JP5105166B2 - Method for producing polyether - Google Patents

Description

  The present invention relates to a polyether having pendant carboxymethyl groups introduced in the side chain, a drug delivery carrier and a drug complex using the polyether. Moreover, it is related with the manufacturing method of the polyether for making a pharmaceutical compound reach | attain a target site.

For example, drugs (pharmaceutical compounds) such as antitumor drugs, when administered systemically by intravenous or oral administration routes, migrate to a specific tumor site and inhibit or suppress the growth of cancer cells. To exert a therapeutic effect. However, systemically administered antitumor drugs are taken from the blood into the reticulo-organs such as the liver, spleen, and bone marrow, or are excreted in the urine via the kidneys, resulting in a decrease in the blood concentration, leading to the tumor site. May not be sufficiently transferred, and in some cases, toxicity may develop.
In addition, when normal anti-tumor drugs are administered systemically alone, the anti-tumor drugs are evenly distributed to various cells, tissues, and organs throughout the body due to low migration selectivity to the tumor site (tumor selectivity). Therefore, it has been pointed out that it also acts as a cytotoxin on normal cells and tissues and causes side effects such as vomiting, fever, and hair loss at a very high rate. Therefore, development of a technique for efficiently and selectively transferring an antitumor drug to a tumor site, and a technique for performing controlled drug release from the reached tumor site have been demanded.
As one of such technologies, for example, an antitumor drug is bound to a water-soluble polymer such as a polysaccharide, the disappearance of the antitumor drug from the blood is delayed, and the directivity to cancer tissue is increased. A way to increase it has been proposed. For example, there is an example in which a complex in which adriamycin is bound to various polysaccharides is disclosed (Patent Document 1). There is also an example in which a complex in which an anthracycline antitumor drug is bound to various polysaccharide derivatives including carboxymethyldextran is disclosed (Non-patent Document 1). Furthermore, there is an example in which a complex in which a drug is bound to a carboxyalkyldextran polyalcohol via a peptide is disclosed (Patent Document 2). Although these naturally-derived polysaccharides have a functional group such as a hydroxyl group and are excellent in drug carrying ability, there is a problem that uniform quality cannot be easily obtained because they are natural products.
On the other hand, as an example using a synthetic polymer as a drug delivery carrier, an example in which an angiogenesis inhibitor (TNP-470) is bound to hydroxypropyl methacrylate (HPMA) via a linker has been reported (Non-Patent Document). 2). In addition, there is an example in which a complex in which paclitaxel is bound to polyethylene glycol (PEG) is disclosed (Non-patent Document 3). Although these synthetic polymers are characterized in that they are not decomposed in vivo, they have low hurdles in terms of safety, such as being administered to a living body as a plasma expander.
US Patent No. 5,688,931 JP 2002-30002 JP Biol. Pharm. Bull., 24, 535-543 (2001) Nature Medicine, 10, 255-261 (2004) J. Med. Chem., 39, 424-431 (1996)

  Conventional polyethylene glycol has not been sufficiently satisfactory in terms of drug carrying capacity and drug delivery. Accordingly, it is possible to provide a drug delivery carrier using a polyether that has excellent drug carrying ability and can transfer the drug to a target site, and a drug complex using this carrier, and to simplify the use of such a polyether. It would be desirable to provide a manufacturing method.

（式中、Ｒ^１は−ＯＲ^２または−ＮＨＲ^３で示される基を表し、Ｒ^２およびＲ^３は水素、無機塩基あるいは有機塩基またはC_１〜C_１５の炭化水素を含む基、または薬理作用を有する基を表す。ここで同一の共重合体内でR¹、Ｒ^２またはＲ^３は同一であっても異なっていてもよい。各構造単位のモル比は、（III）／（（III）＋（IV））＝０．００１〜０．５であり、構造単位（III）および（IV）のモル％はそれぞれ（III）=０．１〜９９．９および（IV）=０．１〜９９．９である。ここで、式（III）および式（IV）を構造単位として含むポリエーテルは、ポリエチレングリコール（PEG）換算重量平均分子量が１，０００〜１００，０００である。）
（２）下式（V）および式（VI）を構造単位として含むポリエーテルに対し、薬理作用を有する基を、薬理作用を有する基自身が有するアミノ基を介して、あるいは、アミノ基を有するリンカーを介して結合させることにより得られる式（VII）を含みかつ（V）、（VI）を構造単位として含む前記（１）に記載のポリエーテル。

（式中、Ｒ^４は水素、無機塩基あるいは有機塩基を示し、ＮＨ−Ｒ^５は自身が有するアミノ基を介し、あるいは自身が結合したリンカーが有するアミノ基を介し結合した薬理作用を有する基を示す。各構造単位のモル比は、（VII）／（（V）＋（VI）＋（VII））＝０．００１〜０．５である。各構造単位（V）、（VI）および（VII）のモル％はそれぞれ（V）＝０〜９９．９、（VI）=０．１〜９９．８および（VII）=０．１〜９９．８である。ここで、式（V）、（VI）および式（VII）を構造単位として含むポリエーテルは、PEG換算重量平均分子量が１，０００〜１００，０００である。）
（３）ポリエチレングリコール（ＰＥＧ）換算重量平均分子量／ＰＥＧ換算数平均分子量（Ｍｗ／Ｍｎ）が１．１〜２．７である前記（１）または（2）に記載のポリエーテル。
（４）薬理作用を有する基がリンカーと結合した基である前記（1）〜（３）のいずれかに記載のポリエーテル。
（５）リンカーがアミノ酸もしくはペプチドである前記（１）〜（４）のいずれかに記載のポリエーテル。
（６）リンカーがＧｌｙ，Ａｌａ，Ｌｅｕ，Ｉｌｅ，Ｐｈｅで示されるアミノ酸から選択または組み合わせて用いられる前記（１）〜（５）のいずれかに記載のポリエーテル。
（７）リンカーがＧｌｙ−Ｇｌｙ，Ａｌａ−ＧＬｙ，Ｇｌｙ−Ａｌａ，Ｌｅｕ−Ｇｌｙ，Ｇｌｙ−Ｌｅｕ，Ｉｌｅ−Ｇｌｙ，Ｇｌｙ−Ｉｌｅ，Ｐｈｅ−Ｇｌｙ，Ｇｌｙ−Ｐｈｅ，Ｇｌｙ−Ｇｌｙ−Ｇｌｙ，Ｇｌｙ−Ｐｈｅ−Ｇｌｙ，Ｐｈｅ−Ｇｌｙ−Ｇｌｙ，Ｇｌｙ−Ｇｌｙ−Ｐｈｅ，Ｇｌｙ−Ｇｌｙ−Ｐｈｅ−Ｇｌｙから選択される前記（１）〜（６）のいずれかに記載のポリエーテル。
（８）薬理作用を有する基が分子中に官能基としてアミノ基、またはカルボキシル基、または水酸基を有する前記（１）〜（７）のいずれかに記載のポリエーテル。
（９）薬理作用を有する基が抗悪性腫瘍薬、抗炎症薬、抗リウマチ薬、酵素阻害薬または核酸である前記（１）〜（８）のいずれかに記載のポリエーテル。
（１０）抗悪性腫瘍薬がアルキル化薬、代謝拮抗薬、抗腫瘍性抗生物質、微小管阻害薬、ホルモン類似薬、白金製剤、トポイソメラーゼ阻害薬、生物製剤（サイトカイン）、分子標的治療薬、非特異的免疫賦活薬およびその誘導体から選択される前記（９）に記載のポリエーテル。
（１１）アルキル化薬がシクロフォスファミドなどのマスタード薬、ニムスチンなどのニトロソウレア類およびその誘導体から選択される前記（１０）に記載のポリエーテル。
（１２）代謝拮抗薬が葉酸代謝拮抗薬であるメトトレキセート、ピリミジン代謝拮抗薬である５−フルオロウラシル、プリン代謝拮抗薬の６−ＭＰおよびその誘導体から選択される前記（１０）に記載のポリエーテル。
（１３）抗腫瘍性抗生物質がアンスラサイクリン系のドキソルビシン、ダウノルビシン、マイトマイシンＣおよびそれらの誘導体から選択される前記（１０）に記載のポリエーテル。
（１４）微小管阻害薬がタキサン系のパクリタキセル、ドセタキセルおよびその誘導体から選択される前記（１０）に記載のポリエーテル。
（１５）ホルモン類似薬がタモキシフェンおよびその誘導体から選択される前記（１０）に記載のポリエーテル。
（１６）白金製剤がシスプラチン、カルボプラチン、オキサリプラチンおよびその誘導体から選択される前記（１０）に記載のポリエーテル。
（１７）トポイソメラーゼ阻害薬がトポイソメラーゼＩ阻害薬のイリノテカン、ノギテカンおよびその誘導体から選択される前記（１０）に記載のポリエーテル。
（１８）生物製剤がインターフェロンおよびその誘導体から選択される前記（１０）に記載のポリエーテル。
（１９）分子標的治療薬がイマチニブ、ゲフィチニブ、リツキシマブおよびその誘導体から選択される前記（１０）に記載のポリエーテル。
（２０）抗炎症薬が副腎皮質ステロイド薬、非ステロイド性抗炎症薬およびその誘導体から選択される前記（９）に記載のポリエーテル。
（２１）副腎皮質ステロイド薬がヒドロコルチゾン、コルチゾン、プレドニゾロン、メチルプレドニゾロン、デキサメタゾン、ベタメタゾンおよびその誘導体から選択される前記（２０）に記載のポリエーテル。
（２２）非ステロイド性抗炎症薬がサリチル酸系のアスピリン、アントラニル系のメフェナム酸、プロピオン酸系のイブプロフェン、ロキソプロフェン、アリール酢酸系のジクロフェナク、インドメタシンおよびその誘導体から選択される前記（２０）に記載のポリエーテル。
（２３）抗リウマチ薬が免疫調節薬、免疫抑制薬、生物学的製剤およびその誘導体から選択される前記（９）に記載のポリエーテル。
（２４）免疫調節薬が金チオリンゴ酸ナトリウム、ペニシラミン、ロベンザリットおよびその誘導体から選択される前記（２３）に記載のポリエーテル。
（２５）免疫抑制薬がミゾリビン、メトトレキサート、レフルノミド、タクロリムスおよびその誘導体から選択される前記（２３）に記載のポリエーテル。
（２６）生物学的製剤がインフリキシマブ、エタネルセプトおよびその誘導体から選択される前記（２３）に記載のポリエーテル。
（２７）酵素阻害薬がファスジルおよびその誘導体から選択される前記（９）に記載のポリエーテル。
（２８）前記（１）〜（２７）のいずれかに記載のポリエーテルを含む製剤。
（２９）前記（１）におけるＲ^２が水素又は無機塩基、または有機塩基であるポリエーテルを用いた生体内組織への薬物、遺伝子、あるいは生体内イメージング用造影剤に対する送達用担体。
（３０）前記（１）におけるＲ^２が水素又は無機塩基、または有機塩基であるポリエーテルを用いた薬剤、ペプチド、核酸または蛋白を担持あるいは固定化するための担体。
（３１）前記（１）〜（２７）のいずれかに記載のポリエーテルを架橋してなる構造体を疾患部位に投与することによる治療方法。
（３２）薬物送達製剤の製造における前記（１）〜（２７）のいずれかに記載のポリエーテルの使用。
（３３）式（I）および式（II）で表される化合物をランダム共重合して、式（III）および式（IV）で表される繰り返し単位を有するポリエーテルを製造する方法。

（式中、Ｒ^１は−ＯＲ^２または−ＮＨＲ^３で示される基を表し、Ｒ^２およびＲ^３は水素、無機塩基あるいは有機塩基またはC_１〜C_１５の炭化水素を含む基、または−ＮＨＲ^３は薬理作用を有する基を表す。ここで同一の共重合体内でR¹、Ｒ^２またはＲ^３は同一であっても異なっていてもよい。各構造単位のモル比は、（III）／（（III）＋（IV））＝０．００１〜０．５であり、構造単位（III）および（IV）のモル％はそれぞれ（III）=０．１〜９９．９および（IV）=０．１〜９９．９である。ここで、式（III）および式（IV）を構造単位として含むポリエーテルは、ポリエチレングリコール（PEG）換算重量平均分子量が１，０００〜１００，０００である。）
（３４）重合に用いる触媒が塩基およびルイス酸から選択される前記（３３）に記載のポリエーテルを製造する方法。
（３５）重合に用いる触媒が水酸化カリウム、水酸化ナトリウム、水酸化セシウムやナトリウムメトキシド、ナトリウムエトキシド、ナトリムプロポキシド、ナトリウムｔ−ブトキシド、カリウムプロポキシド、カリウムｔ−ブトキシド、カリウム−ｔ−２−メチル−２−ブトシキドおよび三フッ化ホウ素エーテル錯体、トリアルキルアルミニウム、トリエチルアルミニウム、トリフェニルアルミニウム、トリブチルアルミニウムから選択され、単独もしくは混合して使用される前記（３３）に記載のポリエーテルを製造する方法。
（３６）前記（１）〜（２７）のいずれかに記載のポリエーテルを静脈内より生体内に投与することによる治療方法。
（３７）前記（１）〜（２７）のいずれかに記載のポリエーテルを疾患部位に投与することによる治療方法。 As a result of intensive studies in view of the above problems, the present invention succeeded in solving this problem by using, as a carrier for drug delivery, a polyether comprising a specific structural unit having a carboxymethyl group introduced in a pendant form. That is, the present invention has the following configuration.
(1) A polyether having repeating units represented by formula (III) and formula (IV), obtained by copolymerizing the compounds represented by formula (I) and formula (II).

(In the formula, R ¹ represents a group represented by —OR ² or —NHR ³ , and R ² and R ³ represent hydrogen, an inorganic base, an organic base, a group containing a C _{1 to} C ₁₅ hydrocarbon, or a pharmacological action. Wherein R ¹ , R ² or R ³ may be the same or different in the same copolymer, and the molar ratio of each structural unit is (III) / ((III) + (IV)) = 0.001 to 0.5, and the molar percentages of the structural units (III) and (IV) are (III) = 0.1 to 99.9 and (IV) = 0.1, respectively. Here, the polyether containing formula (III) and formula (IV) as structural units has a weight average molecular weight in terms of polyethylene glycol (PEG) of 1,000 to 100,000.)
(2) A polyether having the following formula (V) and formula (VI) as a structural unit, having a pharmacological group via an amino group of the pharmacological group itself or having an amino group The polyether according to the above (1), which contains the formula (VII) obtained by bonding via a linker and contains (V) and (VI) as structural units.

(Wherein R ⁴ represents hydrogen, an inorganic base or an organic base, and NH—R ⁵ represents a group having a pharmacological action bonded via an amino group possessed by itself or via an amino group possessed by a linker bound thereto. The molar ratio of each structural unit is (VII) / ((V) + (VI) + (VII)) = 0.001 to 0.5 Each structural unit (V), (VI) and ( The mole percentages of VII) are (V) = 0 to 99.9, (VI) = 0.1 to 99.8 and (VII) = 0.1 to 99.8, respectively, wherein the formula (V) , (VI) and a polyether containing formula (VII) as structural units have a weight average molecular weight in terms of PEG of 1,000 to 100,000.)
(3) Polyether as described in said (1) or (2) whose polyethylene glycol (PEG) conversion weight average molecular weight / PEG conversion number average molecular weight (Mw / Mn) is 1.1-2.7.
(4) The polyether according to any one of (1) to (3), wherein the group having a pharmacological action is a group bonded to a linker.
(5) The polyether according to any one of (1) to (4), wherein the linker is an amino acid or a peptide.
(6) The polyether according to any one of (1) to (5), wherein the linker is selected or used in combination from amino acids represented by Gly, Ala, Leu, Ile, and Phe.
(7) Linker is Gly-Gly, Ala-GLy, Gly-Ala, Leu-Gly, Gly-Leu, Ile-Gly, Gly-Ile, Phe-Gly, Gly-Phe, Gly-Gly-Gly, Gly-Phe -Polyether in any one of said (1)-(6) selected from Gly, Phe-Gly-Gly, Gly-Gly-Phe, Gly-Gly-Phe-Gly.
(8) The polyether according to any one of (1) to (7), wherein the group having a pharmacological action has an amino group, a carboxyl group, or a hydroxyl group as a functional group in the molecule.
(9) The polyether according to any one of (1) to (8), wherein the group having a pharmacological action is an antineoplastic agent, an anti-inflammatory agent, an anti-rheumatic agent, an enzyme inhibitor or a nucleic acid.
(10) Antineoplastic drugs are alkylating drugs, antimetabolites, antitumor antibiotics, microtubule inhibitors, hormone analogs, platinum preparations, topoisomerase inhibitors, biologics (cytokines), molecular targeted therapeutics, non The polyether according to (9), which is selected from a specific immunostimulant and a derivative thereof.
(11) The polyether according to (10), wherein the alkylating drug is selected from mustard drugs such as cyclophosphamide, nitrosoureas such as nimustine, and derivatives thereof.
(12) The polyether according to (10), wherein the antimetabolite is selected from methotrexate, which is an antifolate, 5-fluorouracil, which is a pyrimidine antimetabolite, 6-MP, an antipurine antimetabolite, and derivatives thereof.
(13) The polyether according to (10), wherein the antitumor antibiotic is selected from anthracycline doxorubicin, daunorubicin, mitomycin C and derivatives thereof.
(14) The polyether according to (10), wherein the microtubule inhibitor is selected from taxane-based paclitaxel, docetaxel and derivatives thereof.
(15) The polyether according to (10), wherein the hormone analog is selected from tamoxifen and derivatives thereof.
(16) The polyether according to (10), wherein the platinum preparation is selected from cisplatin, carboplatin, oxaliplatin and derivatives thereof.
(17) The polyether according to the above (10), wherein the topoisomerase inhibitor is selected from the topoisomerase I inhibitor irinotecan, nogitane and derivatives thereof.
(18) The polyether according to (10), wherein the biologic is selected from interferon and derivatives thereof.
(19) The polyether according to the above (10), wherein the molecular target therapeutic agent is selected from imatinib, gefitinib, rituximab and derivatives thereof.
(20) The polyether according to (9) above, wherein the anti-inflammatory drug is selected from corticosteroids, non-steroidal anti-inflammatory drugs and derivatives thereof.
(21) The polyether according to (20), wherein the corticosteroid is selected from hydrocortisone, cortisone, prednisolone, methylprednisolone, dexamethasone, betamethasone and derivatives thereof.
(22) The non-steroidal anti-inflammatory drug is selected from salicylic acid aspirin, anthranilic mefenamic acid, propionic acid ibuprofen, loxoprofen, arylacetic acid diclofenac, indomethacin and derivatives thereof Polyether.
(23) The polyether according to (9), wherein the anti-rheumatic drug is selected from immunomodulators, immunosuppressants, biological preparations and derivatives thereof.
(24) The polyether according to the above (23), wherein the immunomodulator is selected from gold sodium thiomalate, penicillamine, lobenzalit and derivatives thereof.
(25) The polyether according to (23), wherein the immunosuppressive drug is selected from mizoribine, methotrexate, leflunomide, tacrolimus and derivatives thereof.
(26) The polyether according to the above (23), wherein the biologic is selected from infliximab, etanercept and derivatives thereof.
(27) The polyether according to (9), wherein the enzyme inhibitor is selected from fasudil and derivatives thereof.
(28) A preparation comprising the polyether according to any one of (1) to (27).
(29) A carrier for delivery to a drug, gene, or in vivo imaging contrast agent for in vivo tissue using a polyether in which R ² in (1) is hydrogen, an inorganic base, or an organic base.
(30) A carrier for supporting or immobilizing a drug, peptide, nucleic acid or protein using a polyether in which R ² in (1) is hydrogen, an inorganic base, or an organic base.
(31) A treatment method comprising administering a structure obtained by crosslinking the polyether according to any one of (1) to (27) to a disease site.
(32) Use of the polyether according to any one of (1) to (27) in the production of a drug delivery preparation.
(33) A method for producing a polyether having repeating units represented by formula (III) and formula (IV) by random copolymerization of compounds represented by formula (I) and formula (II).

(Wherein R ¹ represents a group represented by —OR ² or —NHR ³ , R ² and R ³ represent hydrogen, an inorganic base or an organic base, a group containing a C _{1 to} C ₁₅ hydrocarbon, or —NHR; ³ represents a group having a pharmacological action, wherein R ¹ , R ² or R ³ may be the same or different in the same copolymer, and the molar ratio of each structural unit is (III) / ((III) + (IV)) = 0.001 to 0.5, and the molar percentages of the structural units (III) and (IV) are (III) = 0.1-99.9 and (IV) = Here, the polyether containing formula (III) and formula (IV) as a structural unit has a weight average molecular weight in terms of polyethylene glycol (PEG) of 1,000 to 100,000. .)
(34) The method for producing a polyether according to the above (33), wherein the catalyst used for the polymerization is selected from a base and a Lewis acid.
(35) The catalyst used for polymerization is potassium hydroxide, sodium hydroxide, cesium hydroxide, sodium methoxide, sodium ethoxide, sodium propoxide, sodium t-butoxide, potassium propoxide, potassium t-butoxide, potassium-t- The polyether according to (33), which is selected from 2-methyl-2-butoxide and boron trifluoride ether complex, trialkylaluminum, triethylaluminum, triphenylaluminum, tributylaluminum, and used alone or in combination. How to manufacture.
(36) A method of treatment by administering the polyether according to any one of (1) to (27) into a living body from intravenously.
(37) A method of treatment by administering the polyether according to any one of (1) to (27) to a disease site.

(1) The polyether of the present invention is a polyether in which an arbitrary number of carboxyl groups are introduced in a pendant manner in the side chain, as in the case of conventional polyethylene glycol (hereinafter sometimes referred to as PEG). Not only the drug is supported on both ends of the main chain of the polymer, but also compounds and drugs can be easily bonded to the side chain of the polymer. As a result, the amount of compound or drug introduced can be controlled, and as a result, a complex or preparation having a high introduction amount or high drug content can be prepared. In the case of administration to a living body as a preparation, the total dose can be reduced, and the burden on the patient, such as a decrease in administration time and a decrease in the administration liquid volume, is reduced.
(2) Since the polyether of the present invention has a pendant side chain carboxyl group, it is possible to easily carry a drug in a high yield by utilizing an amide bond.
(3) The polyether of the present invention has a higher solubility in water by converting the pendant side chain carboxyl group into an inorganic salt or an organic salt, and is excellent in, for example, solubilization of poorly water-soluble compounds. It is particularly useful as a drug delivery carrier for injections.
(4) The polyether of the present invention is highly safe for living bodies, particularly when used as a drug delivery carrier. This is due to the negative charge due to a moderate amount of carboxyl groups introduced into the side chain, which may reduce the distribution to metabolic organs such as liver, spleen, and bone marrow, and side effect organs, causing organ damage and cytotoxicity. It is because it is thought that is reduced. Therefore, when the polyether of the present invention is used for medical purposes, it can be said to be a safe material with very little interaction with a living body.
(5) Especially when the polyether of the present invention is used as a drug delivery carrier, it is possible to selectively accumulate the loaded drug in a target tumor organ or inflammatory organ.

  The polyether of the present invention comprises a random copolymer polyether and contains structural units represented by the following formulas (III) and (IV). The molecular weight of the polyether of the present invention is 1,000 to 200,000 in terms of weight average molecular weight in terms of PEG. If the weight average molecular weight is less than 1,000, the polyether is very soluble in water, but the excretion rate from the body is also extremely high. If it exceeds 200,000, the polyether is soluble in water, but the viscosity increases. . Considering safety to the living body as a drug delivery carrier, the weight average molecular weight of the polyether is preferably 1,000 to 150,000, more preferably 1,000 to 100,000. Furthermore, if it judges from the retentivity improvement in blood and the ease of renal excretion, 1,000-70,000 are preferable. This is because when the molecular weight of plasma albumin in the living body is lower than the molecular weight (about 67,000), the excretion of the polyether outside the body proceeds rapidly. Therefore, in particular, it is more preferable that the molecular weight is about 10,000 to 70,000 with high safety to the living body while the excretion outside the body is controlled. Further, in order to further improve the blood retention when administered in vivo, 30,000 to 70,000 is more preferable.

  Here, the PEG-converted weight average molecular weight is obtained by converting PEG into a standard sample (TSK standard POLY (ETHYLENE OXIDE), manufactured by TOSOH Co., Ltd.) when the molecular weight is measured by GPC. Refers to the obtained molecular weight. Since PEG is soluble in water-soluble solvents and organic solvents, it is used as a molecular weight standard substance when measuring the molecular weight of hydrophilic polymers and hydrophobic polymers.

  The polyether of the present invention is characterized by being produced by random copolymerization in order to randomly disperse the hydrophilic portion and the hydrophobic portion in the same molecule. When the polyether of the present invention is produced by block copolymerization, a polyether acting as a surfactant may be generated, and when such a polyether is administered in vivo, a hemolytic action may be exhibited. This is because there is a risk of adversely affecting the living body.

  The molar ratio of each structural unit of the polyether of the present invention is (III) / ((III) + (IV)) = 0.001 to 0.5, and the mol% of the structural units (III) and (IV). Are (III) = 0.1-99.9 and (IV) = 0.1-99.9, respectively.

  The molecular weight distribution represented by the weight average molecular weight / number average molecular weight (Mw / Mn) in the drug conjugate and drug delivery carrier of the present invention is the drug conjugate and drug delivery in order to sufficiently exhibit the intended pharmacological effect. Since the uniformity of the quality of the carrier is important, the molecular weight distribution is preferably 1.2 to 2.7. In order to ensure the quality of the drug complex and the drug delivery carrier, it is preferably 1.2 to 2.2, more preferably 1.1 to 1.8, and further 1.0 to 1 Still more preferably, .5.

In the general formulas (III) and (IV), R ¹ represents a group represented by —OR ² or —NHR ³ , and R ² and R ³ are each an inorganic or organic base such as hydrogen or sodium, or C ₁ -C _It represents a group containing ₁₅ hydrocarbons or a group having a pharmacological action. Here, R ¹ , R ² or R ³ may be the same or different in the same copolymer.

  Examples of the general formula (III) include the general formulas (V) and (VII). The monomer raw materials for producing the polyethers represented by the general formulas (III), (V), and (VII) Examples include glycidyl ether derivatives (general formula (VIII)) in which the hydroxyl groups of ethylene oxide and glycidol are substituted with organic substituents, and glycidyl ether derivatives (general formula (IX)) into which amino acids or peptides are introduced. .

（式中、Ｒ^４は水素、ナトリウム等の無機塩基あるいは有機塩基を示す。）

（式中、Ｒ^５は自身が有するアミノ基を介し、あるいは自身が結合したリンカーが有するアミノ基を介し結合した薬理作用を有する基を示す。

（式中、Ｒ^６はC_１〜C_１５の炭化水素を含む基を表し、同一の共重合体においてＲ^６は同一であっても異なっていてもよい。）

（式中、Ｒ^７はC_１〜C_２０の脂肪族炭化水素を有する基、又はC_６〜C_２０の芳香族炭化水素誘導体を有する基を示し、同一の共重合体内でＲ^７は同一であっても異なっていても良い。）

(Wherein R ⁴ represents an inorganic base or an organic base such as hydrogen or sodium)

(In the formula, R ⁵ represents a group having a pharmacological action bonded via an amino group contained in itself or via an amino group contained in a linker bonded thereto.

(Wherein R ⁶ represents a group containing a C _{1 to} C ₁₅ hydrocarbon, and R ⁶ in the same copolymer may be the same or different.)

(Wherein R ⁷ represents a group having a C _{1 to} C ₂₀ aliphatic hydrocarbon or a group having a C _{6 to} C ₂₀ aromatic hydrocarbon derivative, and R ⁷ is the same in the same copolymer) It may or may not be.)

Here, the glycidyl ether derivative represented by the general formula (VIII) can be produced, for example, according to the following scheme, but is not limited to the following method. For example, when R ⁶ in the general formula (VIII) is C _3, a synthesis example for obtaining 2,3-epoxypropyl epoxypropoxyacetate using hydroxyacetic acid and epichlorohydrin as starting materials is disclosed in 1960 (Patent Document 3). ). Also in the case R ⁶ is an ethyl group in the general formula (VIII), Synthesis Example of glycidol as a raw material has been reported (Non-Patent Document 4). In this invention, it manufactured according to the scheme as shown below. That is, allyl alcohol (A) as a raw material is reacted with chloroacetic acid in the presence of a base such as sodium hydride, and then extracted with an organic solvent under acidic conditions to obtain a carboxymethyl group-introduced product (B) and dicyclohexyl. An ester (C) is obtained by reacting an alcohol in the presence of a condensing agent such as carbodiimide. Thereafter, the olefin moiety is oxidized with an oxidizing agent such as 3-chloroperbenzoic acid or peracetic acid to obtain the intended glycidyl ether derivative (D) represented by the above general formula (VIII).
US 2925426 Petrochemistry, 36, 874-881 (1987)

For example, as the glycidyl ether derivative, R ⁶ is an aliphatic hydrocarbon such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, t-butyl, pentyl, hexyl, heptyl, octyl, or phenyl, p- Aromatic hydrocarbons and derivatives such as nitrophenyl, benzyl, p-methoxybenzyl, glycidyl, and succinic anhydride bonded by a nitrogen atom can be mentioned. With regard to these R ⁶ , after the ring-opening polymerization of the epoxy is completed, it is possible to form a carboxylic acid by removing it under the deprotection conditions used in usual organic synthesis such as acid treatment, base treatment, hydrogenation and the like. is there. Alternatively, as R ⁶ , for example, when a group called an active ester used in peptide synthesis such as p-nitrophenyl or succinic anhydride is introduced, a compound having an amino group with respect to R ⁶ without passing through a carboxylic acid It is also possible to form an amide bond (peptide bond) by directly reacting.

  A monomer raw material into which an amino acid or peptide is introduced as a glycidyl ether derivative as shown by the following formula (IX) is also a monomer raw material of the present invention.

（式中、Ｒ^７はC_１〜C_２０の脂肪族炭化水素を有する基、C_６〜C_２０の芳香族炭化水素誘導体を有する基を示し、同一の共重合体内でＲ^７は同一であっても異なっていても良い。）

(In the formula, R ⁷ represents a group having a C _{1 to} C ₂₀ aliphatic hydrocarbon and a group having a C _{6 to} C ₂₀ aromatic hydrocarbon derivative, and R ⁷ was the same in the same copolymer. Or different.)

  Here, the glycidyl ether derivative represented by the general formula (IX) can be produced according to the following scheme. For example, the above carboxymethyl group-introduced product (B) is used as a raw material, and an amide compound (E) is obtained by reacting amines in the presence of a condensing agent such as dicyclohexylcarbodiimide. Thereafter, the olefin moiety is oxidized with an oxidizing agent such as 3-chloroperbenzoic acid to obtain the desired glycidyl ether derivative (F) represented by the general formula (IX).

  For example, as the above-mentioned amines, aminocarboxylic acids such as amino acids, peptides, and beta-alanine whose side chains and carboxyl groups are protected as necessary may be mentioned. Here, specific examples of the formula (IX) include glycidyl ether derivatives into which glycine benzyl ester represented by the following formula (X) is introduced.

The greatest feature in the production of the polyether of the present invention is that a carboxymethyl group is introduced as a monomer, so that a carboxyl group is directly introduced into the polymer in a one-step polymerization process. Furthermore, it is also characterized in that the amount of carboxymethyl group introduced can be controlled by adjusting the amount of monomer used with respect to ethylene oxide.
Thus, the polyether of the present invention obtained by random polymerization using a monomer having a carboxyl group is superior in various respects as compared with PEG which is a conventional technique. That is, since the highly reactive carboxymethyl group is introduced into the side chain, the polyether of the present invention is excellent in water solubility, hydration ability, water retention ability, and functional group introduction ability. Further, by effectively using a side chain carboxyl group whose introduction amount is controlled, it becomes possible to bind a drug or the like to obtain a drug complex having a desired drug content. More specifically, in an argon atmosphere, a solvent such as THF or toluene, or in some cases, a water-organic solvent mixed system using DMF or DMSO, or a condensing agent used in peptide synthesis in an aqueous solution. In the presence, it is possible to bind a drug, peptide, biological imaging contrast agent, nucleic acid and protein to the carboxymethyl group of the polyether of the present invention.

  The functional group-introduced polyether thus obtained is very excellent as a medical material used for medical practice such as a drug delivery medicine and a protein purification carrier. In addition, when a carboxymethyl group is converted into an active ester, in the presence of a condensing agent, in an argon atmosphere, in an organic solvent such as THF, ether, DMF, DMSO, or toluene, or in a water-containing organic solvent, or in some cases Drugs, peptides, nucleic acids and proteins can be bound in a buffer solution such as PBS or in an aqueous solution. Such functional group-introduced polyethers are excellent as medical materials used for medical practice such as drug delivery drugs and protein purification carriers.

As a conventional technique, PEG is widely used for pharmaceutical use, but since there is no hydroxyl group that can be used for introducing a functional group only at both ends, the drug-carrying ability is extremely low and it is not suitable as a carrier for drug delivery. . In addition, when attempting to deliver a drug to a target organ by improving the retention of the drug in the blood, there arises a problem that the molecular weight of PEG must be increased to prevent renal excretion. For example, when the PEG molecular weight is 60,000 and the drug is 500, only two molecules of drug are introduced into the hydroxyl groups at both ends in one molecule of PEG, so the amount of drug introduced is 1.6% in weight%. Furthermore, for example, in the prodrug formulation PROTHECAN in which camptothecin is introduced with a PEG molecular weight of 40,000, it has been reported that the drug content is 1.7% (w / w) (Non-patent Document 5).
TRENDS in Biotechnology, 24 (1), 39-47 (2006)

  In these cases, for example, when trying to administer a drug formulated using PEG as a carrier to a patient, it is necessary to administer a PEG preparation 60 times the required drug amount, and the burden on the patient during drug administration is extremely large. On the other hand, since the polyether of the present invention can introduce a large number of carboxyl groups into the side chain, the amount of drug introduced can be increased freely. For example, the amount of drug introduced to the complex can be increased by 10%. The above is also possible. Therefore, the total dose of the drug conjugate of the present invention can be greatly reduced as compared with the conventional PEG conjugate dose, and the burden upon administration to the patient can be greatly reduced.

  About the composition of the polyether of the present invention, the type of raw material used in the reaction, the weight of the raw material, the type of polymerization initiator, the addition amount of the polymerization initiator, the presence or absence of a reaction solvent, the selection of the reaction solvent, the reaction temperature, the reaction time, Polyethers having various composition ratios are produced by adjusting the raw material concentration, the polymerization initiator concentration, the reaction atmosphere, the reaction pressure, the stirring state, the stirring speed, and the like. For example, various glycidyl ethers together with ethylene oxide in a reaction solvent such as toluene, hexane, heptane, 2-methoxyethyl ether, ethylene glycol dimethyl ether, ethylene glycol diethyl ether, or without solvent, for example, boron trifluoride ether complex, Metal polymerization initiators such as alkylaluminum, triethylaluminum, Lewis acid such as triphenylaluminum and tributylaluminum, or potassium hydroxide, sodium hydroxide, cesium hydroxide, sodium methoxide, sodium ethoxide, sodium propoxide, sodium t- A polymerization initiator such as butoxide, potassium propoxide, potassium t-butoxide, potassium-t-2-methyl-2-butoxide and the like is used alone or in combination, Cold, can be obtained by ring-opening polymerization under normal pressure or elevated pressure under heating, if room temperature or required.

  After polymerization, an acid such as hydrochloric acid or sulfuric acid, a base such as sodium hydroxide, potassium hydroxide or calcium hydroxide, or an operation such as hydrogenation in the presence of a catalyst such as palladium carbon may be used to produce the target carboxylic acid. Can do. Alternatively, in the case of an active ester type, there is no need for post-polymerization acid, base and hydrogenation treatment, and it is possible to bind a drug, peptide, nucleic acid and protein as they are.

  As described above, the weight average molecular weight of the polyether in the present invention is important to be a molecular weight that can be easily excreted from the living body to the body from the viewpoint of safety. For example, PEG in GPC analysis In order to achieve the converted weight average molecular weight of 1,000 to 100,000, for example, the starting material may be purified and the high molecular weight fraction or the low molecular weight fraction may be separated and removed after the reaction, if necessary. For example, specific methods for molecular weight fractionation include size exclusion chromatography (SEC), chromatography such as GPC, ultrafiltration including UF module, ultracentrifugation, organic solvents such as ether and ethanol, etc. It can be carried out by various techniques such as precipitation fraction used.

  As described above, when the polyether of the present invention is used as, for example, a drug complex and a drug delivery carrier, an optimal molecular weight is selected, and a side chain carboxymethyl group is introduced in an optimal amount according to the purpose of use. Is possible.

In the drug complex included in the present invention, a drug (pharmaceutical compound) is bound to a drug delivery carrier directly or via a linker. That is, in the general formula (I) representing the structural unit of the polyether of the present invention, R ² , R ³ and R ⁵ are groups having a pharmacological action (including a group having a pharmacological action bonded to a linker). It corresponds to. The drug is not particularly limited as long as it is a pharmaceutical compound having a carboxyl group or a functional group capable of reacting with the linker. For example, a metal atom, amino group, carboxyl group, hydroxy group, mercapto group, carboxamide group, urethane group, etc. Pharmaceutical compounds having are preferred. Among these, a pharmaceutical compound having an amino group can be easily bonded to the polyether of the present invention with or without a linker. Moreover, about such a pharmaceutical compound which does not have an amino group, it becomes possible to couple | bond with a support | carrier by introduce | transducing a functional group with a linker. For example, paclitaxel, dexamethasone, camptothecin and the like can be easily bonded to the polyether of the present invention by converting a hydroxyl group present in a drug into an amino group by a linker.

  Examples of the pharmaceutical compound include antineoplastic drugs, anti-inflammatory drugs, anti-rheumatic drugs, enzyme inhibitors or nucleic acids.

  Antineoplastic agents include alkylating drugs, antimetabolites, antitumor antibiotics, microtubule inhibitors, hormone analogs, platinum preparations, topoisomerase inhibitors, biologics (cytokines), molecular targeted therapeutics, non-specific Specific immunostimulants and derivatives thereof. Examples of the alkylating drug include mustard drugs such as cyclophosphamide and nitrosoureas such as nimustine. Antimetabolites include folate antimetabolite methotrexate, pyrimidine antimetabolite 5-FU, and purine antimetabolite 6-MP. Antitumor antibiotics include anthracycline doxorubicin and daunorubicin, as well as mitomycin C. Examples of microtubule inhibitors include paclitaxel and docetaxel. Examples of hormone analogs include tamoxifen. Examples of platinum preparations include cisplatin, carboplatin, and oxaliplatin. Examples of topoisomerase inhibitors include topoisomerase I inhibitors irinotecan, nogitecan, etoposide and the like. Examples of biologics include interferon. Examples of molecular targeted therapeutic agents include imatinib, gefitinib, rituximab and the like.

  Anti-inflammatory drugs include corticosteroids and non-steroidal anti-inflammatory drugs. Examples of corticosteroids include hydrocortisone, cortisone, prednisolone, methylprednisolone, dexamethasone, betamethasone and the like. Non-steroidal anti-inflammatory drugs include salicylic acid-based aspirin, anthranilic mefenamic acid, propionic acid-based ibuprofen, loxoprofen, arylacetic acid-based diclofenac, and indomethacin. In addition, these nonsteroidal anti-inflammatory drugs are classified according to COX selectivity.

  Anti-rheumatic drugs include immunomodulators, immunosuppressants, biologics and the like. Examples of immunomodulators include sodium gold thiomalate, penicillamine, and lobenzalit. Examples of immunosuppressants include mizoribine, methotrexate, leflunomide, tacrolimus and the like. Biologics include infliximab, etanercept and the like.

  In order to bind the drug delivery carrier and the drug to produce the drug complex of the present invention, for example, a means for forming a complex forming coordination bond, an ester bond, a thioester bond, an acid amide bond, an acid anhydride, etc. It is manufactured by bonding with. For example, in the case of a complex formation reaction, the reaction may usually be performed in an aqueous solution. In the case of an ester bond forming reaction, the reaction is carried out in the presence of a dehydrating condensing agent such as various carbodiimides, and an activator such as dimethylaminopyridine may be added as necessary. In the case of an acid amide bond forming reaction, the reaction may be performed in the presence of a dehydrating condensing agent such as various carbodiimides.

  In the present invention, the drug delivery carrier and the drug may be bound via a linker in some cases. The linker of the present invention includes a so-called spacer that adjusts the distance from the carrier. Examples of the linker include 1 amino acid, 2 to 4 amino acids with peptide bonds, and the like. More specifically, a residue of one amino acid (meaning a residue obtained by removing one hydrogen atom and one hydroxyl group from the amino group and carboxyl group of an amino acid, respectively), or peptide-bonded 2 to 4 Residue of oligopeptide consisting of one amino acid (meaning a residue obtained by removing one hydrogen atom and one hydroxyl group from the N-terminal amino group and the C-terminal carboxyl group, respectively) . Preferred linkers are 1 amino acid residue or oligopeptide residue consisting of 2 to 4 amino acids. Such a difference in steric hindrance of amino acids makes it possible to adjust the release rate of the drug bound to the carrier.

  The type of amino acid constituting the linker is not particularly limited. For example, L- or D-amino acid, preferably L-amino acid can be used. In addition to α-amino acid, β-alanine, ε-aminocaproic acid, γ- Aminobutyric acid or the like may be used. Such amino acids other than α-amino acids are preferably arranged at positions close to the drug delivery carrier. The binding direction of the intervening group is not particularly limited, but generally, the N-terminus of the intervening group is bound to the carboxyl group of the carboxyalkyl polyether by an acid amide bond, and the C-terminus of the intervening group is bound to the amino group or hydroxyl group of the pharmaceutical compound. Can be combined. In addition, for example, if a lysine residue is included as a structural unit of a peptide linker and an α-amino group and an ε-amino group of the lysine residue are bonded to a carboxyl group of another amino acid, respectively, an acid amide bond, Since the terminal is the N terminal, it becomes possible to bond the carboxyl group of the pharmaceutical compound. Furthermore, one or two or more diamine compounds or dicarboxylic acid residues (for example, diamine residues such as ethylenediamine or dicarboxylic acid residues such as succinic acid) are included as structural units in the linker, An intervening group having N terminals at both ends and a linker having C ends at both ends may be used.

  The amino acid sequence of the linker is not particularly limited, but when a linker containing a dipeptide structure is used, hydrolysis is promoted at a tumor site or an inflammation site considered to be rich in peptidases, and a pharmaceutical compound is released at a high concentration at that site. . The partial structure formed by combining a linker containing the dipeptide and a pharmaceutical compound is the partial structure of the drug complex of the present invention. For example, when a concentration-dependent antitumor drug (for example, doxorubicin) is used as a residue of a pharmaceutical compound, it is particularly preferable to use a linker containing a dipeptide residue as a partial peptide sequence. In addition, when using a time-dependent antitumor drug (for example, paclitaxel) that requires a duration of action at a certain concentration or higher as a residue of a pharmaceutical compound, a high antitumor can be obtained by using the above linker. The effect may be achieved. In general, the linker is not limited to the above-mentioned linkers, and it is necessary to select a preferable linker from the viewpoint of the mechanism of action of antitumor drugs, pharmacokinetics and characteristics of toxicity, release of antitumor drugs in the body, etc. There is.

  Specific examples of the amino acid sequence of the linker include Gly, Ala, Leu, Ile, Phe, Gly-Gly, Ala-GLy, Gly-Ala, Leu-Gly, Gly-Leu, Ile-Gly, Gly-Ile, A sequence selected from Phe-Gly, Gly-Phe, Gly-Gly-Gly, Gly-Phe-Gly, Phe-Gly-Gly, Gly-Gly-Phe, and Gly-Gly-Phe-Gly is desirable.

  Regarding a drug conjugate that binds via a linker, the drug conjugate of the present invention can be produced by linking a linker that is bonded to a residue of a pharmaceutical compound to the carboxyl group of the polyether of the present invention. it can. The bond between the linker and the carboxyl group of the polyether of the present invention can be generally formed by bonding the N-terminal amino group of the linker and the carboxyl group of the polyether of the present invention by an acid amide bond. But the coupling | bonding of the linker and the carboxyl group of the polyether of this invention is not limited to the above thing, The other chemical bond and the coupling | bonding using 1 or 2 or more linkers may be sufficient. For example, an acid anhydride may be formed by the C-terminal carboxyl group of the linker and the carboxyl group of the polyether of the present invention, and a diamine compound such as ethylenediamine may be used as a linker to convert each carboxyl group to each amino acid of the diamine. An acid amide bond may be bonded to the group. When the N-terminal amino group of the linker and the carboxyl group of the polyether of the present invention are bonded by an acid amide bond, a conventional dehydration condensing agent used for the synthesis of peptides, for example, N, N'-dicyclohexylcarbodiimide (DCC) ), N, N′-dicycloalkylcarbodiimides, carbodiimide derivatives such as 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide (EDAPC), 1-ethoxycarbonyl-2-ethoxy-1, 2-dihydroxyquinoline (EEDQ) or the like can be used. Moreover, you may react by the active ester method, the acid halide method, etc. In the case of the active ester type of the polyether of the present invention, it is not necessary to use a dehydrating condensing agent.

Specific examples of the group having a pharmacological action are shown below.
When a drug compound represented by the following formula (X) is bonded with glycine (H ₂ N—CH ₂ —COOH) as a linker, a compound having an amino group represented by the following formula (Y) is obtained. . This can be used by bonding to a polyether as a group having pharmacological activity.

（X）

（Y）

(X)

(Y)

  Further, in the compound represented by the following formula, the group having pharmacological activity itself has an imino group, and is bonded to the polyether via the imino group.

  Furthermore, a compound represented by the following formula in which glycine is bound to the above drug as a linker can also be used as a group exhibiting pharmacological action.

  The amount of the pharmaceutical compound residue to be introduced into the polyether of the present invention is not particularly limited, but from the viewpoints of the physicochemical properties of the pharmaceutical compound residue and the pharmacokinetics, efficacy and toxicity of the drug complex of the present invention. It should be selected as appropriate. In general, a range of about 0.1 to 40% by weight, preferably 1 to 20% by weight, more preferably 2 to 20% by weight, and particularly preferably about 3 to 15% by weight can be selected. The ratio of the pharmaceutical compound residue introduced into the polyether can be easily determined by, for example, ultraviolet absorption analysis, fluorescence analysis, absorbance analysis, or the like.

The drug conjugate of the present invention can be polymer-modified using a functional group of a pharmaceutical compound (for example, a functional group of a pharmaceutical compound such as an antitumor agent or anti-inflammatory agent). Can deliver a pharmaceutical compound to a desired target organ. Therefore, the desired pharmaceutical activity of the pharmaceutical compound can be specifically expressed locally such as a tumor site or an inflammatory site. Furthermore, due to the negative charge due to the appropriate amount of carboxyl group introduced into the side chain, the distribution to the liver, spleen, and bone marrow, which are metabolic / side effect (toxicity) organs, is reduced and the side effect (toxicity) expression of the pharmaceutical compound is reduced. Therefore, it has the characteristic that the toxicity which a pharmaceutical compound itself has can be reduced.
The polyether of the present invention is very useful as a carrier for drug delivery because it has extremely excellent blood retention and accumulation at a tumor / inflammatory site, and is a drug complex in which a carrier comprising this polyether and a drug are bound. Can be delivered to the target organ very effectively and exert the action of the drug. In addition, since the drug carrier is structurally different from conventional PEG and has a large number of carboxyl groups in the side chain, it has an excellent ability to carry a pharmaceutical compound. Furthermore, since the drug complex of the present invention has particularly improved solubility in water, it is particularly useful when used as an injection in a poorly water-soluble drug or the like. In particular, the drug conjugate of the present invention has a particularly excellent ability to carry a pharmaceutical compound per mole compared to the case where PEG is used as a drug delivery carrier, so that the burden on the administered patient can be reduced. .

  The medicament containing the drug conjugate of the present invention can be filled into a vial or the like in the form of a freeze-dried product or the like, and clinically used as a parenteral preparation such as a dissolution preparation for injection or infusion preparation. Although provided, the pharmaceutical dosage form of the present invention is not limited to the above embodiment. For the preparation of the above preparation, for example, additives for preparation available in the art such as a solubilizer, pH adjuster, stabilizer and the like can be used.

The dosage of the pharmaceutical agent of the present invention is not particularly limited. Usually, however, the dosage of the pharmaceutical compound constituting the pharmaceutical compound residue, the amount of the pharmaceutical compound residue introduced into the drug complex of the present invention, The decision should be made taking into account the condition and type of disease. For example, when the drug conjugate of the present invention into which camptothecin is introduced at a rate of about 5% by weight is administered parenterally, it is generally about 0.1 to 100 mg, preferably about 0.1 to 1 m ² of body surface area per day. It is preferable to administer once in the range of 1 to 30 mg and repeat every 3 to 4 weeks.

The present invention relates to a method for treating a drug complex by administering it into a living body from a vein or at a disease site. Furthermore, the present invention also relates to a method for treating a diseased site by administering a structure obtained by crosslinking the polyether, which is a drug complex, to a disease site.

Hereinafter, the present invention will be described more specifically with reference to examples. However, the scope of the present invention is not limited to the following examples.
The conditions for GPC in this example are as follows.
Column: G4000PWXL (manufactured by Tosoh Corporation)
Mobile phase: 20% acetonitrile in 50 mM lithium chloride Flow rate: 0.8 ml / min
Column temperature: 40 ° C
Pump: L-6200 (manufactured by Hitachi, Ltd.)
Detector: L-3300 (RI: differential refractometer, manufactured by Hitachi, Ltd.)
The calibration curve for calculating the molecular weight and molecular weight distribution of the compound is standard polyethylene oxide (made by TOSOH, weight average molecular weight 2.4 × 10 ⁴ , 5.00 × 10 ⁴ , 1
・ 7 × 10 ⁴ , 14.0 × 10 ⁴ )
In the following examples, the copolymer (number) may be simply referred to as a compound (same number).

[Example 1]
A solution of allyl alcohol (20.4 ml) in THF (THF 200 ml) was added to a suspension of sodium hydride (13.2 g) in tetrahydrofuran (300 ml) under ice cooling, and the mixture was stirred for 30 minutes. Thereafter, a solution of bromoacetic acid (34.7 g) in tetrahydrofuran (150 ml) was added dropwise over 90 minutes. The reaction was stirred at room temperature for 5 days. The solvent was distilled off under reduced pressure, water was added and the mixture was washed with ethyl acetate. Thereafter, the pH was adjusted to 2 with 5N hydrochloric acid and the aqueous phase was extracted with ethyl acetate. The organic layer was dried over magnesium sulfate to obtain 26.7 g of a crude product (1).
This crude product (1) (15 g) was dissolved in 200 ml of methylene chloride, and ethanol (11.4 ml), N-methylmorpholine (NMM; 14.2 ml) and dimethylaminopyridine (DMAP; 1.5 g) were added. After cooling with ice, a methylene chloride solution of water-soluble carbodiimide (EDC; 24.2 g) was added dropwise over 30 minutes. The reaction was stirred at room temperature for 2 days. The reaction solution was washed with a saturated aqueous sodium bicarbonate solution and then with a saturated saline solution and dried over magnesium sulfate to obtain 26.3 g of a crude product. Purification by silica gel chromatography (silica gel 60, 240 g, hexane / ether 5: 1 from Merck) gave the title compound (2) (11.2 g).
Compound (2) (11.0 g) was dissolved in 150 ml of methylene chloride, and a solution of 3-chloroperbenzoic acid (MCPBA; 15.9 g) in methylene chloride was added dropwise over 90 minutes under ice cooling. The mixture was stirred at room temperature for 2 days, washed with 2N aqueous sodium hydroxide solution and then with saturated brine, dried over magnesium sulfate, and the solvent was distilled off under reduced pressure to obtain a crude product (19.3 g). Purification by silica gel chromatography (silica gel 60, 140 g, hexane / ether 2: 1, manufactured by Merck) gave the title compound (3) (11.1 g).

The compound analysis data regarding the title compound (3) is shown below. As the NMR standard, 3-trimethylsilylpropionic acid-2,2,3,3-d4 sodium salt was used.
Measurement conditions: ¹ H-NMR (400 MHz, JEOL JNM-LA400), ¹³ C-NMR (150 MHz, Bruker Biospin Advance 600), GC / MS (HP HP9773)
¹ H-NMR (CDCl ₃ ): δ 1.29 (3H, t, J = 7.2 Hz), 2.64 (1H, dd, J = 5.1, 2.7 Hz), 2.82 (1H, dd , J = 5.1, 4.2 Hz), 3.21 (1H, m), 3.50 (1H, dd, J = 11.6, 6.0 Hz), 3.91 (1H, dd, J = 11.6, 2.8 Hz), 4.14 (1H, d, J = 16.4 Hz), 4.19 (1H, d, J = 16.4 Hz), 4.23 (2H, q, J = 7) .2Hz)
¹³ C-NMR (CDCl ₃ ): δ 14.2, 44.1, 50.6, 61.0, 68.5, 72.1, 170.2
MS: m / z 161 [M + H] ⁺

[Example 2]
The crude product (1) of Example 1 (11.6 g) was dissolved in 200 ml of methylene chloride, and isopropanol (15.3 ml), N-methylmorpholine (11.0 ml) and dimethylaminopyridine (1.2 g) were added. After cooling with ice, a methylene chloride solution of water-soluble carbodiimide (18.6 g) was added dropwise over 30 minutes. The reaction was stirred at room temperature for 2 days. The reaction solution was washed with a saturated aqueous sodium bicarbonate solution and then with a saturated saline solution and dried over magnesium sulfate to obtain 22.5 g of a crude product. The title compound (4) (9.7 g) was obtained by silica gel chromatography (silica gel 60, 240 g, hexane / ether 5: 1) from Merck.
Compound (4) (7.9 g) was dissolved in 150 ml of methylene chloride, and a methylene chloride solution of 3-chloroperbenzoic acid (16.1 g) was added dropwise over 90 minutes under ice cooling. The mixture was stirred at room temperature for 2 days, washed with 2N aqueous sodium hydroxide solution and then with saturated brine, dried over magnesium sulfate, and the solvent was distilled off under reduced pressure to obtain a crude product (14.5 g). Purification by silica gel chromatography (silica gel 60, 150 g, hexane / ether 2: 1, manufactured by Merck) gave the title compound (5) (9.2 g).

The compound analysis data regarding the title compound (5) is shown below.
Measurement conditions: ¹ H-NMR (400 MHz, JEOL JNM-LA400), ¹³ C-NMR (150 MHz, Bruker Biospin Advance 600), GC / MS (HP HP9773)
¹ H-NMR (CDCl ₃ ): δ 1.27 (6H, t, J = 6.3 Hz), 2.63 (1H, dd, J = 4.9, 2.7 Hz), 2.81 (1H, dd) , J = 4.9, 4.3 Hz), 3.21 (1H, m), 3.50 (1H, dd, J = 11.6, 6.0 Hz), 3.90 (1H, dd, J = 11.6, 2.9 Hz), 4.10 (1 H, d, J = 16.4 Hz), 4.15 (1 H, d, J = 16.4 Hz), 5.10 (1 H, m)
¹³ C-NMR (CDCl ₃ ): δ 21.8, 44.1, 50.6, 68.6, 72.1, 169.7
MS: m / z 175 [M + H] ⁺

[Example 3]
A solution of allyl alcohol (16.3 ml) in THF (THF 200 ml) was added to a tetrahydrofuran suspension (200 ml) of sodium hydride (10.5 g) under ice cooling, and the mixture was stirred for 30 minutes. Thereafter, a tetrahydrofuran solution (150 ml) of bromoacetic acid (27.8 g) was added dropwise over 60 minutes. The reaction solution was stirred at room temperature for 3 days. The solvent was distilled off under reduced pressure, water was added and the mixture was washed with ethyl acetate. Thereafter, the pH was adjusted to 2 with 5N hydrochloric acid and the aqueous phase was extracted with ethyl acetate. The organic layer was dried over magnesium sulfate to obtain 22.1 g of a crude product (6).
This crude product (6) (22 g) was dissolved in 150 ml of methylene chloride, benzyl alcohol (21.6 ml), N-methylmorpholine (20.9 ml) and dimethylaminopyridine (1.1 g) were added, and the mixture was ice-cooled. Thereafter, a methylene chloride solution of water-soluble carbodiimide (35.3 g) was added dropwise over 60 minutes. The reaction was stirred at room temperature for 2 days. The reaction solution was washed with a saturated aqueous sodium bicarbonate solution and then with a saturated saline solution and dried over magnesium sulfate to obtain 66.0 g of a crude product. Purification by silica gel chromatography (silica gel 60, 240 g, hexane / ether 5: 1 from Merck) gave the title compound (7) (31.5 g).
Compound (7) (31.5 g) was dissolved in 200 ml of methylene chloride, and a solution of 3-chloroperbenzoic acid (28.9 g) in methylene chloride was added dropwise over 90 minutes under ice cooling. The mixture was stirred at room temperature for 2 days, washed with 2N aqueous sodium hydroxide solution and then with saturated brine, dried over magnesium sulfate, and the solvent was distilled off under reduced pressure to obtain a crude product (38.4 g). Purification by silica gel chromatography (silica gel 60, 150 g, hexane / ether 2: 1 from Merck) gave the title compound (8) (30.0 g).

The compound analysis data regarding the title compound (8) is shown below.
Measurement conditions: ¹ H-NMR (400 MHz, JEOL JNM-LA400), ¹³ C-NMR (150 MHz, Bruker Biospin Advance 600), GC / MS (HP HP9773)
¹ H-NMR (CDCl ₃ ): 2.62 (1H, dd, J = 4.9, 2.7 Hz), 2.79 (1H, dd, J = 4.9, 4.4 Hz), 3.20 (1H, m), 3.49 (1H, dd, J = 11.6, 6.0 Hz), 3.91 (1H, dd, J = 11.6, 2.8 Hz), 4.18 (1H, d, J = 16.6 Hz), 4.24 (1H, d, J = 16.6 Hz), 5.20 (2H, s), 7.31-7.39 (5H)
¹³ C-NMR (CDCl ₃ ): δ 44.0, 50.6, 66.6, 68.4, 72.1, 128.4-128.6, 135.3, 170.0
MS: m / z 223 [M + H] ⁺

[Example 4]
Under the same conditions as in Example 3, allyl alcohol (20.4 ml) was added to a suspension of 60% pure sodium hydride (22.0 g) in tetrahydrofuran under ice-cooling and stirred for 30 minutes. Thereafter, a tetrahydrofuran solution of bromoacetic acid (34.7 g) was added dropwise over 60 minutes. The reaction solution was stirred at room temperature for 3 days. The solvent was distilled off under reduced pressure, water was added and the mixture was washed with ethyl acetate. Thereafter, the pH was adjusted to 2 with 5N hydrochloric acid and the aqueous phase was extracted with ethyl acetate. The organic layer was dried over magnesium sulfate to obtain 26.4 g of a crude product (9).
This crude product (9) (3.0 g) was dissolved in 50 ml of methylene chloride, and glycine / benzyl ester / p-toluenesulfonate (11.4 g), dimethylaminopyridine (317 mg) and N-methylmorpholine (3 7 ml) was added, and after ice cooling, a methylene chloride solution of water-soluble carbodiimide (4.0 g) was added dropwise over 60 minutes. The reaction was stirred at room temperature for 2 days. The reaction solution was washed with a saturated aqueous sodium bicarbonate solution and then with a saturated saline solution and dried over magnesium sulfate to obtain 6.8 g of a crude product. Purification by silica gel chromatography (silica gel 60,140 g, methylene chloride / acetonitrile = 90: 10 manufactured by Merck) gave the title compound (10) (6.2 g).
Compound (10) (6.1 g) was dissolved in 100 ml of methylene chloride, and a solution of 3-chloroperbenzoic acid (4.6 g) in methylene chloride was added dropwise over 90 minutes under ice cooling. The mixture was stirred at room temperature for 2 days, washed with 2N aqueous sodium hydroxide solution and then with saturated brine, dried over magnesium sulfate, and the solvent was distilled off under reduced pressure to obtain a crude product (8.6 g). The product was purified by silica gel chromatography (silica gel 60,140 g, methylene chloride / acetonitrile = 90: 10), and the compound was again purified by silica gel chromatography (silica gel 60,120 g, methylene chloride / acetonitrile = 80: 20). Purification gave the title compound (11) (5.5 g).

The compound analysis data regarding the title compound (11) are shown below.
Measurement conditions: ¹ H-NMR (400 MHz, JEOL JNM-LA400 ;; 600 MHz Bruker Advance 600), ¹³ C-NMR (100 MHz, JEOL JNM-LA400), GC / MS (Thermo Electron Voyager)
¹ H-NMR (CDCl ₃ ): 2.67 (1H, dd, J = 2.7, 4.9 Hz), 2.82 (1H, t, J = 4.3, 4.9), 3.18 (1H, m), 3.49 (1H, dd, J = 5.9, 11.6 Hz), 3.87 (1H, dd, J = 2.6, 11.6 Hz), 4.04 (1H, d, J = 15.5 Hz), 4.10 (1H, d, J = 15.5 Hz), 4.11 (1H, d, J = 5.6, 18.2 Hz), 4.16 (1H, d , J = 5.6, 18.2 Hz), 5.19 (2H, s), 7.32-7.40 (5H)
¹³ C-NMR (CDCl ₃ ): δ 40.7, 44.1, 50.4, 67.2, 70.4, 71.8, 128.4-128.6, 135.2, 169.5, 169 .8
MS: m / z 280 [M + H] ⁺

[Example 5] Polymerization reaction 1
Under an argon atmosphere, 10.6 g of the compound (3) obtained in Example 1, 70 ml of ethylene oxide, 1 ml of potassium t-butoxide 1M tetrahydrofuran solution and 10 ml of triisobutylaluminum 1M hexane solution, and 300 ml of hexane as a solvent in a pressure resistant reactor. In addition, the reaction was performed at 30 ° C. for 4.5 hours. After completion of the reaction, the reaction product was recovered, and the solvent was removed under reduced pressure to obtain 41 g of the desired random copolymer (12) as a white solid. The result of measuring the PEG conversion molecular weight of the obtained copolymer by GPC was 80,000. Furthermore, according to ¹ H-NMR analysis, the introduction rate of carboxylic acid ethyl ester measured in heavy water using TSP (3-trimethylsilylpropionic acid-2,2,3,3-d4 deuterated sodium salt) as a standard is It was 2.9 mol%.
10.2 g of the copolymer (12) was dissolved in 30 ml dimethyl sulfoxide (DMSO), 30 ml of 1N sodium hydroxide solution was added, and the mixture was stirred at 50 ° C. for 24 hours. After completion of the reaction, dialysis was performed for 2 days using purified water as an external solution using a dialysis membrane (Spectra / Por2, molecular weight fraction 12,000-14,000) manufactured by Spectrapore, USA. Next, after filtration using a membrane filter (DURAPORE, 0.45 μm) manufactured by Millipore Corporation, a random copolymer (13) 9 in which the white amorphous target ethyl group is deprotected through a freeze-drying step. .4 g was obtained. In the following formula, a and b represent integers.

^{From the 1} H-NMR analysis, it was confirmed that the ethyl group was removed, and the introduction rate of the carboxymethyl group was 2.7 mol% in molar ratio. The result of measuring the PEG conversion molecular weight of the obtained copolymer by GPC was 89,000, and Mw / Mn = 2.7. The result of GPC analysis is shown in FIG.

[Example 6] Polymerization reaction 2
Under an argon atmosphere, 9.3 g of the compound (8) obtained in Example 3 in a pressure-resistant reaction vessel, 70 ml of ethylene oxide, 1.1 ml of potassium 2-methyl-2-butoxide 1M tetrahydrofuran solution and triisobutylaluminum 1M hexane solution 11. 1 ml and 300 ml of hexane as a solvent were added and reacted at 25 ° C. for 5 hours. After completion of the reaction, the reaction product was collected and the solvent was removed under reduced pressure to obtain 50 g of the desired random copolymer (14) as a white solid. The result of measuring the PEG conversion molecular weight of the obtained copolymer by GPC was 39,000. Furthermore, the introduction ratio of carboxylic acid benzyl ester by ¹ H-NMR analysis was 2.0 mol% in molar ratio.
20.1 g of the copolymer (14) was dissolved in 60 ml DMSO, 60 ml of 1N sodium hydroxide solution was added, and the mixture was stirred at 50 ° C. for 20 hours. After completion of the reaction, dialysis was performed for 2 days using purified water as an external solution using a dialysis membrane (Spectra / Por2, molecular weight fraction 12,000-14,000) manufactured by Spectrapore, USA. Next, after filtration using a membrane filter (DURAPORE, 0.45 μm) manufactured by Millipore, a random copolymer (15) 16 in which the target amorphous benzyl group is deprotected through a freeze-drying step. 0.0 g was obtained. In the following formula, c and d represent integers.

^{From 1} H-NMR analysis, it was confirmed that the benzyl group was removed, and the carboxymethyl group introduction rate was 2.1 mol% in terms of molar ratio. The result of measuring the PEG conversion molecular weight of the obtained copolymer by GPC was 58,000, and Mw / Mn = 1.6. The results of GPC analysis are shown in FIG.

[Example 7] Polymerization reaction 3
In an argon atmosphere, 9.3 g of the compound (8) obtained in Example 3, 70 ml of ethylene oxide, 1 ml of potassium tert-butoxide 1M tetrahydrofuran solution and 10 ml of triisobutylaluminum 1M hexane solution, and 300 ml of hexane as a solvent were placed in a pressure resistant reactor. In addition, the reaction was performed at 30 ° C. for 4.5 hours. After completion of the reaction, the reaction product was collected and the solvent was removed under reduced pressure to obtain 50 g of the desired random copolymer (16) as a white solid. The result of measuring the PEG conversion molecular weight of the obtained copolymer by GPC was 35,000. Furthermore, the introduction ratio of carboxylic acid benzyl ester by ¹ H-NMR analysis was 2.1 mol% in molar ratio.
20.1 g of the copolymer (16) was dissolved in 60 ml DMSO, 60 ml of 1N sodium hydroxide solution was added, and the mixture was stirred at 50 ° C. for 20 hours. After completion of the reaction, dialysis was performed for 2 days using purified water as an external solution using a dialysis membrane (Spectra / Por2, molecular weight fraction 12,000-14,000) manufactured by Spectrapore, USA. Next, after filtration using a membrane filter (DURAPORE, 0.45 μm) manufactured by Millipore, a random copolymer (17) 16 in which the target amorphous benzyl group is deprotected through a freeze-drying step. .9 g was obtained. In the following formulae, e and f represent integers.

It was confirmed by ¹ H-NMR analysis that the benzyl group was removed, and the carboxymethyl group introduction rate was 2.1 mol% in terms of molar ratio. The result of measuring the molecular weight in terms of PEG of the obtained copolymer by GPC was 54,000, and Mw / Mn = 1.7. The results of GPC analysis are shown in FIG.

[Example 8] Toxicity evaluation Regarding the compound (15) in Example 6 above, physiological saline was used as an administration solvent, and physiological saline was used as a comparative control, and the compound (15) was administered at 1000 mg / kg and 500 mg / kg. From the tail vein of 6-week-old BALB / c female mice (each group n = 5, purchased from Japan SLC Co., Ltd.) on the 0th, 3rd, and 6th days under the condition of a dose of 25 ml / kg Intermittent administration was performed over several times. The result is shown in FIG.
Evaluation was performed using weight loss as an index. The change in body weight when the weight of the mouse on the start day of the experiment was taken as 100% was evaluated, and it was judged that there was toxicity when a weight loss of 10% occurred. As a result, body weight loss of 10% or more after the administration start date was not observed, and no significant difference was observed on the 25th day after administration, and it was determined that there was no toxicity. Each point in the figure indicates the average body weight ± standard deviation (SD) in each administration group. The upward arrow in the figure indicates the day of administration (Day 0, Day 3, Day 6).

The following abbreviations are used in the following examples.
DMF: N, N-dimethylformamide, Trt: triphenylmethyl group (trityl group), Z: benzyloxycarbonyl group, Fmoc: 9-fluorenylmethyloxycarbonyl group, DMAP: N, N-dimethylaminopyridine, HBTU : 2- (1-benzotriazol-1-yl) -1,1,3,3-tetramethylaminium hexafluorophosphate, DIPC: N, N'-diisopropylcarbodiimide, tBuOK: Potassium t-butoxide, DMSO- d6: deuterated dimethyl sulfoxide

Example 9
(Step 1) Production of 2′-Gly-paclitaxel hydrochloride (18) Using paclitaxel manufactured by Hauser, USA as a raw material, an amino acid linker was introduced into the 2′-position hydroxyl group of paclitaxel, and 2′-Gly-paclitaxel hydrochloride (18 ) Was prepared.
The structure of the product was confirmed by ¹ H-NMR and HRMS (high-resolution mass spectrometry). That is, Fmoc-Gly (178 mg, 0.6 mmol), dimethylaminopyridine (73 mg, 0.6 mmol), and paclitaxel (made by HAUSER, USA, 427 mg, 0.5 mmol) were dissolved in methylene chloride (20 ml). Then, N, N′-diisopropylcarbodiimide (76 mg, 0.6 mmol) was added and stirred at room temperature overnight. The reaction solvent was distilled off, and silica gel column chromatography (silica gel: Merck, Germany, Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / acetonitrile = 70 / 30) to obtain 2′-Fmoc-Gly-paclitaxel (489 mg).
This compound (420 mg) was dissolved in N, N-dimethylformamide (10 ml), piperidine (2 ml) was added at room temperature, the mixture was stirred for 5 minutes, the solvent was distilled off to remove Fmoc, and silica gel column chromatography (silica gel : Purified by Merck, Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 50 cm, eluent: acetonitrile / methylene chloride = 80/20), title compound (18) (145 mg) Got. The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 1.01 (s, 3H, Me-17) 1.05 (s, 3H, Me-16) 1.52 (dd, 1H, J = 14.6, 9. 2 Hz, H-14b) 1.51 (s, 3H, Me-19) 1.65 (t, 1H, J = 11.6 Hz, H-6b) 1.81 (dd, 1H, J = 15.5) 9.6 Hz, H-14a) 1.86 (s, 3H, Me-18) 2.11 (s, 3H, Ac-10) 2.23 (s, 3H, Ac-4) 2.32 (m, 1H, H-6a) 3.58 (d, 1H, J = 7.0 Hz, H-3) 3.96-4.07 (m, 3H, GlyCH2, H-20) 4.10 (dd, 1H, J = 6.7, 10.7, H-7) 4.63 (s, 1H, OH-1) 4.90 (brs, 1H, OH-7) 4.91 (dd, 1H, J = 4. 9 Hz, H-5) 5.43 (d, 1H, J = 7.0, H-2) 5.46 (d, 1H, J = 8.2 Hz, H-2 ′) 5.58 (t, 1H , J = 8.4 Hz, H-3 ′) 5.87 (t, 1H, J = 8.6 Hz, H-13) 6.30 (s, 1H, H-10) 7.19-8.00 ( aromatic, 15H) 8.40 (brs, 2H, GlyNH2) 9.25 (d, 1H, J = 8.6 Hz, CONH-3 ')
HRMS: m / z 911.3604
(M + H) +: Calculated value as C49H55O15N2 911.3602

(Step 2) Production of polyether-2′-Gly-paclitaxel (19) The copolymer (15) (100 mg) obtained in Example 6 was dissolved in 5 ml of DMF, and this solution was added to this solution under ice cooling. DMF (0.5 ml) in which 2′-Gly-paclitaxel (18) (10 mg) obtained in (Step 1) was dissolved and DMF (0.5 ml) in which HBTU (52 mg) was dissolved were added, and the mixture was stirred at room temperature for 2 hours. Stir. This reaction solution was dialyzed against purified water at 4 ° C. for 2 days using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). The internal solution was filtered through a membrane filter (0.45 μm) and then freeze-dried to obtain the title compound (19) (102 mg). The amount of drug introduced into this complex was 6.1% (% by weight) as calculated from the ultraviolet absorbance at 254 nm and the total weight of the complex.

Example 10
(Step 1) Preparation of 2'-Ala-paclitaxel (20) Z-Ala (145 mg, 0.65 mmol), DMAP (79 mg, 0.65 mmol) and paclitaxel (427 mg, 0.5 mmol) in methylene chloride (20 ml) Dissolved in. Then DIPC (82 mg, 0.65 mmol) was added and stirred at room temperature overnight. The reaction solvent was distilled off, and silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / acetonitrile = 70/30 ) To obtain 2′-Z-Ala-paclitaxel (435 mg). This compound (400 mg) was dissolved in dioxane (20 ml), palladium-carbon catalyst (200 mg) was added, and the mixture was stirred under a hydrogen atmosphere for 4 hours. The catalyst was removed by filtration, and then the solvent was distilled off under reduced pressure. Is purified by silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400mesh, column: 2.0 × 30 cm, eluent: methylene chloride / methanol / acetonitrile = 95/5/5). The title compound (20) (220 mg) was obtained. The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 1.01 (s, 3H, Me-17) 1.03 (s
, 3H, Me-16) 1.14 (s, 3H, Me-Ala) 1.51 (s, 3H, Me-
19) 1.61 (dd, 1H, J = 15.6, 9.2 Hz, H-14b) 1.64 (t,
1H, J = 12.8 Hz, H-6b) 1.81 (s, 3H, Me-18) 1.88 (dd
, 1H, J = 15.3, 9.5 Hz, H-14a) 2.11 (s, 3H, Ac-10)
2.27 (s, 3H, Ac-4) 2.33 (m, 1H, H-6a) 3.52 (q, 1H,
J = 7.0 Hz, H-Ala) 3.60 (d, 1H, J = 7.3 Hz, H-3) 4.02
(D, 1H, J = 15.0 Hz, H-20) 4.03 (d, 1H, J = 15.0 Hz, H
-20) 4.12 (ddd, 1H, J = 6.6, 6.6, 17.4 Hz, H-7) 4.6
6 (s, 1H, OH-1) 4.91 (d, 1H, J = 6.6, OH-7) 4.92 (dd
, 1H, J = 9.8 Hz, H-5) 5.35 (d, 1H, J = 8.6, H-2 ′) 5.4
3 (d, 1H, J = 7.0 Hz, H-2) 5.64 (t, 1H, J = 8.6 Hz, H-3)
') 5.87 (t, 1H, J = 9.2 Hz, H-13) 6.30 (s, 1H, H-10)
7.20-8.00 (aromatic, 15H) 9.17 (d, 1H, J = 8.9 Hz)
, CONH-3 ')
HRMS: m / z 925.3797
(M + H) +: Calculated value as C50H57O15N2 925.3759

(Step 2) Production of polyether-2′-Ala-paclitaxel (21) The copolymer (15) (100 mg) obtained in Example 6 was obtained in Example 10 (Step 1). The title compound (21) (103 mg) was obtained in the same manner as in Example 9 (Step 2) except that '-Ala-paclitaxel (20) (25 mg) was used. The amount of drug introduced into this complex was 3.7% (% by weight) as calculated from the ultraviolet absorbance at 254 nm and the total weight of the complex.

Example 11
(Step 1) Production of 2′-Leu-paclitaxel (22) 2′-Z-Leu-paclitaxel (Step 1) was performed in the same manner as in Example 10 (Step 1) except that Z-Leu (172 mg, 0.65 mmol) was used. 450 mg) was obtained. This compound (400 mg) was dissolved in dioxane (20 ml), palladium-carbon catalyst (200 mg) was added, and the mixture was stirred under a hydrogen atmosphere for 4 hours. The catalyst was removed by filtration, and the solvent was distilled off under reduced pressure. Purification by silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / methanol / acetonitrile = 95/5/5) The title compound (22) (280 mg) was obtained. The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 0.66 (d, 3H, Me-Leu) 0.70 (d, 3H, Me-Leu) 1.01 (s, 3H, Me-17) 1.03 ( s, 3H, Me-16) 1.26 (ddd, 1H, J = 6.4, 8.5, 13.4 Hz, H-Leu) 1.36 (ddd, 1H, J = 5.8, 7. 6, 13.4 Hz, H-Leu) 1.51 (s, 3H, Me-19) 1.56 (dd, 1H, J = 15.3, 9.0 Hz, H-14b) 1.64 (m, 1H, H-6b) 1.67 (m, 1H, H-Leu) 1.79 (s, 3H, Me-18) 1.84 (dd, 1H, J = 15.3, 9.5 Hz, H- 14a) 2.10 (s, 3H, Ac-10) 2.25 (s, 3H, Ac-4) 2.33 (ddd, 1H, J = 14.7, 9.5, 6.4H) z, H-6a) 3.38 (dd, 1H, J = 8.6, 5.8 Hz, H-Leu) 3.59 (d, 1H, J = 7.0 Hz, H-3) 4.01 ( d, 1H, J = 16.8 Hz, H-20) 4.03 (d, 1H, J = 16.8 Hz, H-20) 4.12 (ddd, 1H, J = 6.9, 6.9, 11.0 Hz, H-7) 4.64 (s, 1H, OH-1) 4.90 (d, 1H, J = 7.0, OH-7) 4.92 (d, 1H, J = 10. 1 Hz, H-5) 5.34 (d, 1H, J = 9.2, H-2 ′) 5.42 (d, 1H, J = 7.0 Hz, H-2) 5.62 (t, 1H , J = 9.0 Hz, H-3 ′) 5.86 (t, 1H, J = 9.2 Hz, H-13) 6.30 (s, 1H, H-10) 7.20-8.00 ( aromatic, 15H) 9.16 (d, 1H J = 8.9Hz, CONH-3 ')
HRMS: m / z 967.4321
(M + H) +: Calculated value as C53H63O15N2 967.4228

(Process 2)
Production of polyether-2′-Leu-paclitaxel (23) 2′-Leu- obtained in Example 11 (Step 1) with respect to copolymer (15) (100 mg) obtained in Example 6 The title compound (23) (97 mg) was obtained in the same manner as in Example 9 (Step 2) except that paclitaxel (22) (25 mg) was used. The amount of drug introduced into this complex was 4.0% (% by weight) as calculated from the ultraviolet absorbance at 254 nm and the total weight of the complex.

Example 12
(Step 1) Preparation of 2′-Ile-paclitaxel (24) Fmoc-Ile (212 mg, 0.6 mmol), DMAP (73 mg, 0.6 mmol) and paclitaxel (427 mg, 0.5 mmol) in methylene chloride (20 ml) Dissolved in. Then, DIPC (76 mg, 0.6 mmol) was added and stirred overnight at room temperature. The reaction solvent was distilled off, and silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 2.0 × 30 cm, eluent: methylene chloride / acetonitrile = 70 / 30) to obtain 2′-Fmoc-Ile-paclitaxel (553 mg). This compound (470 mg) was dissolved in DMF (10 ml), piperidine (2 ml) was added at room temperature, and the mixture was stirred for 5 minutes. The solvent was distilled off under reduced pressure, and the residue was purified by silica gel column chromatography (silica gel: Merck). Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / methanol / acetonitrile = 95/5/5) and purified by the title compound (24) (350 mg) Got. The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 0.62 (t, 3H, J = 7.5 Hz, Me-Ile) 0.81 (d, 3H, J = 6.7 Hz, Me-Ile) 1.01 ( s, 3H, Me-17) 1.03 (s, 3H, Me-16) 1.07 (ddd, 1H, J = 14.4, 7.3, 4.9 Hz, H-Ile) 1.32 ( ddd, 1H, J = 13.4, 7.6, 4.6 Hz, H-Ile) 1.51 (s, 3H, Me-19) 1.56 (dd, 1H, J = 15.3, 9 .. 2Hz, H-14b) 1.56-1.61 (m, 1H, H-Ile) 1.64 (dd, 1H, J = 13.7, 3.1 Hz, H-6b) 1.79 (s, 3H, Me-18) 1.87 (dd, 1H, J = 15.3, 9.8 Hz, H-14a) 2.10 (s, 3H, Ac-10) 2.29 (s, 3H, Ac-4) 2.33 (ddd, 1H, J = 14.4, 9.6, 6.4 Hz, H-6a) 3.60 (d, 1H, J = 7.3 Hz, H-3) 3.60-3.67 (m, 1H, H-Ile) 4.02 (d, 1H, J = 16.6 Hz, H-20) 4.03 (d, 1H, J = 16.6 Hz, H− 20) 4.12 (ddd, 1H, J = 10.8, 6.7, 6.7 Hz, H-7) 4.64 (s, 1H, OH-1) 4.90 (d, 1H, J = 7.0, OH-7) 4.92 (d, 1H, J = 9.8 Hz, H-5) 5.37 (d, 1H, J = 8.9 Hz, H-2 ′) 5.43 (d , 1H, J = 7.3 Hz, H-2) 5.64 (t, 1H, J = 8.7 Hz, H-3 ′) 5.85 (dt, 1H, J = 0.9, 9.2 Hz, H-13) 6.30 (s, 1H, H-10) 7.20-8.00 (aromatic, 15H) 9.15 (d, 1H, J = 9.2Hz, CONH-3 ')
HRMS: m / z 967.4234
(M + H) +: Calculated value as C53H63O15N2 967.4228

(Step 2) Production of polyether-2′-Ile-paclitaxel (25) The copolymer (15) (100 mg) obtained in Example 6 was obtained in Example 12 (Step 1). '-Ile-paclitaxel (24) (30 mg) was used in the same manner as in Example 9 (Step 2) to give the title compound (25) (92 mg). The amount of drug introduced into this complex was 2.5% (% by weight) as calculated from the ultraviolet absorbance at 254 nm and the total weight of the complex.

Example 13
(Step 1) Production of 2′-Phe-Gly-paclitaxel hydrochloride (26) Phe-Gly (Peptide Institute, 1.1 g, 5 mmol) was added to water (2 ml), 2-propanol (2 ml) and diethylamine (1 0.5 ml), trityl chloride (1.8 g, 6.5 mmol) was gradually added to the reaction solution and stirred for 1 hour. Water was added to the reaction solution, and the resulting precipitate was washed with water. Next, 5 ml of acetic acid was added to the precipitate to make it acidic, and then the solvent was distilled off under reduced pressure to obtain 1.4 g of Trt-Phe-Gly. The obtained Trt-Phe-Gly (604 mg, 1.3 mmol), DMAP (158 mg, 1.3 mmol) and paclitaxel (853 mg, 1.0 mmol) were dissolved in methylene chloride (20 ml). Next, DIPC (164 mg, 1.3 mmol) was added and stirred overnight at room temperature. The reaction solvent was distilled off, and silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / acetonitrile = 80/20 ) To obtain 2′-Trt-Phe-Gly-paclitaxel (990 mg). This compound (800 mg) was treated with 90% acetic acid (10 ml) to be de-N-tritylated, and silica gel column chromatography (silica gel: Art No. 9385, Silica gel 60, 200-400 mesh, manufactured by Merck, Germany), column: 4.0 × 30 cm, eluent: methylene chloride / methanol / acetonitrile = 95/5/5), and then the acetate is converted to the hydrochloride with an ion exchange resin to give the title compound (26) (450 mg). Obtained. The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6); δ 1.01 (s, 3H, Me-17) 1.03 (s, 3H, Me-16) 1.42 (dd, 1H, J = 15.5, 9 .. 1 Hz, H-14b) 1.50 (s, 3H, Me-19) 1.63 (t, 1H, J = 12.2 Hz, H-6b) 1.75 (dd, 1H, J = 12.3) 9.5 Hz, H-14a) 1.81 (s, 3H, Me-18) 2.12 (s, 3H, Ac-10) 2.23 (s, 3H, Ac-4) 2.29 (ddd, 1H, J = 14.4, 9.2, 7.0 Hz, H-6a) 2.90 (dd, 1H, 14.2, 7.8 Hz, PheCH2) 3.08 (dd, 1H, 14.4 5.2 Hz, PheCH2) 3.56 (d, 1H, J = 7.0 Hz, H-3) 4.05-4.10 (m, 2H, H-7, PheCH ) 4.15 (dd, 1H, J = 18.0, 5.8 Hz, Gly) 4.61 (brs, 1H, OH-1) 4.90 (brs, 1H, OH-7) 4.90 (d , 1H, J = 5.3 Hz, H-5) 5.38 (d, 1H, J = 8.9 Hz, H-2 ′) 5.41 (d, 1H, J = 7.0 Hz, H-2) 5.53 (t, 1H, J = 8.6 Hz, H-3 ′) 5.83 (t, 1H, J = 8.8 Hz, H-13) 6.29 (s, 1H, H-10) 7 16 to 8.00 (m, 20H, aromatic) 8.15 (brs, 2H, NH2) 9.02 (t, 1H, J = 5.8 Hz, Gly-NH) 9.29 (d, 1H, J = 8.9Hz, CONH-3 ')
HRMS: m / z 1058.241
(M + H) +: Calculated value as C58H64O16N3 10588.4287

(Process 2)
Production of polyether-2′-Phe-Gly-paclitaxel (27)
Except that the 2′-Phe-Gly-paclitaxel (26) (30 mg) obtained in Example 13 (Step 1) was used for the copolymer (15) (100 mg) obtained in Example 6. In the same manner as in Example 9 (Step 2), the title compound (27) (95 mg) was obtained. The amount of drug introduced into this complex was 3.5% (% by weight) as calculated from the ultraviolet absorbance at 254 nm and the total weight of the complex.

Example 14
Preparation of Trt-Gly-Gly-Phe-Gly (28) (1) Synthesis of Phe-Gly-OBn (29) Phe-Gly.H2O (produced by Kokusan Chemical Co., Ltd., 25 g, 104 mmol) was hydrated with paratonin sulfonic acid The product (19.8 g, 104 mmol), benzyl alcohol (25 ml) and toluene (200 ml) are dissolved and heated to reflux with a Dean-Stark apparatus for 5 hours. After the reaction, the solvent was distilled off, and diethyl ether was added to obtain the paratoluenesulfonic acid salt (34 g) of Phe-Gly-OBn (29) as the title compound.
(2) Synthesis of Trt-Gly-Gly (30) Gly-Gly (Peptide Institute, 6.6 g, 50 mmol) was added to a mixed solution of purified water (20 ml), 2-propanol (40 ml) and diethylamine (15 ml). Dissolve, and gradually add trityl chloride (18.1 g, 65 mmol) to the reaction mixture and stir for 1 hour. Purified water was added to the reaction solution, and the resulting precipitate was washed with water. Next, 5 ml of acetic acid was added to the precipitate to make it acidic, and then the solvent was distilled off under reduced pressure to obtain 13.3 g of the title compound (30).
(3) Synthesis of Trt-Gly-Gly-Phe-Gly-OBn (31) To dry DMF (10 ml), Trt-Gly-Gly (30) (1.54 g), N-hydroxysuccinimide (0.52 g) ) And DCC (0.93 g) were added and reacted at 4 ° C. for 3 hours. To the reaction solution was added a DMF solution (DMF 10 ml) in which p-toluenesulfonate (2.0 g) of Phe-Gly-OBn (29) and N-methylmorpholine (0.41 g) were dissolved, and the mixture was reacted at 4 ° C. for 15 hours. . The precipitate was removed, the solvent was distilled off, and the residue was purified by silica gel column chromatography (silica gel: Merck, Germany, Art No. 9365, Silica gel 60, 200-400 mesh, eluent: chloroform / methanol = 20/1). Purification gave the title product (31) (1.6 g).
(4) Synthesis of Trt-Gly-Gly-Phe-Gly (28) Compound (31) Trt-Gly-Gly-Phe-Gly-OBn (1.3 g) was dissolved in DMF (20 ml) and 10% palladium-carbon (0.5 g) and 1,4-cyclohexadiene (0.4 g) are added and reacted at room temperature for 30 minutes. The reaction solution was filtered, the catalyst was removed, the solvent was distilled off, and the residue was subjected to silica gel column chromatography (silica gel: Merck Art No. 9365, Silica gel 60, 200-400 mesh, eluent: chloroform / methanol = 7 / The title compound (28) (1.1 g) was obtained by purification in 1).
Anal. Calcd for: C34H34N4O5: C, 70.57; H, 5.92; N, 9.68
Found: C, 70.03; H, 6.07; N, 9.67
Amino acid analysis: Phe (1) 1.00, Gly (3) 2.91
Hydrolysis conditions: 6N HCl, 110 ° C., 22 hrs

Example 15
Preparation of 2′-Gly-Gly-Phe-Gly-paclitaxel hydrochloride (32) (28) Trt-Gly-Gly-Phe-Gly (739 mg, 1.3 mmol) obtained in Example 14 and DMAP (158 mg, 1.3 mmol) and paclitaxel (853 mg, 1.0 mmol) were dissolved in methylene chloride (20 ml). Next, DIPC (164 mg, 1.3 mmol) was added and stirred at room temperature for 4 hours. The reaction solvent was distilled off, silica gel column chromatography (silica gel: Merck, Germany, Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 50 cm, eluent: methylene chloride / methanol / acetonitrile. = 95/5/30) to obtain 2′-Nα-Trt-Gly-Gly-Phe-Gly-paclitaxel (33) (1250 mg).
HRMS: m / z 1414.5763
(M + H) +: Calculated value as C81H84O18N5 1414.5811
This compound (33) (1100 mg) was treated with 75% acetic acid (10 ml) for de-N-tritylation, and silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column : 4.0 × 50 cm, eluent: methylene chloride / methanol / acetonitrile = 85/15/5), and then converted to hydrochloride with anion exchange resin to give the title compound (32) (533 mg) It was. The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ1.00 (s, 3H, Me-17) 1.03 (s, 3H, Me-16) 1.42 (dd, 1H, J = 15.4,9. 2Hz, H-14b) 1.49 (s, 3H, Me-19) 1.63 (brt, 1H, J = 12.1 Hz, H-6b) 1.74 (dd, 1H, J = 15.4, 9.2 Hz, H-14a) 1.80 (s, 3H, Me-18) 2.11 (s, 3H, Ac-10) 2.23 (s, 3H, Ac-4) 2.30 (m, 1H, H6a) 2.72 (dd, 1H, J = 13.9, 10.2 Hz, PheCH2Hb) 3.02 (dd, 1H, J = 13.9, 3.8 Hz, PheCH2CHa) 3.52 (brs, 2H, GlyCH2) 3.56 (d, 1H, J = 7.2 Hz, H-3) 3.66 (dd, 1H, 1 .9, 5.4 Hz, GlyCH2b) 3.84 (dd, 1H, 16.9, 5.4 Hz, GlyCH2a) 4.01 (dd, 2H, J = 14.5, 8.4 Hz, H-20a, H -20b) 4.01 (2H, GlyCH2) 4.09 (m, 1H, H-7) 4.55 (ddd, 1H, J = 10.2, 8.5, 3.8 Hz, PheCH2CH) 4.61 (S, 1H, OH-1) 4.89 (dd, 1H, J = 8.9, 1.3 Hz, H-5) 4.92 (brs, 1H, OH-7) 5.41 (d, 1H , J = 7.2, H-2) 5.43 (d, 1H, J = 6.3 Hz, H-2 ′) 5.51 (t, 1H, J = 8.5 Hz, H-3 ′) 5 .83 (t, 1H, J = 9.2 Hz, H-13) 6.29 (s, 1H, H-10) 7.10-8.00 (aromatic, 20 ) 8.33 (d, 1H, PheCONH) 8.51 (t, 1H, J = 5.5 Hz, GlyCONH) 8.69 (t, 1H, J = 6.0 Hz, GlyCONH) 9.34 (d, 1H , J = 8.5 Hz, CONH-3 ′)
HRMS: m / z 1172.4711
(M + H) +: Calculated value as C62H70O18N5 1172.4716
Anal. Calcd for: C62H69O18N5.HCl.2.5H2O: C, 59.40; H, 6.
03; N, 5.59.
Found: C, 59.55; H, 6.04; N, 5.60

Example 16
Preparation of polyether-2′-Gly-Gly-Phe-Gly-paclitaxel (34) Copolymer (15) (103 mg) obtained in Example 6 was dissolved in 5 ml of DMF, and this solution was obtained in Example 15. 2′-Gly-Gly-Phe-Gly-paclitaxel (32) (10.3 mg) dissolved in DMF (0.5 ml) and HBTU (53 mg) dissolved in DMF (0.5 ml) were added at room temperature. Stir for 2 hours. This reaction solution was dialyzed against purified water at 4 ° C. for 2 days using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). The internal solution was filtered through a membrane filter (0.45 μm) and then freeze-dried to obtain the title compound (34) (113 mg). The amount of drug introduced into this complex was 6.4% (% by weight) as calculated from the ultraviolet absorbance at 254 nm and the total weight of the complex.

Example 17
Production of 20-Gly-camptothecin hydrochloride (35) (Step 1) Production of 20-BOC-Gly-camptothecin (36)
BOC-Gly (263 mg, 1.5 mmol), dimethylaminopyridine (122 mg, 1.0 mmol) and (S)-(+)-camptothesin (manufactured by Tokyo Chemical Industry, 174 mg, 0.5 mmol) in methylene chloride (20 ml) Dissolved. Next, DIPC (189 mg, 1.5 mmol) was added and stirred at room temperature for 3.5 hours. The reaction solution was washed with 0.1N hydrochloric acid and saturated brine, and then the solvent was distilled off under reduced pressure. Purification by silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / acetonitrile = 70/30), 20− BOC-Gly-camptothecin (36) (315 mg) was obtained.

(Step 2) Production of 20-Gly-camptothecin hydrochloride (35) Compound (36) (300 mg) was dissolved in methylene chloride (5 ml), trifluoroacetic acid (TFA; 2 ml) was added at room temperature, and the mixture was stirred for 5 minutes. Then, the solvent was distilled off. The de-BOC compound was subjected to silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 50 cm, eluent: acetonitrile / methanol / methylene chloride = 10 / 10/90) and then treated with an anion exchange resin to give the title compound (35) (214 mg). The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 0.97 (s, 3H) 2.20 (m, 2H) 4.08, 4.34 (dd, 2H) 5.32 (s, 2H) 5.56 ( s, 2H) 7.32 (s, 1H) 7.73 (t, 1H) 7.88 (t, 1H) 8.15 (d, 1H) 8.17 (d, 1H) 8.56 (brs, 2H) 8.72 (s, 1H)
¹³ C-NMR (DMSO-d6): 7.53, 30.18, 38.6-40.2, 50.10, 66.29, 77.42, 95.51, 118.78, 127.63, 127.82, 128.45, 128.66, 129.49, 130.43, 131.58, 144.72, 145.97, 147.77, 152.13, 156.39, 166.77

Example 18
Production of polyether-20-Gly-camptothecin (37) 20-Gly-camptothecin hydrochloride obtained in Example 17 was dissolved in 5 ml of copolymer (15) (103 mg) DMF obtained in Example 6. DMF (0.5 ml) in which salt (35) (20 mg) was dissolved and DMF (0.5 ml) in which HBTU (108 mg) was dissolved were added, and the mixture was stirred at room temperature for 2 hours. This reaction solution was dialyzed against purified water at 4 ° C. for 2 days using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). The internal solution was filtered through a membrane filter (0.22 μm) and then lyophilized to obtain the title compound (37) (105 mg). The amount of drug introduced into this complex was 3.5% (% by weight) as calculated from the ultraviolet absorbance at 254 nm and the total weight of the complex.

Example 19
Preparation of 20-GlyGlyPheGly-camptothecin hydrochloride (38) (step 1) Preparation of 20-TrtGlyGlyPheGly-camptothecin (39) (28) Trt-GlyGlyPheGly (1736 mg, 3 mmol) obtained in Example 14 and dimethylaminopyridine (1736 mg) 244 mg, 2 mmol) and (S)-(+)-Campothesin (348 mg, 1 mmol) were dissolved in methylene chloride (100 ml). Then, DIPC (378 mg, 3 mmol) was added and stirred at room temperature overnight. The reaction solution was washed with 0.1N hydrochloric acid and saturated brine, and then the solvent was distilled off under reduced pressure. Purification by silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / acetonitrile = 70/30), 20− TrtGlyGlyPheGly-camptothecin (39) (754 mg) was obtained.

(Step 2) Production of 20-GlyGlyPheGly-camptothecin hydrochloride (38) Compound (39) (700 mg) was dissolved in 75% acetic acid (8 ml), stirred for 20 minutes, and then the solvent was distilled off. The detrtrified compound was converted into hydrochloride with an anion exchange resin (Bio-Rad AG1-X8), and silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400). Purification with mesh, column: 4.0 × 50 cm, eluent: methanol / methylene chloride = 15/85) gave the title compound (38) (535 mg). The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 0.93 (t, 3H) 2.32 (m, 2H) 2.77, 3.06 (dd, dd, 2H) 3.27 (s, 2H) 61, 3.76 (d, d, 2H) 4.11, 4.21 (dd, dd, 2H) 4.55 (m, 1H) 5.27 (s, 2H) 5.51 (s, 2H) 7.15 (m, 1H) 7.19 (s, 1H) 7.22 (m, 4H) 7.71 (t, 1H) 7.87 (t, 1H) 8.12 (d, 1H) 17 (d, 1H) 8.23 (brs, 1H) 8.31 (d, 1H) 8.68 (s, 1H) 8.68 (brs, 2H)
¹³ C-NMR (DMSO-d6): 7.52, 30.42, 37.43, 40.41, 41.64, 42.21, 50.08, 53.96, 66.28, 76.27, 95.20, 118.87, 126.20, 127.59, 127.80, 128.01, 128.39, 128.79, 129.06, 129.50, 130.37, 131.49, 137. 82,145.11,145.87,147.72,152.12,156.41,167.00,168.35,168.84,171.68

Example 20
Preparation of polyether-20-GlyGlyPheGly-camptothecin (40) Copolymer (15) (100 mg) obtained in Example 6 was dissolved in DMF (5 ml), and 20- GlyGlyPheGly-camptothecin hydrochloride (38) (20 mg) was added with DMF (0.5 ml) and DMF (0.5 ml) in which HBTU (100 mg) was dissolved, and the mixture was stirred at room temperature for 2 hours. This reaction solution was dialyzed against purified water at 4 ° C. for 2 days using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). The internal solution was filtered through a membrane filter (0.22 μm) and then lyophilized to obtain the title compound (40) (101 mg). The amount of drug introduced into this complex was 4.5% (% by weight) as calculated from the ultraviolet absorbance at 254 nm and the total weight of the complex.

Example 21
Production of 21-Ala-dexamethasone hydrochloride (41) (Step 1) Production of 21-BOC-Ala-dexamethasone (42)
BOC-Ala (568 mg, 3 mmol), dimethylaminopyridine (244 mg, 2 mmol) and dexamethasone (392 mg, 1 mmol) were dissolved in dry methylene chloride (30 ml). Then, DIPC (252 mg, 2 mmol) was added and stirred at room temperature for 0.5 hour. The reaction solution was washed with 0.1N hydrochloric acid and saturated brine, and then the solvent was distilled off under reduced pressure. Purified by silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / acetonitrile = 80/20), 21- BOC-Ala-dexamethasone (42) (553 mg) was obtained.

(Step 2) Preparation of 21-Ala-dexamethasone hydrochloride (41) Compound (42) (500 mg) was dissolved in methylene chloride (10 ml), trifluoroacetic acid (TFA; 1 ml) was added at room temperature, and the mixture was stirred for 2 hours. Was distilled off. The de-BOC compound was subjected to silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 50 cm, eluent: acetonitrile / methanol / methylene chloride = 5 / 15/85), and then treated with an anion exchange resin to give the title compound (41) (412 mg). The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 0.80 (d, 3H) 0.90 (s, 3H) 1.09 (s, 1H) 1.33-1.79 (m, 4H) 1.49 ( s, 3H) 1.50 (d, 3H) 2.08-2.91 (m, 6H) 3.35 (brs, 1H) 4.17 (m, 1H) 4.24 (q, 1H) 95 (d, 1H) 5.20 (d, 1H) 5.27 (s, 1H) 5.53 (d, 1H) 6.01 (s, 1H) 6.23 (dd, 1H) 7.33 ( d, 1H) 8.46 (brs, 1H)
¹³ C-NMR (DMSO-d6): 15.20, 16.09, 16.29, 23.08, 27.41, 30.41, 32.05, 33.70, 35.61, 35.83, 43.42, 47.80, 48.05, 48.25, 69.38, 70.41, 90.61, 101.42, 124.19, 129.03, 152.94, 167.17, 169. 98, 185.41, 204.30

[Example 22]
Production of polyether-21-Ala-dexamethasone (43) The copolymer (15) (100 mg) obtained in Example 6 was dissolved in DMF (5 ml). To this solution was added DMF (0.5 ml) in which 21-Ala-dexamethasone hydrochloride (41) (20 mg) obtained in Example 21 was dissolved and DMF (0.5 ml) in which HBTU ((100 mg) was dissolved, The reaction solution was dialyzed for 2 days at 4 ° C. against purified water using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). Was filtered through a membrane filter (0.22 μm) and lyophilized to obtain the title compound (43) (92 mg) .The amount of drug introduced into this complex was the UV absorbance at 240 nm and the total weight of the complex. From the above, it was 3.5% (% by weight).

Example 23 Production of 21-Leu-dexamethasone hydrochloride (44) (Step 1) Production of 20-BOC-Leu-dexamethasone (45) BOC-Leu · H 2 O (747 mg, 3 mmol) and dimethylaminopyridine (244 mg) 2 mmol) and dexamethasone (392 mg, 1 mmol) were dissolved in methylene chloride (30 ml). Then, DIPC (252 mg, 2 mmol) was added and stirred at room temperature for 13 hours. The reaction solution was washed with 0.1N hydrochloric acid and saturated brine, and then the solvent was distilled off under reduced pressure. Purification by silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / acetonitrile = 80/20), 20− BOC-Leu-dexamethasone (45) (605 mg) was obtained.

(Step 2) Production of 21-Leu-dexamethasone hydrochloride (44) Compound (45) (500 mg) was dissolved in methylene chloride (5 ml), trifluoroacetic acid (TFA; 1 ml) was added at room temperature, and the mixture was stirred for 2 hours. The solvent was distilled off. The de-BOC compound was subjected to silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 50 cm, eluent: acetonitrile / methanol / methylene chloride = 5 / 15/85), and then treated with an anion exchange resin to obtain the title compound (44) (510 mg). The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 0.79 (d, 3H) 0.88 (d, 3H) 0.89 (s, 3H) 0.92 (d, 3H) 1.08 (m, 1H) 1.34-1.81 (m, 6H) 1.49 (s, 3H) 1.86 (m, 1H) 2.1-2.88 (m, 6H) 3.35 (s, 2H) 44 (q, 1H) 4.16 (m, 1H) 4.85 (d, 1H) 5.02 (d, 1H) 5.17 (s, 1H) 5.47 (d, 1H) 6.01 ( s, 1H) 6.23 (dd, 1H) 7.31 (d, 1H)
¹³ C-NMR (DMSO-d6): 15.27, 16.30, 21.87, 23.11, 23.15, 24.15, 27.46, 30.50, 32.13, 33.79, 35.52, 35.79, 43.46, 43.56, 48.15, 48.16, 52.16, 68.21, 70.62, 90.67, 101.47, 124.24, 129. 06, 153.04, 167.27, 175.47, 185.48, 204.90

Example 24
Production of polyether-21-Leu-dexamethasone (46) The copolymer (15) (100 mg) obtained in Example 6 was dissolved in 20 ml of water, and 5 ml of DMF was added to this solution under ice cooling. To this solution was added DMF (0.5 ml) in which 21-Leu-dexamethasone hydrochloride (44) (20 mg) obtained in Example 23 was dissolved and DMF (0.5 ml) in which HBTU (100 mg) was dissolved. For 2 hours. This reaction solution was dialyzed against purified water at 4 ° C. for 2 days using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). The internal solution was filtered through a membrane filter (0.22 μm) and then lyophilized to obtain the title compound (46) (97 mg). The amount of drug introduced into this complex was 4.1% (% by weight) as calculated from the ultraviolet absorbance at 240 nm and the total weight of the complex.

Example 25
Production of 21-Ile-dexamethasone hydrochloride (47) (Step 1) Production of 20-BOC-Ile-dexamethasone (48)
BOC-Ile (1040 mg, 4.5 mmol), dimethylaminopyridine (366 mg, 3 mmol) and dexamethasone (588 mg, 1.5 mmol) were dissolved in methylene chloride (30 ml). Then, DIPC (378 mg, 3 mmol) was added and stirred at room temperature for 14 hours. The reaction solution was washed with 0.1N hydrochloric acid and saturated brine, and then the solvent was distilled off under reduced pressure. Purification by silica gel column chromatography (silica gel: Merck, Germany, Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 30 cm, eluent: methylene chloride / acetonitrile = 80/20), 20-BOC-Ile-dexamethasone (48) (620 mg) was obtained.

(Step 2) Production of 21-Ile-dexamethasone hydrochloride (47) Compound (48) (600 mg) was dissolved in methylene chloride (5 ml), trifluoroacetic acid (TFA; 1 ml) was added at room temperature, and the mixture was stirred for 2 hours. The solvent was distilled off. The de-BOC compound was subjected to silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 4.0 × 50 cm, eluent: acetonitrile / methanol / methylene chloride = 5 / 15/85) and then treated with an anion exchange resin to obtain the title compound (47) (584 mg). The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 0.80 (d, 3H) 0.90 (s, 3H) 0.93 (t, 3H) 1.03 (d, 3H) 1.08 (m, 1H) 1.33-1.86 (m, 6H) 1.49 (s, 3H) 1.50 (d, 1H) 2.13-2.91 (m, 6H) 4.11 (d, 1H) 16 (m, 1H) 4.99 (d, 1H) 5.17 (d, 1H) 5.26 (s, 1H) 5.55 (d, 1H) 6.01 (s, 1H) 6.23 ( dd, 1H) 7.34 (d, 1H) 8.44 (s, 1H)
¹³ C-NMR (DMSO-d6): 11.69, 13.69, 15.21, 16.21, 23.09, 25.02, 27.44, 30.45, 32.08, 33.72, 35.81, 35.61, 36.34, 43.44, 48.08, 48.22, 56.15, 69.54, 70.41, 90.72, 101.46, 124.19, 129. 00, 152.96, 167.18, 168.66, 185.41, 204.40

Example 26
Preparation of polyether-21-Ile-dexamethasone (49) The copolymer (15) (102 mg) obtained in Example 6 was dissolved in 5 ml of DMF. To this solution, DMF (0.5 ml) in which 21-Ile-dexamethasone hydrochloride (47) (20 mg) obtained in Example 25 was dissolved and DMF (0.5 ml) in which HBTU (100 mg) was dissolved were added. For 2 hours. This reaction solution was dialyzed against purified water at 4 ° C. for 2 days using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). The internal solution was filtered through a membrane filter (0.22 μm) and then lyophilized to obtain the title compound (49) (0.97 g). The amount of drug introduced into this complex was 3.0% (% by weight) as calculated from the ultraviolet absorbance at 240 nm and the total weight of the complex.

Example 27
Production of Ala-fasudil (50) (Step 1) Production of BOC-Ala-fasudil (51)
BOC-Ala (756 mg, 4 mmol), dimethylaminopyridine (122 mg, 1 mmol) and fasudil hydrochloride hydrate (673 mg, 2 mmol) were dissolved in dry methylene chloride (30 ml). Next, WSCD · HCl (water-soluble carbodiimide, Peptide Laboratories, 766 mg, 4 mmol) was added, and the mixture was stirred at room temperature overnight. The reaction solution was washed with saturated aqueous sodium hydrogen carbonate and saturated brine, and then the solvent was evaporated under reduced pressure. Purification by silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 2.0 × 30 cm, eluent: methylene chloride / methanol / acetonitrile = 95/5/5) As a result, BOC-Ala-fasudil (51) (713 mg) was obtained.

(Step 2) Production of Ala-fasudil (50) To compound (51) (650 mg), trifluoroacetic acid (TFA, 5 ml) was added at room temperature, and the solvent was distilled off after 10 minutes. Silica gel column chromatography (Silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 2.0 × 30 cm, eluent: acetonitrile / methanol / methylene chloride = 85 / 15/5) and then treated with an anion exchange resin to obtain the title compound (50) (504 mg). The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 1.30 (dd, 2H) 1.7-2.0 (2H) 3.2-3.8 (8H) 4.3 (1H) 7.9 (1H) 8.1 (2H) 8.3-8.4 (2H) 8.5 (1H) 8.7 (1H) 9.5 (1H)
¹³ C-NMR (DMSO-d6): 16.4, 16.7, 27.6, 29.5, 44.4-48.9, 117.0, 126.6, 128.8, 130.6, 132.3, 132.6, 133.5, 133.9, 144.8, 153.5, 169.4

Example 28
Production of polyether-Ala-fasudil (52) The copolymer (15) (104 mg) obtained in Example 6 was dissolved in 5 ml of DMF. To this solution were added DMF (0.5 ml) dissolved in Ala-fasudil (50) (10 mg) and DMF (0.5 ml) dissolved in HBTU (52 mg) obtained in Example 27, and the mixture was stirred at room temperature for 2 hours. did. This reaction solution was dialyzed against purified water at 4 ° C. for 2 days using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). The internal solution was filtered through a membrane filter (0.45 μm) and then lyophilized to obtain the title compound (52) (110 mg). The amount of drug introduced into this complex was 3.4% (% by weight) as calculated from the ultraviolet absorbance at 270 nm and the total weight of the complex.

Example 29
Production of Leu-Fasil (53) (Step 1) Production of BOC-Leu-Fasil (54)
BOC-Leu.H ₂ O (997 mg, 4 mmol), dimethylaminopyridine (122 mg, 1 mmol) and fasudil hydrochloride hydrate (673 mg, 2 mmol) were dissolved in dry methylene chloride (30 ml). Next, WSCD · HCl (water-soluble carbodiimide, Peptide Laboratories, 766 mg, 4 mmol) was added, and the mixture was stirred at room temperature overnight. The reaction solution was washed with saturated aqueous sodium hydrogen carbonate and saturated brine, and then the solvent was evaporated under reduced pressure. Purification by silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 2.0 × 30 cm, eluent: methylene chloride / methanol / acetonitrile = 95/5/5) To obtain BOC-Leu-fasudil (54) (718 mg).

(Step 2) Production of Leu-fasudil (53) To compound (54) (650 mg), trifluoroacetic acid (TFA, 5 ml) was added at room temperature, and the solvent was distilled off after 10 minutes. Silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 2.0 × 30 cm, eluent: acetonitrile / methanol / methylene chloride = 85 / 15/5) and then treated with an anion exchange resin to give the title compound (53) (525 mg). The NMR data in DMSO-d6 solvent using tetramethylsilane as a standard is shown below.
¹ H-NMR (DMSO-d6): δ 0.9 (6H) 1.4-2.0 (5H) 3.2-3.8 (8H) 4.3 (1H) 7.9 (1H) 8. 2 (2H) 8.3-8.4 (2H) 8.5 (1H) 8.7 (1H) 9.5 (1H)
¹³ C-NMR (DMSO-d6): 21.0, 21.2, 23.1, 23.4, 27.8, 29.4, 40.0, 45.9-48.6, 117.1 126.7, 128.8, 130.6, 132.5, 132.7, 133.5, 133.9, 144.6, 153.4, 169.1

Example 30
Production of polyether-Leu-fasudil (55) The copolymer (15) (101 mg) obtained in Example 6 was dissolved in 5 ml of DMF. To this solution were added DMF (0.5 ml) in which Leu-Fasdil (53) (10.6 mg) obtained in Example 29 was dissolved and DMF (0.5 ml) in which HBTU (52 mg) was dissolved. Stir. This reaction solution was dialyzed against purified water at 4 ° C. for 2 days using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). The internal solution was filtered through a membrane filter (0.45 μm) and then lyophilized to obtain the title compound (55) (106 mg). The amount of drug introduced into this complex was 3.4% (% by weight) as calculated from the ultraviolet absorbance at 270 nm and the total weight of the complex.

Example 31
Production of Ile-fasudil (56) (Step 1) Production of BOC-Leu-fasudil (57)
BOC-Ile (766 mg, 4 mmol), dimethylaminopyridine (122 mg, 1 mmol) and fasudil hydrochloride hydrate (673 mg, 2 mmol) were dissolved in dry methylene chloride (30 ml). Next, WSCD · HCl (water-soluble carbodiimide, Peptide Laboratories, 766 mg, 4 mmol) was added, and the mixture was stirred at room temperature overnight. The reaction solution was washed with saturated aqueous sodium hydrogen carbonate and saturated brine, and then the solvent was evaporated under reduced pressure. Purification by silica gel column chromatography (silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 2.0 × 30 cm, eluent: methylene chloride / acetonitrile = 70/30), BOC- Leu-fasudil (57) (826 mg) was obtained.

(Step 2) Production of Ile-fasudil (56) To the compound (57) (800 mg), trifluoroacetic acid (TFA, 5 ml) was added at room temperature, and the solvent was distilled off after 10 minutes. Silica gel column chromatography (Silica gel: Merck Art No. 9385, Silica gel 60, 200-400 mesh, column: 2.0 × 30 cm, eluent: acetonitrile / methanol / methylene chloride = 85 / 15/5), and then treated with an anion exchange resin to obtain the title compound (56) (763 mg). The NMR data and mass spectrometry data in DMSO-d6 solvent with tetramethylsilane as the standard are shown below.
¹ H-NMR (DMSO-d6): δ 0.85 (tt, 3H) 0.96 (dd, 3H) 1.14 (1H) 1.46 (1H) 1.7-2.0 (3H) 2-3.8 (8H) 4.2 (1H) 7.9 (1H) 8.1 (2H) 8.3-8.4 (2H) 8.5 (dd, 1H) 8.7 (dd, 1H) 9.5 (1H)
¹³ C-NMR (DMSO-d6): 11.2, 14.8, 22.9, 27.5, 29.7, 35.7, 35.9, 44.4, 45.9, 46.2, 46.7, 47.1, 47.3, 49.2, 53.8, 117.1, 126.6, 128.8, 130.6, 132.4, 132.6, 133.5, 133. 9, 144.6, 153.4, 168.2
MS: m / z 405 [M + H] ⁺

[Example 32]
Preparation of polyether-Ile-fasudil (58) The copolymer (15) (101 mg) obtained in Example 6 was dissolved in 5 ml of DMF. To this solution were added DMF (0.5 ml) in which Ile-fasudil (56) (10.6 mg) obtained in Example 29 was dissolved and DMF (0.5 ml) in which HBTU (52 mg) was dissolved. Stir. This reaction solution was dialyzed against purified water at 4 ° C. for 2 days using a dialysis membrane (molecular weight cut-off 12,000-14,000, manufactured by Spectrum, USA). The internal solution was filtered through a membrane filter (0.45 μm) and then freeze-dried to obtain the title compound (58) (109 mg). The amount of drug introduced into this complex was 3.7% (% by weight) as calculated from the ultraviolet absorbance at 270 nm and the total weight of the complex.

Example 33
Solubility of compounds (19), (21), (23), (25), (27) and (34) in physiological saline Compounds (19), (21), (23), (25) and (27) 10 mg each of (34) and (34) were weighed and added to 0.1 ml of physiological saline, and the added compounds were completely dissolved. In terms of paclitaxel, compound (19) had a solubility of 6.1 mg / ml (saline). Similarly, (21) is 3.7 mg / ml (saline), (23) is 4.0 mg / ml (saline), and (25) is 2.5 mg / ml (saline). (27) was 3.5 mg / ml (saline) and (34) was 6.4 mg / ml (saline).
Furthermore, 1 mg of paclitaxel (manufactured by HAUSER, USA) did not dissolve in 10 ml of physiological saline.

Example 34
Paclitaxel release evaluation experiment from compounds (19), (21), (23) and (25) in mouse and human plasma Compound (19) obtained in Example 9, Compound (21) obtained in Example 10 The compound (23) obtained in Example 11 and the compound (25) obtained in Example 12 were each dissolved in physiological saline and prepared so that the concentration in terms of paclitaxel was 125 μg / ml. 20 μl of these solutions were added to 200 μl of mouse and human plasma, respectively, and the amount of paclitaxel released from the drug complex at 37 ° C. was measured. In accordance with the method described in Pharmaceutical Journal, 114, 351-355 (1994), paclitaxel was collected from plasma and released from plasma from compounds (19), (21), (23) and (25) by HPLC. The amount of paclitaxel was evaluated.
FIG. 5 and FIG. 6 show changes with time in the release of paclitaxel. As a result, the release rate of paclitaxel from the drug complex showed the same tendency in mouse and human plasma. The order was compound (19)>(21)>(23)> (25) in descending order of release rate, and this was correlated with the size of the steric hindrance of the amino acid that is the linker.

Example 35
Antitumor effect evaluation experiment (1)
Colon 26 tumor cell 4% suspension was transplanted subcutaneously into the flank of a Balb / C female mouse (6 weeks old), and the compound (19) obtained in Example 9 and obtained in Example 10 were used as test compounds. The compound (21), the compound (23) obtained in Example 11, the test solution obtained by dissolving the compound (25) obtained in Example 12 in physiological saline, and paclitaxel were mixed with ethanol-Cremophor EL (Sigma, USA). (Manufactured)-The test solution dissolved in physiological saline was administered into the tail vein as 5 mice per group. The dose was 50 mg / kg in terms of paclitaxel. The untreated group was 9 per group. On the second day after cell transplantation, the first test solution was administered, and then every 7 days, the test solution was administered into the tail vein four times in total. Antitumor effects were determined by measuring the tumor volume of mice. The time-dependent change of the average tumor volume of the untreated group and the test liquid administration group was shown. The tumor volume V was determined by the following formula (XI) when the tumor was measured from the outside and the major axis was a (mm) and the minor axis was b (mm).

  The change over time in tumor volume after administration of the test solution was as shown in FIG. The antitumor effect of the drug conjugates (19) and (21) according to the present invention in the 50 mg / kg administration group was significantly superior to the antitumor effect of paclitaxel in the 50 mg / kg administration group. The strength of the antitumor effect was in the order of [compounds (19) and (21)> (23)> (25) and paclitaxel].

Example 36
Antitumor effect evaluation experiment (2)
Colon 26 tumor cell 4% suspension was transplanted subcutaneously into the flank of a Balb / C female mouse (6 weeks old), and the compound (34) obtained in Example 16 as a test compound was dissolved in physiological saline. The test solution prepared by dissolving paclitaxel in ethanol-Cremophor EL (manufactured by Sigma, USA) -physiological saline as a control was administered into the tail vein as a group of 3 mice. The dose was 50 mg / kg in terms of paclitaxel. There were 5 untreated groups per group. On the second day after tumor cell transplantation, the first test solution was administered, and then the test solution was administered into the tail vein seven times in total every four days. Antitumor effects were determined by measuring and comparing tumor volumes in mice. The tumor volume V was determined by the formula (XI) in Example 35 when the tumor was measured from the outside and made the major axis a (mm) and the minor axis b (mm). The time-dependent change of the average tumor volume of the untreated group and the test liquid administration group was shown.
The change over time in tumor volume after administration of the test solution was as shown in FIG. The antitumor effect of the drug conjugate (34) administration group of the present invention in the 50 mg / kg administration group was significantly superior to the antitumor effect of paclitaxel in the 50 mg / kg administration group.

Example 37
Solubility evaluation of compound (37), (40), (43), (46), (49), (52), (55) and (58) in physiological saline Compound (37), (40), (43 ), (46), (49), (52), (55) and (58) were weighed 10 mg each and added to 0.1 ml of physiological saline, and the added compounds were completely dissolved. Compound (37) had a solubility of 3.5 mg / ml (saline) in terms of camptothecin. Similarly, compound (40) has a solubility of 4.5 mg / ml (saline) in terms of camptothecin, compound (43) has a solubility of 3.5 mg / ml (saline) in terms of dexamethasone, and compound (46) has the same. In addition, the solubility of 4.1 mg / ml (physiological saline) in terms of dexamethasone and the compound (49) similarly had a solubility of 3.0 mg / ml (physiological saline) in terms of dexamethasone. Further, compound (52) has a solubility of 3.4 mg / ml (saline) in terms of fasudil, compound (55) has a solubility of 3.4 mg / ml (saline) in terms of fasudil, and compound (58) has fasudil. It was a solubility of 3.7 mg / ml (saline) in terms of conversion.

Example 38
Dexamethasone release evaluation experiment from compounds (43), (46) and (49) in mouse plasma Compound (43) obtained in Example 22, compound (46) obtained in Example 24 and obtained in Example 26 Compound (49) was dissolved in physiological saline to prepare 80 μg / ml in terms of dexamethasone. 50 μl of these solutions were added to 250 μl of mouse plasma, respectively, and the amount of dexamethasone released from each drug complex at 37 ° C. was measured. Dexamethasone in plasma was collected and quantified as follows. That is, 250 μl of phosphate buffer (pH 7.4) was added to 250 μl of a plasma sample, and acetonitrile-methanol solution (CH ₃ CN / MeOH = 4/1, hydrocortisone acetate concentration = 10 ng / concentration) containing hydrocortisone acetate as an internal standard. (3 ml), centrifuged (3000 rpm, 10 minutes, 4 ° C.), 700 μl of distilled water was added to 700 μl of the obtained supernatant, filtered through a membrane filter (0.4 μm), and then quantified by HPLC. did.
FIG. 9 shows the time course of dexamethasone release from mouse plasma. The order of the release rate of dexamethasone from the drug conjugate was compound (43)>(46)> (49) in the descending order of release rate, which was correlated with the steric hindrance magnitude of the linker amino acid. .
<HPLC conditions>
Column: Asahipak HIKARISIL C18 (4.6 x 150mm) Flow rate: 1.0ml / min
Column temperature: 25 ° C
Detection wavelength: 240 nm
Mobile phase: Linear gradient
0min: 20% acetonitrile aqueous solution (20% CH ₃ CN / H ₂ O)
20min: 50% acetonitrile aqueous solution (50% CH ₃ CN / H ₂ O)

Since the polyether of the present invention is excellent in biotissue compatibility and body fluid compatibility, when this is used as a medical material, high safety of the entire material is realized. In particular, the polyether of the present invention has very little interaction with a living body and is difficult to be recognized by living tissues, so it is rarely distributed in metabolic organs such as liver, spleen, and bone marrow, and side effect organs, resulting in organ damage and cytotoxicity. Can be accumulated in the target organ.
In addition, when a compound having a pharmacological activity is bound to the polyether via a linker such as an amino acid or a peptide and used as a drug complex, the drug is not recognized by the living tissue when it is administered in vivo. Can be delivered to the target tissue. The drug complex can control the drug release rate, and has extremely improved solubility in physiological saline compared to the drug before binding, and enables intravenous administration without a solubilizing agent.

6 is a graph showing an elution curve by GPC of the compound (13) of the present invention in Example 5. FIG. 6 is a graph showing an elution curve by GPC of the compound (15) of the present invention in Example 6. FIG. It is a figure which shows the elution curve by GPC of the compound (17) of this invention of Example 7. 4 is a graph showing toxicity evaluation of the compound (15) of the present invention in Example 8. 2 is a graph of drug release in mouse plasma of compounds (19), (21), (23) and (25) of Example 34 of the present invention. 2 is a graph of drug release in human plasma of compounds (19), (21), (23) and (25) of the present invention of Example 34. The compound (19) of the present invention of Example 9, the compound (21) of the present invention of Example 10, the compound (23) of the present invention of Example 11, and the compound (25) of the present invention of Example 12 against Colon 26 It is a graph showing an anti-tumor effect. 2 is a graph showing the antitumor effect of the compound (34) of the present invention of Example 16 on Colon 26. FIG. 3 is a graph of drug release in mouse plasma of the compound (43) of the present invention of Example 22, the compound (46) of the present invention of Example 24, and the compound (49) of the present invention of Example 26.

Claims (24)

A polyether comprising a repeating unit represented by the formula (III) and the formula (IV) obtained by copolymerizing the compounds represented by the formula (I) and the formula (II), and having the formula (A) The polyether which does not have a repeating unit represented by these.

(Wherein R ¹ represents a group represented by —OR ² or —NHR ³ , R ² and R ³ represent hydrogen, an inorganic base or an organic base, a group containing a C _{1 to} C ₁₅ hydrocarbon, an amino acid or Represents a compound having a pharmacological action to which a linker which is a peptide is bonded, wherein R ¹ , R ² or R ³ may be the same or different in the same copolymer, but in the same copolymer, A compound having a pharmacological action to which a linker, which is an amino acid or a peptide, is bonded is included, and the molar ratio of each structural unit is (III) / ((III) + (IV)) = 0. The molar percentages of the structural units (III) and (IV) are (III) = 0.1 to 99.9 and (IV) = 0.1 to 99.9, respectively. Formula (III) and Formula (IV) Polyethers containing as a structural unit include polyethylene glycol (PEG) equivalent weight average molecular weight of 1,000 to 100,000.) The polyether according to claim 1, wherein the polyethylene glycol (PEG) equivalent weight average molecular weight / PEG equivalent number average molecular weight (Mw / Mn) is 1.1 to 2.7. The polyether according to claim 1 or 2, wherein the linker is selected or used in combination from amino acids represented by Gly, Ala, Leu, Ile, and Phe. The linker is Gly-Gly, Ala-GLy, Gly-Ala, Leu-Gly, Gly-Leu, Ile-Gly, Gly-Ile, Phe-Gly, Gly-Phe, Gly-Gly-Gly, Gly-Phe-Gly, The polyether according to any one of claims 1 to 3, selected from Phe-Gly-Gly, Gly-Gly-Phe and Gly-Gly-Phe-Gly. The polyether according to any one of claims 1 to 4 , wherein the compound having a pharmacological action is an antineoplastic agent, an anti-inflammatory agent, an anti-rheumatic agent, an enzyme inhibitor or a nucleic acid. Antineoplastic agent is an alkylating agent, antimetabolite, antitumor antibiotics, microtubule inhibitors, hormone analogs agents, preparative topoisomerase inhibitors, biologic (cytokines), targeted therapy, nonspecific immunostimulation The polyether of claim 5 selected from drugs. The polyether according to claim 6, wherein the alkylating agent is selected from mustard agents or nitrosoureas. 7. The polyether according to claim 6, wherein the antimetabolite is selected from methotrexate, which is an antifolate, 5-fluorouracil, which is a pyrimidine antimetabolite, and 6-MP, which is an antipurine antimetabolite. The polyether according to claim 6, wherein the antitumor antibiotic is selected from anthracycline doxorubicin, daunorubicin and mitomycin C. The polyether according to claim 6, wherein the microtubule inhibitor is selected from taxane-based paclitaxel and docetaxel. 7. The polyether according to claim 6, wherein the topoisomerase inhibitor is selected from the topoisomerase I inhibitor irinotecan, nogitecan. Polyether of claim 6, targeted therapy imatinib is selected Gefichini blanking or al. The polyether according to claim 5, wherein the anti-inflammatory drug is selected from corticosteroids and non-steroidal anti-inflammatory drugs. 14. The polyether of claim 13 , wherein the corticosteroid is selected from hydrocortisone, cortisone, prednisolone, methylprednisolone, dexamethasone, betamethasone. 14. The polyether according to claim 13 , wherein the non-steroidal anti-inflammatory drug is selected from salicylic acid type aspirin, anthranilic mefenamic acid, propionic acid type ibuprofen, loxoprofen, arylacetic acid type diclofenac, and indomethacin. Polyether according to claim 5, antirheumatic drugs immunomodulatory agents are selected immunosuppressant or al. The polyether according to claim 16 , wherein the immunomodulator is selected from sodium gold thiomalate, penicillamine, lobenzarit. Polyether of claim 16, immunosuppressant mizoribine, Metotorekisa bets are selected from data Kurorimusu. The polyether according to claim 5, wherein the enzyme inhibitor is selected from fasudil or fasudil hydrochloride. Formulations containing polyether according to any of claims 1 to 19. The carrier for drug delivery to in vivo tissues using the polyether according to claim 1 , wherein R ² is hydrogen, an inorganic base, or an organic base. The method for producing a polyether according to any one of claims 1 to 19, comprising a repeating unit represented by formula (III) and formula (IV) and not having a repeating unit represented by formula (A). Because
The said manufacturing method including the process of carrying out the random copolymerization of the compound represented by Formula (I) and Formula (II).

(Wherein R ¹ represents a group represented by —OR ² or —NHR ³ , R ² and R ³ represent hydrogen, an inorganic base or an organic base, a group containing a C _{1 to} C ₁₅ hydrocarbon, an amino acid or The compound which has the pharmacological action which the linker which is a peptide couple | bonded is represented.
Here, R ¹ , R ^2, or R ³ may be the same or different in the same copolymer, but in the same copolymer, at least one pharmacology to which a linker that is an amino acid or a peptide is bound. The compound which has an effect | action shall be included . The molar ratio of each structural unit is (III) / ((III) + (IV)) = 0.001 to 0.5, and the molar percentage of the structural units (III) and (IV) is (III) = 0.1-99.9 and (IV) = 0.1-99.9. Here, the polyether containing formula (III) and formula (IV) as a structural unit has a weight average molecular weight in terms of polyethylene glycol (PEG) of 1,000 to 100,000. ) The method for producing a polyether according to claim 22 , wherein the catalyst used for the polymerization is selected from a base and a Lewis acid. The catalyst used for polymerization is potassium hydroxide, sodium hydroxide, cesium hydroxide, sodium methoxide, sodium ethoxide, sodium propoxide, sodium t-butoxide, potassium propoxide, potassium t-butoxide, potassium-t-2-methyl The method for producing a polyether according to claim 22 , wherein the polyether is selected from -2-butoxide and boron trifluoride ether complex, trialkylaluminum, triethylaluminum, triphenylaluminum, tributylaluminum, and used alone or in combination.

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