DE2319472A1 - Abrin contg. anticancer compsns - esp. for treating uterine tumours - Google Patents

Abstract

Compsn. contain abrin (I) together with conventional auxiliaries and carriers (I), a toxic protein from Abrus precatorius is applied intratumourally (0.0005-0.05 mg/day); i.p. or intra-arterially (0.05-0.5 mg/day) or topically (0.5-5 mg/day). A suitable soln. for injection comprises 10 mg(I) in 10 ml. 0.01 N cold acetic acid, 1 ml. soln. freeze-dried and packaged in an ampoule for making up with normal saline.

Description

Anti-cancer pharmaceutical preparations containing abrin The present invention relates to pharmaceutical preparations which have anti-cancer effects and contain abrin as an active ingredient. By giving Abrin to cancer sufferers A procedure for cancer treatment is provided to patients.

According to the present invention there is an anti-cancer pharmaceutical preparation provided which contains abrin and a pharmaceutically acceptable carrier.

It is known that abrin is a toxic protein (toxalbumin), which has a molecular weight of about 65,000, and of course in the seeds the paternoster pea, Abrus precatorius.

The acute toxicity of Abrin as LD 50 is 0.02 mg per kg of body weight (in mice, intraperitoneally). As a result of its highly toxic properties were Try to use.

The preparation of abrin for medical purposes has not yet been carried out.

However, it has been found, in a completely surprising manner, that the administration of abrin in much smaller amounts than its toxic amount is effective in controlling human cancer. In particular, was found that Abrin is used to treat uterine cancer in female patients especially suitable is.

Various pharmaceutical preparations can be produced with advantage which Abrin together with liquid, solid or semi-solid diluents contain. Suitable preparations include injectable solid preparations and semi-solid preparations. The injectable solid preparation for extemporaneous Dilution can and can be produced by methods known in the art other pharmaceutically active materials such as local anesthetics and other anti-cancer agents etc. included. The injectable solid preparation can be administered intratumorally, intraperitoneally, retroperitoneally or intra-arterially, after diluting the preparation in suitable concentration with suitable diluents, such as normal saline solution, administered. The semi-solid preparation, i.e. ointment, can be used using known ointment base materials, such as, B. white petrolatum, liquid petrolatum, Lanolin, vegetable oils, waxes, polyethylene glycols etc. or mixtures thereof will. To favor the diffusion of abrin into the tissue, the above preparations described additionally contain a suitable amount of hyaluromidase.

The Abrindose varies depending on the administration method or the type or stage of cancer. The recommended Abrindose is from about 0.0005 mg to about 0.005 mg daily, in the case of intratumoral administration, and from about 0.05 mg to about 0.5 mg daily, in the case of the intraperitoneal, retroperitoneal or intra-arterial administration. In the case of a topical or local application of the ointment, much larger amounts, such as 0.5 to 5 mg or more of abrin, can be used.

For example, a daily dose of Abrin ointment is from about 1 to about 10 g can be administered to the surface of the tumor or crater, each Gram contains about 0.0005-0.005g parts depending on the type and stage of the carcinoma.

The anti-cancer effect of Abrin appears slowly in the case of treatment by topical or local administration in the form of an ointment while. they very much fast and drastic in the case of an intra-arterial, continuous infusion and intraperitoneal administration appears. In the ointment treatment and the continuous intra-arterial infusion, no side effects are observed, while some side effects, such as dizziness, are temporary Fever and a trace of albuminuria in some cases with intraperitoneal and can occur after intratumoral administration.

Although various methods of obtaining abrin from Abrus precatorius have already been known, is an advantageous method of obtaining high purity abrin by J. Y. Lin et al "The Journal of the Formosan Medical Association, 68, 518-521 (1969) off. The details of this procedure are as follows: 100 g of Abrus precartorius kernels are poured into 500 ml of 5% acetic acid at 4 ° C Soaked overnight -and then homogenized in a Waring mixer. The homogenate is placed in 250 ml bottles in an International centrifuge for 20 minutes Centrifuged 5,000 g and discarded the residue. Becomes the protruding phase solid ammonium sulfate slowly added with stirring to a saturation of 45 ffi.

The precipitate in which very little toxic protein was found is filtered off. Solid ammonium sulfate becomes the supernatant phase added up to 100% saturation. The resulting precipitate is distilled in Dissolve water and continue by heating the toxic protein in a water bath cleaned at 600c for 30 minutes. The resulting protein precipitate will abaentrifugiert and the supernatant phase against distilled water at 4 ° C dialyzed for 24 hours with multiple exchanges of the distilled water.

Some precipitation of protein occurred during dialysis, which then removed by centrifugation. The remaining phase is on DEAE-Sephadex A-50 (sold by Pharmacia A. B., Uppsala, Sweden) Column (2 x 50 cm), which was previously equilibrated with 0.005 m sodium acetate.

The samples are collected in a Gilson fraction collector.

The fractions of the protein peak are pooled and measured to approx 20 mg protein per milliliter concentrated by evaporation. With evaporation of the water from the surface of the bag, the crystals appear, which is caused by the Shimmer of the solution is displayed. The crystals are used for recrystallization dissolved in distilled water by adding a few drops of 0.1 N acetic acid. Then this is dialyzed against distilled water and that after several times Change of the distilled water occurring crystals filtered off.

Thus, 120 mg of pure abrin are obtained as fine rod-shaped crystals obtain.

The product shows a single band on disk gel electrophoresis. The absorption (280 nm) of the product is determined to be 12.4 in a 1 cm cell. The ratio of the optical adsorption at 280 and 260 nm is 1.95. It shows ensure that the product is free of protease activity and hemaglutination activity is.

The invention is described below by the examples which serve for illustration purposes only. Abrin was used in these examples, which was produced according to the method described above.

Examples I and II illustrate preparations containing abrin.

Example I Ointment Crystalline abrin (50 mg) is dissolved in 5 ml of a 0.1 Dissolved in acetate buffer, pH 4.3, and then 1,000 units of hyaluronidase added in 2 ml of normal saline solution. 5 ml of polyethylene glycol are added to the solution added. This mixture is mixed homogeneously with 75 g of tramycin ointment 1tPfizer11 - (each Gram- contains Oxitetracycline x HCl, which is equivalent to 30 mg Oxitetracycline, 10,000 units of polymyxin B sulfate, white petrolatum and liquid petrolatum) (Chas Pfizer Corp., USA) mixed.

Depending on the size of the tumor or crater, for example Apply 1 to 10 grams or more of this ointment to the surface of the uterine crater or crater are given.

Example II Injectable Solid Preparation Crystalline Abrin (Jan. mg) are dissolved in 10 ml of 0.01 N cold acetic acid and passed through a Chamberland filter filtered. In each case 1 ml of this solution is filled into a sterilized container and then lyophilized. Each thiol or ampoule contains 1 mg of abrin.

When in use, the contents of the ampoule are in appropriate concentration with suitable liquid diluents, such as normal saline solution (normal saline) diluted. If necessary, the solution obtained in this way can be further pharmaceutically active materials and / or additives such as stabilizers and preservatives etc. can be added. The preparation could be administered intratumorally, intraperitoneally, retroperitoneally, or intra-arterially will.

The following Examples III through VIII illustrate the treatment of uterine cancer with abrin ointment. In these examples abrin ointment was used, which was produced by the method corresponding to Example I.

Example III Case L.S., age 56. Uterus cervical "Cancer of the epidermal" Type in stage 1. A papillomatoma the size of a thumb with external growth on the posterior lip was visible.

The Abrin ointment was applied to the cervical tumor once a day while one month applied. The daily dose of Abrin ointment was from about 1 g to about 10 g varies depending on the size of the cancer. A few days after After the administration of abrin, superficial hemorrhagic spots were found in the tumor observed and the entire tumor changed its color from pink-red to liver-yellow, gradually shrank and flattened completely 10 days after the start of administration from and 4 days afterwards the tumor had completely disappeared.

The surface of the tumor base showed erosion with a slope bleeding to the touch for a week and then became normal mucosa Covered 4 weeks after the start of therapy. A repeated PapanieolaouCest took place negative. A biopsy performed one month after therapy only showed questionable carcinoma in situ in two of five specimens, although facing the cervix looked normal to the normal eye.

Example IV Case C.S.J., age 46. Adenocarcinoma type uterine cervical cancer stage I. A fungoid necrotic tumor the size of a chicken egg Size completely covered the vaginal lumen.

In this case a particular mode of administration was used, i. a diaphragm (contraceptive) that was coated with 2.0 g of Abrin ointment was on the cervical tumor with gauze tightly packed and tightly pressed. The diaphragm was changed daily. Three weeks after starting therapy, the tumor flattened and the external mouth could be made out - 6 weeks after the start of therapy the cancerous tissue nearly disappeared and the cervix showed a broad substantial papillary erosion with cervical hypertrophy.

Example V Case L.S.C, Age 56. Endocervical Cancer, Stage II-b, epidermoid type. The en-docervix formed a thumb-sized crater with inclusion of the upper third of the vagina.

About 1 to 10 g of Abrin ointment were applied to the surface of the crater once applied daily for three weeks. Clarified two weeks after starting therapy the vaginal wound and after another week the crater in the Size decreases and the crater wall is gradually cleaned. It became another Tendency to contact bleeding noted.

Example VI Case H.S.Y., Age 46. Stage II-b. Extremely anemic and cachectic. Epidermoid type. A reflection examination revealed one necrotic crater of a size exceeding the size of a hen's egg, which encircles the cervix and the upper half of the posterior vaginal wall involved, with papillomatus growth, which stretched like a dike along the edge of the crater. A rectal exam revealed an apple-sized tumor that completely occupied the pelvis and the anterior rectal wall squeezed together, and that the paramaterial infiltration was there, such as the bilateral one To reach the pelvis wall. However, the rectal mucosa was still intact. The physical one Inclusion was achieved by enlarging the uterus with its fundus 12 cm above the symphysisjindicated.

It was a month-long application of the Abrin ointment with about 1 to 10 g once a day, depending on the size of the cancer. This resulted to a lowering of the uterine fundus to 7 cm above the symphysis and the decrease the size of the pelvic tumor from apple to hen's egg size. Spontaneous bowel movements which she had never exhibited before therapy was due to the reduction rectal compression has become possible. Anemia and the general conditions had improved a lot.

Example VII Case Y. L. Y., age 58. Existing cancer injuries on the vaginal stump after radical surgery. Papanicolaa test positive. The pathological examination of the surgical specimens also revealed posterior vaginal impacts, whose distal cross-sectional line was provided with carcinomas. Epidermoid Type. After two weeks of administration of the AbrinT ointment with about 1 to 10 g once The cancer injury was granulated daily depending on the size of the cancer Tissue replaced as shown histopathologically.

Example VIII Case L. C. C., age 53. Epidermoid Carcinoma. Recurring Injury in the right lateral vaginal wall, which is removed and subsequently has been irradiated. After three months, the BiopsS showed that the cancer cells were still degenerate.

After local administration of the abrin ointment of about 1 to 10 g once daily, depending on the size of the cancer, for two weeks Cancer cells disappeared, as evidenced by the Papanicol: test, as well as by the biopsy became.

Examples IX and X illustrate the intraperitoneal administration of abrin .. In these examples, the injectable solid preparation of Example II used. When used, an injectable solution was made by dissolving the preparation in 1 ml of 0.1 M acetate buffer (4.3). An aliquot of this, which one contained the amount of abrin required for treatment became more normal at 500 ml Saline solution to which 600 units of hyaluronidase have been added. The solution was introduced to the patient through an intraperitoneal and attached to the abdominal skin fixed polyethylene tube dripped in.

Example IX Case C.Y. S.L., age 44. Primary papular cystadenocarcinoma of the bilateral ovaries with 5,000 ml of ascites and visceral spread. From the day After the operation up to the 13th day, an abrin solution containing 0.) mg was administered intraperitoneally administered at three-day intervals. On the 14th day, 0.28 mg of abrin was administered. A total of 1.78 mg of abrin was used. After the third administration, there was none observed further accumulation of ascites and had general conditions improves considerably.

A control operation (second-look operation) was carried out on the 27th day. The abdominal cavity was from cancerous lesions with the exception of ascites and 700 ml only two tiny grayish degenerated nodules free, which removed and the Laboratotium were sent for examination. The cythological examination revealed no cancer cells in ascites and the pathological examination showed that the two tiny nodules of fat tissue.

Example X Case C.W.S., age 44. Recurrent ovarian cancer developed huge metastatic injuries in the liver after radiation, which was proven by X-rays (increased diaphragm) and radioisotope testing became. After emptying 2,500 ml of ascites, a 0.1 mg abrin was used Solution administered intraperitoneally daily. The 3.5 finger before the start of treatment broad liver tumor palpable below the rib area was within a week after the start of therapy has become almost untouchable. The accumulation of ascites was less pronounced and to the Neuralgia radiating from the shoulder had significantly mitigated, so narcotics only used once a day after treatment were required while the patient had narcotics every two hours prior to therapy had requested.

Examples XI and XII illustrate intratumoral administration. In these examples the injectable solid preparation of Example II was used. In use, an injectable solution was prepared by dissolving the preparation in 1 ml of 0.1 M acetate buffer (pH 4.3) prepared and 0.1 ml of the resulting solution then diluted with 500 ml of normal saline and then 600 units of hyaluronidase added. 10 to 20 ml (0.002 to 0.004 mg of abrin) of this solution, which is immediately Before use, 2 ml of 1% lidocaine was added to the tumor and injected the neighboring tissues.

Example XI Case C. U. Y., Age 44. Recurrent tumor in the Pelvis and vaginal stump after previous evisceration due to uterine cervical cancer of the epidermoid type in stage IV. Pronounced anemic and cachectic. As above described, 0.002 to 0.004 mg of abrin were injected into the tumor.

Various injections resulted in significant shrinkage of the tumor and to relieve neuralgia. Also the general condition had improves.

Example XII Case H, M., Age 59. Recurring and metastatic Adenocarcinoma of the right inguinal node. Multiple injections from 0.002 to 0.004 mg of abrin produced a remarkable shrinkage of the tumor.

Example XIII Retroperitoneal Administration of Abrin Case W.C. C. M., age 47. Epidermoid Cancer.

The preparation according to Example II became more normal in one milliliter Saline solution dissolved and 0.1 ml of the resulting solution was made up to 500 ml with normal saline, and then 600 units of hyaluronidase was added. The total volume (500 ml) of this solution was given to the patient through a polyethylene tube, which was inserted retroperitoneally and attached to the abdominal skin, instilled. The same treatments were given daily for the purpose of destroying the cancer cells, possibly in the pelvis after the previous evisceration of the uterine cervical cancer Stage IV were left over, repeated.

Despite the widespread spread of the cancer, the patient was fine very good.

Example XIV Continuous intra-arterial infusion of Abrin.

Case L.A. S., age 49, stage IV uterine cervical cancer with hemorrhagic ascites, spreads to the uterus serosa, vesicuterine reflection, Dougles pouch (Douglas pouch), rectum and intestinal serosa, thumb-sized to Metastatic injuries in the bladder wall the size of a table tennis ball, widespread Pelvic lymph node metastases detected by biopsy at laparathomy and chicken egg large gyrus-like cervical tumor. Epidermoid Carcinoma.

An umbilical arterial catheter "acidic" (Cat. No. Mar 1602-5, size 5 Fr. length 38.1 cm) the patient was in each case in the bilateral internal ilium artheria on the ventral side between the ligature close to the body and the distal clamp introduced and attached to the vessels. The two catheters were retroperitoneal stirred out through the abdominal wall, where it is fitted with a Y-shaped glass fitting connected by a three-way valve and then further to a SIP-11 pump connected by a silicone tube. The preparation corresponding to Example II was dissolved in 1 ml of normal saline and 0.2 ml (Abrin 0.2 mg) of the resulting Solution was diluted with 500 ml of normal saline. The solution was 600 units of hyaluronidase and 40 mg of heparin added. It became a continuous one intra-arterial infusion of the Abrin solution discontinued. The drip speed was adjusted to a constant infusion of 27 ml of the solution per hour. at the last treatment a solution containing 0.1 mg abrin was used. To to date 1.5 mg of abrin have been used. A speculum examination on the following Day revealed that the gyrus-like external growth of the cervix, the size of a hen's egg in size (hen's egg) with overflowing serous and thin discharge in the Vagina was reduced, and by the second day the tumor was small to the size of a chicken egg decreased. The size of the tumor gradually decreased, almost reaching its original size The shape of his outer mouth was palpable, but it was still somewhat papillomatous external growth around the latter. It was very interesting to have the tumor during the overall course of the infusion had become anemic and brittle. The thin discharge was caused by bloody colored discharge from the external mouth from the fifth postoperative Replaced day on. There was no hematological change and no change observed in blood chemistry. A low level fever was caused by E. Coli infection of the urinary tract, which is well controlled by Wintmylon (nalidixic acid) became.

No dizziness. The patient was doing very well.

Claims (1)

Claims 1. Pharmaceutical preparation with anti-cancer effect g e k e n n z e i c h e t together with a content of abrin as an active ingredient, if necessary with the usual auxiliaries and carriers.