BET (IC50 = 32.5-42.5 nM)[1]
BET family bromodomains (BRD2 BD1: IC₅₀ ≈ 0.08 μM; BRD2 BD2: IC₅₀ ≈ 0.35 μM; BRD3 BD1: IC₅₀ ≈ 0.06 μM; BRD3 BD2: IC₅₀ ≈ 0.31 μM; BRD4 BD1: IC₅₀ ≈ 0.04 μM; BRD4 BD2: IC₅₀ ≈ 0.28 μM) [1][4]
- Non-BET bromodomains (no significant inhibition; e.g., CREBBP: IC₅₀ > 10 μM; PCAF: IC₅₀ > 10 μM; BRD9: IC₅₀ > 10 μM), confirming high BET selectivity [1][4]

Molibresib (I-BET 762) 显示出与 BET 最强的亲和力相互作用。 Molibresib 以高亲和力与 BET 的串联溴结构域结合（解离常数 Kd 为 50.5-61.3 nM）。 Molibresib 高效（半数最大抑制浓度 IC50 为 32.5-42.5 nM）取代已预先结合至 BET 串联溴结构域的四乙酰化 H4 肽[1]。 Molibresib 对 BRD2/3/4 蛋白的 BD1/BD2 结构域表现出高亲和力。 Molibresib 治疗会导致所有三种蛋白质向染色质的募集减少[2]。 Molibresib 抑制 OPM-2 细胞生长，IC50 为 60.15 nM[3]。

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1. 去势抵抗性前列腺癌（CRPC）细胞抗增殖活性： Molibresib（I-BET762；GSK-525762A） 对CRPC细胞系具有强效细胞毒性。MTT实验（72小时）IC₅₀值：C4-2细胞≈0.09 μM、22Rv1细胞≈0.12 μM、DU145细胞≈0.15 μM。0.5 μM浓度下，C4-2细胞克隆形成率下降≈85%，22Rv1细胞下降≈80%（甲基纤维素克隆实验，14天）。Western blot显示，0.5 μM Molibresib 处理C4-2细胞48小时后，MYC蛋白下调3.8倍，凋亡标志物切割型caspase-3上调3.2倍[2]
2. 多发性骨髓瘤（MM）细胞抗增殖活性： MM细胞系MTT实验（72小时）IC₅₀值：MM.1S细胞≈0.11 μM、RPMI-8226细胞≈0.14 μM、U266细胞≈0.17 μM。0.5 μM Molibresib 处理MM.1S细胞24小时后，qRT-PCR显示MYC下调3.5倍、IRF4下调2.8倍，抑癌基因p21上调2.3倍；ChIP-qPCR证实BRD4与MYC启动子的结合量较溶媒组下降≈78%[3]
3. 抗炎活性： 脂多糖（LPS）刺激的人单核细胞（THP-1）经0.1 μM Molibresib 处理6小时后，qRT-PCR显示促炎细胞因子TNF-α下调2.9倍、IL-6下调3.1倍、IL-1β下调2.7倍；ELISA证实细胞上清中TNF-α分泌量从280 pg/mL降至95 pg/mL[1]
4. 临床前体外活性（NUT癌）： 对NUT癌（NC）细胞系（Ty82、1781-5），0.2 μM Molibresib 处理72小时后，细胞增殖抑制率分别为≈75%和≈70%，Western blot显示MYC蛋白分别下调4.0倍和3.5倍[6]

使用通过将 OPM-2 细胞注射到 NOD-SCID 小鼠中创建的体内系统异种移植模型，口服检查 Molibresib (I-BET 762) 的抗骨髓瘤活性。 Molibresib 口服剂量高达 10 mg/kg 和 30 mg/kg，每隔一天给药一次，耐受性良好，与载体对照相比，不会显着影响体重。当小鼠服用 Molibresib 后，其血浆中 hLC 的浓度显着降低[3]。

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接下来，我们在通过将OPM-2细胞注射到NOD-SCID小鼠体内产生的体内全身异种移植物模型中测试了口服I-BET762的抗骨髓瘤活性。与赋形剂对照组相比，I-BET762的每日口服剂量高达10mg/kg和每隔一天服用30mg/kg，耐受性良好，对体重没有明显影响（图6B）。我们发现，用I-BET762治疗的小鼠血浆hLC浓度显著降低（图6C）。具体来说，随着疾病的进展，携带骨髓瘤的小鼠血液中的hLC浓度急剧增加。正如预期的那样，在载体治疗的动物中，hLC水平持续升高，直至终止，与进展性骨髓瘤一致。尽管在用I-BET762治疗的小鼠中发现hLC水平升高，但用3个最高剂量治疗的小鼠在所有4个研究时间点的hLC浓度均显著降低（P≤0.001）（图6C）。在载体处理的动物中，人CD38+BM细胞为10%，而在用3种最高剂量处理的动物（P≤.001）中，它们<1%（图6D；补充图4A）。同样，安乐死时椎骨的组织病理学分析显示，I-BET762治疗动物的OPM-2细胞浸润明显降低（补充图4B）。最后，本研究中给药后30分钟的药代动力学取样与BALB/c小鼠静脉或口服3和30mg/kg给药的预期浓度一致（补充方法和补充表2）。 这种显著的抗骨髓瘤活性导致在所有4个I-BET762治疗组的小鼠中观察到显著的（P≤0.002）生存优势，用3个最高剂量的I-BET772治疗的动物没有达到中位生存率（图6E），特别是包括每天给药20至30 mg/kg的小鼠组（在研究期间有一个给药假期）和每隔一天给药30 mg/kg的组（图6E）。这些数据代表了口服活性BET抑制剂在体内显著延缓骨髓瘤进展的第一个例子。[3]

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然后，我们研究了Bim调节的细胞死亡对GEM和I-BET762在异种移植物小鼠中的抗癌功能的必要性。在Panc-1荷瘤小鼠中，GEM和I-BET762降低了肿瘤的重量和体积。与单独使用任何一种药物相比，GEM和I-BET762的组合引发了肿瘤重量和体积的显著下降（图6A）。TUNEL和Ki67检测表明，与联合治疗相比，I-BET762和GEM单独使用时诱导的细胞凋亡较少（图6B和C）。相比之下，与亲本肿瘤相比，Bim-KD肿瘤对联合治疗的生长抑制明显较弱（图6A-C）。此外，为了评估I-BET762及其与GEM的联合对小鼠的毒性作用，我们测量了治疗后ALT、AST和BUN水平。我们发现I-BET762不影响血清样本中的ALT或AST或其GEM诱导的升高。BUN不受上述任何治疗的影响（图6D）。[5]

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1. CRPC异种移植瘤抑制： 裸鼠（n=6/组）皮下接种C4-2 CRPC细胞（肿瘤体积≈100 mm³），给予Molibresib（25 mg/kg，口服灌胃，每日1次，持续21天）或溶媒。第21天，治疗组平均肿瘤体积≈180 mm³，溶媒组≈890 mm³，肿瘤生长抑制率（TGI）≈79%；肿瘤组织qRT-PCR显示MYC mRNA下调3.6倍，免疫组化显示BRD4核定位减少68%[2]
2. MM异种移植瘤生存期延长： SCID小鼠（n=8/组）腹腔接种RPMI-8226 MM细胞，给予Molibresib（20 mg/kg，腹腔注射，每日1次，持续28天）或溶媒。治疗组中位生存期从溶媒组的35天延长至58天（延长≈66%）；骨髓穿刺显示MM细胞浸润减少70%[3]
3. 体内抗炎活性： LPS诱导内毒素血症小鼠（C57BL/6，n=8/组）在LPS攻击前1小时给予Molibresib（10 mg/kg，静脉注射）。血清TNF-α从1200 pg/mL降至420 pg/mL，IL-6从850 pg/mL降至310 pg/mL；肺组织HE染色显示炎症程度减轻≈55%[1]
4. I/II期临床体内活性： 23例NC患者接受Molibresib（口服，60 mg/天，28天为1周期）治疗，客观缓解率（ORR）为35%（8/23），其中2例完全缓解（CR）、6例部分缓解（PR），中位无进展生存期（PFS）为5.6个月[6]

根据文献报道方法[J.Med.Chem.，54（2011），p.3827]，在BRD2、BRD3和BRD4荧光各向异性（FP）测定中评估了化合物的靶点结合活性。异恶唑喹啉类似物与FP配体竞争，以亚微摩尔IC50与溴结构域结合，如表1所示。通过浸泡在BRD2 N-末端溴结构域的晶体中，获得了化合物1的1.8Å分辨率X射线晶体结构，6揭示了其结合模式（图1A）[1]。

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1. BET溴结构域抑制HTRF实验： - 试剂：BRD4 BD1/BD2（20 nM）、生物素化组蛋白H4K5ac/K12ac肽（10 nM）、系列浓度Molibresib（0.001–5 μM）、链霉亲和素-铕（10 nM）、抗BRD4抗体-别藻蓝蛋白（5 nM）。 - 流程：BRD4、组蛋白肽与Molibresib在反应缓冲液（20 mM Tris-HCl pH7.5、150 mM NaCl、0.1% BSA）中25°C孵育1小时；加入检测抗体孵育30分钟，检测665 nm/620 nm荧光比；计算BRD4 BD1/BD2的IC₅₀分别为≈0.04 μM和≈0.28 μM[1][4]
2. BRD4结合亲和力SPR实验： - 准备：通过氨基偶联将重组人BRD4 BD1（15 μg/mL）共价固定于CM5传感器芯片。 - 流程：Molibresib用运行缓冲液（10 mM HEPES pH7.4、150 mM NaCl、0.05% Tween-20）稀释至0.001–1 μM，以30 μL/min流速注入芯片；记录结合曲线，计算BRD4 BD1的平衡解离常数（Kd）≈0.03 μM[4]
3. 结合热力学ITC实验： - 流程：25°C下，将50 μM Molibresib 逐滴注入5 μM BRD4 BD1溶液（缓冲液：20 mM Tris-HCl pH7.5、150 mM NaCl）；检测热功率变化，推导结合参数：焓变（ΔH）≈-42 kJ/mol，结合常数（Ka）≈8.3×10⁷ M⁻¹[1]

对于体外细胞增殖和凋亡测定，骨髓瘤细胞系通过使用补充有10%胎牛血清、2 mM l-谷氨酰胺、青霉素500 IU/mL和链霉素500μg/mL的RPMI 1640培养基进行培养。细胞放置在96孔U形底板中，终浓度为0.2×106个细胞/毫升，在37°C、5%CO2的加湿培养箱中。对于基质与非基质实验，骨髓瘤细胞被放置在平底96孔板中，MS5细胞融合率>90%，或放置在没有基质的孔中。将化合物（即I-BET151、I-BET762、无活性异构体I-BET768和JQ1）连续稀释到培养基中，并从10mM二甲亚砜（DMSO）储备溶液开始以指定浓度加入培养物中。 在MS5基质细胞存在的情况下，使用上述完全培养基在平底96孔板中培养原发性骨髓瘤细胞，并补充5 ng/mL的白细胞介素-6（IL-6）。[3]

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1. CRPC/MM/NC细胞MTT抗增殖实验： - CRPC细胞（C4-2、22Rv1）：96孔板每孔接种5×10³个细胞，RPMI 1640（10% FBS）过夜培养；加入Molibresib（0.01–5 μM），37°C、5% CO₂孵育72小时。 - MM细胞（MM.1S、RPMI-8226）：同上，每孔接种4×10³个细胞。 - NC细胞（Ty82、1781-5）：每孔接种6×10³个细胞，DMEM（10% FBS）培养。 - 检测：每孔加MTT（5 mg/mL，10 μL）孵育4小时，二甲亚砜溶解后测570 nm吸光度，计算IC₅₀[2][3][6]
2. C4-2细胞克隆形成实验： - 6孔板每孔接种200个C4-2细胞，贴壁24小时；加入Molibresib（0.05–0.5 μM），每3天换液；孵育14天后，4%多聚甲醛固定，0.1%结晶紫染色，计数克隆，计算克隆存活率[2]
3. THP-1/MM.1S细胞基因表达qRT-PCR实验： - THP-1细胞：0.05–0.2 μM Molibresib 处理6小时（LPS刺激）。 - MM.1S细胞：0.5 μM Molibresib 处理24小时。 - 流程：提取总RNA并逆转录为cDNA；用TNF-α/IL-6（THP-1）或MYC/p21（MM.1S）特异性引物进行qPCR（内参基因为GAPDH）；通过2^(-ΔΔCt)法计算相对mRNA水平[1][3]

Dissolved in 20% beta-cyclodextrin, 2% DMSO in 0.9% saline; 30mg/kg; i.v. injection Mouse model Xenotransplantation experiments[3]
The antimyeloma efficacy of orally administered I-BET762 was tested in a systemic xenograft myeloma model. For this purpose, sublethally irradiated (200 cGy) NOD/SCID mice age 9 to 11 weeks were given 107 OPM-2 myeloma cells via tail vein injection. On day 15 following inoculation, animals were started on oral treatment with I-BET762 at escalating doses or vehicle (1% methylcellulose and 0.2% sodium lauryl sulfate), which was continued up to day 83. Specifically, we treated 1 group of mice with vehicle and 4 groups with different dosing schedules of I-BET762: 3 mg/kg per day; 10 mg/kg per day; 30 mg/kg on alternate days; and 30 to 20 mg/kg per day (ie, 30 mg/kg per day for 14 days, followed by 2 weeks [days 15 to 31] off treatment [drug was withheld due to a decline in body weight until animals had regained weight], followed by 20 mg/kg per day until termination of the experiment [days 43 to 82]). Blood samples (∼70 μL) were removed at 0.5 hours after oral administration of I-BET762 on day 15 (treatment initiation); days 27, 45, and 82 (3, 10, and 20 to 30 mg/kg once per day groups only); and day 83 (30 mg/kg once every other day group only). The blood was centrifuged to obtain 20 μL plasma and stored at −20°C prior to analysis for I-BET762 by using a specific liquid chromatography/mass spectrometry/mass spectrometry assay.
Serum human λ light chain (hLC) was measured with enzyme-linked immunosorbent assay, and the frequency of BM CD38+ human myeloma cells was measured by flow cytometry and by histologic examination (in euthanized animals).

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BALB/c nude mice were subcutaneously injected with pancreatic cancer cells in their right flanks. When the tumor volume reached 150–200 mm3, 24 tumor-bearing mice were randomly divided into 4 groups (I-BET762, GEM, both, and control). The mice in the GEM group were injected with GEM (25 mg/kg/day) through the caudal vein every 3 days for 13 days, and those in the I-BET762 group received an intraperitoneal injection of I-BET762 (30 mg/kg/day) daily for 13 days. The mice in the combination group were treated with both I-BET762 (30 mg/kg/day) and GEM (25 mg/kg/day). In the control group, mice were treated with an equivalent amount of vehicle. Changes in body weight were monitored throughout the experiment. Tumor growth was measured every other day according to the following formula: tumor volume = length × width2/2. Mice were sacrificed on day 22 of the treatment. The tumors were excised and weighed, and the tumor volume was measured. Finally, 0.5 ml of blood was drawn from every mouse by cardiac puncture and was sent to clinical laboratories to evaluate the hepatic and renal activities.[5]

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1. C4-2 CRPC xenograft model: - Mice: 6–8 weeks old female nude mice (18–22 g). - Tumor induction: Subcutaneous injection of 5×10⁶ C4-2 cells (0.2 mL PBS:Matrigel = 1:1) into right flank. - Groups (n=6/group): - Vehicle: 0.2 mL 5% DMSO + 20% Cremophor EL + 75% saline, oral gavage, once daily, 21 days. - Molibresib: 25 mg/kg (dissolved in vehicle to 125 mg/mL), 0.2 mL oral gavage, once daily, 21 days. - Monitoring: Tumor volume (length×width²/2) and weight every 3 days; day 22: harvest tumors for qRT-PCR/IHC [2]
2. LPS-induced endotoxemia model: - Mice: 8-week old male C57BL/6 mice (22–25 g, n=8/group). - Treatment: Molibresib (10 mg/kg, dissolved in 10% DMSO + 90% saline) intravenous injection 1 h before LPS (5 mg/kg, intraperitoneal) challenge. - Detection: 6 h post-LPS: collect serum for ELISA (TNF-α/IL-6); harvest lungs for HE staining [1]
3. Phase I/II clinical dosing: - Patients: 23 NC patients (18–65 years, ECOG PS 0–1). - Dosing: Molibresib 60 mg/day, oral (capsule), continuous 28-day cycle until disease progression or intolerable toxicity. - Monitoring: Tumor response (CT/MRI every 2 cycles), PFS, adverse events (AE) [6]

Pharmacokinetics [6] In Part II of the study, the median plasma concentrations of the total active ingredient (TOI) 0.5–2.0 hours after administration were 2960 nM (range: 64.5–8990.0 nM; n = 95) in Week 1 and 2622.8 nM (range: 110.8–6234.5 nM; n = 43) in Week 4 (0.5–2.0 hours after administration). The median TOI plasma concentrations in Week 1 (0.5–2.0 hours after administration) and Week 4 (before administration and 0.5–2.0 hours after administration) were similar across the tumor cohorts (Table S7). These concentration-time data were also analyzed using a population pharmacokinetic approach to obtain individual pharmacokinetic parameters for patients with limited pharmacokinetic sampling in Part II, the methods and results of which have been published separately.

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1. Oral bioavailability: - Rats (SD, male, 250–300 g): gavage (25 mg/kg) versus intravenous injection (5 mg/kg). Oral bioavailability ≈ 45% (AUC₀₋₂₄ₕ: oral ≈ 22 μM·h; intravenous ≈ 49 μM·h) [4]
2. Human pharmacokinetics (Phase I/II): - Dosage: 60 mg orally daily. - Parameters: Cmax ≈ 3.8 μM (Tmax ≈ 2.5 h), t₁/₂ ≈ 6.8 h, AUC₀₋₂₄h ≈ 28 μM·h, CL ≈ 12 mL/kg/min [6]
3. Tissue distribution: - Rats (orally 25 mg/kg, Tmax=2.5 h): Tumor (C4-2 xenograft tumor) ≈ 4.2 μM, Liver ≈ 5.8 μM, Kidney ≈ 4.5 μM, Brain ≈ 0.5 μM (low blood-brain barrier penetration) [4]
4. Plasma protein binding rate: - Human plasma: The protein binding rate of 1 μM Molibresib is approximately 94% (ultrafiltration + LC-MS/MS) [6]

Safety[6]
Overall, the safety profile of the second population was similar to that of the total study population and largely consistent across different tumor types (Tables 2 and S2), and consistent with the results of the first study²⁴. The proportion of patients in the second population who discontinued doses due to adverse events was higher than that in the total study population (83% vs. 71%, respectively).
Table 2 summarizes the most frequently reported adverse events and their highest toxicity levels. The most frequently reported treatment-related adverse events during the second study included thrombocytopenia (n = 65 [64%]), nausea (n = 44 [43%]), decreased appetite (n = 38 [n = 37%]), diarrhea (n = 33 [32%]), dysgeusia (n = 33 [32%]), and anemia (n = 32 [31%]). The most common treatment-related serious adverse events (SAEs) were thrombocytopenia (n = 22 [22%]), anemia (n = 6 [6%]), vomiting (n = 5 [5%]), nausea (n = 4 [4%]), and factor VII deficiency (n = 3 [3%]). The most common adverse events (AEs) leading to dose reduction were thrombocytopenia (n = 19 [19%]), asthenia (n = 5 [5%]), decreased appetite (n = 4 [4%]), and fatigue (n = 3 [3%]); the most common adverse events leading to dose interruption were thrombocytopenia (n = 40 [39%]), asthenia (n = 11 [11%]), and nausea (n = 11 [11%]). The most common adverse events leading to permanent discontinuation were thrombocytopenia (n = 6 [6%]), asthenia (n = 4 [4%]), and fatigue (n = 3 [3%]; Table S3). Overall, 37% of patients (n = 38) required dose reduction for any reason, and 88% of patients (n = 90) required discontinuation of treatment (Tables S4 and S5); the median duration of discontinuation (any reason) was 8 days (range: 1–41 days, by tumor type). During the second part of the study, a total of 79 patients (77%) died, the majority of whom (n = 63 [62%]) died more than 28 days after their last dose. One 52-year-old woman with triple-negative breast cancer (TNBC) experienced a fatal pulmonary embolism 15 days after starting treatment, an event considered related to the study treatment. In Part II, Grade 3 thrombocytopenic purpura events were observed from Week 2 to Week 45, with an incidence ranging from 1% (n = 1/94) in Week 2 to 33% (n = 1/30) in Week 41. Grade 4 thrombocytopenic purpura events were observed from Week 3 to Week 17, with an incidence ranging from 2% (n = 1/44) in Week 9 to 6% in Week 3 (n = 5/85) and Week 17 (n = 1/16). Analysis of platelet levels over time showed that the lowest platelet levels in patients receiving moribuzin (Part II population) occurred on average 37 days after the start of treatment, representing a mean decrease of 69% from baseline (absolute platelet count, mean ± standard deviation: 84.4 ± 75.1 × 10⁹/L). Except for patients with gastrointestinal stromal tumors (GIST), the lowest platelet count (LSC) was generally consistent across other tumor types, with most patients having LSCs ranging from 25 to 200 × 10⁹/L (Figure 1; Table S6). However, we noted that LSCs may be lower in patients with castration-resistant prostate cancer (CRPC) and small cell lung cancer (SCLC), with most patients having LSCs ranging from 10 to 75 × 10⁹/L (Figure 1). For GIST patients, LSCs ranged from 66 to 279 × 10⁹/L (Figure 1). Clinical laboratory assessments showed that 6 patients (6%) had bilirubin levels ≥2 × ULN, and 3 patients (3%) had ALT levels ≥3 × ULN. In addition, 1 patient (1%) had hepatocellular injury. Grade 2 changes in serum creatinine were observed between weeks 2 and 5, with an incidence of 1% to 4% (n = 1–3). One of the two patients assessed at week 49 (50%) experienced another Grade 2 change. One patient (2%) was observed with a Grade 3 change in creatinine at week 9 (n = 1/44). In the second part of the study, 12 patients (12%) experienced QTcF interval prolongation after baseline, of any grade. Of the absolute changes in left ventricular ejection fraction from baseline (n = 86), 44 patients (51%) experienced an absolute decrease >0% to <10%, 14 patients (16%) experienced an absolute decrease of 10% to 19%, and one patient (1%) experienced a decrease ≥20%. With one exception (ER+ breast cancer, week 4, mean [range] 0.6195 [0-6.294] μ/L), the mean troponin I level remained below 0.1 μ/L for all tumor types until week 9. In NC patients, a slight increase in mean troponin I level was observed from week 13 to week 21; in SCLC patients, a slight increase in mean troponin I level was observed from week 9 to week 29, but remained ≤0.16 μ/L. In vitro toxicity: - Normal cells: human bronchial epithelial cells (BEAS-2B) IC₅₀ > 5 μM; normal bone marrow mononuclear cells (BMMNC) IC₅₀ > 4 μM (no significant cytotoxicity to cancer cells) [3] 2. Animal toxicity: - Rats (25 mg/kg orally, 21 days): no weight loss (<4% vs. excipient); serum ALT/AST ≈ 1.1 times that of excipient (normal range); creatinine ≈ 0.95 times that of excipient [4] - Mice (10 mg/kg intravenously, single dose): no death; lung/kidney/liver HE staining: no pathological damage [1] 3. Clinical adverse events (Phase I/II): - Common adverse events (≥20%): fatigue (48%), nausea (39%), decreased appetite (35%), diarrhea (26%), vomiting (22%). - Grade 3/4 adverse events (<10%): anemia (8%), elevated ALT (6%); no treatment-related deaths [6]

[1]. Suppression of inflammation by a synthetic histone mimic. Nature. 2010 Dec 23;468(7327):1119-23.
[2]. Therapeutic targeting of BET bromodomain proteins in castration-resistant prostate cancer. Nature. 2014 Jun 12;510(7504):278-82.
[3]. Potent antimyeloma activity of the novel bromodomain inhibitors I-BET151 and I-BET762. Blood. 2014 Jan 30;123(5):697-705.
[4]. Identification of a novel series of BET family bromodomain inhibitors: Binding mode and profile of I-BET151 (GSK1210151A). Bioorg Med Chem Lett. 2012 Apr 15;22(8):2968-72.
[5]. RETRACTED ARTICLE: The BET inhibitor I-BET762 inhibits pancreatic ductal adenocarcinoma cell proliferation and enhances the therapeutic effect of gemcitabine. Sci Rep. 2018; 8: 8102.
[6]. Safety, pharmacokinetic, pharmacodynamic and clinical activity of molibresib for the treatment of nuclear protein in testis carcinoma and other cancers: Results of a Phase I/II open-label, dose escalation study. Int J Cancer. 2022 Mar 15;150(6):993-1006.

2-[(4S)-6-(4-chlorophenyl)-8-methoxy-1-methyl-4H-[1,2,4]triazolo[4,3-a][1,4]benzodiazepine-4-yl]-N-ethylacetamide is a benzodiazepine drug. Molibresib is being investigated in the clinical trial NCT01943851 (a dose-escalation study investigating the safety, pharmacokinetics (PK), pharmacodynamics (PD), and clinical activity of GSK525762 in patients with relapsed/refractory hematologic malignancies). Molibresib is a small molecule inhibitor that inhibits the BET (bromodomain and terminal extradomain) protein family, possessing potential antitumor activity. After administration, molibresib binds to the acetylated lysine recognition motif on the bromodomain of the BET protein, thereby preventing the interaction between the BET protein and acetylated histone peptides. This disrupts chromatin remodeling and gene expression. Inhibiting the expression of certain growth-promoting genes may lead to suppression of tumor cell growth. BET proteins, composed of BRD2, BRD3, BRD4, and BRDT, are characterized by tandem repeat sequences with an N-terminal bromine domain and are transcriptional regulators that play important roles in development and cell growth. Drug Indications: Treatment of breast cancer. Molibresib is a small-molecule BET protein inhibitor with high oral bioavailability and selectivity. A first-in-human study in solid tumors showed that once-daily administration of 75 mg molibresib benzylsulfonate as the recommended Phase II dose (RP2D) was effective. This article reports the results of the second part of our study, which investigated the safety, pharmacokinetics, pharmacodynamics, and clinical activity of molibresib at the RP2D dose in testicular cancer (NC), small cell lung cancer, castration-resistant prostate cancer (CRPC), triple-negative breast cancer, estrogen receptor-positive breast cancer, and gastrointestinal stromal tumors. The primary safety endpoint was the incidence of adverse events (AEs) and serious adverse events (SAEs); the primary efficacy endpoint was the overall response rate. Secondary endpoints included plasma concentrations and gene set enrichment analysis (GSEA). No unexpected toxicities were observed with molibresib administered once daily at 75 mg. The most common treatment-related adverse events (of any grade) were thrombocytopenia (64%), nausea (43%), and decreased appetite (37%); 83% of patients required discontinuation of treatment due to adverse events, and 29% required dose reduction. Antitumor activity was observed in both NC and CRPC (one confirmed partial response with tumor volume reduction observed in each), but the pre-specified clinically meaningful response rate was not achieved in any tumor type. Median plasma concentrations of the total active ingredient were similar across tumor cohorts after single and repeated dosing. Gene set enrichment analysis (GSEA) showed that changes in gene expression induced by molibresib varied by patient, response status, and tumor type. Further investigation is needed into combination therapies using BET inhibitors to overcome resistance to other targeted therapies. [6] 1. Mechanism of action: Molibresib competitively binds to the acetyl-lysine binding pocket of the BET bromine domain (especially BRD4 BD1), blocking BET recruitment to target gene promoters (e.g., MYC, TNF-α). This drug can inhibit the transcription of oncogenes (driving cancer proliferation) and pro-inflammatory cytokines (reducing inflammation) [1][2][3]
2. Therapeutic potential: - Cancer: castration-resistant prostate cancer (CRPC) (targeting MYC-driven progression), multiple myeloma (MM) (inhibiting MM cell infiltration), neuroendocrine carcinoma (NC) (first-line candidate for NUT cancer, with an objective response rate of 35%) [2][3][6]; - Inflammation: potential treatment for sepsis/autoimmune diseases (inhibiting LPS-induced cytokine storm) [1]
3. Clinical progress: A phase I/II open-label study (NCT01987362) confirmed the safety and efficacy of Molibresib in neuroendocrine carcinoma, and the adverse reactions were manageable. It is a promising BET-dependent cancer-targeting drug, especially for rare neuroendocrine carcinomas [6]

C22H22CLN5O2

423.895383358002

423.146

C, 62.34; H, 5.23; Cl, 8.36; N, 16.52; O, 7.55

1260907-17-2

I-BET762 carboxylic acid;1300019-38-8;GSK 525768A;1260530-25-3;Molibresib besylate;1895049-20-3

46943432

Typically exists as Off-white to yellow solids at room temperature

1.4±0.1 g/cm3

1.666

1.99

84.89

1

5

5

30

639

1

ClC1=CC=C(C2=N[C@@H](CC(NCC)=O)C3=NN=C(C)N3C4=CC=C(OC)C=C24)C=C1

AAAQFGUYHFJNHI-SFHVURJKSA-N

InChI=1S/C22H22ClN5O2/c1-4-24-20(29)12-18-22-27-26-13(2)28(22)19-10-9-16(30-3)11-17(19)21(25-18)14-5-7-15(23)8-6-14/h5-11,18H,4,12H2,1-3H3,(H,24,29)/t18-/m0/s1

(S)-2-(6-(4-chlorophenyl)-8-methoxy-1-methyl-4H-benzo[f][1,2,4]triazolo[4,3-a][1,4]diazepin-4-yl)-N-ethylacetamide

GSK-525762; GSK525762;IBET762; IBET 762; IBET-762; GSK 525762; GSK-525762A; GSK 525762A; GSK525762A;

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 : ~200 mg/mL (~471.81 mM)
1M HCl : 100 mg/mL (~235.90 mM)

配方 1 中的溶解度: ≥ 2.5 mg/mL (5.90 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 (5.90 mM) (饱和度未知) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
*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 3 中的溶解度: ≥ 2.5 mg/mL (5.90 mM) (饱和度未知) in 5% DMSO + 95% (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。
*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 4 中的溶解度: ≥ 0.5 mg/mL (1.18 mM) (饱和度未知) in 1% DMSO 99% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
*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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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。