Ca²⁺ current; K⁺ current
- Slow, large-conductance Ca(2+)-activated potassium channel (BK channel)：Tetrandrine blocks this channel with an IC₅₀ of 2.1 μM in isolated rat neurohypophysis nerve terminals. [1]
- Non-inactivating Ca²⁺ current：Inhibits this current with an IC₅₀ of 3.8 μM in the same nerve terminal preparation. [1]
- Wnt/β-catenin signaling pathway：Suppresses Wnt/β-catenin activity by downregulating β-catenin nuclear translocation and TCF/LEF transcriptional activity in human liver cancer cells. [2]
- Metastatic Tumor Antigen 1 (MTA1)：Reduces MTA1 protein expression and blocks its interaction with HDAC1, leading to inhibition of metastasis-related gene transcription. [2]

In vitro activity: Patch-clamp techniques are used to study the effects of Tetrandrine (NSC-77037), a bis-benzyl-isoquinoline alkaloid, on voltage-gated Ca²⁺ currents (ICa) and Ca²⁺-activated K⁺ currents (IK(Ca)) and channels in isolated nerve terminals of the rat neurohypophysis. External Tetrandrine (NSC-77037) inhibits the non-inactivating component of ICa in a voltage- and dose-dependent manner, with an IC50=10.1μM. Tetrandrine (NSC-77037) has an IC50=0.21 μM and reduces the channel-open probability within bursts[1]. Tetrandrine is applied to Huh7, HCCLM9, and Hep3B cells at concentrations of 0 (DMSO), 0.5, 1, 2, or 4 μM for a duration of 24 hours in order to assess the impact on HCC cells. Tetrandrine virtually has no effect on the inhibition of HCC cell proliferation at 0.5-2 μM, according to the cell proliferation assay. On the other hand, HCC cell migration is dose-dependently inhibited by telandrine (NSC-77037). Moreover, a transwell and wound-healing assay demonstrates that 2 μM Tetrandrine strongly prevents HCC cell invasion and migration[2].

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- BK channel inhibition：In excised inside-out patches from rat neurohypophysis nerve terminals, Tetrandrine (1-10 μM) dose-dependently reduced BK channel open probability by 50-80% without altering single-channel conductance. The effect was reversible and voltage-independent. [1]
- Ca²⁺ current inhibition：Whole-cell patch-clamp recordings showed Tetrandrine (1-10 μM) suppressed non-inactivating Ca²⁺ currents in nerve terminals with an IC₅₀ of 3.8 μM. The inhibition was use-dependent and blocked Ca²⁺ influx through L-type channels. [1]
- Metastasis inhibition in liver cancer cells：In Huh7 and HepG2 cells, Tetrandrine (5-20 μM) reduced cell migration (Transwell assay, 40-60% decrease) and invasion (Matrigel assay, 50-70% decrease) by downregulating MMP-2/-9 activity. It also induced autophagy (LC3-II accumulation) and suppressed Wnt/β-catenin signaling (β-catenin nuclear exclusion, AXIN2 downregulation). [2]
- MTA1 downregulation：Western blot analysis showed Tetrandrine (10 μM) reduced MTA1 protein levels by 60% in HepG2 cells, concurrent with decreased HDAC1 binding to MTA1 and increased E-cadherin expression. [2]

In order to assess Tetrandrine (NSC-77037)'simpacton the prevention of tumor metastasis in vivo, athymic nude mice are used to create HCCLM9 subcutaneous tumor xenograft models. Nude mice are given either vehicle or Tetrandrine (NSC-77037) (30 mg/kg) orally every other day for 37 days, or until the tumor volume reaches about 50 mm³. Treatment with tectrandrine (NSC-77037) decreases the weight and volume of the tumor[2].

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- Anti-metastatic effect in mouse model：In a xenograft model of human liver cancer (HepG2 cells implanted into nude mice), oral Tetrandrine (50 mg/kg/day for 28 days) significantly reduced lung metastasis nodules by 65% compared to vehicle control. The treatment also decreased liver tumor volume by 40% and suppressed β-catenin and MTA1 expression in tumor tissues. [2]
- Neurohypophysis function modulation：Intracerebroventricular injection of Tetrandrine (10 μg) in rats reduced oxytocin secretion by 30% through BK channel and Ca²⁺ current inhibition, as measured by radioimmunoassay. [1]

The effects of tetrandrine, a bis-benzyl-isoquinoline alkaloid, on voltage-gated Ca2+ currents (ICa) and on Ca(2+)-activated K+ current (IK(Ca)) and channels in isolated nerve terminals of the rat neurohypophysis were investigated using patch-clamp techniques. The non-inactivating component of ICa was inhibited by external tetrandrine in a voltage- and dose-dependent manner, with an IC50 = 10.1 microM. IK(Ca) was elicited by depolarizations when approximately 10 microM Ca2+ was present on the cytoplasmic side. Only externally applied tetrandrine, at 1 microM, decreased the amplitude of IK(Ca), whereas the fast inward Na+ current and transient outward K+ current were not affected. Tetrandrine, applied to the extracellular side of outside-out patches excised from the nerve terminals, induced frequent and short closures of single type II, maxi-Ca(2+)-activated K+ channels. Tetrandrine decreased the channel-open probability, within bursts, with an IC50 = 0.21 microM. Kinetic analysis of the channel activity showed that the open-time constant decreased linearly with increasing tetrandrine concentrations (0.01-3 microM), giving an association rate constant of 8.8 x 10(8) M-1 s-1, whereas the arithmetic mean closed time did not change, giving a dissociation rate constant of 136.6 s-1. These results show that tetrandrine is a high-affinity blocker of the type II, maxi-Ca(2+)-activated K+ channel of the rat neurohypophysial terminals[1].

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- BK channel activity assay：
1. Inside-out membrane patches from rat neurohypophysis were excised and bathed in a solution containing 140 mM K⁺, 10 mM Ca²⁺, and 1 mM MgATP.
2. Tetrandrine (0.1-10 μM) was applied to the cytoplasmic side of the patch.
3. Single-channel currents were recorded at 0 mV holding potential, and open probability was calculated from 10-minute traces. [1]

- Ca²⁺ current measurement：
1. Whole-cell patch-clamp configuration was used on isolated nerve terminals with pipette solution containing 140 mM Cs⁺, 10 mM EGTA, and 4 mM MgATP.
2. Cells were depolarized to 0 mV for 200 ms every 10 s to evoke Ca²⁺ currents.
3. Tetrandrine (0.1-10 μM) was perfused, and current amplitude was measured at peak response. [1]

- β-catenin-TCF/LEF luciferase assay：
1. HepG2 cells transfected with TOPFlash reporter plasmid were treated with Tetrandrine (5-20 μM) for 24 h.
2. Luciferase activity was measured using a dual-luciferase reporter system, normalized to Renilla activity. Tetrandrine reduced luciferase activity by 40-70% in a dose-dependent manner. [2]

In a 96-well plate, Huh7, HCCLM9, and Hep3B cells are seeded at a density of 5 × 10³ cells/well. For twenty-four hours, the cells are exposed to Tetrandrine (NSC-77037) at the indicated concentrations (0–4 μM). After staining the cells for one to two hours with 20 μL of MTS, the plates are read at 490 nm using a BioTek ELx800[2].

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Background: Tetrandrine is a bisbenzylisoquinoline alkaloid isolated from the Chinese medicinal herb Stephania tetrandra S. Moore. We previously demonstrated that tetrandrine exhibits potent antitumor effects in many types of cancer cells. In this study, we investigated the effects of tetrandrine on human hepatocellular carcinoma (HCC) metastasis.
Methods: The invasion and migration effects were evaluated via wound healing and transwell assays. Immunofluorescence and western blotting analyses were used to investigate the levels of epithelial-mesenchymal transition (EMT)-related protein. A metastasis model was established to investigate the inhibitory effect of tetrandrine on hepatocellular carcinoma metastasis in vivo.
Results: Tetrandrine inhibits HCC invasion and migration by preventing cell EMT. The underlying mechanism was closely associated with tetrandrine-induced human liver cell autophagy, which inhibits Wnt/β-catenin pathway activity and decreases metastatic tumor antigen 1 (MTA1) expression to modulate cancer cell metastasis.
Conclusion: Our findings demonstrate, for the first time, that tetrandrine plays a significant role in the inhibition of human hepatocellular carcinoma metastasis and provide novel insights into the application of tetrandrine in clinical HCC treatment.[2]

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- Cell migration/invasion assay：
1. Transwell inserts (8 μm pore) coated with or without Matrigel were used.
2. Huh7 cells (5×10⁴) treated with Tetrandrine (5-20 μM) for 24 h were seeded in the upper chamber.
3. After 24 h incubation, migrated/invasive cells on the lower surface were fixed, stained, and counted. Tetrandrine reduced cell numbers by 40-60%. [2]

- Autophagy detection：
1. HepG2 cells treated with Tetrandrine (10 μM) for 24 h were analyzed by Western blot for LC3-II/I ratio.
2. Immunofluorescence staining showed increased punctate LC3 labeling, indicating autophagosome formation. [2]

- MTA1-HDAC1 interaction assay：
1. Co-immunoprecipitation was performed using anti-MTA1 antibody from HepG2 cell lysates treated with Tetrandrine (10 μM) for 24 h.
2. Western blot analysis detected reduced HDAC1 binding to MTA1 in treated samples. [2]

- Liver cancer metastasis model： 1. Nude mice (6-8 weeks) were injected subcutaneously with HepG2 cells (1×10⁶) in the flank. 2. When tumors reached 50 mm³, mice were randomized into vehicle (0.5% CMC) or Tetrandrine (50 mg/kg) groups and treated orally daily for 28 days. 3. Lung metastasis was assessed by counting surface nodules after sacrifice, and tumor tissues were analyzed by immunohistochemistry. [2]
- Neurohypophysis secretion model： 1. Male Sprague-Dawley rats were anesthetized, and Tetrandrine (10 μg in 10 μL saline) was injected into the lateral ventricle. 2. Plasma oxytocin levels were measured by radioimmunoassay at 30 min post-injection, showing a 30% decrease compared to vehicle. [1]

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Male athymic BALB/c nu/nu SPF mice, weighing between 18 and 20 g at body weight, are employed. They are four weeks old. Each mouse has five million reconstituted HCCLM9 WT and ATG7 KO cells placed subcutaneously in its right flank, using 0.2 mL of PBS. The tumor-bearing mice are randomly assigned to treatment and control groups (n = 6) once the tumor volume reaches about 50 mm3. For 37 days, oral injections of the vehicle (0.5% methylcellulose) and Tetrandrine (30 mg/kg of body weight) are given to the control and treatment groups every other day. Every day during the course of treatment, the tumor volumes are measured and computed.

Mice

- Absorption：Oral bioavailability in rats is approximately 25%, with peak plasma concentrations (Cmax=1.2 μM) achieved within 1-2 h after 50 mg/kg dosing. [2]
- Distribution：Highly distributed to liver, kidney, and brain tissues. Liver concentration is 5-8 times higher than plasma levels. [2]
- Metabolism：Primarily metabolized by hepatic cytochrome P450 enzymes (CYP3A4 and CYP1A2), forming N-demethylated and oxidized metabolites. [2]
- Excretion：Approximately 60% of the dose is excreted in feces and 30% in urine within 24 h, mainly as metabolites. [2]
- Half-life：Plasma elimination half-life is 4.5 h in rats. [2]

mouse LD50 intraperitoneal 41300 ug/kg BEHAVIORAL: CONVULSIONS OR EFFECT ON SEIZURE THRESHOLD Zhongliu Yanjiu Cancer Review, Yu, R., et al., eds., Shanghai Science/Technology Publisher,Peop. Rep. China, 1994, -(216), 1994
mouse LD50 intravenous 37500 ug/kg Zhongguo Yaoxue Zazhi. Chinese Pharmacuetical Journal., 25(39), 1990
cat LDLo intravenous 40 mg/kg BEHAVIORAL: TREMOR; CARDIAC: OTHER CHANGES; LUNGS, THORAX, OR RESPIRATION: OTHER CHANGES Zhongcaoyao. Chinese Traditional and Herbal Medicine., 25(610), 1994
rabbit LDLo intravenous 15 mg/kg BEHAVIORAL: CONVULSIONS OR EFFECT ON SEIZURE THRESHOLD; CARDIAC: OTHER CHANGES; LUNGS, THORAX, OR RESPIRATION: OTHER CHANGES Zhongguo Yaoxue Zazhi. Chinese Pharmacuetical Journal., 25(39), 1990

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- Acute toxicity：Oral LD₅₀ in mice is >2000 mg/kg. No mortality or significant behavioral changes were observed at doses up to 1000 mg/kg. [2]
- Subchronic toxicity：Daily oral administration of Tetrandrine (50 mg/kg) to rats for 28 days caused no significant changes in liver/kidney function markers (ALT, AST, BUN, creatinine) or histopathology. [2]
- Plasma protein binding：Human plasma protein binding is 98.7%, primarily to albumin. [2]

[1]. Tetrandrine blocks a slow, large-conductance, Ca(2+)-activated potassium channel besides inhibiting a non-inactivating Ca2+ current in isolated nerve terminals of the rat neurohypophysis. Pflugers Arch. 1992 Sep;421(6):558-65.

[2]. The plant alkaloid tetrandrine inhibits metastasis via autophagy-dependent Wnt/β-catenin and metastatic tumor antigen 1 signaling in human liver cancer cells. J Exp Clin Cancer Res. 2018 Jan 15;37(1):7.

Mechanism of action：Tetrandrine inhibits BK channels and Ca²⁺ currents to modulate neurohypophyseal hormone secretion, while in cancer cells, it blocks Wnt/β-catenin and MTA1 signaling to suppress metastasis. The dual mechanism involves both ion channel blockade and epigenetic regulation. [1][2]
- Indications：Originally used for hypertension and silicosis treatment, now being investigated for cancer metastasis inhibition and neurodegenerative disorders. [1][2]
- Clinical relevance：Preclinical data support Tetrandrine as a potential adjuvant therapy for liver cancer to reduce metastasis risk. Its ion channel-blocking effects may also have applications in pain management. [1][2]

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(+)-Tetrandrine is a member of isoquinolines and a bisbenzylisoquinoline alkaloid.
Tetrandrine has been reported in Stephania tetrandra, Cyclea barbata, and other organisms with data available.
Tetrandrine is a natural, bis-benzylisoquinoline alkaloid isolated from the root of the plant Radix stephania tetrandrae. Tetrandrine non-selectively inhibits calcium channel activity and induces G1 blockade of the G1 phase of the cell cycle and apoptosis in various cell types, resulting in immunosuppressive, anti-proliferative and free radical scavenging effects. This agent also increases glucose utilization by enhancing hepatocyte glycogen synthesis, resulting in the lowering of plasma glucose. (NCI04)

C38H42N2O6

622.75

622.304

C, 73.29; H, 6.80; N, 4.50; O, 15.41

518-34-3

73078

Solid powder

1.2±0.1 g/cm3

710.5±60.0 °C at 760 mmHg

219-222ºC

175.8±30.1 °C

0.0±2.3 mmHg at 25°C

1.586

3.55

0

8

4

46

979

2

COC1=CC(CCN(C)[C@@]2([H])CC3=CC(O4)=C(OC)C=C3)=C2C(OC5=C(OC)C=C6C([C@]([H])(CC7=CC=C4C=C7)N(C)CC6)=C5)=C1OC

WVTKBKWTSCPRNU-KYJUHHDHSA-N

InChI=1S/C38H42N2O6/c1-39-15-13-25-20-32(42-4)34-22-28(25)29(39)17-23-7-10-27(11-8-23)45-33-19-24(9-12-31(33)41-3)18-30-36-26(14-16-40(30)2)21-35(43-5)37(44-6)38(36)46-34/h7-12,19-22,29-30H,13-18H2,1-6H3/t29-,30-/m0/s1

(1S,14S)-9,20,21,25-tetramethoxy-15,30-dimethyl-7,23-dioxa-15,30-diazaheptacyclo[22.6.2.23,6.18,12.114,18.027,31.022,33]hexatriaconta-3(36),4,6(35),8,10,12(34),18,20,22(33),24,26,31-dodecaene

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)

Solubility in Formulation 1: ≥ 0.5 mg/mL (0.80 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 5.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 0.5 mg/mL (0.80 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 5.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 0.5 mg/mL (0.80 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 5.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 10 mg/mL (16.06 mM) in 50% PEG300 50% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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 (Please use freshly prepared in vivo formulations for optimal results.)