Use of isoquinoline derivatives for diabetic wound healing

Abstract

The present invention is related to a method for diabetic wound healing comprising administering to a subject in need thereof a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound having the general Formula I, preferably salsolinol.

Claims

1. A method for diabetic wound healing in a subject comprising applying to the diabetic wound in said subject a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound having the general Formula I: ##STR00006## wherein R, R.sub.1 and R.sub.2 are each independently H, alkyl or acyl (R.sub.aCO) group; R.sub.3 is H, alkyl or substituted benzyl group; wherein R.sub.a is H or alkyl group.

2. The method of claim 1, wherein said substituted benzyl group has the following Formula II: ##STR00007## wherein X and Y are each independently H, OH, methoxy (OMe) or acyloxy (R.sub.bCOO) group; wherein R.sub.b is H or alkyl group.

3. The method of claim 1, wherein the compound is salsolinol.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) The drawings presenting the preferred embodiments of the present invention are aimed at explaining the present invention. It should be understood that the present invention is not limited to the preferred embodiments shown. The data in the figures and examples are shown as meanstandard deviation (SD), determined by the paired t-test. Significant differences are shown as follows: *: P<0.05; **: P<0.01.

(2) FIG. 1 shows the recovery rates of the wounds of normal rats without treatment (NC) and STZ induced diabetic rats without treatment (DC) for 15 days.

(3) FIG. 2 shows the recovery rates of the wounds of STZ induced diabetic rats treated with salsolinol at different concentrations (0.01 mg/g, 0.03 mg/g, 0.1 mg/g, and 0.3 mg/g) for 15 days.

(4) FIG. 3 shows the recovery rates of the wounds of STZ induced diabetic rats without treatment (DC), STZ induced diabetic rats treated with vehicle (DV), Swivazin (DP, positive control), and salsolinol (D+Salsolinol, compound of the invention) for 14 days.

DETAILED DESCRIPTION OF THE INVENTION

(5) Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of skill in the art to which this invention belongs.

(6) Unless clearly specified herein, meanings of the articles a, an, and said all include the plural form of more than one. Therefore, for example, when the term a component is used, it includes multiple said components and equivalents known to those of common knowledge in said field.

(7) As used herein, the term substituted or substitution refers to where a functional group in a chemical compound is replaced by another group.

(8) As used herein, the term subject refers to a human or a mammal, such as a patient, a companion animal (e.g., dog, cat, and the like), a farm animal (e.g., cow, sheep, pig, horse, and the like) or a laboratory animal (e.g., rat, mouse, rabbit, and the like).

(9) The term alkyl group used herein refers to linear or branched monovalent hydrocarbons containing 1-20 carbon atoms, such as alkyl groups with 1-10 carbons, preferably alkyl groups with 1-6 carbons, more preferably alkyl groups with 1-3 carbons. Examples of alkyl groups include, but not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, and t-butyl.

(10) Accordingly, the invention, in one aspect, provides a method for diabetic wound healing. The method comprises administering to a subject in need thereof a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound having the general Formula I:

(11) ##STR00003##
wherein R, R.sub.1 and R.sub.2 are each independently H, alkyl or acyl (R.sub.aCO) group; R.sub.3 is H, alkyl or substituted benzyl group; wherein R.sub.a is H or alkyl group.

(12) In a particular example of the present invention, said substituted benzyl group has the following Formula II:

(13) ##STR00004##
wherein X and Y are each independently H, OH, methoxy (OMe) or acyloxy (R.sub.bCOO) group; wherein R.sub.b is H or alkyl group.

(14) An embodiment of the active compound of the present invention is the compound having the general Formula I, wherein RR.sub.1R.sub.2H and R.sub.3=Me (methyl), which compound is salsolinol having the following formula:

(15) ##STR00005##

(16) As shown in the examples of the present invention, the compound having Formula I of the present invention, such as salsolinol, has an effect of treating diabetic wounds.

(17) According to the present invention, said compound having Formula I can be formulated into any forms of medications that are well known or commonly used in the pharmaceutical field, and can be prepared into a composition, according to any techniques well known in the pharmaceutical field, comprising a therapeutically effective amount of said compound in combination with a commonly used carrier or a pharmaceutically acceptable carrier.

(18) The term carrier or pharmaceutically acceptable carrier used herein includes, but not limited to, pharmaceutically acceptable excipients, fillers, diluents, or the like, including those well known to one of ordinary skills in the pharmaceutical field.

(19) The present invention is explained in the above description of the invention and the following examples, which should not be used to restrict the scope of the present invention.

Example

(20) 1. Preparation of the Compound Having Formula I of the Present Invention

(21) Dopamine (1.6 g), 10 mL methanol, 1 mL 1N hydrochloric acid, and 2 mL 99% acetaldehyde were added sequentially into a 50 mL round-bottom flask and stirred for 6 hours under room temperature. The concentrate obtained by reduced-pressure concentration was loaded into a Lobar RP-18 column (size B, Merck), eluted by a 0.05% formic acid aqueous solution, to give .sup.1HNMR essential pure salsolinol (1.0 g).

(22) Using ESI-TOF mass spectrometry and NMR spectroscopy analysis, the characterization data of salsolinol are as follows:

(23) .sup.1H NMR (CD.sub.3OD, 400 MHz) 6.63 (1H, s), 6.57 (1H, s), 4.30 (1H, q, J=6.8 Hz, H-1), 3.40 (1H, dt, J=12.6, 5.6 Hz, H.sub.a-3), 3.20 (1H, ddd, J=12.6, 8.2, 5.6 Hz, H.sub.b-3), 2.92 (1H, ddd, J=16.8, 8.2, 5.8 Hz, H.sub.a-4), 2.80 (1H, dt, J=16.8, 5.6 Hz, H.sub.b-4), 1.55 (3H, d, J=6.8 Hz, Me-1); ESIMS: m/z 180 ([M+H].sup.+).

(24) 2. Evaluation of the Effect of Salsolinol in Treatment of Diabetic Wounds

(25) Salsolinol ointments at 0.01 mg/g, 0.03 mg/g, 0.1 mg/g, and 0.3 mg/g were prepared respectively by dissolving 0.5 mg, 1.5 mg, 5 mg, and 15 mg of salsoinol in 2.5 mL of glycerol (Sigma Inc., MO, USA) plus 1.0 mL of Creagel emulsifier (First Chemical, TPE, Taiwan) and 45.5 mL of distilled water. Vehicle was composed of glycerol (2.5 mL), Creagel emulsifier (1.0 mL), and distilled water (45.5 mL).

(26) Swivazin-HN cream was used as positive control for treating the wounds of STZ induced diabetic rats, which contained asiaticoside (4 mg/g), isolated from Centella asiatica. It was reported by Shukla et al., that asiaticoside had the effects on ameliorating poor diabetic wound healing. [Shukla et al. In vitro and in vivo wound healing activity of asiaticoside isolated from Centella asiatica. Ethnopharmacology 65 (1999): 1-11].

(27) Adult male Wistar rats were used at four weeks of age, purchased from BioLASCO Taiwan, and were acclimatized in a room with 12-12 h light (7:00 A.M. to 7:00 P.M.)-dark (7:00 P.M. to 7:00 A.M.) cycle, a temperature of 231 C., and a humidity of 655%.

(28) Each rat had free access to both water and standard rodent soft chow ad libitum. STZ was injected intraperitoneally at a dose of 65 mg/Kg body weight. A week later, the rats that had a blood glucose level higher than 250 mg/dL were considered as STZ induced diabetic rats and used for experiments.

(29) All experimental procedures were approved by and performed in compliance with the guidelines of the Institutional Animal Care and Use Committee (IACUC). All surgeries were conducted while animals were under continuous anesthesia with 4% isoflurane.

(30) The dorsal skin was shaved and then disinfected with 10% povidone-iodine before an excisional full-thickness square-shaped skin wound (1.51.5 cm) was induced using a sterile wound maker. Animal was then housed alone in its cage to avoid any further wound damage.

(31) Rats were randomly divided into eight groups (n=6, each group) as follows: normal rats (NC) for comparison, STZ induced diabetic rats without treatment (DC), STZ induced diabetic rats treated with the vehicle only (DV), STZ induced diabetic rats treated with Swivazin-HN cream as positive control (DP, wherein the Swivazin-HN cream contains 4 mg/g asiaticoside from Centella asiatica), and STZ induced diabetic rats treated with salsolinol (D+Salsolinol) at different concentrations of 0.01 mg/g, 0.03 mg/g, 0.1 mg/g, and 0.3 mg/g respectively. The groups were administered at the same amount once daily and the wounds were photopictured on Day 1, Day 3, Day 5, Day 7, Day 10, and Day 15 and the sizes of the wounds were analyzed using Visual Basic 6.0. Wound closure at experimental time points was calculated as percentage of initial wound area, called as recovery rate.

(32) In this example, the rats were injected with STZ intraperitoneally to obtain STZ induced diabetic rats, which had increased blood glucose levels (>250 mg/dL). The wounds of normal rats without treatment (NC) and diabetic rats without treatment (DC) were compared. As shown in FIG. 1, the wounds of NC recovered quickly in the beginning and the recovery rate over 98% at Day 15, while the wound healing of DC showed slower in terms of lower recovery rates as compared with normal rats (NC), and the wounds of DC were found to be even worse at Day 1-3. It was indicated that the wound healing of the STZ induced diabetes rats was poor and delayed.

(33) The STZ induced diabetic rats were divided into four groups and treated with salsolinol at the concentrations of 0.01 mg/g, 0.03 mg/g, 0.1 mg/g and 0.3 mg/g, respectively. As shown in FIG. 2, for the groups treated with salsolinol at all the concentrations, salsolinol could enhance the wound closure and healing. In particular, the group treated with salsolinol at the concentration of 0.01 mg/g showed relatively improved recovery rate at Day 7-15 (over 97% at Day 15).

(34) The groups of STZ induced diabetic rats treated with salsolinol (D+Salsolinol) were compared with the groups without treatment (DC) or the groups treated with vehicle (DV), and Swivazin (DP, positive control). As shown in FIG. 3, the wounds of the DP and D+Salsolinol groups were recovered quickly in the beginning and having the recovery rates over 97% at Day 15. The group treated with salsolinol (D+Salsolinol) showed significantly increased recovery rate, and there is no obvious difference between the group treated with salsolinol (D+Salsolinol) and Swivazin (DP).

(35) In summary, salsolinol, one example of the present invention, provides significantly improved efficacy in diabetic wound healing, and has potential for developing a medicament for diabetic wound healing.