JP2009051784A - Radioactive substance that suppresses cancer and method for adjusting the same - Google Patents
Radioactive substance that suppresses cancer and method for adjusting the same Download PDFInfo
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Abstract
【課題】癌を抑制する放射性物質及びその調整方法の提供。
【解決手段】癌を抑制する放射性物質はM- SOCTA- Zで、Mは放射性核種で、188Re或いは99mTcで、ZはNH或いはNH2基団を含むアミノ酸のタンパク質あるいはペプチド類である。該癌を抑制する放射性物質の調整方法のステップは、SOCTAとタンパク質或いはペプチドを反応させ、SOCTAのエステル類構造(-COOR)とアミン類(-NH、- NH2)のボンド結合を利用し、ペプチドボンド(peptide bomd)のボンド結合を形成し、SOCTA-protein複合物を形成する。さらに該SOCTA-protein複合物と放射性核種Mを反応させ、M- SOCTA-protein或いはM- SOCTA-peptideを形成し、該放射性核種とタンパク質の特性を対応させ、M- SOCTA-protein或いはM- SOCTA-peptideを癌の診断或いは治療に用いることができる。本発明はハーセプチン(Herceptin)単株抗体の運用例であり、SOCTAの結合に、さらに放射線を組合せた治療方式に応用し、癌(乳癌など)の治療効果を高めることができる。
【選択図】図２A radioactive substance that suppresses cancer and a method for adjusting the same are provided.
The radioactive substance that suppresses cancer is M-SOCTA-Z, M is a radionuclide, 188 Re or 99m Tc, and Z is an amino acid protein or peptide containing NH or NH2 group. The step of the method for preparing a radioactive substance that suppresses cancer comprises reacting SOCTA with a protein or peptide, and utilizing the bond bond between the ester structure (-COOR) and amines (-NH, -NH2) of SOCTA, A bond bond of a peptide bomd is formed, and an SOCTA-protein complex is formed. Furthermore, the SOCTA-protein complex and radionuclide M are reacted to form M-SOCTA-protein or M-SOCTA-peptide, the characteristics of the radionuclide and protein are matched, and M-SOCTA-protein or M-SOCTA-peptide is associated. -peptide can be used for diagnosis or treatment of cancer. The present invention is an operation example of a Herceptin single strain antibody, and can be applied to a treatment system that combines SOCTA binding and radiation, thereby enhancing the therapeutic effect of cancer (such as breast cancer).
[Selection] Figure 2
Description
本発明は一種の癌を抑制する放射性物質及びその調整方法に関する。特に一種の188Re- SOCTA-Herceptinの放射性物質とその調整方法で、癌細胞(乳癌細胞など)の生長を阻害し、癌細胞を破壊し、癌(乳癌など)の治療効果を向上させる癌を抑制する放射性物質及びその調整方法に係る。 The present invention relates to a radioactive substance that suppresses a kind of cancer and a method for adjusting the same. In particular, with a kind of radioactive substance of 188 Re- SOCTA-Herceptin and its adjustment method, it inhibits the growth of cancer cells (breast cancer cells etc.), destroys cancer cells and improves the therapeutic effect of cancer (breast cancer etc.) The present invention relates to radioactive materials to be controlled and adjustment methods thereof.
女性は40歳以降、及び閉経後はホルモンの変化により乳癌、卵巣癌、子宮頸癌の発生率が増加する。乳癌は乳腺管細胞或いは腺泡細胞が異常な分裂を起こし繁殖し形成される悪性腫瘍である。これら悪性腫瘍は局部器官(乳房)を犯すだけでなく、離れた器官(骨格、肝臓、肺、脳など)に転移する可能性があり、身体の重要な器官の機能を破壊し、生命に危害を及ぼす。
台湾衛生署の統計データによれば、台湾の乳癌発生率と死亡率は年々増加しており、乳癌は、台湾女性の罹患する癌で2005年は第四位、欧米各国では第一位となっている。乳癌の悪化は多くの患者とその家庭に重大な生理的かつ心理的負担をもたらすため、台湾では既に乳癌は非常に重要な問題となっている。
乳癌の治療は伝統的には外科治療、補助薬物治療、ホルモン療法、放射線治療があるが、最近は乳癌の治療で画期的な進展があった。それが抗体治療である。これはハーセプチン(Herceptin)単株抗体を利用し、乳癌細胞上に位置するHer-2/neu(又はErb B-2)の抗原に対抗するものである。
In women after age 40 and after menopause, hormonal changes increase the incidence of breast, ovarian, and cervical cancer. Breast cancer is a malignant tumor that forms when breast duct cells or glandular foam cells divide and grow abnormally. These malignant tumors not only commit local organs (breasts), but can also metastasize to distant organs (skeletal, liver, lung, brain, etc.), destroying vital organ functions and harming life Effect.
According to statistics from the Taiwan Health Service, the incidence and mortality of breast cancer in Taiwan is increasing year by year. Breast cancer is the fourth most common cancer affected by Taiwanese women and the first in Europe and the United States. ing. Breast cancer has already become a very important issue in Taiwan because the worsening of breast cancer creates a significant physiological and psychological burden on many patients and their families.
Traditionally, there are surgical treatments, adjuvant medications, hormonal treatments, and radiation treatments for the treatment of breast cancer, but recent breakthroughs have been made in the treatment of breast cancer. That is antibody therapy. This uses a Herceptin single-strain antibody and opposes the antigen of Her-2 / neu (or Erb B-2) located on breast cancer cells.
1983年Haymanなど、ニワトリの肉瘤或いは赤芽球症(erythroblastosis)の細胞内において、炭化タンパクを発生する発癌遺伝子を発見し、erb B-2と命名したことをレポートで明らかにした。
1986年にはBargmaなどが、ラットの神経胚芽細胞瘤(neuroblastoma)内において、ニワトリ赤芽球症(erythroblastosis)にフェーズ似の発癌遺伝子を発見し、neu(erb B-2)と命名し、後に人類正常細胞内の上皮生長因子EGFR遺伝子と酷似した発癌遺伝子HER-2(human epidermal growth factor receptor-2)を発見した。このため、正式名称をC-erb-B2/Her-2/neuとし、略称をHer-2/neuと命名した。この遺伝子のタンパク質産物はp185HER-2で、乳癌患者の癌細胞内においては約30%のHer-2/neuの突然変異現象が見られる。よってこの発癌遺伝子は乳癌細胞を確認するための重要な指標の一つとなっている。さらにHer-2/neuの高発癌は非小細胞肺癌、胃癌、前立腺癌、卵巣癌、子宮頸癌などの他の癌でもしばしば見られる。しかもHer-2/neu高発癌は若干の癌では予後の重要な因子と見なされる。
In 1983, Hayman et al. Found in a report that an oncogene that produces carbonized protein was found in cells of chicken lumps or erythroblastosis and named erb B-2.
In 1986, Bargma and others found an oncogene similar to chicken erythroblastosis in a rat neuroblastoma and named it neu (erb B-2). An oncogene HER-2 (human epidermal growth factor receptor-2), which is very similar to the epidermal growth factor EGFR gene in normal human cells, was discovered. For this reason, the official name was C-erb-B2 / Her-2 / neu and the abbreviation was Her-2 / neu. The protein product of this gene is p185HER-2, and about 30% of Her-2 / neu mutations are observed in cancer cells of breast cancer patients. Therefore, this oncogene has become one of the important indicators for confirming breast cancer cells. In addition, high Her-2 / neu carcinogenesis is often seen in other cancers such as non-small cell lung cancer, gastric cancer, prostate cancer, ovarian cancer, cervical cancer. Moreover, high Her-2 / neu carcinogenesis is considered an important prognostic factor in some cancers.
Baselgaなどの学者は先ず、ハーセプチン(Herceptin)を用い実験を行った。46例の転移性乳癌にHer-2/neuの高発癌が合わさった患者に治療を施し、その反応率は11.6%であった。続いて、より規模の大きい第二レベルの臨床実験を行い、222例の類似の患者が化学治療で効果を得られなかった後にハーセプチン(Herceptin)治療を受け、反応率は16%であり、制御期間は9.1ヶ月であった。
16%の反応率は抗癌薬物としては話しにならないほどに低いが、実験に参加したすべての患者は多種の化学治療を受け効果が出なかった訳で、その癌細胞は非常に頑強である。よって、このような結果は重大な進歩と言うことができる。
独特の抗癌治療効果及び転機によりハーセプチン(Herceptin)は既に米国では優先審査薬品に列記されている。また1998年9月末、米国食品薬物管理局(FDA)から販売許可を受け、発癌遺伝子Her-2/neu oncogene高発癌型で、しかも一種(或いは一種以上)の化学療法で効果が見られない転移性乳癌患者の治療に用いられている。本研究はハーセプチン(Herceptin)に放射免疫治療方式を組合せ、188Reが放出するβ粒子を利用し乳癌細胞を破壊し、治療効果を増強する。
ハーセプチン(Herceptin)は乳癌Her-2/neu高発癌型患者に対して非常に高い治療効果を有する。単一薬物は約30%の腫瘍緩解率を示し、Her-2/neu receptorのジメライゼイション (dimerization)及びリン酸化を抑制することができる。こうして癌細胞生長を遅らせ、さらには死亡に至らせる。
ハーセプチン(Herceptin)と伝統的な化学療法の併用は、乳癌に対して高い制御効果を発揮する。アンスラサイクリンとシクロフォスファミド( Anthracyclines and cyclophosphamide)などの伝統的化学療法は臨床報告で転移線乳癌患者の病状制御に非常に有効であることが臨床報告に記載されている。
Scholars such as Baselga first experimented with Herceptin. Forty-six patients with metastatic breast cancer combined with high incidence of Her-2 / neu were treated with a response rate of 11.6%. Following a larger second-level clinical experiment, 222 similar patients were treated with Herceptin after being ineffective with chemotherapy, with a response rate of 16% and control. The period was 9.1 months.
Although the response rate of 16% is low enough to be unspoken as an anticancer drug, all the patients who participated in the experiment did not respond to various chemotherapy treatments, and the cancer cells were very robust . Thus, such a result can be said to be a significant advance.
Herceptin has already been listed as a priority drug in the United States due to its unique anti-cancer therapeutic effect and turning point. In addition, at the end of September 1998, the US Food and Drug Administration (FDA) received marketing permission, and the oncogene Her-2 / neu oncogene is highly carcinogenic and has no effect with one (or more) types of chemotherapy. It is used to treat patients with sexual breast cancer. In this study, Herceptin is combined with a radioimmunotherapy method to destroy breast cancer cells using β particles released by 188 Re and enhance the therapeutic effect.
Herceptin has a very high therapeutic effect on breast cancer Her-2 / neu highly carcinogenic patients. A single drug has a tumor remission rate of about 30% and can inhibit Her-2 / neu receptor dimerization and phosphorylation. In this way, cancer cell growth is delayed and even death is caused.
Herceptin (Herceptin) and traditional chemotherapy are highly effective in controlling breast cancer. Clinical reports describe that traditional chemotherapies such as anthracyclines and cyclophosphamide are very effective in controlling the pathology of patients with metastatic breast cancer.
188Reは188W子核種で、半減期は16.9時間である。放射可能なβ粒子は2.2MeV、γ線155keVに達し、診断と治療効果を兼ね備える数少ない放射性同位元素である。SOCTAは本研究機関核エネルギー研究所化学組の林正憲博士が合成した新双官能基有機配位子で、その化学構造中には活性化したカルボン酸を含み、タンパク質或いはペプチドをボンド結合することができる(本発明中では単株抗体ハーセプチン(Herceptin)をボンド結合する)。また、N2S2配位子を有しレニウムとテクネチウムをボンド結合することができる。よってタンパク質或いはペプチドをレニウム或いはテクネチウムにより標識とする化合物SOCTAは適当な選択である。 188 Re is a 188 W nuclide with a half-life of 16.9 hours. The radiable β particles reach 2.2 MeV and γ-ray 155 keV, and are one of the few radioactive isotopes that have both diagnostic and therapeutic effects. SOCTA is a new bifunctional organic ligand synthesized by Dr. Masanori Hayashi of the Chemical Institute of the Nuclear Energy Research Institute of this research institution. It contains an activated carboxylic acid in its chemical structure and bonds a protein or peptide. (In the present invention, the single antibody Herceptin is bonded with a bond). It also has an N 2 S 2 ligand and can bond bond rhenium and technetium. Therefore, the compound SOCTA that labels proteins or peptides with rhenium or technetium is an appropriate choice.
本発明の主要目的は、単株抗体ハーセプチン(Herceptin)の抗癌効果を高めるために、癌を抑制する放射性物質及びその調整方法を提供し、ハーセプチン(Herceptin)を利用し乳癌細胞上のHer-2/neu発癌遺伝子の作用を抑制し、ハーセプチン(Herceptin)とSOCTAをボンド結合させ、SOCTAを放射性同位元素M(188Reなど)にボンド結合し、188Re-SOCTA-Herceptinの構造を形成し、該構造上の同位元素188Reを利用し2.2MeVのβ粒子を放射し、組織貫通が6.4-11mmである特徴とハーセプチン(Herceptin)の既存の抗癌特性により、癌細胞(乳癌細胞など)の成長を阻害し、癌細胞を破壊し、癌(乳癌など)治療の効果を達成する。 The main object of the present invention is to provide a radioactive substance that suppresses cancer and a method for adjusting the same in order to enhance the anticancer effect of the single antibody Herceptin (Herceptin), and use Herceptin to make Her- 2 / neu suppresses the oncogene action, bonds Herceptin (Herceptin) and SOCTA, bonds SOCTA to radioisotope M ( 188 Re, etc.), forms the structure of 188 Re-SOCTA-Herceptin, Using the structural isotope 188 Re to emit 2.2 MeV β particles, tissue penetration is 6.4-11 mm and the existing anticancer properties of Herceptin (Herceptin), cancer cells (such as breast cancer cells) Inhibits growth, destroys cancer cells and achieves the effect of treating cancer (such as breast cancer).
本発明の次要目的は、癌を抑制する放射性物質及びその調整方法を提供する。
該癌を抑制する放射性物質は放射性元素及び単株抗体ハーセプチン(Herceptin)を備えるため、癌細胞(乳癌細胞など)の生長抑制を強化し癌細胞を破壊し、癌(乳癌など)の治療効果を向上させることができる。
また該癌を抑制する放射性物質の調整方法は一段式の合成ステップを使用し、癌を抑制する放射性物質(188Re-SOCTA-Herceptinなど)の合成時間を短縮可能で、該癌を抑制する放射性物質の調整生産率を高めることができる。
該癌を抑制する放射性物質は以下の化学構造式１を含み、内、Mは188Re及び99mTcのグループの内の一つで、Zはアミン類基団を含むアミノ酸のタンパク質及びアミン類基団を含むアミノ酸のペプチドグループの内の一つである。
該癌を抑制する放射性物質の調整方法のステップは、SOCTAとタンパク質或いはペプチドを反応させ、SOCTA複合物を形成し、該SOCTA複合物はSOCTA-protein複合物及びSOCTA-peptide複合物のグループの内の一つで、及び該SOCTA複合物とMを反応させ、M-SOCTA複合物を形成し、該Mは188Re及び99mTcのグループの内の一つで、しかも該M-SOCTA複合物は188Re-SOCTA-protein複合物、188Re-SOCTA-peptide複合物、99mTc-SOCTA-protein複合物、99mTc-SOCTA-peptide複合物のグループの内の一つで、しかも該M-SOCTA複合物は癌を抑制する放射性物質である。
Since the radioactive substance that suppresses the cancer comprises a radioactive element and the single antibody Herceptin (Herceptin), it suppresses the growth of cancer cells (breast cancer cells, etc.), destroys the cancer cells, and has a therapeutic effect on cancer (breast cancer, etc.). Can be improved.
In addition, the method for preparing a radioactive substance that suppresses cancer uses a one-step synthesis step, and the synthesis time of a radioactive substance that suppresses cancer (such as 188 Re-SOCTA-Herceptin) can be shortened. Increase the production rate of substances.
The radioactive substance that suppresses the cancer includes the following chemical structural formula 1, wherein M is one of the group of 188 Re and 99m Tc, and Z is an amino acid protein and amine group containing an amine group. It is one of the peptide groups of amino acids containing groups.
The step of the method for preparing a radioactive substance that suppresses cancer comprises reacting SOCTA with a protein or peptide to form an SOCTA complex, and the SOCTA complex is selected from the group of SOCTA-protein complex and SOCTA-peptide complex. And reacting the SOCTA complex with M to form an M-SOCTA complex, wherein M is one of the group of 188 Re and 99m Tc, and the M-SOCTA complex is One of the group of 188 Re-SOCTA-protein complex, 188 Re-SOCTA-peptide complex, 99m Tc-SOCTA-protein complex, 99m Tc-SOCTA-peptide complex, and the M-SOCTA complex Things are radioactive substances that suppress cancer.
請求項１の発明は、癌を抑制する放射性物質は以下の化学構造式２を含み、内、Mは188Re及び99mTcのグループの内の一つで、Zはアミン類基団を含むアミノ酸のタンパク質及びアミン類基団を含むアミノ酸のペプチドグループの内の一つであることを特徴とする癌を抑制する放射性物質としている。
請求項３の発明は、請求項１記載の癌を抑制する放射性物質において、前記アミン類基団のアミノ酸はリシン(Lysine)、アルギニン(Arginine)、グルタミン(Glutamine)、アスパラギン(Asparagine) のグループの内の一つであることを特徴とする癌を抑制する放射性物質としている。
請求項４の発明は、請求項１記載の癌を抑制する放射性物質において、前記アミン類基団のアミノ酸のタンパク質は単株抗体であることを特徴とする癌を抑制する放射性物質としている。
請求項５の発明は、請求項４記載の癌を抑制する放射性物質において、前記単株抗体はハーセプチン(Herceptin)単株抗体であることを特徴とする癌を抑制する放射性物質としている。
In the invention of claim 1, the radioactive substance that suppresses cancer comprises the following chemical structural formula 2, wherein M is one of the group of 188 Re and 99m Tc, and Z is an amino acid containing an amine group. It is a radioactive substance that suppresses cancer, which is one of a peptide group of amino acids containing a protein and an amine group.
According to a third aspect of the present invention, in the radioactive substance that suppresses cancer according to the first aspect, the amino acids of the amine group are lysine (Lysine), arginine (Arginine), glutamine (Glutamine), asparagine (Asparagine). It is a radioactive substance that suppresses cancer characterized by being one of them.
According to a fourth aspect of the present invention, there is provided the radioactive substance for suppressing cancer according to the first aspect, wherein the amino acid protein of the amine group is a single strain antibody.
According to a fifth aspect of the present invention, there is provided the radioactive substance that suppresses cancer according to the fourth aspect, wherein the single strain antibody is a Herceptin single strain antibody.
請求項６の発明は、癌を抑制する放射性物質の調整方法のステップは、
SOCTAとタンパク質或いはペプチドを反応させ、SOCTA複合物を形成し、該SOCTA複合物はSOCTA-protein複合物及びSOCTA-peptide複合物のグループの内の一つで、
該SOCTA複合物とMを反応させM- SOCTA複合物を形成し、該Mは188Re及び99mTcのグループの内の一つで、しかも該M- SOCTA複合物は188Re-SOCTA-protein複合物、188Re-SOCTA-peptide複合物、99mTc-SOCTA-protein複合物、99mTc -SOCTA-peptide複合物のグループの内の一つで、
該M- SOCTA複合物は癌を抑制する放射性物質であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項７の発明は、請求項６記載の癌を抑制する放射性物質の調整方法において、前記タンパク質はハーセプチン(Herceptin)単株抗体で、SOCTA-protein複合物はSOCTA-Herceptin複合物で、M- SOCTA-protein複合物は188Re-SOCTA-Herceptin複合物であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項８の発明は、請求項７記載の癌を抑制する放射性物質の調整方法において、前記SOCTAと該ハーセプチン(Herceptin)単株抗体反応が該SOCTA-Herceptin複合物を形成するステップにおいて、該SOCTAと該ハーセプチン(Herceptin)は結合反応を行うことを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項９の発明は、請求７の癌を抑制する放射性物質の調整方法において、前記SOCTAと該ハーセプチン(Herceptin)単株抗体反応が該SOCTA-Herceptin複合物を形成するステップにおいては、さらに該SOCTAをジメチルホルムアミド(DMF)溶液に溶解させ、該ハーセプチン(Herceptin)をN-ジハイドロキエチルピレラジン-N'-ジエタンサルフォニック酸(N-[2-Hydroxyethyl]piperazine-N'-[2-ethanesulfonic acid] )緩衝液(HEPES Buffer)に溶解させるステップを含むことを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１０の発明は、請求項７記載の癌を抑制する放射性物質の調整方法において、前記SOCTAと該ハーセプチン(Herceptin)単株抗体反応が該SOCTA-Herceptin複合物を形成するステップにおいて、該SOCTAと該ハーセプチン(Herceptin)の最適モル数比は1：1であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１１の発明は、請求項７記載の癌を抑制する放射性物質の調整方法において、前記SOCTAと該ハーセプチン(Herceptin)単株抗体反応が該SOCTA-Herceptin複合物を形成するステップは室温下で反応させることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１２の発明は、請求項７記載の癌を抑制する放射性物質の調整方法において、前記SOCTAと該ハーセプチン(Herceptin)単株抗体反応が該SOCTA-Herceptin複合物を形成するステップには透析のステップを含むことを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１３の発明は、請求項７記載の癌を抑制する放射性物質の調整方法において、前記SOCTA-Herceptin複合物と該188Reを反応させ該188Re-SOCTA-Herceptin複合物を形成するステップにおいて、該188Reは188Re高レニウム酸塩溶液(188Re perrhenate solution)により提供することを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１４の発明は、請求項７記載の癌を抑制する放射性物質の調整方法において、前記SOCTA-Herceptin複合物と該188Reを反応させ該188Re-SOCTA-Herceptin複合物を形成するステップにおいては、さらに
先ず該188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)と塩化スズ(SnCl2)を反応させ、188Re-Glucoheptonateを得、
該188Re-Glucoheptonateと該SOCTA-Herceptinを反応させ、該188Re-SOCTA-Herceptinを得る二段式ステップを含むことを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１５の発明は、請求項１４記載の癌を抑制する放射性物質の調整方法において、前記188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)と塩化スズ(SnCl2)を反応させ188Re-Glucoheptonateを得るステップ中の該グルコヘプトネートと該塩化スズの重量費は1.5：1〜5.4：1であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１６の発明は、請求項１５記載の癌を抑制する放射性物質の調整方法において、前記188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)と塩化スズ(SnCl2)を反応させ188Re-Glucoheptonateを得るステップ中の該グルコヘプトネートと該塩化スズの最適重量比は1.6：1であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１７の発明は、請求項１４記載の癌を抑制する放射性物質の調整方法において、前記188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)と塩化スズ(SnCl2)を反応させ188Re-Glucoheptonateを得るステップ中の該グルコヘプトネート(Glucoheptonate)はキレート剤であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１８の発明は、請求項１４記載の癌を抑制する放射性物質の調整方法において、前記188Re-Glucoheptonateと該SOCTA-Herceptinを反応させ該188Re-SOCTA-Herceptinを得るステップの反応温度は室温或いは37℃であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項１９の発明は、請求項７記載の癌を抑制する放射性物質の調整方法において、前記SOCTA-Herceptin複合物と該188Reを反応させ該188Re-SOCTA-Herceptin複合物を形成するステップ中には、該188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)、塩化スズ(SnCl2)及び該SOCTA-Herceptin複合物に一段式反応を行わせ、188Re-SOCTA-Herceptin複合物を得る最適ステップを含むことを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項２０の発明は、請求項１９記載の癌を抑制する放射性物質の調整方法において、前記188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)、塩化スズ(SnCl2)及び該SOCTA-Herceptin複合物に一段式反応を行わせ、188Re-SOCTA-Herceptin複合物を得る最適ステップ中の該グルコヘプトネートと該塩化スズの重量比は1.5：1〜5.4：1であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項２１の発明は、請求項２０記載の癌を抑制する放射性物質の調整方法において、前記188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)、塩化スズ(SnCl2)及び該SOCTA-Herceptin複合物に一段式反応を行わせ、188Re-SOCTA-Herceptin複合物を得る最適ステップ中の該グルコヘプトネートと該塩化スズの重量比は1.6：1であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項２２の発明は、請求項１９記載の癌を抑制する放射性物質の調整方法において、前記188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)、塩化スズ(SnCl2)及び該SOCTA-Herceptin複合物に一段式反応を行わせ、188Re-SOCTA-Herceptin複合物を得る最適ステップ中の反応温度は室温或いは37℃であることを特徴とする癌を抑制する放射性物質の調整方法としている。
請求項２３の発明は、請求項６記載の癌を抑制する放射性物質の調整方法において、前記SOCTAと該タンパク質或いは該ペプチドを反応させ、該SOCTA複合物を形成するステップ中において、該SOCTAのエステル類構造(-COOR)とタンパク質或いはペプチドボンド結合のアミン類(-NH、- NH2)を反応させ、ペプチドボンド(peptide bomd)のボンド結合を形成し、該SOCTA複合物を形成することを特徴とする癌を抑制する放射性物質の調整方法としている。
In the invention of claim 6, the step of the method for adjusting a radioactive substance that suppresses cancer comprises:
SOCTA and protein or peptide are reacted to form an SOCTA complex, which is one of the group of SOCTA-protein complex and SOCTA-peptide complex,
The SOCTA complex is reacted with M to form an M-SOCTA complex, wherein M is one of the group of 188 Re and 99m Tc, and the M-SOCTA complex is 188 Re-SOCTA-protein complex. things, 188 Re-SOCTA-peptide complexes, 99m Tc-SOCTA-protein conjugate, in one of the groups of 99m Tc -SOCTA-peptide complexes,
The M-SOCTA composite is a radioactive substance that suppresses cancer, which is a radioactive substance that suppresses cancer.
The invention of claim 7 is the method for preparing a radioactive substance that suppresses cancer according to claim 6, wherein the protein is a Herceptin single-strain antibody, the SOCTA-protein complex is a SOCTA-Herceptin complex, M- The SOCTA-protein complex is a 188 Re-SOCTA-Herceptin complex, which is a method for preparing a radioactive substance that suppresses cancer.
The invention of claim 8 is the method for preparing a radioactive substance that suppresses cancer according to claim 7, wherein the SOCTA and the Herceptin single-strain antibody reaction form the SOCTA-Herceptin complex in the step of forming the SOCTA-Herceptin complex. And Herceptin (Herceptin) is a method for preparing a radioactive substance that suppresses cancer characterized by performing a binding reaction.
The invention of claim 9 is the method for preparing a radioactive substance that suppresses cancer of claim 7, wherein the SOCTA and Herceptin single-strain antibody reaction further forms the SOCTA-Herceptin complex. Is dissolved in a dimethylformamide (DMF) solution, and the Herceptin is dissolved in N-dihydroxyethylpyrazine-N'-diethanesulfonic acid (N- [2-Hydroxyethyl] piperazine-N '-[2- ethanesulfonic acid]) a method of preparing a radioactive substance that suppresses cancer, comprising a step of dissolving in a buffer solution (HEPES Buffer).
The method of preparing a radioactive substance that suppresses cancer according to claim 7, wherein the SOCTA and the Herceptin single-strain antibody reaction form the SOCTA-Herceptin complex in the step of forming the SOCTA-Herceptin complex. The optimal molar ratio of Herceptin to Herceptin is 1: 1, which is a method for preparing a radioactive substance that suppresses cancer.
The invention according to claim 11 is the method of preparing a radioactive substance that suppresses cancer according to claim 7, wherein the step of forming the SOCTA-Herceptin complex by the SOCTA and Herceptin single-strain antibody reaction at room temperature It is a method for adjusting a radioactive substance that suppresses cancer, characterized by reacting.
The invention of claim 12 is the method for preparing a radioactive substance that suppresses cancer according to claim 7, wherein the step of reacting the SOCTA with the Herceptin single strain antibody to form the SOCTA-Herceptin complex involves dialysis. It is set as the adjustment method of the radioactive substance which suppresses the cancer characterized by including a step.
The invention of claim 13, in the adjustment method of inhibiting radioactive substances cancer according to claim 7, wherein in the step of forming the 188 Re-SOCTA-Herceptin conjugate by reacting the SOCTA-Herceptin composite and the 188 Re The 188 Re is provided by a 188 Re high rhenate solution ( 188 Re perrhenate solution), and is a method for preparing a radioactive substance that suppresses cancer.
The invention of claim 14 is the method of adjusting inhibiting radioactive substances cancer according to claim 7, wherein in the step of forming the 188 Re-SOCTA-Herceptin conjugate by reacting the SOCTA-Herceptin composite and the 188 Re Is further reacted with the 188 rhenium ( 188 Re), glucoheptonate (Glucoheptonate) and tin chloride (SnCl 2 ) to obtain 188 Re-Glucoheptonate,
It is a method for preparing a radioactive substance that suppresses cancer, comprising a two-stage step of reacting the 188 Re-Glucoheptonate and the SOCTA-Herceptin to obtain the 188 Re-SOCTA-Herceptin.
The invention of claim 15 is the method of adjusting inhibiting radioactive substances cancer of claim 14, wherein 188 rhenium (188 Re), glucoheptonate and (Glucoheptonate) is reacted with tin chloride (SnCl 2) 188 Re In the step of obtaining -Glucoheptonate, the weight cost of the glucoheptonate and the tin chloride is 1.5: 1 to 5.4: 1.
The invention of claim 16 is the method of adjusting inhibiting radioactive substances cancer according to claim 15, wherein 188 rhenium (188 Re), glucoheptonate and (Glucoheptonate) is reacted with tin chloride (SnCl 2) 188 Re -The optimal weight ratio of the glucoheptonate to the tin chloride in the step of obtaining -Glucoheptonate is 1.6: 1.
The invention of claim 17, in the adjustment method of inhibiting radioactive substances cancer of claim 14, wherein 188 rhenium (188 Re), glucoheptonate and (Glucoheptonate) is reacted with tin chloride (SnCl 2) 188 Re -The glucoheptonate (Glucoheptonate) in the step of obtaining Glucoheptonate is a chelating agent, which is a method for preparing a radioactive substance that suppresses cancer.
The invention of claim 18 is the method for preparing a radioactive substance that suppresses cancer according to claim 14, wherein the reaction temperature of the step of reacting the 188 Re-Glucoheptonate with the SOCTA-Herceptin to obtain the 188 Re-SOCTA-Herceptin is It is a method for preparing a radioactive substance that suppresses cancer, characterized by being at room temperature or 37 ° C.
The invention of claim 19, in the adjustment method of inhibiting radioactive substances cancer according to claim 7, wherein the step of forming the SOCTA-Herceptin composite and the 188 Re-SOCTA-Herceptin conjugate by reacting the 188 Re The 188 rhenium ( 188 Re), glucoheptonate (Glucoheptonate), tin chloride (SnCl 2 ) and the SOCTA-Herceptin complex are subjected to a one-step reaction to obtain a 188 Re-SOCTA-Herceptin complex. It is the adjustment method of the radioactive substance which suppresses the cancer characterized by including the optimal step.
The invention of claim 20 is the method for preparing a radioactive substance that suppresses cancer according to claim 19, wherein the 188 rhenium ( 188 Re), glucoheptonate, tin chloride (SnCl 2 ) and the SOCTA-Herceptin Wherein the weight ratio of the glucoheptonate to the tin chloride is 1.5: 1 to 5.4: 1 in the optimal step of obtaining a 188 Re-SOCTA-Herceptin complex by performing a one-step reaction on the complex. It is a method for adjusting radioactive substances that suppress cancer.
The invention of claim 21 is the method for preparing a radioactive substance that suppresses cancer according to claim 20, wherein the 188 rhenium ( 188 Re), glucoheptonate, tin chloride (SnCl 2 ) and the SOCTA-Herceptin Inhibiting cancer, characterized in that the weight ratio of the glucoheptonate to the tin chloride is 1.6: 1 during the optimal step of obtaining a 188 Re-SOCTA-Herceptin complex by performing a one-step reaction on the complex. It is a method of adjusting the radioactive material to be used.
The invention of claim 22 is the method for preparing a radioactive substance that suppresses cancer according to claim 19, wherein the 188 rhenium ( 188 Re), glucoheptonate, tin chloride (SnCl 2 ) and the SOCTA-Herceptin It is a method for preparing a radioactive substance that suppresses cancer, characterized in that the reaction temperature during the optimal step of obtaining a 188 Re-SOCTA-Herceptin complex by allowing the complex to undergo a one-step reaction is room temperature or 37 ° C.
23. The method for preparing a radioactive substance for suppressing cancer according to claim 6, wherein in the step of reacting the SOCTA with the protein or the peptide to form the SOCTA complex, an ester of the SOCTA is prepared. Characterized by reacting a similar structure (-COOR) with a protein or peptide bond bond amine (-NH, -NH2) to form a peptide bond (peptide bomd) bond to form the SOCTA complex. It is a method for adjusting radioactive substances that suppress cancer.
本発明癌を抑制する放射性物質及びその調整方法の癌を抑制する放射性物質の調整方法は一段式合成ステップを使用し、癌を抑制する放射性物質(188Re-SOCTA-Herceptinなど)合成の時間を短縮し、その生産率を高めることができる。該癌を抑制する放射性物質は放射性元素及び単株抗体ハーセプチン(Herceptin)を備えるため、癌細胞(乳癌細胞など)の生長抑制を強化し、癌細胞を破壊することができ、癌(乳癌など)の治療効果を向上させることができる。 The method for preparing a radioactive substance that suppresses cancer according to the present invention and a method for adjusting the same use a one-step synthesis step, and the time for synthesizing a radioactive substance that suppresses cancer (such as 188 Re-SOCTA-Herceptin) is used. It can be shortened and the production rate can be increased. Since the radioactive substance that suppresses the cancer comprises a radioactive element and a single strain antibody Herceptin (Herceptin), it can strengthen the growth suppression of cancer cells (such as breast cancer cells), can destroy the cancer cells, cancer (such as breast cancer) Can improve the treatment effect.
本発明癌を抑制する放射性物質は以下の化学構造式３を含み、内、Mは188Re及び99mTcのグループの内の一つで、Zはアミン類基団を含むアミノ酸のタンパク質及びアミン類基団を含むアミノ酸のペプチドグループの内の一つである。該アミン類基団は-NH及び-NH2の内の一つである。該アミン類基団のアミノ酸はリシン(Lysine)、アルギニン(Arginine)、グルタミン(Glutamine)、アスパラギン(Asparagine) のグループの内の一つで、該アミン類基団を含むアミノ酸のタンパク質は単株抗体で、それはハーセプチン(Herceptin)単株抗体である。
SOCTA(図１参照)は本発明合成の新双官能基有機配位子で、その化学構造中には活性化したカルボン酸を含み、タンパク質或いはペプチドをボンド結合することができる(本発明中では単株抗体ハーセプチン(Herceptin)をボンド結合する)。またN2S2配位子を有しレニウムとテクネチウムをボンド結合することができる。よってタンパク質或いはペプチドをレニウム或いはテクネチウムにより標識とする化合物SOCTAは適当な選択である。 SOCTA (see Fig. 1) is a new bifunctional organic ligand synthesized according to the present invention, which contains an activated carboxylic acid in its chemical structure and can bond proteins or peptides (in the present invention). Single-strain antibody Herceptin (bonded to Herceptin). It also has an N 2 S 2 ligand and can bond bond rhenium and technetium. Therefore, the compound SOCTA that labels proteins or peptides with rhenium or technetium is an appropriate choice.
本発明癌を抑制する放射性物質の調整方法は以下のステップを含む(図２参照)。
S1: SOCTAとタンパク質或いはペプチドを反応させSOCTA複合物を形成し、該SOCT
複合物はSOCTA-protein(SOCTA-プロテイン)複合物及びSOCTA-peptide(SOCTA-ペプチド)複合物のグループの内の一つである。
S2:該SOCTA複合物とMを反応させ、M-SOCTA複合物を形成し、該Mは188Re及び99mTcのグループの内の一つで、しかも該M-SOCTA複合物は188Re-SOCTA-protein複合物、188Re-SOCTA-peptide複合物、99mTc-SOCTA-protein複合物、99mTc-SOCTA-peptide複合物のグループの内の一つで、しかも該M-SOCTA複合物は癌を抑制する放射性物質である。
該タンパク質はハーセプチン(Herceptin)単株抗体で、SOCTA-protein複合物はSOCTA-Herceptin複合物及びM-SOCTA- protein複合物は188Re-SOCTA-Herceptin複合物である。
The preparation method of the radioactive substance which suppresses this invention cancer includes the following steps (refer FIG. 2).
S1: SOCTA and protein or peptide react to form a SOCTA complex, and the SOCT
The complex is one of the group of SOCTA-protein (SOCTA-protein) complex and SOCTA-peptide (SOCTA-peptide) complex.
S2: The SOCTA reacted composites and M, to form a M-SOCTA composite, the M is one of the group of 188 Re and 99m Tc, moreover the M-SOCTA composite 188 Re-SOCTA -protein complex, 188 Re-SOCTA-peptide complex, 99m Tc-SOCTA-protein complex, 99m Tc-SOCTA-peptide complex, and the M-SOCTA complex It is a radioactive material to suppress.
The protein is Herceptin single strain antibody, the SOCTA-protein complex is SOCTA-Herceptin complex, and the M-SOCTA-protein complex is 188 Re-SOCTA-Herceptin complex.
ステップS1中でSOCTAとタンパク質或いはペプチドを反応させ、SOCTAのエステル類構造(-COOR)とタンパク質或いはペプチドのアミン類(-NH、-NH2)をボンド結合させペプチドボンド(peptide bond)のボンド結合を生じ、SOCTA-protein複合物或いはSOCTA- peptide複合物を形成する。S1の該SOCTAと該ハーセプチン(Herceptin)を反応させ、該SOCTA-Herceptin複合物を形成するステップにおいて、該SOCTAと該ハーセプチン(Herceptin)は結合反応を行う。しかも、ステップS1は該SOCTAをジメチルホルムアミド(DMF)溶液に溶解させ、該ハーセプチン(Herceptin)をN-ジハイドロキエチルピレラジン-N'-ジエタンサルフォニック酸(N-[2-Hydroxyethyl]piperazine-N'-[2-ethanesulfonic acid] )緩衝液(HEPES Buffer)に溶解させるステップを含む。しかも該SOCTAと該ハーセプチン(Herceptin)の最適モル数比は1：1である。S1の該SOCTAと該ハーセプチン(Herceptin)を反応させ、該SOCTA-Herceptin複合物を形成するステップは室温下で行い、しかもS1ステップ中においては透析のステップを含む。 In step S1, SOCTA reacts with a protein or peptide, and the bond structure of the peptide bond is formed by bonding the ester structure of SOCTA (-COOR) and the amine of the protein or peptide (-NH, -NH2). To form a SOCTA-protein complex or a SOCTA-peptide complex. In the step of reacting the SOCTA of S1 and the Herceptin (Herceptin) to form the SOCTA-Herceptin complex, the SOCTA and the Herceptin (Herceptin) undergo a binding reaction. In addition, in step S1, the SOCTA is dissolved in a dimethylformamide (DMF) solution, and the Herceptin is converted into N-dihydroxyethylpyrazine-N′-dietansulfonic acid (N- [2-Hydroxyethyl] piperazine). -N '-[2-ethanesulfonic acid]) including a step of dissolving in a buffer (HEPES Buffer). In addition, the optimum molar ratio of the SOCTA and the Herceptin is 1: 1. The step of reacting the SOCTA of S1 with the Herceptin to form the SOCTA-Herceptin complex is performed at room temperature, and the S1 step includes a dialysis step.
S2の該SOCTA-Herceptin複合物と該188Reを反応させ、該188Re-SOCTA-Herceptinを形成するステップ中において、該188Reは188Re高レニウム酸塩溶液(188Re perrhenate solution)により提供する。しかもS2の該SOCTA-Herceptin複合物と該188Reを反応させ、該188Re-SOCTA-Herceptinを形成するステップ中においては、先ず該188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)と塩化スズ(SnCl2)を反応させ、188Re-Glucoheptonateを得、該188Re-Glucoheptonateと該SOCTA-Herceptinを反応させ、該188Re-SOCTA-Herceptinを得る二段式ステップを含む。内、先ず該188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)と塩化スズ(SnCl2)を反応させ、188Re-Glucoheptonateを得るステップ中の、該グルコヘプトネートと該塩化スズの重量費は1.5：1〜5.4：1で、該グルコヘプトネートと該塩化スズの最適重量比は1.6：1である。先ず、該188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)と塩化スズ(SnCl2)を反応させ、188Re-Glucoheptonateを得るステップ中の、該グルコヘプトネート(Glucoheptonate)はキレート剤で、しかも該188Re-Glucoheptonateと該該SOCTA-Herceptinを反応させ該188Re-SOCTA-Herceptinを得るステップの反応温度は室温或いは37℃である。 In the step of reacting the 188 Re with the SOCTA-Herceptin complex of S2 to form the 188 Re-SOCTA-Herceptin, the 188 Re is provided by a 188 Re high rhenate solution ( 188 Re perrhenate solution) . Moreover, in the step of reacting the SOCTA-Herceptin complex of S2 with the 188 Re to form the 188 Re-SOCTA-Herceptin, first, the 188 rhenium ( 188 Re), glucoheptonate (Glucoheptonate), and chloride. It includes a two-stage step of reacting tin (SnCl 2 ) to obtain 188 Re-Glucoheptonate, reacting the 188 Re-Glucoheptonate with the SOCTA-Herceptin to obtain the 188 Re-SOCTA-Herceptin. First, the weight of the glucoreptonate and the tin chloride in the step of reacting the 188 rhenium ( 188 Re), glucoheptonate (Glucoheptonate) and tin chloride (SnCl 2 ) to obtain 188 Re-Glucoheptonate. The cost is 1.5: 1 to 5.4: 1 and the optimum weight ratio of the glucoheptonate to the tin chloride is 1.6: 1. First, in the step of reacting the 188 rhenium ( 188 Re), glucoheptonate (Glucoheptonate) and tin chloride (SnCl 2 ) to obtain 188 Re-Glucoheptonate, the glucoheptonate (Glucoheptonate) is a chelating agent. Moreover, the reaction temperature in the step of reacting the 188 Re-Glucoheptonate with the SOCTA-Herceptin to obtain the 188 Re-SOCTA-Herceptin is room temperature or 37 ° C.
本発明はステップS2中において該188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)、塩化スズ(SnCl2)、及び該SOCTA-Herceptin複合物を使用し、一段式反応を行い188Re-SOCTA-Herceptinを得る最適ステップとすることもできる。該188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)、塩化スズ(SnCl2)、及び該SOCTA-Herceptin複合物を使用し、一段式反応を行い188Re-SOCTA-Herceptinを得る最適ステップ中において、該グルコヘプトネートと該塩化スズの重量費は1.5：1〜5.4：1で、しかも最適重量比は1.6：1である。該188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)、塩化スズ(SnCl2)、及び該SOCTA-Herceptin複合物を使用し、一段式反応を行い188Re-SOCTA-Herceptinを得る最適ステップ中において、反応温度は室温或いは37℃である。. The present invention uses the 188 rhenium ( 188 Re), glucoheptonate (Glucoheptonate), tin chloride (SnCl 2 ), and the SOCTA-Herceptin complex in step S2 to perform a one-step reaction and perform 188 Re-SOCTA. -It can also be the optimal step to get Herceptin. Using the 188 rhenium ( 188 Re), glucoheptonate (Glucoheptonate), tin chloride (SnCl 2 ), and the SOCTA-Herceptin complex, performing a one-step reaction to obtain 188 Re-SOCTA-Herceptin , The weight cost of the glucoheptonate and the tin chloride is 1.5: 1 to 5.4: 1, and the optimum weight ratio is 1.6: 1. Using the 188 rhenium ( 188 Re), glucoheptonate (Glucoheptonate), tin chloride (SnCl 2 ), and the SOCTA-Herceptin complex, performing a one-step reaction to obtain 188 Re-SOCTA-Herceptin The reaction temperature is room temperature or 37 ° C. .
[SOCTA-Herceptin(SOCTA-ハーセプチン)結合反応]
適量のSOCTAをジメチルホルムアミド(DMF)溶液に溶解させ、適量のハーセプチン(Herceptin)をN-ジハイドロキエチルピレラジン-N'-ジエタンサルフォニック酸(N-[2-Hydroxyethyl]piperazine-N'-[2-ethanesulfonic acid] )緩衝液(HEPES Buffer)に溶解させる。該HEPES Bufferの組成はジハイドロキエチルピレラジン-N'-ジエタンサルフォニック酸(N-[2-Hydroxyethyl]piperazine-N'-[2-ethanesulfonic acid] 、HEPES)と0.29gの塩化ナトリウム(NaCl)を水(Ph=7.4)に溶かし、それぞれSOCTA溶液とハーセプチン(Herceptin)溶液中から適当な体積を取り出し、両者のモル数比を1：1とする。上記組成に基づき、適量の溶液を取り出し1.5ml遠心管に入れ、室温下で3時間反応させる。反応完成後の溶液を透析袋内に入れ、袋の口を封鎖し、4℃の冷蔵庫に入れ、2日間透析する。透析緩衝液(Dialysis Buffer)の組成はクエン酸ナトリウム(Sodium Citrate)と塩化ナトリウム(NaCl)を2リットルの水に溶かし、pHを5.2に調整する。透析完成後、袋内の液体を4℃で10分間遠心処理し、上澄み液を収集する。
本実験が使用するSOCTAとハーセプチン(Herceptin)のモル比は1：1で、結合反応を経て、透析後のSOCTA-Herceptin(SOCTA-ハーセプチン)複合物を、ELISA測定標準液とサンプルの間の吸収値により、EXCEL内挿方式を使用し求めた平均タンパク質濃度は7.9mg/mlである。
[SOCTA-Herceptin (SOCTA-Herceptin) binding reaction]
An appropriate amount of SOCTA is dissolved in a dimethylformamide (DMF) solution, and an appropriate amount of Herceptin is added to N-dihydroxyethylpyrazine-N'-dietansulfonic acid (N- [2-Hydroxyethyl] piperazine-N ' -[2-ethanesulfonic acid]) Dissolve in buffer (HEPES Buffer). The composition of the HEPES Buffer is dihydroxyethylpyrazine-N′-dietansulfonic acid (N- [2-Hydroxyethyl] piperazine-N ′-[2-ethanesulfonic acid], HEPES) and 0.29 g of sodium chloride ( NaCl) is dissolved in water (Ph = 7.4), and appropriate volumes are taken out from the SOCTA solution and the Herceptin solution, respectively, so that the molar ratio between the two is 1: 1. Based on the above composition, an appropriate amount of the solution is taken out and placed in a 1.5 ml centrifuge tube and reacted at room temperature for 3 hours. Place the solution after completion of the reaction in a dialysis bag, seal the mouth of the bag, place it in a refrigerator at 4 ° C, and dialyze for 2 days. The composition of the dialysis buffer (Sodium Citrate) and sodium chloride (NaCl) is dissolved in 2 liters of water and the pH is adjusted to 5.2. After completion of dialysis, centrifuge the liquid in the bag for 10 minutes at 4 ° C and collect the supernatant.
The molar ratio of SOCTA and Herceptin used in this experiment is 1: 1, and the dialysis SOCTA-Herceptin (SOCTA-Herceptin) complex is absorbed between the ELISA standard solution and the sample after the binding reaction. Depending on the value, the average protein concentration determined using the EXCEL interpolation method is 7.9 mg / ml.
[188Re-SOCTA-Herceptinの二段式調整]
１．188Re-Glucoheptonate標識の調整
400μlの188レニウム高レニウム酸塩溶液(188Re perrhenate solution)を塩化スズ(SnCl2)6-14mgのグルコヘプトネート(Glucoheptonate、GHと略称)に加え、1.5ml遠心管に入れ、室温下で反応させる。混合後15分後にサンプルを取り放射化学純度分析を開始すする。展開システムの固定フェーズはITLC-SGで、移動フェーズは生理食塩水とメチルエチルケトン(Methylethylketone、MEKと略称)である。
今回の実験はグルコヘプトネート(Glucoheptonate(GH))を選択し弱キレート剤(weak chelating agent)とし、反応瓶にGH(32mg)、SnCl2(6mg)、 188Re perrhenate solutionを加え、室温下で反応させ、定時にサンプルを取り放射化学分析(図３参照)を行う。その結果、15分後に188Re-GHの放射化学純度は76%に達し、30分時点では100%に達し、60分まではその完全反応を維持することが分かった。188Re-GHが反応30~60分間で予期の放射化学標準に達したことを確認後、該反応条件を選定し、188Re-GHの次の段階のSOCTA-Herceptin反応に用いる。
２．188Re-SOCTA-Herceptinの調整
上記放射化学純度が既に100%に達した188Re-GH溶液を適量のSOCTA-Herceptinに加え反応を継続させる。反応温度の標識効率に与える影響を検討するため、本実験では2種の温度(室温と37℃)を選択する。混合物の反応を開始後、15、30、60、90分後及び2時間後にサンプルを取り分析する。ITLC-SG/MEK(or NS)などのシステム対応の放射線薄層式フェーズ分析器(Radio-TLC)により放射化学純度分析を行う。
反応が完了した188Re-GHを20μlのSOCTA-Herceptinに加え、2組(室温と37℃)に分け実験を行い、温度の標識効率に対する影響(図４参照)について検討する。その結果、37℃の反応条件下で、製品の標識効率は反応30分後48.99%から反応1時間では66.12%に向上し、2時間時点の標識効率は最高で96.01±0.13%に達することが分かった。
反応を室温で行う時には、反応が完了した188Re-GHを適量のSOCTA-Herceptinに加え、様々な時間点でサンプルを取り分析する。その結果、最高標識効率は1時間時点でわずかに60.83±8.59%であることが分かった。
[ 188 Re-SOCTA-Herceptin two-stage adjustment]
1. 188 Re-Glucoheptonate sign adjustment
Add 400 μl of 188 rhenium high rhenate solution ( 188 Re perrhenate solution) to tin chloride (SnCl 2 ) 6-14 mg glucoheptonate (abbreviated as Glucoheptonate, GH) and place in a 1.5 ml centrifuge tube at room temperature. React. Take a sample 15 minutes after mixing and start the radiochemical purity analysis. The stationary phase of the deployment system is ITLC-SG, and the mobile phase is saline and methyl ethyl ketone (abbreviated as MEK).
In this experiment, glucoheptonate (GH) was selected as a weak chelating agent, GH (32 mg), SnCl 2 (6 mg), and 188 Re perrhenate solution were added to the reaction bottle at room temperature. The sample is taken at a fixed time and subjected to radiochemical analysis (see FIG. 3). As a result, it was found that the radiochemical purity of 188 Re-GH reached 76% after 15 minutes, reached 100% at 30 minutes, and maintained its complete reaction until 60 minutes. After confirming that 188 Re-GH has reached the expected radiochemical standard within 30-60 minutes of reaction, the reaction conditions are selected and used for the SOCTA-Herceptin reaction in the next stage of 188 Re-GH.
2. Preparation of 188 Re-SOCTA-Herceptin The 188 Re-GH solution, whose radiochemical purity has already reached 100%, is added to an appropriate amount of SOCTA-Herceptin and the reaction is continued. To examine the effect of reaction temperature on labeling efficiency, two temperatures (room temperature and 37 ° C) are selected in this experiment. Samples are taken and analyzed after 15, 30, 60, 90 minutes and 2 hours after starting the reaction of the mixture. Radiochemical purity analysis is performed using a radiation thin layer phase analyzer (Radio-TLC) compatible with systems such as ITLC-SG / MEK (or NS).
188 Re-GH after the reaction is completed is added to 20 μl of SOCTA-Herceptin, and experiments are performed in two groups (room temperature and 37 ° C.) to examine the effect of temperature on labeling efficiency (see FIG. 4). As a result, under 37 ° C reaction conditions, the product labeling efficiency improved from 48.99% after 30 minutes to 66.12% in 1 hour of reaction, and the labeling efficiency at 2 hours could reach 96.01 ± 0.13% at the maximum. I understood.
When the reaction is carried out at room temperature, 188 Re-GH, which has completed the reaction, is added to an appropriate amount of SOCTA-Herceptin, and samples are taken and analyzed at various time points. As a result, it was found that the maximum labeling efficiency was only 60.83 ± 8.59% at 1 hour.
[188Re-SOCTA-Herceptinの一段式調整]
本実験が採用する方法は、188Re perrhenate solution、SnCl2、Glucoheptonate、及びSOCTA-Herceptinを同時に試験管内に加え、それぞれ室温と37℃で反応させ、様々な時間点における放射化学純度の変化を観察する。放射化学分析は二段式方法と同様で、ITLC-SG/MEK(or NS)システムを採用する。
二段式調整方法は37℃の条件下において96%の放射化学純度を得ることができるが、反応にかなりの時間を要するため、一段式調整方法を試みる。適量の188Re溶液を直接取り、グルコヘプトネート(Glucoheptonate)、SnCl2及びSOCTA-Herceptinを反応小瓶内に加え作用させる。反応温度、塩化スズ含量及びSOCTA-Herceptin含量などの条件が標識効率に与える影響を検討するため、関連試験を行った。その結果は以下に記載する。
１．温度の標識効率に対する影響
実験では先ず37℃の条件を選択し、反応30分後の標識効率は96.91%に達し、継続し24時間観察したところ、188Re-SOCTA-Herceptinの放射化学純度は97％を維持可能(図４参照)であることが分かった。同時に、室温での反応と比較したところ、図５に示すように、SnCl2の量が6mgである時、反応時間の延長に従い、188Re-SOCTA-Herceptinの放射化学純度も増加し、2時間時点での標識効率は95.18±0.50%に達する。しかし、SnCl2を8mg以上にまで増量すると、反応30分後には95%の放射化学純度に達する。
２．異なる含量のSnCl2が標識効率に与える影響
SnCl2は医薬物で最もよく使用される還元剤である。本実験でもSnCl2を採用し、七価の188Reを五価の188Re(V)に還元し、6、8、10、12、14、16、18、20mgのSnCl2を加え、室温下において反応させる時の標識に対する影響を比較する(図５参照)。その結果、SOCTA-Herceptinを14mg加えた15分間での標識効率は91%以上に達し、2時間の標識効率は98.71%に達し、しかも分析の結果14mgのSnCl2を加えた1時間毎の標識効率はすべて6mgのSnCl2を加えた標識効率より高く、かつ明らかな差異があることが分かった。実験では同時に、加えるSnCl2の量が8mg以上で、室温下で30分反応させた時には95%に達し、しかもその標識効率は2時間以上維持可能であるが、この結果は良好であるが、ジェル質を形成し易いことが後に分かった。よって、6mgのSnCl2及び室温下での反応を188Re-SOCTA-Herceptinの反応条件とすることとした。
３．異なる体積のSOCTA-Herceptin(SOCTA-ハーセプチン)が標識効率に与える影響関係
本研究では10、20及び40μlの３種の体積のSOCTA-Herceptinを選択し使用する(図６参照)。20μlに含有するSOCTA-Herceptinの量は0.158mgで、10μlとするなら質量は0.079mgに半減する。それを室温下で0.25、0.5時間の反応を行った時の標識効率はそれぞれ38.29%及び65.01%であった。この結果は同一時間点の20或いは40μlを加えた反応結果と顕著な差異があり、製品の最高標識効率は3時間時点に出現し、その値は93.30%であった。もし加える体積が20、或いは40μlであるなら、両者の平均標識効率は30分時点で93%以上に達するが、加える体積が20、或いは40μlである場合の同一時間点での標識効率には顕著な差異はない。
[ 188 Re-SOCTA-Herceptin one-stage adjustment]
In this experiment, 188 Re perrhenate solution, SnCl2 , Glucoheptonate, and SOCTA-Herceptin were added to the test tube at the same time, and reacted at room temperature and 37 ° C, respectively, and observed changes in radiochemical purity at various time points. To do. Radiochemical analysis is the same as the two-stage method, using the ITLC-SG / MEK (or NS) system.
The two-stage adjustment method can obtain a radiochemical purity of 96% under the condition of 37 ° C. However, since the reaction takes a considerable time, the one-stage adjustment method is tried. An appropriate amount of 188 Re solution is directly taken and glucoheptonate (Glucoheptonate), SnCl 2 and SOCTA-Herceptin are added to the reaction vial and allowed to act. In order to investigate the effect of conditions such as reaction temperature, tin chloride content and SOCTA-Herceptin content on labeling efficiency, related tests were conducted. The results are described below.
1. Effect of temperature on labeling efficiency In the experiment, the condition of 37 ° C was first selected, the labeling efficiency after 30 minutes of reaction reached 96.91%, and when observed continuously for 24 hours, the radiochemical purity of 188 Re-SOCTA-Herceptin was 97 % Can be maintained (see FIG. 4). At the same time, when compared with the reaction at room temperature, as shown in FIG. 5, when the amount of SnCl 2 was 6 mg, the radiochemical purity of 188 Re-SOCTA-Herceptin increased as the reaction time increased, and 2 hours The labeling efficiency at the time point reaches 95.18 ± 0.50%. However, when SnCl 2 is increased to 8 mg or more, a radiochemical purity of 95% is reached after 30 minutes of reaction.
2. Effects of SnCl 2 of different content has on the labeling efficiency
SnCl 2 is the reducing agent most often used in pharmaceutical products. In this experiment, SnCl 2 was also used to reduce heptavalent 188 Re to pentavalent 188 Re (V), add 6, 8, 10, 12, 14, 16, 18, 20 mg of SnCl 2 at room temperature. Compare the effects on the label when reacting in (see FIG. 5). As a result, the labeling efficiency in 15 minutes after adding 14 mg of SOCTA-Herceptin reached 91% or more, the labeling efficiency in 2 hours reached 98.71%, and the result of analysis showed that the labeling was performed every hour with 14 mg of SnCl 2 added. The efficiencies were all higher than the labeling efficiency with 6 mg SnCl 2 added and were found to be clearly different. At the same time in the experiment, the amount of SnCl 2 to be added is 8 mg or more and reaches 95% when reacted at room temperature for 30 minutes, and its labeling efficiency can be maintained for 2 hours or more, but this result is good, It was later found that it was easy to form a gel. Therefore, the reaction condition of 6 mg of SnCl 2 and room temperature was set as the reaction condition of 188 Re-SOCTA-Herceptin.
3. Effect of different volumes of SOCTA-Herceptin (SOCTA-Herceptin) on labeling efficiency In this study, three types of SOCTA-Herceptin (10, 20 and 40 μl) are selected and used (see FIG. 6). The amount of SOCTA-Herceptin contained in 20 μl is 0.158 mg, and if it is 10 μl, the mass is halved to 0.079 mg. When the reaction was carried out at room temperature for 0.25 and 0.5 hours, the labeling efficiencies were 38.29% and 65.01%, respectively. This result was significantly different from the reaction result when 20 or 40 μl of the same time point was added, and the maximum labeling efficiency of the product appeared at 3 hours, and its value was 93.30%. If the added volume is 20 or 40 μl, the average labeling efficiency of both reaches 93% or more at 30 minutes, but the labeling efficiency at the same time point is significant when the added volume is 20 or 40 μl. There is no significant difference.
[188Re-SOCTA-Herceptinの体外安定性試験]
体外安定性試験は二組に分かれて行う。一組は反応が完了した188Re-SOCTA-Herceptinを室温下に置き、異なる時間点での製品の放射純度変化を観察する。もう一組は、188Re-SOCTA-Herceptinをヒトの血清内に加え、37℃で反応させ、時間の経過に伴う製品の放射純度変化を観察する。
188Re-SOCTA-Herceptinの標識完成後、室温下で24時間、製品の安定性放射化学純度分析を行った。その結果により、188Re-SOCTA-Herceptinの標識効率は2~24時間で、94~96%を安定的に維持することが分かった。
実験では同時に血清の安定性についても検討する。188Re-SOCTA-Herceptinの標識完成後、37℃のヒト血清内に加え、24時間の製品の安定性放射化学純度分析を行った。その結果により、188Re-SOCTA-Herceptinの標識効率は2時間から徐々に90%に下降し、24時間ではなお83％を維持可能であることが分かった(図７参照)。
初期の二段式標識方式は188レニウム(188Re)とグルコヘプトネート(Glucoheptonate)に先にエステル化合物を形成させ、配位体交換(ligand-exchange)によりさらにSOCTA-Herceptinに換える。実験後期には、一段式標識方式は、標識時間上において1〜1.5時間を節減可能で、しかも加熱が不要で、平均室温22℃下において約1時間で標識が完成されることが発見された。これは、半減期を具える治療用放射性方位元素にとっては、治療効果を延長可能である他、さらには標識時間を節減することもできることを示す。また安定性についても、二段式標識方式は標識後2時間で徐々に分解し、24時間時点でなお77.29%を残し、一段式では96.88%を維持可能であることが分かった。
一段式標識方法反応の所要時間は最短で、標識効率を24時間維持可能な安定性を具える。本製品の現在の標識効率と安定性は、濾過プロセスを経なくとも95%以上を達成可能で、しかも24時間の安定を維持し、純度は100％に近い。研究により、高標識効率、24時間安定性、100%に近い純度の188Re-SOCTA-Herceptinが既に完成されたことは明らかである。今後は細胞及び動物に対する治療効果の試験を行い、将来の癌治療(乳癌治療など)に臨床上のより良い選択を提供できることを期する。
[ 188 Re-SOCTA-Herceptin in vitro stability test]
In vitro stability tests are conducted in two groups. One set places the completed 188 Re-SOCTA-Herceptin at room temperature and observes the change in product's radiopurity at different time points. The other set adds 188 Re-SOCTA-Herceptin into human serum, reacts at 37 ° C, and observes the change in radiopurity of the product over time.
After completion of the labeling of 188 Re-SOCTA-Herceptin, the stability radiochemical purity analysis of the product was performed at room temperature for 24 hours. As a result, it was found that the labeling efficiency of 188 Re-SOCTA-Herceptin was stably maintained at 94 to 96% in 2 to 24 hours.
In the experiment, the stability of serum is also examined. After completing the labeling of 188 Re-SOCTA-Herceptin, it was added to human serum at 37 ° C, and the stability radiochemical purity analysis of the product was performed for 24 hours. As a result, it was found that the labeling efficiency of 188 Re-SOCTA-Herceptin gradually decreased from 2 hours to 90% and could still be maintained at 83% in 24 hours (see FIG. 7).
In the initial two-stage labeling method, 188 rhenium ( 188 Re) and glucoheptonate (Glucoheptonate) are first formed into an ester compound, and further converted to SOCTA-Herceptin by ligand-exchange. Later in the experiment, it was discovered that the one-stage labeling method can save 1 to 1.5 hours on the labeling time, does not require heating, and completes labeling in about 1 hour at an average room temperature of 22 ° C. . This indicates that for therapeutic radioactive orientation elements with a half-life, the therapeutic effect can be extended and also the labeling time can be saved. In addition, regarding the stability, it was found that the two-stage labeling method gradually decomposed 2 hours after the labeling, and remained at 77.29% at 24 hours, while the one-stage system could maintain 96.88%.
The time required for the one-step labeling method reaction is the shortest, and it has the stability that can maintain the labeling efficiency for 24 hours. The current labeling efficiency and stability of this product can be over 95% without going through the filtration process, and it remains stable for 24 hours, and the purity is close to 100%. Studies clearly show that 188 Re-SOCTA-Herceptin with high labeling efficiency, 24-hour stability, and purity near 100% has already been completed. In the future, we will test the therapeutic effects on cells and animals and hope to provide better clinical choices for future cancer treatments (such as breast cancer treatment).
Claims (23)
SOCTAとタンパク質或いはペプチドを反応させ、SOCTA複合物を形成し、該SOCTA複合物はSOCTA-protein複合物及びSOCTA-peptide複合物のグループの内の一つで、
該SOCTA複合物とMを反応させM- SOCTA複合物を形成し、該Mは188Re及び99mTcのグループの内の一つで、しかも該M- SOCTA複合物は188Re-SOCTA-protein複合物、188Re-SOCTA-peptide複合物、99mTc-SOCTA-protein複合物、99mTc -SOCTA-peptide複合物のグループの内の一つで、
該M- SOCTA複合物は癌を抑制する放射性物質であることを特徴とする癌を抑制する放射性物質の調整方法。 The steps of the method of adjusting the radioactive material that suppresses cancer are:
SOCTA and protein or peptide are reacted to form an SOCTA complex, which is one of the group of SOCTA-protein complex and SOCTA-peptide complex,
The SOCTA complex is reacted with M to form an M-SOCTA complex, wherein M is one of the group of 188 Re and 99m Tc, and the M-SOCTA complex is 188 Re-SOCTA-protein complex. things, 188 Re-SOCTA-peptide complexes, 99m Tc-SOCTA-protein conjugate, in one of the groups of 99m Tc -SOCTA-peptide complexes,
A method for preparing a radioactive substance that suppresses cancer, wherein the M-SOCTA composite is a radioactive substance that suppresses cancer.
先ず該188レニウム(188Re)、グルコヘプトネート(Glucoheptonate)と塩化スズ(SnCl2)を反応させ、188Re-Glucoheptonateを得、
該188Re-Glucoheptonateと該SOCTA-Herceptinを反応させ、該188Re-SOCTA-Herceptinを得る二段式ステップを含むことを特徴とする癌を抑制する放射性物質の調整方法。 The method for preparing a radioactive substance for suppressing cancer according to claim 7, wherein the step of reacting the SOCTA-Herceptin complex with the 188 Re to form the 188 Re-SOCTA-Herceptin complex further comprises the 188 rhenium. ( 188 Re), reacting glucoheptonate (Glucoheptonate) and tin chloride (SnCl 2 ) to obtain 188 Re-Glucoheptonate,
A method for preparing a radioactive substance for suppressing cancer, comprising a two-step step of reacting the 188 Re-Glucoheptonate and the SOCTA-Herceptin to obtain the 188 Re-SOCTA-Herceptin.
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| JP2002518460A (en) * | 1998-06-22 | 2002-06-25 | イムノメディクス， インコーポレイテッド | Use of bispecific antibodies for pretargeting diagnostics and pretargeting therapy |
| JP2004501093A (en) * | 2000-04-28 | 2004-01-15 | プレジデント・アンド・フェロウズ・オブ・ハーバード・カレッジ | Technetium-99m and rhenium-labeled small agents and tumor imaging methods |
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| JPH11507623A (en) * | 1995-06-07 | 1999-07-06 | イムノメディクス，インコーポレイテッド | Thiolation of peptides for radionuclide-based radiation detection and radiotherapy |
| JP2002518460A (en) * | 1998-06-22 | 2002-06-25 | イムノメディクス， インコーポレイテッド | Use of bispecific antibodies for pretargeting diagnostics and pretargeting therapy |
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