Cdk4/cyclin D1 (IC50 = 2 nM); CDK6/cyclinD1 (IC50 = 10 nM); CDK9/cyclinT1 (IC50 = 57 nM); CDK5/p35 (IC50 = 287 nM); Cdk5/p25 (IC50 = 355 nM); CDK2/cyclinE (IC50 = 504 nM); CDK7/Mat1/cyclinH1 (IC50 = 3910 nM); CDK1/cyclinB1 (IC50 = 1627 nM); PIM1 (IC50 = 39 nM); PIM2 (IC50 = 3400 nM); HIPK2 (IC50 = 31 nM); DYRK2 (IC50 = 61 nM); CK2 (IC50 = 117 nM); GSK3b (IC50 = 192 nM); JNK3 (IC50 = 389 nM); FLT3 (D835Y) (IC50 = 403 nM); FLT3 (IC50 = 3960 nM); DRAK1 (IC50 = 659 nM)

体外活性：Abemaciclib（以前称为 LY2835219）是一种有效、选择性、口服的 CDK4（细胞周期蛋白依赖性激酶）和 CDK6 双重抑制剂，在无细胞测定中 IC50 分别为 2 nM 和 10 nM。 LY2835219 特异性抑制 CDK4 和 6，从而抑制 G1 早期视网膜母细胞瘤 (Rb) 蛋白磷酸化。抑制 Rb 磷酸化可阻止 CDK 介导的 G1-S 相变，从而将细胞周期阻滞在 G1 期，抑制 DNA 合成并抑制癌细胞生长。丝氨酸/苏氨酸激酶 CDK4/6 的过度表达可导致细胞周期失调，如某些类型的癌症中所见。激酶测定：将细胞 (5 × 103) 铺在 96 孔板中。第二天将细胞处理 24 至 48 小时，然后根据制造商的说明和发光板读数器，通过 Caspase-Glo-3/7 测定法评估 caspase-3 活性。细胞测定：将细胞接种到 96 孔板中，使其粘附过夜，并用 DMSO 对照 (0.1% v/v) 或指定化合物处理 72 小时。根据制造商的说明，使用细胞计数试剂盒测定细胞活力和增殖。使用 CompuSyn 确定 LY2835219 和 mTOR 抑制剂之间的相互作用。组合指数 (CI) 值为 1 表示药物相互作用相加，而 CI < 1 表示协同作用，CI > 1 表示拮抗作用。

LY2835219 使 BBB 流出饱和，未结合血浆 IC50 约为 95 nM。 LY2835219-MsOH 在大脑中的剂量百分比为 0.5–3.9%。在皮下和颅内人胶质母细胞瘤模型 (U87MG) 中，LY2835219-MsOH 作为单一药物以及与替莫唑胺联合使用，均以剂量依赖性方式抑制肿瘤生长。

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Abemaciclib单药治疗导致肿瘤生长延迟，这与肿瘤中T细胞炎症特征增加有关。联合抗pd - l1治疗可导致肿瘤完全消退和免疫记忆，并伴有抗原呈递增强、T细胞炎症表型和细胞周期控制增强。[3]

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研究人员在一项多中心研究中评估了abemaciclib的安全性、药代动力学特征、药效学效应和抗肿瘤活性。abemaciclib是一种口服生物可用的细胞周期蛋白依赖性激酶（CDK） 4和6抑制剂，包括乳腺癌、非小细胞肺癌（NSCLC）、胶质母细胞瘤、黑色素瘤和结直肠癌的I期剂量递增和肿瘤特异性队列。共纳入225例患者：33例剂量递增组，192例肿瘤特异性组。剂量限制毒性为3级疲劳。最大耐受剂量为每12小时200毫克。最常见的可能与治疗相关的不良事件包括疲劳和胃肠道、肾脏或造血系统。血浆浓度随剂量增加而增加，在增殖角化细胞和肿瘤中观察到药效学效应。在先前接受过治疗的乳腺癌、非小细胞肺癌和黑色素瘤患者中，放射学反应得到了缓解。对于激素受体阳性的乳腺癌，总有效率为31%；此外，61%的患者达到缓解或病情稳定持续≥6个月。［２］

LY2835219 (abemaciclib)是由礼来公司研究实验室的科学家通过化合物和生化筛选鉴定出来的，并因其生物活性和对CDK4/ cyclin D1复合物(IC50 =2 nmol/L)和CDK6/cyclin D1复合物(IC50 =10 nmol/L)的高度选择性抑制而被选中，在纳摩尔范围内对其他CDK/cyclin复合物或细胞周期相关激酶没有活性，除了CDK9的IC50至少高出5倍(图2)23该化合物被证明是CDK4和CDK6的atp结合域的竞争性抑制剂，对CDK4的抑制作用是对CDK6的14倍。与palbociclib和ribociclib相比，abemaciclib对复合物CDK4/cyclin D1具有更高的选择性，IC50值比其他两种化合物低5倍[1]。

将细胞接种到 96 孔板中，然后让其粘附一整夜，然后用所示化合物或 DMSO 对照（0.1% v/v）处理 72 小时。按照制造商的指示，使用细胞计数试剂盒来评估细胞的活力和增殖。 CompuSyn用于分析mTOR抑制剂和Abemaciclib之间的关系。联合指数 (CI) 值为 1 表示相加药物相互作用，而 CI 值 <1 或 >1 表示协同或拮抗药物相互作用。

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体外治疗人乳腺癌及T细胞[3]
用DMSO和abemaciclib (500 nM)治疗乳腺癌细胞系MCF-7和MDA-MB-46 8天。按照上述方法分离RNA。用抗cd3 /CD28/CD2或thapsigarin刺激Jurkat T细胞或原代人T细胞，并按指示用abemaciclib孵育。采用RosetteSep试剂盒阴性选择从全血中分离人原代T细胞。T细胞在RPMI1640中培养，添加10%胎牛血清（FCS）、100 U/mL青霉素、100 μg/mL链霉素、10 mM HEPES和2 mM l -谷氨酰胺，并用CD3/CD28 DynaBeads按1:3的T细胞/头比刺激。Abemaciclib以0.3 μM的终浓度加入。每2-3天对细胞进行计数并饲喂新鲜培养基。

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体外肿瘤细胞活力测定[3]
肿瘤细胞在37℃下单独培养4小时，然后加入abemaciclib、palbociclib或DMSO对照，按指定浓度在37℃下培养96小时。然后使用CellTiter-Glo 评估细胞活力。细胞活力抑制率（%）按公式[1−（处理样品的平均光度/未处理对照的平均光度）]× 100计算。使用四参数logistic曲线拟合公式计算生长或活力抑制的50%抑制浓度（IC50）。

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NFAT报告试验[3]
将含有NFAT-luc报告基因的Jurkat T细胞与abemaciclib按指定浓度孵育30分钟，然后用250 ng/mL抗cd3 /CD28/CD2刺激6小时。使用荧光素酶测定试剂在Wallac 1420 Victor2多标签计数器 中监测NFAT-luc活性。使用未经处理的对照样品计算NFAT刺激（%）。

Subcutaneous injections of OSC-19 (1×10⁶) cells are given to six-week-old BALB/c female nude mice. Mice are randomized by tumor size and given each treatment when tumor sizes approach 100 mm³. Each treatment group comprises a minimum of 5 mice. Every group of mice receives a daily oral gavage dose of either RAD001 (5 mg/kg/d), Abemaciclib (45 mg/kg/d or 90 mg/kg/d), or a combination of both. In 20 mM phosphate buffer (pH 2.0), 1% HEC is used to dissolve the Abemaciclib. Weight and tumor size are measured twice a week. V=(L×W²)/2 is the formula used to compute tumor volumes. On day 14, mice undergo one last gavage before being sacrificed the next day. In order to perform immunohistochemistry and Western blot, the tumors are removed.

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In Vivo Tumor Studies [2]
BALB/c or C57BL/6 mice were implanted into the flank subcutaneously with 1 × 106 CT26, 5 × 105 MC38, or 5 × 105 EMT6 tumor cells per mouse on day 0. Mice were randomized into individual treatment groups (n = 5–15 mice per group) as indicated. A separate cohort of animals (n = 5 animals per time point) was allocated for mechanistic analyses in some cases. Abemaciclib was used. Rat anti-mouse anti-PD-L1 was generated from a rat of Lou/WS1 strain immunized with recombinant mouse PD-L1-Fc protein. 178G7 was identified on the basis of its binding to PD-L1 (half maximal effective concentration [EC50] = 0.1 nM) and blocking activities against PD-L1 interactions with PD-1 and CD80 (IC50 = 1.5 nM and 2.5 nM, respectively). Tumor volume (TV) was calculated as TV (mm3) = π/6 × length × width2. Animals were sacrificed because of progressive disease if tumor burden was greater than 2,500 mm3 or growth would surpass 2,500 mm3 before the next measurement. For experiments with secondary tumor re-challenge, mice were re-challenged with 1 × 106 CT26 tumors on the opposite flank of the original tumor injection site. Secondary challenge tumor growth was followed for up to 22 days.

Absorption, Distribution and Excretion
The plasma concentration of the drug increases in a dose-proportional manner. Following a single oral dose administration of 200 mg abemaciclib, the mean peak plasma concentration (Cmax) of 158 ng/mL is reached after 6 hours. The median time to reach maximum plasma concentration (Tmax) ranges from 4-6 hours following an oral administration of abemaciclib over a range of 50–275 mg, but may range up to 24 hours. The absolute bioavailability of the drug is reported to be 45%.
Following a single oral dose of 150mg radiolabeled abemaciclib, approximately 81% of the total dose was recovered in feces while 3% of the dose was detected in urine. The majority of the drug is exceted as metabolites.
The geometric mean systemic volume of distribution is approximately 690.3 L (49% CV).
The geometric mean hepatic clearance (CL) of abemaciclib in patients was 26.0 L/h (51% CV).

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Metabolism / Metabolites
Abemaciclib mainly undergoes hepatic metabolism mediated by CYP3A4. The major metabolite formed is N-desethylabemaciclib (M2), while other metabolites hydroxyabemaciclib (M20), hydroxy-N-desethylabemaciclib (M18), and an oxidative metabolite (M1) are also formed. M2, M18, and M20 are equipotent to abemaciclib and their AUCs accounted for 25%, 13%, and 26% of the total circulating analytes in plasma, respectively.

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Biological Half-Life
The mean plasma elimination half-life for abemaciclib in patients was 18.3 hours (72% CV).

Hepatotoxicity
In the large clinical trials, adverse events were common and led to dose reductions in up to one-half of patients and discontinuation in 9%. In preregistration clinical trials, ALT elevations occurred in 31% to 41% of abemaciclib treated subjects which were above 5 times the ULN in 3% to 5%. In one study, several recipients developed clinically apparent liver injury with jaundice and one recipient died of hepatic failure, but these outcomes were considered to be unrelated to abemaciclib therapy. Thus, there were no cases of clinically apparent liver injury that could be attributed to abemaciclib therapy during prelicensure studies. Since the approval and more widescale use of abemaciclib, there have been no published reports of its hepatotoxicity. Nevertheless, the high rate of serum enzyme elevations during therapy and the similarity of abemaciclib to ribociclib and palbociclib makes it an agent that should be suspected of causing rare instances of clinically significant liver injury.
Likelihood score: E* (unproved but suspected, rare cause of clinically apparent liver injury).

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Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
No information is available on the clinical use of abemaciclib during breastfeeding. Because abemaciclib and its metabolites are over 90% bound to plasma proteins, the amount in milk is likely to be low. However, the manufacturer recommends that breastfeeding be discontinued during abemaciclib therapy and for 3 weeks after the final dose.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

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Protein Binding
According to in vitro models using animal brain tissues, the protein binding of abemaciclib is approximately 95-98%. While abemaciclib demonstrated *in vitro* binding to serum albumin, alpha-1-acid glycoprotein and other human plasma proteins in a concentration-depedent manner, its major metabolites are also shown to bind to plasms proteins as well. The approximate bound fractions of M2, M18 and M20 are 93.4%, 96.8% and 97.8%, respectively.

[1]. Drug Des Devel Ther . 2018 Feb 16:12:321-330.

[2]. Cancer Discov . 2016 Jul;6(7):740-53.

[3]. Cell Rep . 2018 Mar 13;22(11):2978-2994.

Pharmacodynamics
In combination with fulvestrant, the progression-free survival for patients with HR-positive, HER2-negative breast cancer was 16.4 months compared to 9.3 months for patients taking a placebo with fulvestrant. As a monotherapy, 19.7% of patients taking abemaciclib achieved complete or partial shrinkage of their tumors for a median 8.6 months after treatment. Abemaciclib induces cell cycle arrest and exerts an antitumor activity in human tumor xenograft models. In patient investigations and a healthy volunteer study, abemaciclib is not shown to induce any clinically significant changes in the QTc interval.

C27H32F2N8

506.59

506.271

C, 64.01; H, 6.37; F, 7.50; N, 22.12

1231929-97-7

Abemaciclib methanesulfonate;1231930-82-7;Abemaciclib-d8;2088650-53-5

46220502

White to off-white solid powder

1.3±0.1 g/cm3

689.3±65.0 °C at 760 mmHg

370.7±34.3 °C

0.0±2.2 mmHg at 25°C

1.656

2.74

75

1

9

7

37

723

0

CC1=NC2=C(F)C=C(C3=NC(NC4=NC=C(CN5CCN(CC)CC5)C=C4)=NC=C3F)C=C2N1C(C)C

UZWDCWONPYILKI-UHFFFAOYSA-N

InChI=1S/C27H32F2N8/c1-5-35-8-10-36(11-9-35)16-19-6-7-24(30-14-19)33-27-31-15-22(29)25(34-27)20-12-21(28)26-23(13-20)37(17(2)3)18(4)32-26/h6-7,12-15,17H,5,8-11,16H2,1-4H3,(H,30,31,33,34)

N-[5-[(4-ethylpiperazin-1-yl)methyl]pyridin-2-yl]-5-fluoro-4-(7-fluoro-2-methyl-3-propan-2-ylbenzimidazol-5-yl)pyrimidin-2-amine

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 本产品在运输和储存过程中需避光。

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: 3.33 mg/mL (6.57 mM) in 0.5% HEC (这些助溶剂从左到右依次添加，逐一添加), 悬浮液；超声助溶。

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配方 2 中的溶解度: 5% DMSO+40% PEG 300+5%Tween80+ 50%ddH2O: 0.2mg/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;
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