Vismodegib (GDC-0449)   中文名称: 维莫德吉

hedgehog ( IC50 = 3 nM ); P-gp ( IC50 = 3.0 μM ); ABCG2 ( IC50 = 1.4 μM )

体外活性：GDC-0449 靶向 Hedgehog 信号通路，阻断 Hedgehog 配体细胞表面受体 PTCH 和/或 SMO 的活性，并抑制 Hedgehog 信号传导。 GDC-0449 可防止多种 ATP 结合盒 (ABC) 转运蛋白。 GDC-0449 还阻断与 MDR 相关的 ABCG2、Pgp 和 MRP1 重要 ABC 转运蛋白。 GDC-0449 是 ABC 转运蛋白、ABCG2/BCRP 和 ABCB1/Pgp 的有效抑制剂，也是 ABCC1/MRP1 的轻度抑制剂。在 ABCG2 过表达 HEK293 细胞中，GDC-0449 增加荧光 ABCG2 底物 BODIPY-哌唑嗪的保留，并使这些细胞对米托蒽醌重新敏感。在经过工程改造以过表达 Pgp 或 MRP1 的 Madin-Darby 犬肾 II 细胞中，GDC-0449 增加了钙黄绿素-AM 的保留，并使它们对秋水仙碱重新敏感。 GDC-0449 还可使人非小细胞肺癌细胞 NCI-H460/par 和 NCI-H460/MX20 重新敏感，这些细胞因响应米托蒽醌、米托蒽醌和拓扑替康或 SN-38 而过度表达 ABCG2。 GDC-0449 预防 ABCG2 和 Pgp 的 IC50 值分别约为 1.4 μM 和 3.0 μM。 GDC-0449 改变细胞内 Ca2+ 稳态并抑制顺铂耐药肺癌细胞的细胞生长。激酶测定：Vismodegib (GDC-0449) 是一种口服活性的 hedgehog 通路抑制剂，IC50 为 3 nM。它还抑制 P-gp、ABCG2，IC50 值分别为 3.0 μM 和 1.4 μM。细胞测定：将 MDCKII 细胞以每孔 3 × 105 个细胞的密度接种到 24 孔板中，并使其贴壁。然后将培养基更换为含有不同药物的培养基（DMSO 中的 50 μM VP、50 μM 吲哚美辛或 20 μM GDC-0449 或单独的 DMSO 作为对照，添加非荧光钙黄绿素-AM 至最终浓度为 1.0 μM，并在 37 ℃下孵育。 °C 2 小时。然后用含 Ca2+、Mg2+ 的 Hanks 平衡盐溶液缓冲液洗涤细胞两次，并通过在 PBS 缓冲液中的 0.01% Triton X-100 中摇动在室温下 1 小时或在 4 °C 下过夜进行裂解。然后将裂解物转移至 96 孔板中，并使用 SpectraMax M5 多重检测读数器使用 495 nm 的激发波长和 515 nm 的发射波长对由细胞来源的钙黄绿素引起的荧光信号进行分光光度定量。所有操作均进行所有读数均表示为标准化至对照的平均 SEM。

GDC-0449 已用于治疗动物模型中的髓母细胞瘤。 GDC-0449 可防止原代胰腺异种移植物的生长，但不会非特异性抑制胰腺细胞增殖。在两种配体依赖性结直肠癌中，口服给药 GDC-0449，剂量≥25 mg/kg，可导致髓母细胞瘤 Ptch(+/-) 同种异体移植模型中的肿瘤消退；剂量高达 92 mg/kg，每日两次，可抑制肿瘤生长模型、D5123 和 1040830。对 Hh 通路活性和 PK/PD 模型的分析表明，GDC-0449 在髓母细胞瘤和 D5123 模型中以相似的 IC50 抑制 Gli1（分别为 0.165 μM 和 0.267 μM）。使用集成 PK/PD 模型将通路调节与功效联系起来，揭示了一种陡峭的关系，其中 > 50% 的 GDC-0449 活性与 > 80% 的 Hh 通路抑制相关。

Vismodegib (GDC-0449) 是一种口服活性刺猬通路抑制剂，IC50 为 3 nM。此外，它对 P-gp 和 ABCG2 抑制的 IC50 值分别为 3.0 μM 和 1.4 μM。

MDCKII 细胞以每孔 3 × 10⁵ 细胞的密度接种到 24 孔板中后粘附。之后，将培养基切换到含有不同药物的培养基（50 μM VP、50 μM 吲哚美辛或 20 μM GDC-0449 的 DMSO 溶液或单独的 DMSO 作为对照）。然后将非荧光钙黄绿素-AM 添加到混合物中，终浓度为 1.0 μM，并将混合物在 37 °C 下孵育两小时。用含有 Ca²⁺ 和 Mg²⁺ 的 Hank 平衡盐溶液缓冲液洗涤两次后，通过在 PBS 缓冲液中的 0.01% Triton X-100 中摇动来裂解细胞室温下 1 小时或 4 °C 下过夜。使用 SpectraMax M5 多重检测读数器以及 495 nm 的激发波长和 515 nm 的发射波长，然后将裂解物转移到 96 孔板中，并使用分光光度法对细胞来源的钙黄绿素产生的荧光信号进行定量。所有操作过程中都是完全黑暗的。标准化对照，所有读数均报告为平均 SEM。

Mice:
Mice bearing tumors are grouped into cohorts based on tumor volume when the tumors grow to a size of 200–350 mm³. A Ptch^+/−, p53^−/− medulloblastoma allograft is periodically dosed suboptimally to generate the vismodegib-resistant allograft, sg274. Vismodegib is taken orally as a suspension made of 0.2% tween-80 (MCT) and 0.5% methylcellulose. Digital calipers are used to calculate tumor volumes using the formula (L×W×W)/2. The percentage of the area under the fitted curve (AUC) for each dose group relative to the vehicle is used to calculate tumor growth inhibition (%TGI), which is expressed as follows: %TGI=100×1-(AUCtreatment/day)/(AUCvehicle/day).

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Rats:
Vismodegib (10 mg/kg) drug was gavaged orally for 14 days in rats to significantly decrease the SHH signaling proteins [SHH, protein patched homolog 1 (PTCH1), smoothened protein (SMO), glioma-associated oncogene homolog 1 (GLI1)], induce damage in SMG tissue, and affect salivary functional markers AQP5 and Keratin5. After that, in conjunction with vismodegib administration, PBM was performed using an 850 nm high-power light-emitting diode (LED) device treated daily for 6 days at varying total energy densities of 60, 120, and 180 J/cm2 in at least 3 rats per group. The test results were confirmed by Western blot, immunofluorescence staining, and hematoxylin and eosin staining, and the statistics were t-test or one-way analysis of variance (ANOVA) with Tukey's multiple comparisons tests.[5]

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Preclinical PK studies used in our study were previously reported (Wong et al., 2009). For the intravenous PK studies in rats, dogs, and monkeys, three male animals of each species were given a single intravenous dose of 1 mg/kg vismodegib in 30%, 80%, and 80% polyethylene glycol (PEG 400), respectively. For oral PK, three male animals for each species were given an oral vismodegib dose at 5 mg/kg (rats) or 2 mg/kg (dogs and monkeys) formulated in 0.5% methylcellulose with 0.2% Tween 80. For all studies, sequential plasma samples were collected following drug administration and vismodegib plasma concentrations were determined by liquid chromatography tandem mass spectrometry (LC/MS/MS)[6].

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Based on the results of in vitro and in vivo studies, vismodegib is not mutagenic. No evidence of carcinogenicity was found in mice and rats given vismodegib. A 26-week rat fertility study found that at doses of 100 mg/kg/day, vismodegib has no effects on male reproductive organs or fertility. In female rats, the administration of vismodegib was associated with decreased implantations, increased percent preimplantation loss, and decreased numbers of dams with viable embryos [7].

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Absorption, Distribution and Excretion
Vismodegib appears to have a nonlinear pharmacokinetic profile following daily oral dosing, and steady state is achieved within 7 days. A dose increase from 150 mg to 540 mg (1 to 3.6 times the recommended dose) does not lead to an increase in steady-state plasma concentrations. With a once-daily dose of 150 mg, the average plasma concentration of vismodegib at steady state is approximately 23 µM. The absolute bioavailability of a single dose of vismodegib is 31.8%. Absorption is saturable and is not affected by food.
Vismodegib is excreted mostly unchanged. Vismodegib and its metabolites are mainly eliminated through feces. Approximately 82% and 4.4% of the administered dose are recovered in feces and urine, respectively.
The volume of distribution of vismodegib ranges between 16.4 and 26.6 L.
The volume of distribution of vismodegib ranges from 16.4 to 26.6 L. Vismodegib plasma protein binding in patients is greater than 99%. Vismodegib binds to both human serum albumin and alpha-1-acid glycoprotein (AAG) and binding to AAG is saturable.
The single dose absolute bioavailability of vismodegib is 31.8%. Absorption is saturable as evidenced by the lack of dose proportional increase in exposure after a single dose of 270 mg or 540 mg vismodegib. Erivedge capsule may be taken without regard to meals because the systemic exposure of vismodegib at steady state is not affected by food.
Vismodegib and its metabolites are eliminated primarily by the hepatic route with 82% of the administered dose recovered in the feces and 4.4% recovered in urine.
While recent publications have suggested the pharmacokinetics (PK) of vismodegib appear to be non-linear, there has not been a report describing the mechanisms of non-linearity. This study provides evidence that two separate processes, namely, solubility-limited absorption and concentration-dependent plasma protein binding, can explain the non-linear PK of vismodegib. This study provides quantitative results which can account for the lower than expected accumulation of vismodegib with continuous daily dosing. Vismodegib has demonstrated clinical activity in patients with advanced basal cell carcinoma. The pharmacokinetics (PK) of vismodegib are non-linear. The objective of this study was to determine whether vismodegib PK change following repeated dosing by administering a tracer intravenous (iv) dose of (14) C-vismodegib with single and multiple oral doses. Healthy post menopausal female subjects (n= 6/group) received either a single or daily 150 mg vismodegib oral dose with a (14) C-labelled 10 ug iv bolus dose administered 2 hr after the single or last oral dose (day 7). Plasma samples were assayed for vismodegib by LC-MS/MS and for (14) C-vismodegib by accelerator mass spectrometry. Following a single i.v. dose, mean clearance, volume of distribution and absolute bioavailability were 43.4 mL hr(-1) , 16.4 l and 31.8%, respectively. Parallel concentration-time profiles following single oral and i.v. administration of vismodegib indicated elimination rate limited PK. Following iv administration at steady-state, mean clearance and volume of distribution were 78.5 mL hr(-1) and 26.8 L, respectively. Comparison of iv PK parameters after single and multiple oral dosing showed similar half-life, increased clearance and volume of distribution (81% and 63% higher, respectively) and decreased bioavailability (77% lower) after repeated dosing. Relative to single dose, the unbound fraction of vismodegib increased 2.4-fold with continuous daily dosing. Vismodegib exhibited a long terminal half-life after oral and iv administration, moderate absolute bioavailability and non-linear PK after repeated dosing. Results from this study suggest that the non-linear PK of vismodegib result from two separate, non-linear processes, namely solubility limited absorption and high affinity, saturable plasma protein binding.
For more Absorption, Distribution and Excretion (Complete) data for Vismodegib (7 total), please visit the HSDB record page.

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Metabolism / Metabolites
Vismodegib is mainly metabolized by CYP2C9 and CYP3A4 in the liver; however, more than 98% of total systemic vismodegib is not metabolized. Metabolic pathways of vismodegib in humans include oxidation, glucuronidation, and pyridine ring cleavage. The two most abundant oxidative metabolites recovered in feces are produced _in vitro_ by recombinant CYP2C9 and CYP3A4/5.
Greater than 98% of the total circulating drug-related components are the parent drug. Metabolic pathways of vismodegib in humans include oxidation, glucuronidation, and pyridine ring cleavage. The two most abundant oxidative metabolites recovered in feces are produced in vitro by recombinant CYP2C9 and CYP3A4/5.
2-Chloro-N-(4-chloro-3-(pyridin-2-yl)-phenyl)-4-(methylsulfonyl)-benzamide (GDC-0449, vismodegib) is a potent and selective first-in-class small-molecule inhibitor of the Hedgehog signaling pathway and is currently in clinical development. In this study, we investigated the metabolic fate and disposition of GDC-0449 in rats and dogs after a single oral administration of (14)C-GDC-0449. ... GDC-0449 underwent extensive metabolism in rats and dogs with the major metabolic pathways being oxidation of the 4-chloro-3-(pyridin-2-yl)-phenyl moiety followed by phase II glucuronidation or sulfation. Three other metabolites resulting from an uncommon pyridine ring opening were found, mainly in feces, representing 1.7 to 17.7% of the dose in total in rats and dogs. ...
... Proposed metabolites from exploratory metabolite identification in vitro (rat, dog and human liver microsomes) and in vivo (dog and rat urine) include three primary oxidative metabolites (M1-M3) and three sequential glucuronides (M4-M6). Oxidative metabolites identified in microsomes M1 and M3 were formed primarily by P4503A4/5 (M1) and P4502C9 (M3). GDC-0449 was not a potent inhibitor of P4501A2, P4502B6, P4502D6, and P4503A4/5 with IC50 estimates greater than 20 uM. K(i)'s estimated for P4502C8, P4502C9 and P4502C19 and were 6.0, 5.4 and 24 uM, respectively. An evaluation with Simcyp suggests that GDC-0449 has a low potential of inhibiting P4502C8 and P4502C9. Furthermore, GDC-0449 (15 uM) was not a potent P-glycoprotein/ABCB1 inhibitor in MDR1-MDCK cells.

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Biological Half-Life
The half-life of vismodegib after a single dose is 12 days, and after continuous daily dosing is 4 days.
The estimated elimination half-life of vismodegib is 4 days after continuous once-daily dosing and 12 days after a single dose.

Hepatotoxicity
Most clinical trials of vismodegib included few patients and rates of liver tests abnormalities were usually not reported. The product label for vismodegib includes no mention serum enzyme elevations or hepatotoxicity. However, a subsequent review of all published studies of vismodegib mentions that liver enzyme elevations occurred in 1.4% of a total of 363 patients treated. Since its approval and more general use, reports of clinically apparent liver injury linked to vismodegib have appeared. In one report, an elderly man presented with fatigue, nausea and jaundice 41 days after starting vismodegib with a cholestatic pattern of serum enzyme elevations and rapid improvement on stopping (Case 1). In addition, review of 7 years of spontaneous adverse event reporting to the FDA revealed 94 reports of hepatotoxicity during vismodegib therapy, including 20 that were considered serious and 4 that resulted in hepatic failure. Thus, clinically apparent liver injury from vismodegib occurs, but is somewhat rare.
Likelihood score: C (probable 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 vismodegib during breastfeeding. Because vismodegib is more than 99% bound to plasma proteins, the amount in milk is likely to be low. However, its half-life is 4 days and it might accumulate in the infant. The manufacturer recommends that breastfeeding be discontinued during vismodegib therapy and for 24 months 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
Vismodegib has high plasma protein binding (>99%). Vismodegib binds to plasma albumin and alpha-1-acid glycoprotein (saturable binding).

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Interactions
Drugs that alter the pH of the upper GI tract (e.g. proton pump inhibitors, H2-receptor antagonists, and antacids) may alter the solubility of vismodegib and reduce its bioavailability. However, no formal clinical study has been conducted to evaluate the effect of gastric pH altering agents on the systemic exposure of vismodegib. Increasing the dose of Erivedge when coadministered with such agents is not likely to compensate for the loss of exposure. When Erivedge is coadministered with a proton pump inhibitor, H2-receptor antagonist or antacid, systemic exposure of vismodegib may be decreased and the effect on efficacy of Erivedge is unknown.
In vitro studies indicate that vismodegib is a substrate of the efflux transporter P-glycoprotein (P-gp). When Erivedge is coadministered with drugs that inhibit P-gp (e.g. clarithromycin, erythromycin, azithromycin), systemic exposure of vismodegib and incidence of adverse events of Erivedge may be increased.
Vismodegib elimination involves multiple pathways. Vismodegib is predominantly excreted as an unchanged drug. Several minor metabolites are produced by multiple CYP enzymes. Although vismodegib is a substrate of CYP2C9 and CYP3A4 in vitro, CYP inhibition is not predicted to alter vismodegib systemic exposure since similar steady-state plasma vismodegib concentrations were observed in patients in clinical trials concomitantly treated with CYP3A4 inducers (i.e., carbamazepine, modafinil, phenobarbital) and those concomitantly treated with CYP3A4 inhibitors (i.e., erythromycin, fluconazole).
Vismodegib, a first-in-class oral hedgehog pathway inhibitor, is an effective treatment for advanced basal cell carcinoma. Based on in vitro data, a clinical drug-drug interaction (DDI) assessment of cytochrome P450 (CYP) 2C8 was necessary; vismodegib's teratogenic potential warranted a DDI study with oral contraceptives (OCs). This single-arm, open-label study included two cohorts of patients with locally advanced or metastatic solid malignancies [Cohort 1: rosiglitazone 4 mg (selective CYP2C8 probe); Cohort 2: OC (norethindrone 1 mg/ethinyl estradiol 35 ug; CYP3A4 substrate)]. On Day 1, patients received rosiglitazone or OC. On Days 2-7, patients received vismodegib 150 mg/day. On Day 8, patients received vismodegib plus rosiglitazone or OC. The effect of vismodegib on rosiglitazone and OC pharmacokinetic parameters (primary objective) was evaluated through pharmacokinetic sampling over a 24-h period (Days 1 and 8). RESULTS: The mean + or - SD vismodegib steady-state plasma concentration (Day 8, N = 51) was 20.6 + or - 9.72 uM (range 7.93-62.4 uM). Rosiglitazone AUC(0-inf) and C(max) were similar with concomitant vismodegib [=8% change in geometric mean ratios (GMRs); N = 24]. Concomitant vismodegib with OC did not affect ethinyl estradiol AUC(0-inf) and C(max) (=5% change in GMRs; N = 27); norethindrone C(max) and AUC(0-inf) GMRs were higher (12 and 23%, respectively) with concomitant vismodegib. CONCLUSIONS: This DDI study in patients with cancer demonstrated that systemic exposure of rosiglitazone (a CYP2C8 substrate) or OC (ethinyl estradiol/norethindrone) is not altered with concomitant vismodegib. Overall, there appears to be a low potential for DDIs when vismodegib is co-administered with other medications.

[1]. Trends Pharmacol Sci. 2009 Jun;30(6):303-12.

[2]. Neoplasia. 2009 Jan;11(1):96-101.

[3]. Anticancer Res. 2012 Jan;32(1):89-94.

[4]. Clin Cancer Res. 2011 Jul 15;17(14):4682-92.

[5]. Photobiomodulation Recovers the Submandibular Gland in Vismodegib-Treated Rats. https://doi.org/10.1089/photob.2023.0063.

[6]. Drug Metab Dispos. 2022 Sep;50(9):1170-1181. doi: 10.1124/dmd.122.000885.

[7]. https://go.drugbank.com/drugs/DB08828.

Therapeutic Uses
Erivedge capsule is indicated for the treatment of adults with metastatic basal cell carcinoma, or with locally advanced basal cell carcinoma that has recurred following surgery or who are not candidates for surgery, and who are not candidates for radiation. /Included in US product label/

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Drug Warnings
/BOXED WARNING/ WARNING: EMBRYO-FETAL DEATH AND SEVERE BIRTH DEFECTS ERIVEDGE (vismodegib) capsule can result in embryo-fetal death or severe birth defects. Erivedge is embryotoxic and teratogenic in animals. Teratogenic effects included severe midline defects, missing digits, and other irreversible malformations. Verify pregnancy status prior to the initiation of Erivedge. Advise male and female patients of these risks. Advise female patients of the need for contraception and advise male patients of the potential risk of Erivedge exposure through semen.
Advise patients not to donate blood or blood products while receiving Erivedge and for at least 7 months after the last dose of Erivedge.
Dysregulated hedgehog signaling is the pivotal molecular abnormality underlying basal-cell carcinomas. Vismodegib is a new orally administered hedgehog-pathway inhibitor that produces objective responses in locally advanced and metastatic basal-cell carcinomas. /The researchers/ tested the anti-basal-cell carcinoma efficacy of vismodegib in a randomized, double-blind, placebo-controlled trial in patients with the basal-cell nevus syndrome at three clinical centers from September 2009 through January 2011. The primary end point was reduction in the incidence of new basal-cell carcinomas that were eligible for surgical resection (surgically eligible) with vismodegib versus placebo after 3 months; secondary end points included reduction in the size of existing basal-cell carcinomas. In 41 patients followed for a mean of 8 months (range, 1 to 15) after enrollment, the per-patient rate of new surgically eligible basal-cell carcinomas was lower with vismodegib than with placebo (2 vs. 29 cases per group per year, P<0.001), as was the size (percent change from baseline in the sum of the longest diameter) of existing clinically significant basal-cell carcinomas (-65% vs. -11%, P=0.003). In some patients, all basal-cell carcinomas clinically regressed. No tumors progressed during treatment with vismodegib. Patients receiving vismodegib routinely had grade 1 or 2 adverse events of loss of taste, muscle cramps, hair loss, and weight loss. Overall, 54% of patients (14 of 26) receiving vismodegib discontinued drug treatment owing to adverse events. At 1 month, vismodegib use had reduced the hedgehog target-gene expression by basal-cell carcinoma by 90% (P<0.001) and diminished tumor-cell proliferation, but apoptosis was not affected. No residual basal-cell carcinoma was detectable in 83% of biopsy samples taken from sites of clinically regressed basal-cell carcinomas. Vismodegib reduces the basal-cell carcinoma tumor burden and blocks growth of new basal-cell carcinomas in patients with the basal-cell nevus syndrome. The adverse events associated with treatment led to discontinuation in over half of treated patients. Comment in The following popper user interface control may not be accessible. Tab to the next button to revert the control to an accessible version. Destroy user interface controlVismodegib in advanced basal-cell carcinoma.
FDA Pregnancy Risk Category: D /POSITIVE EVIDENCE OF RISK. Studies in humans, or investigational or post-marketing data, have demonstrated fetal risk. Nevertheless, potential benefits from the use of the drug may outweigh the potential risk. For example, the drug may be acceptable if needed in a life-threatening situation or serious disease for which safer drugs cannot be used or are ineffective./
For more Drug Warnings (Complete) data for Vismodegib (8 total), please visit the HSDB record page.

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Pharmacodynamics
Vismodegib selectively binds to and inhibits the transmembrane protein smoothened homologue (SMO) to inhibit the Hedgehog signalling pathway. Following 7 days of 150 mg once-daily dosing, the use of vismodegib was not associated with a clinically significant QT interval prolongation. Vismodegib can cause embryo-fetal death or severe birth defects, as well as severe cutaneous adverse reactions and musculoskeletal adverse reactions. In pediatric patients given vismodegib, premature fusion of the epiphyses has been reported.

C19H14CL2N2O3S

421.3

420.01

C, 54.17; H, 3.35; Cl, 16.83; N, 6.65; O, 11.39; S, 7.61

879085-55-9

24776445

Off-white to light yellow solid powder

1.4±0.1 g/cm3

561.6±50.0 °C at 760 mmHg

293.4±30.1 °C

0.0±1.5 mmHg at 25°C

1.641

2.98

84.51

1

4

4

27

625

0

O=C(C1C(Cl)=CC(S(C)(=O)=O)=CC=1)NC1C=C(C2C=CC=CN=2)C(Cl)=CC=1

BPQMGSKTAYIVFO-UHFFFAOYSA-N

InChI=1S/C19H14Cl2N2O3S/c1-27(25,26)13-6-7-14(17(21)11-13)19(24)23-12-5-8-16(20)15(10-12)18-4-2-3-9-22-18/h2-11H,1H3,(H,23,24)

2-chloro-N-(4-chloro-3-pyridin-2-ylphenyl)-4-methylsulfonylbenzamide

RG3616; GDC0449; RG 3616; GDC 0449; RG-3616; GDC-0449; trade name: Erivedge

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 (5.93 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 (5.93 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), 悬浮液； 超声和加热处理
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 (5.93 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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配方 4 中的溶解度: 2% DMSO+30% PEG 300+5% Tween 80+ddH2O: 10mg/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。