PRMT1 (IC50 = 30 nM); PRMT3 (IC50 = 119 nM); PRMT4 (IC50 = 83 nM); PRMT6 (IC50 = 4 nM); PRMT8 (IC50 = 5 nM)

MS023 二盐酸盐（1-1000 nM；48 小时）可抑制 MCF7 细胞中的 PRMT1 甲基转移酶活性 [1]。 MS023 二盐酸盐（1-1000 nM；20 小时）可抑制 HEK293 细胞中的 PRMT6 甲基转移酶活性 [1]。

MS023 diHClide (160 mg/kg, ip) 与 PKC412 (100 mg/kg, ig) 结合通过减少功能性 MLL-r ALL 起始细胞的支持来防止 MLL-r 及时循环 (ALL)。 ）

PRMT生化测定[1]
闪烁邻近度测定（SPA）用于评估试验化合物对抑制PRMT催化的甲基转移反应的影响，如前所述。27简言之，氚化的S-腺苷-L-甲硫氨酸被用作甲基的供体。将（3H）甲基化生物素标记肽捕获在链亲和素/闪烁体包被的微孔板中，使掺入的3H甲基和闪烁体接近，从而产生发光，通过追踪TopCount NXT™微孔板闪烁和发光计数器测量的放射性信号（每分钟计数）来量化发光。必要时，使用非氚化SAM来补充反应。在平衡条件下，通过滴定反应混合物中的测试化合物，在底物和辅因子的Km浓度下测定IC50值。

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细胞PRMT1测定[1]
MCF7细胞在补充有10%FBS、青霉素（100单位mL−1）和链霉素（100μg mL−1。将40%汇合的细胞用不同浓度的MS023和化合物4-6以指定浓度或DMSO对照处理48小时。将细胞在100μL总裂解缓冲液（20 mM Tris-HCl pH 8、150 mM NaCl、1 mM EDTA、10 mM MgCl2、0.5%TritonX-100、12.5 U mL−1苯并酶、完全不含EDTA的蛋白酶抑制剂混合物）中裂解。在RT下孵育3分钟后，将SDS添加到最终1%的浓度。在SDS-PAGE上运行裂解物，并如下所述进行免疫印迹以在蛋白质印迹中测定H4R3me2a、精氨酸不对称二甲基化、精氨酰对称二甲基化和精氨酸单甲基化。

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细胞PRMT6测定[1]
HEK293细胞在补充有10%FBS、青霉素（100 U mL−1）和链霉素（100μg mL−1）的DMEM中的12孔板中生长。按照制造商的说明，使用jetPRIME®转染试剂（Polyplus Transferion），用FLAG标记的PRMT6或突变体V86K/D88A PRMT6（每孔1μg DNA）转染50%的融合细胞。4小时后移除培养基，并用指示浓度的MS023或DMSO对照处理细胞。20小时后，取出培养基，并在100μL的总裂解缓冲液中裂解细胞。

蛋白质印迹分析 [1]
细胞类型： MCF7 和 HEK293 细胞
测试浓度： 1.4、4、12、37、111、333 和 1000 nM
孵育持续时间：MCF7细胞为48小时； HEK293 细胞 20 小时 (hrs (hours))
实验结果：：治疗有效且浓度依赖性地降低 H4R3me2a 的细胞水平 (IC50=9±0.2 nM)。处理浓度依赖性地减少 H3R2me2a 标记 (IC50=56±7 nM)。

Animal/Disease Models: NOD-scid IL2Rgnull (NSG) mice carrying primary MLL-r ALL cells [2]
Doses: 160 mg/kg
Route of Administration: intraperitoneal (ip) injection; spread of [2]. Results of 4 weeks of PKC412 (100 mg/kg, ig), MS023 (160 mg/kg, ip) or combination treatment: Combination treatment prolonged the survival of leukemia mice relative to single treatment.

[1]. Eram MS, et al. A Potent, Selective, and Cell-Active Inhibitor of Human Type I Protein Arginine Methyltransferases. ACS Chem Biol. 2016 Mar 18;11(3):772-81.
[2]. Yinghui Zhu, et al. Targeting PRMT1-mediated FLT3 methylation disrupts maintenance of MLL-rearranged acute lymphoblastic leukemia. Blood. 2019 Oct 10;134(15):1257-1268.

Protein arginine methyltransferases (PRMTs) play a crucial role in a variety of biological processes. Overexpression of PRMTs has been implicated in various human diseases including cancer. Consequently, selective small-molecule inhibitors of PRMTs have been pursued by both academia and the pharmaceutical industry as chemical tools for testing biological and therapeutic hypotheses. PRMTs are divided into three categories: type I PRMTs which catalyze mono- and asymmetric dimethylation of arginine residues, type II PRMTs which catalyze mono- and symmetric dimethylation of arginine residues, and type III PRMT which catalyzes only monomethylation of arginine residues. Here, we report the discovery of a potent, selective, and cell-active inhibitor of human type I PRMTs, MS023, and characterization of this inhibitor in a battery of biochemical, biophysical, and cellular assays. MS023 displayed high potency for type I PRMTs including PRMT1, -3, -4, -6, and -8 but was completely inactive against type II and type III PRMTs, protein lysine methyltransferases and DNA methyltransferases. A crystal structure of PRMT6 in complex with MS023 revealed that MS023 binds the substrate binding site. MS023 potently decreased cellular levels of histone arginine asymmetric dimethylation. It also reduced global levels of arginine asymmetric dimethylation and concurrently increased levels of arginine monomethylation and symmetric dimethylation in cells. We also developed MS094, a close analog of MS023, which was inactive in biochemical and cellular assays, as a negative control for chemical biology studies. MS023 and MS094 are useful chemical tools for investigating the role of type I PRMTs in health and disease.[1]

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Relapse remains the main cause of MLL-rearranged (MLL-r) acute lymphoblastic leukemia (ALL) treatment failure resulting from persistence of drug-resistant clones after conventional chemotherapy treatment or targeted therapy. Thus, defining mechanisms underlying MLL-r ALL maintenance is critical for developing effective therapy. PRMT1, which deposits an asymmetric dimethylarginine mark on histone/non-histone proteins, is reportedly overexpressed in various cancers. Here, we demonstrate elevated PRMT1 levels in MLL-r ALL cells and show that inhibition of PRMT1 significantly suppresses leukemic cell growth and survival. Mechanistically, we reveal that PRMT1 methylates Fms-like receptor tyrosine kinase 3 (FLT3) at arginine (R) residues 972 and 973 (R972/973), and its oncogenic function in MLL-r ALL cells is FLT3 methylation dependent. Both biochemistry and computational analysis demonstrate that R972/973 methylation could facilitate recruitment of adaptor proteins to FLT3 in a phospho-tyrosine (Y) residue 969 (Y969) dependent or independent manner. Cells expressing R972/973 methylation-deficient FLT3 exhibited more robust apoptosis and growth inhibition than did Y969 phosphorylation-deficient FLT3-transduced cells. We also show that the capacity of the type I PRMT inhibitor MS023 to inhibit leukemia cell viability parallels baseline FLT3 R972/973 methylation levels. Finally, combining FLT3 tyrosine kinase inhibitor PKC412 with MS023 treatment enhanced elimination of MLL-r ALL cells relative to PKC412 treatment alone in patient-derived mouse xenografts. These results indicate that abolishing FLT3 arginine methylation through PRMT1 inhibition represents a promising strategy to target MLL-r ALL cells.[2]

C17H28CL3N3O

396.78272

287.19976

C, 51.46; H, 7.11; Cl, 26.80; N, 10.59; O, 4.03

2108631-19-0

MS023;1831110-54-3;MS023 dihydrochloride;1992047-64-9

92136227

Typically exists as solids (or liquids in special cases) at room temperature

1.1±0.1 g/cm3

437.8±45.0 °C at 760 mmHg

218.6±28.7 °C

0.0±1.1 mmHg at 25°C

1.567

2.3

54.3Ų

C(C1=CNC=C1C1C=CC(OC(C)C)=CC=1)N(C)CCN.Cl

VEUUSCXROKBMMJ-UHFFFAOYSA-N

InChI=1S/C17H25N3O.3ClH/c1-13(2)21-16-6-4-14(5-7-16)17-11-19-10-15(17)12-20(3)9-8-18;;;/h4-7,10-11,13,19H,8-9,12,18H2,1-3H3;3*1H

N1-((4-(4-isopropoxyphenyl)-1H-pyrrol-3-yl)methyl)-N1-methylethane-1,2-diamine trihydrochloride

MS023 trihydrochloride); MS023 triHCl

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

注意: 如下所列的是一些常用的体内动物实验溶解配方，主要用于溶解难溶或不溶于水的产品（水溶度<1 mg/mL）。 建议您先取少量样品进行尝试，如该配方可行，再根据实验需求增加样品量。

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注射用配方1: DMSO : Tween 80： Saline = 10 : 5 : 85 (如： 100 μL DMSO → 50 μL Tween 80 → 850 μL Saline)
*生理盐水/Saline的制备：将0.9g氯化钠/NaCl溶解在100 mL ddH ₂ O中，得到澄清溶液。
注射用配方 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (如： 100 μL DMSO → 400 μL PEG300 → 50 μL Tween 80 → 450 μL Saline)
注射用配方 3: DMSO : Corn oil = 10 : 90 (如： 100 μL DMSO → 900 μL Corn oil)
示例: 以注射用配方 3 (DMSO : Corn oil = 10 : 90) 为例说明, 如果要配制 1 mL 2.5 mg/mL的工作液, 您可以取 100 μL 25 mg/mL 澄清的 DMSO 储备液，加到 900 μL Corn oil/玉米油中, 混合均匀。

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注射用配方 4: DMSO : 20% SBE-β-CD in Saline = 10 : 90 [如：100 μL DMSO → 900 μL (20% SBE-β-CD in Saline)]
*20% SBE-β-CD in Saline的制备（4°C，储存1周）：将2g SBE-β-CD (磺丁基-β-环糊精) 溶解于10mL生理盐水中，得到澄清溶液。
注射用配方 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (如： 500 μL 2-Hydroxypropyl-β-cyclodextrin (羟丙基环胡精)→ 500 μL Saline)
注射用配方 6: DMSO : PEG300 : Castor oil : Saline = 5 : 10 : 20 : 65 (如： 50 μL DMSO → 100 μL PEG300 → 200 μL Castor oil → 650 μL Saline)
注射用配方 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (如： 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
注射用配方 8: 溶解于Cremophor/Ethanol (50 : 50), 然后用生理盐水稀释。
注射用配方 9: EtOH : Corn oil = 10 : 90 (如： 100 μL EtOH → 900 μL Corn oil)
注射用配方 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (如： 100 μL EtOH → 400 μL PEG300 → 50 μL Tween 80 → 450 μL Saline)

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口服配方 1: 悬浮于0.5% CMC Na (羧甲基纤维素钠)
口服配方 2: 悬浮于0.5% Carboxymethyl cellulose (羧甲基纤维素)
示例: 以口服配方 1 (悬浮于 0.5% CMC Na)为例说明, 如果要配制 100 mL 2.5 mg/mL 的工作液, 您可以先取0.5g CMC Na并将其溶解于100mL ddH2O中，得到0.5%CMC-Na澄清溶液；然后将250 mg待测化合物加到100 mL前述 0.5%CMC Na溶液中，得到悬浮液。

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口服配方 3: 溶解于 PEG400 (聚乙二醇400)
口服配方 4: 悬浮于0.2% Carboxymethyl cellulose (羧甲基纤维素)
口服配方 5: 溶解于0.25% Tween 80 and 0.5% Carboxymethyl cellulose (羧甲基纤维素)
口服配方 6: 做成粉末与食物混合

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注意: 以上为较为常见方法，仅供参考， InvivoChem并未独立验证这些配方的准确性。具体溶剂的选择首先应参照文献已报道溶解方法、配方或剂型，对于某些尚未有文献报道溶解方法的化合物，需通过前期实验来确定（建议先取少量样品进行尝试），包括产品的溶解情况、梯度设置、动物的耐受性等。

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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。