KR100632250B1 - Anticancer agent containing arsenic compound as an active ingredient - Google Patents

Abstract

The present invention relates to a novel arsenic compound and an anticancer agent comprising the same as an active ingredient, wherein the NaH ₂ As ₃ O ₉ (sodium hydrogen arsenate; sodium arsenate trioxide; Asolveren) of the present invention is less toxic than As ₂ O ₃ . While it is about one third and the anti-cancer effect is more than doubled, the side effects are much weaker and are expected to be useful for some malignancies of humans.

Arsenic compounds, sodium triarsenite, anticancer

Description

Anticancer agent containing arsenic compound as an active ingredient {ANTICANCER AGENT COMPRISING ARSENIC COMPOUND}

1 schematically illustrates a lattice structure of NaH ₂ As ₃ O ₉ , wherein FIGS. 1A to 1C are NaH ₂ As ₃ O ₉ , and FIG. 1D is As ₂ O ₃ .

2 is an X-ray lattice analysis chart of NaH ₂ As ₃ O ₉ .

The present invention relates to a novel arsenic compound and an anticancer agent comprising the same as an active ingredient, and specifically, a new arsenic compound having low toxicity and superior anticancer effect compared to arsenic acid, NaH ₂ As ₃ O ₉ (sodium hydrogen arsenate; triarsenic oxide 9). Sodium; Asolveren) and methods for their preparation, and their use as anticancer agents.

Among the arsenic (As) compounds, arsenic trioxide (As ₂ O ₃ ) is toxic and is mainly used for insecticides, herbicides, wood preservatives, and the coloring of glass. It has been used as a therapeutic agent for diseases and as a treatment for leukemia, etc. until the middle of the 20th century (Seas, DA, History of the treatment of chronic myelocytic leukemia. Am. J. Med. Sci, 296 : 85-86, 1988; Sir William Osler's The principles and practice medicine, 1892; 李 時珍, Herbal Wood, 春陽 堂, 1928. However, by the 1980s, even small amounts of As ₂ O ₃ Recognition that prolonged exposure (skin contact) can cause cancer (Leonard, A., Carcinogenicity, teratogenicity and mutagenicity of arsenic. Mutat. Res. 75 : 49-62, 1980) ceased use as a treatment for disease. .

Meanwhile, Shen et al. In China ( Blood, 89 : 3354-3360, 1999) reported that acute promyelocytic leukemia (APL) caused resistance to the treatment of retinoic acid (As ₂ O ₃ ). Since the use of injectables has been reported, many hospitals and researchers around the world have used arsenic acid for the treatment of several solid type tumors as well as acute promyelocytic leukemia.

Regarding such a mechanism of action of As ₂ O _{3 as} an anticancer agent, at present (1) the blocking of blood vessels to cancer tissue; (2) inhibition of telomerase transcription ( Cancer Research, 59 , 6033-6037, December 15, 1999); And (3) apoptosis of cancer cells ( The J. of Clinical Investigation, Nov. 2001, 108: 10, 1541-1547).

It is generally known that the LD ₅₀ of the _trivalent oxide As ₂ O ₃ and its derivative oxides is 39.4 mg / kg for mice and the LD ₅₀ for mice is 15.1 mg / kg (The Merck Index: 11th Ed. Published by Merck Co., Inc. USA, 1989.).

In addition, the agonists at the cellular unit of arsenic acid (As ₂ O ₃ ) are not As ₂ O ₃ molecules but there is a study showing that AsO ₃ -ion (The Pharmacological Basis of Therapeutics, Goodman and Gilman, Macmillan Pub. Co. New York, 1985).

The present inventors have focused on the above-described mechanism of action of arsenic acid, and focused on finding and researching As-based compounds that deteriorate cancer tissues of patients and minimize side effects. The problem was the toxicity of arsenic acid (As ₂ O ₃ ), which produced NaH ₂ As ₃ O ₉ , which is thought to emit the best AsO ₃ -ion of several As-based oxides, and studied its anticancer activity. As a result, the present invention was completed by discovering that the anticancer effect was about 2 times higher than that of As ₂ O ₃ and the toxicity was low.

An object of the present invention is toxic than As ₂ O ₃ from the As-based compound is to provide a low antitumor effect, while the As-based compound is excellent for new and their preparation.

In order to achieve the above object, the present invention provides an arsenic compound having the formula: NaH ₂ As ₃ O ₉

In addition, the present invention provides an anticancer agent comprising NaH ₂ As ₃ O ₉ (sodium hydrogen arsenate; sodium triarsenite oxoxide; Asolveren) as an active ingredient.

NaH ₂ As ₃ O ₉ according to the present invention is mixed with As ₂ O ₅ and Na ₂ O and heated at about 400 ℃ for about 2 to 4 days, or by adding water to As ₂ O ₃ and Na ₂ O It is prepared by heating at 30 ° C. for about 6 to 10 hours.

Arsenic (As) has an atomic number of 33 and an electron arrangement is 2 in the 1S orbit of the K shell, 8 in the L and 18 in the M, and 2 and 4P orbits in the 4S orbit of the outer N. Three are present, and as for the oxide of As, a trivalent material (As ₂ O ₃ ) and a pentavalent material (As ₂ O ₅ ) are present. In general, pentavalent As compounds are less toxic than pentavalent As compounds, have better penetration into tissues, and can inhibit cell division of cancer tissue by releasing Arsenoxide radicals such as AsO ₃ ^- slowly. (Cutting's Hand Book of Pharmacology, Fourth Ed. Appleton-Century-Crofts Educational Division Menedith Co. New York, USA, 1969.)

NaH ₂ As ₃ O ₉ of the present invention may be prepared according to any one of the following methods:

(1) Reaction method at 400 ° C. (As ₂ O ₅ is used as substrate)

As ₂ O ₅ (arsenic pentoxide) was added to the crucible used for high thermal chemical reaction, Na ₂ O (sodium oxide) was added to the upper layer, mixed and heated at about 400 ° C. for about 2-4 days. Fibrous NaH ₂ As ₃ O ₉ is obtained.

(2) Reaction method at 30 ° C. (As ₂ O ₃ is used as substrate)

Put As ₂ O ₃ into a crucible and then add water to make a solution. Then, Na ₂ O is added and heated at about 30 ℃ for about 6 to 10 hours to separate NaH ₂ As ₃ O ₉ . . As ₂ O ₃ is poorly soluble, but 2 g is dissolved when 100 ml of water is heated to 90 ° C. and can be dissolved even at 20 to 10 ° C.

The physicochemical properties of NaH ₂ As ₃ O ₉ thus prepared are as follows:

NaH ₂ As ₃ O ₉ (sodium hydrogen arsenate; sodium triarsenic oxide 9; Asolveren)

Molecular Weight: 393.77

Shape of crystal: white fibrous

Melting Point (mp): 280 ± 5 ℃

Boiling Point (bp): 500 ± 10 ℃

Solubility:

1. Solubility in water (H ₂ O): insoluble at 0-50 ° C., 2.0 g dissolved per 100 ml at 70 ° C. Once the solution is formed, it is dissolved even below 20 ℃.

2. Insoluble in ethanol or chloroform.

1 schematically illustrates a lattice structure of NaH ₂ As ₃ O ₉ , wherein FIGS. 1A to 1C are NaH ₂ As ₃ O ₉ , and FIG. 1D is As ₂ O ₃ . 2A and 2B are X-ray lattice analysis charts of NaH ₂ As ₃ O _{9, and} the high resolution triple axis x-ray diffractometer of the Daegu branch of the Korea Research Institute of Basic Science (Philips); Phillips, X. Pert MRD, USA).

An anticancer agent comprising NaH ₂ As ₃ O ₉ according to the present invention as an active ingredient is administered parenterally, such as intravenous administration by dividing the amount of 0.15-0.75 mg / kg into one to three times per day for adults, or 0.2 to 1.0 per day. An amount of mg / kg may be administered orally.

In addition, an anticancer agent comprising NaH ₂ As ₃ O ₉ according to the present invention as an active ingredient may be formulated according to conventional preparations such as injections, powders, granules, capsules, tablets and solutions. However, when using As-based anticancer drugs such as As ₂ O ₃ or NaH ₂ As ₃ O ₉ , it is recommended to use a method of administering intravenous or oral administration for a certain period and then administering again after a rest period.

Hereinafter, the present invention will be described in more detail with reference to Examples. However, these Examples are only illustrative of the present invention, and the scope of the present invention is not limited thereto.

Preparation Example: NaH ₂ As ₃ O ₉ Manufacture

(1) Reaction method at 400 ° C. (As ₂ O ₅ is used as substrate)

10 g of As ₂ O ₅ (arsenic pentoxide) was added to a 200 ml capacity crucible used for high thermal chemical reaction, and 3 g of Na ₂ O (sodium oxide) was added to the upper layer and mixed for 72 hours at 400 ° C. heating to obtain a fiber of _{NaH 2 As 3 O 9 7 g} ( yield 70%).

(2) Reaction method at 30 ° C. (As ₂ O ₃ is used as substrate)

5 g of As ₂ O ₃ was added to a crucible (crucible, 800 ml capacity) and 400 ml of H ₂ O was added to make a solution. Then, 1 g of Na ₂ O was added thereto at 30 ° C. for 8 hours. 3 g of NaH ₂ As ₃ O ₉ was separated by heating (yield 60%).

Toxicity Test: NaH ₂ As ₃ O ₉ With As ₂ O ₃ LD intraperitoneally ₅₀ Chi

(1) LD ₅₀ value of NaH ₂ As ₃ O ₉ was measured by the following method. That is, 72 Swiss-Webster mice (mouse, male, average weight 28 g) were divided into 9 groups of 8 animals each, physiological saline for group 1 animals, and NaH in physiological saline for groups 2 to 9 animals. ₂ As ₃ O ₉ was administered by dissolving 10, 20, 30, 40, 50, 60, 70 or 80 mg / kg, respectively.

The number of animals who died 7 days prior to intraperitoneal administration was 1 (13%) in the 30 mg / kg group, 2 (25%) in the 40 mg / kg group, or 4 (50%) in the 50 mg / kg group. ), And 6 animals (75%) in the 60 mg / kg administration group. The dose of NaH ₂ As ₃ O ₉ (mg / kg), the number of dead animals, the mortality rate and the like in each animal group are shown in Table 1 below.

Fauna The number of animals Dosage of NaH ₂ As ₃ O ₉ (mg / kg) Number of animals killed in 14 days of observation % Mortality One 8 0 0 0 2 8 10 0 0 3 8 20 0 0 4 8 30 One 13 5 8 40 2 25 6 8 50 4 50 7 8 60 6 75 8 8 70 8 100 9 8 80 8 100

As a result of the above experiment, the LD ₅₀ value in the intraperitoneal administration of NaH ₂ As ₃ O ₉ can be estimated to about 50 mg / kg. On the other hand, Na (Arsenate, Sodium) of As (arsenic) has been reported to be not carcinogenic in primates (monkey, nonhuman primates) (Glod, LS, Environ. Health Perspect. 107 (suppl4) : 527-600, 1999).

(2) Determination of LD ₅₀ value by intraperitoneal administration of As ₂ O ₃ was followed by the test of NaH ₂ As ₃ O ₉ .

That is, 56 Swiss-Webster species mice (mouse, male, 28 g average weight) were divided into 7 groups of 8 animals, and 0.5 ml of saline was injected into the abdominal cavity for the control group, and As ₂ O ₃ was added to 10 saline each. , Lysed to 13, 15, 20, 25 or 30 mg / kg and dissolved and injected intraperitoneally of each animal group.

The number of animals killed up to 7 days after intraperitoneal injection of As ₂ O ₃ was 2 (25%) in the 13 mg / kg group, 4 (50%) in the 15 mg / kg group, and 20 mg / kg in the group. Six (75%) and eight (100%) were administered in the 25 mg / kg group. The experimental results are shown in Table 2 below.

Fauna The number of animals Dose of As ₂ O ₃ (mg / kg) Animals killed during the 14-day observation period Lethality One 8 0 0 0 2 8 10 0 0 3 8 13 2 25 4 8 15 4 50 5 8 20 6 75 6 8 25 8 100 7 8 30 8 100

As a result of the above experiment, in case of Swiss-Webster mice, the LD ₅₀ value was 15 mg / kg upon intraperitoneal injection of As ₂ O ₃ saline solution.

Efficacy test 1: NaH on prolonged survival in mouse-induced Ehrlich Ascites Carcinoma ₂ As ₃ O ₉ Effect

Forty female Swiss Webster species (mouse, purchased through Kyungpook National University) are divided into five groups of eight dogs with an average body weight of 28 g. The animals are divided into five groups, and Kleim ( Exper. Cell Research 2 : 528-573, 1851) and the like by injecting 0.2 ml (containing 1.4 × 10 ³ cells) of 10% ascites cancer solution into the abdominal cavity to induce ascites cancer.

Ascites-induced mice were administered with As ₂ O ₃ or NaH ₂ As ₃ O ₉ for a period of time to observe the life prolonging effect of each animal. The allocation of experimental animals and the dosages of As ₂ O ₃ and NaH ₂ As ₃ O ₉ are shown in Table 3 below. Each drug was administered six times every 72 hours, starting 24 hours after intraperitoneal injection of cancer cells.

Fauna The number of animals Medication: Intraperitoneal Administration 1. Control group 8 6 times 0.3 ml saline 2. As ₂ O ₃ 5 mg / kg administration group 8 5 mg / kg of As ₂ O ₃ was dissolved in 0.3 ml of saline at 36 ° C. and administered 6 times. 3. As ₂ O ₃ 10 mg / kg administration group 8 10 mg / kg of As ₂ O ₃ was dissolved in 0.3 ml of saline solution at 36 ° C. and administered 6 times. 4. NaH ₂ As ₃ O ₉ 5 mg / kg administration group 8 5 mg / kg of NaH ₂ As ₃ O ₉ was dissolved in 0.3 ml of saline at 36 ° C. and administered 6 times. 5. NaH ₂ As ₃ O ₉ 10 mg / kg administration group 8 10 mg / kg of NaH ₂ As ₃ O ₉ was dissolved in 0.3 ml of saline at 36 ° C. and administered 6 times.

Ehrlich Ascites Carcinoma was induced in mice, and As ₂ O ₃ and NaH ₂ As ₃ O ₉ were administered to observe the life prolonging effect of each animal.

Laboratory animals The number of animals weight Drug dosage Average survival time (days) Survival rate (%) 1. Control group 8 28.7 ± 0.8 0 14.2 ± 0.5 100 2. As ₂ O ₃ administration group 8 27.6 ± 0.6 5 mg / kg 47.2 ± 4.3 336 3. As ₂ O ₃ administration group 8 29.0 ± 1.1 10 mg / kg 80.1 ± 5.2 571 4. NaH ₂ As ₃ O ₉ administration group 8 28.1 ± 1.2 5 mg / kg 92.2 ± 4.8 657 5. NaH ₂ As ₃ O ₉ administration group 8 28.9 ± 1.0 10 mg / kg 124 ± 5.9 886

As shown in Table 4, the control animals that did not receive the drug survived for 14 days on average after cancer cell transplantation, and the animals that received 5 mg / kg of As ₂ O ₃ averaged 47 days and 10 mg / kg As _2. O ₃ administered animals survived on average 80 days. On the other hand, animals administered 5 mg / kg of NaH ₂ As ₃ O ₉ survived on average 92 days and animals receiving 10 mg / kg NaH ₂ As ₃ O _{9 on} average 124 days. In the animals treated with 10 mg / kg of As ₂ O ₃ , the movement was slowed 10 to 30 minutes after injection, and there was a sleeping animal, but returned to normal before 5 hours. No particular abnormalities were observed in the animal group administered As ₂ O ₃ of 5 mg / kg. On the other hand, in all the animal groups administered NaH ₂ As ₃ O ₉ , no side effects as seen in the 10 mg / kg As ₂ O ₃ administration group was observed.

In summary the above experimental _{results, As 2 O 3 5 ㎎ /} ㎏ was the survival time when As ₂ O ₃ 10 ㎎ / ㎏ administration extended three times compared to when _{administered, NaH 2 As 3 O 9 10} ㎎ / ㎏ administered is Survival was extended by nine times compared to control. In addition, the NaH ₂ As ₃ O ₉ 5 mg / kg administration was longer than the As ₂ O ₃ 10 mg / kg administration. Moreover, when 10 mg / kg of NaH ₂ As ₃ O ₉ was administered, the degree of side effects observed when As ₂ O ₃ 10 mg / kg was administered was also weak.

In conclusion, the anticancer effect of NaH ₂ As ₃ O ₉ is more than twice that of As ₂ O ₃ and the side effects are much weaker.

The use of As ₂ O ₃ in several human cancers (malignancy) was 0.15 mg / kg / iv / day as of 2004 (Solgnet SL, et al., United States multicenter study arsenic trioxide in relapsed acute promyelocytic leucemia, J Clin.Oncol . 19 : 3852-3860, 2001). In the case of NaH ₂ As ₃ O ₉ of the present invention, in consideration of the above experimental results, it would be preferable to use it in an amount of about 0.15 mg / kg / iv / day to 0.5 mg / kg / iv / day. It is thought that it can be increased up to 1 mg / kg / iv / day.

As described above, the arsenic compound NaH ₂ As ₃ O ₉ of the present invention (sodium hydrogen arsenate; sodium triarsenite nitrate; Asolveren) is about one third the toxicity compared to As ₂ O ₃ and anticancer effect While more than two times, the side effects are much milder and are expected to be useful for some malignancies in humans.

Claims (3)

delete An anticancer agent comprising NaH ₂ As ₃ O ₉ (sodium hydrogen arsenate; sodium triarsenic oxide 9; Asolveren) as an active ingredient. Method of producing NaH ₂ As ₃ O ₉ (sodium hydrogen arsenate; sodium arsenate trioxide; Asolveren), characterized in that water is added to As ₂ O ₃ and Na ₂ O is added and heated at 30 ℃ for 6 to 10 hours. .

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