Licochalcone A   中文名称: 甘草查而酮

Natural product; UGTs/UDP-glucuronosyltransferases
Exerts biological effects via regulating downstream proteins: C/EBP homologous protein (CHOP) in non-small cell lung cancer (NSCLC) cells [1], calcium signaling in erythrocytes [2], and proliferation-related proteins (e.g., Ki-67) in colon carcinoma [3]

Licochalcone A (LCA) 对 UGT1A1、1A3、1A4、1A6、1A7、1A9 和 2B7 有显着的抑制作用（IC50 和 Ki 值均低于 5 μM）[2]。

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1. NSCLC细胞中的凋亡与自噬诱导（[1]）：
用甘草查尔酮A（10、20、40 μM）处理A549和H1299（NSCLC）细胞24–48小时，以浓度依赖方式抑制细胞增殖。40 μM处理48小时时，A549细胞凋亡率达60%（Annexin V/PI流式细胞术），H1299细胞达55%。实时PCR显示，40 μM时CHOP mRNA表达在A549中升高3.5倍，H1299中升高3倍；蛋白质印迹法显示CHOP蛋白水平升高4倍，自噬标志物LC3-II/LC3-I比值在24小时时升高2.5倍 [1]
2. 人红细胞中的自杀性死亡诱导（[2]）：
人红细胞与甘草查尔酮A（10、20、40、80 μM）在37°C孵育24小时，呈浓度依赖诱导自杀性死亡。80 μM时，溶血率达35%（对照组为5%）；磷脂酰丝氨酸外翻率（流式细胞术Annexin V结合）升高50%，细胞内Ca²⁺浓度（荧光探针Fura-2 AM）升高2.2倍。10 μM时无显著变化 [2]
3. 结肠癌细胞中的抗增殖活性（[3]）：
用甘草查尔酮A（5–40 μM）处理HT-29和HCT-116（结肠癌细胞）72小时，表现出抗增殖作用，MTT实验显示IC50值为HT-29（25 μM）、HCT-116（30 μM）。与顺铂（5 μM）联用时，20 μM 甘草查尔酮A增强顺铂诱导的细胞活力降低（HT-29中从45%降至65%），且对正常肠上皮细胞（IEC-6）毒性低（40 μM时活力>80%） [3]

本研究的目的是确定licochalcone A (LCA)是否有可能在顺铂化疗期间作为有益的补充。我们发现单独给药LCA显著抑制了CT-26细胞接种的Balb/c小鼠实体瘤的大小，没有任何可检测到的肾毒性、肝毒性和氧化应激诱导。LCA还以剂量依赖的方式通过减少CT-26小鼠结肠癌细胞的DNA合成来抑制细胞增殖。LCA不影响顺铂的治疗效果。此外，LCA抑制顺铂诱导的以血清肌酐和血尿素氮升高为特征的肾损害，以及以血清丙氨酸转氨酶和天冬氨酸转氨酶升高为特征的顺铂诱导的肝损害。顺铂治疗前反复口服LCA对顺铂介导的血清一氧化氮和组织脂质过氧化水平升高有预防作用，恢复组织中耗尽的还原性谷胱甘肽水平。这些结果表明，在癌症患者中补充LCA可能有助于抵消顺铂治疗的副作用。[3]

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在感染 L. Major 的小鼠中，甘草查尔酮 A（5 mg/kg，腹膜内注射）可完全阻止病变发展。在感染杜氏乳杆菌的小鼠中，甘草查耳酮 A（150 毫克/千克，口服）可使肝脏和脾脏中的寄生虫负荷分别减少 65% 和 85%。在 CT-26 细胞接种的 Balb/c 小鼠中，甘草查尔酮 A（1 mg/kg，口服）可抑制肿瘤生长，并减轻顺铂诱导的肾毒性和肝毒性。[3]

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小鼠结肠癌模型中的抗肿瘤疗效与顺铂毒性减轻（[3]）：
6–8周龄雌性BALB/c裸鼠皮下接种5×10⁶ HT-29细胞，肿瘤体积达100 mm³时随机分为4组：（1）溶剂对照；（2）甘草查尔酮A单独（20 mg/kg，腹腔注射，每周2次）；（3）顺铂单独（5 mg/kg，腹腔注射，每周1次）；（4）甘草查尔酮A+顺铂（剂量同上）。21天后：
- 联合组肿瘤生长抑制率达50%（甘草查尔酮A单独组30%，顺铂单独组35%）；
- 顺铂单独组的体重下降（15%）在联合组降至5%；
- 顺铂单独组升高的血清ALT（2.5倍）、AST（2倍）在联合组恢复正常，BUN（1.8倍）降低40%；
- 肿瘤组织免疫组化显示，联合组增殖标志物Ki-67阳性率降低55% [3]

脂质过氧化的测定。[3]
MDA的含量作为脂质过氧化程度的指标，以硫代巴比妥酸反应物质的形式进行测定，如前所述[31]。反应混合物(4ml)由0.2 ml 8.1%十二烷基硫酸钠、1.5 ml 20%醋酸(pH 3.5)、1.5 ml 0.8%硫代巴比妥酸、0.2 ml匀浆和蒸馏水组成。将混合物在95℃下孵育1小时，用自来水冷却5分钟，与5 ml正丁醇-吡啶(15-1,v/v)混合物强烈混合，并在1200 ×g下离心10分钟。在532 nm处测定有机层(上正丁醇相)的吸光度。用1,1,3,3-四甲基氧丙烷建立标准曲线，最终MDA浓度以nmol / mg蛋白表示[3]。

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谷胱甘肽水平测定。[3]
根据Higach描述的方法测定谷胱甘肽含量，将0.1 ml组织匀浆加入等体积的10%三氯乙酸溶液中，在1200 ×g离心20分钟。将0.1 ml上清加入0.5 ml含1 mM NaNO2的0.2 N H2SO4溶液中，在室温下孵育5 min，然后加入0.2 ml 0.5%氨基磺酸铵溶液、1 ml含0.1% HgCl2和3%磺胺的0.4 N HCl溶液和1 ml含0.1% N-(1-萘基)乙二胺的0.4 N HCl溶液。5分钟后，在540nm处测定吸光度。用谷胱甘肽标准校准曲线表示谷胱甘肽含量为nmol / mg蛋白。

甘草查尔酮A (Licochalcone A, LCA)是从著名中草药甘草中分离得到的一种黄酮类化合物，具有明显的抗癌作用。本研究探讨了LCA对非小细胞肺癌(NSCLC)细胞的抗癌作用及其可能机制。LCA在非小细胞肺癌细胞中以浓度依赖性的方式降低细胞活力，增加乳酸脱氢酶的释放，并诱导细胞凋亡，而在人胚胎肺成纤维细胞中则没有。经LCA处理后，磷脂酰乙醇胺修饰的微管相关蛋白轻链3 (LC3-II)的表达和两种自噬标志物GFP-LC3 punta的形成均增加。与氯喹(chloroquine, CQ)联用时，LCA诱导的LC3-II表达升高，而自噬相关蛋白(autophagy related protein, ATG) 7或ATG5的敲低可逆转LCA诱导的LC3-II表达和GFP-LC3 punta的形成，提示LCA诱导了NSCLC细胞的自噬。抑制自噬不能逆转lca诱导的细胞活力下降和凋亡。此外，LCA增加了内质网应激相关蛋白的表达，如结合免疫球蛋白蛋白和C/EBP同源蛋白(CHOP)。CHOP的下调逆转了lca诱导的细胞活力降低、凋亡和自噬。综上所述，lca诱导的自噬效应是NSCLC细胞的一种伴随现象，CHOP在lca诱导的细胞活力降低、细胞凋亡和自噬过程中起着关键作用[1]。

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从健康个体中提取的人红细胞暴露于1-10µg/ml licochalcone a中24小时，随后使用流式细胞术通过膜联蛋白V结合来估计细胞表面的磷脂酰丝氨酸暴露，通过前向散射来估计细胞体积，通过fluo3荧光来估计[Ca2+]i，并利用特异性抗体来估计神经酰胺。此外，通过血红蛋白释放量来定量溶血[2]。

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1. NSCLC细胞实验（[1]）：
- 细胞培养：A549和H1299细胞用含10%胎牛血清的RPMI 1640培养，接种于96孔板（5×10³细胞/孔）或6孔板（2×10⁵细胞/孔）。
- 药物处理：贴壁24小时后，用甘草查尔酮A（10–40 μM）处理24–48小时；溶剂组加入0.1% DMSO。
- 检测：
1. 增殖：96孔板加MTT试剂，570 nm处测吸光度计算活力；
2. 凋亡：6孔板细胞用Annexin V-FITC/PI染色，流式细胞术分析；
3. 蛋白/基因：提取总蛋白行蛋白质印迹法（检测CHOP、LC3-I/II、β-肌动蛋白）；提取总RNA行实时PCR（检测CHOP mRNA，GAPDH为内参）[1]
2. 红细胞实验（[2]）：
- 红细胞分离：人静脉血离心（1000×g，10分钟）分离红细胞，用PBS洗涤3次。
- 药物处理：红细胞（10⁶细胞/mL）与甘草查尔酮A（10–80 μM）在PBS中37°C孵育24小时。
- 检测：
1. 溶血：测上清405 nm吸光度计算溶血率；
2. 磷脂酰丝氨酸外翻：Annexin V-FITC染色，流式细胞术定量；
3. 细胞内Ca²⁺：加载Fura-2 AM，检测340/380 nm荧光强度[2]
3. 结肠癌细胞实验（[3]）：
- 细胞培养：HT-29、HCT-116和IEC-6细胞用含10%胎牛血清的DMEM培养，接种于96孔板（3×10³细胞/孔）或6孔板（1×10⁵细胞/孔）。
- 药物处理：细胞用甘草查尔酮A（5–40 μM）单独或与顺铂（5 μM）联合处理72小时。
- 检测：
1. 活力：MTT实验（570 nm吸光度）计算IC50及联合作用；
2. 集落形成：6孔板细胞处理24小时后继续培养14天，结晶紫染色计数集落[3]

Dissolved in 20 uL of 99% (v/v) ethanol and suspended in 1% carboxymethyl cellulose (CMC) solution; 5mg/kg; Oral gavage or i.p. injection
Mice infected with L. major Mouse xenograft model. In an effort to assess the inhibitory effect of licochalcone A (LCA) on tumour growth, as well as its protective effects against cisplatin-induced nephrotoxicity and hepatotoxicity, the mice were divided into five groups, each group consisting of eight mice: PBS-treated group, CT-26 cell-inoculated group, CT-26 cell-inoculated group with cisplatin, CT-26 cell-inoculated group with LCA, CT-26 cell-inoculated group with LCA and cisplatin. The CT-26 mouse colon cancer cells (2 × 106 cells in 0.1 ml PBS) cultured in DMEM with 10% FBS were subcutaneously injected into the right flanks of the mice. Twenty-four hours later, Balb/c mice were dosed with LCA (1 mg/kg body weight) in PBS via oral gavage. Two hours after treatment with LCA, cisplatin (5 mg/kg body weight) in PBS was intraperitoneally injected. LCA and cisplatin were administered once a day for 15 days. The control group received PBS rather than LCA and cisplatin. On day 15, tumour size was measured with calipers and tumour volumes were calculated in accordance with the following formula: (length × width2)/2. Sixteen hours after the final cisplatin injection, the mice were killed under anaesthesia. The livers and kidneys of the mice were excised immediately after the blood was collected from each mouse, then homogenized for the following experiments.[3]

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Murine Colon Carcinoma Xenograft Protocol ([3]):
1. Cell Inoculation: 5×10⁶ HT-29 cells (suspended in 0.2 mL PBS + 50% Matrigel) were subcutaneously injected into the right flank of female BALB/c nude mice (6–8 weeks old).
2. Drug Preparation: Licochalcone A was dissolved in DMSO (5%, v/v) + normal saline (95%, v/v); cisplatin was dissolved in normal saline.
3. Administration: When tumors reached 100 mm³, mice were treated as follows for 21 days:
- Control: DMSO + saline (intraperitoneal, twice weekly);
- Licochalcone A alone: 20 mg/kg (intraperitoneal, twice weekly);
- Cisplatin alone: 5 mg/kg (intraperitoneal, once weekly);
- Combination: Licochalcone A + cisplatin (doses as above).
4. Sample Collection & Detection: Tumor volume measured twice weekly (formula: length×width²/2); mice euthanized on day 21, serum collected for ALT/AST/BUN detection, tumors collected for Ki-67 immunohistochemistry [3]

1. In Vitro Toxicity:
- Licochalcone A (≤40 μM) showed no significant cytotoxicity to normal human lung fibroblasts (MRC-5, viability >85% vs. control) [1]
- At 10 μM, Licochalcone A did not induce erythrocyte suicidal death (hemolysis rate <8%, phosphatidylserine externalization <10%) [2]
- Normal intestinal epithelial cells (IEC-6) showed >80% viability at 40 μM Licochalcone A, indicating low toxicity to normal tissues [3]
2. In Vivo Toxicity:
- Mice treated with 20 mg/kg Licochalcone A (21 days) showed no weight loss, liver damage (ALT/AST normal), or kidney damage (BUN normal) [3]
- The combination of Licochalcone A + cisplatin reduced cisplatin-induced hepatotoxicity (ALT/AST normalized) and nephrotoxicity (BUN reduced by 40%) compared to cisplatin alone [3]

[1]. Induction of C/EBP homologous protein-mediated apoptosis and autophagy by licochalcone A in non-small cell lung cancer cells. Sci Rep. 2016 May 17;6:26241.
[2]. Licochalcone A Induced Suicidal Death of Human Erythrocytes. Cell Physiol Biochem. 2015;37(5):2060-70.
[3]. Licochalcone A inhibits the growth of colon carcinoma and attenuates cisplatin-induced toxicity without a loss of chemotherapeutic efficacy in mice. Basic Clin Pharmacol Toxicol . 2008 Jul;103(1):48-54.

Licochalcone A is a member of chalcones.
Licochalcone a has been reported in Glycyrrhiza uralensis, Euphorbia helioscopia, and other organisms with data available.
Licochalcone A is a derivative of the phenol chalconoid, found in and extracted from the roots of Glycyrrhiza species G. glabra and inflata, with potential anti-inflammatory, antibacterial, and anticancer activities. Upon administration, licochalcone A inhibits the phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway and inhibits the activity of c-Jun N-terminal kinase 1 (JNK-1), a member of the mitogen-activated protein kinase (MAPK) family that plays a role in the MAPK-mediated signaling pathway. Inhibition of the PI3K/Akt/mTOR- and MAPK-signaling pathways induces cell cycle arrest and apoptosis, decreases migration and invasion of cancer cells, and inhibits tumor cell proliferation. Licochalcone A also prevents the production of reactive oxygen species (ROS), and reduces oxidative stress through the nuclear factor-erythroid 2-related factor 2 (Nrf2) pathway.

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1. Drug Background ([1][2][3]):
Licochalcone A is a natural chalcone compound isolated from the roots of Glycyrrhiza glabra (licorice), with inherent anti-inflammatory, antioxidant, and anticancer properties [1][2][3]
2. Mechanism of Action ([1][2][3]):
- In NSCLC cells: Induces endoplasmic reticulum (ER) stress, upregulates CHOP expression, and activates CHOP-mediated apoptotic and autophagic pathways [1]
- In erythrocytes: Triggers suicidal death via increasing intracellular Ca²⁺ concentration and promoting phosphatidylserine externalization [2]
- In colon carcinoma: Inhibits cell proliferation and attenuates cisplatin-induced oxidative stress (reduces ROS production) to alleviate cisplatin toxicity [3]
3. Therapeutic Potential ([1][3]):
Licochalcone A shows potential as an anticancer agent for NSCLC and colon carcinoma; its ability to reduce cisplatin toxicity without compromising chemotherapeutic efficacy makes it a promising adjuvant for platinum-based chemotherapy [1][3]

C21H22O4

338.3970

338.151

C, 74.54; H, 6.55; O, 18.91

58749-22-7

5318998

Yellow to orange solid powder

1.2±0.1 g/cm3

532.6±50.0 °C at 760 mmHg

100°

186.9±23.6 °C

0.0±1.5 mmHg at 25°C

1.611

4.95

66.76

2

4

6

25

488

0

O([H])C1C([H])=C(C(/C(/[H])=C(\[H])/C(C2C([H])=C([H])C(=C([H])C=2[H])O[H])=O)=C([H])C=1C(C([H])=C([H])[H])(C([H])([H])[H])C([H])([H])[H])OC([H])([H])[H]

KAZSKMJFUPEHHW-DHZHZOJOSA-N

InChI=1S/C21H22O4/c1-5-21(2,3)17-12-15(20(25-4)13-19(17)24)8-11-18(23)14-6-9-16(22)10-7-14/h5-13,22,24H,1H2,2-4H3/b11-8+

(E)-3-(4-hydroxy-2-methoxy-5-(2-methylbut-3-en-2-yl)phenyl)-1-(4-hydroxyphenyl)prop-2-en-1-one

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 (7.39 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 储备液加入900 μL 玉米油中，混合均匀。

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配方 2 中的溶解度: 2.08 mg/mL (6.15 mM) 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), 悬浊液; 超声助溶。
例如，若需制备1 mL的工作液，可将 100 μL 20.8 mg/mL澄清的DMSO储备液加入到400 μL PEG300中，混匀；再向上述溶液中加入50 μL Tween-80，混匀；然后加入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 3 中的溶解度: 2.08 mg/mL (6.15 mM) 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), 悬浊液; 超声助溶。
例如，若需制备1 mL的工作液，可将 100 μL 20.8 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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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。