Akt1 (IC50 = 3 nM); Akt2 (IC50 = 7 nM); Akt3 (IC50 = 7 nM); ROCK2 (IC50 = 60 nM); ROCK1 (IC50 = 470 nM); PKA (IC50 = 7 nM); P70S6K (IC50 = 6 nM); Autophagy

AZD5363 是一种有效的 Akt 抑制剂，对 Akt1、Akt2 和 Akt3 的 IC50 分别为 3 nM、8 nM 和 8 nM。 [1] AZD5363 的效力约为 0.3 至 0.8 μM，可防止细胞中 AKT 底物的磷酸化。 AZD5363 的效力小于 < 3 μM，可阻止 182 种实体瘤和血液肿瘤细胞系中 41 种的生长。[2]显着预测 AZD5363 反应性的因素包括 PIK3CA 的激活突变、肿瘤抑制因子 PTEN 的丢失或失活以及 HER2 扩增。此外，细胞系的 RAS 突变状态与其对 AZD5363 的耐药性之间存在联系。[1]

裸鼠口服 AZD5363 (100, 300 mg/kg) 会导致 U87 中血糖水平可逆性升高，2[18F]氟-2-脱氧-d-葡萄糖 (18F-FDG) 摄取呈剂量依赖性下降-MG 异种移植物，以及 BT474c 异种移植物中 PRAS40、GSK3 和 S6 磷酸化的剂量依赖性减少。长期口服 AZD5363（130、200 和 300 mg/kg）会对各种肿瘤类型（包括曲妥珠单抗耐药 HER2+ 乳腺癌模型）的异种移植物产生剂量依赖性生长抑制。此外，在乳腺癌异种移植物中，AZD5363 显着增加多西他赛、拉帕替尼和曲妥珠单抗的抗肿瘤活性。 [2]

通过 Caliper 片外孵化迁移率变化测定评估 AZD5363 和其他化合物抑制 AKT1、AKT2 和 AKT3 活性的能力。将活性重组 AKT1、AKT2 或 AKT3 与 5-FAM 标记的定制合成肽底物以及浓度不断增加的抑制剂一起孵育。最终反应包含 1 至 3 nM AKT1、AKT2 或 AKT3 酶、1.5 mM 肽底物、每个 AKT 同工型的 Km ATP、10 mM MgCl2、4 mM DTT、100 mM HEPES 和 0.015% Brij-35。反应在室温下进行一小时，然后添加含有 40 mM EDTA、5% DMSO、0.1% 涂层试剂、0.1% Brij-35 溶液和 100 mM HEPES 的缓冲液来停止反应。之后，在 Caliper LC3000 上检查板，从而实现肽底物和磷酸化产物的电泳分离，以及随后激光诱导荧光的检测和定量。

MTS 和 Sytox Green 是用于测量细胞增殖的两种方法。简而言之，将细胞铺板于 96 孔培养皿中，并在 37°C、5% CO2 下孵育整夜。随后，将细胞置于浓度范围为 30 至 0.003 μM 的 AZD5363 中 72 小时。根据制造商的说明，使用 CellTiter 水性非放射性细胞增殖测定试剂评估细胞增殖的 MTS 终点。 Sytox Green 终点，Sytox Green 核酸染料以 0.13 μM 的终浓度对细胞进行染色，并使用 Acumen Explorer 计算死细胞的数量。皂苷透化（最终浓度为 0.03%，在 TBS-EDTA 缓冲液中稀释）后，将细胞孵育过夜以确定总细胞计数。对 Sytox Green 和 MTS 终点进行给药前测量，并使用活细胞计数或 MTS 吸光度读数计算将处理的细胞生长与未处理的细胞生长相比减少一半所需的浓度。

Mice: Specific, pathogen-free, female nude mice (nu/nu: Alpk) and male SCID mice (SCID/CB17; 786-0 xenograft studies) are used. The mice are randomly assigned to control and treatment groups once the mean tumor sizes reach about 0.2 cm3. The treatment groups received RP-56976, which was dissolved in 2.6% ethanol in injectable water, once on day 1, at 15 or 5 mg/kg once a week, and Capivasertib (AZD5363), which was dissolved in a 10% DMSO 25% w/v Kleptose HPB (Roquette) buffer by oral gavage. When used in conjunction with Capivasertib (AZD5363), RP-56976 is given an hour before the oral dose. The control group received the DMSO/Kleptose buffer alone, twice daily by oral gavage. For the duration of the study, tumor volumes (as determined by caliper), animal weight, and tumor condition are noted twice a week. By using CO2 euthanasia, mice are sacrificed. Using the formula: (length×width)×√(length×width)×(π/6), the tumor volume is calculated by considering length to be the longest diameter across the tumor and width to be the corresponding perpendicular diameter. By comparing the variations in tumor volume between the control and treated groups, growth inhibition from the onset of treatment is evaluated.

Absorption, Distribution and Excretion
The capivasertib steady-state AUC is 8,069 h·ng/mL (37%) and Cmax is 1,371 ng/mL (30%). Steady-state concentrations are predicted to be attained on the 3rd and 4th dosing day of each week, starting week 2. Capivasertib plasma concentrations are approximately 0.5% to 15% of the steady-state Cmax during the off-dosing days. Capivasertib AUC and Cmax are proportional with doses over a range of 80 to 800 mg (0.2 to 2 times the approved recommended dosage). Tmax is approximately 1-2 hours. The absolute bioavailability is 29%. No clinically meaningful differences in capivasertib pharmacokinetics were observed following the administration of capivasertib with a high-fat meal (approximately 1,000 kcal; fat 60%) or a low-fat meal (approximately 400 kcal; fat 26%).
Following a single radiolabeled oral dose of 400 mg, the mean total recovery was 45% from urine and 50% from feces.
The steady-state oral volume of distribution is 1,847 L (36%).
The steady-state oral clearance of capivasertib is 50 L/h (37% CV), and renal clearance was 21% of total clearance.

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Metabolism / Metabolites
Capivasertib is primarily metabolized by CYP3A4 and UGT2B7.

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Biological Half-Life
The half-life of capivasertib is 8.3 hours.

Protein Binding
Capivasertib plasma protein binding is 22% and the plasma-to-blood ratio is 0.71.

[1]. J Med Chem. 2013 Mar 14;56(5):2059-73.

[2]. Mol Cancer Ther. 2012 Apr;11(4):873-87.

Capivasertib is an aminopiperidine that is piperidine substituted by 7H-pyrrolo[2,3-d]pyrimidin-4-yl, amino, and [(1S)-1-(4-chlorophenyl)-3-hydroxypropyl]aminocarbonyl groups at positions 1, 4, and 4, respectively. It is a pan-AKT kinase inhibitor used in combination with fulvestrant for the treatment of adult patients with hormone receptor (HR)-positive, human epidermal growth factor receptor 2 (HER2)-negative, locally advanced or metastatic breast cancer with one or more PIK3CA/AKT1/PTEN-alterations. It has a role as an antineoplastic agent and an EC 2.7.11.1 (non-specific serine/threonine protein kinase) inhibitor. It is a pyrrolopyrimidine, an aminopiperidine, a piperidinecarboxamide, a member of monochlorobenzenes, a primary alcohol and a secondary carboxamide.
Hormone receptor (HR) positive, especially estrogen receptor-positive, HER2-negative breast cancer is the most common subtype of metastatic breast cancer, resulting in more than 400,000 deaths annually. Although endocrine-based therapy is the first line of treatment, resistance eventually emerges, leaving chemotherapy the only but often ineffective treatment left. Therefore, significant research has been put into developing genetically targeted treatments. The PIK3/AKT pathway is one of the most commonly activated pathways in breast cancer, mainly through the constitutively active mutation in AKT1, loss of function mutation in PTEN, a negative regulator of the PIK3/AKT pathway, or PIK3CA mutations. Therefore, targeting the PIK3/AKT pathway presents a promising approach for the treatment of breast cancer, leading to the development of capivasertib, a pan-AKT kinase inhibitor. On November 17th, 2023, capivasertib, under the brand name TRUQAP, was approved by the FDA for the treatment of adult patients HR-positive, HER2-negative locally advanced or metastatic breast cancer with one or more alterations in PIK3CA/AKT1/PTEN gene(s) in combination with [fulvestrant]. This approval is based on favorable results obtained from the CAPItello-291 trial, where the combination of capivasertib and [fulvestrant] reduced the risk of disease progression or death by 50% versus [fulvestrant] alone.
Capivasertib is a novel pyrrolopyrimidine derivative, and an orally available inhibitor of the serine/threonine protein kinase AKT (protein kinase B) with potential antineoplastic activity. Capivasertib binds to and inhibits all AKT isoforms. Inhibition of AKT prevents the phosphorylation of AKT substrates that mediate cellular processes, such as cell division, apoptosis, and glucose and fatty acid metabolism. A wide range of solid and hematological malignancies show dysregulated PI3K/AKT/mTOR signaling due to mutations in multiple signaling components. By targeting AKT, the key node in the PIK3/AKT signaling network, this agent may be used as monotherapy or combination therapy for a variety of human cancers.

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Drug Indication
Capivasertib, in combination with fulvestrant, is indicated for the treatment of adult patients with hormone receptor (HR)-positive, human epidermal growth factor receptor 2 (HER2)-negative, locally advanced or metastatic breast cancer with one or more PIK3CA/AKT1/PTEN-alteration as detected by an FDA-approved test following progression on at least one endocrine-based regimen in the metastatic setting or recurrence on or within 12 months of completing adjuvant therapy.
Treatment of breast cancer , Treatment of prostate cancer

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Mechanism of Action
Capivasertib is an inhibitor of all 3 isoforms of serine/threonine kinase AKT (AKT1, AKT2, and AKT3) and inhibits phosphorylation of downstream AKT substrates. AKT activation in tumors is a result of activation of upstream signaling pathways, mutations in AKT1, loss of phosphatase and tensin homolog (PTEN) function, and mutations in the catalytic subunit alpha of phosphatidylinositol 3-kinase (PIK3CA).

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Pharmacodynamics
In vitro, capivasertib reduced the growth of breast cancer cell lines including those with relevant PIK3CA or AKT1 mutations or PTEN alteration. In vivo, capivasertib alone and in combination with fulvestrant inhibited tumor growth of mouse xenograft models including estrogen receptor-positive breast cancer models with alterations in PIK3CA, AKT1, and PTEN. The exposure-response relationship and time course of pharmacodynamic response for the effectiveness of capivasertib has not been fully characterized. Exposure-response relationships were observed for diarrhea (CTCAE Grade 2 to 4), rash (CTCAE Grade 2 to 4), and hyperglycemia (CTCAE Grades 3 or 4) at doses of 80 to 800 mg (0.2 to 2 times the approved recommended dosage). At the recommended capivasertib dose, a mean increase in the QTc interval > 20 ms was not observed.

C21H25CLN6O2

428.9152

428.172

C, 58.81; H, 5.87; Cl, 8.27; N, 19.59; O, 7.46

1143532-39-1

(R)-Capivasertib;1143532-51-7

25227436

white solid powder

1.4±0.1 g/cm3

1.670

1.04

123.65

4

6

6

30

580

1

ClC1C([H])=C([H])C(=C([H])C=1[H])[C@]([H])(C([H])([H])C([H])([H])O[H])N([H])C(C1(C([H])([H])C([H])([H])N(C2C3C([H])=C([H])N([H])C=3N=C([H])N=2)C([H])([H])C1([H])[H])N([H])[H])=O

JDUBGYFRJFOXQC-KRWDZBQOSA-N

InChI=1S/C21H25ClN6O2/c22-15-3-1-14(2-4-15)17(6-12-29)27-20(30)21(23)7-10-28(11-8-21)19-16-5-9-24-18(16)25-13-26-19/h1-5,9,13,17,29H,6-8,10-12,23H2,(H,27,30)(H,24,25,26)/t17-/m0/s1

4-amino-N-[(1S)-1-(4-chlorophenyl)-3-hydroxypropyl]-1-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide

Capivasertib; AZD-5363; AZD5363; AZD 5363; Truqap

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: 86~125 mg/mL (200.5~291.4 mM)
Ethanol: ~2 mg/mL (~4.0 mM)

配方 1 中的溶解度: ≥ 2.08 mg/mL (4.85 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 (4.85 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 (4.85 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。