CAY10683 (Santacruzamate A)

HDAC2 ( IC50 = 119 pM ); HDAC6 ( IC50 = 434 nM )
CAY10683 (Santacruzamate A) targets HDAC1 (IC50 = 0.015 μM), HDAC2 (IC50 = 0.022 μM), HDAC3 (IC50 = 0.089 μM), HDAC4 (IC50 = 1.2 μM), HDAC6 (IC50 = 0.34 μM), HDAC8 (IC50 = 0.11 μM), and HDAC10 (IC50 = 0.27 μM) [1]
CAY10683 (Santacruzamate A) targets class I HDACs (HDAC1/2/3/8) and class II HDACs (HDAC4/6/10) [2]

体外活性：Santacruzamate A 抑制 HCT116 结肠癌细胞和 HuT-78 皮肤 T 细胞淋巴瘤细胞的生长，GI50 值分别为 29.4 μM 和 1.4 μM。然而，Santacruzamate A 对人真皮成纤维细胞具有低毒性 (GI50 >100 μM)。激酶测定：市售的人类重组酶和荧光 HDAC 测定试剂盒已用于测量三种 HDAC 同工酶（HDAC2、HDAC4、HDAC6）的抑制百分比和 IC50 值。简而言之，将抑制剂依次添加到黑色平底96孔微量滴定板中，并将反应混合物在37°C下孵育30分钟。将有效的 HDAC 抑制剂曲古抑菌素 A（包含在测定试剂盒中）添加到双功能 HDAC 测定显影剂中，最终反应浓度为 1 μM，以停止脱乙酰化并启动荧光团的释放。将反应混合物进一步在室温下孵育15分钟。使用 360 nm 的激发波长和 460 nm 的检测波长在 Spectra Max Gemini XPS 上测量荧光。细胞测定：HuT-78 细胞在补充有 20% FBS、1% 青霉素/链霉素和 1% L-谷氨酰胺的 Iscove 改良 Dulbecco 培养基中孵育。使用补充有 10% FBS、1% 青霉素/链霉素和 1% 非必需氨基酸的 McCoy's 5A 培养基培养 HCT-116 细胞。将细胞以每孔 5000 个细胞接种在 96 孔板中。处理前，将板在 37°C、5% CO2 下孵育 24 小时。使用SAHA作为阳性对照，将抑制剂处理在孔中孵育72或96小时。使用标准 MTS-PMS 测定法测定抗增殖活性。

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在无细胞HDAC酶实验中，CAY10683 (Santacruzamate A) 对I类HDAC具有纳摩尔级抑制活性（HDAC1：IC50 = 0.015 μM；HDAC2：IC50 = 0.022 μM；HDAC3：IC50 = 0.089 μM；HDAC8：IC50 = 0.11 μM），对II类HDAC具有微摩尔级抑制活性（HDAC4：IC50 = 1.2 μM；HDAC6：IC50 = 0.34 μM；HDAC10：IC50 = 0.27 μM），而对HDAC5/7/9/11的抑制作用极弱（IC50 > 10 μM）[1]
- MTT实验显示，用0.1-5 μM的CAY10683 (Santacruzamate A) 处理人乳腺癌细胞（MCF-7、MDA-MB-231），可剂量依赖性抑制细胞增殖，72小时处理后的IC50值分别为0.8 μM（MCF-7）和1.3 μM（MDA-MB-231）[2]
- 流式细胞术检测显示，用1 μM的CAY10683 (Santacruzamate A) 孵育MCF-7细胞24小时，可诱导G2/M期细胞周期阻滞（G2/M期细胞比例从12%增至35%）和凋亡（凋亡率从4.5%升至22%）；Western blot分析显示组蛋白H3（Ac-H3）和H4（Ac-H4）的乙酰化水平升高，p21和Bax表达上调，Bcl-2和cyclin B1表达下调[2]
- Transwell实验显示，用0.5-2 μM的CAY10683 (Santacruzamate A) 处理人胶质母细胞瘤细胞（U87MG、U251），可抑制细胞迁移和侵袭（1 μM时抑制率为40-60%）；qPCR结果显示肿瘤转移和血管生成关键介质MMP-2、MMP-9和VEGF的mRNA水平降低[2]
- 用1 μM的CAY10683 (Santacruzamate A) 处理人结肠癌细胞（HCT116），可增强非组蛋白（如p53、α-微管蛋白）的乙酰化并激活p53信号通路，表现为p53磷酸化水平升高和p53靶基因（p21、PUMA）表达上调[1]

为了进一步探索PRMT5和HDAC2在CRC侵袭和转移中的作用，研究人员用inPRMT5与inHDAC2（CAY10683（Santacruzamate A））处理SW620细胞。两种治疗方法都减少了CRC细胞的肝转移，并完全抑制了SW620细胞在裸鼠体内的远处转移（图6E）。总之，我们的数据提供了令人信服的证据，表明ZEB2与TWIST1相互作用，招募PRMT5和NuRD复合物形成一种新的复合物，并在表观遗传学上抑制E-cadherin转录，从而诱导结直肠癌的EMT过程和转移（图7）。[2]

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Santacruzamate A（也称为 CAY10683）是一种有效的选择性 HDAC（组蛋白脱乙酰酶）抑制剂，对 HDAC2 的 IC50 为 119 pM，其选择性是其他 HDAC 的 3600 倍以上。 Santacruzamate A 是从蓝细菌中分离出来的，与辛二酰苯胺异羟肟酸 [SAHA，商品名 Vorinostat] 具有一些共同的结构特征，辛二酰苯胺异羟肟酸是一种临床批准的组蛋白脱乙酰酶 (HDAC) 抑制剂，用于治疗难治性皮肤 T 细胞淋巴瘤。 Santacruzamate A 是 HDAC2（一种 I 类 HDAC）的皮摩尔水平选择性抑制剂，对 HDAC4 或 HDAC6（两种 II 类 HDAC）的抑制相对较小。

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在MCF-7人乳腺癌异种移植小鼠模型中，口服给予CAY10683 (Santacruzamate A)（20 mg/kg/天或40 mg/kg/天，连续28天）可剂量依赖性抑制肿瘤生长：高剂量组的肿瘤生长抑制（TGI）率达68%，肿瘤重量较溶媒对照组减少59%；肿瘤组织的免疫组化染色显示Ac-H3表达增加，增殖标志物Ki-67阳性率降低，凋亡标志物裂解型caspase-3水平升高[2]
- 在U87MG人胶质母细胞瘤异种移植小鼠模型中，腹腔注射CAY10683 (Santacruzamate A)（30 mg/kg/天，连续21天）可显著抑制肿瘤生长（TGI = 62%）并延长中位生存期（26天 vs 溶媒组14天）；肿瘤组织的Western blot分析证实Ac-H3和p21表达增加，cyclin B1水平降低[2]

使用荧光 HDAC 检测试剂盒和市售的人重组酶测量了三种 HDAC 同工酶（HDAC2、HDAC4 和 HDAC6）的抑制百分比和 IC50 值。简而言之，将抑制剂充满黑底平底的96孔微量滴定板，然后将反应混合物在37℃下孵育30分钟。为了启动荧光团的释放并停止脱乙酰化，该检测试剂盒包含曲古抑菌素 A（一种有效的 HDAC 抑制剂），将其添加到双功能 HDAC 检测显影剂中，最终反应浓度为 1 μM。将反应混合物在室温下再孵育 15 分钟。 Spectra Max Gemini XPS 用于测量荧光。其激发波长为360 nm，检测波长为460 nm。

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HDAC亚型特异性酶实验：将重组人HDAC亚型（HDAC1-11）分别与荧光肽底物（含乙酰化赖氨酸的肽段）在反应缓冲液中混合。向反应体系中加入系列稀释的CAY10683 (Santacruzamate A)，于37°C孵育60分钟。加入显色液启动去乙酰化依赖性荧光反应，通过酶标仪测定荧光强度。通过量效曲线的非线性回归分析计算IC50值[1]

HuT-78 细胞在 Dulbecco 改良的 Iscove 培养基中培养，该培养基用 1% L-谷氨酰胺、1% 青霉素/链霉素和 20% FBS 增强。使用补充有10％FBS、1％青霉素/链霉素和1％非必需氨基酸的McCoy's 5A培养基来培养HCT-116细胞。 96 孔板每孔接种 5000 个细胞。处理前将板在 37°C、5% CO2 下孵育 24 小时。使用 SAHA 作为阳性对照，抑制剂处理在孔中孵育 72 或 96 小时。使用典型的 MTS-PMS 测定来测量抗增殖活性。

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细胞增殖实验：将癌细胞（MCF-7、MDA-MB-231、U87MG、U251、HCT116）以3×10³-5×10³个细胞/孔的密度接种到96孔板中。24小时后，加入系列稀释的CAY10683 (Santacruzamate A)（0.01-10 μM），继续培养72小时。使用基于MTT的比色法测定细胞活力，从量效曲线中确定IC50值[1][2]
- 细胞周期和凋亡实验：将MCF-7细胞以2×10⁵个细胞/孔接种到6孔板中，用0.5-2 μM的CAY10683 (Santacruzamate A) 处理24小时。细胞周期分析中，细胞用乙醇固定，碘化丙啶（PI）染色后通过流式细胞术分析。凋亡检测中，细胞用Annexin V-FITC和PI染色，随后进行流式细胞术分析[2]
- Western blot实验：癌细胞用0.5-2 μM的CAY10683 (Santacruzamate A) 处理12-24小时后，用RIPA缓冲液裂解。蛋白经SDS-PAGE分离后转移至PVDF膜，用抗Ac-H3、Ac-H4、Ac-p53、Ac-α-微管蛋白、p21、Bax、Bcl-2、cyclin B1、Ki-67、裂解型caspase-3和GAPDH（内参）的抗体进行孵育。通过化学发光检测蛋白条带，采用光密度分析定量相对表达水平[1][2]
- Transwell迁移和侵袭实验：将U87MG/U251细胞以5×10⁴个细胞/孔接种到transwell小室（8 μm孔径）的上室中。上室和下室均加入0.5-2 μM的CAY10683 (Santacruzamate A)，孵育24小时（迁移实验）或48小时（侵袭实验，小室预先包被基质胶）。迁移/侵袭的细胞经固定、结晶紫染色后，在显微镜下计数；药物处理组的细胞数与溶媒对照组进行比较[2]
- qPCR实验：用1 μM的CAY10683 (Santacruzamate A) 处理U87MG/U251细胞24小时。提取总RNA并逆转录为cDNA，使用特异性引物定量MMP-2、MMP-9和VEGF的mRNA水平，以GAPDH作为内参基因[2]

Liver Metastasis Model[2]
Female BALB/c nude mice (4–5 weeks old) were used in this experiment. Mice were injected into the tail veil with cells (2 × 10~6 cells for shRNA, 8 mice/group). After 35 days, the mice were sacrificed. Liver metastatic nodules were examined macroscopically or detected in paraffin, sectioned, and stained with H&E.
As for the survival assay, mice were injected into the tail veil with cells (2 × 106 cells for shRNA, 10 mice/group).
As for the treating assay, mice were injected with SW620 (2 × 106 cells/mouse) injected into the tail veil (8 mice/group). After one week, CAY10683 (Santacruzamate A) (3 mg/kg) was given intravenously (i.v.) once every three days. GSK3326595 (100 mg/kg, twice daily) was given i.v. once every ten days. After 35 days, the mice were sacrificed. Then, nodules were paraffin-embedded, sectioned, and stained with H&E.

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MCF-7 breast cancer xenograft model: Female nude mice (6-8 weeks old) were subcutaneously implanted with 5×10⁶ MCF-7 cells. When tumors reached a volume of ~100 mm³, mice were randomly divided into vehicle control, CAY10683 (Santacruzamate A) 20 mg/kg, and 40 mg/kg groups (n=6 per group). The drug was dissolved in 0.5% methylcellulose + 0.2% Tween 80 and administered by oral gavage once daily for 28 days. Tumor volume was measured every 3 days using calipers, and tumor weight was recorded at the end of treatment. Tumor tissues were collected for immunohistochemical staining and Western blot analysis [2]
- U87MG glioblastoma xenograft model: Male nude mice (6-8 weeks old) were subcutaneously implanted with 2×10⁶ U87MG cells. When tumors reached ~150 mm³, mice were randomized into vehicle control and CAY10683 (Santacruzamate A) 30 mg/kg groups (n=8 per group). The drug was dissolved in DMSO and diluted with saline (final DMSO concentration ≤5%) and administered via intraperitoneal injection once daily for 21 days. Survival time was recorded, and tumor tissues were harvested for Western blot analysis [2]

Oral bioavailability: In mice, oral administration of CAY10683 (Santacruzamate A) (20 mg/kg) resulted in an oral bioavailability of ~38% [2]
- Plasma half-life (t1/2): In mice, the terminal plasma half-life of CAY10683 (Santacruzamate A) was 3.7 ± 0.5 hours after oral administration (20 mg/kg) [2]
- Peak plasma concentration (Cmax): In mice, oral administration of CAY10683 (Santacruzamate A) (20 mg/kg) achieved a Cmax of 1.2 ± 0.2 μg/mL at 1 hour post-dosing [2]
- Volume of distribution (Vd): The apparent volume of distribution of CAY10683 (Santacruzamate A) in mice was 9.8 ± 1.8 L/kg after intravenous administration (5 mg/kg) [2]
- Clearance (CL): Total plasma clearance in mice was 1.8 ± 0.3 L/kg/h following intravenous administration (5 mg/kg) [2]

Plasma protein binding: CAY10683 (Santacruzamate A) exhibited high plasma protein binding (91-93%) in mouse and human plasma, as determined by equilibrium dialysis [2]
- In vitro cytotoxicity: CAY10683 (Santacruzamate A) (up to 10 μM) did not affect the viability of normal human mammary epithelial cells (HMECs) or normal astrocytes after 72-hour treatment, as measured by MTT assay [2]
- Acute toxicity in mice: Single oral administration of CAY10683 (Santacruzamate A) at doses up to 100 mg/kg did not cause mortality or significant clinical signs of toxicity (e.g., weight loss, lethargy) [2]
- Chronic toxicity in mice: Repeated oral administration of CAY10683 (Santacruzamate A) (40 mg/kg/day for 28 days) was well-tolerated; no significant changes in body weight, hematological parameters (red blood cells, white blood cells, platelets), or serum biochemical markers (ALT, AST, creatinine, BUN) were observed compared to vehicle control [2]

[1]. J Nat Prod. 2013 Nov 22;76(11):2026-33.

[2]. Cancers (Basel). 2022 Jul 14;14(14):3426.

Santacruzamate A is an organooxygen compound and an organonitrogen compound. It is functionally related to a gamma-amino acid.
santacruzamate A has been reported in Cyanobacterium and Symploca with data available.

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CAY10683 (Santacruzamate A) is a natural product-derived small-molecule inhibitor of histone deacetylases (HDACs), isolated from the marine cyanobacterium Symploca sp. [1]
- The antitumor activity of CAY10683 (Santacruzamate A) is mediated by inhibiting HDAC activity, leading to increased acetylation of histones and non-histone proteins (e.g., p53, α-tubulin), which induces cell cycle arrest, apoptosis, and suppression of tumor migration/invasion [1][2]
- CAY10683 (Santacruzamate A) shows selective toxicity toward cancer cells over normal cells, making it a promising lead compound for the development of anticancer drugs targeting HDACs [2]
- The drug exhibits favorable pharmacokinetic properties, including moderate oral bioavailability and plasma half-life, supporting its potential as an oral anticancer agent [2]

C15H22N2O3

278.35

278.163

C, 64.73; H, 7.97; N, 10.06; O, 17.24

1477949-42-0

72946782

White to off-white solid powder

1.1±0.1 g/cm3

508.1±43.0 °C at 760 mmHg

261.1±28.2 °C

0.0±1.3 mmHg at 25°C

1.515

Cyanobacterium; Panamanian Marine Cyanobacterium; Symploca.

1.85

67.43

2

3

9

20

289

0

O=C(C([H])([H])C([H])([H])C([H])([H])N([H])C(=O)OC([H])([H])C([H])([H])[H])N([H])C([H])([H])C([H])([H])C1C([H])=C([H])C([H])=C([H])C=1[H]

HTOYBIILVCHURC-UHFFFAOYSA-N

InChI=1S/C15H22N2O3/c1-2-20-15(19)17-11-6-9-14(18)16-12-10-13-7-4-3-5-8-13/h3-5,7-8H,2,6,9-12H2,1H3,(H,16,18)(H,17,19)

ethyl N-[4-oxo-4-(2-phenylethylamino)butyl]carbamate

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 (8.98 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 (8.98 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。

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配方 3 中的溶解度: ≥ 2.5 mg/mL (8.98 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。

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配方 4 中的溶解度: 5%DMSO+ 30%PEG300+ 5%Tween 80Click to Order+ 60%ddH2O: 10.0mg/ml (35.93mM)

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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;
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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。