With IC50 values less than 10 nM, valemetostat tosylate (1-1000 nM) substantially and selectively inhibits EZH1 and EZH2 [3]. H3K27me3 is efficiently increased and unintentional increases are prevented by valemetostat tosylate (100 nM; 7 d) [3]. 0.1-100 nM; 7 d) Uses low-dose therapy to effectively block the H3K27me3 pore size model in sensitive cells [3].

甲苯磺酸伐美司他 (valemetostat tosylate) (1-1000 nM) 的 IC50 值小于 10 nM，可显着且选择性地抑制 EZH1 和 EZH2 [3]。甲苯磺酸伐美司他 (valemetostat tosylate)（100 nM；7 d）可有效增加 H3K27me3 并防止意外增加 [3]。 0.1-100nM； 7 d) 使用低剂量疗法有效阻断敏感细胞中的 H3K27me3 孔径模型 [3]。

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Valemetostat（也称为 (R)-OR-S2 或 DS-3201）是一种有效的 SAM 竞争性 EZH1 和 EZH2 甲基转移酶双重抑制剂，对两种酶的 IC₅₀ 值均 ≤ 10 nM。在使用 HCT116 细胞的检测中，它能降低全局 H3K27me3 水平。在一组 23 种血液恶性肿瘤细胞系（包括成人 T 细胞白血病淋巴瘤 (ATL)、外周 T 细胞淋巴瘤 (PTCL)、野生型和突变型 EZH2 的弥漫性大 B 细胞淋巴瘤 (DLBCL)、伯基特淋巴瘤 (BL)、原发性渗出性淋巴瘤 (PEL) 和 MLL 重排白血病）中，与 EZH2 选择性抑制剂（GSK126, E7438）相比，它显示出更优越的抗增殖活性。生长抑制 IC₅₀ 值均低于 1000 nM，其中 ATL 和其他 T 细胞淋巴瘤特别敏感。在 ATL 细胞系（TL-Om1）和原代 ATL 细胞中，用 100 nM 的 valemetostat 处理 7 天能有效去除 H3K27me3 标记，将 EZH1 和 EZH2 从染色质上驱逐，重新激活沉默基因的表达（如 SLA、PAG1、BTG1、BTG2），使 NF-κB 信号失活，并诱导细胞凋亡。在去除 H3K27me3 和抑制细胞生长方面，这种双重抑制比 10 倍剂量的 EZH2 选择性抑制剂 GSK126 更有效。在染色质相关基因（ARID1A、KDM6A、BAP1、SMARCB1）失活或被致癌病毒（HTLV-1、EBV）感染的癌症细胞模型中也观察到了合成致死效应。处理不会破坏核心 PRC2 复合物的组成，但减少了 EZH1/EZH2 在染色质上的占据。[3]

在皮下接种 ATL 来源的 TL-Om1 细胞的免疫缺陷（NOD/SCID）小鼠中，每日口服 200 mg/kg 的 valemetostat（以 OR-S1 形式，同一系列的相关 EZH1/2 双重抑制剂）完全阻止了肿瘤植入。在治疗模型中，从接种后第 14 天开始每日口服 OR-S1（100-200 mg/kg/天）显著抑制了已建立的 TL-Om1 异种移植瘤的生长并抑制了肝转移，且未引起显著的体重减轻。[3]

使用闪烁亲近分析法进行 PRC2-EZH2 和 PRC2-EZH1 的体外甲基转移酶活性检测。反应混合物包含 Tris-HCl 缓冲液 (pH 8.8)、MgCl₂、DTT、BSA、未标记的 S-腺苷甲硫氨酸 (SAM)、氚标记的 SAM ([³H]-SAM) 以及生物素化的组蛋白 H3 肽段作为底物。加入重组 PRC2-EZH2 或 PRC2-EZH1 复合物以启动反应。在链霉亲和素包被的板中于室温孵育 2 小时后，测量氚标记甲基基团从 [³H]-SAM 到生物素化 H3 肽段的转移。为了测定 IC₅₀，将 valemetostat 等测试化合物进行系列稀释并加入反应混合物。使用 S 型剂量反应模型分析数据。[3]

对于抗增殖实验，将淋巴瘤或实体瘤细胞系接种到多孔板中，并用溶解于 DMSO 的 valemetostat 或其他 EZH 抑制剂的一系列浓度进行处理。细胞培养 14 天，每 3-4 天传代至含有化合物的新鲜培养基中。在处理期结束时评估细胞活力/生长。对于原代 ATL 细胞活力实验，将患者来源的 PBMCs 与 IL-2 一起培养，并用抑制剂处理 7 天，然后进行细胞计数检测。为了评估细胞中 H3K27me3 的减少，用化合物处理 HCT116 细胞 3 天，并使用基于 AlphaLISA 技术的特定细胞检测试剂盒定量全局 H3K27me3 水平。对于染色质免疫沉淀 (ChIP) 实验，细胞用甲醛交联，裂解，染色质通过微球菌核酸酶消化和超声处理进行剪切。用针对 H3K27me3、EZH1、EZH2 或其他组蛋白标记的抗体对剪切后的染色质进行免疫沉淀。纯化沉淀的 DNA，并通过定量 PCR 或微阵列 (ChIP-chip) 分析以确定在特定基因组位点的富集情况。[3]

Animal/Disease Models: Male C57BL/6J mice performed chronic and acute running exercise or no exercise [1]
Doses: 0.01 mg/g.
Route of Administration: intraperitoneal (ip) injection; 0.01 mg/g; 30 minutes before the start of running.
Experimental Results: H3K27me3 levels increased Dramatically after exercise, EZH1 levels diminished slightly, EZH2 levels increased, and phosphorylated AMPK levels increased. Inhibits myonuclear H3K27me3 accumulation during training and leads to failure of adaptive changes.

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For tumor prevention studies, NOD/SCID mice were subcutaneously inoculated with TL-Om1 cells in the post-auricular region. Starting from the day of inoculation, mice were orally administered valemetostat (as OR-S1) suspended in a 0.5% (w/v) methylcellulose solution at a dose of 200 mg/kg daily. Tumor formation was monitored. For therapeutic efficacy studies, NOG mice were subcutaneously inoculated with TL-Om1 cells. After tumors were established (day 14), mice were randomized into groups and treated orally with OR-S1 at doses of 100 or 200 mg/kg daily. Tumor dimensions were measured regularly to calculate volume, and body weight was monitored. [3]

Repeated dosing of valemetostat (as part of the OR-S series) in preclinical models did not cause any critical or severe toxicity in vitro or in vivo. In the mouse xenograft studies, daily oral administration of OR-S1 at 100-200 mg/kg did not lead to significant body weight loss in the treated mice. [3]

[1]. Daiichi Sankyo’s EZH1/2 Dual Inhibitor Valemetostat (DS-3201) Receives SAKIGAKE Designation for Treatment of Patients with Relapsed/Refractory Peripheral T-Cell Lymphoma from Japan MHLW.

[2]. Shimizu J, Kawano F. Exercise-induced histone H3 trimethylation at lysine 27 facilitates the adaptation of skeletal muscle to exercise in mice. J Physiol. 2022 Jul;600(14):3331-3353.

[3]. Targeting Excessive EZH1 and EZH2 Activities for Abnormal Histone Methylation and Transcription Network in Malignant Lymphomas. Cell Rep. 2019 Nov 19;29(8):2321-2337.e7.

Valemetostat Tosylate is the tosylate form of valemetostat, an orally available selective inhibitor of the histone lysine methyltransferases enhancer of zeste homolog 1 (EZH1) and 2 (EZH2), with potential antineoplastic activity. Upon oral administration, valemetostat selectively inhibits the activity of both wild-type and mutated forms of EZH1 and EZH2. Inhibition of EZH1/2 specifically prevents the methylation of lysine 27 on histone H3 (H3K27). This decrease in histone methylation alters gene expression patterns associated with cancer pathways, enhances transcription of certain target genes, and results in decreased proliferation of EZH1/2-expressing cancer cells. EZH1/2, histone lysine methyltransferase (HMT) class enzymes and catalytic subunits of the polycomb repressive complex 2 (PRC2), are overexpressed or mutated in a variety of cancer cells and play key roles in tumor cell proliferation, progression, stem cell self-renewal and migration.

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Valemetostat is an EZH1 and EZH2 dual inhibitor. The study provides a mechanistic rationale for dual targeting, showing that EZH1 and EZH2 are co-expressed in many lymphomas and have reciprocal/interfering functions on chromatin. Inhibition of only EZH2 can lead to compensatory recruitment of EZH1 and residual H3K27me3, whereas dual inhibition effectively remodels the oncogenic epigenome. The drug shows potent activity against a wide range of hematological malignancies, particularly those overexpressing EZH2 (e.g., ATL, PTCL) or harboring mutations in epigenetic modifier genes (ARID1A, SMARCA4, SMARCB1, KDM6A, BAP1, KMT2D). It is also effective against pre-malignant cells epigenetically perturbed by oncovirus infections (HTLV-1, EBV). Based on these results, a phase 1 clinical trial against T- and B-cell non-Hodgkin’s lymphomas, including ATL, was initiated. [3]

C33H42CLN3O7S

660.220487117767

659.243

1809336-93-3

Valemetostat;1809336-39-7

126482037

White to yellow solid powder

143

3

8

6

45

1090

1

ClC1=CC(C(NCC2C(NC(C)=CC=2C)=O)=O)=C(C)C2=C1O[C@](C)(C1CCC(CC1)N(C)C)O2.S(C1C=CC(C)=CC=1)(=O)(=O)O

JSBKGJUYSLVFPF-RRKMXGHKSA-N

InChI=1S/C26H34ClN3O4.C7H8O3S/c1-14-11-15(2)29-25(32)20(14)13-28-24(31)19-12-21(27)23-22(16(19)3)33-26(4,34-23)17-7-9-18(10-8-17)30(5)6;1-6-2-4-7(5-3-6)11(8,9)10/h11-12,17-18H,7-10,13H2,1-6H3,(H,28,31)(H,29,32);2-5H,1H3,(H,8,9,10)/t17?,18?,26-;/m1./s1

(2R)-7-chloro-2-[4-(dimethylamino)cyclohexyl]-N-[(4,6-dimethyl-2-oxo-1H-pyridin-3-yl)methyl]-2,4-dimethyl-1,3-benzodioxole-5-carboxamide;4-methylbenzenesulfonic acid

DS-3201 tosylateDS3201 tosylate Ezharmia

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)

DMSO : ~100 mg/mL (~151.46 mM)

配方 1 中的溶解度: ≥ 2.08 mg/mL (3.15 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 20.8 mg/mL澄清DMSO储备液加入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: ≥ 2.08 mg/mL (3.15 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 20.8 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: ≥ 2.08 mg/mL (3.15 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 20.8 mg/mL 澄清 DMSO 储备液加入到 900 μL 玉米油中并混合均匀。

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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。