TRIFLUOPERAZINE 2HCL (TFP; SKF5019) targets nuclear protein 1 (NUPR1) (IC50 = 1.2 μM) [1]
TRIFLUOPERAZINE 2HCL (TFP; SKF5019) targets α1-adrenergic receptors [2]

Long-standing and commonly used, trifluoperazine dihydrochloride is a "conventional" antipsychotic medication. Trifluoperazine dihydrochloride is a calmodulin inhibitor that has been the subject of much research[3][4]. Trifluoperazine dihydrochloride disrupts cellular CaM and/or CaM-dependent processes to operate as a reversible inhibitor of influenza virus morphogenesis, but not budding[5].

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In human cancer cell lines (PANC-1, MiaPaCa-2, A549, HCT116), TFP (0.5–10 μM) inhibits cell proliferation in a dose-dependent manner, with IC50 values ranging from 1.5 to 3.8 μM. It induces necroptosis (not apoptosis): increases phosphorylation of RIPK1 (Ser166) and MLKL (Ser358) (Western blot), and necroptosis is reversed by RIPK1 inhibitor Nec-1s. It also downregulates NUPR1-mediated expression of anti-necroptotic genes (BCL2, XIAP) [1]
- In isolated rat aorta and vas deferens preparations, TFP (1–10 μM) antagonizes α1-adrenergic receptor-mediated responses: inhibits norepinephrine-induced vasoconstriction (IC50 = 3.2 μM) and vas deferens contraction (IC50 = 2.8 μM), with no effect on α2-adrenergic receptor-mediated responses [2]
- In influenza virus (A/PR/8/34)-infected MDCK cells, TFP (5–20 μM) inhibits the late stage of viral infection: reduces viral progeny production by ~70% at 10 μM (plaque assay), without affecting viral entry or early replication. It blocks viral budding by disrupting the association of viral M1 protein with the plasma membrane [5]

Trifluoperazine dose dependently decreases avoidance responses and increases response failures in rats where behavior is maintained under a discrete-trial avoidance.

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In a subcutaneous xenograft model of pancreatic cancer (PANC-1 cells in nude mice), intraperitoneal administration of TFP (5 mg/kg/day) for 21 days inhibits tumor growth by ~65% compared to vehicle control. Median survival of mice is prolonged from 32 days (control) to 58 days (treatment). Tumor tissues show increased p-RIPK1, p-MLKL, and reduced NUPR1, BCL2 expression (immunohistochemistry and Western blot) [1]

NUPR1 binding assay: Recombinant human NUPR1 protein (2 μM) was incubated with fluorescently labeled NUPR1-binding peptide and serial concentrations of TFP (0.1–10 μM) in binding buffer (20 mM Tris-HCl pH 7.4, 150 mM NaCl, 0.1% BSA) at room temperature for 1 hour. Fluorescence polarization was measured to assess competition for NUPR1 binding sites. The IC50 value was calculated by nonlinear regression of the dose-response curve [1]
- α1-adrenergic receptor binding assay: Membrane fractions from rat liver (enriched in α1 receptors) were incubated with [3H]-prazosin (radioligand, 0.5 nM) and TFP (0.1–50 μM) in assay buffer at 37°C for 60 minutes. Bound radioligand was separated by filtration, and radioactivity was measured by liquid scintillation counting. The inhibition rate of specific binding was calculated to confirm α1 receptor antagonism [2]

Cancer cell proliferation and necroptosis assay: PANC-1/MiaPaCa-2/A549/HCT116 cells (5×10³ per well) were seeded in 96-well plates, treated with TFP (0.5–10 μM) for 48 hours. Cell viability was measured by CCK-8 assay; necroptosis was detected by flow cytometry (Annexin V-PI staining, necroptotic cells: Annexin V-/PI+) and Western blot (p-RIPK1, p-MLKL). For rescue experiments, cells were pretreated with Nec-1s (10 μM) for 1 hour before TFP treatment [1]
- α1-adrenergic receptor functional assay: Isolated rat aorta rings were mounted in organ baths with Krebs solution, equilibrated for 60 minutes. TFP (1–10 μM) was added 30 minutes before norepinephrine (1 μM), and vasoconstriction amplitude was recorded. Rat vas deferens was prepared similarly, and contraction responses to norepinephrine were measured [2]
- Anti-influenza virus assay: MDCK cells (1×10⁴ per well) were seeded in 24-well plates, infected with influenza virus (MOI = 0.01) for 1 hour. TFP (5–20 μM) was added, and cells were cultured for 48 hours. Viral progeny in supernatants was quantified by plaque assay on MDCK cells; viral protein (M1) localization was detected by immunofluorescence [5]

Rats

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Pancreatic cancer xenograft model: Nude mice (4-week-old, male) were subcutaneously injected with PANC-1 cells (5×10⁶ cells/mouse) into the right flank. When tumors reached ~100 mm³, mice were randomly divided into control (n = 6) and TFP treatment (n = 6) groups. TFP was dissolved in DMSO (5%) + saline (95%) and administered via intraperitoneal injection at 5 mg/kg once daily for 21 days. Tumor volume was measured every 3 days (volume = length × width² / 2), and mouse body weight was recorded weekly. At the end of treatment, mice were euthanized, tumors were excised for weight measurement, immunohistochemistry (NUPR1, p-MLKL), and Western blot analysis [1]

Absorption, Distribution and Excretion
Phenothiazine drugs have an extremely long final residence time in the body. /Phenothiazines/

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Metabolism/Metabolites
Hepatic metabolism.
Prolonged administration of piperazine-substituted phenothiazine drugs to rats resulted in the presence of drug metabolites in their tissues. These metabolites are substituted ethylenediamines formed by the cleavage of the piperazine ring through multiple oxidative dealkylation reactions. ...2-Trifluoromethyl analogues are generated from trifluoperazine in a similar manner.
In vivo degradation of the trifluoperazine piperazine ring leads to the formation of γ-(phenothiazinyl-10)-propylamine and its ring-substituted analogues CF3- and Cl-. The sulfoxides of these metabolites have been identified as biotransformation products in rat (chronic) urine.
Hepatic metabolism.
Half-life: 10-20 hours

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Biological half-life
10-20 hours

Toxicity Summary
Trifluoperazine blocks postsynaptic dopamine D1 and D2 receptors in the mesolimbic system of the brain; it inhibits the release of hypothalamic and pituitary hormones and is thought to suppress the reticular activating system, thereby affecting basal metabolism, body temperature, arousal, vasomotor activity, and vomiting. Hepatotoxicity
A higher incidence of liver dysfunction has been reported in patients taking phenothiazines long-term, but elevations of liver function indicators exceeding three times the upper limit of normal are uncommon. Transaminase abnormalities are usually mild, asymptomatic, and transient, and can be reversed with continued use. There have been reports of rare, clinically significant acute liver injury caused by trifluoperazine, similar to that caused by chlorpromazine. Jaundice usually appears within 1 to 4 weeks, and serum enzyme elevations are typically cholestatic or mixed. Some cases have experienced immune allergic reactions (rash, fever, and eosinophilia), but these are mild and self-limiting; autoantibodies are rare.
Probability Score: D (Possibly a rare cause of clinically significant liver damage).

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Effects during Pregnancy and Lactation
◉ Overview of Use During Lactation
Limited information suggests that maternal administration of up to 10 mg of trifluoperazine daily does not affect breastfed infants. Very limited long-term follow-up data suggest no adverse developmental effects when used alone with other phenothiazines. The safety rating system recommends against the use of trifluoperazine during lactation. Due to the scarcity of reported experience with trifluoperazine use during lactation, other antipsychotic medications may be preferable, especially in breastfed newborns or preterm infants.
◉ Effects on Breastfed Infants
Two mothers received oral trifluoperazine at 5 mg/day and 10 mg/day, respectively, and began breastfeeding their infants at 1 week and 8 weeks of age, respectively. Mental and motor development was assessed at various time points before the infants reached 30 months of age, and the results showed normal development.
One mother was breastfeeding from birth while taking trifluoperazine 10 mg/day, clonazepam 0.25 mg/day, and valproic acid 500 mg/day. The mother did not report any adverse reactions in the infant (follow-up time not specified).
One mother started taking trifluoperazine (dosage not specified) 2 months postpartum while breastfeeding. She also started taking olanzapine 10 mg/day, paroxetine, and procyclophosphamide (dosage not specified). The infant did not experience any adverse reactions.
◉ Effects on Lactation and Breast Milk
Phenothiazines can cause galactorrhea in 26% to 40% of female patients. Hyperprolactinemia appears to be the cause of galactorrhea. Hyperprolactinemia is caused by the drug blocking the action of dopamine in the tuberous-infundibular pathway.

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Drug Interactions
When probucol is used in combination with phenothiazines, the cumulative QT interval prolongation may increase the risk of ventricular tachycardia.
Phenothiazines
Phenothiazines may produce additive photosensitizing effects when used in combination with other photosensitizing drugs. Furthermore, concomitant use of systemic methoxsalen, trisalicyline, or tetracyclines with phenothiazines may exacerbate intraocular photochemical damage to the choroid, retina, or lens.
Pre-use of phenothiazines may reduce the pressor effect of phenylephrine and shorten its duration of action.
In addition to increasing central nervous system and respiratory depression, concomitant use of opioid (narcotic) analgesics/phenothiazines may increase orthostatic hypotension and the risk of severe constipation, which may lead to paralytic ileus and/or urinary retention.
/Phenothiazines/
For more complete data on interactions of trifluoperazines (29 in total), please visit the HSDB record page.

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Non-human toxicity values
Mouse intraperitoneal LD50 175 mg/kg

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In vitro toxicity: At concentrations up to 10 μM of TFP, there was no significant cytotoxicity to normal human pancreatic ductal epithelial cells (HPDE) or MDCK cells (cell viability >80% vs. control group) [1,5]
-In vivo toxicity: Mice treated with TFP (5 mg/kg/day, intraperitoneal injection, for 21 days) did not show significant weight loss, lethargy, or organ damage. Serum biochemical analysis (ALT, AST, BUN, creatinine) and histological examination of liver, kidney and heart tissues showed no abnormalities [1]
- Clinical toxicity:TFP may cause extrapyramidal symptoms (dystonia, Parkinson's syndrome) at doses used to treat schizophrenia, which can be reversed by anticholinergic drugs. It has minimal effect on human cardiovascular parameters (heart rate, blood pressure) [3,4]
- Plasma protein binding rate:TFP has a plasma protein binding rate of up to about 90-95% in human plasma (data from literature review) [4]

[1]. Ligand-based design identifies a potent NUPR1 inhibitor exerting anticancer activity via necroptosis. J Clin Invest. 2019;129(6):2500-2513. Published 2019 Mar 28.

[2]. Huerta-Bahena J, Villalobos-Molina R, García-Sáinz JA. Trifluoperazine and chlorpromazine antagonize alpha 1- but not alpha2- adrenergic effects. Mol Pharmacol. 1983;23(1):67-70.

[3]. Marques LO, Lima MS, Soares BG. Trifluoperazine for schizophrenia. Cochrane Database Syst Rev. 2004;2004(1):CD003545.

[4]. Howland RH. Trifluoperazine: A Sprightly Old Drug. J Psychosoc Nurs Ment Health Serv. 2016;54(1):20-22.

[5]. Influence of trifluoperazine on the late stage of influenza virus infection in MDCK cells. Antiviral Res. 1991;15(2):149-160.

Trifluoperazine belongs to the phenothiazine class of compounds. Its structural characteristic is the introduction of a trifluoromethyl substituent at the 2-position of the phenothiazine and a 3-(4-methylpiperazin-1-yl)propyl substituent at the N-10 position. It possesses various pharmacological activities, including dopamine antagonist, antiemetic, EC 1.8.1.12 (trypanositol disulfide reductase) inhibitor, EC 5.3.3.5 (cholesterol delta-isomerase) inhibitor, calmodulin antagonist, and phenothiazine antipsychotic. It is an N-alkylpiperazine, N-methylpiperazine compound, belonging to the phenothiazine class, and is also an organofluorine compound. It is a phenothiazine drug with effects similar to chlorpromazine. It is used as an antipsychotic and antiemetic. Trifluoperazine is a phenothiazine drug. Trifluoperazine is a phenothiazine antipsychotic drug, currently no longer commonly used in clinical practice. Trifluoperazine is a rare cause of clinically significant acute cholestatic liver injury. It has been reported to have been found in Crotalaria pallida, with supporting data. Trifluoperazine is a phenothiazine derivative and a dopamine antagonist with antipsychotic and antiemetic activities. It exerts its antipsychotic effect by blocking central dopamine receptors, thus preventing symptoms such as delusions and hallucinations caused by dopamine overdose. It also acts as a calmodulin inhibitor, leading to increased cytoplasmic calcium ion concentration. Trifluoperazine is only found in individuals who have previously used or taken the drug. It is a phenothiazine drug with a mechanism of action similar to chlorpromazine. It is used as an antipsychotic and antiemetic. [PubChem] Trifluoperazine blocks postsynaptic mesolimbic dopamine D1 and D2 receptors in the brain; it inhibits the release of hypothalamic and pituitary hormones and is thought to inhibit the reticular activating system, thereby affecting basal metabolism, body temperature, arousal, vasomotor activity, and vomiting. A phenothiazine drug with a mechanism of action similar to chlorpromazine. It is used as an antipsychotic and antiemetic. See also: phenothiazines (subclass); trifluoperazine hydrochloride (salt form); trifluoperazine dimaleate (its active ingredient). Indications: Used to treat anxiety disorders, anxiety-induced depression, and agitation. Mechanism of Action: Trifluoperazine blocks postsynaptic mesolimbic dopamine D1 and D2 receptors in the brain; it inhibits the release of hypothalamic and pituitary hormones and is thought to inhibit the reticular activating system, thereby affecting basal metabolism, body temperature, arousal, vasomotor activity, and vomiting.
...Phenothiazines block dopamine receptors, increasing dopamine turnover in the striatum. This increased turnover is thought to be a result of neural feedback mechanisms. ...Dopaminergic neurons have been identified in the substantia nigra and ventral tegmentum. Spontaneous firing of these cells increases... /Phenothiazines/

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Therapeutic Uses
Antiemetics; Antipsychotics, phenothiazines; Dopamine antagonists
In the treatment of acute psychosis, the dose of antipsychotics is increased in the first few days to control symptoms. Then, in the following weeks, the dose is adjusted according to the patient's condition. /Phenothiazines/
Antipsychotics have achieved some success in the treatment of mania and depression. /Phenothiazines/
...Trifluoperazine... has been reported to be effective in children with severe autism, and differential diagnosis of these children from so-called minimal brain damage (MBD) syndrome is crucial.
For more (complete) data on the therapeutic uses of trifluoperazine (10 in total), please visit the HSDB records page.

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Drug Warnings
Phenothiazines should be used with extreme caution and are contraindicated in untreated epilepsy patients and patients who are withdrawing from central nervous system depressants (such as alcohol and barbiturates). Phenothiazines
…A small number of patients with angina have reported increased pain during treatment with trifluoperazine. Therefore, patients with angina should be closely monitored, and the drug should be discontinued immediately if adverse reactions occur. Hydrochloride drugs
…The potential adverse effects on fetal development from use during pregnancy have not been established. …Phenothiazines…should be used with extreme caution in patients with a history of glaucoma or benign prostatic hyperplasia. /Phenothiazines/
Most antipsychotic regimens should avoid routine use of anti-Parkinson's drugs. /Phenothiazines/
For more complete data on drug warnings for trifluoperazine (28 in total), please visit the HSDB records page.

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Pharmacodynamics
Trifluoperazine is a trifluoromethylphenothiazine derivative used to treat schizophrenia and other psychotic disorders. Trifluoperazine has not been proven effective in treating behavioral complications in patients with intellectual disabilities. Trifluoperazine hydrochloride (TFP; SKF5019) is a first-generation phenothiazine antipsychotic with anticancer and antiviral activity [1,4,5]. Its anticancer mechanism involves binding to NUPR1 (a stress-responsive nucleoprotein), inhibiting its transcriptional activity, and inducing necroptosis in cancer cells by activating the RIPK1-MLKL pathway [1]. As an antipsychotic, it exerts its therapeutic effect by antagonizing α1-adrenergic receptors and dopamine D2 receptors (its dopamine antagonistic effect has not been reported in the literature) [2,3,4]. It inhibits influenza virus replication by interfering with the transport and budding of the viral M1 protein on the plasma membrane [5]. It has received clinical approval. Used to treat schizophrenia and other psychotic disorders; has potential for treating pancreatic cancer, lung cancer, and influenza virus infection [1,3,4,5]

C21H24F3N3S.2HCL

480.42

479.117

440-17-5

Trifluoperazine;117-89-5;Trifluoperazine dimaleate;605-75-4

5566

White to off-white solid powder

506ºC at 760 mmHg

243 °C (dec.)(lit.)

259.8ºC

2.32E-10mmHg at 25°C

6.49

0

7

4

28

510

0

ZEWQUBUPAILYHI-UHFFFAOYSA-N

InChI=1S/C21H24F3N3S/c1-25-11-13-26(14-12-25)9-4-10-27-17-5-2-3-6-19(17)28-20-8-7-16(15-18(20)27)21(22,23)24/h2-3,5-8,15H,4,9-14H2,1H3

10-[3-(4-methylpiperazin-1-yl)propyl]-2-(trifluoromethyl)phenothiazine

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

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: ≥ 2.5 mg/mL (5.20 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 25.0 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: ≥ 2.5 mg/mL (5.20 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 25.0 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: ≥ 2.5 mg/mL (5.20 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 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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