VS-5584 (SB-2343)   中文名称: VS-5584

PI3Kα (IC50 = 16 nM); PI3Kγ (IC50 = 25 nM); PI3Kδ (IC50 = 42 nM); PI3Kβ (IC50 = 68 nM); Vps34 (IC50 = 7470 nM); mTOR (IC50 = 37 nM); DNA-PK (IC50 = 1270 nM); mTORC1; mTORC2

VS-5584 是一种 ATP 竞争性抑制剂，可选择性抑制 PI3K/mTOR 信号传导，对所有人类 I 类 PI3K 同工型和 mTOR 激酶具有同等的低纳摩尔效力。 VS-5584 在 HMLE 乳腺癌细胞中的 EC50 为 15 nM，对癌症干细胞的选择性约为 10 倍。在 HMLER 永生化乳腺癌细胞系中，VS-5584 优先减少 CD44Hi/CD24Lo 细胞。 VS-5584 成功根除 SUM159 细胞中的癌症干细胞侧群。在大型人类癌细胞系筛选（436 个细胞系）中发现了广泛的抗增殖敏感性，并且具有 PI3KCA 突变的细胞通常对 VS-5584 治疗更敏感。

在携带三阴性乳腺癌肿瘤的小鼠中，口服 VS-5584 可以减少肿瘤干细胞，并在紫杉烷耐药模型中诱导肿瘤消退。在 PTENnull 人前列腺 PC3 异种移植模型中，用 VS-5584 治疗，11 毫克/公斤和 25 毫克/公斤分别显着抑制肿瘤生长 (TGI) 79% 和 113%。在 FLT3-ITD AML 异种移植模型中，VS-5584 治疗可对肿瘤生长产生剂量依赖性抑制（3.7 mg/kg 时抑制 28%，11 mg/kg 时抑制 76%）。 [1]

反应混合物由以下成分组成，溶于 10 μL 测定缓冲液（50 mM Hepes pH 7.5、10 mM MgCl 2、3 mM MnCl 2、1 mM EGTA、2 mM DTT、0.01% Tween-20）：10 μM ATP、0.05 μM ULight-eIF4E 结合蛋白 1 (Thr37/46) 肽和 0.10 μg/mL 自制 mTOR 酶。在室温下，将混合物孵育 60 分钟。然后，添加 1X LANCE® 检测缓冲液和 10 μL 检测混合物，其中包括 16 mM EDTA、0.004 mM Eu-W1024 标记的抗 Phospho-eIF4E 结合蛋白 1-(Thr37/46) 抗体。然后将混合物孵育 60 分钟。

对于 96 孔板中的增殖测定，使用 SET-2、SNU-478、SNU-1196、SNU-245、SNU-1079、SNU-308、SNU-869 和 MKN7 细胞。使用多发性骨髓瘤细胞（H929、MM1.S、MM1.R、R8226、U266）和鼻咽细胞（CNE-1、CNE-2、HONE1、HK1）。第二天，贴壁细胞以 30% 至 50% 汇合度接种，悬浮细胞以 2,000 至 6,000 个细胞接种后，用浓度高达 10 μM 的 VS-5584（一式三份）处理细胞 48 小时。 CellTiter-Glo 测定用于追踪细胞活力。使用 XL-fit 程序绘制剂量反应曲线以确定化合物的 IC50 值。

Mice: Athymic BALB/c nude mice (BALB/cOlaHsd-Foxn1nu) are used. Fox-Chase severe combined immunodeficient (SCID) mice (CB17/Icr-Prkdcscid/CrlBltw) are used. The right flank of male (PC3 and COLO 205), female (MV4-11 and HuH7), or male (NCI-N87) BALB/c nude mice is intradermally implanted with 5106 (PC3, COLO205, HuH7, NCI-N87), or 1107 (MV4-11) cells. Using a 27.5-gauge needle, cells are resuspended in 70% (v/v; only for COLO205 and HuH7) or 50% (v/v) serum-free growth medium/Matrigel and injected in a total volume of 100 L. Dosing began seven to fourteen days after tumor implantation. Oral doses of VS-5584 (11 and 25 mg/kg) are administered daily[1].

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Dosing started 7 to 14 days after tumor implantation. VS-5584 was dosed daily orally. The reference compounds everolimus and gefitinib were dosed p.o. at 5 and 150 mg/kg, respectively, with everolimus dosed daily and gefitinib dosed for 5 day-on and 2 day-off in cycles. 5-FU was administered intraperitoneally, at 25 mg/kg, every Tuesday, Thursday, and Saturday. All statistics conducted were done using GraphPad Prism (v5).

Pharmacokinetic and pharmacodynamic properties of VS-5584 To investigate the efficacy of VS-5584 in disease models, the pharmacokinetic and pharmacodynamic profile of VS-5584 was determined to enable the selection of an optimal dosing schedule. A single oral dose of VS-5584 was rapidly absorbed with a tmax of 0.9 hours and an elimination half-life of 10 hours (Supplementary Fig. S3). To determine the pharmacokinetic and pharmacodynamic relationship in tumors, PC3-tumor–bearing mice were treated with a single dose of VS-5584 and plasma and tumors were harvested after 6 hours and analyzed for concentrations of VS-5584 and effects on target efficacy biomarkers. Plasma levels of VS-5584 increased dose-dependently (Fig. 2A). Plasma pharmacokinetic was not significantly different to tumor pharmacokinetic. Drug levels exceeded the IC50 for inhibition of the target kinases in the enzymatic and cell-based assays starting from 3.7 mg/kg. Dose-dependent inhibition of pAkt(S473) and pS6 (S240/244) was observed in tumor tissue with complete inhibition from 33 mg/kg (EC50 of 4.2 and 1.7 mg/kg; Fig. 2B). To study the time course of drug levels and inhibition of target kinase signaling in plasma and tumor, PC3-tumor–bearing mice were treated with a single oral dose of 33 mg/kg VS-5584 and the tissues harvested 1, 6, and 24 hours postdosing. The plasma concentration of VS-5584 following the 33 mg/kg dose of VS-5584 was highest 1 hour after dosing (1221 ng/mL or 3.55 μmol/L) and was still above concentrations required to block the targets in in vitro assays after 24 hours (15 ng/mL or 43 nmol/L; Fig. 2C). pAkt(S473) and pS6(S240/244) were blocked by 90% or more within 1 hour of VS-5584 treatment and remained inhibited by 60% to 70% after 24 hours (Fig. 2D).[1]
In summary, VS-5584 shows good oral bioavailability with dose-linear pharmacokinetic and a profound and long-lasting pharmacodynamic response in tumor tissue following a single oral dose in tumor bearing mice.[1]

[1]. VS-5584, a novel and highly selective PI3K/mTOR kinase inhibitor for the treatment of cancer. Mol Cancer Ther, 2013, 12(2), 151-161.

VS-5584 has been used in trials studying the treatment of Lymphoma, Metastatic Cancer, and Non Hematologic Cancers.
PI3K/mTOR Kinase Inhibitor VS-5584 is a potent and selective inhibitor of both phosphatidylinositol 3 kinase (PI3K) and mammalian target of rapamycin (mTOR) kinase in the PI3K/mTOR signaling pathway, with potential antineoplastic activity. PI3K/mTOR kinase inhibitor VS-5584 inhibits mTOR kinase and all class I PI3K isoforms. Consequently, this disrupts phosphorylation of substrates downstream of PI3K and mTOR and may result in apoptosis and growth inhibition in susceptible tumor cells. Activation of the PI3K/mTOR pathway promotes cell growth, survival, and resistance to chemotherapy and radiotherapy. mTOR is a serine/threonine kinase downstream of PI3K, which also has PI3K-independent activity. Consequently, this agent may potentially be more potent than an agent that inhibits either PI3K kinase or mTOR kinase.

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Dysregulation of the PI3K/mTOR pathway, either through amplifications, deletions, or as a direct result of mutations, has been closely linked to the development and progression of a wide range of cancers. Moreover, this pathway activation is a poor prognostic marker for many tumor types and confers resistance to various cancer therapies. Here, we describe VS-5584, a novel, low-molecular weight compound with equivalent potent activity against mTOR (IC(50) = 37 nmol/L) and all class I phosphoinositide 3-kinase (PI3K) isoforms IC(50): PI3Kα = 16 nmol/L; PI3Kβ = 68 nmol/L; PI3Kγ = 25 nmol/L; PI3Kδ = 42 nmol/L, without relevant activity on 400 lipid and protein kinases. VS-5584 shows robust modulation of cellular PI3K/mTOR pathways, inhibiting phosphorylation of substrates downstream of PI3K and mTORC1/2. A large human cancer cell line panel screen (436 lines) revealed broad antiproliferative sensitivity and that cells harboring mutations in PI3KCA are generally more sensitive toward VS-5584 treatment. VS-5584 exhibits favorable pharmacokinetic properties after oral dosing in mice and is well tolerated. VS-5584 induces long-lasting and dose-dependent inhibition of PI3K/mTOR signaling in tumor tissue, leading to tumor growth inhibition in various rapalog-sensitive and -resistant human xenograft models. Furthermore, VS-5584 is synergistic with an EGF receptor inhibitor in a gastric tumor model. The unique selectivity profile and favorable pharmacologic and pharmaceutical properties of VS-5584 and its efficacy in a wide range of human tumor models supports further investigations of VS-5584 in clinical trials.[1]

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In summary, the favorable target selectivity profile, pharmacokinetic and pharmacodynamic properties of VS-5584 and as a consequence its efficacy in a range of tumors resistant to rapalogs and standard of care, provide a compelling rationale for the clinical evaluation of this drug in a range of liquid and solid tumor indications. The genetic markers of sensitivity and resistance identified in the large cell panel screen provide a rationale for patient selection for single-agent therapy as well as for drug combinations.[1]

C17H22N8O

354.40958

354.19166

C, 57.61; H, 6.26; N, 31.62; O, 4.51

1246560-33-7

1246560-33-7

46912230

White to off-white crystalline solid

1.7±0.1 g/cm3

1.796

2.41

90.88

1

8

3

26

466

0

NC1=NC=C(C2=C3N=C(C)N(C(C)C)C3=NC(N4CCOCC4)=N2)C=N1

QYBGBLQCOOISAR-UHFFFAOYSA-N

InChI=1S/C17H22N8O/c1-10(2)25-11(3)21-14-13(12-8-19-16(18)20-9-12)22-17(23-15(14)25)24-4-6-26-7-5-24/h8-10H,4-7H2,1-3H3,(H2,18,19,20)

5-(8-methyl-2-morpholin-4-yl-9-propan-2-ylpurin-6-yl)pyrimidin-2-amine

VS5584; VS5584; VS 5584; 1246560-33-7; VS-5584; 5-(9-Isopropyl-8-methyl-2-morpholino-9H-purin-6-yl)pyrimidin-2-amine; VS-5584 (SB2343); 5-(8-methyl-2-morpholin-4-yl-9-propan-2-ylpurin-6-yl)pyrimidin-2-amine; UNII-W71J4X250V; SB2343; SB2343; SB 2343

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

DMSO: ~71 mg/mL (~200.3 mM)
Water: <1 mg/mL
Ethanol: ~3 mg/mL (~8.5 mM)

配方 1 中的溶解度: ≥ 2.5 mg/mL (7.05 mM) (饱和度未知) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 25.0 mg/mL澄清DMSO储备液加入到400 μL PEG300中，混匀；然后向上述溶液中加入50 μL Tween-80，混匀；加入450 μL生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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配方 2 中的溶解度: ≥ 2.5 mg/mL (7.05 mM) (饱和度未知) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 mg/mL澄清DMSO储备液加入900 μL 20% SBE-β-CD生理盐水溶液中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

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配方 3 中的溶解度: ≥ 2.5 mg/mL (7.05 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 mg/mL 澄清 DMSO 储备液添加到 900 μL 玉米油中并混合均匀。

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配方 4 中的溶解度: 0.5% methylcellulose+0.2%Tween 80: 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。