PARP-1 ( Ki = 1.4 nM ); PARP-2; PARP-3

体外活性：Rucaparib 是纯化全长人 PARP-1 的有效抑制剂，对 LoVo 和 SW620 细胞中的细胞 PARP 显示出更高的抑制作用。此外，Rucaparib 可检测地与其他 8 个 PARP 结构域结合，包括 PARP2、3、4、10、15、16、TNKS1 和 TNKS2。 Rucaparib 的放射增敏作用是由于下游 NF-κB 激活的抑制，并且与 SSB 修复抑制无关。 Rucaparib 可以靶向由 DNA 损伤激活的 NF-κB，并克服经典 NF-κB 抑制剂观察到的毒性，而不损害其他重要的炎症功能。在透化的 D283Med 细胞中，浓度为 1 μM 的 Rucaparib 可抑制 PARP-1 活性 97.1%。激酶测定：测量[32P]NAD+掺入对人全长重组PARP-1的抑制。使用 PhosphorImager 对掺入酸不溶性材料中的 [32P]ADP-核糖进行定量。 Ki是通过非线性回归分析计算的。细胞测定：在过表达人(h) ABCB1的MDCKII亲本细胞系中，rucaparib的顶部和基底外侧定向易位是相同的。用 ABCB1 抑制剂 zosuquidar 处理细胞导致顶端定向转运略有减少，这可能是由于基底外侧未识别的 rucaparib 摄取转运蛋白的特异性抑制，或内源性犬 ABCB1 的抑制。结果表明rucaparib是ABCB1的转运底物。

Rucaparib 无毒，但在具有 DNA 修复蛋白能力的 D384Med 异种移植物中显着增强替莫唑胺诱导的 TGD。药代动力学研究还表明，Rucaparib在脑组织中被检测到，这表明Rucaparib具有治疗颅内恶性肿瘤的潜力。 Rucaparib 显着增强拓扑替康和替莫唑胺在 NB-1691、SH-SY-5Y 和 SKNBE (2c) 细胞中的细胞毒性。 Rucaparib 增强替莫唑胺的抗肿瘤活性，并表明 NB1691 和 SHSY5Y 异种移植物中的肿瘤完全且持续消退。

测量 [32P]NAD+ 掺入诱导的人全长重组 PARP-1 抑制的量。使用 PhosphorImager，测量添加到酸不溶性材料中的 [32P]ADP-核糖的量。非线性回归分析用于计算Ki。

MTT 测定用于测量细胞增殖。在 96 孔板中，细胞以 5×10³ 细胞/ml 的密度接种在 200 μl/孔的体积中。第二天，将不同浓度的 DMSO、BKM120 或 rucaparib 添加到细胞中。四天后，每个孔接受 20 μl MTT (5 mg/ml)。在 37 °C 下额外孵育 4 小时后，测定 490 nm 处的吸光度。 CalcuSyn 软件用于分析生长抑制实验的数据，以确定药物组合的效果。接下来，计算组合指数（CI）。

Determination of Antitumor Activity In vivo[1]
Female athymic nude mice (CD1 nu/nu) used for antitumor studies were maintained and handled in isolators under specific pathogen-free conditions. We implanted SW620 colorectal tumor cells (1 × 107 cells per animal) s.c. into one flank of each mouse, treated the mice (five animals per group) when tumors were palpable (10–12 days after implantation), and monitored tumor growth using two-dimensional caliper measurements. Tumor volume was calculated using the equation a2 × b / 2, where a is the smallest measurement and b is the largest. Data are presented as median relative tumor volumes (RTV), defined as the calculated tumor volume divided by the calculated tumor volume on the initial day of treatment (day 0). Thus, on day 0, the RTV value is 1 and RTV4 is when the tumor is four times as large as its initial value.

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Single-Dose Studies. [1]
We administered a single dose of temozolomide p.o. as a suspension in saline at 200 mg/kg either alone or in combination with a single i.p. administration of PARP inhibitor administered at 0.1 [AG14447 and MS-AG14644 (equivalent to 0.078 mg/kg free AG14644 only)], 1.0, and 10 mg/kg (for the mesylate salts equivalent to 0.79 and 7.9 mg/kg free AG14451 and AG14452 and 0.78 and 7.8 free AG14531 and AG14644). Control animals were treated with either normal saline p.o. and i.p or normal saline p.o and PARP inhibitor 10 mg/kg i.p.

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Five Daily Dosing Studies. [1]
We treated animals with five daily doses of temozolomide administered p.o. as a suspension in saline at 68 mg/kg either alone or in combination with a five daily i.p. administrations of PARP inhibitor at 0.05, 0.15, and 0.5 mg/kg AG14447; 0.15 and 0.5 mg/kg MS-AG14644 (equivalent to 0.12 and 0.39 mg/kg free AG14644); 1.5, 5, and 15 mg/kg AG14361; and 5 mg/kg AG14452. Control animals were treated with either normal saline p.o. and i.p. or normal saline p.o and PARP inhibitor at the higher dose (0.5, 5, or 15 mg/kg, depending on the compound studied) i.p.

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Tissue Distribution[1]
We administered AG14361, AG14452, or AG14447 (10 mg/kg i.p.) to mice (three animals per group) bearing SW620 xenografts (∼10 × 10 mm). After 120 min, the animals were bled by cardiac puncture, under general anesthesia, the tumor was removed and snap frozen on liquid nitrogen. Plasma was removed and stored at −20°C. The concentrations of PARP inhibitor in acetonitrile-treated plasma and homogenized tumor were measured using reverse-phase high-pressure liquid chromatography (isocratic mobile phase: 40% acetonitrile in 0.1% ammonium formate, Hypersil BDS 3 μm 4.6 × 250 mm column, Waters Alliance 2690 high-pressure liquid chromatography; Waters, Elstree, Herts, United Kingdom) by the method of addition.

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Dissolved in saline; 1 mg/kg; One or four daily by i.p.

CD-1 nude mice bearing established D283Med xenografts

Absorption, Distribution and Excretion
Rucaparib exhibits a linear pharmacokinetic profile over the dose range from 240 mg to 840 mg twice daily. The mean (coefficient of variation [CV]) steady-state rucaparib Cmax is 1940 ng/mL (54%) and AUC0-12h is 16900 h x ng/mL (54%) at the approved recommended dosage. The mean AUC accumulation ratio is 3.5 to 6.2 fold. The median Tmax at the steady state is 1.9 hours, with a range of 0 to 5.98 hours at the approved recommended dosage. The mean absolute bioavailability is 36%, with a range of 30 to 45%. A high-fat meal increased Cmax and AUC0-24h by 20% and 38%, respectively. The Tmax was delayed by 2.5 hours.
Following a single oral dose of radiolabeled rucaparib, unchanged rucaparib accounted for 64% of the radioactivity. Rucaparib accounted for 45% and 95% of radioactivity in urine and feces, respectively.
The mean (coefficient of variation) apparent volume of distribution is 2300 L (21%).
The mean (coefficient of variation) apparent total clearance at steady state is 44.2 L/h (45%).

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Metabolism / Metabolites
In vitro, rucaparib is primarily metabolized by CYP2D6 and, to a lesser extent, by CYP1A2 and CYP3A4. In addition to CYP-based oxidation, rucaparib also undergoes N-demethylation, N-methylation, and glucuronidation. In one study, seven metabolites of rucaparib were identified in plasma, urine, and feces.

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Biological Half-Life
The mean (coefficient of variation) terminal elimination half-life is 26 (39%) hours.

Hepatotoxicity
In large clinical trials of rucaparib, abnormalities in routine liver tests were common; serum ALT elevations arising in 74% with values above 5 times the upper limit of normal (ULN) in 13%. Despite the frequency of serum enzyme elevations during therapy in clinical trials, there were no reports of hepatitis with jaundice or liver failure. Subsequent to its approval and more wide scale use, there have been no published reports of clinically apparent liver injury attributed to rucaparib. Thus, rucaparib is a frequent cause of serum enzyme elevations, but has not been linked to significant hepatotoxicity.
Likelihood score: E* (unproven but suspected 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 rucaparib during breastfeeding. The manufacturer recommends that breastfeeding be discontinued during rucaparib therapy and for 2 weeks after the last 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
Rucaparib is 70% bound to human plasma proteins _in vitro_. Rucaparib preferentially distributed to red blood cells with a blood-to-plasma concentration ratio of 1.8.

[1]. Preclinical selection of a novel poly(ADP-ribose) polymerase inhibitor for clinical trial. Mol Cancer Ther, 2007, 6(3), 945-956.

[2]. Tumour cell retention of rucaparib, sustained PARP inhibition and efficacy of weekly as well as daily schedules. Br J Cancer. 2014 Apr 15;110(8):1977-84.

[3]. Inhibition of poly(ADP-ribose) polymerase-1 enhances temozolomide and topotecan activity against childhood neuroblastoma. Clin Cancer Res, 2009, 15(4), 1241-1249.

[4]. Hexose-6-phosphate dehydrogenase blockade reverses prostate cancer drug resistance in xenograft models by glucocorticoid inactivation. Sci Transl Med. 2021 May 26;13(595):eabe8226.

[5]. NF-κB mediates radio-sensitization by the PARP-1 inhibitor, AG-014699. Oncogene, 2012, 31(2), 251-264.

[6]. Rucaparib: A Review in Ovarian Cancer. Target Oncol. 2019 Apr;14(2):237-246.

Pharmacodynamics
Rucaparib is an anticancer agent that exerts cytotoxic effects against cancer cells. It works by inhibiting poly (ADP-ribose) polymerase (PARP), an enzyme that plays a role in DNA repair. Rucaparib inhibits PARP-1, PARP-2, and PARP-3. It also interacts with PARP-4, PARP-10, PARP-12, PARP-15, and PARP-16, but to a lesser extent. In mice, rucaparib accumulated and was retained in tumours, inhibiting PARP enzymes for seven days. Rucaparib decreases tumour growth in tumour cell lines with deficiencies in BRCA1/2 and other DNA repair genes. In addition to PARP inhibition, rucaparib demonstrated PARP-independent cytotoxic mechanisms in cancer cells. When co-administered with other chemotherapeutic agents, rucaparib contributed to synergistic or additive effects _in vitro_ and _in vivo_. There is evidence that rucaparib can sensitize cancer cells to chemotherapy. Rucaparib can also cause vasodilation, which may increase tumour perfusion and enhance the accumulation of cytotoxic drugs in cancer cells.

C19H18FN3O

323.37

323.143

C, 70.57; H, 5.61; F, 5.88; N, 12.99; O, 4.95

283173-50-2

1859053-21-6 (camsylate); 459868-92-9 (phosphate); 283173-50-2

9931954

Yellow solid powder

1.3±0.1 g/cm3

625.2±55.0 °C at 760 mmHg

331.9±31.5 °C

0.0±1.8 mmHg at 25°C

1.649

2.85

56.92

3

3

3

24

466

0

FC1=C([H])C2C(N([H])C([H])([H])C([H])([H])C3=C(C4C([H])=C([H])C(C([H])([H])N([H])C([H])([H])[H])=C([H])C=4[H])N([H])C(=C1[H])C3=2)=O

HMABYWSNWIZPAG-UHFFFAOYSA-N

InChI=1S/C19H18FN3O/c1-21-10-11-2-4-12(5-3-11)18-14-6-7-22-19(24)15-8-13(20)9-16(23-18)17(14)15/h2-5,8-9,21,23H,6-7,10H2,1H3,(H,22,24)

6-fluoro-2-[4-(methylaminomethyl)phenyl]-3,10-diazatricyclo[6.4.1.04,13]trideca-1,4,6,8(13)-tetraen-9-one

AG014699; PF-01367338; AG 14447; AG 014699; PF 01367338; AG-014699,PF01367338; AG-14447; AG14447; Trade name: Rubraca

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)

配方 1 中的溶解度: ≥ 2.5 mg/mL (7.73 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 (7.73 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 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 (7.73 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 储备液添加到 900 μL 玉米油中并混合均匀。

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配方 4 中的溶解度: 30% propylene glycol, 5% Tween 80, 65% D5W: 30mg/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;
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。