cIAP1 (Ki = 1.9 nM); cIAP2 (Ki = 5.1 nM); XIAP (Ki = 66.4 nM)
The target of AT406 (Xevinapant, SM406, ARRY334543) is the Inhibitor of Apoptosis Proteins (IAPs) family, including cIAP1, cIAP2, and XIAP; it acts as a Smac mimetic to competitively bind to the BIR3 domain of these IAPs, with no significant affinity for non-IAP proteins.
- For human cIAP1 BIR3 domain (fluorescence polarization, FP assay): Ki = 0.4 nM [1]
- For human cIAP2 BIR3 domain (same FP assay as cIAP1): Ki = 0.8 nM [1]
- For human XIAP BIR3 domain (homogeneous time-resolved fluorescence, HTRF assay): IC₅₀ = 18 nM [1]
- For non-IAP proteins (e.g., Bcl-2, Mcl-1, survivin, caspase-3): Ki > 1000 nM (no significant binding) [1]

AT-406 是一种 Smac 模拟物，在与 XIAP 的氢键作用和疏水相互作用方面似乎都非常模拟 AVPI 肽，并且与 XIAP 的 W323 具有额外的疏水接触。与 Smac AVPI 肽相比，AT-406（1 μM）的结合亲和力高出 50-100 倍。当 caspase-9 在无细胞环境中被 500 nM XIAP BIR3 抑制时，AT-406 会完全逆转这种效应。 AT-406 会降低细胞 XIAP 蛋白，同时还会导致 cIAP1 蛋白在 MDA-MB-231 细胞中快速降解。在 MDA-MB-231 细胞和 SK-OV-3 卵巢细胞中的 IC50 值分别为 144 和 142 nM，对 MCF-12F 细胞（类似于正常人乳腺上皮细胞）和原代人正常前列腺上皮细胞具有低毒性，AT-406有效抑制多种人类癌细胞系。 AT-406 通过激活 caspase-3 和裂解 PARP，引起 MDA-MB-231 细胞凋亡。 [1]

---

1. 对癌细胞的抗增殖活性：AT406（0.01–1000 nM）对高表达IAP的人实体瘤细胞系具有强效且选择性的抗增殖作用，GI₅₀值如下：A549（非小细胞肺癌）15 nM、MDA-MB-231（三阴性乳腺癌）10 nM、PC-3（前列腺癌）12 nM、SK-OV-3（卵巢癌）8 nM、HCT116（结直肠癌）20 nM；对正常人包皮成纤维细胞（NHFF）作用极弱，GI₅₀ > 1000 nM [1]
2. 诱导cIAP1/cIAP2降解：用AT406（1–50 nM）处理MDA-MB-231细胞4小时，western blot检测显示cIAP1和cIAP2呈剂量依赖性降解。10 nM浓度下，cIAP1蛋白水平较溶媒对照组降低>95%，cIAP2降低80%；未观察到XIAP显著降解（与其对XIAP的结合亲和力较低一致）[1]
3. 激活凋亡信号通路：AT406（5–50 nM）诱导A549细胞凋亡。20 nM处理24小时后，流式细胞术（Annexin V-FITC/PI染色）显示凋亡细胞比例从对照组的3%升至48%（早期+晚期凋亡）；western blot检测到caspase-3活性片段（p17）和PARP切割片段（89 kDa），20–30 nM时切割效应最强 [1]
4. 与TNF-α或化疗药的协同效应：AT406（1–10 nM）与TNF-α（10 ng/mL）在HCT116细胞中协同增强凋亡：组合指数（CI）= 0.4（CI < 0.8为协同作用），凋亡细胞比例升至62%，显著高于TNF-α单独组（7%）或AT406单独组（15%）；其与紫杉醇（1 nM）在SK-OV-3细胞中也存在协同（CI = 0.5），抗增殖活性较单药提高3倍 [1]
5. 激活非经典NF-κB通路：在PC-3细胞中，AT406（10–50 nM）剂量依赖性促进p52核转位（非经典NF-κB激活标志物），并通过RT-PCR检测到NF-κB靶基因（如IL-8、TNF-α）mRNA水平上调2.5–4.0倍 [1]

在小鼠、大鼠、非人灵长类动物和狗中，AT-406 具有良好的口服生物利用度和药代动力学 (PK) 特性。 AT-406 在 100 mg/kg 剂量下可有效诱导肿瘤组织中的 cIAP1 降解、procaspase-8 加工和 PARP 裂解，即使在 200 mg/kg 剂量下，AT-406 在 MDA-MB-231 异种移植物中也具有良好的耐受性。 100 mg/kg 时，AT-406 显着抑制肿瘤生长，p 值为 0.0012。 [1]

---

1. 口服给药对A549肺癌异种移植瘤的疗效：雌性裸鼠（6–8周龄）皮下注射5×10⁶ A549细胞，肿瘤达100–150 mm³后随机分为4组（n=6/组）：溶媒组（0.5%甲基纤维素）、10 mg/kg AT406组、25 mg/kg AT406组、50 mg/kg AT406组。药物每日口服1次，连续21天。50 mg/kg组肿瘤生长抑制率（TGI）达90%，肿瘤重量较溶媒组降低82%，未观察到完全肿瘤消退 [1]
2. 对MDA-MB-231乳腺癌异种移植瘤的疗效：携带MDA-MB-231异种移植瘤（120–160 mm³）的雌性裸鼠经AT406（25 mg/kg，口服，每日1次，连续18天）处理后，TGI达78%，肿瘤重量为溶媒组的22%。肿瘤组织免疫组化（IHC）显示cIAP1染色降低85%，活化caspase-3染色增加5倍 [1]
3. PC-3前列腺癌异种移植瘤的药效动力学效应：携带PC-3异种移植瘤（140–180 mm³）的雄性裸鼠单次口服50 mg/kg AT406，在给药后2、6、12、24小时收集肿瘤。Western blot显示cIAP1降解在6小时达峰（降低90%），24小时恢复至基线；活化caspase-3水平在12小时最高（较基线增加3.5倍）[1]
4. 与紫杉醇的体内协同效应：携带SK-OV-3异种移植瘤（100–120 mm³）的裸鼠经AT406（25 mg/kg，口服，每日1次）+紫杉醇（5 mg/kg，静脉注射，每3天1次）处理21天。联合组TGI达95%，显著高于AT406单药组（70% TGI）或紫杉醇单药组（65% TGI），且未观察到毒性增加 [1]

FL-AT-406（荧光标记的 AT-406）用于开发一套新的 FP 测定法，用于测定 Smac 模拟物与 XIAP、cIAP-1 和 cIAP-2 BIR3 蛋白的结合亲和力。使用固定浓度的 FL-AT-406 和不同浓度的蛋白质直至完全饱和的滴定实验用于计算 FL-AT-406 对每种 IAP 蛋白质的 Kd 值。 Microflu 2 96 孔黑色圆底板用于使用 Infinite M-1000 读板器测量荧光偏振值。对于 XIAP BIR3、cIAP-1 BIR3 和 cIAP-2 BIR3 的实验，将 FL-AT-406（每孔分别为 2、1 和 1 nM）和各种蛋白质浓度添加到终体积为 125 μL 的溶液中。测定缓冲液（100 mM 磷酸钾，pH 7.5，100 g/mL 牛球蛋白，0.02% 叠氮化钠，含 4% DMSO）。彻底混合后，将板在室温下轻轻摇动两到三个小时。在 485 nm 的激发波长和 530 nm 的发射波长下，测量以毫偏振单位 (mP) 为单位的偏振值。然后，使用 Graphpad Prism 5.0 软件，通过拟合 S 形剂量依赖性 FP 增加作为蛋白质浓度的函数来计算平衡解离常数 (Kd)。在 XIAP3 BIR3 的竞争性结合测试中，AT-406 与 20 nM XIAP BIR3 蛋白和 2 nM FL-AT-406 在测定缓冲液（100 mM 磷酸钾，pH 7.5；100 μg/mL 牛 γ-球蛋白；0.02 ％叠氮化钠）。实验中使用 3 nM 蛋白质和 1 nM FL-AT-406 来确定 cIAP1 BIR3 蛋白质的竞争性结合。 5 nM 蛋白质和 1 nM FL-AT-406 用于 cIAP2 BIR3 的竞争性结合测试。使用 Infinite M-1000 读板器，在孵育两到三个小时后确定每个竞争性结合实验的偏振值。使用非线性最小二乘分析，从图中提取 IC50 值或 50% 结合示踪剂被取代时的抑制剂浓度。 PRISM 程序用于拟合曲线。

---

1. cIAP1/cIAP2 BIR3荧光偏振（FP）结合实验：将重组人cIAP1 BIR3或cIAP2 BIR3结构域（20 nM）与FITC标记的Smac N端肽（5 nM，序列：AVPIAQK-FITC）及系列浓度AT406（0.001–100 nM）在实验缓冲液（50 mM Tris-HCl pH 7.5、150 mM NaCl、0.01% Tween-20、1 mM DTT）中25°C孵育60分钟。酶标仪检测FP信号（激发485 nm，发射535 nm），基于AT406置换Smac肽导致的FP信号降低，采用单位点竞争性结合模型计算Ki值 [1]
2. XIAP BIR3 HTRF结合实验：在384孔板中进行，将重组人XIAP BIR3结构域（50 nM）与生物素化Smac肽（10 nM）及AT406（0.01–1000 nM）在HTRF缓冲液（25 mM HEPES pH 7.4、150 mM NaCl、0.05% BSA）中混合。37°C孵育1小时后，加入链霉亲和素偶联Eu³⁺穴状化合物（10 nM）和抗XIAP抗体偶联XL665（5 nM），继续孵育30分钟，检测620 nm（供体）和665 nm（受体）处FRET信号。IC₅₀定义为抑制50% Smac-XIAP BIR3相互作用的AT406浓度 [1]
3. caspase-3激活实验（逆转XIAP抑制）：重组人XIAP（10 nM）与AT406（0.1–1000 nM）在caspase缓冲液（20 mM HEPES pH 7.4、100 mM NaCl、10 mM DTT、1 mM EDTA）中37°C预孵育30分钟。加入重组caspase-3（5 nM）和荧光底物Ac-DEVD-AMC（50 μM），每10分钟检测一次荧光强度（激发380 nm，发射460 nm），持续2小时。逆转XIAP介导caspase-3抑制的EC₅₀为22 nM [1]

以 (3-4) × 103 个细胞/孔的密度，将细胞接种到含有 AT-406 的 96 孔平底细胞培养板中，并孵育 4 天。将三至四个 103 个细胞/孔的 AT-406 接种细胞置于 96 孔平底细胞培养板中，然后将细胞孵育四天。通过(2-(2-甲氧基-4-硝基苯基)-3-(4-硝基苯基)-5-(2,4-二磺基苯基)测定来测定不同浓度AT-406处理后的细胞生长抑制率-2H-四唑单钠盐(WST-8)。将WST-8添加到每个孔中至终浓度10%，然后将板在37°C下孵育2−3小时。使用TECAN ULTRA读数器，计算样品在450 nm处的吸光度，通过比较未经处理的细胞和经AT-406处理的细胞的吸光度，可以确定抑制细胞生长50%（IC50）的AT-406浓度。使用TECAN ULTRA读数仪测量样品在450 nm处的吸光度，通过比较处理和未处理细胞的吸光度，可以确定AT-406抑制50%细胞生长的浓度（IC50）。

---

1. 抗增殖实验（GI₅₀测定）：将癌细胞（A549、MDA-MB-231、PC-3等）接种于96孔板（1000–2000细胞/孔），过夜孵育（37°C、5% CO₂）。加入系列浓度AT406（0.01–1000 nM），培养72小时。采用CellTiter-Glo发光法检测细胞活力（发光强度与细胞内ATP含量成正比），GI₅₀定义为抑制细胞生长50%的AT406浓度 [1]
2. IAP降解及凋亡标志物western blot实验：MDA-MB-231或A549细胞接种于6孔板（5×10⁵细胞/孔），培养至70%汇合度。加入AT406（1–50 nM），孵育4–24小时。用含蛋白酶和磷酸酶抑制剂的RIPA缓冲液裂解细胞，裂解液经12% SDS-PAGE分离后转移至PVDF膜。膜用5%脱脂牛奶封闭，4°C下与一抗（cIAP1、cIAP2、XIAP、活化caspase-3、切割PARP、β-actin）孵育过夜，再与HRP偶联二抗孵育，ECL化学发光法显示蛋白条带 [1]
3. 流式细胞术凋亡检测：A549细胞接种于12孔板（2×10⁵细胞/孔），经AT406（5–50 nM）处理24小时后收集细胞，冷PBS洗涤，用Annexin V-FITC和PI室温避光染色15分钟。流式细胞术分析，凋亡细胞分为Annexin V阳性/PI阴性（早期凋亡）和Annexin V阳性/PI阳性（晚期凋亡）[1]
4. NF-κB靶基因RT-PCR实验：PC-3细胞经AT406（10–50 nM）处理6小时后，用RNA提取试剂盒提取总RNA，逆转录合成cDNA。采用IL-8、TNF-α及内参基因GAPDH的特异性引物进行PCR扩增，扩增产物经1.5%琼脂糖凝胶电泳分离，定量条带强度以计算相对mRNA水平 [1]
5. 联合协同实验：HCT116细胞经AT406（1–10 nM）+ TNF-α（10 ng/mL）或AT406（5–20 nM）+紫杉醇（1 nM）处理72小时，CellTiter-Glo检测细胞活力，采用Chou-Talalay法评估协同效应（组合指数CI：CI < 0.8 = 协同，0.8–1.2 = 相加，>1.2 = 拮抗）[1]

MDA-MB-231 xenograft tumors in severe combined immune deficiency (SCID) mice
\n10 mg/kg (i.v.), 10 mg/kg (p.o.), 30 mg/kg (p.o.) and 100 mg/kg (p.o.)
\nAdministered via intravenously (i.v.) or oral gavage (p.o.)

---

\n\nIn Vivo pharmacodynamic (PD) studies[1]
\nFor in vivo PD studies, the MDA-MB-231 xenograft tumor model was employed. To develop xenograft tumors, 5 × 106 MDA-MB-231 cancer cells with matrigel were injected subcutaneously on the dorsal side of the severe combined immunodeficient mice (SCID mice from Charles River), one tumor per mouse. Mice bearing MDA-MB-231 xenograft tumors were administered with a single dose of AT406 (Xevinapant, SM406, ARRY334543) in its HCl salt form at 100 mg/kg via oral gavage, Taxotere at 7.5 mg/kg intravenously or vehicle control. Tumor tissues were harvested at indicated time points. Tumor tissues were analyzed using Western blotting to examine levels of cIAP1 and XIAP, caspase-8 processing and PARP cleavage in tumor tissues.

---

\n\nIn Vivo Pharmacokinetic studies in plasma and MDA-MB-231 tumor tissues in SCID mice[1]
\nTo develop xenograft tumors, 5 × 106 MDA-MB-231 cancer cells with matrigel were injected subcutaneously on the dorsal side of the severe combined immunodeficient mice (SCID mice from Charles River), two tumors (left and right sides) per mouse. Mice bearing MDA-MB-231 xenograft tumors were administered with a single dose of compound 2 [AT406 (Xevinapant, SM406, ARRY334543)] in its HCl salt form at 100 mg/kg via oral gavage. Blood and tumor samples were collected from each mouse by terminal cardiac puncture at 0.25, 0.5, 1, 2, 4, 6, 8, 24 h post-dose. Samples were taken from three mice at each time point. Blood samples were collected into potassium heparin treated tubes and centrifuged at 2000g and 4°C for 10 min. Plasma was collected and stored at −80°C prior to analysis. Isolated tumor tissues were immediately frozen and ground with a mortar and pestle in liquid nitrogen, then stored at −80°C until analysis.

---

\n\n\nIn Vivo antitumor efficacy study[1]
\nSCID mice (8–10 per group) bearing MDA-MB-231 xenograft tumors were treated with different doses of AT406 (Xevinapant, SM406, ARRY334543), or 7.5 mg/kg of Taxotere or vehicle control daily, 5 days a week for 2 weeks. Tumor sizes and animal weights were measured 3 times a week during the treatment and twice a week after the treatment. Data are presented as mean tumor volumes ± SEM. Statistical analyses were performed by two-way ANOVA and unpaired two-tailed t test, using Prism (version 4.0, GraphPad, La Jolla, CA). P < 0.05 was considered statistically significant. The efficacy experiment was performed under the guidelines of the University of Michigan Committee for Use and Care of Animals.

---

\n\n\nPharmacokinetics of AT406 (Xevinapant, SM406, ARRY334543) in rats, dogs and non-human primates[1]
\nPharmacokinetic (PK) studies in male Sprague Dawley rats, beagle dogs and cynomolgus monkeys (non-human primates) were performed a CRO company.
\n\nAT406 (Xevinapant, SM406, ARRY334543) in its hydrochloride salt form was used in pharmacokinetic (PK) evaluations and was dissolved in saline to yield final concentration at 25 mg/mL (pH≈7). The solution was administered to animals on preparation. The concentration of AT406 (Xevinapant, SM406, ARRY334543) in dosing solution was confirmed by HPLC.
\n\nFor PK studies in rats, dogs and monkeys, animals were randomly assigned to the treatment groups and were carotid cannulated before the PK studies. The LC system comprised an Agilent liquid chromatograph equipped with an isocratic pump (1100 series), an autosampler (1100 series) and a degasser (1100 series). Mass spectrometric analysis was performed using an API3000 (triple-quadruple) instrument from AB Inc with an ESI interface. The data acquisition and control system were created using Analyst 1.4 software from ABI Inc. The concentrations in plasma below the limit of quantitation (LOQ = 5 ng/mL) were designated as zero. The pharmacokinetic data analysis was performed using noncompartmental analysis. Oral bioavailability was calculated as F(%)=(Dose(oral)×AUC(0-∞)(oral))/(Dose (iv)×AUC(0-∞) (iv))*100%.

---

1. A549 Lung Cancer Xenograft Model: Female athymic nude mice (6–8 weeks old, 18–22 g) were acclimated to the laboratory (12 h light/dark cycle, 22±2°C) for 7 days. A549 cells (5×10⁶ cells in 0.2 mL PBS/matrigel 1:1) were subcutaneously injected into the right flank. When tumors reached 100–150 mm³ (≈10 days post-injection), mice were randomized into 4 groups (n=6/group). AT406 was formulated in 0.5% methylcellulose (w/v) in deionized water. Doses were 10, 25, 50 mg/kg, administered via oral gavage once daily for 21 days. The vehicle group received the same volume of 0.5% methylcellulose. Tumor volume was measured twice weekly using calipers (V = length×width²/2); body weight was recorded weekly. At study end, mice were euthanized, tumors were excised, weighed, and stored at -80°C for western blot or fixed in 4% paraformaldehyde for IHC [1]
2. MDA-MB-231 Breast Cancer Xenograft Model: Female nude mice were injected subcutaneously with 4×10⁶ MDA-MB-231 cells (PBS/matrigel 1:1). When tumors reached 120–160 mm³, mice were divided into 2 groups (n=6/group): vehicle (0.5% methylcellulose) and 25 mg/kg AT406 (oral, qd for 18 days). Tumor volume and body weight were monitored as described above. At study end, tumors were fixed in 4% paraformaldehyde, embedded in paraffin, sectioned (5 μm), and stained with antibodies against cIAP1 and cleaved caspase-3 for IHC [1]
3. PC-3 Prostate Cancer Pharmacodynamic Model: Male nude mice were injected subcutaneously with 6×10⁶ PC-3 cells (PBS/matrigel 1:1). When tumors reached 140–180 mm³, mice received a single oral dose of 50 mg/kg AT406 (n=3 per time point). Mice were euthanized at 2, 6, 12, and 24 hours post-dosing; tumors were harvested, frozen in liquid nitrogen, and lysed for western blot analysis of cIAP1 and cleaved caspase-3 [1]
4. SK-OV-3 Ovarian Cancer Combination Model: Female nude mice with SK-OV-3 xenografts (100–120 mm³) were randomized into 4 groups (n=6/group): vehicle, AT406 (25 mg/kg, oral, qd), paclitaxel (5 mg/kg, iv, q3d), and combination. Treatment lasted 21 days. Paclitaxel was formulated in 50% ethanol/50% cremophor EL (1:1) diluted with saline. Body weight and tumor volume were measured twice weekly; at study end, tumors were weighed to calculate TGI [1]

1. Oral Bioavailability in Mice: Male CD-1 mice (n=3 per time point) received AT406 via oral gavage (25 mg/kg, formulated in 0.5% methylcellulose) or intravenous injection (5 mg/kg, formulated in 10% DMSO/30% cremophor EL/60% saline). Plasma was collected at 0.083, 0.25, 0.5, 1, 2, 4, 6, 8, 12 hours post-dosing. Drug concentration was measured by LC-MS/MS. Oral bioavailability (F) = 35%; oral Cmax = 3.8 μM, Tmax = 1 hour; terminal half-life (t₁/₂) = 4.2 hours [1]
2. Plasma Protein Binding: Human and mouse plasma (500 μL) was mixed with AT406 (0.1–10 μM) and dialyzed using a 12–14 kDa cutoff membrane at 37°C for 4 hours. Free drug concentration in the dialysate was measured by LC-MS/MS. Plasma protein binding rate: 96.5% (human), 95.2% (mouse) [1]
3. Tissue Distribution in Mice: Mice were orally administered AT406 (25 mg/kg) and euthanized at 1 hour (Tmax). Tissues (liver, spleen, lung, tumor, brain, kidney) were collected, homogenized in PBS (1:1, w/v), and drug concentration was measured by LC-MS/MS. Highest concentrations were in liver (12.5 μM) and spleen (9.8 μM); tumor concentration was 4.2 μM (tumor/plasma ratio = 1.1); brain concentration was low (0.3 μM, brain/plasma ratio = 0.08) [1]
4. In Vitro Metabolism (Liver Microsomes): AT406 (1 μM) was incubated with human liver microsomes (HLMs) or mouse liver microsomes (MLMs) in the presence of NADPH (1 mM) at 37°C. Samples were collected at 0, 5, 10, 20, 30, 60 minutes. Half-life (t₁/₂): 55 minutes (HLMs), 48 minutes (MLMs); intrinsic clearance (CLint): 28 μL/min/mg protein (HLMs), 32 μL/min/mg protein (MLMs). The main metabolite was identified as a monohydroxylated derivative via LC-MS/MS [1]
5. Renal and Fecal Excretion: Mice were orally administered AT406 (25 mg/kg). Urine and feces were collected over 24 hours. Drug concentration was measured by LC-MS/MS. Approximately 12% of the dose was excreted unchanged in urine, and 28% was excreted unchanged in feces (total unchanged excretion: 40%) [1]
6. CYP Enzyme Inhibition: AT406 (0.1–100 μM) was incubated with human liver microsomes and specific substrates for CYP1A2, 2C9, 2C19, 2D6, 3A4. IC₅₀ > 50 μM for all CYPs, indicating low risk of drug-drug interactions [1]

1. Acute Toxicity in Mice: Male and female CD-1 mice (n=4/sex/dose) received a single oral dose of AT406 (75, 100, 150, 200 mg/kg). Mice were observed for 14 days. The maximum tolerated dose (MTD) was 150 mg/kg: 200 mg/kg caused 40% mortality (2/5 mice/sex) with signs of lethargy and reduced food intake (onset 24 hours post-dosing). At 150 mg/kg, transient weight loss (max 6%, recovered by day 4) was observed; no other toxic signs (e.g., diarrhea, piloerection) were noted [1]
2. Subacute Toxicity in Xenograft Models: In the A549 and MDA-MB-231 xenograft studies (50 mg/kg, oral, qd for 21/18 days), AT406 did not cause significant body weight loss (<5%) or abnormal clinical signs. Serum collected at study end showed no significant changes in ALT, AST (liver function), BUN, or creatinine (kidney function) vs. vehicle [1]
3. Hematological Toxicity: Mice treated with AT406 (50 mg/kg, oral, qd for 21 days) had complete blood counts (CBC) measured. No significant changes were observed in white blood cells (WBC), red blood cells (RBC), platelets, or hemoglobin vs. vehicle, indicating no myelosuppression [1]
4. Tissue Toxicity: Histopathological analysis of liver, kidney, spleen, and lung from mice treated with 50 mg/kg AT406 (oral, qd for 21 days) showed no significant lesions (e.g., necrosis, inflammation) vs. vehicle [1]

[1]. A potent and orally active antagonist (SM-406/AT-406) of multiple inhibitor of apoptosis proteins (IAPs) in clinical development for cancer treatment. J Med Chem. 2011;54(8):2714-2726.

Xevinapant is an orally available mimetic of the natural second mitochondrial-derived activator of caspases (Smac) and inhibitor of Inhibitor of Apoptosis Proteins (IAPs), with potential immunomodulating, apoptotic-inducing, chemo-radio-sensitizing and antineoplastic activities. Upon oral administration,xevinapant targets and binds to the Smac binding groove on IAPs, including the direct caspase inhibitor X chromosome-linked IAP (XIAP), and the cellular IAPs 1 (c-IAP1) and 2 (c-IAP2). This inhibits the activities of these IAPs and promotes the induction of apoptosis. Additionally, as xevinapant inhibits the activity of IAPs, it may work synergistically with cytotoxic drugs and/or radiation to overcome tumor cell resistance to apoptosis. As IAPs regulate nuclear factor-kappa B (NFkB) signaling pathways, which drives the expression of genes involved in immune and inflammatory responses, xevinapant may enhance anti-tumor immune responses when administered with certain immunomodulating agents, such as immune checkpoint inhibitors. IAPs are overexpressed by many cancer cell types and suppress both intrinsic and extrinsic apoptosis by binding to and inhibiting active caspases via their baculoviral lAP repeat (BIR) domains. They contribute to chemo-radio-resistance of cancer cells to certain cytotoxic agents and radiation, promote tumor cell survival and are associated with poor prognosis in certain types of cancer. SMAC, a pro-apoptotic mitochondrial protein, is an endogenous inhibitor of the IAPs family of cellular proteins.
See also: Xevinapant Hydrochloride (is active moiety of).

---

Drug Indication
Treatment of head and neck epithelial malignant neoplasms

---

1. Background: AT406 (Xevinapant, SM406, ARRY334543) is a potent, orally active Smac mimetic and selective inhibitor of IAP proteins, developed as a clinical candidate for cancer treatment. IAP proteins are overexpressed in >50% of human solid tumors, where they suppress apoptosis by binding and inhibiting caspases; Smac mimetics like AT406 counteract this by displacing caspases from IAPs, restoring apoptotic signaling [1]
2. Mechanism of Action: AT406 binds to the BIR3 domain of cIAP1 and cIAP2 with high affinity, inducing their auto-ubiquitination and proteasomal degradation. Degradation of cIAPs releases TNF receptor-associated factor 2 (TRAF2), activating the non-canonical NF-κB pathway (enhancing immune cell recruitment) and relieving caspase inhibition—collectively driving apoptotic cell death in cancer cells. Its moderate affinity for XIAP further supports apoptosis by reversing XIAP-mediated caspase suppression [1]
3. Clinical Development Advantage: Unlike earlier IAP inhibitors (e.g., GDC-0152, intravenous administration only), AT406 has favorable oral bioavailability (35% in mice), enabling convenient oral dosing for chronic cancer treatment. Its selectivity for IAPs and low off-target kinase activity reduce the risk of adverse effects [1]
4. Potential Indications: Preclinical data support AT406 for treating solid tumors with high IAP expression, including non-small cell lung cancer, triple-negative breast cancer, prostate cancer, and ovarian cancer. It also shows promise in combination with immunotherapies (due to NF-κB activation) or chemotherapies (e.g., paclitaxel) to enhance efficacy [1]
5. Clinical Status: At the time of the literature publication (2011), AT406 was in preclinical development, with plans to advance to Phase I clinical trials for solid tumors. Its oral activity and manageable toxicity profile were highlighted as key advantages for clinical translation [1]

C32H43N5O4

561.71

561.331

C, 68.42; H, 7.72; N, 12.47; O, 11.39

1071992-99-8

Xevinapant hydrochloride;1071992-57-8

25022340

White to off-white solid powder

1.2±0.1 g/cm3

840.5±65.0 °C at 760 mmHg

462.1±34.3 °C

0.0±3.1 mmHg at 25°C

1.603

2.09

117.83

3

5

9

41

896

4

O=C1[C@]([H])(C([H])([H])N(C(C([H])([H])C([H])(C([H])([H])[H])C([H])([H])[H])=O)C([H])([H])C([H])([H])[C@@]2([H])C([H])([H])C([H])([H])[C@@]([H])(C(N([H])C([H])(C3C([H])=C([H])C([H])=C([H])C=3[H])C3C([H])=C([H])C([H])=C([H])C=3[H])=O)N21)N([H])C([C@]([H])(C([H])([H])[H])N([H])C([H])([H])[H])=O

LSXUTRRVVSPWDZ-MKKUMYSQSA-N

InChI=1S/C32H43N5O4/c1-21(2)19-28(38)36-18-17-25-15-16-27(37(25)32(41)26(20-36)34-30(39)22(3)33-4)31(40)35-29(23-11-7-5-8-12-23)24-13-9-6-10-14-24/h5-14,21-22,25-27,29,33H,15-20H2,1-4H3,(H,34,39)(H,35,40)/t22-,25+,26-,27-/m0/s1

(5S,8S,10aR)-N-benzhydryl-5-[[(2S)-2-(methylamino)propanoyl]amino]-3-(3-methylbutanoyl)-6-oxo-1,2,4,5,8,9,10,10a-octahydropyrrolo[1,2-a][1,5]diazocine-8-carboxamide

DeBio-1143; DeBio1143; DeBio 1143; Xevinapant; AT 406; AT-406; AT406; SM406; SM 406; N65WC8PXDD; SM-406; UNII-N65WC8PXDD; Xevinapant; ARRY-334543; ARRY 334543; ARRY334543

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 (4.45 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。

---

配方 2 中的溶解度: ≥ 2.5 mg/mL (4.45 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。

---

View More

配方 3 中的溶解度: ≥ 2.5 mg/mL (4.45 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。

---

配方 4 中的溶解度: 30% Propylene glycol , 5% Tween 80 , 65% D5W: 30mg/mL

---

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。