JPH07304732A - Fluorine-containing vitamin d3 analogue - Google Patents

Abstract

PURPOSE:To obtain a new compound having low hypercalcemic action and high differentiating-inducing action of tumor cells and useful as a carcinostatic agent. CONSTITUTION:A compound of formula I (R is a protecting group of H and OH). This compound is obtained by carrying out coupling reaction of a ketone substance of formula II with an anion derived from a phosphine oxide of formula III (Ph is phenyl) and then as necessary, deprotecting the reaction product. The induction of phosphine oxide to anion is carried out in the presence of a base (e.g. n-butyl lithium) and the coupling reaction is carried out under inert atmosphere (argon) in a solvent (e.g. THF), at -100 to 50 deg.C for 10min to 24hr.

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention relates to a novel vitamin D ₃ analog useful as an anticancer agent because it has a low blood calcium-elevating action and a high tumor cell differentiation-inducing action.

[0002]

2. Description of the Related Art An in vivo metabolite of vitamin D ₃ ,
1α, known as active-form vitamin D _3, 25-dihydroxyvitamin D ₃ has a calcium absorption promoting action from the intestine, is known to be effective for the treatment of such bone lesions. In addition, recently, this active vitamin D ₃ and its analogs have an action of inducing differentiation to return cancerous cells to normal cells (Tanaka Hirofumi et al .: Biochemistry 55, 1323, 1923).
1983), and in fact, some of them have an action to prevent the progression of cancer (K. W. Colton et al., Lancet , Jan. 28 , p. 188, 1989). It recognized. However, active vitamin D ₃ has a strong effect on calcium metabolism, and this effect causes an undesirable side effect of increasing blood calcium when using vitamin D ₃ as an anticancer agent.

Vitamin D ₃ which exhibits biological activity similar to vitamin D ₃
As a derivative, the general formula:

[Chemical 2] The compound represented by (generic name: OCT) is Murayam
a), E. et al . Chem. Pharm . Bull ., 34, 4410 (1986) .
Year) and other general formulas:

[Chemical 3] The compound represented by (product name: KH1060) is WO90 /
09991 (PCT / DK90 / 00036).

[0004]

DISCLOSURE OF THE INVENTION Problems to be Solved by the Invention The inventors of the present invention have shown an excellent effect on calcium metabolism in vivo (that is, they have no side effects and therefore do not show hypercalcemia). The present invention was completed by conducting research for the purpose of creating a novel vitamin D ₃ analog compound showing an excellent cell differentiation-inducing action for cancerous cells, finding vitamin D ₃ having desired properties. An object of the present invention is to provide a novel vitamin D ₃ analog having a carcinostatic action based on a pharmacological action, particularly a cell differentiation inducing action. Another object of the present invention is to provide a method for producing the vitamin D ₃ analog. These objects of the invention and the advantages afforded by the invention will be apparent to those skilled in the art from the following description.

The novel vitamin D ₃ analogs provided by the present invention have the general formula:

[Chemical 4] (In the formula, R is a hydrogen atom or a hydroxyl-protecting group). Here, as the hydroxyl-protecting group, acetal-based protecting groups such as methoxymethyl, ethoxyethyl, methoxyethoxymethyl, and tetrahydropyranyl, silyl ethers such as trimethylsilyl, triethylsilyl, t-butyldimethylsilyl, and t-butyldiphenylsilyl. Examples thereof include a system protecting group, an acyl group such as acetyl, propionyl, n-octanoyl, and benzoyl.

Specific examples of the compound represented by the general formula [I] include general formula [IA]:

[Chemical 5] (Compound A); 1α, 3-bis (methoxymethyl) ether of Compound A; 1α, 3-bis (trimethylsilyl) ether of Compound A; 1α, 3-bis (t-butyldimethylsilyl) ether of Compound A; Compound 1α, 3-bis of A
(Tetrahydropyranyl) ether; Compound A 1α, 3
-Bis (acetyl) ester; 1α, 3-bis of compound A
(Methoxyethoxymethyl) ether; Compound A 1α,
3-bis (ethoxyethyl) ether; 1α of compound A,
3-bis (triethylsilyl) ether; 1 of compound A
α, 3-bis (propionyl) ester; 1α of compound A,
3-bis (n-octanoyl) ester; 1 of compound A
Examples include, but are not limited to, α, 3-bis (benzoyl) ester.

The compound [I] of the present invention can be produced by various methods. An example of the best method is shown below. That is, the formula
[II]:

[Chemical 6] And a ketone represented by the general formula [III]:

[Chemical 7] (Wherein R represents a hydroxyl-protecting group and Ph means phenyl) is subjected to a coupling reaction with an anion derived from a phosphine oxide and, if necessary, deprotected. can get. The induction of phosphine oxide to the anion is achieved in the presence of a base,
The base used is preferably alkyl lithium such as n-butyl lithium.

The above coupling reaction of the compound [II] and the compound [III] is carried out, for example, at -100 ° C to 50 ° C, preferably at -80 ° C to 20 ° C, under an inert atmosphere (for example, argon atmosphere). 10 minutes ~ in an ether solvent (eg, diethyl ether, tetrahydrofuran (THF), etc.)
It is carried out for 24 hours, preferably 30 minutes to 2 hours. The obtained product [I] can be purified by a known method such as silica gel chromatography. If necessary, deprotection of the hydroxyl group from compound [I] can be performed by a known method.

The production method of the starting compound [III] used in the above coupling reaction is described by EG Baggio.
lini et al.), J. Am. Chem. Soc., 104, 294.
Page 5, 1982 and JP-A-2-250844. Another starting material [II] can be produced by the following reaction steps. The following reaction steps also show an example of the above-mentioned method for converting the starting materials [II] and [III] into the compound [I] of the present invention.

[Chemical 8] (In the formula, Bn means benzyl, MOM means methoxymethyl, TBDMS means t-butyldimethylsilyl)

According to the above reaction step, first, the aldehyde compound (1) is converted into the formic acid ester compound (2), and the compound (2) is passed through the alcohol compound (3) and the ketone compound.
Get (4). Reduction to the compound (4) gives the compound (5). Compound (5) is reacted with bromide (9) to give ether compound (6) and then methoxymethyl protecting group is removed to give compound (7). The benzyl protecting group is removed from compound (7) to give diol compound (8). The diol (8) is oxidized to obtain a ketone compound (starting material [II]). The other starting material R was t-butyldimethylsilyl.
(TBDMS) which is a phosphine oxide [III] to obtain a compound [IB] of the present invention in which R is TBDMS, the protecting group is removed from this compound, and a compound [IA] of the present invention in which R is hydrogen. (Compound A) is obtained. The detailed reaction conditions of the above reaction steps are described in Reference Examples 1 to 8 and Examples 1 and 2 below.
Described in. Hereinafter, the present invention will be specifically described with reference to Reference Examples, Examples, and Test Examples, but the present invention is not limited thereto. Further, each compound listed in Reference Examples and Examples is numbered, and this number corresponds directly to the compound number given to each compound in the reaction step.

[0011]

EXAMPLES Reference Example 1 Synthesis of Compound (2) from Compound (1): Aldehyde Compound
(1) (1.80 g, 6 mmol) of dichloromethane (32 ml)
To the solution was added m-chloroperbenzoic acid (1.60 g, 9.3 mmol) at room temperature and the mixture was stirred for 18 hours. After the insoluble matter is filtered off, the filtrate is extracted with dichloromethane, and the organic layer is washed with an aqueous sodium thiosulfate solution, a saturated aqueous sodium hydrogen carbonate solution and saturated saline. After drying over anhydrous magnesium sulfate, the solvent was distilled off, and the formate ester compound (2)
(1.9 g) is obtained.

Reference Example 2 Synthesis of compound (3) from compound (2): The above compound (2) was dissolved in methanol (20 ml) and anhydrous potassium carbonate (1.7) was added.
g, 12.3 mmol), and stirred at room temperature for 1 hour. The mixture is extracted with diethyl ether and water, the organic layer is washed with water and saturated brine, and dried over anhydrous magnesium sulfate. The solvent was evaporated, the residue was purified by column chromatography, and the alcohol compound (3) (1.35 g,
Yield 78%). The physical properties of compound (3) are as follows. IR (neat): 3396, 2932, 2873, 1454,
1109,735,696 cm ^-1 . ¹ H-NMR (CDCl ₃ ) δ: 0.97 (s, 3H), 1.22
(d, J = 5.4Hz, 3H), 3.62-3.80 (m, 2H), 4.
36 (d, J = 12.1Hz, 1H), 4.62 (d, J = 12.1
Hz, 1H), 7.20-7.40 (m, 5H).

Reference Example 3 Synthesis of compound (4) from compound (3): Oxalyl chloride (3.
To a solution of 6 ml, 41.3 mmol) in dichloromethane (212 ml) was added dimethyl sulfoxide (DMSO) (3.9 ml, 5 ml).
A solution of 5.0 mmol) in dichloromethane (14 ml) was added to -78.
Add dropwise at 0 ° C. and stir the mixture for 10 minutes. Then, a solution of the alcohol compound (3) (1.0 g, 3.47 mmol) in dichloromethane (70 ml) was added dropwise to the mixture, and the mixture was stirred at -78 ° C for 15 minutes and at -45 ° C for 1 hour. Triethylamine (22 ml, 158 mmol) was added to the reaction mixture, the mixture was stirred at 0 ° C. for 20 minutes, saturated aqueous ammonium chloride solution was added, and the mixture was extracted with diethyl ether. The organic layer is dried over anhydrous magnesium sulfate. The solvent was distilled off to obtain the ketone compound (4) (0.79 g).

Reference Example 4 Synthesis of compound (5) by hydrogenation of compound (4): The above ketone compound (4) was dissolved in ethanol (30 ml) and sodium borohydride (500 mg, 13.2 mmol) was added. , Stir at room temperature for 24 hours. A saturated aqueous ammonium chloride solution is added to the reaction mixture, and the mixture is extracted with diethyl ether. The organic layer is washed with water and saturated brine and dried over anhydrous magnesium sulfate. The solvent was evaporated, the residue was purified by column chromatography, and the alcohol compound (5) (6
02 mg, yield 60%) and alcohol compound (3) (13
5 mg, 14% yield) are obtained. The physical properties of compound (5) are as follows. IR (neat): 3419, 2928, 2864, 1454,
1090,735,696 cm ^-1 . ¹ H-NMR (CDCl ₃ ) δ: 1.06 (s, 3H), 1.13
(d, J = 6.1Hz, 3H), 3.65-3.85 (m, 2H), 4.
35 (d, J = 12.3Hz, 1H), 4.53 (d, J = 12.3)
Hz, 1H), 7.18-7.38 (m, 5H).

Reference Example 5 Synthesis of Compound (6) from Compound (5): Alcohol Compound (5) (288 mg, 1.0 mmol) obtained in Reference Example 4, Bromide (9) (1.33 g, 3.99) To a mixture of (mmol) and potassium hydride (35% oil suspension, 2 ml) was added crown ether (18-C-6, 264 mg) in THF (4 ml) and the mixture was stirred at room temperature for 18 hours. The reaction mixture is extracted with diethyl ether, the organic layer is washed with water and saturated brine, and dried over anhydrous magnesium sulfate. The solvent was distilled off to give a crude ether compound (6) (0.25
g) is obtained.

Reference Example 6 Synthesis of compound (7) by deprotection of compound (6): The above-mentioned crude ether compound (6) was dissolved in dioxane (5 ml), concentrated hydrochloric acid (0.5 ml) was added, and a mixed solution was prepared. Is stirred at 65 ° C. for 24 hours. The reaction mixture is returned to room temperature, water is added, and the mixture is extracted with diethyl ether. The organic layer is washed with water, saturated aqueous sodium hydrogen carbonate solution and saturated brine, and dried over anhydrous magnesium sulfate. The solvent was distilled off to obtain the crude compound (7) (0.1).
22 g) are obtained.

Reference Example 7 Synthesis of Compound (8) by Deprotection of Compound (7): Reference Example 6
Dissolve the compound (7) obtained in step 5 in methanol (20 ml) and add 5%
Hydrogen atmosphere (1 atm) in the presence of Pd-C catalyst (100 mg)
Stir at room temperature for 18 hours. The catalyst was filtered off, the solvent was distilled off and the residue was purified by column chromatography to obtain the diol compound (8) (184 mg, yield 45%). The physical properties of compound (8) are as follows. IR (neat): 3435, 2935, 1455, 1206,
720 cm ^-1 . MS (EI) 391 [M + 1]: 253,209,163,1
07,81. ¹ H-NMR (CDCl ₃ ) δ: 0.95 (s, 3H), 1.13
(d, J = 6.0Hz), 3.32-3.63 (m, 3H), 4.10
(brs, 1H), 6.81 (s, 1H).

Reference Example 8 Synthesis of Compound [II] by Oxidation of Compound (8): Reference Example 7
A solution of the diol compound (8) (180 mg, 0.44 mmol) obtained in 1) in dichloromethane (5 ml) was added to a suspension of pyridinium chlorochromate (PCC, 300 mg, 1.39 mmol) in dichloromethane (15 ml), and the mixture was added. Stir at room temperature for 2 hours. The reaction mixture is extracted with diethyl ether,
After concentrating the ether layer, the residue is subjected to column chromatography.
Purified with (Florisil), ketone compound [II] (179 m
g, yield 100%). The physical properties of the compound [II] are as follows. IR (neat): 3238, 1713, 1205, 718cm
^-1 . MS (EI) 404 [M ⁺ ]: 253,209,179,15
2,111,81. ¹ H-NMR (CDCl ₃ ) δ: 0.65 (s, 3H), 1.17
(d, J = 5.9Hz, 3H), 3.34-3.62 (m, 4H), 6.
61 (s, 1H).

Example 1 Synthesis of compound [IB] by Wittig reaction of compound [II] and compound [III] in which R is TBDMS: R is TB
DMS, a phosphine oxide compound [III] (1.1
To a solution of 5 g (1.97 mmol) in THF (10 ml) was added -78 n-butyllithium (hexane solution, 1.97 mmol).
Add at 0 ° C. and stir the mixture at the same temperature for 5 minutes. The reaction mixture was charged with TH of compound [II] (90 mg, 0.22 mmol).
Add F (5 ml) solution. The mixture is gradually warmed to room temperature, saturated aqueous ammonium chloride solution is added, and the mixture is extracted with diethyl ether. The solvent is distilled off and the residue is purified by column chromatography to obtain the compound of the present invention [IB] (171 mg, yield 100%) in which R is TBDMS.

Example 2 Synthesis of compound [IA] by deprotecting compound [IB]: Compound [IB] obtained in Example 1 was ion-exchange resin (50 W).
X 4.3 g) in methanol (30 ml) suspension at room temperature
Stir for hours. After removing the ion exchange resin by filtration, methanol was distilled off under reduced pressure, and the residue was purified by column chromatography to obtain the compound of the present invention [IA] (114 mg, yield 95%) in which R is hydrogen. [α] ¹⁸ _D : -39.0 ° (c = 0.97, methanol). UV (ethanol) λmax ²² °: 264 nm (ε16564). ¹ H-NMR (CD ₃ OD) δ: 0.58 (s, 3H), 1.09
(d, J = 5.9Hz, 3H), 4.12 (m, 1H), 4.35 (t,
J = 5.8Hz, 1H), 4.92 (brs, 1H), 5.28 (brs,
1H), 6.07 (d, J = 11.6Hz, 1H), 6.33 (d, J
= 11.6 Hz, 1 H).

Test Example 1 Effect of the Compound of the Present Invention on Serum Calcium Concentration Test Method: Three-week-old Wistar rat (male, Japan SL)
(Company C) is a low calcium-low vitamin D ₃ product of CLEA Japan.
The rats were raised on a diet to produce low-calcium serum rats. The rats were subcutaneously administered with the solvent and the test compound in 0.1 ml for 5 days. From the day after the first administration, the calcium concentration in the serum was measured every day, and the control rat to which only the solvent was administered and the rat to which the solvent and the test compound were administered were compared to assay the blood calcium elevating effect. The calcium concentration increase value was put on the dose action curve of the standard product (1α, 25-dihydroxyvitamin D ₃ ) and expressed by the magnification with the standard product. Test compound: 1α, 25- (OH) ₂ D ₃ compound A (compound of the present invention) OCT (known compound represented by the chemical formula 2) KH1060 (known compound represented by the chemical formula 3) Test result: The test results are shown in the following table. Shown in 1.

Table 1 Test compounds Magnification 1α, 25- (OH) ₂ D ₃ 1 Compound A 0.1-0.2 OCT <0.01 KH1060 1.3 As is apparent from the above results, the compound of the present invention is 1
Compared with α, 25-dihydroxyvitamin D ₃ , the blood calcium elevation action is weak.

Test Example 2 Cell Differentiation Inducing Action Test Method: Human leukemia cells (HL-60) were used for tissue culture.
A 6-well plate was inoculated at a concentration of 1.5 × 10 ⁵ cells / ml, 20 μl of each test compound was added, and the cells were cultured for 3 days. Take 100 μl of the supernatant, add 100 μl of NBT reagent, and incubate at 37 ° C. for 40 minutes. Counting black stained cells was calculated standard intensity of differentiation-inducing effects on (1,25-dihydroxyvitamin D ₃₎ (relative intensity). Test compound: The same compound as in Test Example 1 above was used. Test results:

Table 2 Test compounds Relative strength 1α, 25- (OH) ₂ D ₃ 1 Compound A 11.0 OCT 0.6 KH1060 150.0 From the results in Table 2 above, according to the compound A of the present invention, 1α of the standard product, 25 compared to dihydroxyvitamin D _3, it is understood that more HL-60 cells are induced to differentiate.
Furthermore, from the results of Table 1 and Table 2 above, the known compound O
CT has a very weak blood calcium-inducing action and cell differentiation-inducing action, while the known compound KH1060 has a strong cell differentiation-inducing action, but blood calcium-elevating action is 1α, 2.
It is about the same as 5- (OH) ₂ D ₃ .

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Katsuhiko Izeki, Miyukigaoka, Tsukuba, Ibaraki Prefecture, Daikin Industry Co., Ltd. (72) Inventor, Yunwa Yuten, Miyukigaoka, Tsukuba, Ibaraki Prefecture, Daikin Industry Co., Ltd.

Claims (2)

[Claims] 1. A compound represented by the general formula [I]: (In the formula, R is a hydrogen atom or a hydroxyl-protecting group.) A fluorine-containing vitamin D ₃ analog. 2. A hydroxyl-protecting group is methoxymethyl, ethoxyethyl, methoxyethoxymethyl, tetrahydropyranyl, trimethylsilyl, triethylsilyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, acetyl, propionyl, n-octanoyl and benzoyl. The compound of claim 1 selected from the group consisting of: