BAY-1436032

BAY-1436032 is a brand-new pan-mutant inhibitor of isocitrate dehydrogenase 1 (IDH1). In mouse hematopoietic cells expressing IDH1R132H or IDH1R132C, BAY-1436032 reduces intracellular (R)-2-hydroxyglutarate (R-2HG) synthesis with IC50 values of 60 and 45 nM, respectively. Even at dosages up to 10 μM, BAY-1436032 does not lower R-2HG levels in mouse hematopoietic cells expressing IDH2R140Q and IDH2R172K. At doses as high as 100 μM, BAY-1436032 did not impede the colony expansion of patient-derived IDH1 wild-type AML cells, but it did at 50% when it came to that growth. BAY-1436032 significantly enhanced myelomonocytic differentiation of myeloid progenitor cells in morphological assessment [1].

BAY-1436032是一种全新的异柠檬酸脱氢酶1（IDH1）全突变抑制剂。在表达 IDH1R132H 或 IDH1R132C 的小鼠造血细胞中，BAY-1436032 减少细胞内 (R)-2-羟基戊二酸 (R-2HG) 的合成，IC50 值分别为 60 和 45 nM。即使剂量高达 10 μM，BAY-1436032 也不会降低表达 IDH2R140Q 和 IDH2R172K 的小鼠造血细胞中的 R-2HG 水平。在高达 100 μM 的剂量下，BAY-1436032 不会阻碍患者来源的 IDH1 野生型 AML 细胞的集落扩张，但在增长时会阻碍 50%。 BAY-1436032 在形态学评估中显着增强了骨髓祖细胞的骨髓单核细胞分化 [1]。

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BAY-1436032 选择性抑制表达 IDH1R132H（IC₅₀ 60 nM）或 IDH1R132C（IC₅₀ 45 nM）的永生化小鼠造血细胞内的 R-2HG 产生。[1]
BAY-1436032 能有效抑制携带各种 IDH1R132 突变的人原代 AML 细胞中 R-2HG 的产生（IC₅₀ 在 3 到 16 nM 之间），但在浓度高达 1 μM 时对 IDH2 突变细胞几乎没有影响。[1]
BAY-1436032 在甲基纤维素克隆形成实验中抑制人原代 IDH1 突变 AML 细胞的集落形成，在 0.1 μM 浓度下抑制约 50%。即使浓度高达 100 μM，也不抑制 IDH1 野生型 AML 细胞的集落生长。[1]
BAY-1436032 在原代 IDH1 突变 AML 细胞的体外培养中诱导髓系分化，表现为表面分化标志物 CD14 和 CD15 的上调，以及形态学上显示单核细胞/粒细胞成熟的改变。[1]
BAY-1436032 处理（500 nM，14 天）降低了原代 IDH1R132C 和 IDH1R132H 突变 AML 细胞中组蛋白 H3 在 H3K4、H3K9、H3K27 和 H3K36 位点的全局三甲基化水平，但对 IDH1 野生型细胞无此作用。[1]
使用 Illumina Infinium HumanMethylation450 平台进行的 DNA 甲基化分析表明，BAY-1436032 处理（500 nM，14 天）改变了特定的甲基化模式，包括降低 IDH1 突变 AML 细胞中 PLU1 启动子的甲基化和增加 E2F1 启动子的甲基化。[1]

在 150 mg/kg 剂量下，每天一次长期口服 BAY-1436032 治疗显示几乎完全抑制 (R)-2-羟基戊二酸 (R-2HG) 合成。接受载体治疗的小鼠的白细胞计数稳步上升；在给予 45 mg/kg BAY-1436032 的小鼠中，增加速度减慢；在 150 mg/kg 组中，数字没有变化。第 60 天，媒介物组和 45 mg/kg 组的血红蛋白水平略低于 150 mg/kg 组，而用 BAY-1436032 以 45 mg/kg 组和媒介物组治疗的小鼠的血红蛋白水平略低于 150 mg/kg 组。 150毫克组的动物。第 60 天，该队列的血小板计数/公斤显着减少。虽然用媒介物治疗的动物死亡，但所有给予 150 mg/kg BAY-1436032 的小鼠均存活，并且外周血中的 hCD45+ 细胞负荷最低，直至治疗开始后第 150 天观察结束（P<0.001）。位的使用寿命为 91 天。 45 mg/kg 剂量的 BAY-1436032 在小鼠体内产生中等表达的 CD14/CD15 [1]。

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在 IDH1R132C 突变 AML 的患者来源异种移植（PDX1）小鼠模型中，移植后 17 天开始，每日一次口服 150 mg/kg 的 BAY-1436032，持续 150 天，导致血清 R-2HG 水平持续抑制，外周血中人 CD45⁺ (hCD45⁺) 白血病母细胞显著减少，并延长了生存期（所有小鼠存活至第 150 天，而溶剂组中位生存期为 91 天）。[1]
在第二个独立的、伴有 MLL-PTD 共突变的 IDH1R132C 突变 AML PDX 模型（PDX2）中，移植后 90 天开始，每日一次口服 150 mg/kg BAY-1436032 治疗 100 天，抑制了血清 R-2HG，降低了外周血 hCD45⁺ 母细胞计数，并显著延长了生存期（10 只小鼠中有 6 只存活至治疗停止的第 100 天，中位生存期未达到，而溶剂组为 62 天）。[1]
BAY-1436032 在体内诱导白血病细胞髓系分化，表现为经治疗的 PDX 小鼠外周血中表达 CD14 和 CD15 的 hCD45⁺ 细胞比例增加，以及 CD34⁺ 祖细胞比例减少。[1]
BAY-1436032 在体内耗竭了白血病干细胞（LSCs）。有限稀释移植实验显示，与溶剂处理的小鼠相比，用 BAY-1436032（150 mg/kg）治疗的小鼠骨髓细胞中 LSC 频率降低了约 100 倍。[1]
对来自治疗后的 PDX 小鼠的 hCD45⁺ 细胞进行的基因表达谱分析显示，干性相关基因（如 SPARC、CD69、DNMT3B）下调，而髓系分化和免疫反应相关基因上调。细胞周期分析显示，BAY-1436032 治疗后，处于 G₀/G₁ 期的 hCD45⁺ 细胞增加，S 期细胞减少。[1]

为了评估 R-2HG 产生的抑制，用逆转录病毒工程化表达突变 IDH1 或 IDH2 的 HoxA9 永生化小鼠骨髓细胞用 BAY-1436032 或 DMSO 处理 8 天。收获细胞，通过质谱法测量细胞裂解液中 R-2HG 与 S-2-羟基戊二酸（S-2HG）的比值。[1]
从患者新鲜分离的原代人 AML 细胞用 BAY-1436032 或 DMSO 处理 24 小时。测量细胞裂解液中 R-2HG 与 S-2HG 的比值以确定 IC₅₀ 值。[1]
对于克隆形成祖细胞实验，将人原代 AML 单核细胞接种在补充了细胞因子以及 BAY-1436032 或溶剂的甲基纤维素培养基中。10-14 天后在显微镜下计数集落。[1]
为了评估分化，将原代 IDH1 突变 AML 细胞在含有 BAY-1436032 或 DMSO 的悬浮培养基中培养数天。然后通过流式细胞术分析 CD14 和 CD15 的表达，并在染色后通过细胞涂片进行形态学评估。[1]
对于组蛋白甲基化分析，原代 AML 细胞用 500 nM BAY-1436032 或 DMSO 处理 14 天。提取组蛋白，通过 Western blotting 分析 H3K4me3、H3K9me3、H3K27me3 和 H3K36me3 的全局水平，并定量条带强度相对于总组蛋白 H3 的比值。[1]

For pharmacokinetic and pharmacodynamic studies, mice were administered BAY-1436032 orally by gavage at doses of 45, 90, or 150 mg/kg. Blood was collected at various time points to measure plasma drug concentrations and serum R-2HG levels. [1]
For efficacy studies in PDX models, NOD-scid IL2Rγnull (NSG) mice were transplanted intravenously with primary human IDH1 mutant AML cells. Once engraftment was established (as measured by hCD45⁺ cells in peripheral blood), treatment was initiated. [1]
In the PDX1 model, treatment started 17 days post-transplantation. Mice (n=10 per group) received vehicle, 45 mg/kg, or 150 mg/kg BAY-1436032 by oral gavage once daily for 150 days. [1]
In the PDX2 model, treatment started 90 days post-transplantation to mimic a higher disease burden. Mice (n=10 per group) received vehicle or 150 mg/kg BAY-1436032 by oral gavage once daily for 100 days, after which treatment was stopped and mice were monitored for relapse. [1]
Mice were monitored for survival, body weight, and peripheral blood counts. Peripheral blood was periodically analyzed by flow cytometry for hCD45⁺ cells and differentiation markers (CD14, CD15, CD34, CD33). Serum R-2HG levels were measured by mass spectrometry or enzymatic assay. [1]
For limiting dilution assays to assess LSC frequency, bone marrow cells were harvested from PDX mice treated with vehicle or BAY-1436032 (150 mg/kg) for 4 weeks. Serially diluted cells were transplanted into secondary NSG recipients, and engraftment was assessed to calculate LSC frequency using Poisson statistics. [1]

Plasma exposure of BAY-1436032 was almost dose-linear between 45 and 150 mg/kg in mice. [1]
Unbound plasma concentrations covered the in vitro IC₅₀ for R-2HG inhibition for 24 hours after dosing. [1]
Oral administration of BAY-1436032 led to a rapid decline in serum R-2HG levels, detectable as early as 3 hours post-dose. [1]
Long-term once-daily oral dosing with 150 mg/kg BAY-1436032 resulted in nearly complete suppression of R-2HG production in PDX mice. [1]

[1]. Pan-mutant-IDH1 inhibitor BAY1436032 is highly effective against human IDH1 mutant acute myeloid leukemia in vivo. Leukemia. 2017 Oct;31(10):2020-2028.

Pan-Mutant-IDH1 Inhibitor Bay-1436032 is an orally available pan-inhibitor of mutant forms of the metabolic enzyme isocitrate dehydrogenase type 1 (IDH1; IDH-1; IDH1 [NADP+] soluble), including forms with mutations of arginine 132 (IDH1(R132)), with potential antineoplastic activity. Upon administration, pan-mutant-IDH-1 inhibitor BAY-1436032 specifically inhibits the activity of IDH1 mutant forms, which prevents the formation of the oncometabolite 2-hydroxyglutarate (2HG) from alpha-ketoglutarate (a-KG). This prevents 2HG-mediated signaling and leads to both an induction of cellular differentiation and an inhibition of cellular proliferation in tumor cells expressing IDH1 mutant forms. IDH1 mutations, including IDH1(R132) mutations, are highly expressed in certain malignancies; they initiate and drive cancer growth by both blocking cell differentiation and catalyzing the formation of 2HG.

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BAY-1436032 is a novel, orally bioavailable pan-mutant IDH1 inhibitor developed to target all major IDH1R132 mutations found in cancer. [1]
Its mechanism of action involves inhibiting the neomorphic activity of mutant IDH1, thereby reducing the production of the oncometabolite R-2HG. This leads to reversal of histone and DNA hypermethylation, release of the differentiation block, inhibition of proliferation, and depletion of leukemic stem cells. [1]
The study demonstrates that BAY-1436032 is highly effective against human IDH1 mutant acute myeloid leukemia in vivo, leading to blast clearance, differentiation, and prolonged survival in preclinical PDX models. [1]
Based on this preclinical work, a clinical phase I trial for patients with IDH1R132 mutant AML was being initiated at the time of publication. [1]

C26H30F3N3O3

489.529917240143

489.22

C, 63.79; H, 6.18; F, 11.64; N, 8.58; O, 9.80

1803274-65-8

118310260

White to gray solid powder

7.2

76.4

2

8

7

35

726

2

C[C@H]1C[C@H](CC(C1)(C)C)N2C3=C(C=C(C=C3)CCC(=O)O)N=C2NC4=CC=C(C=C4)OC(F)(F)F

RNMAUIMMNAHKQR-QFBILLFUSA-N

InChI=1S/C26H30F3N3O3/c1-16-12-19(15-25(2,3)14-16)32-22-10-4-17(5-11-23(33)34)13-21(22)31-24(32)30-18-6-8-20(9-7-18)35-26(27,28)29/h4,6-10,13,16,19H,5,11-12,14-15H2,1-3H3,(H,30,31)(H,33,34)/t16-,19+/m0/s1

3-(2-((4-(trifluoromethoxy)phenyl)amino)-1-((1R,5R)-3,3,5-trimethylcyclohexyl)-1H-benzo[d]imidazol-5-yl)propanoic acid

BAY-1436032; BAY 1436032; BAY1436032.

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 : ~125 mg/mL (~255.35 mM)

配方 1 中的溶解度: ≥ 2.5 mg/mL (5.11 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 (5.11 mM) 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), 悬浊液; 超声助溶。
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 (5.11 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。