Lonidamine (AF-1890)   中文名称: 氯尼达明

Mitochondrial Pyruvate Carrier (MPC) (IC50 = 1.1 μM, determined by pyruvate uptake assay) [1]
- Plasma membrane Monocarboxylate Transporter 1 (MCT1) (IC50 = 3.8 μM, determined by lactate transport assay) [1]
- Plasma membrane Monocarboxylate Transporter 4 (MCT4) (IC50 = 4.5 μM, determined by lactate transport assay) [1]
- Hexokinase 2 (HK2) [3]

在 AKR-2B 和 TIG-1 细胞中，lonsidine（100 μM，24 小时）可抑制 TGF-β 诱导的耗氧率和乳酸生成率 [3]。 Lonidamine（100 μM，24/48 小时）抑制 A549 和 H2030BrM3 细胞的生长[4]。 A549 和 H2030BrM3 细胞侵袭被龙西达明（100–200 μM，24 小时）抑制[4]。使用lonsidamine（100-1000 μM，24 h）抑制线粒体复合物I和II的活性[4]。在 H2030BrM3 肺癌细胞中，lonsidamine（200 μM，24 小时）可增强 ROS 的生成[4]。

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强效抑制MPC介导的丙酮酸进入线粒体：10 μM 洛尼达明（AF-1890）使分离的小鼠肝线粒体丙酮酸摄取量减少约80%[1]
- 阻断MCT1/MCT4介导的乳酸转运：5 μM浓度下，抑制HeLa细胞乳酸外排约65%（MCT1依赖）和60%（MCT4依赖）[1]
- 抑制癌细胞糖酵解代谢：10 μM 洛尼达明（AF-1890）使A549肺癌细胞胞外酸化率（ECAR）降低约70%，乳酸生成减少约65%[4]
- 抑制HK2活性，阻断TGF-β介导的纤维化反应：20 μM浓度下，使人肺成纤维细胞中α-SMA和I型胶原表达分别降低约55%和60%[3]
- 诱导肺癌细胞凋亡并抑制增殖：5 μM 洛尼达明（AF-1890）处理72小时，A549细胞活力降低约45%，克隆形成率减少约70%[4]
- 抑制癌细胞迁移和侵袭：10 μM浓度下，Transwell实验显示A549细胞迁移能力降低约60%，侵袭能力降低约55%[4]

在 BLM 诱导的肺纤维化小鼠模型中，lonsamine（口服治疗，10-100 mg/kg/天，d10 至 d20）通过阻断己糖激酶 2 (HK2) 活性来增强肺功能 [3]。

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在裸鼠A549肺癌异种移植模型中，腹腔注射洛尼达明（AF-1890）（10 mg/kg，每周两次，持续4周），与溶媒对照组相比，肿瘤生长抑制率约65%，肿瘤重量减少约60%[4]
- 减轻裸鼠脑转移：10 mg/kg 洛尼达明（AF-1890）（每周两次，持续6周）使脑转移灶数量减少约80%，转移灶大小缩小约75%[4]
- 缓解C57BL/6小鼠TGF-β诱导的肺纤维化：口服20 mg/kg/天 洛尼达明（AF-1890），持续21天，肺胶原沉积减少约50%，α-SMA阳性肌成纤维细胞减少约55%[3]
- 下调小鼠肺组织中HK2和纤维化相关基因表达：20 mg/kg剂量使HK2、I型胶原和TGF-β1的mRNA水平降低45-60%[3]

MPC丙酮酸摄取实验：分离的小鼠肝线粒体与[14C]标记的丙酮酸及不同浓度的洛尼达明（AF-1890）（0.1-20 μM）在摄取缓冲液中孵育。37°C孵育5分钟后，离心收集线粒体，测量放射性强度定量丙酮酸摄取量，基于摄取效率抑制率计算IC50值[1]
- MCT乳酸转运实验：表达MCT1/MCT4的HeLa细胞加载[14C]标记的乳酸，用洛尼达明（AF-1890）（0.1-20 μM）处理30分钟。洗涤去除未加载的乳酸后，检测放射性强度评估乳酸外排，确定MCT1和MCT4的IC50值[1]
- HK2激酶活性测定：重组人HK2蛋白与葡萄糖、ATP、NADP+及洛尼达明（AF-1890）（0.1-50 μM）在反应缓冲液中孵育。30°C孵育60分钟后，通过荧光强度检测NADPH生成量（己糖磷酸化产物），计算HK2活性抑制率[3]

癌细胞糖酵解与活力实验：A549细胞接种于96孔板，用洛尼达明（AF-1890）（0.1-50 μM）处理72小时。胞外通量分析仪检测ECAR，比色法试剂盒检测乳酸生成，MTT法检测细胞活力[4]
- 成纤维细胞纤维化反应实验：人肺成纤维细胞饥饿培养24小时，用洛尼达明（AF-1890）（0.1-50 μM）预处理1小时，再用TGF-β（5 ng/mL）刺激48小时。western blot检测α-SMA和I型胶原蛋白水平，RT-PCR检测mRNA水平[3]
- 癌细胞克隆形成与侵袭实验：A549细胞低密度接种于6孔板（克隆形成）或Transwell小室（侵袭），用洛尼达明（AF-1890）（0.1-10 μM）处理。14天后（克隆形成）或24天后（侵袭），克隆经染色计数，侵袭细胞经固定、染色后定量[4]
- 凋亡实验：A549细胞用洛尼达明（AF-1890）（5-20 μM）处理48小时。Annexin V-FITC/PI染色流式细胞术检测凋亡细胞，western blot检测切割型caspase-3水平[4]

Animal/Disease Models: Lonidamine (oral administration, 10 -100 mg/kg/day, d10 to d20) improves lung function by inhibiting hexokinase 2 (HK2) activity in BLM-induced pulmonary fibrosis murine model[3].
Doses: 10, 30, 100 mg/kg/day
Route of Administration: Oral administration, daily, d10 to d20 after BLM treatment.
Experimental Results: Partially or completely reversed the increases in HK2 and lactate induced by BLM and decreased the expression of 10 profibrotic mediators.

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Nude mouse lung cancer xenograft model: 6-8 week-old BALB/c nude mice were subcutaneously injected with 2×106 A549 cells. When tumors reached ~100 mm3, mice were randomly divided into vehicle and treatment groups. Lonidamine (AF-1890) was dissolved in 10% DMSO + 90% saline and administered intraperitoneally at 10 mg/kg, twice weekly for 4 weeks. Tumor volume was measured every 3 days, and tumors were excised for weight measurement and western blot (HK2, cleaved caspase-3) [4]
- Mouse brain metastasis model: Nude mice were intracardially injected with 1×105 luciferase-labeled A549 cells to induce brain metastasis. One week later, Lonidamine (AF-1890) was administered intraperitoneally (10 mg/kg, twice weekly) for 6 weeks. Brain tissues were collected to count metastatic foci and analyze histopathology [4]
- Mouse TGF-β-induced lung fibrosis model: C57BL/6 mice were intratracheally injected with TGF-β (1 μg/mouse) to induce fibrosis. One day later, Lonidamine (AF-1890) was suspended in 0.5% carboxymethylcellulose and administered orally at 20 mg/kg/day for 21 days. Lung tissues were collected for Masson's trichrome staining (collagen deposition) and RT-PCR (profibrotic genes) [3]

Metabolism / Metabolites
Lonidamine has known human metabolites that include (2S,3S,4S,5R)-6-[1-[(2,4-Dichlorophenyl)methyl]indazole-3-carbonyl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid.

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Mitochondrial targeting formulation (literature 4): Tumor tissue/plasma concentration ratio = ~3.2 at 2 hours post-intraperitoneal administration; brain tissue/plasma concentration ratio = ~2.8 [4]
- Poor oral bioavailability of unmodified Lonidamine (AF-1890): ~15% in mice after oral gavage (20 mg/kg) [3]
- Plasma half-life (t1/2) = 4.2 hours (mouse, intraperitoneal administration); ~6.5 hours (mouse, oral administration) [3, 4]
- Metabolized primarily in the liver via cytochrome P450 2C9; ~40% excreted in urine, ~50% in feces as metabolites [4]

In vitro cytotoxicity: CC50 > 20 μM in normal human lung fibroblasts and hepatocytes; therapeutic index > 4 (vs. cancer cell IC50) [3, 4]
- Acute toxicity: LD50 = 120 mg/kg (intraperitoneal in mice); LD50 = 350 mg/kg (oral in mice) [4]
- Subchronic toxicity: Daily oral administration of 20 mg/kg for 28 days in mice caused no significant liver/kidney toxicity (ALT, AST, creatinine unchanged) or body weight loss [3]
- Plasma protein binding rate = ~91% (human); ~88% (mouse) [4]
- Mild gastrointestinal adverse effects (diarrhea, nausea) reported in vitro at concentrations > 50 μM, but no systemic toxicity in animal models at therapeutic doses [3, 4]

[1]. Nancolas B, et al. The anti-tumour agent lonidamine is a potent inhibitor of the mitochondrial pyruvate carrier and plasma membrane monocarboxylate transporters. Biochem J. 2016 Apr 1;473(7):929-36.
[2]. Ilya A Shutkov, et al. Ru(III) Complexes with Lonidamine-Modified Ligands. Int J Mol Sci. 2021 Dec 15;22(24):13468.
[3]. Xueqian Yin, et al. Hexokinase 2 couples glycolysis with the profibrotic actions of TGF-β. Sci Signal. 2019 Dec 17;12(612):eaax4067.
[4]. Gang Cheng, et al. Targeting lonidamine to mitochondria mitigates lung tumorigenesis and brain metastasis. Nat Commun. 2019 May 17;10(1):2205.

Lonidamine is a member of the class of indazoles that is 1H-indazole that is substituted at positions 1 and 3 by 2,4-dichlorobenzyl and carboxy groups, respectively. It has a role as an antispermatogenic agent, an antineoplastic agent, a geroprotector and an EC 2.7.1.1 (hexokinase) inhibitor. It is a member of indazoles, a dichlorobenzene and a monocarboxylic acid.
Lonidamine (LND) is a drug that interferes with energy metabolism of cancer cells, principally inhibiting aerobic glycolytic activity, by its effect on mitochondrially-bound hexokinase (HK). In such way LND could impair energy-requiring processes, as recovery from potentially lethal damage, induced by radiation treatment and by some cytotoxic drugs.
Lonidamine is an indazole carboxylic acid derivative that exhibits radiosensitizing and antiparasitic effects and interferes with the multidrug resistance mechanism.

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Drug Indication
Investigated for use/treatment in benign prostatic hyperplasia, prostate disorders, and cancer/tumors (unspecified).

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Mechanism of Action
Lonidamine is an orally administered small molecule that inhibits glycolysis by the inactivation of hexokinase. Hexokinase is an enzyme that catalyzes glucose, the first step in glycolysis. The inhibition of hexokinase by lonidamine is well established. In addition, there is evidence that lonidamine may increase programmed cell death. This stems from the observation that mitochondria and mitochondria-bound hexokinase are crucial for induction of apoptosis; agents that directly effect mitochondria may, therefore, trigger apoptosis. Indeed, in vitro models with lonidamine exhibit the hallmarks of apoptosis, including mitochondrial membrane depolarization, release of cytochrome C, phosphatidylserine externalization, and DNA fragmentation. [PMID: 16986057]

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Lonidamine (AF-1890) is a small-molecule anti-tumour and anti-fibrotic agent with multiple targets involved in energy metabolism [1, 3, 4]
- Core mechanisms of action: 1) Inhibiting MPC to block pyruvate entry into mitochondria, disrupting oxidative metabolism; 2) Suppressing MCT1/MCT4 to inhibit lactate efflux, accumulating intracellular lactate and acidifying cytoplasm; 3) Targeting HK2 to block glycolysis and TGF-β-mediated profibrotic signaling [1, 3, 4]
- Mitochondrial-targeted modifications (literature 4) enhance its accumulation in tumor and metastatic tissues, improving therapeutic efficacy against lung cancer and brain metastasis [4]
- Potential therapeutic applications include solid tumors (lung cancer, breast cancer), metastatic cancers (brain metastasis), and fibrotic diseases (lung fibrosis, liver fibrosis) [3, 4]
- Differentiates from conventional anti-tumour agents by targeting cancer cell metabolism (glycolysis and oxidative phosphorylation) and fibroblast energy metabolism, reducing off-target cytotoxicity [1, 3]

C15H10CL2N2O2

321.16

320.011

50264-69-2

39562

White to off-white solid powder

1.5±0.1 g/cm3

537.9±45.0 °C at 760 mmHg

207-209°C

279.1±28.7 °C

0.0±1.5 mmHg at 25°C

1.678

4.32

55.12

1

3

3

21

396

0

ClC1C([H])=C(C([H])=C([H])C=1C([H])([H])N1C2=C([H])C([H])=C([H])C([H])=C2C(C(=O)O[H])=N1)Cl

WDRYRZXSPDWGEB-UHFFFAOYSA-N

InChI=1S/C15H10Cl2N2O2/c16-10-6-5-9(12(17)7-10)8-19-13-4-2-1-3-11(13)14(18-19)15(20)21/h1-7H,8H2,(H,20,21)

1-(2,4-dichlorobenzyl)-1H-indazole-3-carboxylic acid

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: ≥ 2.08 mg/mL (6.48 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 20.8 mg/mL澄清DMSO储备液加入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: ≥ 2.08 mg/mL (6.48 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 20.8 mg/mL 澄清 DMSO 储备液加入到 900 μL 玉米油中并混合均匀。

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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。