| 名稱 | INCB3344 |
| 描述 | INCB3344 is an effective, specific and orally bioavailable CCR2 antagonist with IC50 values of 9.5 nM (mCCR2) and 5.1 nM (hCCR2) in binding antagonism and 7.8 nM (mCCR2) and in 3.8 nM (hCCR2) antagonism of chemotaxis activity. |
| 細(xì)胞實驗 | WEHI-274.1 cells (5×10^5) in RPMI 1640 (VWR) with or without various concentrations of INCB3344 in RPMI 1640 are loaded in the wells on top of an 8-μm polycarbonate filter in a 96-well-modified Boyden chamber. Beneath the filter, 30 nM mCCL2 with or without INCB3344 or media is placed in a corresponding 96-well plate. The sealed chambers are incubated for 45 min at 37°C, 5% CO2. Filters are washed, stained with Wright-Giemsa, and the number of cells that migrate toward mCCL2 in the bottom chamber counted by microscopy. The ability of INCB3344 to antagonize CCR2-mediated chemotaxis is reported as the inhibitor concentration required for IC50 values of specific migration to mCCL2. Specific migration is defined as the total migration minus the background migration. A similar assay is used to determine the impact of INCB3344 on CCR1-mediated chemotaxis of WEHI-274.1 cells, by using mouse MIP-1α as a ligand. In addition C5a, FMLP and RANTES are similarly tested in the presence of INCB3344 for migration of WEHI-274.1 cells. For the studies on the impact of INCB3344 on CCR5-mediated chemotaxis, murine T cells are used as the cell system with mouse MIP-1β as the ligand [2]. |
| 激酶實驗 | Cells (5×10^5) in RPMI 1640 (VWR), +0.1% BSA+20 mM HEPES (VWR), are added to various concentrations of INCB3344 in RPMI 1640 followed immediately by the addition of 150 pM 125I-labeled mCCL2 (mouse CCL2(JE)) and incubated for 30 min at room temperature (RT). For the nonspecific control, 0.3 μM mCCL2 is added in place of INCB3344. Cells are then harvested through 1.2-μm polyvinylidene difluoride filters, the filters are air-dried, and binding is determined by counting in a gamma counter. Antagonist activity is reported as the inhibitor concentration required for IC50 of specific binding. Specific binding is defined as the total binding minus the nonspecific binding and typically represents 97% of the total binding [2]. |
| 動物實驗 | In a subset of experiments, DOCA/salt-treated mice are further randomly assigned to receive the CCR2 antagonist, INCB3344 (30 mg/kg per day) or vehicle (10% DMSO/0.9% carboxymethylcellulose) via daily intraperitoneal injections commencing 10 days after induction of hypertension and continuing until the end of the 21-day treatment period. The normotensive control group for these experiments consists of sham-treated mice that receive a vehicle from days 10 to 21 [3]. Adult male Sprague-Dawley rats (200-250 g) are used. After t=0 baseline measurement, rats are lightly anesthetized under an isoflurane/oxygen (5%; 2 L/min) flow and 25 μL of either saline (vehicle), 1 μg of CCL2 and/or 1 mM of INCB3344 is administered intrathecally between L5 and L6 vertebrae. Animals are tested once at 30, 60, 90, 120, and 240 min following drug administration. The percentage of maximal potential effect is calculated for every time point [4]. |
| 體外活性 | INCB3344的藥理活性首先通過利用小鼠單核細(xì)胞系WEHI-274.1，通過全細(xì)胞結(jié)合試驗，評估其抑制CCL2與CCR2結(jié)合的能力來進(jìn)行表征。在該試驗中，INCB3344的結(jié)合IC50被確定為10 nM，而在90 nM的濃度下觀察到>90%的結(jié)合抑制[1]。INCB3344還是對大鼠和獼猴CCR2的有效拮抗劑，其結(jié)合拮抗IC50值分別為7.3和16 nM，化學(xué)趨化活性拮抗的IC50值分別為2.7和6.2 nM。此外，INCB3344對超過50種離子通道、轉(zhuǎn)運蛋白、趨化因子受體及其他選定的GPCRs展示出>1 μM的IC50值。它也是一種選擇性的mCCR2拮抗劑，對最與mCCR2同源的兩種趨化因子受體，即murine CCR1和murine CCR5表現(xiàn)出>1 μM和>3 μM的IC50值[4]。 |
| 體內(nèi)活性 | INCB3344能夠防止去氧皮質(zhì)酮醋酸酯(DOCA)/鹽誘導(dǎo)的血管CCR2表達(dá)變化。在一系列獨立實驗中，從DOCA/鹽處理周期的第7天到第21天接受INCB3344的小鼠主動脈中CCR2表達(dá)提高約1.5倍，與安慰劑動物相比明顯較低；但是，這一CCR2表達(dá)水平顯著低于僅接受載體處理組的水平。同樣，接受INCB3344的小鼠中DOCA/鹽處理引起的其受體配體CCL2表達(dá)增加被減弱。相比之下，接受載體或INCB3344的DOCA/鹽處理小鼠中CCL7、CCL8和CCL12的水平均有類似程度的提高[2]。通過靜脈給藥給CD-1小鼠時，INCB3344表現(xiàn)出高清除率和適中的分布體積，導(dǎo)致半衰期短，僅為1小時。盡管清除率高，但通過口服給藥仍能獲得良好的暴露效果，10 mg/kg劑量的AUC為2664 nM h?？诜锢枚葹?7%。相比之下，以相同劑量口服給Balb/c小鼠時獲得略好的口服暴露（AUC=3888 nM h）[4]。 |
| 存儲條件 | Powder: -20°C for 3 years | In solvent: -80°C for 1 year | Shipping with blue ice. |
| 溶解度 | H2O : Insoluble
DMSO : 210 mg/mL (363.58 mM), Sonication is recommended.
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| 關(guān)鍵字 | CC chemokine receptor | INCB 3344 | INCB3344 | INCB-3344 | CCR | Inhibitor | inhibit |
| 相關(guān)產(chǎn)品 | BMS-817399 | Pirfenidone | WAY-639418 | PF-4136309 | AZD2098 | Aplaviroc | Vercirnon | CCR2 antagonist 5 | MLN-3897 TFA | CCR3 antagonist 1 | INCB-9471 | CCR6 antagonist 1 |
| 相關(guān)庫 | 抑制劑庫 | 腫瘤免疫治療小分子化合物庫 | 經(jīng)典已知活性庫 | 已知活性化合物庫 | 膜蛋白靶向化合物庫 | 口服活性化合物庫 |