αvβ3 (IC50 = 4 nM, αvβ3-Vitronectin interaction); αvβ5 (IC50 = 79 nM, αvβ5-Vitronectin interaction); αvβ3 (IC50 = 0.61 nM); αvβ5 (IC50 = 8.4 nM); α5β1 (IC50 = 14.9 nM); STAT3

αvβ3 和 αvβ5 整合素受体的拮抗剂是西仑吉肽 (EMD 121974)。在分别评估人肺癌细胞系 UCLA-P3 或黑色素瘤 M21 细胞粘附的研究中，克仑吉肽可减少整合素介导的玻连蛋白结合，IC50 分别为 0.4 和 0.4 μM [1]。在体外，西仑吉肽在浓度高于 1 μM 时表现出浓度和时间依赖性细胞毒性作用 [2]。

将西来吉肽（10、50和250μg）每周3次腹腔注射给患有M21-L黑色素瘤的裸鼠；这些剂量被证明可以抑制肿瘤生长，同时缩小肿瘤体积（分别为 55%、75% 和 55%）。肿瘤重量（分别为 23%、38% 和 61%）和 89% [2]。在所研究的大鼠模型中，ILP单独与西仑吉肽联合给药、ILP与西仑吉肽联合美法仑、TNF或两者联合给药并不影响腹膜内施用的西仑吉肽的全身药代动力学。腹腔注射治疗10分钟后，西仑吉肽的全身水平达到约20μg/mL（约35μM），并在第一小时内持续上升至约40μg/mL（约70μM）。此后，赛利必肽血清水平的消除半衰期为 2.1 小时 [3]。

整合素结合试验[Sci Rep. 2017 Jan 11;7:39805.]
整合素配体的活性和选择性通过固相结合试验确定，根据先前报道的方案，使用包被的细胞外基质蛋白和可溶性整合素。以下列化合物为内标:Cilengitide/西伦吉肽，c(RGDf(NMe)V) (αvβ3-0.54 nM， αvβ5-8 nM， α5β1-15.4 nM)，线性肽RTDLDSLRT4 (αvβ6-33 nM;8 - 100 nM)和vαβtirofiban5 (IIbαβ3 - 1.2海里)。
用ecm蛋白(1)(每孔100 μL)在碳酸缓冲液(15 mM Na2CO3, 35 mM NaHCO3, pH 9.6)中在4°C下包被96孔平底ELISA板过夜。然后用pbs - t缓冲液(磷酸盐缓冲盐水/Tween20, 137 mM NaCl, 2.7 mM KCl, 10 mM Na2HPO4, 2 mM KH2PO4, 0.01% Tween20, pH 7.4)洗涤每孔;3 × 200 μL)，室温下用ts -b缓冲液(Tris-saline/BSA缓冲液;150μL /;20 mM Tris-HCl, 150 mM NaCl, 1 mM CaCl2, 1 mM MgCl2, 1 mM MnCl2, pH 7.5, 1% BSA)。同时，将化合物和内标品以1:5的稀释步骤，从20 μM到6.4 nM，在另一个板上配制稀释系列。用PBS-T (200 μL)洗涤三次后，从B-G中每孔转移50 ul稀释系列。A孔填入100 ul tsb溶液(空白)，H孔填入50 ul ts -b缓冲液。将人整合素(2)在ts -b缓冲液中的溶液50 ul转移到h -b孔中，rt孵育1 h, PBS-T缓冲液洗涤3次，然后加入一抗(3)(每孔100 μL)。rt孵育1 h后，用PBS-T洗涤3次。然后，在板中加入二次过氧化物酶标记抗体(4)(100 μL/孔)，rt孵育1 h。PBS-T洗涤三次后，快速加入SeramunBlau (50 μL/孔，Seramun Diagnostic GmbH, Heidesee, Germany)， rt孵育5 min。用3 M H2SO4 (50 μL/孔)停止反应，在450 nm处用平板仪测定吸光度。每个化合物的IC50分两份进行测试，用OriginPro 7.5G软件分析得到的抑制曲线。拐点表示IC50值。测定的IC50均参照内标活度。

蛋白质印迹分析[Bioengineered. 2022 Feb;13(2):4557-4572.]
细胞类型： B16 和 A375 细胞
测试浓度： 0、5、10 和 20 μg/mL
孵育持续时间： 12 小时
实验结果： 在浓度大于 5 μg/mL 时抑制 PD-L1 表达和 STAT3 磷酸化。

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细胞凋亡分析[3]
细胞类型： B16 和 A375 细胞
测试浓度： 5 μg/mL
孵育时间：12小时
实验结果：B16和A375细胞的凋亡率分别为15.27%和14.89%。

Animal/Disease Models: Nude mice bearing M21-L melanoma tumors[1]
Doses: 10, 50, and 250 μg
Route of Administration: Dosed ip three times per week
Experimental Results: Demonstrated inhibition of tumor growth with a reduction in both tumor volume (55%, 75%, and 89%, respectively) and tumor weight (23%, 38%, and 61%, respectively), when compared to controls.

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Animal/Disease Models: Female C57BL/6 mice (6-8 weeks old) with B16 cells sc[Bioengineered. 2022 Feb;13(2):4557-4572.]
Doses: 50 mg/kg; with or without 10 mg/kg Anti-PD1 monoclonal antibody or isotype control ip every 3 days;
Route of Administration: intraperitoneal (ip)injection; daily
Experimental Results: Downregulated the expression of PD-L1 via STAT3 pathway and diminished the expression of PD-L1.

Dissolved in PBS; 100μg; i.p. injection

Human glioblastoma xenografts U87 MG

[1]. Assessment of the biological and pharmacological effects of the alpha nu beta3 and alpha nu beta5 integrinreceptor antagonist, Cilengitide (EMD 121974), in patients with advanced solid tumors. Ann Oncol. 2007 Aug;18(8):1400-7.

[2]. Combination therapy of cilengitide with belotecan against experimental glioblastoma. Int J Cancer. 2013 Aug 1;133(3):749-56.

[3]. The αVβ3/αVβ5 integrin inhibitor cilengitide augments tumor response to melphalan isolated limb perfusion in a sarcoma model. Int J Cancer. 2012 Nov 13.

Background: Cilengitide, an antiangiogenic agent that inhibits the binding of integrins alpha(nu)beta(3) and alpha(nu)beta(5) to the extracellular matrix, was studied at two dose levels in cancer patients to determine the optimal biological dose. Patients and methods: The doses of cilengitide were 600 or 1200 mg/m(2) as a 1-h infusion twice weekly every 28 days. A novel dose escalation scheme was utilized that relied upon the biological activity rate. Results: Twenty patients received 50 courses of cilengitide with no dose-limiting toxic effects. The pharmacokinetic (PK) profile revealed a short elimination half-life of 4 h, supporting twice weekly dosing. Of the six soluble angiogenic molecules assessed, only E-selectin increased significantly from baseline. Analysis of tumor microvessel density and gene expression was not informative due to intrapatient tumor heterogeneity. Although several patients with evaluable tumor biopsy pairs did reveal posttreatment increases in tumor and endothelial cell apoptosis, these results did not reach statistical significance due to the aforementioned heterogeneity. Conclusions: Cilengitide is a well-tolerated antiangiogenic agent. The biomarkers chosen in this study underscore the difficulty in assessing the biological activity of antiangiogenic agents in the absence of validated biological assays.[1]

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The prognosis of patients diagnosed with glioblastoma remains dismal in spite of the current concomitant chemoradiotherapy with temozolomide. In particular, the resistance to temozolomide appears to be the greatest obstacle to the treatment of glioblastoma. In the present study, we evaluated in vitro and in vivo the antitumor effects of combination therapy of cilengitide with belotecan, a camptothecin derivate, to treat experimental glioblastoma. The therapeutic effects of the drugs on the U87MG and U251MG human glioblastoma cell lines were assessed using in vitro cell viability and apoptosis assays. The combination treatment group with cilengitide and belotecan enhanced the cytotoxic effects to the glioblastoma cell lines and increased the apoptosis of the tumor cells compared to monotherapy with either drug alone in vitro. Nude mice with established U87MG glioblastoma were assigned to the following four groups: control, cilengitide, belotecan and combination treatment. The volume of tumors and length of survival were also measured. Animals in the combination therapy group demonstrated a significant reduction of tumor volume and an increase in survival (p < 0.05). Immunohistochemistry revealed a decrease in angiogenesis by cilengitide and an increase in apoptosis by cilengitide and belotecan in vivo. The combination therapy of cilengitide with belotecan presented more cytotoxic effects compared to the monotherapy of either drug in vitro and in vivo. This combination protocol may serve as an alternative treatment option for glioblastoma.[2]

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Isolated limb perfusion (ILP) with melphalan and tumor necrosis factor (TNF)-α is used to treat bulky, locally advanced melanoma and sarcoma. However, TNF toxicity suggests a need for better-tolerated drugs. Cilengitide (EMD 121974), a novel cyclic inhibitor of alpha-V integrins, has both anti-angiogenic and direct anti-tumor effects and is a possible alternative to TNF in ILP. In this study, rats bearing a hind limb soft tissue sarcoma underwent ILP using different combinations of melphalan, TNF and cilengitide in the perfusate. Further groups had intra-peritoneal (i.p.) injections of cilengitide or saline 2 hr before and 3 hr after ILP. A 77% response rate (RR) was seen in animals treated i.p. with cilengitide and perfused with melphalan plus cilengitide. The RR was 85% in animals treated i.p. with cilengitide and ILP using melphalan plus both TNF and cilengitide. Both RRs were significantly greater than those seen with melphalan or cilengitide alone. Histopathology showed that high RRs were accompanied by disruption of tumor vascular endothelium and tumor necrosis. Compared with ILP using melphalan alone, the addition of cilengitide resulted in a three to sevenfold increase in melphalan concentration in tumor but not in muscle in the perfused limb. Supportive in vitro studies indicate that cilengitide both inhibits tumor cell attachment and increases endothelial permeability. Since cilengitide has low toxicity, these data suggest the agent is a good alternative to TNF in the ILP setting.[3]

C29H41F3N8O9

702.68

702.295

C, 55.09; H, 6.85; N, 19.04; O, 19.03

199807-35-7

Cilengitide;188968-51-6; 199807-35-7 (TFA); 188969-00-8 (HCl)

129626550

cyclo[L-arginyl-glycyl-L-alpha-aspartyl-D-phenylalanyl-N-methyl-L-valyl] trifluoroacetic acid

cyclo[Arg-Gly-Asp-D-Phe-N(Me)Val].TFA

White to off-white solid powder

1.111

273.21

8

13

9

49

1110

4

CC(C)[C@H]1C(=O)N[C@H](C(=O)NCC(=O)N[C@H](C(=O)N[C@@H](C(=O)N1C)CC2=CC=CC=C2)CC(=O)O)CCCN=C(N)N.C(=O)(C(F)(F)F)O

WHJCSACXAPYNTG-LOPTWHKWSA-N

InChI=1S/C27H40N8O7.C2HF3O2/c1-15(2)22-25(41)33-17(10-7-11-30-27(28)29)23(39)31-14-20(36)32-18(13-21(37)38)24(40)34-19(26(42)35(22)3)12-16-8-5-4-6-9-16;3-2(4,5)1(6)7/h4-6,8-9,15,17-19,22H,7,10-14H2,1-3H3,(H,31,39)(H,32,36)(H,33,41)(H,34,40)(H,37,38)(H4,28,29,30);(H,6,7)/t17-,18-,19+,22-;/m0./s1

2-[(2S,5R,8S,11S)-5-benzyl-11-[3-(diaminomethylideneamino)propyl]-7-methyl-3,6,9,12,15-pentaoxo-8-propan-2-yl-1,4,7,10,13-pentazacyclopentadec-2-yl]acetic acid;2,2,2-trifluoroacetic acid

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 请将本产品存放在密封且受保护的环境中，避免吸湿/受潮。

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

配方 1 中的溶解度: 16.67 mg/mL (23.72 mM) in PBS (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液; 超声助溶。

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配方 2 中的溶解度: 30% Propylene glycol , 5% Tween 80 , 65% D5W: 30 mg/mL

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。