JP6966763B2 - Improvement or treatment of chondromalacia patella - Google Patents

Description

The present invention relates to a novel pharmaceutical use of a vaccinia virus-inoculated inflammatory tissue extract (hereinafter, may be referred to as "the present extract"). More specifically, the present invention relates to an ameliorating or therapeutic agent for chondromalacia patella containing the present extract as an active ingredient.

Patella chondrosis is a common clinical bone joint disease in which the cartilage on the back of the patella softens and pain in the patellofemoral joint is the main symptom. Many patients with patellar cartilage have a clear history of trauma or a buildup of chronic minor injuries, and pathological features include softening, fissures, shedding, and degeneration of the patella cartilage. The characteristic of joint pain due to patellar chondrosis is that it starts with dull pain and discomfort on the anterior side of the knee joint and gradually develops into knee joint pain, especially when exercising such as climbing up and down stairs. ..

Patients with moderate patellar chondrosis are treated with oral non-steroidal anti-inflammatory drugs and physiotherapy. However, long-term physiotherapy interferes with work and is costly, so patients tend to be less compliant. In addition, since most patients refuse surgery, it is common clinically to choose intra-articular injection therapy with hyaluronic acid preparations. Intra-articular injection therapy is widely used in clinical practice because of its reliable effect and few complications. In such a situation, a method for further improving the effect of intra-articular injection therapy is required in clinical practice.

The vaccinia virus-inoculated inflammatory tissue extract, which is an active ingredient for improving or treating patellar cartilage softening according to the present invention, has an analgesic effect, a sedative effect, an antistress effect, an antiallergic effect (see Patent Document 1), and an immunostimulatory effect. Action, anticancer action, liver cirrhosis inhibitory action (see Patent Document 2), therapeutic effect on idiopathic thrombocytopenic purpura (see Patent Document 3), post-herpes zoster nerve pain, cerebral edema, dementia, spinal cerebral degeneration, etc. Therapeutic effect (see Patent Document 4), Reynaud syndrome, diabetic neuropathy, therapeutic effect on Smon aftereffects (see Patent Document 5), calicrane production inhibitory effect, peripheral circulatory disorder improving effect (see Patent Document 6), bone atrophy Improving action (see Patent Document 7), effective nitrogen monoxide production inhibitory action for the treatment of septicemia and endotoxin shock (see Patent Document 8), therapeutic effect for osteoporosis (see Patent Document 9), Nef action inhibitory action and chemokine production inhibitory action. Action-based AIDS therapeutic effect (see Patent Documents 10 and 11), therapeutic effect on ischemic diseases such as cerebral infarction (see Patent Document 12), therapeutic effect on fibromyalgia (see Patent Document 13), treatment for infectious diseases Effect (see Patent Document 14), preventive or alleviating effect of peripheral neuropathy by anticancer agent (see Patent Document 15), therapeutic effect on chronic prostatic inflammation, interstitial cystitis and / or urinary disorder (see Patent Document 16), BDNF It is known that there are a wide variety of actions and effects such as the action of promoting the production of neurotrophic factors such as (see Patent Document 17) and the action of promoting the synthesis of collagen and proteoglycan in cartilage cells (see Patent Document 18). However, it has not been known so far that this extract is effective in improving or treating chondromalacia patella.

Japanese Unexamined Patent Publication No. 53-101515 Japanese Unexamined Patent Publication No. 55-87724 Japanese Unexamined Patent Publication No. 1-265028 Japanese Unexamined Patent Publication No. 1-319422 Japanese Unexamined Patent Publication No. 2-28119 Japanese Unexamined Patent Publication No. 7-97336 Japanese Unexamined Patent Publication No. 8-291707 Japanese Unexamined Patent Publication No. 10-194978 Japanese Unexamined Patent Publication No. 11-80005 Japanese Unexamined Patent Publication No. 11-139977 Japanese Unexamined Patent Publication No. 2000-336034 Japanese Unexamined Patent Publication No. 2000-16942 International Publication WO2004 / 039383 Gazette Japanese Unexamined Patent Publication No. 2004-300146 International Publication WO2009 / 028605 Gazette International Publication WO2011 / 111770 International Publication WO2011 / 162317 International Publication WO2012 / 051173

An object of the present invention is to provide a highly safe drug effective for improving or treating chondromalacia patella.

As a result of diligent research on the drug treatment of chondromalacia patella, for which an effective treatment method is required, the present inventors have found that the preparation containing this extract as an active ingredient is excellent for chondromalacia patellae. The present invention has been completed by finding that it exhibits an improving or therapeutic effect.

This extract has an excellent pharmacological effect of improving or treating chondromalacia patella. Further, since the drug containing the extract as an active ingredient is a highly safe drug with few problems such as side effects, the present invention is extremely useful.

This extract is an extract containing a non-protein active substance extracted and isolated from the inflamed tissue of an animal inoculated with vaccinia virus and smallpox. The present extract is a liquid in the extracted state, but can be made into a solid by drying. A drug containing the present extract as an active ingredient (hereinafter referred to as "the present pharmaceutical product") is extremely useful as a pharmaceutical product. In this case, since this extract is the active ingredient of this preparation, this extract is the drug substance of this preparation. As a specific product manufactured and sold by the applicant in Japan as this preparation, there is "Vaccinia virus-inoculated rabbit inflamed skin extract-containing preparation" (trade name: neurotropin / NEUROTROPIN [registered trademark]). This formulation includes injections and tablets, both of which are ethical drugs.

The indications for injection of neurotropin are "lower back pain, cervicobrachial syndrome, symptomatic nerve pain, pruritus associated with skin diseases (eczema, dermatitis, urticaria), allergic rhinitis, cold sensation of SMON aftereffects. Abnormal perception / pain ". The indications for neurotropin tablets are "postherpetic neuralgia, lumbar pain, cervicobrachial syndrome, periarthritis of the shoulder, osteoarthritis". This product was created by the applicant and developed as a pharmaceutical product, and has been marketed for many years due to its excellent efficacy and safety characteristics, and has established a solid position in the Japanese pharmaceutical market. It is a thing.

The vaccinia virus-inoculated inflammatory tissue extract in the present invention is inoculated with vaccinia virus to crush the inflamed tissue, and an extraction solvent is added to remove tissue fragments, followed by deproteinization treatment, which is adsorbed on an adsorbent. It can be obtained by allowing the virus to elute and then elution the active ingredient. That is, for example, the process is as follows.
(A) Collect skin tissues such as rabbits and mice inoculated with vaccinia virus and crush smallpox, crush the smallpox tissue, and add an extraction solvent such as water, phenol water, physiological saline or phenol-added glycerin water. After that, an extract (filtrate or supernatant) is obtained by filtration or centrifugation.
(B) The extract is adjusted to an acidic pH and heated to remove protein. Then, the deproteinized solution is adjusted to alkaline, heated, and then filtered or centrifuged.
(C) The obtained filtrate or supernatant is acidified and adsorbed on an adsorbent such as activated carbon or kaolin.
(D) An extraction solvent such as water is added to the adsorbent to adjust the pH to alkaline, and the adsorbed component is eluted to obtain a vaccinia virus-inoculated inflamed tissue extract. Then, if desired, the eluate may be evaporated to dryness or freeze-dried under reduced pressure to form a dry solid.

Various animals infected with the vaccinia virus such as rabbits, cows, horses, sheep, goats, monkeys, rats, and mice can be used as animals for inoculating the vaccinia virus to obtain inflammatory tissue, and rabbits can be used as the inflammatory tissue. Inflamed skin tissue is preferred. The rabbit may be anything that belongs to the order Lagomorpha. Examples include rabbits, rabbits (domesticated rabbits), hares (Japanese hares), pikas, and mountain hares. Of these, rabbits are suitable for use. In Japan, there is a rabbit (rabbit) that has been bred for a long time and is often used as livestock or experimental animals, but this is also another name for rabbits. There are many varieties (bleeds) of rabbits, but varieties such as Japanese white rabbits and New Zealand white rabbits (New Zealand white) can be preferably used.

The vaccinia virus may be of any strain. Examples include Lister strains, Dalian strains, Ikeda strains, EM-63 strains, New York City Board of Health strains, and the like.

As a basic extraction step of the present extract, for example, the following steps are used.
Step (A) Collect the inflamed skin tissue that has been inoculated intradermally with vaccinia virus into the skin of rabbits and caused smallpox. The collected skin tissue is washed and disinfected with a phenol solution or the like. Crush this inflamed skin tissue and add 1 to 5 times the amount of extraction solvent. Here, crushing means finely crushing into minced meat using a mincing machine or the like. Further, as the extraction solvent, distilled water, physiological saline, a weakly acidic to weakly basic buffer solution and the like can be used, and a bactericidal / preservative such as phenol, a stabilizer such as glycerin, sodium chloride and potassium chloride are used. , Salts such as magnesium chloride may be added as appropriate. At this time, it is also possible to destroy the cell tissue and facilitate the extraction by treatment with freeze-thaw, ultrasonic waves, cell membrane lytic enzyme, surfactant or the like. The resulting suspension is left for 5-12 days. During that time, the mixture may be heated to 30 to 45 ° C. with or without stirring as appropriate. Tissue pieces are removed from the obtained liquid by solid-liquid separation (filtration or centrifugation, etc.) to obtain a crude extract (filtrate or supernatant).

About step (B) The crude extract obtained in step (A) is subjected to deproteinization treatment. Deproteinization can be carried out by a known method that is usually performed, such as heat treatment, treatment with a protein denaturing agent (for example, an organic solvent such as acid, base, urea, guanidine, or acetone), isoelectric point precipitation, and salting out. Etc. can be applied. Next, the insoluble protein precipitated by a usual method for removing insoluble matter, for example, filtration using filter paper (cellulose, nitrocellulose, etc.), glass filter, Celite, Zytz filter plate, etc., ultrafiltration, centrifugation, etc. Obtain the removed filtrate or supernatant.

Step (C) The filtrate or supernatant obtained in step (B) is adjusted to acidity, preferably pH 3.5 to 5.5, and adsorbed to an adsorbent is performed. Examples of the adsorbent that can be used include activated carbon and kaolin, and the adsorbent is added to the extract and stirred, or the extract is passed through an adsorbent-filled column to add the active ingredient to the adsorbent. Can be adsorbed. When the adsorbent is added to the extract, the solution can be removed by filtration, centrifugation or the like to obtain an adsorbent having the active ingredient adsorbed.

About step (D) In order to elute (desorb) the active component from the adsorbent obtained in step (C), an elution solvent is added to the adsorbent, and the temperature is adjusted to basic, preferably pH 9 to 12, at room temperature. Alternatively, it is appropriately heated or stirred to elute, and the adsorbent is removed by a usual method such as filtration or centrifugation. As the elution solvent used, a basic solvent such as water adjusted to basic pH, methanol, ethanol, isopropanol or the like or an appropriate mixed solution thereof can be used, and water adjusted to pH 9 to 12 is preferable. Can be used. The amount of elution solvent can be set as appropriate. In order to use the eluate thus obtained as a drug substance, the pH is appropriately adjusted to near neutral to finally obtain a vaccinia virus-inoculated rabbit inflamed skin extract (this extract). Can be done.

Since the present extract is a liquid at the time of preparation, it can be appropriately concentrated and diluted to obtain a desired concentration. When the pharmaceutical product is produced from the present extract, it is preferable to perform heat sterilization treatment. In order to make an injection, for example, sodium chloride or the like can be added to prepare an isotonic solution with a physiological saline solution. In addition, it is also possible to produce an oral solid preparation such as a tablet by performing an operation such as appropriate concentration to dryness on the present extract in a liquid state. Specific methods for producing such an oral solid preparation from the present extract are described in Japanese Patent No. 3818657 and No. 4883798. This preparation is an injection or an oral solid preparation thus obtained.

Methods of administering a pharmaceutically effective amount to a patient in need of treatment include subcutaneous, intramuscular, intravenous administration, etc. in addition to oral administration, and the dose depends on the type of inflammatory tissue extract inoculated with vaccinia virus. It can be set as appropriate. The doses permitted for over-the-counter formulations are basically indicated as ethical drugs to administer 16 NU daily for oral administration and 3.6 to 7.2 NU daily for injectables, but the type of disease, The dose can be increased or decreased as appropriate depending on the severity of injury, individual differences of patients, administration method, administration period, etc. (NU: neurotropin unit. Neurotropin unit is a chronic stress animal whose pain threshold is lower than that of normal animals, using SART stress mice. The test is performed according to the modified Randall-Selitto method, and the ED ₅₀ value of the analgesic efficacy is specified. 1NU is the activity showing the analgesic activity-containing component 1 mg of the neurotropin preparation when the _{ED 50 value is 100 mg / kg.} )

The following are examples of methods for producing the present extract, novel pharmacological actions of the present extract, and pharmacological test results on chondromalacia patella, but the present invention is not limited by the description of these examples. No.

Example 1 (Production of this extract)
Vaccinia virus was intradermally inoculated into the skin of a healthy mature rabbit, and the smallpoxed skin was cut out and collected. The collected skin is washed and disinfected with a phenol solution, then the excess phenol solution is removed, crushed, the phenol solution is added and mixed, left for 3 to 7 days, and then stirred for another 3 to 4 days. It was heated to 35-40 ° C. Then, the extract obtained by solid-liquid separation was adjusted to pH 4.5 to 5.2 with hydrochloric acid, heat-treated at 90 to 100 ° C. for 30 minutes, and then filtered to remove protein. Further, the filtrate was adjusted to pH 9.0 to 9.5 with sodium hydroxide, heat-treated at 90 to 100 ° C. for 15 minutes, and then solid-liquid separated.

The obtained deproteinized solution was adjusted to pH 4.0 to 4.3 with hydrochloric acid, activated carbon in an amount of 2% of the mass of the deproteinized solution was added, and the mixture was stirred for 2 hours and then solid-liquid separated. Water was added to the collected activated carbon, the pH was adjusted to 9.5 to 10 with sodium hydroxide, the mixture was stirred at 60 ° C. for 90 to 100 minutes, and then centrifuged to obtain a supernatant. Water was added again to the activated carbon precipitated by centrifugation, the pH was adjusted to 10.5 to 11 with sodium hydroxide, the mixture was stirred at 60 ° C. for 90 to 100 minutes, and then centrifuged to obtain a supernatant. Both supernatants were combined and neutralized with hydrochloric acid to obtain the present extract.

Example 2 (test method)
Next, an example of the test result regarding the action (improvement or therapeutic effect) of the preparation (this preparation) containing the present extract obtained in Example 1 as an active ingredient on chondromalacia patella is shown.

In order to investigate the effect of this product on chondromalacia patella, the following tests were conducted in humans.

(1) Exam (Exam period: January 2011 to September 2012)
Thirty patients with a diagnosis of chondromalacia patella were randomly divided into two groups, a group receiving this drug and a control group. To the control group, 2.0 mL of hyaluronic acid was administered once a week and 5 times by intra-articular injection. To the group administered with this product, 2.0 mL of hyaluronic acid and 1 tube of "neurotropin injection 3.6 units" (trade name of the injection of this product) were administered once a week and 5 times by intra-articular injection. For intra-articular injection, the lateral puncture method of the patellofemoral joint was used. In the supine position, the patient's knee joint is flexed 15 to 20 degrees, the skin is disinfected in the usual way, and a 5 mL injector is used as the puncture site between the superior outer margin of the patella and the lateral condyle of the femur. Place it in the knee joint sac along the patellofemoral joint space, pull the injector to confirm that there was joint fluid, then absorb the joint fluid as completely as possible, and gradually inject it into the joint with the needle left. bottom. Basically, no other drug was used in combination. After the treatment was completed, the therapeutic effect was judged.

(2) Judgment of therapeutic effect The following diagnostic criteria were used to judge the therapeutic effect.
"Healing" was a state of complete recovery.
The "significant effect" was that the main symptoms such as joint pain and swelling disappeared, the joint function was almost recovered, and normal activity was possible.
"Effective" means that the main symptoms such as joint pain and swelling have almost disappeared, joint function has almost recovered or clearly improved, the degree of independence in daily life has increased, and an independent life can be almost achieved, or work or work. The ability to work was partially restored.
"Ineffective" means that the main symptoms such as arthralgia and swelling did not improve, and the joint activity function did not improve compared to before the treatment.
The therapeutic effect was determined using the above diagnostic criteria, and the cure rate, marked efficiency, efficacy rate, and total efficacy rate for the total number of cases in each group were calculated. The results are shown in Table 1.
The two groups were evaluated by knee osteoarthritis scores before and 1 month after treatment. The results are shown in Table 2.

(3) Statistical analysis The chi-square test was used to determine the therapeutic effect between the two groups. A paired t-test was used for the Lysholm scores before and after treatment in the same group, and an independent sample t-test was used for the Lysholm scores before and after treatment in the two groups. In all the tests, the case where p <0.05 was regarded as "significant".

The test results in Tables 1 and 2 above were statistically analyzed. As a result, in the group to which this preparation was administered to patients with chondromalacia patella, the overall efficacy rate was significantly improved as compared with the control group. From this, it was shown that this pharmaceutical product has an effect of generally improving or treating the symptoms of chondromalacia patella.

As described above, it was confirmed that this preparation has an excellent improvement or therapeutic effect on the symptom of chondromalacia patella. In addition, this product has been used for many years and is recognized as a highly safe drug. Therefore, this preparation is extremely useful as an ameliorating or therapeutic agent for chondromalacia patella.

Claims (6)

An ameliorating or therapeutic agent for chondromalacia patella for intra-articular injection, which contains an inflammatory tissue extract inoculated with vaccinia virus as an active ingredient and is used in combination with a hyaluronic acid preparation. Improving or therapeutic agent according to claim 1 symptom of patellar chondromalacia is Ru pain der knee. The improving or therapeutic agent according to claim 1 or 2, wherein the inflamed tissue is an inflamed skin tissue of a rabbit. Use of vaccinia virus-injected inflammatory tissue extract for the production of ameliorating or therapeutic agents for chondromalacia patella for intra-articular injection in combination with hyaluronic acid preparations. Use of the symptoms of patellar chondromalacia is claim 4 Ru pain der knee. The use according to claim 4 or 5 , wherein the inflamed tissue is an inflamed skin tissue of a rabbit.