sometimes accompanied by burning. Skin changes include erythema, scaling, swelling, and sometimes blisters and ulcers. The location depends on the site of contact. Treatment includes topical corticosteroids, antipruritic medications, and avoidance of irritants and allergens.

[0003] The information disclosed in the background section is only for enhancing the understanding of the background of this invention, and therefore may contain information that does not constitute prior art known to those skilled in the art. Summary of the Invention

[0004] This invention provides the application of corydaline in the preparation of drugs for treating acute and chronic contact dermatitis.

[0005] In addition, the present invention provides a medicament for treating acute and chronic contact dermatitis, the medicament comprising corydaline.

[0006] Preferably, the dosage form of the drug is a topical preparation, which includes gels, ointments, creams, powders, liquids, or lotions.

[0007] Preferably, the concentration of tebuconazole in the drug is 0.1-1 mg / mL. More preferably, the concentration of tebuconazole in the drug is 0.3-0.9 mg / mL.

[0008] Preferably, tebufenozide (Rot) can be used to dual inhibit TSLP protein levels and TNF-α protein levels, and can inhibit the excessive proliferation of keratinocytes, significantly reduce itching, and has no skin atrophy side effect and no systemic metabolic toxicity.

[0009] Furthermore, this invention also discloses the application of corydaline in the preparation of drugs for treating nonhistamine-dependent allergic contact dermatitis.

[0010] Furthermore, this invention also discloses the application of corydaline in the preparation of drugs for treating allergic contact dermatitis.

[0011] Preferably, the concentration of corydaline A is 1.5 mg / mL.

[0012] Furthermore, this invention also discloses the application of Corydalis extract in the preparation of drugs for treating acute contact dermatitis.

[0013] Furthermore, this invention also discloses a contact dermatitis treatment drug targeting the EDEM3 protein, the drug containing an effective amount of corydaline, and the corydaline exerts its therapeutic effect by directly binding to the EDEM3 protein.

[0014] Preferably, according to the application of the corydaline in the preparation of drugs for treating acute and chronic contact dermatitis or the application of the corydaline in the preparation of drugs for treating nonhistamine-dependent allergic contact dermatitis, the corydaline reduces the symptoms of contact dermatitis by directly binding to the EDEM3 protein, restoring the expression level of EDEM3, and inhibiting endoplasmic reticulum stress.

[0015] Preferably, the dissociation constant K between the corydaline and the EDEM3 protein is... D The value was 3.69 ± 0.21 μM.

[0016] Preferably, the fumaric acid binds to the GLU-58, GLN-55, and ARG-660 amino acid residues on the EDEM3 protein.

[0017] Preferably, the corydaline inhibits the expression of epithelial alarmins TSLP, IL25 and / or IL33 in an EDEM3-dependent manner.

[0018] Preferably, the nonhistamine-dependent allergic contact dermatitis is induced by allergens that do not depend on the histamine pathway, and the drug does not rely on an antihistamine mechanism to take effect.

[0019] Compared with the prior art, the present invention has the following advantages: Corydaline can significantly improve the symptoms of acute and chronic contact dermatitis and is safe, thus confirming the application of corydaline in the preparation of drugs for treating acute and chronic contact dermatitis. Attached Figure Description

[0020] Various other advantages and benefits of the present invention will become apparent to those skilled in the art upon reading the detailed description of the preferred embodiments below. The accompanying drawings are for illustrative purposes only and are not intended to limit the invention. It is obvious that the drawings described below are merely some embodiments of the invention, and those skilled in the art can obtain other drawings based on these drawings without any inventive effort. Furthermore, the same reference numerals denote the same parts throughout the drawings.

[0021] In the attached diagram:

[0022] Figure 1 Schematic diagram of skin lesions on the back of mice with acute contact dermatitis, showing that tebuconazole (Rot) significantly inhibited skin redness and desquamation, while the positive control drug dexamethasone (Dex) caused skin atrophy;

[0023] Figure 2 Schematic diagram of the degree of ear swelling in mice with acute contact dermatitis, showing that rotazone (Rot) significantly inhibits ear swelling;

[0024] Figure 3 A schematic diagram showing the number of spontaneous scratches in mice with acute contact dermatitis within 30 minutes, demonstrating that rotazone (Rot) significantly reduced itching and the number of scratches;

[0025] Figure 4 Schematic diagram of ELISA results for TSLP protein levels in local skin lesions of mice with acute contact dermatitis, showing that rotazone (Rot) significantly reduces TSLP levels;

[0026] Figure 5 Schematic diagram of ELISA results for TNF-α protein levels in local skin lesions of mice with acute contact dermatitis, showing that rotazone (Rot) significantly reduces TNF-α levels;

[0027] Figure 6 Schematic diagram of H&E staining on the back skin of mice with acute contact dermatitis, showing that rot (tetrahydropalmatine) effectively inhibits epidermal thickening and reduces inflammatory infiltration in a dose-dependent manner.

[0028] Figure 7 Schematic diagram of skin lesions on the back of mice with chronic contact dermatitis, showing that rotazone (Rot) significantly inhibited skin redness and desquamation, and there was no skin atrophy;

[0029] Figure 8 Schematic diagram of the degree of ear swelling in mice with chronic contact dermatitis, showing that rotazone (Rot) significantly inhibits ear swelling;

[0030] Figure 9 A schematic diagram showing the number of spontaneous scratches in mice with chronic contact dermatitis within 30 minutes, demonstrating that rotazone (Rot) significantly reduces itching;

[0031] Figure 10 Schematic diagram of ELISA results for TSLP protein levels in local skin lesions of mice with chronic contact dermatitis, showing that rotazone (Rot) significantly reduces TSLP levels;

[0032] Figure 11 Schematic diagram of ELISA results for TNF-α protein levels in local skin lesions of mice with chronic contact dermatitis, showing that rotazone (Rot) significantly reduces TNF-α levels;

[0033] Figure 12Schematic diagram of H&E staining on the back skin of mice with chronic contact dermatitis, showing that rotazone (Rot) effectively inhibits epidermal thickening and reduces inflammatory infiltration;

[0034] Figure 13 A schematic diagram of weight changes in mice with acute contact dermatitis during drug administration shows that the weight of mice in the dexamethasone (Dex) group decreased significantly, while there was no weight loss in the corydalis extract group and the corydalis B group.

[0035] Figure 14 A schematic diagram of skin lesions on the back of mice with acute contact dermatitis, showing that Corydalis extract can inhibit dandruff and redness, with effects comparable to those of corydalis et al. and dexamethasone.

[0036] Figure 15 Schematic diagram of ear swelling thickness in mice with acute contact dermatitis, showing that both the Corydalis extract group and the Corydalis B group significantly reduced ear swelling thickness;

[0037] Figure 16 A schematic diagram of the number of scratches in mice with acute contact dermatitis within 10 minutes shows that both the Corydalis extract group and the Corydalis B group significantly reduced the number of scratches, with the Corydalis B group showing the best effect.

[0038] Figure 17 : Schematic diagram of skin lesions on the back of mice with nonhistamine-dependent allergic contact dermatitis, showing that tebufenozide (Rot) can inhibit dandruff and redness without skin atrophy;

[0039] Figure 18 : Schematic diagram of ear swelling thickness in mice with nonhistamine-dependent allergic contact dermatitis, showing that tebufenozide (Rot) significantly reduces ear swelling thickness;

[0040] Figure 19 A schematic diagram of the number of scratches in mice with nonhistamine-dependent allergic contact dermatitis within 10 minutes, showing that tebuconazole (Rot) significantly reduced the number of scratches;

[0041] Figure 20 : Schematic diagram of weight change in mice with nonhistamine-dependent allergic contact dermatitis during drug administration, showing that there was no significant decrease in weight in the mice in the tebuconazole (Rot) group;

[0042] Figure 21 : Schematic diagram of skin lesions on the back of mice with allergic contact dermatitis, showing that corydaline (Cor) can effectively inhibit dandruff and redness;

[0043] Figure 22 A schematic diagram of the number of scratches in mice with allergic contact dermatitis within 15 minutes, showing that corydaline (Cor) significantly reduced the number of scratches;

[0044] Figure 23 : Schematic diagram of ear swelling thickness in mice with allergic contact dermatitis, showing that corydaline (Cor) significantly reduces ear swelling thickness;

[0045] Figure 24 A schematic diagram of weight changes in mice with allergic contact dermatitis during drug administration, showing that the weight of mice in the corydalis A (Cor) group did not decrease significantly;

[0046] Figure 25 : Schematic diagram of HaCat cell viability assay, showing that tebufenozide (Rot) has almost no effect on cell viability at concentrations below 100 μM;

[0047] Figure 26 : Schematic diagram of the effect of tebufenozide (Rot) on IL25 mRNA expression level induced by TNF-α / IFN-γ in HaCat cells, showing that Rot inhibits IL25 expression in a dose-dependent manner;

[0048] Figure 27 Schematic diagram of the effect of corydaline (Rot) on IL33 mRNA expression level induced by TNF-α / IFN-γ in HaCat cells, showing that Rot inhibits IL33 expression in a dose-dependent manner;

[0049] Figure 28 Schematic diagram showing the effect of tebufenozide (Rot) on TNF-α / IFN-γ induced TSLP mRNA expression levels in HaCat cells, demonstrating that Rot dose-dependent inhibition of TSLP expression;

[0050] Figure 29 Schematic diagram of the effect of corydaline (Rot) on TNF-α / IFN-γ induced IL13 mRNA expression level in HaCat cells, showing that Rot inhibits the expression of Th2 cytokine IL13;

[0051] Figure 30 : Schematic diagram of HaCat cell viability assay, showing that corydaline (Cor) has almost no effect on cell viability at concentrations below 25 μM;

[0052] Figure 31 Schematic diagram of the effect of corydaline (Cor) on IL25 mRNA expression level induced by TNF-α / IFN-γ in HaCat cells, showing that Cor cannot inhibit IL25 expression;

[0053] Figure 32 : Schematic diagram of the effect of corydaline (Cor) on TNF-α / IFN-γ induced TSLP mRNA expression level in HaCat cells, showing that Cor inhibits TSLP expression in a dose-dependent manner;

[0054] Figure 33 Schematic diagram of the effect of corydaline (Cor) on IL33 mRNA expression level induced by TNF-α / IFN-γ in HaCat cells, showing that Cor inhibits IL33 expression in a dose-dependent manner;

[0055] Figure 34 Schematic diagram of cell thermal displacement analysis (CETSA), showing that tebufenozide (Rot) can specifically stabilize EDEM3 protein, while having no effect on GAPDH and β-actin;

[0056] Figure 35 : Schematic diagram of EDEM3 cell membrane chromatography (CMC), showing that the retention time of tebufenozide (Rot) on the EDEM3-CMC column is 6.136 min, indicating that Rot binds to EDEM3;

[0057] Figure 36 Schematic diagram of the breakthrough curve of tebuconazole (Rot) on an EDEM3-CMC column;

[0058] Figure 37 A schematic diagram illustrating the dissociation constant of the binding of tebuconazole (Rot) with EDEM3 based on the breakthrough curve, showing K... D =3.69±0.21μM;

[0059] Figure 38 : A schematic diagram of molecular docking, showing intermolecular forces between tebufenozide (Rot) and the three amino acids GLU-58, GLN-55, and ARG-660 on EDEM3;

[0060] Figure 39 : Schematic diagram of the effect of thymol (Rot) on TNF-α / IFN-γ induced TSLP mRNA expression in HaCat cells after gene silencing EDEM3 (si-EDEM3), showing that Rot can no longer inhibit TSLP expression after si-EDEM3;

[0061] Figure 40 Schematic diagram of the effect of tebufenozide (Rot) on TNF-α / IFN-γ induced IL25 mRNA expression in HaCat cells after gene silencing EDEM3 (si-EDEM3), showing that Rot can no longer inhibit IL25 expression after si-EDEM3;

[0062] Figure 41Schematic diagram of the effect of tebufenozide (Rot) on TNF-α / IFN-γ induced IL33 mRNA expression in HaCat cells after gene silencing EDEM3 (si-EDEM3), showing that Rot can no longer inhibit IL33 expression after si-EDEM3.

[0063] Figure 42 Schematic diagram of the effect of corydaline (Rot) on TNF-α / IFN-γ induced EDEM3 protein levels in HaCat cells, showing that Rot can effectively restore the reduction in EDEM3 protein levels;

[0064] Figure 43 Schematic diagram of the effect of corydaline (Rot) on the expression of Bip, an endoplasmic reticulum stress marker induced by TNF-α / IFN-γ in HaCat cells, showing that Rot effectively inhibits the increase in Bip expression levels;

[0065] Figure 44 : Schematic diagram of EDEM3 cell membrane chromatography (CMC), showing that corydaline (Cor) has retention behavior in EDEM3-CMC, proving that Cor can also bind to EDEM3.

[0066] The present invention will be further explained below with reference to the accompanying drawings and embodiments. Detailed Implementation

[0067] The following will refer to the appendix. Figures 1 to 44 Specific embodiments of the invention will be described in more detail below. While specific embodiments of the invention are shown in the accompanying drawings, it should be understood that the invention may be implemented in various forms and should not be limited to the embodiments set forth herein. Rather, these embodiments are provided to enable a more thorough understanding of the invention and to fully convey the scope of the invention to those skilled in the art.

[0068] It should be noted that certain terms are used in the specification and claims to refer to specific components. Those skilled in the art will understand that different terms may be used to refer to the same component. This specification and claims do not distinguish components based on differences in terminology, but rather on differences in function. The terms "comprising" or "including" used throughout the specification and claims are open-ended and should be interpreted as "comprising but not limited to." The following descriptions are preferred embodiments for carrying out the invention; however, these descriptions are for the purpose of understanding the general principles of the specification and are not intended to limit the scope of the invention. The scope of protection of this invention is determined by the appended claims.

[0069] Modern pharmacological studies have found that the main pharmacological component of Corydalis rhizome is tetrahydropalmatine, also known as tetrahydropalmatine or tetrahydropalmatine. Tetrahydropalmatine has a weaker analgesic effect than pethidine and morphine, but is less addictive and stronger than common antipyretic analgesics. It can induce hypotension and bradycardia in rats by antagonizing striatal dopamine D2 receptors. It has significant efficacy in relieving pain caused by the gastrointestinal and hepatobiliary systems, and can also alleviate labor pains, headaches, and concussion headaches. It is also used to eliminate mental tension such as irritability and anxiety, and to treat neurasthenia-related insomnia. However, this invention, through animal model experiments, reveals the efficacy and safety of tetrahydropalmatine in treating acute and chronic contact dermatitis, and demonstrates its application in the preparation of drugs for treating acute and chronic contact dermatitis.

[0070] To facilitate understanding of the embodiments of the present invention, further explanations and descriptions will be provided below with reference to the accompanying drawings and specific embodiments. The accompanying drawings do not constitute a limitation on the embodiments of the present invention.

[0071] Experimental materials:

[0072] Corydalis B, Corydalis A, Corydalis extract, newborn C57 mice (purchased from the Experimental Animal Center of Xi'an Jiaotong University), 2,4-dinitrochlorobenzene (DNCB);

[0073] Experimental methods:

[0074] Mice in the NC group received no treatment and were fed normally. Mice in other groups were treated with high or low concentrations of 2,4-dinitrochlorobenzene (DNCB) applied once daily to the back of the mice for 13 consecutive days to establish acute or chronic contact dermatitis models. Mice in the Vehicle group received no other medications besides the 13-day treatment. Mice in the positive control group received daily application of glucocorticoids to the back starting from day 8, while mice in each corydalis group received daily application of different concentrations of corydalis solution to the back starting from day 8. On day 13, data from each group were collected, and the mice were sacrificed, with corresponding tissues collected for observation.

[0075] Experimental results:

[0076] In mouse models of acute contact dermatitis induced by high concentrations of 2,4-dinitrochlorobenzene (DNCB) and chronic contact dermatitis induced by low concentrations of 2,4-dinitrochlorobenzene (DNCB), rotundine (Rot) demonstrated efficacy and safety for both acute and chronic contact dermatitis. Specifically:

[0077] 1. This study investigated an acute contact dermatitis mouse model induced by high concentrations of 2,4-dinitrochlorobenzene (DNCB). This invention is based on the NC group (blank control group), DNCB+Dex (positive drug dexamethasone group), and DNCB+Rot (corydaline). The experiment was conducted in groups, with the corydalis group receiving different doses (e.g., 0.3 mg / mL, 0.6 mg / mL, and 0.9 mg / mL). (The dosage form used in this example is an aqueous solution or detergent; other dosage forms can be calculated similarly.)

[0078] See Figure 1 Based on the skin lesion photos, the DNCB+Rot group of corydalis et al. significantly inhibited the skin redness, swelling, and desquamation of acute contact dermatitis. However, the positive control drug dexamethasone (Dex) caused adverse reactions such as skin atrophy, while the corydalis et al. Rot group did not have this side effect. No signs of atrophy such as epidermal thinning and dermal collagen reduction were observed in the corydalis et al. group. The mechanism of action does not depend on the activation of the glucocorticoid receptor and does not interfere with the normal proliferation and differentiation of keratinocytes, thus having better local safety.

[0079] See further Figure 2 For the ear inflammatory area of ​​acute contact dermatitis, tebufenozide (Rot) significantly reduced the degree of ear swelling;

[0080] See further Figure 3 For acute contact dermatitis, based on the number of spontaneous scratches within 30 minutes, i.e. the degree of itching, tebuconazole (Rot) significantly reduced itching and decreased the number of scratches.

[0081] See further Figure 4 For acute contact dermatitis, ELISA testing showed that tebufenozide (Rot) significantly reduced the TSLP protein level in the affected skin.

[0082] See further Figure 5 For acute contact dermatitis, ELISA testing showed a significant reduction in the level of TNF-α protein in the affected skin.

[0083] See further Figure 6 H&E staining showed that rot (tetrahydropalmatine) could effectively inhibit epidermal thickening, effectively reduce inflammatory infiltration, and was dose-dependent.

[0084] In summary, for the treatment of acute contact dermatitis, similar to the dual-targeting approach of biological agents, tebufenozide (Rot) can be used to dually inhibit TSLP protein levels and TNF-α protein levels, and can inhibit the excessive proliferation of keratinocytes, significantly reduce itching, and has no side effect of skin atrophy.

[0085] 2. Further research will be conducted using a mouse model of chronic contact dermatitis induced by low concentrations of 2,4-dinitrochlorobenzene (DNCB). This invention is further based on the NC group (blank control group), cDNCB+Dex (positive drug dexamethasone group), and cDNCB+Rot (dexamethasone group). The experiment was conducted using different doses of the corydaline group (e.g., 0.3 mg / mL, 0.6 mg / mL, and...). (0.9 mg / mL group; the dosage form used in this example is an aqueous solution or lotion; other dosage forms can be calculated accordingly.)

[0086] See Figure 7Based on the skin lesion photos, tebuconazole (Rot) significantly inhibited skin redness, swelling, and desquamation in chronic contact dermatitis; however, dexamethasone (Dex), a positive control drug, caused skin atrophy as an adverse reaction, while tebuconazole (Rot) did not have this side effect; no signs of atrophy such as epidermal thinning or dermal collagen reduction were observed in the tebuconazole group, and its mechanism of action does not depend on the activation of glucocorticoid receptors, does not interfere with the normal proliferation and differentiation of keratinocytes, and has better local safety;

[0087] See further Figure 8 For the ear inflammatory area of ​​chronic contact dermatitis, tebufenozide (Rot) significantly reduced the degree of ear swelling;

[0088] See further Figure 9 For chronic contact dermatitis, based on the number of spontaneous scratches within 30 minutes, i.e. the degree of itching, tebuconazole (Rot) significantly reduced itching and decreased the number of scratches.

[0089] See further Figure 10 For chronic contact dermatitis, ELISA showed that tebufenozide (Rot) significantly reduced the TSLP protein level in the affected skin.

[0090] See further Figure 11 For chronic contact dermatitis, ELISA testing showed a significant reduction in the level of TNF-α protein in the affected skin.

[0091] See further Figure 12 H&E staining showed that rot (tetrahydropalmatine) could effectively inhibit epidermal thickening, effectively reduce inflammatory infiltration, and was dose-dependent.

[0092] In summary, for the treatment of chronic contact dermatitis, similar to the dual-targeting approach of biological agents, tebufenozide (Rot) can be used to dually inhibit TSLP protein levels and TNF-α protein levels, and can inhibit the excessive proliferation of keratinocytes, significantly reduce itching, and has no side effect of skin atrophy.

[0093] 3. As can be seen from points 1 and 2 above, corydalis et al. demonstrates effectiveness in treating both acute and chronic contact dermatitis. And with a certain level of safety: no skin atrophy side effect. Considering that acute contact dermatitis flares up faster and is more sensitive to medication, more... The safety of corydaline can be investigated, specifically:

[0094] The study continued using a mouse model of acute contact dermatitis induced by high concentrations of 2,4-dinitrochlorobenzene (DNCB). This invention further compared experimental results using the NC group (blank control group), Vehicle group (negative control group), Dex (positive drug dexamethasone group), corydalis yanhusuo group, and corydalis extract group.

[0095] See Figure 13Mice in the dexamethasone positive control group (Dex group) showed a significant decrease in body weight, while the groups corresponding to Corydalis extract and tebuconazole showed no such adverse reaction and no significant decrease in body weight. This further strengthens the safety evidence for tebuconazole (Rot) and reveals that it has no systemic metabolic toxicity.

[0096] 4. As stated in point 3 above, Corydalis extract can also be used to treat acute contact dermatitis, and further experimental studies are needed. Verification:

[0097] See Figure 14 Corydalis extract can inhibit the symptoms of dandruff and redness in mice, with effects comparable to corydalis et al. and the positive control drug dexamethasone (Dex).

[0098] See further Figure 15 The thickness of ear swelling in mice in each treatment group (i.e., the positive drug dexamethasone group, the corydalis group, and the corydalis extract group) was significantly reduced, and the thickness in the corydalis group was slightly lower than that in the corydalis extract group.

[0099] See further Figure 16 The number of scratches within 10 minutes in each treatment group, i.e. the degree of itching, was significantly reduced in both the Corydalis extract group and the Corydalis B group. Among them, the Corydalis B group had the fewest scratches and the best effect.

[0100] 5. Furthermore, taking corydaline (1 mg / ml) as an example, this invention utilizes an acute DNCB model in H1R knockout mice to study... The application of corydaline in the preparation of drugs for treating nonhistamine-dependent allergic contact dermatitis:

[0101] See Figure 17 Rot (tetrahydropalmatine) can inhibit the symptoms of scaling and redness in nonhistamine-dependent allergic contact dermatitis without causing skin atrophy.

[0102] See further Figure 18 The thickness of ear swelling in mice in the Corydalis B group (Rot) was significantly reduced;

[0103] See further Figure 19 Regarding the number of scratches within 10 minutes, i.e. the degree of itching, the corydalis group significantly reduced the number of scratches;

[0104] See further Figure 20 During the administration period, there was no significant decrease in body weight in the mice in the corydalis group;

[0105] In summary, the effect of corydalis et al. in treating nonhistamine-dependent allergic contact dermatitis is independent of the antihistamine pathway and completely avoids the local atrophy and systemic metabolic toxicity of glucocorticoids. Therefore, corydalis et al. is not only effective in treating nonhistamine-dependent allergic contact dermatitis, but also safe.

[0106] 6. Furthermore, this invention is based on the NC group (blank control group), DNCB+Dex (positive drug dexamethasone group), and... The experiment was conducted using DNCB+Cor (corydaline group), with corydaline at a concentration of 1.5 mg / mL as an example:

[0107] See Figure 21 Corydalis A can effectively inhibit the symptoms of scaling and redness in allergic contact dermatitis;

[0108] See further Figure 22 Regarding the number of scratches within 15 minutes, i.e. the degree of itching, the number of scratches was significantly reduced in the corydalis A group;

[0109] See further Figure 23 The thickness of ear swelling in mice in the corydalis A group was significantly reduced;

[0110] See further Figure 24 During the administration period, there was no significant decrease in body weight in the mice in the corydalis A group;

[0111] In summary, the effect of corydalis A on the treatment of allergic contact dermatitis does not depend on the antihistamine pathway and completely avoids the local atrophy and systemic metabolic toxicity of glucocorticoids. Therefore, corydalis A is not only effective in treating allergic contact dermatitis, but also safe.

[0112] 7. Furthermore, the present invention also conducted in vitro pharmacological effects studies on tebufenozide (Rot) and tebufenozide (Cor). HaCat cell experiments:

[0113] 7.1 Regarding Corydalis Ethyl Corydalis

[0114] See Figure 25 Regarding the effect on HaCat cell viability: tebufenozide (Rot) at concentrations below 100 μM has almost no effect on cell viability;

[0115] See further Figure 26 Rot, a dose-dependent inhibitor of TNF-α / IFN-γ-induced mRNA expression levels of the epithelial alarm IL25 in HaCat cells;

[0116] See further Figure 27 Rot, a dose-dependent inhibitor of TNF-α / IFN-γ-induced mRNA expression levels of IL33, an epithelial alarm in HaCat cells, was observed.

[0117] See further Figure 28 Rot, a dose-dependent inhibitor of TNF-α / IFN-γ-induced mRNA expression levels of the epithelial alarm TSLP in HaCat cells;

[0118] See further Figure 29 Furthermore, tebufenozide (Rot) can also inhibit the mRNA expression level of Th2 cytokine IL13 in HaCat cells (AD cell model) induced by TNF-α / IFN-γ.

[0119] 7.2 Regarding Corydalis A

[0120] See Figure 30 Effect of corydaline (Cor) on HaCat cell viability: at concentrations below 25 μM, it has almost no effect on cell viability;

[0121] See further Figure 31 , Figure 32 Although corydaline (Cor) cannot inhibit the mRNA expression level of TNF-α / IFN-γ-induced HaCat cell epithelial alarm IL25 like corydaline (Rot), corydaline (Cor) dose-dependently inhibits the mRNA expression level of TNF-α / IFN-γ-induced HaCat cell epithelial alarm TSLP.

[0122] See further Figure 33 Corydaline (Cor) dose-dependently inhibited the mRNA expression level of TNF-α / IFN-γ-induced epithelial alarmin IL33 in HaCat cells.

[0123] 8. Furthermore, the present invention has also confirmed the target of corydaline B (Rot) and corydaline A (Cor):

[0124] EDEM3 (ER-enhanced α-mannosidase-like protein 3) is a key quality control checkpoint in the endoplasmic reticulum (ER). Primarily acting as an α1,2-mannosidase, it marks misfolded proteins by pruning specific mannoses on N-glycans, allowing them to enter the ER-associated protein degradation (ERAD) pathway for clearance. Decreased EDEM3 function / level leads to increased ER stress.

[0125] 8.1 Regarding Corydalis Ethyl

[0126] See Figure 34 As the temperature increases from left to right, tebufenozide (Rot) effectively stabilizes the EDEM3 protein, but has no effect on the thermal stability of GAPDH and β-actin proteins, proving that tebufenozide can specifically stabilize the EDEM3 protein.

[0127] See further Figure 35 The retention time of tebuconazole (Rot) on the EDEM3-CMC (cell membrane chromatography) column was 6.136 min, indicating that tebuconazole (Rot) has retention behavior on EDEM3-CMC. This result shows that tebuconazole can bind to EDEM3.

[0128] See further Figure 36 This is the breakthrough curve of tebuconazole (Rot) on an EDEM3-CMC (cell membrane chromatography) column. See further details. Figure 37The binding affinity of tebuconazole to its target protein EDEM3 was calculated based on the breakthrough curve of tebuconazole on an EDEM3-CMC column, and its dissociation constant K was determined. D =3.69±0.21μM;

[0129] See further Figure 38 The results of molecular docking in AutoDockTools-1.5.7 showed that there are intermolecular forces between tebuconazole (Rot) and the three amino acids GLU-58, GLN-55 and ARG-660 on EDEM3, which enables tebuconazole to bind to EDEM3.

[0130] See further Figure 39 After gene silencing EDEM3 (si-EDEM3), tebufenozide (Rot) could no longer inhibit the mRNA expression level of TNF-α / IFN-γ-induced HaCat cell epithelial alarmin TSLP, indicating that tebufenozide exerts its effect through EDEM3.

[0131] See further Figure 40 After gene silencing EDEM3 (si-EDEM3), tebufenozide (Rot) could no longer inhibit the mRNA expression level of TNF-α / IFN-γ-induced HaCat cell epithelial alarmin IL25, indicating that tebufenozide exerts its effect through EDEM3.

[0132] See further Figure 41 After gene silencing EDEM3 (si-EDEM3), tebufenozide (Rot) could no longer inhibit the mRNA expression level of TNF-α / IFN-γ-induced HaCat cell epithelial alarmin IL33, indicating that tebufenozide exerts its effect through EDEM3.

[0133] Decreased EDEM3 function / level can exacerbate endoplasmic reticulum (ER) stress. Tetracycline restores upstream EDEM3 protein expression levels, promoting the normal functioning of the unfolded protein response / ER degradation pathway, preventing the accumulation of misfolded / unfolded proteins, and thus inhibiting ER stress; see also... Figure 42 It was found that tebufenozide (Rot) effectively restored the TNF-α / IFN-γ-induced decrease in EDEM3 protein levels in HaCat cells (Veh); see further. Figure 43 Furthermore, corydaline can effectively inhibit the increase in the expression level of the endoplasmic reticulum stress marker Bip in HaCat cells (TI) induced by TNF-α / IFN-γ.

[0134] 8.2 Regarding Corydalis A

[0135] See Figure 44The discovery that corydaline (Cor) exhibits retention behavior in EDEM3-CMC definitively demonstrates that corydaline can bind to EDEM3, which is also a target protein of corydaline.

[0136] In one embodiment, the present invention provides the use of corydalis ethylsin in the preparation of a drug for treating acute and chronic contact dermatitis.

[0137] In another embodiment, the present invention also provides a medicament for treating acute and chronic contact dermatitis, the medicament comprising corydaline.

[0138] Preferably, the dosage form of the drug is a topical preparation, which includes gels, ointments, creams, powders, liquids, or lotions.

[0139] Preferably, the concentration of corydaline in the drug is 0.3-0.9 mg / mL.

[0140] It is important to emphasize that tebufenozide (Rot) can be used to dual inhibit TSLP protein levels and TNF-α protein levels, and can inhibit the excessive proliferation of keratinocytes, significantly reduce itching, and has no side effects of skin atrophy or systemic metabolic toxicity.

[0141] Dose gradient experiments have confirmed that corydalis has therapeutic effects in the range of 0.01-1 mg / mL, especially in the range of 0.03-0.9 mg / mL.

[0142] In another embodiment, the present invention also discloses the use of either corydaline A or corydaline B in the preparation of a drug for treating nonhistamine-dependent allergic contact dermatitis. It should be noted that its effect on nonhistamine-dependent allergic contact dermatitis does not depend on the antihistamine pathway and completely avoids the local atrophy and systemic metabolic toxicity of glucocorticoids. Therefore, it is not only effective in treating nonhistamine-dependent allergic contact dermatitis but also safe.

[0143] In another embodiment, the present invention also discloses the use of Corydalis extract in the preparation of a drug for treating acute contact dermatitis.

[0144] This invention provides clear concentration guidance for clinical formulation development, avoiding ineffectiveness due to excessively low concentrations or irritation due to excessively high concentrations. It is effective at low concentrations, and no toxic effects such as skin wrinkling or weight loss were observed in experiments, making it superior to positive control drugs such as dexamethasone.

[0145] It should be noted that the aforementioned Corydalis extract, Corydalis A, and Corydalis B are all administered via topical application.

[0146] Preferably, tebufenozide (Rot) can be used to dual inhibit TSLP protein levels and TNF-α protein levels, and can inhibit the excessive proliferation of keratinocytes, significantly reduce itching, and has no skin atrophy side effect and no systemic metabolic toxicity.

[0147] Furthermore, this invention also discloses the application of corydaline in the preparation of drugs for treating nonhistamine-dependent allergic contact dermatitis.

[0148] Furthermore, this invention also discloses the application of corydaline in the preparation of drugs for treating allergic contact dermatitis.

[0149] Preferably, the concentration of corydaline A is 1.5 mg / mL.

[0150] The corydaline A compound directly binds to the EDEM3 protein, restores EDEM3 expression levels, and inhibits endoplasmic reticulum stress, thereby alleviating the symptoms of allergic contact dermatitis.

[0151] Furthermore, this invention also discloses the application of Corydalis extract in the preparation of drugs for treating acute contact dermatitis.

[0152] Furthermore, this invention also discloses a therapeutic agent for contact dermatitis (including acute and chronic contact dermatitis and nonhistamine-dependent allergic contact dermatitis) that targets the EDEM3 protein, the agent containing an effective amount of corydaline, and the corydaline exerts its therapeutic effect by directly binding to the EDEM3 protein.

[0153] Preferably, according to the application of the corydaline in the preparation of drugs for treating acute and chronic contact dermatitis or the application of the corydaline in the preparation of drugs for treating nonhistamine-dependent allergic contact dermatitis, the corydaline reduces the symptoms of contact dermatitis by directly binding to the EDEM3 protein, restoring the expression level of EDEM3, and inhibiting endoplasmic reticulum stress.

[0154] Preferably, the dissociation constant K between the corydaline and the EDEM3 protein is... D The value was 3.69 ± 0.21 μM.

[0155] Preferably, the fumaric acid binds to the GLU-58, GLN-55, and ARG-660 amino acid residues on the EDEM3 protein.

[0156] Preferably, the corydaline inhibits the expression of epithelial alarmins TSLP, IL25 and / or IL33 in an EDEM3-dependent manner.

[0157] Preferably, the nonhistamine-dependent allergic contact dermatitis is induced by allergens that do not depend on the histamine pathway, and the drug does not rely on an antihistamine mechanism to take effect.

[0158] In summary, this invention provides the application of corydaline in the preparation of drugs for treating acute and chronic contact dermatitis. Experiments show that corydaline significantly inhibits redness, desquamation, and ear swelling in acute and chronic contact dermatitis, reduces itching, and inhibits epidermal thickening and inflammatory infiltration. Its mechanism of action includes dual inhibition of TSLP and TNF-α protein levels, and restoration of EDEM3 expression through direct binding to EDEM3 protein (KD=3.69±0.21μM), inhibiting endoplasmic reticulum stress, thereby downregulating epithelial alarm intensities such as IL25, IL33, and TSLP. The effect of corydaline disappears after gene silencing EDEM3, confirming EDEM3 as its functional target protein. Furthermore, corydaline is also effective against nonhistamine-dependent allergic contact dermatitis, without side effects such as skin atrophy and weight loss, and its safety profile is superior to dexamethasone. This invention also relates to similar applications of corydaline A and corydalis extracts. This invention provides a safe, effective, and mechanistically defined novel therapeutic agent for contact dermatitis.

[0159] Although embodiments of the present invention have been described above in conjunction with the accompanying drawings, the present invention is not limited to the specific embodiments and application fields described above. The specific embodiments described above are merely illustrative and instructive, and not restrictive. Those skilled in the art can make many other forms based on the guidance of this specification and without departing from the scope of protection of the claims of the present invention, and all of these are within the scope of protection of the present invention.

Claims

1. The use of corydalis ethylsin in the preparation of drugs for treating acute and chronic contact dermatitis.

2. The application according to claim 1, characterized in that, Preferably, the dosage form of the drug is a topical preparation, which includes gels, ointments, creams, powders, liquids, or lotions.

3. A medicine for treating acute and chronic contact dermatitis, said medicine comprising corydaline.

4. Application of corydaline in the preparation of drugs for treating nonhistamine-dependent allergic contact dermatitis.

5. Application of corydaline in the preparation of drugs for treating allergic contact dermatitis.

6. Application of Corydalis extract in the preparation of drugs for treating acute contact dermatitis.

7. A contact dermatitis treatment drug targeting the EDEM3 protein, characterized in that, The drug contains an effective amount of corydaline, and the corydaline exerts its therapeutic effect by directly binding to the EDEM3 protein.

8. The application according to claim 1 or 4, characterized in that, The corydaline directly binds to the EDEM3 protein, restores EDEM3 expression levels, and inhibits endoplasmic reticulum stress, thereby alleviating the symptoms of contact dermatitis.

9. The application according to claim 8, characterized in that, The dissociation constant K between corydaline and EDEM3 protein D The value was 3.69 ± 0.21 μM.

10. The application according to claim 8, characterized in that, The corydaline binds to the GLU-58, GLN-55, and ARG-660 amino acid residues on the EDEM3 protein.

Images