Abstract:
Administering to a patient of muscular dystrophy a pharmaceutical agent containing glycyrrhizin and/or a pharmaceutically acceptable salt thereof as effective components is effective against muscular dystrophy, particularly, Duchenne or Becker muscular dystrophy and is highly safe with less side effect.

Description:
BACKGROUND OF THE INVENTION 
     This invention relates to a method of treatment for muscular dystrophy and, more particularly, to a method of treatment for muscular dystrophy such as Duchenne muscular dystrophy, Becker muscular dystrophy and the like. 
     The Duchenne muscular dystrophy is a sex-linked recessive disease occuring in childhood, in which muscle weakness and muscular wasting in the proximal parts of extremities and trunk are progressed, resulting in a death at about twenty. The Becker muscular dystrophy is also a sex-linked recessive disease showing the same symptoms, although its onset age is older and tile progression is slower, compared with the Duchenne type. 
     Other muscular dystrophies include limb-girdle muscular dystrophy, facioscapulohumeral (FSH) muscular dystrophy, congenital muscular dystrophy, and the like. 
     Studies on these muscular dystrophies were widely made through many years in developed countries as leaders, with respect to the etiological cause, mechanism of incidence, medical treatment and the like, but, an effective medical treatment so far has not been established yet. Thus the earliest possible development of an effective remedy has long been awaited. Up to the present time, number of medicines were examined for their possibilities as the remedy, but prednisolone, a known 11-betahydroxy corticosteroid, and dantrolene are merely reported to be slightly effective. 
     Glyeyrrhizin has been used as a remedy for liver diseases and an anti-allergic medicine for many years, and its effectiveness is recognized. Although it causes hypokalemia as a side effect, this is relatively easy to control by the combined use with spironolactone. Further, glycyrrhizin is known to enhance the effects of adrenal cortical hormones by the suppression of metabolism of them, and it is also expected to reduce the side effect of prednisolone by the combined use with prednisolone. 
     However, glycyrrhizin has not been examined as a remedy for muscular dystrophy, and there has been no report on its effectiveness yet. 
     With respect to the prednisolone and dantrolene as the remedy for muscular dystrophy, it is the present state that troublesome adverse effects by their long-term use such as the production of a difficult side effect such as Cushing syndrome at a high ratio in case of the prednisolone and the induction of a liver damage in case of the dantrolene can not be avoided. The single use of these medicines can not provide a sufficient effect, and a new drug is desired which is well tolerated to the patients with muscular dystrophies for a long period, and can be used as an adjunctive therofy to prednisolone or dantrolene. 
     SUMMARY OF THE INVENTION 
     The present invention has a purpose to provide an effective and safe medical treatment for muscular dystrophy with less side effect. 
     As a result of the earnest studies to solve the above problems, the present inventors have found that glycyrrhizin known to be highly safe is effective for muscular dystrophy. 
     The present invention provides a method of treatment for muscular dystrophy which comprises the step of administering a pharmaceutical agent containing glycyrrhizin and/or pharmaceutically acceptable salts thereof as effective components to a patient. The method according to the present invention is particularly effective for Duchenne and Becker muscular dystrophies. 
     The present invention is further illustrated in detail. 
     The pharmaceutical agent used in the method of treatment according to the present invention contain glycyrrhizin and/or pharmacologically allowable salts thereof. Glycyrrhizin can be obtained by the extraction from licorice root. Commercially available ones may be also used. The pharmaceutically acceptable salts of glycyrrhizin include glycyrrhizin ammonium salt, glycyrrhizin alkali metal salts, glycyrrhizin choline salt and the like. These salts are obtained by reacting glycyrrhizin with inorganic or organic bases in molar ratio. 
     The drug form of the parmaceutically agent is not particularly limited, but, in general, they may be used in the form of injections and internal medicines such as powders, tablets, granules, capsules, solutions and the like. These can be manufactured by the use of conventionally known techniques. 
     Further, one or several kinds of pharmaceutically acceptable nontoxic fillers, for example, lactose, potato starch, calcium carbonate, and sodium alginate and the like may be added to these drugs. In the injection form, distilled water for injection, polyethylene glycol and the like may be used as a solvent, and a dispersing agent may be added thereto. 
     The method of treatment according to the present invention comprises the step of administering the pharmaceutical agent mentioned above to a patient of muscular dystrophy. As the administering method, either of oral and parenteral methods may be selected. The dose depends on the age and symptom of the patient, and in general, the dose in oral administration preferably ranges in an adult from 25-500 mg a day as the quantity of glycyrrhizin or its salt and, more preferably, a sufficient effect can be expected by the use within the range of 150-225 mg. The dose in paranteral administration is properly 40-400 mg a day in an adult. 
     The medical treatment for muscular dystrophy according to the present invention is effective for muscular dystrophies such as Duchenne type and Becker type muscular dystrophies, and safe with less side effect. It is expected to be effectively usable even in the combination with prednisolone and dantrolene, well-known cortisteroids. 
    
    
     DESCRIPTION OF PREFERRED EMBODIMENTS 
     Described below are preferred embodiments of the present invention. Examples of formulations used in the treatment for muscular dystrophy of the present invention are first illustrated. 
     Formulation Example 1 &lt;Tablet&gt; 
     A mixture of the following compositions is formed into a tablet in the usual way. 
     
         ______________________________________Glycyrrhizin      25 mgPotato starch     270 mgMagnesium stearate              5 mgTotal             300 mg______________________________________ 
    
     Formulation Example 2 &lt;Sugar-coated tablet&gt; 
     A mixture of the following compositions is formed into a sugar-coated tablet in the usual way. 
     
         ______________________________________Glycyrrhizin        25 mgAminoacetic acid    25 mgMethionine          25 mgPrecipitated calcium carbonate               Proper quantityLactose             Proper quantityCarboxymethylcellulose               Proper quantityTotal               300 mg______________________________________ 
    
     Formulation Example 3 &lt;Granule&gt; 
     A mixture of the following compositions is formed into a granule in the usual way. 
     
         ______________________________________Glycyrrhizin   25 mgLactose        210 mgStarch         60 mgGelatin         5 mgTotal          300 mg______________________________________ 
    
     Formulation Example 4 &lt;Injection&gt; 
     To an isotonic sodium chloride solution, 200 mg of glycyrrhizin is dissolved into 100 ml, and formed into an injection in the usual way. 
     Formulation Example 5 &lt;Injection&gt; 
     To an isotonic sodium chloride solution, 200 mg of glycyrrhizin, 2000 mg of aminoacetic acid, and 100 mg of cysteine are dissolved into 100 ml, and formed into an injection in the usual way. 
     Pharmacological Test 
     The effect of the treatment for muscular dystrophy of the present invention is illustrated in reference to a clinical test. 
     1. Subject 
     Subjects were 17 patients of Duchenne muscular dystrophy (age 13-22, average age 17.6) and 4,patients of Becker muscular dystrophy (age 15-64, average age 44.5). The subjects was diagnosed by five doctors approved by the Japanese Society of Neurology on the basis of the present history, clinical symptoms, neurological examinations, neurophysiological tests and muscle biopsy. 
     2. Used agents 
     As a muscular dystrophic remedy containing glycyrrhizin and/or its pharmaceutically acceptable salt, Glycyrrhon as internal tablets and Stronger Neo-Minophagen C as an intravenous injection (the both are made by MINOPHAGEN PHARMACEUTICAL CO., Trademarks) were employed. The prescriptions of Glycyrrhon and Stronger Neo-Minophagen C are as follows. &lt;Glycyrrhon tablet&gt; 
     A white sugar-coated tablet containing the following components in one tablet. 
     
         ______________________________________Glycyrrhizin         25 mgDL-Methionine        25 mgAminoacetic acid     25 mgPrecipitated calcium carbonate                25 mg______________________________________ &lt;Stronger Neo-Minophagen C&gt; 
    
     The following components are contained in one ample (20 ml). 
     
         ______________________________________Glycyrrhizin ammonium            40 mg            (as glycyrrhizin)Aminoacetic acid 400 mgL-cysteine hydrochloride            20 mg______________________________________ 
    
     As additives, 16 mg of sodium sulfite and proper amount of 10% of ammonia water are added. 
     3. Method for the test 
     Administration dose hereinbelow shows quantity of glycyrrhizin. 
     The 17 patients of Duchenne type muscular dystrophy were divided into 12 of medication group and 5 of nomedication group at random. All the patients of the medication group were subjected to continued administration of tablets (150 mg/day), or continued administration of tablet (150 mg/day) after intravenous injection (40 mg/day) for 2 weeks, and the results were observed. 
     On the other hand, the patients with Becker muscular dystrophy were subjected to continued administration of tablets (225 mg/day), or continued administration of tablets (225 mg/day) after intravenous injection (80 mg/day) for 4 weeks, and the results were observed. Informed consents were obtained from all the medicated patients and their families. 
     The evaluation was carried out with respect to muscule strength and function by using the criteria by Brooke et al. (Brooke, M. H. et al., Muscle Nerve 4, 186-197, 1981). The 18 muscle groups shown in Table 1 were evaluated for muscle strength according to the criteria shown in Table 2 before starting the administration and 4, 8 and 12 weeks after administration, and respective mean muscle strength scores were calculated. As an index evaluating extremity functions, a 3×3 inch square clipping test was conducted, and the required time therefor was taken as the index. The reason for using this test for the evaluation of extremity functions is that the abilities for locomotion of the subject patients were low, and this is the only test that all the patients can undergo for evaluating the extremity functions. 
     The mean muscle strength score and the change ratio of the required time in the 3×3 inch square clipping test were compared between the medication group and the no-medication group. The assessment was performed by the five doctors approved by the Japanese Society of Neurology. 
     Further, routine laboratory tests such as peripheral blood picture, liver function test, and urinalysis were similarly carried out. 
     
                       TABLE 1______________________________________Evaluated Muscle GroupsMuscle Groups       Muscle Groups______________________________________shoulder abduction  hip abductionshoulder external rotation               knee flexionelbow flexion       knee extentionelbow extention     ankle inversionwrist flexion       ankle eversionwrist extention     ankle plantar flexionthumb abduction     ankle dorsiflexionhip flexion         neck flexionhip extention       neck extention______________________________________ 
    
     
                       TABLE 2______________________________________Strength Grading for MusclesGrade Degree of Strength        Score______________________________________5     Normal muscular strength  105-    Slightly hyposthenic      94s    Middle between 5- and 4   74     Hyposthenic, but movable against combination                           7 of gravity and resistance4w    Middle between 4 and 3+   73+    Temporarily movable against resistance, but                           5 becomes unmovable soon3     Unmovable to addition of resistance, but                           4 movable against gravity3-    Movable against gravity, but the movable                           3 range is not perfect2     Movable in the state having no gravity                           21     Slightly movable          10     Entirely unmovable        0______________________________________ 
    
     The results are shown in Tables 3 and 4. A significant difference was recognized between the no-medication group and the medication group with a P value less than 5% (P&lt;0.05). For the test, the Wilcoxon&#39;s two-sample rank test (Wilcoxon, F. Individual comparison by ranking methods. Biometrics, 1, 80-83, (1945); Wilcoxon, F. Probability tables for individual companions by ranking methods. Biometrics, 3, 119-122, (1947)) was adapted. 
     
                                           TABLE 3__________________________________________________________________________Change Ratio of Mean Muscle StrengthDuchenne type   Becker type                    Duchenne typemedication group           medication group                    no-medication group(%)             (%)      (%)__________________________________________________________________________Before 0     0   0     0  0         0medication    0         0          04th week 0     38.2*           0     2.2*                    -8.0      -2.0     8.9      0.6        -4.38th week 0     62.2*           0     2.1*                    -12.2     -4.5    17.7      0.9        -8.412th week 0     62.2*           0     6.2*                    -18.7     -5.7    19.7      3.6        -11.2__________________________________________________________________________ Upper numbers show range, and lower numbers show mean. Minus change ratio means deterioration, and plus change ratio means improvement. *: significantly different with P value less than 5%, compared with nomedication group. 
    
     
                                           TABLE 4__________________________________________________________________________Change Ratio of Required Time in 3 × 3&#34; Square Clipping TestDuchenne          Becker         Duchennemedication group  medication group                            no-medication group(%)               (%)            (%)__________________________________________________________________________Before 0      0    0         0    0      0medication   0              0            04th week 7.1    -42.6*             -9.5      -44.4*                            34.6   -12.2   -20.6          -31.4        9.48th week 1.8    -40.0#             -27.3     -47.2*                            12.7   -20.5   -22.2          -38.3        -7.712th week 7.1    -49.3*             -36.1     -57.8*                            23.6   -14.4   -28.0          -46.1        -1.4__________________________________________________________________________ Upper numbers show range, and lower numbers show mean. Minus change ratio means improvement, and plus change ratio means deterioration. *: significantly different with a risk less than 5%, compared with nomedication group. #: not significant different with a risk of 5-10% compared with nomedication group. 
    
     The results shows that the treatment according to the present invention has therapeutic effects to the reduction in muscular strength and/or reduction in extremity function of the Duchenne and Becker muscular dystrophies. 
     Since this effect is supposed to be due to the action of stabilizing cell membranes and action of suppressing calcium separation from sarcoplasmic reticula possessed by the glycyrrhizin which is the effective component of the remedy used in the treatment of the present invention, the similar effect to other muscular dystrophies of the same morbid state can be expected. Hypokalemia emerged in two of twelve patients in Duchenne muscular dystrophy and in two of four patients in Becker muscular dystrophy, but it was improved by the combined use with 25 mg-50 mg/week of spironolactone. No side effect emerged in the others. Acute Toxicity Test for Glycyrrhizin 
     Glycyrrhizin shows the following LD 50 , and is low toxic. 
     
         ______________________________________(1) Oral administration (rat)                       3        g/kg&lt;(2) Subcutaneous injection - 5% aq. soln.                       1873.3   mg/kg    (mouse)(3) Intravenous injection - 2% aq. soln.                       682.5    mg/kg    (mouse)(4) Intraperitoneal injection - 0.2% aq. soln.                       225-244  mg/kg    (mouse)______________________________________