JP2006219459A - Cancer cell proliferation inhibitor composed of alkoxytetrahydrofuran derivative, apoptosis inducing agent and medicine containing the same - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a cancer cell proliferation inhibitor and apoptosis inducing agent effective for the treatment of diseases such as malignant tumor and cancer and provide a medicine containing the cancer cell proliferation inhibitor and/or apoptosis inducing agent. <P>SOLUTION: The invention provides a cancer cell proliferation inhibitor and an apoptosis inducing agent composed of an alkoxytetrahydrofuran derivative and a medicine containing the inhibitor, etc. Examples of the alkoxytetrahydrofuran derivative are alkylidenealkoxytetrahydrofuran derivative and benzylidenealkoxytetrahydrofuran derivative. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to a cancer cell growth inhibitor and an apoptosis inducer comprising an alkoxytetrahydrofuran derivative, and a drug containing the same, and the object thereof is a cancer cell proliferation inhibitor and apoptosis effective for the treatment of diseases such as malignant tumors and cancers. Another object is to provide a pharmaceutical containing an inducer and the cancer cell growth inhibitor and / or apoptosis inducer.

  Because cancer is the leading cause of death, people are very interested in preventing it. In general, cancer cells grow very slowly at an early stage of malignant transformation from normal cells. However, when cancer cells are generated, it is observed that new blood vessels newly branched from the surrounding blood vessels migrate toward the cancer cells. Once the blood vessel reaches this lesion site, inexhaustible nutrients and oxygen are supplied to the cancer cells, which not only starts to explode, but also causes remote metastasis through the blood vessel. That is, since malignant tumor / cancer cells have disordered cell growth, it can be said that suppressing their cell growth is indispensable in cancer treatment.

  In recent years, research related to apoptosis has become active in the field of cancer research. Apoptosis is a physiological cell death caused by normal cell replacement in a living body, and is distinguished from necrosis (necrosis) which is a pathological cell death. So-called programmed cell death is cell death observed in certain types of cells that have been programmed to die in vivo, and apoptosis is one of them. In apoptosis, phenomena such as nuclear chromatin condensation and chromosomal DNA fragmentation are characteristically observed.

  In general, all cells are normally differentiated, become specialized cells, and as a result have individual functions, but at this point, they are responsible for biological functions. Unnecessary cells undergo apoptosis and undergo a process of being phagocytosed and removed by macrophages and the like.

  In contrast to the normal differentiation of such cells, cancer cells (so-called tumors) are thought to arise because cells that should have died no longer die. That is, since cancer cells cause abnormal growth without being controlled for growth, one method for suppressing the growth of cancer cells is to induce apoptosis. Therefore, for example, in Patent Document 1, an apoptosis inducer containing an evening primrose extract as an active ingredient, and in Patent Document 2, a brain tumor therapeutic agent containing a protein containing a specific peptide as an active ingredient can induce apoptosis. Are listed.

In view of the above, if there is a drug that can induce apoptosis of cancer cells in order to suppress the growth of cancer cells, it can be said that it can be an ideal anticancer agent. That is, cancer cells are effectively eliminated through a normal cell death process while suppressing abnormal growth. Today, the development of such effective drugs is expected.
JP 2002-307663 A JP 2003-128575 A

An object of the present invention is to provide a cancer cell growth inhibitor and an apoptosis inducer effective for treatment of diseases such as malignant tumors and cancers, and pharmaceuticals containing them.

The invention according to claim 1 relates to a cancer cell growth inhibitor comprising an alkoxytetrahydrofuran derivative represented by the following formula (Formula 1).

（式中Ｒ_１及びＲ_２は、同一又は異なって、水素原子、芳香族置換基又は炭素数１〜１０のアルキル基を表す。Ｒ_３は炭素数１〜１０のアルキル基、或は次式（化２）又は（化３）で表される置換基である。）

(In the formula, R ₁ and R ₂ are the same or different and each represents a hydrogen atom, an aromatic substituent or an alkyl group having 1 to 10 carbon atoms. R ₃ is an alkyl group having 1 to 10 carbon atoms, or (It is a substituent represented by (Chemical Formula 2) or (Chemical Formula 3).)

（式中ｎは１〜１０の整数、ｍは１〜１０の整数であり、式中Ｘは、‐ＣＯＯ‐（エステル）、‐Ｏ‐、‐ＮＲ_４‐（Ｒ_４は、水素原子、炭素数１〜６のアルキル基又は芳香族置換基を表す）又は‐Ｓ‐を表す。）

(Wherein n is an integer of 1 to 10, m is an integer of 1 to 10, and X is —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon Represents an alkyl group of 1 to 6 or an aromatic substituent) or -S-).

（式中ｎは４〜１０の整数であり、式中Ｘは、‐ＣＨ_２‐、‐ＣＯＯ‐（エステル）、‐Ｏ‐、‐ＮＲ_４‐（Ｒ_４は、水素原子、炭素数１〜６のアルキル基又は芳香族置換基を表す）又は‐Ｓ‐を表す。）
請求項２に係る発明は、次式（化４）で示されるアルコキシテトラヒドロフラン誘導体からなるアポトーシス誘導剤に関する。

(In the formula, n is an integer of 4 to 10, wherein X is —CH ₂ —, —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon number 1 to 6 represents an alkyl group or an aromatic substituent) or -S-.
The invention according to claim 2 relates to an apoptosis inducer comprising an alkoxytetrahydrofuran derivative represented by the following formula (Formula 4).

（式中Ｒ_１及びＲ_２は、同一又は異なって、水素原子、芳香族置換基又は炭素数１〜１０のアルキル基を表す。Ｒ_３は炭素数１〜１０のアルキル基、或は次式（化５）又は（化６）で表される置換基である。）

(In the formula, R ₁ and R ₂ are the same or different and each represents a hydrogen atom, an aromatic substituent or an alkyl group having 1 to 10 carbon atoms. R ₃ is an alkyl group having 1 to 10 carbon atoms, or (It is a substituent represented by (Chemical Formula 5) or (Chemical Formula 6).)

（式中ｎは１〜１０の整数、ｍは１〜１０の整数であり、式中Ｘは、‐ＣＯＯ‐（エステル）、‐Ｏ‐、‐ＮＲ_４‐（Ｒ_４は、水素原子、炭素数１〜６のアルキル基又は芳香族置換基を表す）又は‐Ｓ‐を表す。）

(Wherein n is an integer of 1 to 10, m is an integer of 1 to 10, and X is —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon Represents an alkyl group of 1 to 6 or an aromatic substituent) or -S-).

（式中ｎは４〜１０の整数であり、式中Ｘは、‐ＣＨ_２‐、‐ＣＯＯ‐（エステル）、‐Ｏ‐、‐ＮＲ_４‐（Ｒ_４は、水素原子、炭素数１〜６のアルキル基又は芳香族置換基を表す）又は‐Ｓ‐を表す。）
請求項３に係る発明は、前記アルコキシテトラヒドロフラン誘導体が、前記式（化１）中のＲ_１がフェニル基、Ｒ_２が水素原子、或はＲ_１が水素原子、Ｒ_２がフェニル基であるベンジリデンアルコキシテトラヒドロフラン誘導体であることを特徴とする請求項１記載の癌細胞増殖阻害剤に関する。
請求項４に係る発明は、前記アルコキシテトラヒドロフラン誘導体が、前記式（化４）中のＲ_１がフェニル基、Ｒ_２が水素原子、或は（化４）中のＲ_１が水素原子、Ｒ_２がフェニル基であるベンジリデンアルコキシテトラヒドロフラン誘導体であることを特徴とする請求項２記載のアポトーシス誘導剤に関する。
請求項５に係る発明は、前記アルコキシテトラヒドロフラン誘導体が、前記式（化１）中のＲ_３がブチル基であるブトキシテトラヒドロフラン誘導体であることを特徴とする請求項１又は３記載の癌細胞増殖阻害剤に関する。
請求項６に係る発明は、前記アルコキシテトラヒドロフラン誘導体が、前記式（化４）中のＲ_３がブチル基であるブトキシテトラヒドロフラン誘導体であることを特徴とする請求項２又は４記載のアポトーシス誘導剤に関する。
請求項７に係る発明は、前記アルコキシテトラヒドロフラン誘導体が、次式（化７）で示される（Z）‐４‐ベンジリデン‐２‐ｎ‐ブトキシテトラヒドロフランであることを特徴とする請求項５記載の癌細胞増殖阻害剤に関する。

(In the formula, n is an integer of 4 to 10, wherein X is —CH ₂ —, —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon number 1 to 6 represents an alkyl group or an aromatic substituent) or -S-.
In the invention according to claim 3, the alkoxytetrahydrofuran derivative is a benzylidene wherein R ₁ in the formula (Formula 1) is a phenyl group, R ₂ is a hydrogen atom, or R ₁ is a hydrogen atom, and R ₂ is a phenyl group. 2. The cancer cell growth inhibitor according to claim 1, which is an alkoxytetrahydrofuran derivative.
The invention according to claim 4, wherein the alkoxy tetrahydrofuran derivative, R ₁ is a phenyl group of the Formula (4), R ₂ is a hydrogen atom, or (Formula 4) R ₁ is a hydrogen atom in, R ₂ 3. The apoptosis-inducing agent according to claim 2, wherein is a benzylidenealkoxytetrahydrofuran derivative having a phenyl group.
The invention according to claim 5 is the inhibition of cancer cell proliferation according to claim 1 or 3, wherein the alkoxytetrahydrofuran derivative is a butoxytetrahydrofuran derivative in which R ₃ in the formula (Formula 1) is a butyl group. It relates to the agent.
The invention according to claim 6 relates to the apoptosis inducer according to claim 2 or 4, wherein the alkoxytetrahydrofuran derivative is a butoxytetrahydrofuran derivative in which R ₃ in the formula (Formula 4) is a butyl group. .
The invention according to claim 7 is the cancer according to claim 5, wherein the alkoxytetrahydrofuran derivative is (Z) -4-benzylidene-2-n-butoxytetrahydrofuran represented by the following formula (Formula 7): The present invention relates to a cell growth inhibitor.

請求項８に係る発明は、前記アルコキシテトラヒドロフラン誘導体が、次式（化８）で示される（Z）‐４‐ベンジリデン‐２‐ｎ‐ブトキシテトラヒドロフランであることを特徴とする請求項６記載のアポトーシス誘導剤に関する。

The invention according to claim 8 is the apoptosis according to claim 6, wherein the alkoxytetrahydrofuran derivative is (Z) -4-benzylidene-2-n-butoxytetrahydrofuran represented by the following formula (Formula 8): It relates to an inducer.

請求項９に係る発明は、乳癌、前立腺癌、膵癌、胃癌、肺癌、大腸癌（結腸癌、直腸癌、肛門癌）、食道癌、十二指腸癌、頭頚部癌（舌癌、咽頭癌）、脳腫瘍、神経鞘腫、非小細胞肺癌、肺小細胞癌、肝臓癌、腎臓癌、胆管癌、子宮癌（子宮体癌、子宮頸癌）、卵巣癌、膀胱癌、皮膚癌、血管腫、悪性リンパ腫、悪性黒色腫、甲状腺癌、骨腫瘍、血管腫、血管線維腫、網膜肉腫、陰茎癌、小児固形癌、カポジ肉腫、ＡＩＤＳに起因するカポジ肉腫、上顎洞腫瘍、線維性組織球腫、平滑筋肉腫、横紋筋肉腫のうちいずれかの治療或は予防のために用いられることを特徴とする請求項１，３，５又は７に記載の癌細胞増殖阻害剤に関する。
請求項１０に係る発明は、乳癌、前立腺癌、膵癌、胃癌、肺癌、大腸癌（結腸癌、直腸癌、肛門癌）、食道癌、十二指腸癌、頭頚部癌（舌癌、咽頭癌）、脳腫瘍、神経鞘腫、非小細胞肺癌、肺小細胞癌、肝臓癌、腎臓癌、胆管癌、子宮癌（子宮体癌、子宮頸癌）、卵巣癌、膀胱癌、皮膚癌、血管腫、悪性リンパ腫、悪性黒色腫、甲状腺癌、骨腫瘍、血管腫、血管線維腫、網膜肉腫、陰茎癌、小児固形癌、カポジ肉腫、ＡＩＤＳに起因するカポジ肉腫、上顎洞腫瘍、線維性組織球腫、平滑筋肉腫、横紋筋肉腫のうちいずれかの治療或は予防のために用いられることを特徴とする請求項２，４，６又は８に記載のアポトーシス誘導剤に関する。
請求項１１に係る発明は、請求項１，３，５，７又は９記載の癌細胞増殖阻害剤を含有することを特徴とする医薬品に関する。
請求項１２に係る発明は、請求項２，４，６，８又は１０記載のアポトーシス誘導剤を含有することを特徴とする医薬品に関する。

The invention according to claim 9 is breast cancer, prostate cancer, pancreatic cancer, stomach cancer, lung cancer, colon cancer (colon cancer, rectal cancer, anal cancer), esophageal cancer, duodenal cancer, head and neck cancer (tongue cancer, pharyngeal cancer), brain tumor. , Schwannoma, non-small cell lung cancer, small cell lung cancer, liver cancer, kidney cancer, bile duct cancer, uterine cancer (uterine body cancer, cervical cancer), ovarian cancer, bladder cancer, skin cancer, hemangioma, malignant lymphoma , Malignant melanoma, thyroid cancer, bone tumor, hemangioma, angiofibroma, retinal sarcoma, penile cancer, childhood solid cancer, Kaposi sarcoma, Kaposi sarcoma caused by AIDS, maxillary sinus tumor, fibrous histiocytoma, smooth muscle The cancer cell proliferation inhibitor according to claim 1, 3, 5, or 7, which is used for the treatment or prevention of any one of tumor and rhabdomyosarcoma.
The invention according to claim 10 includes breast cancer, prostate cancer, pancreatic cancer, stomach cancer, lung cancer, colon cancer (colon cancer, rectal cancer, anal cancer), esophageal cancer, duodenal cancer, head and neck cancer (tongue cancer, pharyngeal cancer), brain tumor. , Schwannoma, non-small cell lung cancer, small cell lung cancer, liver cancer, kidney cancer, bile duct cancer, uterine cancer (uterine body cancer, cervical cancer), ovarian cancer, bladder cancer, skin cancer, hemangioma, malignant lymphoma , Malignant melanoma, thyroid cancer, bone tumor, hemangioma, angiofibroma, retinal sarcoma, penile cancer, childhood solid cancer, Kaposi sarcoma, Kaposi sarcoma caused by AIDS, maxillary sinus tumor, fibrous histiocytoma, smooth muscle The apoptosis-inducing agent according to claim 2, 4, 6, or 8, which is used for treatment or prevention of any one of tumor and rhabdomyosarcoma.
The invention according to claim 11 relates to a pharmaceutical comprising the cancer cell proliferation inhibitor according to claim 1, 3, 5, 7, or 9.
The invention according to claim 12 relates to a pharmaceutical comprising the apoptosis inducer according to claim 2, 4, 6, 8 or 10.

The cancer cell growth inhibitor comprising the alkoxytetrahydrofuran derivative of the present invention has a concentration-dependent growth inhibitory activity against cancer cells. Moreover, the apoptosis inducer comprising the alkoxytetrahydrofuran derivative of the present invention induces apoptosis in cancer cells.
Since the drug containing the cancer cell proliferation inhibitor and / or apoptosis inducer of the present invention has a growth inhibitory activity and / or apoptosis induce activity against cancer cells, it is effective for treating diseases such as malignant tumors and cancers. is there.

  The cancer cell proliferation inhibitor and apoptosis inducer of the present invention comprise an alkoxytetrahydrofuran derivative (shown by the following formula (Chemical Formula 9)).

（式中Ｒ_１及びＲ_２は、同一又は異なって、水素原子、芳香族置換基又は炭素数１〜１０のアルキル基を表す。Ｒ_３は炭素数１〜１０のアルキル基、或は次式（化１０）又は（化１１）で表される置換基である。）

(In the formula, R ₁ and R ₂ are the same or different and each represents a hydrogen atom, an aromatic substituent or an alkyl group having 1 to 10 carbon atoms. R ₃ is an alkyl group having 1 to 10 carbon atoms, or (It is a substituent represented by (Chemical Formula 10) or (Chemical Formula 11).)

（式中ｎは１〜１０の整数、ｍは１〜１０の整数であり、式中Ｘは、‐ＣＯＯ‐（エステル）、‐Ｏ‐、‐ＮＲ_４‐（Ｒ_４は、水素原子、炭素数１〜６のアルキル基又は芳香族置換基を表す）又は‐Ｓ‐を表す。）

(Wherein n is an integer of 1 to 10, m is an integer of 1 to 10, and X is —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon Represents an alkyl group of 1 to 6 or an aromatic substituent) or -S-).

（式中ｎは４〜１０の整数であり、式中Ｘは、‐ＣＨ_２‐、‐ＣＯＯ‐（エステル）、‐Ｏ‐、‐ＮＲ_４‐（Ｒ_４は、水素原子、炭素数１〜６のアルキル基又は芳香族置換基を表す）又は‐Ｓ‐を表す。）

(In the formula, n is an integer of 4 to 10, wherein X is —CH ₂ —, —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon number 1 to 6 represents an alkyl group or an aromatic substituent) or -S-.

The aromatic substituents of R ₁ and R _{2 in} the above formula (Chemical Formula 9) may be monocyclic or condensed rings, and may be hydrocarbon rings or heterocyclic aromatic substituents. . Further, the number of carbon atoms contained in the aromatic substituent is preferably 6 to 10, and the aromatic substituent may have a substitutable substituent. Examples of the substituent include an alkyl group and a hydroxyl group. Groups, alkoxyl groups, ester groups, nitro groups and amino groups.
A benzylidenealkoxytetrahydrofuran derivative in which R ₁ is a phenyl group, R ₂ is a hydrogen atom, R ₁ is a hydrogen atom, and R ₂ is a phenyl group in the formula (Formula 9) is more preferably used.
The alkyl group represented by R ₁ and R ₂ in the formula (Chemical Formula 9) has 1 to 10 carbon atoms, and particularly preferably 2 to 8 carbon atoms.

In the formula (Formula 9), R ₃ is an alkyl group or a substituent represented by the formula (Formula 10) or (Formula 11). The alkyl group has 1 to 10 carbon atoms, and particularly preferably 2 to 8 carbon atoms. Moreover, n in the said Formula (Formula 10) is an integer 1-10, and the integer 2-8 is especially preferable. M in the formula (Formula 10) is an integer of 1 to 10, and an integer of 2 to 8 is particularly preferable. Moreover, n in the said Formula (Formula 11) is an integer 4-10, and the integer 5-8 is especially preferable.
In the present invention, among the alkoxytetrahydrofuran derivatives, a benzylidenetetrahydrofuran derivative in which R ₁ in the formula (Chemical Formula 9) is a phenyl group, R ₂ is a hydrogen atom, or R ₁ is a hydrogen atom, and R ₂ is a phenyl group. Preferably used.
Further, among the benzylidenetetrahydrofuran derivatives, (Z) -4-benzylidene-2-n-butoxytetrahydrofuran (C ₁₅ H ₂₀ O ₂ ) in which R ₃ in the formula (8) is a butyl group is preferably used. It is done. The following formula (Chemical formula 12) shows (Z) -4-benzylidene-2-n-butoxytetrahydrofuran.

  The method for obtaining the alkoxytetrahydrofuran derivative according to the present invention is not particularly limited. For example, it can be obtained by chemical synthesis, and the synthesis method is not limited.

  The cancer cell proliferation inhibitor and apoptosis inducer of the present invention are cancer (eg, breast cancer, prostate cancer, pancreatic cancer, gastric cancer, lung cancer, colon cancer (colon cancer, rectal cancer, anal cancer), esophageal cancer, duodenal cancer, head and neck. Cancer (tongue cancer, pharyngeal cancer), brain tumor, schwannoma, non-small cell lung cancer, small cell lung cancer, liver cancer, kidney cancer, bile duct cancer, uterine cancer (uterine body cancer, cervical cancer), ovarian cancer, bladder Cancer, skin cancer, hemangioma, malignant lymphoma, melanoma, thyroid cancer, bone tumor, hemangioma, hemangiofibroma, retinal sarcoma, penile cancer, childhood solid cancer, Kaposi sarcoma, Kaposi sarcoma due to AIDS, maxillary sinus Tumor, fibrous histiocytoma, leiomyosarcoma, rhabdomyosarcoma, etc.) and malignant tumors such as leukemia.

  The effective amount of the cancer cell growth inhibitor and apoptosis inducer of the present invention to cancer cells is adjusted according to the activity of the solvent used together, the administration route, the severity of the pathological condition to be treated, the patient's condition and medical history. The However, it is desirable to start with a lower level of dose than is required to achieve the desired therapeutic effect and gradually increase the dose until the desired effect is achieved. The specific dose level in each patient can be determined by various factors such as body weight, general health, diet, time, route of administration, concomitant medications, or severity of the disease being treated. desirable.

  The cancer cell proliferation inhibitor and apoptosis inducer of the present invention contain additives such as pharmaceutically acceptable carriers, and can be made into pharmaceutical products. Specific additives include various organic or inorganic carrier substances that are commonly used as pharmaceutical materials. Excipients, lubricants, binders, disintegrants, thickeners in solid preparations; solvents in liquid preparations, dispersion It is added as an agent, solubilizer, suspending agent, isotonic agent, buffering agent, soothing agent and the like. If necessary, additives such as preservatives, antioxidants, colorants, sweeteners and the like can be used.

The pharmaceutical containing the cancer cell proliferation inhibitor and apoptosis inducer of the present invention is administered by systemic routes such as intravenous infusion or injection in addition to oral administration of tablets, capsules, granules, drinks and the like, Alternatively, parenteral administration such as intramuscular injection, enteral administration, and local administration may be used. Specific examples of the pharmaceutical product of the present invention are shown below.
(1) Tablets, powders, granules, and chaos cells: The cancer cell growth inhibitor and apoptosis inducer of the present invention are compression-molded by adding, for example, an excipient, a disintegrant, a binder, or a lubricant, Then, if necessary, it can be produced by coating for taste masking, enteric or sustained purposes.
(2) Injection: The cancer cell growth inhibitor and apoptosis inducer of the present invention, for example, as an aqueous injection together with a dispersing agent, preservative, tonicity agent and the like, or vegetable oils such as olive oil, sesame oil, cottonseed oil and corn oil It can be produced by dissolving, suspending or emulsifying in propylene glycol or the like and molding it as an oily injection.
(3) External preparation: manufactured by making the cancer cell proliferation inhibitor and apoptosis inducer of the present invention into a solid, semi-solid or liquid composition. For example, the solid composition is produced by using the pharmaceutical composition as it is, or by adding and mixing excipients, thickeners and the like into a powder form.

The liquid pharmaceutical is almost the same as in the case of injections, and is produced by making an oily or aqueous suspension. The semi-solid pharmaceutical is preferably an aqueous or oily gel or ointment. In addition, any of these compositions may contain a buffering agent, a preservative, and the like.
(4) Suppository: produced by making the cancer cell growth inhibitor and apoptosis inducer of the present invention into an oily or aqueous solid, semi-solid or liquid composition. Examples of the oily base used in such a composition include glycerides of higher fatty acids (for example, cacao butter, vitepsols), intermediate fatty acids (for example, miglyols), or vegetable oils (for example, sesame oil, soybean oil, Cottonseed oil, etc.). Examples of the aqueous gel base include natural gums, cellulose derivatives, vinyl polymers, and acrylic acid polymers.

  The cancer cell proliferation inhibitor and apoptosis inducer of the present invention may be used in combination with other therapeutic agents including known antitumor agents conventionally used in the art. Suitable examples of such conventionally used antitumor agents include adriamycin, cisplatin, colchicine, CCNU (Lomastine), BCNU (Carmustine), actinomycin D, 5-fluorouracil, thiotepa, cytosine arabinoside, cyclophosphine One kind or a mixture of two or more kinds such as famide and mitomycin C may be mentioned.

<Synthesis of alkoxytetrahydrofuran derivative [(Z) -4-benzylidene-2-n-butoxytetrahydrofuran] according to the present invention>
Palladium acetate (22.5 mg, 0.1 mmol), copper acetate (18.2 mg, 0.1 mmol), and catechol (0.1 mmol, 11.0 mg) were added to a 25 mL branch flask containing a stir bar. A balloon containing oxygen was attached to the flask, and the inside of the flask was purged with oxygen. Acetonitrile (1.0 mL) was added to the flask using a syringe and the solution was stirred at room temperature for about 30 minutes. Thereafter, n-butyl vinyl ether (283 mg, 4.0 mmol) was added using a syringe under an oxygen atmosphere, followed by trans-cinnamyl alcohol (134 mg, 1.0 mmol). After the solution was stirred at room temperature for 3 hours, the reaction mixture was filtered using a florisil column (florisil, 30 g: column, 20 mm × 20.5 mm: developing solvent, ethyl acetate). When the solvent was removed, (Z) -4-benzylidene-2-n-butoxytetrahydrofuran (212.0 mg, 91.2%), an alkoxytetrahydrofuran derivative according to the present invention, was obtained.
The chemical reaction formula of the above synthesis is represented by the following formula (Formula 13).

The following are the results of instrumental analysis for identifying the (Z) -4-benzylidene-2-n-butoxytetrahydrofuran. FIG. 1 shows a ¹ H NMR chart, and FIG. 2 shows a ¹³ C NMR chart.
bp (bulb-to-bulb) 173-175 ° C. (5 mmHg); ¹ H NMR (400 MHz, CDCl ₃ ) δ 0.91 (t, J = 6.60 Hz, 3H, CH ₃ ), 1.36 (sex, J = 6.60 Hz, 2 H, CH ₂ CH ₃ ), 1.56 (qui, J = 6.60 Hz, 2 H, CH ₂ CH ₂ CH ₃ ), 2.71 (d, J = 16.48, 1 H, C * HCH ₂ ), 2.93 (dd, J = 16.48, 5.15 Hz, 1 H, C * HCH ₂ ), 3.44 (dt, J = 9.51, 6.60) Hz, 1 H, —OCH ₂ CH ₂ ), 3.71 (dt, J = 9.51, 6.60 Hz, 1H, —OCH ₂ CH ₂ ), 4.68 (s, 2 H, —OCH ₂ C =), 5.21 (d, J = 5.15 Hz, 1 H, C * H), 6.42 (s, 1H, Ph—CH), 7.13 (d, J = 7.33 Hz , 2 H, Ph), 7.20 (t, J = 7.33 Hz, 1 H, Ph), 7.34 (d, J = 7.33 Hz, 2 H, Ph); ¹³ C {1H} ( 100 MHz, CDCl ₃ ) δ 13.9 (CH ₃ ), 19.3 (CH ₂ CH ₃ ), 31.7 (—OCH ₂ CH ₂ ), 41.1 (C * HCH ₂ ), 67.0 (− _{OCH 2 CH 2), 67.9 (} OCH 2 C =), 102.2 (C * H), 121.5 (Ph-CH), 126.4 (Ph), 127.8 (Ph), 128. 4 (Ph), 137.4 (Ph), 139.1 (CH = C); GCMS m / e 232 (M ⁺ ); Calcd for C ₁₅ H ₂₀ O ₂ : C, 77.55; H, 8. 68. Found: C, 77.53; H, 8.62.

<Test Example 1> Cancer Cell Growth Inhibition Test In this test, U937 cells (human mononuclear leukemia cells) were used.
First, U937 cells were seeded at 3 × 10 ⁴ cells / well / ml in a 12-well plate, and (Z) -4-benzylidene-2-n-butoxytetrahydrofuran obtained in the above synthesis was dissolved in dimethyl sulfoxide (DMSO). The product was added to a concentration of 100 μM / well and 200 μM / well and incubated for 72 hours.
Next, 30 μl of medium was collected from each hole into an Eppendorf tube, trypan blue solution was added, and the number of viable cells was counted using a hemocytometer. Similarly, at the start of the experiment, the number of cells was similarly counted for the control (no addition) and those with only DMSO added, and the results are shown in Table 1 and FIG.
In FIG. 3, the cell growth rate is shown with the control as 100%. The cell growth rate was obtained by adding the cancer cell growth inhibitor of the present invention at a concentration of DMSO and 100 μM and 200 μM and dividing the number of cells after 72 hours by the number of control (no addition) cells after 72 hours. 100 times.

  As is apparent from Table 1 and FIG. 3, it can be seen that the cancer cell growth inhibitor of the present invention has growth inhibitory activity on cancer cells in a concentration-dependent manner.

<Test Example 2> Apoptosis-inducing activity test
1) Test method a) Detection of DNA ladder U937 cells were seeded at 2 × 10 ⁵ cells / well / 500 μl in a 24-well plate. Next, 20 μm of the apoptosis-inducing agent of the present invention (5 μl (final concentration: 200 μM)) was added, and U937 cells incubated for 30 minutes, 1 hour, 3 hours, and 6 hours were used. Detected.

First, DNA was extracted by the following protocol using the following reagents and enzymes.
(reagent)
Cell Lysis Buffer
1M tris-HCl pH 7.4 0.1ml
0.5 MEDTA pH 8.0 0.2ml
10% Triton X-100 0.5ml
Sterile distilled water 9.2ml
Total 10.0ml
TE Buffer
1 Mtris-HCl pH 7.4 1.0 ml
0.5 MEDTA pH 8.0 0.2ml
Sterile distilled water 98.8ml
Total 100.0ml
(enzyme)
RNase A 20mg / ml
Proteinase K 20mg / ml

(Protocol)
U937 cells treated according to the present invention were collected in a 1.5 ml Eppendorf tube, centrifuged at 2000 rpm for 3 minutes, washed with PBS, and further centrifuged at 2000 rpm for 3 minutes. After removing the supernatant in the Eppendorf tube, 100 μl of Cell Lysis Buffer was added and gently pipetted to lyse the cells. The mixture was placed at 4 ° C. for 15 minutes and centrifuged at 15,000 rpm for 20 minutes. The entire amount of the obtained supernatant was transferred to a new tube, and 2 μl of RNase (20 mg / ml) was added. This was incubated at 37 ° C. for 1 hour. Furthermore, 2 μl of Proteinase K (20 mg / ml) was added, incubated at 37 ° C. for 1 hour, 20 μl of 5M NaCl and 120 μl of isopropanol were added, and the mixture was left at −20 ° C. overnight. Next, the mixture was centrifuged at 15,000 rpm for 15 minutes, and the supernatant was discarded, and further centrifuged at 15,000 rpm for 5 minutes to completely remove the supernatant, and the pellet was dissolved in 10 μl of TE Buffer. .
The resulting solution was subjected to 2% agarose gel electrophoresis. After electrophoresis, it was stained with SYBR Green I (Molecular Probes) for 30 minutes, and a gel image was captured with FluoroImager (Filter: 530) (Mokecular Dynamics). The resulting DNA ladder is shown in FIG.

  As shown in FIG. 4, the U937 cells added with the apoptosis inducer of the present invention form a DNA ladder as compared with the U937 cells not added with the apoptosis inducer of the present invention. It can be seen that the inducer induces apoptosis.

b) Chromatin aggregation U937 cells were seeded at 2 × 10 ⁵ cells / well / 500 μl in a 24-well plate. Next, 20 μm of apoptosis-inducing agent of the present invention (5 μl (final concentration: 200 μM)) was added, and U937 cells incubated for 30 minutes, 1 hour and 6 hours were used to confirm chromatin aggregation by the following methods.

  First, as described above, U937 cells treated according to the present invention were washed with PBS. Next, it was fixed with 1% glutaraldehyde for 2 hours or more, glutaraldehyde was removed by centrifugation, and further washed with PBS. The pellet was dissolved in 20 μl of PBS, 10 μl was taken, mixed with 2 μl of 1 mM Hoechst 33342 solution (Calbiochem), and dropped onto a slide glass. The slide glass was observed with a fluorescence microscope U region (Ultra violet: excitation light 330-380 nm, observation light 420 nm or more). The obtained fluorescence micrograph is shown in FIG.

  As shown in the fluorescence micrograph in the control of FIG. 5, the fluorescence micrograph (FIG. 5) of the U937 cells treated according to the present invention, compared with the case where the apoptosis inducer of the present invention was not added ((d) in FIG. 5). In (5), (a) to (c)) are aggregated with chromatin fragments and a granular image is seen, which indicates that apoptosis has occurred.

  From the results of FIGS. 4 and 5, it was found that the apoptosis inducer of the present invention is a substance that induces apoptosis in cancer cells, that is, has an apoptosis-inducing action.

(Z)-4-a diagram showing a ¹ H NMR spectrum of the benzylidene -2-n-butoxy tetrahydrofuran. It is a figure which shows the ¹³ C NMR spectrum of (Z) -4-benzylidene-2-n-butoxytetrahydrofuran. It is a graph which shows the U937 cell growth rate after the cancer cell growth inhibitor addition of this invention in the cell growth inhibition test of Test Example 1. 3 is an electrophoretic image showing a DNA ladder in an apoptosis-inducing activity test of Test Example 2. 4 is a fluorescence micrograph showing a chromatin aggregated image in an apoptosis-inducing activity test of Test Example 3. FIG.

Claims (12)

A cancer cell growth inhibitor comprising an alkoxytetrahydrofuran derivative represented by the following formula (Formula 1):

(In the formula, R ₁ and R ₂ are the same or different and each represents a hydrogen atom, an aromatic substituent or an alkyl group having 1 to 10 carbon atoms. R ₃ is an alkyl group having 1 to 10 carbon atoms, or (It is a substituent represented by (Chemical Formula 2) or (Chemical Formula 3).)

(Wherein n is an integer of 1 to 10, m is an integer of 1 to 10, and X is —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon Represents an alkyl group of 1 to 6 or an aromatic substituent) or -S-).

(In the formula, n is an integer of 4 to 10, wherein X is —CH ₂ —, —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon number 1 to 6 represents an alkyl group or an aromatic substituent) or -S-. An apoptosis inducer comprising an alkoxytetrahydrofuran derivative represented by the following formula (Formula 4):

(In the formula, R ₁ and R ₂ are the same or different and each represents a hydrogen atom, an aromatic substituent or an alkyl group having 1 to 10 carbon atoms. R ₃ is an alkyl group having 1 to 10 carbon atoms, or (It is a substituent represented by (Chemical Formula 5) or (Chemical Formula 6).)

(Wherein n is an integer of 1 to 10, m is an integer of 1 to 10, and X is —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon Represents an alkyl group of 1 to 6 or an aromatic substituent) or -S-).

(In the formula, n is an integer of 4 to 10, wherein X is —CH ₂ —, —COO— (ester), —O—, —NR ₄ — (R ₄ is a hydrogen atom, carbon number 1 to 6 represents an alkyl group or an aromatic substituent) or -S-. The alkoxytetrahydrofuran derivative is a benzylidenealkoxytetrahydrofuran derivative in which R ₁ in the formula (Chemical Formula 1) is a phenyl group, R ₂ is a hydrogen atom, R ₁ is a hydrogen atom, and R ₂ is a phenyl group. The cancer cell proliferation inhibitor according to claim 1. The alkoxy tetrahydrofuran derivatives, the Formula (4) R ₁ is a phenyl group in, R ₂ is a hydrogen atom, or (reduction 4) R ₁ is a hydrogen atom in, benzylidene alkoxy tetrahydrofuran R ₂ is a phenyl group 3. The apoptosis inducer according to claim 2, which is a derivative. 4. The cancer cell growth inhibitor according to claim 1, wherein the alkoxytetrahydrofuran derivative is a butoxytetrahydrofuran derivative in which R ₃ in the formula (Formula 1) is a butyl group. The apoptosis-inducing agent according to claim 2 or 4, wherein the alkoxytetrahydrofuran derivative is a butoxytetrahydrofuran derivative in which R ₃ in the formula (Formula 4) is a butyl group. 6. The cancer cell growth inhibitor according to claim 5, wherein the alkoxytetrahydrofuran derivative is (Z) -4-benzylidene-2-n-butoxytetrahydrofuran represented by the following formula (Formula 7).

The apoptosis-inducing agent according to claim 6, wherein the alkoxytetrahydrofuran derivative is (Z) -4-benzylidene-2-n-butoxytetrahydrofuran represented by the following formula (Formula 8).

  Breast cancer, prostate cancer, pancreatic cancer, stomach cancer, lung cancer, colon cancer (colon cancer, rectal cancer, anal cancer), esophageal cancer, duodenal cancer, head and neck cancer (tongue cancer, pharyngeal cancer), brain tumor, schwannoma, non-small cell Lung cancer, small cell lung cancer, liver cancer, kidney cancer, bile duct cancer, uterine cancer (uterine body cancer, cervical cancer), ovarian cancer, bladder cancer, skin cancer, hemangioma, malignant lymphoma, malignant melanoma, thyroid cancer, Bone tumor, hemangioma, angiofibroma, retinal sarcoma, penile cancer, childhood solid cancer, Kaposi sarcoma, Kaposi sarcoma caused by AIDS, maxillary sinus tumor, fibrous histiocytoma, leiomyosarcoma, rhabdomyosarcoma The cancer cell proliferation inhibitor according to claim 1, 3, 5, or 7, which is used for any treatment or prevention.   Breast cancer, prostate cancer, pancreatic cancer, stomach cancer, lung cancer, colon cancer (colon cancer, rectal cancer, anal cancer), esophageal cancer, duodenal cancer, head and neck cancer (tongue cancer, pharyngeal cancer), brain tumor, schwannoma, non-small cell Lung cancer, small cell lung cancer, liver cancer, kidney cancer, bile duct cancer, uterine cancer (uterine body cancer, cervical cancer), ovarian cancer, bladder cancer, skin cancer, hemangioma, malignant lymphoma, malignant melanoma, thyroid cancer, Bone tumor, hemangioma, angiofibroma, retinal sarcoma, penile cancer, childhood solid cancer, Kaposi sarcoma, Kaposi sarcoma caused by AIDS, maxillary sinus tumor, fibrous histiocytoma, leiomyosarcoma, rhabdomyosarcoma The apoptosis inducer according to claim 2, 4, 6 or 8, which is used for any treatment or prevention.   A pharmaceutical comprising the cancer cell proliferation inhibitor according to claim 1, 3, 5, 7 or 9. A pharmaceutical comprising the apoptosis-inducing agent according to claim 2, 4, 6, 8 or 10.

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