【正文】 的補(bǔ)充。Figure 2. Comparison of the effects of MOMIPP (pound 19) and MIPP (pound 2) on growth and viability of U251 GBM cells. (A). One day after plating the cells in 96well dishes, MOMIPP (●) or MIPP (■) was added at the indicated concentrations. Controls consisted of cells in parallel wells treated with an equivalent volume of vehicle (DMSO). Medium containing fresh pound was added after 24 h, and MTT assays were performed after a total 48 h of treatment. Each point represents the mean (177。到48小時(shí)，細(xì)胞活力減少約40％，而在類(lèi)似的細(xì)胞密度下同期處理的MCF7乳腺癌細(xì)胞減少了70％（圖5F）。 SD), with the results expressed as percent of the mean from parallel control dishes treated with an equivalent volume of vehicle alone (DMSO). (C) Parental and doxorubicinresistant (DoxR) MCF7 breast cancer cells(56) were treated with 10 μM MOMIPP for 24 h and then examined by phasecontrast microscopy. (D) Longterm viability of parental or DoxR MCF7 cells was assessed by colonyforming assay after a 48 h of treatment with the indicated concentrations of MOMIPP. The results are the mean 177。然而，在這一點(diǎn)上，我們的SAR研究還沒(méi)有發(fā)現(xiàn)任何非本質(zhì)的數(shù)據(jù)庫(kù)，可以鏈接到一個(gè)龐大的親和標(biāo)簽庫(kù)（例如，生物素）或拴了沒(méi)有失去活性的堅(jiān)實(shí)基體。這個(gè)區(qū)域選擇性的特點(diǎn)是一維的Overhauser核效應(yīng)（NOE），這在試驗(yàn)階段也有陳述。 (2) NaN3, 75% AcOH, 0 176?；衔?6（5疊氮MIPP）的合成基于2烷基吲哚5位的直接硝化和進(jìn)一步的疊氮結(jié)構(gòu)重氮化轉(zhuǎn)換（26）（見(jiàn)計(jì)劃3B）。Figure 6. Comparison of the effects of MOMIPP (pound 19), 5azidoMIPP (pound 36), and 6azidoMOMIPP (pound 41) on morphology and viability of U251 glioblastoma cells. (A) The indicated pounds were added to the cells at concentrations of or 10 μM, and the cells were observed by phasecontrast microscopy after 48 h. (B and C) U251 cells were treated with varying concentrations of the indicated pounds for 48 h, and longterm cell viability was assessed by colonyforming assays. Each represents the counts from three dishes (mean 177。在幾乎所有情況中，產(chǎn)品從溶液中析出，然后用簡(jiǎn)單沖洗的方法很容易的從多余的催化劑（或起始原料）中純化出來(lái)。像2 吡啶（化合物10），3 吡啶（化合物11和16），吡嗪（化合物17），或苯基類(lèi)似物（化合物9）這些類(lèi)似物是無(wú)效的。這些類(lèi)型的觀(guān)測(cè)促使了人們對(duì)那些通過(guò)對(duì)其特定受體的共價(jià)性改造而起效的藥物的興趣的普遍高漲。在文獻(xiàn)中描述的所有具有細(xì)胞毒性的查耳酮類(lèi)化合物中，與MOMIPP最相似的那個(gè)化合物含有一個(gè)連接到3,4,5 三甲氧基苯基基取代的取代吲哚環(huán)。歸根結(jié)底，這一機(jī)制的澄清將取決于MOMIPP及相關(guān)化合物（S）的特定的分子靶點(diǎn)的鑒定。因此，如果MOMIPP是以共價(jià)鍵結(jié)合到靶蛋白（S），有可能光起始劑反應(yīng)中間體的形成是沒(méi)有必要的。第二個(gè)挑戰(zhàn)涉及到MOMIPP和其活性類(lèi)似物的弱水溶性。后者可能提供額外的好處：通過(guò)對(duì)腫瘤特異性肽或抗體運(yùn)載工具的表面改性允許非特異性藥物有選擇性地選擇受體。在抗TMZ的GBM細(xì)胞以及耐阿霉素藥乳腺癌細(xì)胞中MOMIPP有效地誘導(dǎo)methuosis。在這方面， MIPP吲哚環(huán)的5位引入光活性疊氮化合物后產(chǎn)生了一種保留了良好的methuosis誘導(dǎo)活性（圖6）的衍生物，我們的這項(xiàng)發(fā)現(xiàn)提供了幾種按照確定的步驟鑒定蛋白質(zhì)受體的途徑。在10μM濃度下，我們用這種化合物處理U251細(xì)胞48小時(shí)，但沒(méi)有引起明顯的細(xì)胞空泡化或死亡。后者的化合物被設(shè)計(jì)成許多秋水仙堿的和能結(jié)合β微管蛋白的天然產(chǎn)品bretastatins的類(lèi)似物。親電基團(tuán)使具有親電基團(tuán)的內(nèi)在基質(zhì)變?yōu)榧?xì)胞親核試劑，他們?cè)谒幬镌O(shè)計(jì)上不常使用，因?yàn)樗麄兛梢噪S意修改許多生物大分子。首先，關(guān)于吲哚環(huán)，氮的甲基化（化合物20）或去除2 甲基（化合物13）降低但并未消除活性。這里，我們描述了類(lèi)似查耳酮的小分子MIPP，它能夠在GBM和其他腫瘤細(xì)胞株中誘導(dǎo)methuosis。溫和重氮化條件下，氨基吲哚轉(zhuǎn)化為芳香疊氮化合物34。C, 88%. Compound 36: 4acetylpyridine, piperidine, MeOH, reflux, 39%. Compound 37: NaNO3, H2SO4, 0 176。隨后的形態(tài)和可行性研究顯示，與芐衍生物（化合物14）不同的是，當(dāng)在U251細(xì)胞（數(shù)據(jù)未顯示）測(cè)試時(shí)，沒(méi)有一個(gè)苯甲酮類(lèi)似物（30日和31日）能誘導(dǎo)methuosis。（18）最初，我們?cè)O(shè)計(jì)了一個(gè)MIPP的苯甲類(lèi)似物作為潛在的光親和探針。SD) from four separate wells. (G) Normal human skin fibroblasts were treated with 10 μM MOMIPP or an equivalent volume of DMSO (control) and examined by phasecontrast microscopy after 24 h. (H) Normal human fibroblasts were seeded in 96well plates at subconfluent density (1000 cells/well) or confluent density (10000 cells/well). After 24 h, fresh medium containing 10 μM MOMIPP or an equal volume of vehicle (DMSO) was added. MTT viability assays were performed at a 48 h end point. Values are the means (177。在所有其他細(xì)胞測(cè)試實(shí)驗(yàn)中，到24小時(shí)10μMMOMIPP明顯誘導(dǎo)成纖維細(xì)胞空泡化（圖5G）。 SD from three separate cultures.Figure 3. Comparison of the abilities of MOMIPP and MIPP to induce the morphological hallmarks of methuosis. One day after plating, U251 GBM cells were treated with MOMIPP or MIPP at final concentrations of (A) or 10 μM (B). Controls received an equivalent vo