| 名稱 | Raltegravir |
| 描述 | Raltegravir (MK-0518) is a pyrrolidinone derivative and HIV INTEGRASE INHIBITOR that is used in combination with other ANTI-HIV AGENTS for the treatment of HIV INFECTION. |
| 細(xì)胞實(shí)驗(yàn) | Human MT-4 cells are infected for 2 hours with the SIVmac251, HIV-1 (IIIB) and HIV-2 (CDC 77618) stocks at a multiplicity of infection of, approximately, 0.1. Cells are then washed three times in phosphate buffered saline, and suspended at 5 × 105/mL in fresh culture medium (to primary cells 50 units/mL of IL-2 are added) in 96-well plates, in the presence or absence of a range of triplicate raltegravir concentrations (0.0001 μM -1 μM). Untreated infected and mock-infected controls are prepared too, in order to allow comparison of the data derived from the different treatments. Viral cytopathogeniciy in MT-4 cells is quantitated by the methyl tetrazolium (MTT) method (MT-4/MTT assay) when extensive cell death in control virus-infected cell cultures is detectable microscopically as lack of capacity to re-cluster. The capability of MT-4 cells to form clusters after infection. Briefly, clusters are disrupted by pipetting; and, after 2 hours of incubation at 37 °C, the formation of new clusters is assessed by light microscopy (100 × magnification). Cell culture supernatants are collected for HIV-1 p24 and HIV-2/SIVmac251 p27 core antigen measurement by ELISA. In CEMx174-infected cell cultures, which show a propensity to form syncytia induced by the virus envelope glycoproteins, syncytia are counted, in blinded fashion, by light microscopy for each well at 5 days following infection. (Only for Reference) |
| 激酶實(shí)驗(yàn) | PFV integration assay: For quantitative strand transfer assays, donor DNA substrate is formed by annealing HPLC grade oligonucleotides 5′-GACTCACTATAGGGCACGCGTCAAAATTCCATGACA and 5′-ATTGTCATG GAATTTTGACGCGTGCCCTATAGTGAGTC. Reactions (40 μL) contains 0.75 μM PFV IN, 0.75 μM donor DNA, 4 nM (300 ng) supercoiled pGEM9-Zf(?) target DNA, 125 mM NaCl, 5 mM MgSO4, 4 μM ZnCl2, 10 mM DTT, 0.8% (vol/vol) DMSO, and 25 mM BisTris propane–HCl, pH 7.45. Raltegravir is added at indicated concentrations. Reactions are initiated by addition of 2 μL PFV IN diluted in 150 mM NaCl, 2 mM DTT, and 10 mM Tris-HCl, pH 7.4, and stopped after 1 hour at 37 °C by addition of 25 mM EDTA and 0.5% (wt/vol) SDS. Reaction products, deproteinized by digestion with 20 μg proteinase K for 30 minutes at 37 °C followed by ethanol precipitation, are separated in 1.5% agarose gels and visualized by staining with ethidium bromide. Integration products are quantified by quantitative real-time PCR, using Platinum SYBR Green qPCR SuperMix and three primers: 5′-CTACTTACTCTAGCTTCCCGGCAAC, 5′-TTCGCCAGTTAATAGTTTGCGCAAC, and 5′-GACTCACTATAGGGCACGCGT. PCR reactions (20 μL) contained 0.5 μM of each primer and 1 μL diluted integration reaction product. Following a 5-min denaturation step at 95 °C, 35 cycles are carried out in a CFX96 PCR instrument, using 10 seconds denaturation at 95 °C, 30 seconds annealing at 56 °C and 1 minutes extension at 68 °C. Standard curves are generated using serial dilutions of WT PFV IN reaction in the absence of INSTI. |
| 體外活性 | Raltegravir誘導(dǎo)非人靈長(zhǎng)類病毒的免疫學(xué)改善,并伴有進(jìn)展性SIVmac251感染. |
| 體內(nèi)活性 | Raltegravir在體外作用于人類T淋巴細(xì)胞培養(yǎng)物,有效作用于HIV-1,抑制達(dá)95%時(shí)濃度為31±20 nM。S217Q PFV IN對(duì)Raltegravir的敏感性至少與WT酶相似。在急性感染的人類淋巴CD4 + T細(xì)胞系MT-4和CEMx174中,Raltegravir高效抑制SIVmac251復(fù)制,EC90處于低納摩爾范圍。Raltegravir微弱抑制肝細(xì)胞色素P450。Raltegravir不會(huì)誘導(dǎo)CYP3A4 RNA表達(dá)或CYP3A4依賴的睪丸激素6-β-羥化酶活性。Raltegravir在鎂和鈣存在時(shí),細(xì)胞擴(kuò)散性降低。Raltegravir和相關(guān)的HIV-1整合酶鏈轉(zhuǎn)移抑制劑高效阻斷病毒復(fù)制。 |
| 存儲(chǔ)條件 | Powder: -20°C for 3 years | In solvent: -80°C for 1 year | Shipping with blue ice. |
| 溶解度 | H2O : < 1 mg/mL (insoluble or slightly soluble)
DMSO : 82 mg/mL (184.5 mM)
Ethanol : < 1 mg/mL (insoluble or slightly soluble)
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| 關(guān)鍵字 | HIV | inhibit | HIV Integrase | Raltegravir | MK0518 | Inhibitor | MK 0518 | Human immunodeficiency virus |
| 相關(guān)產(chǎn)品 | Tenofovir | Valproic Acid | Chloroquine phosphate | Emtricitabine | Dolutegravir intermediate-1 | Lamivudine | 5-Fluorouracil | Decanedioic acid | Dextran sulfate sodium salt (MW 4500-5500) | Stavudine | Kaempferol | Dimethyl fumarate |
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