| アッセイID： B151 | 報告日： - | 所属： 化学療法基盤支援活動 | アッセイ者名： 深澤秀輔 | アッセイ名： プロテインキナーゼ阻害の検定 II-1 |
| 概要 チロシンキナーゼおよびその下流のシグナル伝達系は，多くのがん細胞で活性化しており，抗がん剤の有力な分子標的である．ここでは細胞を血小板由来増殖因子(platelet-derived growth factor, PDGF) 刺激して，活性化されるPDGFレセプターチロシンキナーゼおよびその下流の主要な細胞内シグナル伝達経路に対する検体の阻害効果を評価するとともに，４種類のチロシンキナーゼに対する阻害効果をcell based ELISAを用いて評価する． 実験系１：細胞内シグナル伝達 NRK細胞を96ウェルプレートにまき込み，3日間培養後，サンプルを添加，3時間処理した後にPDGFで刺激する．電気泳動用サンプルを調製し，リン酸化されたシグナル伝達分子（AKT，ERK，PKD，PLCγ1，S6 ribosomal protein）及びphosphotyrosineに対する抗体によるウェスタンブロットを行い，PDGFレセプターからの細胞内シグナル伝達に対する影響を評価する． |
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| 評価のクライテリア 10 μMで50％以上の阻害活性が認められたものを有効とする．+++, 1?M; ++, 10 ?M で50％以上の阻害 |
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発表論文 | レポート提出者によるレポート内容の最終確認日： - | |||
| Well | KIT 1 | KIT 2 | KIT 3 | |||
| compound | 結果 | compound | 結果 | compound | 結果 | |
| 1 - A | none (DMSO) | none (DMSO) | none (DMSO) | |||
| 1 - B | 5-FU | - | Pifithrin-a (cyclic) | - | ABT-702 | - |
| 1 - C | Bestatin Hydrochloride | - | PRIMA-1 | - | Akt Inhibitor IV | - |
| 1 - D | Bleomycin sulfate | - | Finasteride | - | Akt Inhibitor VIII, Isozyme-Selective, Akti-1/2 | AKT++ |
| 1 - E | Cisplatin | - | Aminoglutethimide | - | Akt Inhibitor XI | - |
| 1 - F | Methotrexate | - | Formestane | - | compound C | AKT++, ERK++, PKD++, S6R++, phosphotyrosine++ |
| 1 - G | Mitomycin C | - | Mifepristone | - | ATM/ATR kinase inhibitor | - |
| 1 - H | Vinblastine sulfate | phosphotyrosine ++ | TOFA | - | ATM kinase inhibitor | - |
| 2 - A | Paclitaxel | - | Amastatin | - | Aurora kinase/cdk inhibitor | - |
| 2 - B | Flutamide | - | Actinonin | - | Aurora kinase inhibitor II | - |
| 2 - C | Daunorubicin, HCl | ERK ++, phosphotyrosine ++ | Oligomycin | - | Aurora kinase inhibitor III | - |
| 2 - D | Doxorubicin, HCl | ERK ++, phosphotyrosine ++ | Bafilomycin A1 | ERK++, S6R++ | AG957 | - |
| 2 - E | Tamoxifen, citrate | - | HA 14-1 | - | LFM-A13 | - |
| 2 - F | Actinomycin D | - | BH3I-1 | - | Terreic acid | - |
| 2 - G | Camptothecin | - | LFM-A13 | - | KN-93 | - |
| 2 - H | Aclarubicin | - | Terreic acid | - | KN-62 | - |
| 3 - A | Etoposide (VP-16) | - | E-64d | - | Lavendustin C | - |
| 3 - B | Cytochalasin D | AKT++, ERK ++, PKD ++, PLCγ1 ++, S6R ++, phosphotyrosine ++ | ALLN | - | Kenpaullone | - |
| 3 - C | 2',5'-dideoxyadenosine | - | CA-074 | - | purvalanol A | - |
| 3 - D | AKT inhibitor | - | Pepstatin A | - | Olomoucine | - |
| 3 - E | NL-71-101 | Z-GLF-CMK | AKT++, ERK++, PKD++, PLCγ1++, S6R++, phosphotyrosine++ | Alsterpaullone, 2-cyanoethyl | - | |
| 3 - F | AG957 | - | RS 102895 | - | Cdk1/2 inhibitor III | PKD++, PLCγ1++, phosphotyrosine++ |
| 3 - G | KN93 | - | SB 328437 | - | Cdk2/9 inhibitor | - |
| 3 - H | Z-VAD-FMK | - | SB 225002 | - | NU6102 | - |
| 4 - A | Kenpaullone | - | AMD3100 octahydrochloride | - | Cdk4 inhibitor | AKT++, ERK ++ |
| 4 - B | Purvalanol A | - | NSC95397 | AKT++, ERK++, PKD++, PLCγ1++, S6R++ | NSC625987 | - |
| 4 - C | 3-ATA | - | SC-ααδ9 | - | SB218078 | AKT++, ERK++, PKD++, PLCγ1++, S6R++, phosphotyrosine++ |
| 4 - D | Olomoucine | - | Amiloride | - | isogranulatimide | - |
| 4 - E | TBB | - | Lidocaine | - | Chk2 inhibitor | - |
| 4 - F | Sulindac sulfide | - | Monensin | - | Chk2 inhibitor II | - |
| 4 - G | Valeryl salicylate | - | Ouabain | - | Ellagic acid (Dihydrate) | - |
| 4 - H | NS-398 | - | Sanguinarine | AKT++, ERK++, PKD++, S6R++, phosphotyrosine++ | TBB | - |
| 5 - A | Sodium salicylate | - | Glibenclamide | - | DMAT | - |
| 5 - B | Theophylline | - | Dequalinium | - | D4476 | - |
| 5 - C | Azacytidine | - | Diazoxide | - | TG003 | - |
| 5 - D | Aphidicolin | - | Valinomycin | - | Diacylglycerol kinase inhibitor II | - |
| 5 - E | AG1478 | - | Nigericin | - | IC60211 | - |
| 5 - F | Genistein | - | Diltiazem | - | TX-1918 | - |
| 5 - G | Manumycin A | AKT++, ERK++, PKD++, PLCγ1++, S6R++, phosphotyrosine++ | Nifedipine | - | BPIQ-Ⅱ | - |
| 5 - H | FTI-276 | - | Verapamil | - | AG1478 | - |
| 6 - A | SU1498 | - | PGP-4008 | - | AG490 | - |
| 6 - B | GGTI-286 | - | Fumitremorgin C | - | SU4984 | AKT++, ERK++, PKD++, PLCγ1++, phosphotyrosine++ |
| 6 - C | Dexamethasone | - | A23187 | - | SU5402 | - |
| 6 - D | GSK-3 inhibitor II | - | Ionomycin | AKT++, ERK++, PKD++, PLCγ1++, S6R++, phosphotyrosine++ | Flt-3 Inhibitor | - |
| 6 - E | Scriptaid | - | Thapsigargin | - | cFMS Receptor Tyrosine Kinase Inhibitor | - |
| 6 - F | Trichostatin A | - | t-Butylhydroquinone (BHQ) | - | SU6656 | - |
| 6 - G | AG825 | - | N-phenylanthranilic acid | - | GSK-3 inhibitor IX | - |
| 6 - H | Cycloheximide | - | DIDS | - | 1-Azakenpaullone | - |
| 7 - A | Lovastatin | - | SB 218078 | PKD++, phosphotyrosine++ | Indirubin-3'-monoxime | - |
| 7 - B | Radicicol | - | Debromohymenialdisine (DBH) | - | AG825 | - |
| 7 - C | 17-AAG | - | Rotenone | - | AG1024 | - |
| 7 - D | AG1024 | - | Antimycin A1 | - | AGL 2263 | - |
| 7 - E | 1400W, HCl | - | Leptomycin B* | PKD++ | BMS-345541 | - |
| 7 - F | AMT, HCl | - | R59022 | - | IKK-2 inhibitor VI | - |
| 7 - G | AG490 | - | Dioctanoylglycol | - | IRAK-1/4 inhibitor | - |
| 7 - H | Cucurbitacin I | AKT+++, ERK+++, PKD+++, PLCγ1+++, S6R+++, phosphotyrosine+++ | RHC80267 | - | JAK Inhibitor I | - |
| 8 - A | SP600125 | - | Xanthohumol | - | JAK3 Inhibitor VI | PKD++, PLCγ1++, phosphotyrosine++ |
| 8 - B | Damnacanthal | - | C75 | - | SP600125 | - |
| 8 - C | PD 98059 | - | Cerulenin | - | JNK inhibitor VIII | phosphotyrosine+++ |
| 8 - D | U0126 | - | Tunicamycin | - | Damnacanthal | - |
| 8 - E | Fumagillin | - | Deoxynojirimycin | - | PP2 | - |
| 8 - F | GM 6001 | - | Swainsonine | - | ERK inhibitor II | - |
| 8 - G | N-Acetyl-L-cysteine | phosphotyrosine++ | LY 83583 | AKT++, ERK++, PKD++, PLCγ1++, S6R++, phosphotyrosine++ | PD98059 | - |
| 8 - H | Aminoguanidine, HCl | phosphotyrosine++ | ODQ | - | U-0126 | - |
| 9 - A | L-NMMA | - | Anacardic acid | - | MEK inhibitor I | - |
| 9 - B | PD169316 | - | Chetomin | AKT++, ERK++, PKD++, PLCγ1++, S6R++, phosphotyrosine++ | SU11274 | - |
| 9 - C | SB 203580 | - | Dimethyloxalylglycine | - | ML-7 | - |
| 9 - D | Rapamycin | phosphotyrosine++, S6R+++ | HR22C16 | - | SB202190 | - |
| 9 - E | NU1025 | - | Monastrol | - | SB239063 | - |
| 9 - F | Benzamide | - | Nordihydroguaiaretic acid (NDGA) | - | AG1296 | - |
| 9 - G | D609 | - | ETYA | - | SU11652 | AKT++, ERK++, PKD++, PLCγ1++, S6R++, phosphotyrosine++ |
| 9 - H | IBMX | - | Baicalein | - | PDGF receptor tyrosine kinase inhibitor V | AKT++, ERK+++, PKD +++, PLCγ1++, S6R++, phosphotyrosine+++ |
| 10 - A | Ro-20-1724 | - | Nutlin-3 | - | PDGF receptor tyrosine kinase inhibitor IV | AKT++ |
| 10 - B | Zaprinast | - | MDM2 inhibitor | - | LY-294002 | AKT++ |
| 10 - C | AG1296 | phosphotyrosine++ | Phenelzine | - | Wortmannin | AKT++ |
| 10 - D | LY294002 | ATK++, S6R++ | Deprenyl | - | H-89 | - |
| 10 - E | Wortmannin | ATK++ | Decylubiquinone | - | 4-cyano-3-methylisoquinoline | - |
| 10 - F | H-89, HCl | - | Ro 5-4864 | - | Bisindolymaleimide I, HCl | PKD++ |
| 10 - G | Bisindolymaleimide I, HCl | phosphotyrosine++, PKD++, PLCγ1++ | Lonidamine | - | Go7874 | - |
| 10 - H | H-7 | - | ML-7 | - | Rp-8-CPT-cGMPS | - |
| 11 - A | Staurosporine | AKT+++, ERK+++, PKD+++, PLCγ1+++, S6R+++, phosphotyrosine+++ | Benzylguanine | - | KT5823 | - |
| 11 - B | cPLA2inhibitor | - | DFMO | - | PKR inhibitor | PLCγ1++ |
| 11 - C | OBAA | - | KT 5823 | - | RAF1 kinase inhibitor I | ERK+++ |
| 11 - D | Cantharidin | - | Rp-8-CPT-cGMPS | - | ZM 336372 | PKD++, PKD++, phosphotyrosine++ |
| 11 - E | Cytostatin | MK 886 | - | H-1152 | - | |
| 11 - F | Cyclosporin A | - | Clofibrate | - | Y-27632 | - |
| 11 - G | FK-506 | - | BADGE | - | radicicol | - |
| 11 - H | MG-132 | - | Troglitazone | - | PP1 analog | - |
| 12 - A | Lactacystin | - | AZT | - | Syk inhibitor | ERK ++ |
| 12 - B | Hydroxyurea | - | Nalidixic acid | - | SB431542 | ERK ++ |
| 12 - C | HA1077 | - | a-Amanitin | - | TGF-b RI kinase inhibitor II | ERK ++ |
| 12 - D | Y27632 | - | MST-312 | - | Tpl2 kinase inhibitor | - |
| 12 - E | PP1 (analog) | - | b-Rubromycin | phosphotyrosine++ | TrkA inhibitor | - |
| 12 - F | PP-H | SB 431542 | - | VEGFR receptor tyrosine kinase inhibitor II | AKT++, ERK+++, PK ++, PLCγ1++, S6R++, phosphotyrosine+++ | |
| 12 - G | Nocodazole | - | N1,N12-Diethylspermine (BESpm) | - | VEGF recptor 2 kinase inhibitor I | - |
| 12 - H | Dephostatin | - | Fumonisin B1 | - | SU1498 | ERK++ |