Combination of AAV­mediated NUPR1 knockdown and trifluoperazine induces premature senescence in human lung adenocarcinoma A549 cells in nude mice.

Nuclear protein 1 (NUPR1)/p8, a transcriptional regulator, has the ability to facilitate lung cancer cell survival. Adeno­associated virus (AAV)­based vectors are efficient vehicles for gene transfer and expression. In this study, an AAV­mediated NUPR1 shRNA vector was constructed that effectively inhibited the expression of NUPR1 in a tumor xenograft model derived from lung adenocarcinoma A549 cells. Trifluoperazine (TFP), which is an antipsychotic drug, has the ability to bind to NUPR1 and mimic NUPR1 deficiency in cancer cells. It was also found that the combination of TFP and AAV­mediated NUPR1 shRNA delivery led to significant tumor growth inhibition in nude mice bearing human lung cancer xenografts. Moreover, AAV­mediated NUPR1 shRNA therapy induced premature senescence in vitro and in vivo. Collectively, the findings of this study suggest a putative role for the combination of AAV­NUPR1 shRNA and TFP in lung cancer therapy.

Trifluoperazine as an alternative strategy for the inhibition of tumor growth of colorectal cancer.

The development of cancer in patients with schizophrenia is affected by genetic and environmental factors and antipsychotic medication. Several studies found that schizophrenia was associated with decreased risk of some cancers, and the neuroleptic medication might help to reduce the risk of colorectal cancer (CRC). Phenothiazine drugs including trifluoperazine (TFP) are widely used antipsychotic drugs and showed some antitumor effects, we here investigated the potential application of TFP in the treatment of colon cancer. A series doses of TFP were treated to the colon cancer cell line HCT116 and the inhibitory concentration (IC50 ) of TFP for HCT116 was determined by cell counting kit-8. The results indicated that the treatment of TFP impaired the cell vitality of HCT116 in a dose- and time-dependent manner. Meanwhile, the Edu assay demonstrated that the proliferation was also inhibited by TFP, which was accompanied with the induction of apoptosis and autophagy. The expression of CCNE1, CDK4, and antiapoptosis factor BCL-2 was downregulated but the proapoptosis factor BAX was upregulated. The autophagy inhibitor chloroquine could significantly reverse the TFP-induced apoptosis. Moreover, the ability of migration and invasion of HCT116 was found to be suppressed by TFP, which was associated with the inhibition of epithelial-mesenchymal transition (EMT). The function of TFP in vivo was further confirmed. The results showed that the administration of TFP remarkably abrogated the tumor growth with decreased tumor volume and proliferation index Ki-67 level in tumor tissues. The EMT phenotype was also confirmed to be inhibited by TFP in vivo, suggesting the promising antitumor effects of TFP in CRC.

Photochemical and Pharmacokinetic Characterization of Orally Administered Chemicals to Evaluate Phototoxic Risk.

This study aimed to verify the applicability of a proposed photosafety screening system based on a reactive oxygen species (ROS) assay and a cassette-dosing pharmacokinetic (PK) study to chemicals with wide structural diversity. The orally taken chemicals, erythromycin, gatifloxacin, 8-methoxypsoralen (MOP), pirfenidone (PFD), trifluoperazine (TFP), and voriconazole (VRZ), were selected as test compounds. The ROS assay was conducted to evaluate their photoreactivity, and all test compounds excluding erythromycin generated significant ROS under simulated sunlight exposure. According to the ROS data, TFP had potent photoreactivity, and the photoreactivity of 4 other compounds was judged to be moderate. Regarding the oral cassette-dosing PK test in rats, the skin deposition of MOP, PFD, and VRZ was relatively high, and gatifloxacin and TFP exhibited moderate skin deposition properties. Based on the ROS and PK data of test compounds, PFD and TFP were judged to be potent phototoxic compounds, and MOP and VRZ were deduced to have phototoxic risk. The predicted phototoxic risk of test compounds by proposed screening was mostly in agreement with observed in vivo phototoxicity in the rat skin. The proposed screening system could provide reliable photosafety information on orally administered compounds with wide structural diversity.

A chemical screen identifies trifluoperazine as an inhibitor of glioblastoma growth.

Glioblastoma (GBM) is regarded as the most common malignant brain tumor but treatment options are limited. Thus, there is an unmet clinical need for compounds and corresponding targets that could inhibit GBM growth. We screened a library of 80 dopaminergic ligands with the aim of identifying compounds capable of inhibiting GBM cell line proliferation and survival. Out of 45 active compounds, 8 were further validated. We found that the dopamine receptor D2 antagonist trifluoperazine 2HCl inhibits growth and proliferation of GBM cells in a dose dependent manner. Trifluoperazine's inhibition of GBM cells is cell line dependent and correlates with variations in dopamine receptor expression profile. We conclude that components of the dopamine receptor signaling pathways are potential targets for pharmacological interventions of GBM growth.

Trifluoperazine, a novel autophagy inhibitor, increases radiosensitivity in glioblastoma by impairing homologous recombination.

BACKGROUND: Resistance to adjuvant radiotherapy is a major cause of treatment failure in patients with glioblastoma (GBM). Autophagy inhibitors have been shown to enhance the efficacy of radiotherapy for certain solid tumors. However, current inhibitors do not penetrate the blood-brain-barrier (BBB). Here, we assessed the radiosensitivity effects of the antipsychotic drug trifluoperazine (TFP) on GBM in vitro and in vivo. METHODS: U251 and U87 GBM cell lines as well as GBM cells from a primary human biopsy (P3), were used in vitro and in vivo to evaluate the efficacy of TFP treatment. Viability and cytotoxicity was evaluated by CCK-8 and clonogenic formation assays. Molecular studies using immunohistochemistry, western blots, immunofluorescence and qPCR were used to gain mechanistic insight into the biological activity of TFP. Preclinical therapeutic efficacy was evaluated in orthotopic xenograft mouse models. RESULTS: IC50 values of U251, U87 and P3 cells treated with TFP were 16, 15 and 15.5 µM, respectively. TFP increased the expression of LC3B-II and p62, indicating a potential disruption of autophagy flux. These results were further substantiated by a decreased Lysotracker Red uptake, indicating impaired acidification of the lysosomes. We show that TFP and radiation had an additive effect when combined. This effect was in part due to impaired TFP-induced homologous recombination. Mechanistically we show that down-regulation of cathepsin L might explain the radiosensitivity effect of TFP. Finally, combining TFP and radiation resulted in a significant antitumor effect in orthotopic GBM xenograft models. CONCLUSIONS: This study provides a strong rationale for further clinical studies exploring the combination therapy of TFP and radiation to treat GBM patients.

Different mechanisms of risperidone result in improved interpersonal trust, social engagement and cooperative behavior in patients with schizophrenia compared to trifluoperazine.

AIM: Atypical antipsychotic treatment (e.g. risperidone) has been found to improve social functioning more than standard antipsychotic treatment. However, it is unclear which specific social behaviors are implicated in this improvement. The current study employed an interactive puzzle game to examine how social behaviors contribute to the improvement of social functioning by comparing patients receiving risperidone with those receiving trifluoperazine. METHODS: Scores on the Positive and Negative Syndrome Scale, executive functioning, and social functioning were obtained from 24 patients with schizophrenia receiving either risperidone (n = 12) or trifluoperazine (n = 12), before their social behavior was measured in the interactive Tangrams Game. Immediately after the Tangrams Game, participants filled in two questionnaires measuring their interpersonal trust and rejection toward their game partner. RESULTS: Patients receiving risperidone showed more social engagement, cooperative behavior and interpersonal trust toward their game partners than those receiving trifluoperazine. Additional multivariate analysis of variance revealed that lower affiliative behavior was a function of positive symptoms; interpersonal trust had an impact on social engagement but executive functioning did not explain lower interpersonal trust or social disengagement. CONCLUSION: Improvement of social competence by risperidone might be related to the enhancement of both social behaviors and interpersonal trust as well as better symptom resolution.

Combinatorial therapy with tamoxifen and trifluoperazine effectively inhibits malignant peripheral nerve sheath tumor growth by targeting complementary signaling cascades.

Chemotherapeutic agents effective against malignant peripheral nerve sheath tumors (MPNSTs) are urgently needed. We recently found that tamoxifen potently impedes xenograft growth. In vitro, tamoxifen inhibits MPNST proliferation and survival in an estrogen receptor-independent manner; these effects are phenocopied by the calmodulin inhibitor trifluoperazine. The present study was performed to establish the mechanism of action of tamoxifen in vivo and optimize its therapeutic effectiveness. To determine if tamoxifen has estrogen receptor-dependent effects in vivo, we grafted MPNST cells in castrated and ovariectomized mice; xenograft growth was unaffected by reductions in sex hormones. To establish whether tamoxifen and trifluoperazine additively or synergistically impede MPNST growth, mice xenografted with neurofibromatosis type 1-associated or sporadic MPNST cells were treated with tamoxifen, trifluoperazine, or both drugs for 30 days. Both monotherapies inhibited graft growth by 50%, whereas combinatorial treatment maximally reduced graft mass by 90% and enhanced decreases in proliferation and survival. Kinomic analyses showed that tamoxifen and trifluoperazine have both shared and distinct targets in MPNSTs. In addition, trifluoperazine prevented tamoxifen-induced increases in serum/glucocorticoid regulated kinase 1, a protein linked to tamoxifen resistance. These findings suggest that combinatorial therapy with tamoxifen and trifluoperazine is effective against MPNSTs because these agents target complementary pathways that are essential for MPNST pathogenesis.

Delusional disorder-somatic type (or body dysmorphic disorder) and schizophrenia: a case report.

With regard to delusional disorder-somatic subtype there may be a relationship with body dysmorphic disorder. There are reports that some delusional disorders can evolve to become schizophrenia. Similarly, the treatment of such disorders with antipsychotics has been documented. This report describes a case of delusional disorder - somatic type - preceding a psychotic episode and its successful treatment with an antipsychotic drug, thus contributing to what has been documented on the subject.

Inhibition of nitric oxide-activated guanylyl cyclase by calmodulin antagonists.

BACKGROUND AND PURPOSE: Nitric oxide (NO) controls numerous physiological processes by activation of its receptor, guanylyl cyclase (sGC), leading to the accumulation of 3'-5' cyclic guanosine monophosphate (cGMP). Ca(2+)-calmodulin (CaM) regulates both NO synthesis by NO synthase and cGMP hydrolysis by phosphodiesterase-1. We report that, unexpectedly, the CaM antagonists, calmidazolium, phenoxybenzamine and trifluoperazine, also inhibited cGMP accumulation in cerebellar cells evoked by an exogenous NO donor, with IC(50) values of 11, 80 and 180 microM respectively. Here we sought to elucidate the underlying mechanism(s). EXPERIMENTAL APPROACH: We used cerebellar cell suspensions to determine the influence of CaM antagonists on all steps of the NO-cGMP pathway. Homogenized tissue and purified enzyme were used to test effects of calmidazolium on sGC activity. KEY RESULTS: Inhibition of cGMP accumulation in the cells did not depend on changes in intracellular Ca(2+) concentration. Degradation of cGMP and inactivation of NO were both inhibited by the CaM antagonists, ruling out increased loss of cGMP or NO as explanations. Instead, calmidazolium directly inhibited purified sGC (IC(50)= 10 microM). The inhibition was not in competition with NO, nor did it arise from displacement of the haem moiety from sGC. Calmidazolium decreased enzyme V(max) and K(m), indicating that it acts in an uncompetitive manner. CONCLUSIONS AND IMPLICATIONS: The disruption of every stage of NO signal transduction by common CaM antagonists, unrelated to CaM antagonism, cautions against their utility as pharmacological tools. More positively, the compounds exemplify a novel class of sGC inhibitors that, with improved selectivity, may be therapeutically valuable.

Trifluoperazine: a rynodine receptor agonist.

Trifluoperazine (TFP), a phenothiazine, is a commonly used antipsychotic drug whose therapeutic effects are attributed to its central anti-adrenergic and anti-dopaminergic actions. However, TFP is also a calmodulin (CaM) antagonist and alters the Ca(2+) binding properties of calsequestrin (CSQ). The CaM and CSQ proteins are known modulators of sarcoplasmic reticulum (SR) Ca(2+) release in ventricular myocytes. We explored TFP actions on cardiac SR Ca(2+) release in cells and single type-2 ryanodine receptor (RyR2) channel activity in bilayers. In intact and permeabilized ventricular myocytes, TFP produced an initial activation of RyR2-mediated SR Ca(2+) release and over time depleted SR Ca(2+) content. At the single channel level, TFP or nortryptiline (NRT; a tricyclic antidepressant also known to modify CSQ Ca(2+) binding) increased the open probability (Po) of CSQ-free channels with an EC(50) of 5.2 microM or 8.9 microM (respectively). This Po increase was due to elevated open event frequency at low drug concentrations while longer mean open events sustained Po at higher drug concentrations. Activation of RyR2 by TFP occurred in the presence or absence of CaM. TFP may also inhibit SR Ca uptake as well as increase RyR2 opening. Our results suggest TFP and NRT can alter RyR2 function by interacting with the channel protein directly, independent of its actions on CSQ or CaM. This direct action may contribute to the clinical adverse cardiac side effects associated with these drugs.

Consideration of use of phenothiazines in particular trifluorperazine for epidermal growth factor receptor associated cancers.

Papers from a generation ago suggested that phenothiazines--in particular trifluorperazine (Stelazine) a medicinal approved by the FDA and still commonly used for schizophrenia--downregulate the epidermal growth factor receptor. As numerous cancers--e.g., colon cancer, breast cancer, pancreatic cancer and glioblastoma--are dependent on signaling via this receptor, we here suggest that phenothiazines such as trifluorperazine be considered for use in epidermal growth factor receptor associated cancers.

Topical verapamil HCl, topical trifluoperazine, and topical magnesium sulfate for the treatment of Peyronie's disease--a placebo-controlled pilot study.

INTRODUCTION: Transdermal and intralesional verapamil has been reported to be useful in the treatment of Peyronie's Disease. This study evaluates a topically applied calcium channel blocker (verapamil hydrochloride 15% gel), a topically applied calmodulin blocker (trifluoperazine), and a topically applied weak calcium channel blocker (magnesium sulfate), each incorporated in a transdermal vehicle. AIM: This pilot study was conducted to assess the efficacy of a 15% verapamil gel applied topically to the penile shaft twice daily for the treatment of Peyronie's Disease. MAIN OUTCOME MEASURE: To assess improvement in curvature, plaque size, resolution of painful erections, and improvement in erection quality. METHODS: Two simultaneous, three armed, double blinded, placebo-controlled studies were conducted. After randomization into one of four groups, patients were treated for 3 months. At the end of 3 months' treatment using blinded drug, each patient was treated with open label topical verapamil for 6 months. The studies were completed after each patient had been treated and evaluated for 9 months after randomization. RESULTS: Fifty-seven patients were randomized. In total, 94.4% of patients treated for 9 months with topical verapamil experienced improvement in curvature with an average percent curvature change of 61.1% compared with 43.6% curvature improvement at 3 months. At 9 months the average percent plaque change was 84.7% compared with 55% at 3 months. Pain resolution at 9 months was 100% compared with 87.5% at 3 months. Patient perception of erection quality also increased at 9 months to 81.8% compared with 72.7% at 3 months. CONCLUSIONS: Topical verapamil gel proved effective in eliminating pain on erection, decreasing the size of plaque, decreasing curvature, and improving erection quality in patients with Peyronie's Disease. Treatment results improved significantly after 9 months' treatment as compared with 3 months' treatment.

[Prescription of traditional neuroleptics in the remission period for schizophrenic patients with excess of body mass caused by atypical antipsychotics].

A sample included 61 patients, 53 men and 8 women, with ICD-10 episodic schizophrenia in the remission after treatment with atypical neuroleptics (risperidon, olanzapine, clozapine). All patients were featured by therapeutically caused excess of body mass (obesity of different degrees) that hampered the further treatment. In 31 cases (the main group) atypical neuroleptics were substituted for traditional drugs that exerted lesser influence on body mass. Haloperidol (mean dosage 4,1 mg daily) was administered to 17 patients and trifluoperazine (mean dosage 7,1 mg daily) to 14 patients. Other 30 patients (a control group) continued to receive atypical neuroleptics. Between group differences of patient's mental and somatic state were assessed using quantitative scales. It was shown that the substitution of atypical neuroleptics for traditional neuroleptic drugs allowed to stop further body mass gain and even decreased it without significant influence on psychopathological symptoms and other side-effects in patients with excess of body mass.

Investigations of calsequestrin as a target for anthracyclines: comparison of functional effects of daunorubicin, daunorubicinol, and trifluoperazine.

Anthracycline therapy is associated with a life-threatening but poorly understood cardiotoxicity. Effects of treatment are consistent with drug-induced disruption of cardiac sarcoplasmic reticulum (SR) calcium homeostasis, including inhibition of calcium release by anthracyclines. This effect, which depends on luminal SR calcium concentration, is hypothesized to involve interactions of anthracyclines with the calcium binding protein calsequestrin (CSQ). This study was designed to test the hypothesis that an interaction between CSQ and anthracyclines could be related to alterations in SR calcium release and cardiac function. The effects of the anthracycline, daunorubicin, and its metabolite daunorubicinol were compared with those of a known CSQ inhibitor, trifluoperazine (TFP). Protein fluorescence quenching studies demonstrated that TFP, daunorubicin, and daunorubicinol bind to CSQ with apparent binding affinities in the low micromolar range. The presence of calcium decreases the drug-dependent fluorescence quenching, probably because of calcium-induced CSQ conformational changes. TFP also inhibited SR calcium release. Although the TFP IC50 value is somewhat larger than for anthracyclines, the TFP effect is also dependent on luminal SR calcium concentration. In a muscle preparation, micromolar TFP decreased cardiac contractility in a manner that implicates the involvement of SR calcium and resembles the effects of anthracyclines. These data are consistent with a mechanism in which TFP or anthracyclines impair SR calcium release and cardiac function through a mechanism involving disruption of CSQ function. Such a mechanism may contribute to anthracycline cardiotoxicity.

The role of calmodulin in the regulation of osteoclastogenesis.

Calmodulin plays an important role in regulating the function of mature osteoclasts. However, its role in osteoclastogenesis has not been investigated. In the present study, we examined the role of calmodulin in osteoclastogenesis using in vivo and in vitro systems. Calmodulin antagonists, trifluoperazine (TFP), W7, and tamoxifen, dose-dependently inhibited osteoclast formation, which occurred only in the last 24 h of a 4-d osteoclastogenesis culture using mouse bone marrow macrophages. Inhibitory effects were quantitated by measuring tartrate-resistant acid phosphatase activity and counting osteoclast numbers. In contrast, bis indolylmaleimide, a protein kinase C inhibitor, showed no such inhibitory effect even when applied at a concentration that was 10-fold greater than its IC50. Overexpressing calmodulin by recombinant retrovirus reversed the inhibitory effect of TFP on osteoclast-like differentiation in RAW264.7 cells. Furthermore, administration of TFP to mice was as effective as estrogen in abolishing the ovariectomy-induced increment of osteoclastogenesis as determined by quantitative assessment of tartrate-resistant acid phosphatase activity in tibias, which led to the recovery of the ovariectomy-induced decrement in trabecular bone volume. To investigate potential cellular and molecular mechanisms by which calmodulin antagonists inhibit osteoclastogenesis, Z-VAD-FMK, a broad caspase inhibitor, failed to block the inhibitory effect of TFP on mouse osteoclast formation, indicating that apoptosis is not the underlying mechanism. Pretreatment of RAW264.7 cells with different concentrations of TFP dose-dependently inhibited receptor activator of nuclear factor kappaB ligand-stimulated phosphorylation of c-Jun N-terminal kinase and inhibitory kappaBalpha but not that of p38. Taken together, our data indicate that calmodulin mediates osteoclast differentiation, possibly via modulating specific receptor activator of NF-kappaB-signaling pathways.

Successful treatment of a complicated case of neuroleptic malignant syndrome.

Neuroleptic malignant syndrome (NMS) is a life-threatening reaction often related to neuroleptic drugs, characterized by rigidity, hyperthermia, altered consciousness, and fluctuating blood pressure. We present a case of NMS that followed a doubled oral dose of a drug compound: tranylcypromine sulfate, a monoamine oxidase inhibitor, and trifluoperazine (neuroleptic). The case was complicated by rhabdomyolisis and disseminated intravascular coagulation. It was treated successfully with dantrolene sodium and generous fluid therapy without using neuromuscular blocking agents or dopamine agonists.

Effect of calmodulin antagonists on phospholipase D activity in SH-SY5Y cells.

The aim of this study was to investigate the involvement of calmodulin in phospholipase D activation in SH-SY5Y cells. Cells prelabelled with [3H]-palmitic acid were incubated with calmodulin antagonists and/or other compounds. Phosphatidylethanol, a specific marker for phospholipase D activity, and phosphatidic acid were analysed. The calmodulin antagonists, calmidazolium and trifluoperazine, induced an extensive increase in phosphatidylethanol formation, and thus increased basal phospholipase D activity, in a dose- and time-dependent manner. The effect of calmidazolium on carbachol-induced activation of muscarinic receptors was also studied. Calmidazolium did not significantly affect the amount of phosphatidylethanol formed following carbachol addition. However, taking into account the increase in basal activity observed after calmidazolium addition, calmidazolium probably inhibits the muscarinic receptor-induced phospholipase D activation. In addition to phosphatidylethanol, basal phosphatidic acid levels were also increased after calmidazolium and trifluoperazine addition. Incubation with calmidazolium (10 microM) for 10 min induced a two-fold increase in phosphatidic acid. The calmidazolium-induced increase in basal phospholipase D activity was not affected by the protein kinase inhibitors H7 and staurosporine. On the other hand tyrosine kinase inhibitors abolished the calmidazolium-induced activation of phospholipase D. Calmidazolium also induced tyrosine phosphorylation in parallel to the phospholipase D activation. In conclusion, our data indicate that calmodulin antagonists induce phospholipase D activity in SH-SY5Y cells via a tyrosine kinase dependent pathway. This may point to a negative control of phospholipase D by calmodulin although a calmodulin-independent mechanism cannot be excluded. Calmodulin antagonists may be useful tools to further elucidate the mechanisms of phospholipase D regulation.