Ibrutinib (PCI-32765) is a selective, irreversible Btk inhibitor with an IC₅₀ of 0.5 nM^[1].

IC50: 0.5 nM (Btk)

Ibrutinib (PCI-32765) selectively inhibits B-cell signaling and activation. It inhibits autophosphorylation of Btk (IC₅₀=11 nM), phosphorylation of Btk’s physiological substrate PLCγ (IC₅₀=29 nM), and phosphorylation of a further downstream kinase, ERK (IC₅₀=13 nM)^[1].
Ibrutinib (PCI-32765) inhibits BCR-activated primary B cell proliferation (IC₅₀=8 nM). Following FcγR stimulation, Ibrutinib (PCI-32765) inhibits TNFα, IL-1β and IL-6 production in primary monocytes (IC₅₀=2.6, 0.5, 3.9 nM, respectively)^[3].
Ibrutinib binds C481 (Cysteine481) of BTK with an ideal IC₅₀ of 0.5 nM. Ibrutinib cannot form a covalent bond with the hydroxyl group of serine, C481S mutation increases the IC₅₀ against BTK-C481S phosphorylation from 2.2 nM to 1 μM^[4].

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.

Ibrutinib (PCI-32765) (3.125-50 mg/kg, p.o.) reduces the level of circulating autoantibodies and completely suppresses disease in mice with collagen-induced arthritis. Ibrutinib (PCI-32765) inhibits autoantibody production and the development of kidney disease in the MRL-Fas(lpr) lupus model. Ibrutinib (PCI-32765) (3.125-50 mg/kg, p.o.) reduces renal disease and autoantibody production in MRL-Fas(lpr) mice^[1]. Ibrutinib (PCI-32765) (0.1 μM) inhibits activation-induced proliferation of CLL cells, induces selective cytotoxicity in B cells compared with T cells, but alters activation induced T-cell cytokine production^[2]. Ibrutinib (PCI-32765) dose-dependently and potently reverses arthritic inflammation in a therapeutic CIA model with an ED₅₀ of 2.6 mg/kg/day. Ibrutinib (PCI-32765) also prevents clinical arthritis in CAIA models^[3].

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.

440.50

C₂₅H₂₄N₆O₂

936563-96-1

依鲁替尼；伊布鲁替尼；伊布替尼

Room temperature in continental US; may vary elsewhere.

4°C, protect from light

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light)

DMSO : 100 mg/mL (227.01 mM; Need ultrasonic)

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 (protect from light)。-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： 0.5% MC  0.5% Tween-80

  Solubility: 3.33 mg/mL (7.56 mM); Suspension solution; Need ultrasonic
• 2.

  请依序添加每种溶剂： 5% DMSO    95% (20% SBE-β-CD in saline)

  Solubility: ≥ 2.75 mg/mL (6.24 mM); Clear solution
• 3.

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

  Solubility: ≥ 2.5 mg/mL (5.68 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (5.68 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 25.0 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

• 4.

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

  Solubility: 2.5 mg/mL (5.68 mM); Suspended solution; Need ultrasonic

  此方案可获得 2.5 mg/mL (5.68 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 25.0 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
• 5.

  请依序添加每种溶剂： 10% DMSO    90% corn oil

  Solubility: ≥ 2.5 mg/mL (5.68 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (5.68 mM，饱和度未知) 的澄清溶液，此方案不适用于实验周期在半个月以上的实验。

  以 1 mL 工作液为例，取 100 μL 25.0 mg/mL 的澄清 DMSO 储备液加到 900 μL玉米油中，混合均匀。

• [1]. Honigberg LA, et al. The Bruton tyrosine kinase inhibitor PCI-32765 blocks B-cell activation and is efficacious in models of autoimmune disease and B-cell malignancy. Proc Natl Acad Sci U S A. 2010 Jul 20;107(29):13075-80.

  [2]. Herman SE, et al. Bruton tyrosine kinase represents a promising therapeutic target for treatment of chronic lymphocytic leukemia and is effectively targeted by PCI-32765. Blood. 2011 Jun 9;117(23):6287-96.

  [3]. Chang BY, et al. The Bruton tyrosine kinase inhibitor PCI-32765 ameliorates autoimmune arthritis by inhibition of multiple effector cells. Arthritis Res Ther. 2011 Jul 13;13(4):R115.

  [4]. Sun Y, et al. PROTAC-induced BTK degradation as a novel therapy for mutated BTK C481S induced ibrutinib-resistant B-cell malignancies. Cell Res. 2018 Jul;28(7):779-781.

Primary human B cells are isolated from peripheral blood mononuclear cell using human Miltenyl human B cell Isolation Kit II. In 0.2 mL RPMI plus 10% FBS, 100,000 B cells are treated with Ibrutinib (PCI-32765) (0.3 nM-10 μM) in triplicate wells or vehicle control in 0.1% DMSO final concentration for 30 minutes at 37°C, 5% CO₂, then cells are stimulated with 10 μg/mL anti-IgM F(ab’)₂, 5 μg/mL anti-CD3/CD28 as a negative control or 0.5 μg/mL PMA (Phorbal 12-myristate 13-acetate) as a positive control. B cells are stimulated for 72 hours at 37°C, 5% CO₂. Proliferation is measured with Cell Titer Glo reagent and measured on a luminometer.

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.

Male DBA1/1OlaHsd mice are injected on days 0 and 21 with Freunds’ Complete Adjuvant containing bovine type II collagen. On days 21 to 35, mice are randomized into treatment groups when the average clinical score of each animal is 1.5 (in a scale of 5). Ibrutinib (PCI-32765) treatment (1.56-12.5 mg/kg, p.o.) is initiated following enrollment and continues for 18 days. Clinical scores are given to each mouse daily for each paw. Clinical score assessment is made using the following criteria: 0=normal; 1=one hind paw or fore paw joint affected or minimal diffuse erythema and swelling; 2=two hind or fore paw joints affected or mild diffuse erythema and swelling; 3=three hind or fore paw joints affected or moderate diffuse erythema and swelling; 4=marked diffuse erythema and swelling or four digit joints affected; 5=severe diffuse erythema and severe swelling of entire paw, unable to flex digits.

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]. Honigberg LA, et al. The Bruton tyrosine kinase inhibitor PCI-32765 blocks B-cell activation and is efficacious in models of autoimmune disease and B-cell malignancy. Proc Natl Acad Sci U S A. 2010 Jul 20;107(29):13075-80.

  [2]. Herman SE, et al. Bruton tyrosine kinase represents a promising therapeutic target for treatment of chronic lymphocytic leukemia and is effectively targeted by PCI-32765. Blood. 2011 Jun 9;117(23):6287-96.

  [3]. Chang BY, et al. The Bruton tyrosine kinase inhibitor PCI-32765 ameliorates autoimmune arthritis by inhibition of multiple effector cells. Arthritis Res Ther. 2011 Jul 13;13(4):R115.

  [4]. Sun Y, et al. PROTAC-induced BTK degradation as a novel therapy for mutated BTK C481S induced ibrutinib-resistant B-cell malignancies. Cell Res. 2018 Jul;28(7):779-781.