Retaspimycin Hydrochloride is a potent inhibitor of Hsp90 with EC₅₀s of 119 nM for both Hsp90 and Grp9.

Retaspimycin (IPI-504) is a novel and highly soluble analog of 17AAG, an inhibitor of Hsp90. Retaspimycin can abrogate both the unfolded protein response element (UPRE) and ERSE-driven luciferase activity in non-treated U266 and MM.1s cells as well as in Tunicamycin (Tm)-treated cells. The IC₅₀s for the inhibition of reporter gene activity by Retaspimycin are 196±56 nM in U266 and 472±177 nM in MM.1s for UPRE-luc activity and 213±140 nM for the ERSE-driven activity in MM.1s cells. Retaspimycin treatment leads to a dose-dependent decrease of p50ATF6 with EC₅₀ of 237 nM, consistent with the reporter-gene assay. The level of sXBP1 is decreased in the presence of Retaspimycin with an apparent EC₅₀ between 300 nM and 1 μM^[1]. Incubation of Retaspimycin (IPI-504) potently suppresses both Akt and MAPKs phosphorylation in both sensitive and Trastuzumab-resistant cells. Total levels of Akt decreased in all 4 cell lines (BT474, SKBR-3, HCC1569, and HCC1569) in a dose-dependent manner. However, levels of total MAPKs are not significantly altered with Retaspimycin treatment^[2].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

Retaspimycin (IPI-504) and Trastuzumab independently induce tumor regression of Trastuzumab-sensitive BT474 cell-derived xenografts. Xenografts derived from BT474R cells continue to grow in the presence of Trastuzumab but are still sensitive to Retaspimycin. When used in combination, Retaspimycin and Trastuzumab add only marginal benefits to Retaspimycin monotherapy. Retaspimycin (100 mg/kg) as a single agent is more efficacious than Trastuzumab in inhibiting tumor growth in HCC1569 xenografts. The combination is not significantly superior to Retaspimycin used as a single agent^[2].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

624.17

C₃₁H₄₆ClN₃O₈

857402-63-2

瑞他霉素盐酸盐

Room temperature in continental US; may vary elsewhere.

-20°C, sealed storage, away from moisture

*In solvent : -80°C, 6 months; -20°C, 1 month (sealed storage, away from moisture)

1M HCl : 100 mg/mL (160.21 mM; ultrasonic and adjust pH to 1 with 1M HCl)

DMSO : 60 mg/mL (96.13 mM; Need ultrasonic)

H₂O : < 0.1 mg/mL (ultrasonic) (insoluble)

请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。
储备液的保存方式和期限：-80°C, 6 months; -20°C, 1 month (sealed storage, away from moisture)。-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。

请根据您的实验动物和给药方式选择适当的溶解方案。以下溶解方案都请先按照 In Vitro 方式配制澄清的储备液，再依次添加助溶剂：

——为保证实验结果的可靠性，澄清的储备液可以根据储存条件，适当保存；体内实验的工作液，建议您现用现配，当天使用； 以下溶剂前显示的百
分比是指该溶剂在您配制终溶液中的体积占比；如在配制过程中出现沉淀、析出现象，可以通过加热和/或超声的方式助溶

• 1.

  请依序添加每种溶剂： 10% DMSO    40% PEG300    5% Tween-80    45% saline

  Solubility: ≥ 3 mg/mL (4.81 mM); Clear solution

  此方案可获得 ≥ 3 mg/mL (4.81 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 30.0 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀；向上述体系中加入50 μL Tween-80，混合均匀；然后继续加入 450 μL生理盐水定容至 1 mL。

  将 0.9 g 氯化钠，完全溶解于 100 mL ddH₂O 中，得到澄清透明的生理盐水溶液

• 2.

  请依序添加每种溶剂： 10% DMSO    90% (20% SBE-β-CD in saline)

  Solubility: 3 mg/mL (4.81 mM); Suspended solution; Need ultrasonic

  此方案可获得 3 mg/mL (4.81 mM) 的均匀悬浊液，悬浊液可用于口服和腹腔注射。

  以 1 mL 工作液为例，取 100 μL 30.0 mg/mL 的澄清 DMSO 储备液加到 900 μL 20% 的 SBE-β-CD 生理盐水水溶液中，混合均匀。

  将 2 g 磺丁基醚 β-环糊精加入 5 mL 生理盐水中，再用生理盐水定容至 10 mL，完全溶解，澄清透明

• [1]. Patterson J, et al. IPI-504, a novel and soluble HSP-90 inhibitor, blocks the unfolded protein response in multiple myeloma cells. Cancer Chemother Pharmacol. 2008 May;61(6):923-32.

  [2]. Scaltriti M, et al. Antitumor Activity of the Hsp90 Inhibitor IPI-504 in HER2-Positive Trastuzumab-Resistant Breast Cancer. Mol Cancer Ther. 2011 May;10(5):817-24.

  [3]. Sydor JR, et al. Development of 17-allylamino-17-demethoxygeldanamycin hydroquinone hydrochloride (IPI-504), an anti-cancer agent directed against Hsp90. Proc Natl Acad Sci U S A. 2006 Nov 14;103(46):17408-13. Epub 2006 Nov 7.

Hela cells are grown in Dulbecco’s modified Eagle’s medium containing 10% fetal bovine serum, 1 ug/mL streptomycin and 1 ug/mL penicillin. U266 and MM.1s are cultured in RPMI 1640 medium containing 15% fetal bovine serum, 1 mM pyruvate, 1 ug/mL streptomycin, and 1 ug/mL penicillin. All the cell lines are maintained at 37°C in a humidified 5% CO₂ atmosphere. Viability studies are performed using the vital mitochondrial function stain Alamar Blue. After cells are incubated in 96-well plates (200 μL) ± Retaspimycin, 20 μL of Alamar Blue is added and incubated for 4-6 h at 37°C. The Alamar Blue reduction is monitored using an Envision plate reader at λ_EM=544 nm and λ_EM=590 nm. The ratios obtained from drug-treated cells versus vehicle treated cells are quantified and plotted against drug concentration to give EC₅₀ values. Caspase-3 and 7 activities are detected using the Caspase Glow kit^[1].

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

Mice^[2]
For all the experiments, 2×10⁷ cells are injected into the right flanks of 10 mice for each experimental condition. Established tumors are treated with Trastuzumab, Retaspimycin, or the combination as following: Trastuzumab (10 mg/kg in sterile PBS) or sterile PBS (control) is given intraperitoneally twice weekly. Retaspimyci (100 mg/kg) is administered intraperitoneally thrice weekly. Retaspimyci, Trastuzumab, and the combination treatments are tolerable. No significant toxicity is noticed among the treatment arms. Tumor growth is measured with digital calipers as indicated and tumor volume is determined using the formula: (length×width²)×(π/6). At the end of the experiments, the animals are anesthetized with 1.5% isofluorane-air mixture and killed by cervical dislocation. Results are depicted as means of tumor volume±SE.

上海金畔生物科技有限公司 has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Patterson J, et al. IPI-504, a novel and soluble HSP-90 inhibitor, blocks the unfolded protein response in multiple myeloma cells. Cancer Chemother Pharmacol. 2008 May;61(6):923-32.

  [2]. Scaltriti M, et al. Antitumor Activity of the Hsp90 Inhibitor IPI-504 in HER2-Positive Trastuzumab-Resistant Breast Cancer. Mol Cancer Ther. 2011 May;10(5):817-24.

  [3]. Sydor JR, et al. Development of 17-allylamino-17-demethoxygeldanamycin hydroquinone hydrochloride (IPI-504), an anti-cancer agent directed against Hsp90. Proc Natl Acad Sci U S A. 2006 Nov 14;103(46):17408-13. Epub 2006 Nov 7.