JPH0588720B2 - - Google Patents
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- Publication number
- JPH0588720B2 JPH0588720B2 JP27062486A JP27062486A JPH0588720B2 JP H0588720 B2 JPH0588720 B2 JP H0588720B2 JP 27062486 A JP27062486 A JP 27062486A JP 27062486 A JP27062486 A JP 27062486A JP H0588720 B2 JPH0588720 B2 JP H0588720B2
- Authority
- JP
- Japan
- Prior art keywords
- structural formula
- hydroxyguanine
- salt
- represented
- compound
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Lifetime
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- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Saccharide Compounds (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Description
（産業上の利用分野）
本発明は、構造式()で表される新規な７−ヒ
ドロキシグアニン誘導体およびその造塩可能なも
のの塩、及びその製造方法、並びにこれを有効成
分とする抗腫瘍剤に関するものである。
(Industrial Application Field) The present invention relates to a novel 7-hydroxyguanine derivative represented by the structural formula (), a salt of the salt-formable salt thereof, a method for producing the same, and an antitumor agent containing the same as an active ingredient. It is related to.
【化】 （発明が解決しようとする問題点） 本発明者等は、下記構造式()[ka] (Problem that the invention attempts to solve) The inventors have proposed the following structural formula ()
【式】
で示される７−ヒドロキシグアニンを化学的ある
いは酵素的に修飾することによつて、いくつかの
誘導体を得ることができ、その薬理作用を広く試
験した結果、構造式()で示される化合物及びそ
の塩が優れた抗腫瘍作用を有することを見出し、
本発明に到達した。
以下に本発明を詳細説明する。
（問題点を解決するための手段及び作用効果）
本発明による新規化合物は、下記の構造式()
で表わされる。By chemically or enzymatically modifying 7-hydroxyguanine represented by the formula, several derivatives can be obtained, and as a result of extensive testing of their pharmacological effects, the derivatives represented by the structural formula () discovered that the compound and its salts have excellent antitumor effects,
We have arrived at the present invention. The present invention will be explained in detail below. (Means and effects for solving the problems) The new compound according to the present invention has the following structural formula ()
It is expressed as
【化】
本発明による化合物は酸性物質であり、塩基と
塩を作ることが可能であるが、本発明による化合
物の塩としては造塩可能な任意のものが対象にな
る。例えば、アルカリ金属、アルカリ土類金属
との塩、アンモニウム塩、アミン塩、特にエ
チルアミン、ジメチルアミン、ピペリジン、モル
フイリンなどとの塩があげられる。これらの塩を
抗腫瘍剤として使用する場合には、生理的に許容
されるものを選ぶべきである。
本発明化合物の２ナトリウム塩の性状は次の通
りである。
融点（分解点） 135〜140℃
分子量 423
元素分析値
実験値 C23.51 H4.40 N13.48 O43.75
C10JH12N5O9PNa2・5H2Oとしての理論値（％）
C23.40 H4.32 N13.65 O43.64
比旋光度〔α〕25 D＝−44.6゜（ｃ＝0.5、H2O）
本発明化合物の紫外吸収スペクトル、赤外部吸
収スペクトル（臭化カリウム錠剤）を夫々図１、
図２に示す。また本発明化合物はFABマススペ
クトルにおいて、水素化分子イオン（（Ｍ＋Ｈ）+）
がｍ／Ｚ＝424に観察された。また試料のナトリ
ウムが水素と置換したイオンがｍ／Ｚ＝402，380
に観測された。ｍ／Ｚ＝402，380のイオンについ
て高分解能マススペクトルを行つた結果、それぞ
れのイオンに対して、組成式C10H14N5O9PNa，
C10H15N5O9Pが、それぞれ10.9mmμ，12.3mmμ
の誤差で得られた。
次に本発明の構造式()に示される７−ヒドロ
キシグアニン誘導体の製法を説明する。
本発明の化合物を製造するにあたつては、基質
として構造式()で示される７−ヒドロキシグア
ニンと、構造式()で示される５−ホスホリボー
ス−１−ピロリン酸に、ホスホリボシルトランス
フエラーゼを作用させることにより、構造式()
に示される７−ヒドロキシグアニン誘導を得るこ
とができる。
本方法に基質として用いられる７−ヒドロキシ
グアニンは、ストレプトミセス・スピーシズ
（Streptomyces sp.）Ａ−347微工研寄第541号
（FERM.BP−541）の培養液から、特開昭61−
178984号に記載された方法に従つて調製できる。
また５−ホスホリボシル−１−ピロリン酸又はそ
れらの塩としては市販品を用いることができる。
本発明に用いるヒポキサンチン・グアニン・ホ
スホリボシルトランスフエラーゼは、動物、微生
物など広く生物界から得られる精製ヒポキサンチ
ン・グアニン・ホスホリボシルトランスフエラー
ゼ，粗ヒポキサンチン・グアニン・ホスホリボシ
ルトランスフエラーゼ，ヒポキサンチン・グアニ
ン・ホスホリボシルトランスフエラーゼ含有物な
どのヒポキサンチン・グアニン・ホスホリボシル
トランスフエラーゼ活性を有するものであり、例
えばヒポキサンチン−グアニンホスホリボシルト
ランスフエラーゼ（米国シグマ社製、酵母由来）
などが挙げられる。
酵素反応は、基質の化合物（７−ヒドロキシ
グアニン）を0.05〜0.2％（ｗ／ｖ）、および化合
物（５−ホスホリボース−１−ピロリン酸）を
0.05〜0.4％（ｗ／ｖ）の範囲で反応液に懸濁し、
酵素を適量、例えば酵素と基質との重量比１：20
乃至１：1000の割合で加え、温度10〜60℃、好ま
しくは25〜35℃の範囲で反応を行ない、高速液体
クロマトグラフイー（HPLC）によつて残基質
（化合物）と生成物（化合物）の量を測定し、
生成物の増加が止つた時点で反応を止めれば良
い。また、酵素反応を行なう際の溶媒のPH範囲は
６〜９、好ましくは7.5〜8.5であり、これらの範
囲でPHの調整を行えばよい。
反応溶媒としては緩衝液の使用が望ましく、ト
リス−塩酸の如き無機酸塩の緩衝液、酢酸ナトリ
ウム、クエン酸ナトリウムの如き有機酸塩の緩衝
液を適宜使用することができる。濃度は緩衝液の
種類にもよるが10mM〜200mM、好ましくは
20mM〜100mMを使用すると良い。
反応液中から生成物を分離する方法としては、
その水溶性、酸性の性質を利用して行えば良い。
反応液中の生成した化合物は、種々の吸着剤
を用いて採取することができる。吸着剤として、
活性炭、弱塩基性カチオン交換樹脂などを利用で
きる。そのほか、セフアデツクスＧ−10，セフア
デツクスLH−20（フアルマシア製）などのゲル
過用担体によるカラムクロマトグラフイーも有
用な精製手段である。
７−ヒドロキシグアニン誘導体（化合物）は
上記の分離法または精製法を適宜組合わせ、ある
いは繰り返すことによつてて純粋に採取できる。
純粋な化合物を得る方法として、例えば水−ア
ルコールなどの混合溶媒系から結晶を析出させる
方法が適している。
次に本発明の抗腫瘍剤について説明する。
本発明による抗腫瘍剤は前記の構造式()で示
される７−ヒドロキシグアニン誘導体またはその
造塩可能なものの塩を有効成分とするものであ
る。本化合物の抗腫瘍作用は以下の試験例に示さ
れる通りである。抗腫瘍作用は以下の如く試験し
た。
BDF1系雌性マウス（体重18〜23g）を用い１
群３匹とした。このマウスに、マウス白血病Ｌ−
1210 １×105個を腹腔内に移植し、被験化合物を
50mM燐酸緩衝液（PH7.4）に溶解したものを、
翌日より連続５日間腹腔内投与して延命率を測定
した。なお、50mMのリン酸緩衝液を腹腔内投与
したマウスを対照とした。
表１に示された結果から、本発明による化合物
は優れた抗腫瘍作用を有することが明らかであ
る。なお、ICR系雄性マウス（体重18〜23g）を
用いた急性毒性試験（腹腔内投与）でのLD50
（mg／Kg）値は150〜300mg／Kgであつた。[Chemical formula] The compound according to the present invention is an acidic substance and can form a salt with a base, and the salt of the compound according to the present invention can be any salt-formable substance. Examples include salts with alkali metals and alkaline earth metals, ammonium salts, amine salts, especially salts with ethylamine, dimethylamine, piperidine, morphiline, and the like. When using these salts as antitumor agents, physiologically acceptable salts should be selected. The properties of the disodium salt of the compound of the present invention are as follows. Melting point (decomposition point) 135-140℃ Molecular weight 423 Elemental analysis value Experimental value C23.51 H4.40 N13.48 O43.75 C 10 JH 12 N 5 O 9 PNa 2・5H 2 Theoretical value as O (%) C23 .40 H4.32 N13.65 O43.64 Specific rotation [α] 25 D = -44.6° (c = 0.5, H 2 O) Ultraviolet absorption spectrum, infrared absorption spectrum of the compound of the present invention (potassium bromide tablet) Figure 1, respectively.
Shown in Figure 2. In addition, the compound of the present invention has hydrogenated molecular ions ((M+H) + ) in the FAB mass spectrum.
was observed at m/Z=424. Also, the ion in which sodium in the sample was replaced with hydrogen is m/Z = 402, 380
was observed. As a result of performing high-resolution mass spectra on ions with m/Z = 402,380, the composition formula for each ion is C 10 H 14 N 5 O 9 PNa,
C 10 H 15 N 5 O 9 P are 10.9mmμ and 12.3mmμ, respectively.
was obtained with an error of Next, a method for producing the 7-hydroxyguanine derivative represented by the structural formula () of the present invention will be explained. In producing the compound of the present invention, 7-hydroxyguanine represented by the structural formula () as a substrate and 5-phosphoribose-1-pyrophosphate represented by the structural formula () are combined with a phosphoribosyl transferer. By acting with ze, the structural formula ()
The 7-hydroxyguanine derivative shown in can be obtained. 7-Hydroxyguanine used as a substrate in this method was obtained from the culture solution of Streptomyces sp.
It can be prepared according to the method described in No. 178984.
Moreover, commercially available products can be used as 5-phosphoribosyl-1-pyrophosphoric acid or salts thereof. The hypoxanthine guanine phosphoribosyltransferase used in the present invention includes purified hypoxanthine guanine phosphoribosyltransferase, crude hypoxanthine guanine phosphoribosyltransferase, which is obtained from a wide range of living organisms such as animals and microorganisms, Hypoxanthine-guanine-phosphoribosyltransferase-containing substances having hypoxanthine-guanine-phosphoribosyltransferase activity, such as hypoxanthine-guanine phosphoribosyltransferase (manufactured by Sigma, USA, derived from yeast)
Examples include. The enzyme reaction was performed using 0.05-0.2% (w/v) of the substrate compound (7-hydroxyguanine) and the compound (5-phosphoribose-1-pyrophosphate).
Suspended in the reaction solution in the range of 0.05 to 0.4% (w/v),
Use an appropriate amount of enzyme, for example, a weight ratio of enzyme to substrate of 1:20.
The reaction is carried out at a temperature of 10 to 60°C, preferably 25 to 35°C, and the residual substances (compounds) and products (compounds) are separated by high performance liquid chromatography (HPLC). measure the amount of
The reaction may be stopped when the product stops increasing. Further, the PH range of the solvent used in the enzymatic reaction is 6 to 9, preferably 7.5 to 8.5, and the PH may be adjusted within these ranges. It is desirable to use a buffer as the reaction solvent, and buffers of inorganic acid salts such as Tris-hydrochloric acid and buffers of organic acid salts such as sodium acetate and sodium citrate can be used as appropriate. The concentration depends on the type of buffer, but is preferably 10mM to 200mM.
It is best to use 20mM to 100mM. As a method for separating the product from the reaction solution,
This can be done by taking advantage of its water-soluble and acidic properties. Compounds produced in the reaction solution can be collected using various adsorbents. As an adsorbent,
Activated carbon, weakly basic cation exchange resin, etc. can be used. In addition, column chromatography using gel filtration carriers such as Cephadex G-10 and Cephadex LH-20 (manufactured by Pharmacia) is also a useful purification method. The 7-hydroxyguanine derivative (compound) can be purified in a pure manner by appropriately combining or repeating the above separation or purification methods.
A suitable method for obtaining a pure compound is, for example, a method in which crystals are precipitated from a mixed solvent system such as water-alcohol. Next, the antitumor agent of the present invention will be explained. The antitumor agent according to the present invention contains as an active ingredient a 7-hydroxyguanine derivative represented by the above-mentioned structural formula () or a salt-formable salt thereof. The antitumor effect of this compound is as shown in the following test example. The antitumor effect was tested as follows. 1 using BDF 1 female mice (weight 18-23 g).
There were 3 animals in the group. This mouse was infected with murine leukemia L-
Five 1210 1×10 cells were implanted intraperitoneally, and the test compound was administered.
Dissolved in 50mM phosphate buffer (PH7.4),
Starting from the next day, the drug was administered intraperitoneally for 5 consecutive days to measure the survival rate. Note that mice to which 50 mM phosphate buffer was intraperitoneally administered were used as controls. From the results shown in Table 1, it is clear that the compounds according to the invention have excellent antitumor effects. In addition, LD 50 in an acute toxicity test (intraperitoneal administration) using ICR male mice (body weight 18-23 g)
(mg/Kg) values were between 150 and 300 mg/Kg.
【表】
治療マウスの平均生存日数
＊ 延命率％＝[Table] Average survival days of treated mice
* Life extension rate % =
Claims (1)
シグアニン誘導体およびその塩。 【式】 ２ 構造式() 【式】 で示される７−ヒドロキシグアニン又はその塩
と、構造式() 【化】 で示される５−ホスホリボース−１−ピロリン酸
を基質とし、ヒポキサンチン・グアニン・ホスホ
リボシルトランスフエラーゼを作用させることを
特徴とする、構造式() 【化】 で表わされる７−ヒドロキシグアニン誘導体の製
造方法。 ３ ヒポキサンチン・グアニン・ホスホリボシル
トランスフエラーゼが酵母由来である特許請求の
範囲第２項記載の製造方法。 ４ 下記の構造式()で表される７−ヒドロキシ
グアニン誘導体またはその生理的に許容される塩
を有効成分とする抗腫瘍剤。 【化】[Claims] 1. A 7-hydroxyguanine derivative represented by the following structural formula () and a salt thereof. [Formula] 2 Using 7-hydroxyguanine or its salt represented by the structural formula () [Formula] and 5-phosphoribose-1-pyrophosphate represented by the structural formula () [Chemical] as substrates, hypoxanthine guanine - A method for producing a 7-hydroxyguanine derivative represented by the structural formula (), which is characterized by allowing phosphoribosyltransferase to act. 3. The production method according to claim 2, wherein the hypoxanthine guanine phosphoribosyltransferase is derived from yeast. 4. An antitumor agent containing a 7-hydroxyguanine derivative represented by the following structural formula () or a physiologically acceptable salt thereof as an active ingredient. [ka]
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP27062486A JPS63122698A (en) | 1986-11-13 | 1986-11-13 | 7-hydroxyguanine derivative and production thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP27062486A JPS63122698A (en) | 1986-11-13 | 1986-11-13 | 7-hydroxyguanine derivative and production thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS63122698A JPS63122698A (en) | 1988-05-26 |
| JPH0588720B2 true JPH0588720B2 (en) | 1993-12-24 |
Family
ID=17488681
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP27062486A Granted JPS63122698A (en) | 1986-11-13 | 1986-11-13 | 7-hydroxyguanine derivative and production thereof |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS63122698A (en) |
-
1986
- 1986-11-13 JP JP27062486A patent/JPS63122698A/en active Granted
Also Published As
| Publication number | Publication date |
|---|---|
| JPS63122698A (en) | 1988-05-26 |
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