Phenethyl Isothiocyanate Research Articles

Inhibitory Effect of Phenethyl Isothiocyanate on the Adhesion and Biofilm Formation of Staphylococcus aureus and Application on Beef

This study aimed to explore the mechanism by which phenethyl isothiocyanate (PEITC) inhibited the adhesion and biofilm formation of Staphylococcus aureus (S. aureus). PEITC exhibited antimicrobial efficacy against S. aureus, demonstrating a minimum inhibition concentration (MIC) of 1 mmol/L. PEITC exerted its antibacterial effect by disrupting cell membrane integrity, and it decreased total adenosine triphosphate (ATP) production after 1 and 4 h treatment. PEITC at 0.5 mmol/L increased the level of intracellular reactive oxygen species (ROS) by 26.39% compared to control. The mature biofilm of S. aureus was destroyed by 86.4% after treatment with PEITC for 24 h. Adhesion tests revealed that PEITC at 0.5 mmol/L reduced 44.51% of the S. aureus that adhered to NCM460 cells. Furthermore, at the genetic level, PEITC significantly downregulated the related genes by 31.26% to 97.04%, including agrB, agrD, isdA, ebh, luxS, fnbA, and icaR. Moreover, PEITC markedly inhibited S. aureus proliferation in beef preserved at temperatures of 25 and 4 °C, respectively. In summary, the present study suggests that PEITC effectively inhibits the adhesion and biofilm formation of S. aureus by affecting the relevant genes of S. aureus and holds promise for microbial management in meat products.

The Potential of Glucosinolates and Their Hydrolysis Products as Inhibitors of Cytokine Storms

A cytokine storm is an intense inflammatory response characterized by the overproduction of proinflammatory cytokines, resulting in tissue damage, and organ dysfunction. Cytokines play a crucial role in various conditions, such as coronavirus disease, in which the immune system becomes overactive and releases excessive levels of cytokines, including interleukins, tumor necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ). This anomalous response often leads to acute respiratory distress syndrome (ARDS), disseminated intravascular coagulation (DIC), and multiple organ injury (MOI). Glucosinolates are plant secondary metabolites predominantly found in Brassica vegetables, but are also present in other species, such as Moringa Adens and Carica papaya L. When catalyzed by the enzyme myrosinase, glucosinolates produce valuable products, including sulforaphane, phenethyl isothiocyanate, 6-(methylsulfinyl) hexyl isothiocyanate, erucin, goitrin, and moringin. These hydrolyzed products regulate proinflammatory cytokine production by inhibiting the nuclear factor kappa-light-chain-enhancer of activated B-cell (NF-κB) signaling pathway and stimulating the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway. This action can alleviate hyperinflammation in infected cells and modulate cytokine storms. In this review, we aimed to examine the potential role of glucosinolates in modulating cytokine storms and reducing inflammation in various conditions, such as coronavirus disease. Overall, we found that glucosinolates and their hydrolysis products can potentially attenuate cytokine production and the onset of cytokine storms in diseased cells. In summary, glucosinolates could be beneficial in regulating cytokine production and preventing complications related to cytokine storms.

PEITC Induces DNA Damage and Inhibits DNA Repair-Associated Proteins in Human Retinoblastoma Cells In Vitro.

Phenethyl isothiocyanate (PEITC), a natural product, exists in biological activities, including anticancer activity in many human cancer cells. No information shows that PEITC affects DNA damage in human retinoblastoma (RB) cells in vitro. In this study, the aim of experiments was to determine whether PEITC decreased total viable cell number or not by inducing protein expressions involved in DNA damage and repair in Y79 RB cells in vitro. Total cell viability was measured by PI exclusion assay, and PEITC reduced the total Y79 viable cell numbers in a dose-dependent manner. DNA condensation and DNA impairment were conducted by DAPI staining and comet assays, respectively, in Y79 cells. The findings show that PEITC induced DNA condensation dose-dependently based on the brighter fluorescence of cell nuclei stained by DAPI staining. PEITC-induced DNA damage showed a more extended DNA migration smears than that of the control, which was performed by a comet assay. Western blotting was performed to measure the protein expressions involved in DNA damage and repair, which showed that PEITC at 2.5-10 μM increased NRF2, HO-1, SOD (Mn), and catalase; however, it decreased SOD (Cu/Zn) except 10 μM PEITC treatment, and decreased glutathione, which were associated with oxidativestress. Furthermore, PEITC increased DNA-PK, MDC1, H2A.XpSer139, ATMpSer1981, p53, p53pSer15, PARP, HSP70, and HSP90, but decreased TOPIIα, TOPIIβ, and MDM2pSer166 that were associated with DNA damage and repair mechanism in Y79 cells. The examination from confocal laser microscopy shows that PEITC increased H2A.XpSer139 and p53pSer15, and decreased glutathione and TOPIIα in Y79 cells. In conclusion, the cytotoxic effects of PEITC on reducing the number of viable cells may be due to the induction of DNA damage and the alteration of DNA repair proteins in Y79 cells in vitro.

Effects of microencapsulated phenethyl isothiocyanate on gastrointestinal cancer cells and pathogenic bacteria

Gastrointestinal cancers remain a global health burden, demanding more effective prevention and treatments. Phenethyl isothiocyanate (PEITC), a compound derived from cruciferous vegetables, stands out as a promising nutraceutical agent due to its chemopreventive and therapeutic properties. However, its therapeutic translation remains limited mainly due to its poor water solubility and rapid metabolism. Herein, we encapsulated PEITC into biocompatible chitosan-based microparticles with an extra virgin olive oil core to improve its bioavailability and stability. Pure PEITC's biocompatibility and microencapsulated PEITC's stability and antibacterial activity were evaluated. The antibacterial activity analysis showed microencapsulated PEITC as a promising antibacterial agent against gastrointestinal pathogenic bacteria (two Gram-positive and two Gram-negative). The impact of both pure and microencapsulated PEITC was assessed on gastrointestinal cancer cells (MKN45 gastric cancer and SW48 colon cancer cell lines). PEITC exhibited threshold or hormetic dose-dependent toxicity in colon fibroblasts and decreased gastric cancer cells' migration capacity, enhanced upon encapsulation into microparticles. In addition, microencapsulated PEITC induced downregulation of phosphorylated AKT, FAK, and ERK1/2 proteins, disrupting motility signaling pathways and tubulin expression. These findings suggest that the delivery of PEITC via chitosan-based microparticles holds promise as a nutraceutical delivery strategy against gastrointestinal disorders that predispose to cancer.

“All-in-one” metal polyphenol network nanocapsules integrated microneedle patches for lipophagy fueled ferroptosis-mediated multimodal therapy

Ferroptosis-mediated multimodal therapy has emerged as a promising strategy for tumor elimination, with lipid peroxide (LPO) playing a pivotal role. However, the therapeutic efficiency is limited due to insufficient intracellular levels of free fatty acids (FFA), which severely hinder the production of LPO. To address this limitation, we proposed a lipophagy strategy aimed at degrading lipid droplets (LDs) to release FFA, serving as the essential “fuel” for LPO production. In this study, the lipophagy inducer epigallocatechin gallate (EGCG) was self-assembled with reactive oxygen species (ROS)-producer phenethyl isothiocyanate (PEITC) mediated by Fe2+ to form EFP nanocapsules, which were further integrated into microneedle patches to form a “all-in-one” EFP@MNs. The metal-polyphenol network structure of EFP endow it with photothermal therapy capacity. Upon insertion into tumors, the released EFP nanocapsules were demonstrated to induce lipophagy through metabolic disturbance, thereby promoting LPO production and facilitating ferroptosis. When combined with photothermal therapy, this approach significantly remolded the tumor immune microenvironment by driving tumor-associated macrophages toward M1 phenotype and enhancing dendritic cell maturation. Encouragingly, in conjunction with αPD-L1 treatment, the proposed EFP@MNs exhibited remarkable efficacy in tumor ablation. Our study presents a versatile framework for utilizing microneedle patches to power ferroptosis-mediated multimodal therapy.

Naturally-derived phenethyl isothiocyanate modulates apoptotic induction through regulation of the intrinsic cascade and resulting apoptosome formation in human malignant melanoma cells

Malignant melanoma is the most aggressive type of skin cancer with increasing incidence rates worldwide. On the other hand, watercress is a rich source of phenethyl isothiocyanate (PEITC), among others, which has been widely investigated for its anticancer properties against various cancers. In the present study, we evaluated the role of a watercress extract in modulating apoptotic induction in an in vitro model of human malignant melanoma consisting of melanoma (A375, COLO-679, COLO-800), non-melanoma epidermoid carcinoma (A431) and immortalized, non-tumorigenic keratinocyte (HaCaT) cells. Moreover, the chemical composition of the watercress extract was characterized through UPLC MS/MS and other analytical methodologies. In addition, cytotoxicity was assessed by the alamar blue assay whereas apoptosis was determined, initially, by a multiplex activity assay kit (measuring levels of activated caspases −3, −8 and −9) as well as by qRT-PCR for the identification of major genes regulating apoptosis. In addition, protein expression levels were evaluated by western immunoblotting. Our data indicate that the extract contains various phytochemicals (e.g. phenolics, flavonoids, pigments, etc.) while isothiocyanates (ITCs; especially PEITC) were the most abundant. In addition, the extract was shown to exert a significant time- and dose-dependent cytotoxicity against all malignant melanoma cell lines while non-melanoma and non-tumorigenic cells exhibited significant resistance. Finally, expression profiling revealed a number of genes (and corresponding proteins) being implicated in regulating apoptotic induction through activation of the intrinsic apoptotic cascade. Overall, our data indicate the potential of PEITC as a promising anti-cancer agent in the clinical management of human malignant melanoma.

Covalent isothiocyanate inhibitors of macrophage migration inhibitory factor as potential colorectal cancer treatments.

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine with roles in innate and adaptive human immune responses, as well as inflammation. MIF exerts its biological activity by binding to the cell surface receptor CD74 as well as intracellular signalling proteins. MIF also possesses keto-enol tautomerase activity. Inhibition of the tautomerase activity has been associated with loss of biological activity of MIF and a potential anticancer target. Isothiocyanates (ITCs) are a class of compounds present in cruciferous vegetables that inhibit the MIF tautomerase activity via covalent modification of the N-terminal proline. A range of substituted ITCs featuring benzyl, phenethyl and phenyl propyl isothiocyanates were designed, synthesised and tested to determine any structure activity relationship for inhibiting MIF. Crystal structures of covalent compounds 8 and 9 in complex with rhMIF revealed key hydrogen bonding and edge-to-face π stacking interactions. Compound 9 and 11 with sub micromolar activity were tested in the NCI60 cancer cell lines panel. Both compounds showed tissue-specific reduced growth in colon and renal cancer cell lines, while one of these showed potent, dose-dependent inhibition of growth against all seven colon cancer cell lines (GI50 < 2.5 µM) and all eight renal cancer cell lines (GI50 < 2.2 µM).

MAFF mediates PEITC-induced enhancement of sensitivity to carboplatin in ovarian cancer cell lines via activating ZNF711 transcription in vivo and invitro

Enhanced drug resistance poses a significant challenge in treating ovarian cancer (OC). Phenylethyl isothiocyanate (PEITC) is involved in drug resistance in OC, but the mechanism remains unclear. In this study, we investigated the molecular regulatory mechanism of carboplatin sensitivity in OC associated with PEITC, MAF BZIP Transcription Factor F (MAFF), and Zinc finger proteins (ZNF) 711. The carboplatin sensitivity was significantly increased in OC cells after PEITC treatment. Knockdown of MAFF significantly enhanced the carboplatin sensitivity of OC cells, promoted apoptosis, inhibited colony-forming efficiency in vitro, and suppressed tumor growth in vivo. The binding site of MAFF to the ZNF711 promoter was predicted, and the knockdown of MAFF significantly increased the ZNF711 expression. Results of the dual luciferase assay and ChIP-PCR confirmed the binding of MAFF to the ZNF711 promoter. Immunofluorescence and CoIP results demonstrated the colocalization and the binding of MAFF and its interacting protein, BZIP Transcription Factor ATF-like 3 (BATF3). Similarly, we confirmed the binding of BATF3 to the ZNF711 promoter. Knockdown of BATF3 alone and simultaneous knockdown of BATF3 and MAFF showed similar regulatory effects on ZNF711 transcription and apoptosis. These suggested that the binding of MAFF to BATF3 inhibited ZNF711 transcription and reduced carboplatin sensitivity in OC.

Phenethyl isothiocyanate inhibits the carcinogenic properties of hepatocellular carcinoma Huh7.5.1 cells by activating MAPK/PI3K-Akt/p53 signaling pathways.

Hepatocellular carcinoma (HCC) is an aggressive malignancy with limited effective treatment options. Phenethyl isothiocyanate (PEITC) is a bioactive substance present primarily in the cruciferous vegetables. PEITC has exhibited anti-cancer properties in various cancers, including lung, bile duct, and prostate cancers. It has been demonstrated that PEITC can inhibit the proliferation, invasion, and metastasis of SK-Hep1 cells, while effectively inducing apoptosis and cell cycle arrest in HepG2 cells. However, knowledge of its anti-carcinogenic effects on Huh7.5.1 cells and its underlying mechanism remains elusive. In the present study, we aim to evaluate the anti-carcinogenic effects of PEITC on human HCC Huh7.5.1 cells. MTT assay and colony formation assay was performed to investigate the anti-proliferative effects of PEITC against Huh7.5.1 cells. The pro-apoptosis effects of PEITC were determined by Annexin V-FITC/PI double staining assay by flow cytometry (FCM), mitochondrial transmembrane potential (MMP) measurement, and Caspase-3 activity detection. A DAPI staining and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay was conducted to estimate the DNA damage in Huh7.5.1 cells induced by PEITC. Cell cycle progression was determined by FCM. Transwell invasion assay and wound healing migration assay were performed to investigate the impact of PEITC on the migration and invasion of Huh7.5.1 cells. In addition, transcriptome sequencing and gene set enrichment analysis (GSEA) were used to explore the potential molecular mechanisms of the inhibitory effects of PEITC on HCC. Quantitative real-time PCR (qRT-PCR) analysis was performed to verify the transcriptome data. MTT assay showed that treatment of Huh7.5.1 cells with PEITC resulted in a dose-dependent decrease in viability, and colony formation assay further confirmed its anti-proliferative effect. Furthermore, we found that PEITC could induce mitochondrial-related apoptotic responses, including a decrease of mitochondrial transmembrane potential, activation of Caspase-3 activity, and generation of intracellular reactive oxygen species. It was also observed that PEITC caused DNA damage and cell cycle arrest in the S-phase in Huh7.5.1 cells. In addition, the inhibitory effect of PEITC on the migration and invasion ability of Huh7.5.1 cells was assessed. Transcriptome sequencing analysis further suggested that PEITC could activate the typical MAPK, PI3K-Akt, and p53 signaling pathways, revealing the potential mechanism of PEITC in inhibiting the carcinogenic properties of Huh7.5.1 cells. PEITC exhibits anti-carcinogenic activities against human HCC Huh7.5.1 cells by activating MAPK/PI3K-Akt/p53 signaling pathways. Our results suggest that PEITC may be useful for the anti-HCC treatment.

Nutritional Sources and Anticancer Potential of Phenethyl Isothiocyanate: Molecular Mechanisms and Therapeutic Insights.

Phenethyl isothiocyanate (PEITC), a compound derived from cruciferous vegetables, has garnered attention for its anticancer properties. This review synthesizes existing research on PEITC, focusing on its mechanisms of action in combatting cancer. PEITC has been found to be effective against various cancer types, such as breast, prostate, lung, colon, and pancreatic cancers. Its anticancer activities are mediated through several mechanisms, including the induction of apoptosis (programmed cell death), inhibition of cell proliferation, suppression of angiogenesis (formation of new blood vessels that feed tumors), and reduction of metastasis (spread of cancer cells to new areas). PEITC targets crucial cellular signaling pathways involved in cancer progression, notably the Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-κB), Protein Kinase B (Akt), and Mitogen-Activated Protein Kinase (MAPK) pathways. These findings suggest PEITC's potential as a therapeutic agent against cancer. However, further research is necessary to determine the optimal dosage, understand its bioavailability, and assess potential side effects. This will be crucial for developing PEITC-based treatments that are both effective and safe for clinical use in cancer therapy.

Targeted anti-cancer therapy: Co-delivery of VEGF siRNA and Phenethyl isothiocyanate (PEITC) via cRGD-modified lipid nanoparticles for enhanced anti-angiogenic efficacy

Anti-tumor angiogenesis therapy, targeting the suppression of blood vessel growth in tumors, presents a potent approach in the battle against cancer. Traditional therapies have primarily concentrated on single-target techniques, with a specific emphasis on targeting the vascular endothelial growth factor, but have not reached ideal therapeutic efficacy. In response to this issue, our study introduced a novel nanoparticle system known as CS-siRNA/PEITC&L-cRGD NPs. These chitosan-based nanoparticles have been recognized for their excellent biocompatibility and ability to deliver genes. To enhance their targeted delivery capability, they were combined with a cyclic RGD peptide (cRGD). Targeted co-delivery of gene and chemotherapeutic agents was achieved through the use of a negatively charged lipid shell and cRGD, which possesses high affinity for integrin αvβ3 overexpressed in tumor cells and neovasculature. In this multifaceted approach, co-delivery of VEGF siRNA and phenethyl isothiocyanate (PEITC) was employed to target both tumor vascular endothelial cells and tumor cells simultaneously. The co-delivery of VEGF siRNA and PEITC could achieve precise silencing of VEGF, inhibit the accumulation of HIF-1α under hypoxic conditions, and induce apoptosis in tumor cells. In summary, we have successfully developed a nanoparticle delivery platform that utilizes a dual mechanism of action of anti-tumor angiogenesis and pro-tumor apoptosis, which provides a robust and potent strategy for the delivery of anti-cancer therapeutics.

Efficacy of outer membrane permeabilization in promoting aromatic isothiocyanates-mediated eradication of multidrug resistant Gram-negative bacteria and bacterial persisters.

Multidrug resistant (MDR) bacteria are recognized to be one of the most important problems in public health. The outer membrane permeability is a critical intrinsic mechanism of bacterial resistance. In addition, bacteria produce a small number of dormant persister cells causing multidrug tolerance that reduces antimicrobial efficacy. This study aimed to evaluate the inhibitory effects of the combination of aromatic isothiocyanates (ITCs) with membrane-active agents on bacterial persisters and MDR Gram-negative bacteria. Our study demonstrated that membrane-active agents, particularly ethylenediaminetetraacetic acid (EDTA) synergistically enhanced the inhibitory activity of aromatic benzyl ITC and phenethyl ITC against most Gram-negative bacteria strains with fractional inhibitory concentration index values ranging from 0.18 to 0.5 and 0.16 to 0.5, respectively, and contributed to an 8- to 64-fold minimal inhibitory concentration reduction compared with those of aromatic ITCs alone. The EDTA-aromatic ITCs combination effectively reduced the survival rates of tested bacteria and significantly eradicated bacterial persisters (p = 0.033 and 0.037, respectively). The growth kinetics analysis also supported the enhanced inhibitory effect of EDTA-aromatic ITCs combination against tested bacteria. Our results suggested an alternate treatment strategy against Gram-negative bacteria, promoting the entry of aromatic ITCs into bacterial cytoplasm to facilitate bacterial clearance and thus preventing the development of bacterial resistance.

Determination of Phenethyl Isothiocyanate, Erucin, Iberverin, and Erucin Nitrile Concentrations in Healthy and Pieris rapae-Infected Broccoli Tissues Using Gas Chromatography-Mass Spectrometry

Cruciferous vegetables (Brassicaceae family) are a rich source of phytochemicals, in particular glucosinolates (GLS) and their hydrolysis products, isothiocyanates and nitriles. These phytochemicals may act as chemosensors, attracting insects, such as Pieris rapae, and stimulating oviposition. There is a lack of information on the concentrations of isothiocyanates and nitriles when an insect affects a Brassicaceae plant. In the current study, some GLS hydrolysis products were determined in healthy and Pieris rapae-infected organic cultivated broccoli plants, as well as the infesting insects’ larvae, using gas chromatography-mass spectrometry (GC-MS). This study investigated the following phytochemicals: phenethyl isothiocyanate (PEITC), erucin (ER), 3-(methylthio)propyl isothiocyanate (3MIC), and 1-cyano-4-(methylthio)butane (5MITN). All these components were quantified in the aerial and underground parts of the plants and were found in high concentrations in the roots. Among the phytochemicals studied, 5MITN presented the highest concentration in all the broccoli samples but was especially high in the stalks of the infected plants. Moreover, the analysis of a sample of Pieris rapae larvae, fed from the hosted broccoli, revealed the presence of PEITC and ER. These findings indicate that the infestation of broccoli with Pieris rapae may affect the distribution of PEITC, ER, 3MIC, and 5MITN throughout the plant. An extension of our study to conventional cultivated broccoli showed that the roots are indeed rich in GLS hydrolysis products.

Naturally Derived Phenethyl Isothiocyanate Modulates Induction of Oxidative Stress via Its N-Acetylated Cysteine Conjugated form in Malignant Melanoma.

Phenethyl isothiocyanate (PEITC) is a secondary metabolic product yielded upon the hydrolysis of gluconasturtiin and it is highly accumulated in the flowers of watercress. The aim of the current study was to assess the role of a naturally derived PEITC-enriched extract in the induction of oxidative stress and to evaluate its anti-melanoma potency through the regulation of its metabolism with the concurrent production of the N-acetyl cysteine conjugated by-product. For this purpose, an in vitro melanoma model was utilized consisting of human primary (A375) cells as well as metastatic (COLO-679) malignant melanoma cells together with non-tumorigenic immortalized keratinocytes (HaCaT). Cytotoxicity was assessed via the Alamar Blue assay whereas the antioxidant/prooxidant activity of PEITC was determined via spectrophotometric assays. Finally, kinetic characterization of the end-product of PEITC metabolism was monitored via UPLC coupled to a tandem mass spectrometry (MS/MS). Our results indicate that although PhEF showed very minor antioxidant activity in a cell-free system, in a cell-based system, it can modulate the activity of key enzyme(s) involved in cellular antioxidant defense mechanism(s). In addition, we have shown that PhEF induces lipid and protein oxidation in a concentration-dependent manner, while its cytotoxicity is not only dependent on PEITC itself but also on its N-acetylated cysteine conjugated form.

Architecture and potential roles of a delta-class glutathione S-transferase in protecting honey bee from agrochemicals

The European honey bee, Apis mellifera, serves as the principle managed pollinator species globally. In recent decades, honey bee populations have been facing serious health threats from combined biotic and abiotic stressors, including diseases, limited nutrition, and agrochemical exposure. Understanding the molecular mechanisms underlying xenobiotic adaptation of A. mellifera is critical, considering its extensive exposure to phytochemicals and agrochemicals present in the environment. In this study, we conducted a comprehensive structural and functional characterization of AmGSTD1, a delta class glutathione S-transferase (GST), to unravel its roles in agrochemical detoxification and antioxidative stress responses. We determined the 3-dimensional (3D) structure of a honey bee GST using protein crystallography for the first time, providing new insights into its molecular structure. Our investigations revealed that AmGSTD1 metabolizes model substrates, including 1-chloro-2,4-dinitrobenzene (CDNB), p-nitrophenyl acetate (PNA), phenylethyl isothiocyanate (PEITC), propyl isothiocyanate (PITC), and the oxidation byproduct 4-hydroxynonenal (HNE). Moreover, we discovered that AmGSTD1 exhibits binding affinity with the fluorophore 8-Anilinonaphthalene-1-sulfonic acid (ANS), which can be inhibited with various herbicides, fungicides, insecticides, and their metabolites. These findings highlight the potential contribution of AmGSTD1 in safeguarding honey bee health against various agrochemicals, while also mitigating oxidative stress resulting from exposure to these substances.

Synergistic Anti-Cancer Potential of Phenethyl Isothiocyanate and Curcumin Induces Apoptosis and G2/M Cell Cycle Arrest in HER2-Positive Breast Cancer Cells

Synergistic Anti-Cancer Potential of Phenethyl Isothiocyanate and Curcumin Induces Apoptosis and G2/M Cell Cycle Arrest in HER2-Positive Breast Cancer Cells

Phenethyl isothiocyanate induces cytotoxicity and apoptosis of porcine kidney cells through Mitochondrial ROS-associated ERS pathway

Glucosinolates (GLS) in cruciferous vegetables are anti-nutritional factors. Excessive or long-term intake of GLS-containing feed is harmful to animal health and may cause kidney damage. Phenethyl isothiocyanate (PEITC) is a GLS. In this study, we investigated the inhibitory effect of PEITC on a porcine kidney (PK-15) cell line and explored the mechanism of PEITC-induced apoptosis. We found that PEITC could affect cell viability and induce cell apoptosis after incubating cells for 24 h. High concentrations of PEITC can induce intracellular ROS accumulation, resulting in impaired mitochondrial function (decreased MMP, decreased ATP) and DNA damage (increased 8-OHdG), cytochrome c in mitochondria is released into the cytoplasm and activates mitochondrial pathway apoptosis-related proteins (Bcl-2 family and caspase-9, -3). Meanwhile, PEITC could induce intracellular Ca2+ accumulation, disrupt ER homeostasis, and activate the expression levels of three ER-resident transmembrane proteins orchestrating the UPR (PERK, IRE-1α and ATF6) and ER-related proteins (GRP78 and CHOP), thereby activating ERS-pathway apoptosis-related proteins (caspase-12, -7). Our results showed that low concentration (2.5 μM) of PEITC had no damaging effect on cells. In comparison, a high concentration (10 μM) of PEITC could induce cell damage in porcine kidney cells and induce apoptosis in PK-15 cells via the Mitochondrial ROS-associated ERS pathway.

Phenethyl isothiocyanate inhibits metastasis potential of non-small cell lung cancer cells through FTO mediated TLE1 m6A modification.

N6-methyladenosine (m6A) modification is a prevalent RNA epigenetic modification, which plays a crucial role in tumor progression including metastasis. Isothiocyanates (ITCs) are natural compounds and inhibit the tumorigenesis of various cancers. Our previous studies show that ITCs inhibit the proliferation and metastasis of non-small cell lung cancer (NSCLC) cells, and have synergistic effects with chemotherapy drugs. In this study, we investigated the molecular mechanisms underlying the inhibitory effects of ITCs on cancer cell metastasis. We showed that phenethyl isothiocyanate (PEITC) dose-dependently inhibited the cell viability of both NSCLC cell lines H1299 and H226 with IC50 values of 17.6 and 15.2 μM, respectively. Furthermore, PEITC dose-dependently inhibited the invasion and migration of H1299 and H226 cells. We demonstrated that PEITC treatment dose-dependently increased m6A methylation levels and inhibited the expression of the m6A demethylase fat mass and obesity-associated protein (FTO) in H1299 and H226 cells. Knockdown of FTO significantly increased m6A methylation in H1299 and H226 cells, impaired their abilities of invasion and migration in vitro, and enhanced the inhibition of PEITC on tumor growth in vivo. Overexpression of FTO promoted the migration of NSCLC cells, and also mitigated the inhibitory effect of PEITC on migration of NSCLC cells. Furthermore, we found that FTO regulated the mRNA m6A modification of a transcriptional co-repressor Transducin-Like Enhancer of split-1 (TLE1) and further affected its stability and expression. TCGA database analysis revealed TLE1 was upregulated in NSCLC tissues compared to normal tissues, which might be correlated with the metastasis status. Moreover, we showed that PEITC suppressed the migration of NSCLC cells by inhibiting TLE1 expression and downstream Akt/NF-κB pathway. This study reveals a novel mechanism underlying ITC's inhibitory effect on metastasis of lung cancer cells, and provided valuable information for developing new therapeutics for lung cancer by targeting m6A methylation.

Phenethyl isothiocyanate and irinotecan synergistically induce cell apoptosis in colon cancer HCT 116 cells in vitro.

Irinotecan (IRI), an anticancer drug to treat colon cancer patients, causes cytotoxic effects on normal cells. Phenethyl isothiocyanate (PEITC), rich in common cruciferous plants, has anticancer activities (induction of cell apoptosis) in many human cancer cells, including colon cancer cells. However, the anticancer effects of IRI combined with PEITC on human colon cancer cells in vitro were unavailable. Herein, the aim of this study is to focus on the apoptotic effects of the combination of IRI and PEITC on human colon cancer HCT 116 cells in vitro. Propidium iodide (PI) exclusion and Annexin V/PI staining assays showed that IRI combined with PEITC decreased viable cell number and induced higher cell apoptosis than that of IRI or PEITC only in HCT 116 cells. Moreover, combined treatment induced higher levels of reactive oxygen species (ROS) and Ca2+ than that of IRI or PEITC only. Cells pre-treated with N-acetyl-l-cysteine (scavenger of ROS) and then treated with IRI, PEITC, or IRI combined with PEITC showed increased viable cell numbers than that of IRI or PEITC only. IRI combined with PEITC increased higher caspase-3, -8, and -9 activities than that of IRI or PEITC only by flow cytometer assay. IRI combined with PEITC induced higher levels of ER stress-, mitochondria-, and caspase-associated proteins than that of IRI or PEITC treatment only in HCT 116 cells. Based on these observations, PEITC potentiates IRI anticancer activity by promoting cell apoptosis in the human colon HCT 116 cells. Thus, PEITC may be a potential enhancer for IRI in humans as an anticolon cancer drug in the future.

Phenethyl isothiocyanate and dasatinib combination synergistically reduces hepatocellular carcinoma growth via cell cycle arrest and oxeiptosis.

Introduction: Hepatocellular carcinoma (HCC) is the most common type of liver cancer, which is among the most lethal tumours. Combination therapy exploits multiple drugs to target key pathways synergistically to reduce tumour growth. Isothiocyanates have been shown to possess anticancer potential and to complement the anticancer activity of other compounds. This study aimed to investigate the potential of phenethyl isothiocyanate (PEITC) to synergise with dasatinib, improving its anticancer potential in HCC. Methods: MTT, 3D spheroids and clonogenic assays were used to assess the combination anti-tumour effect in vitro, whereas a murine syngeneic model was employed to evaluate the combination efficacy in vivo. DCFDA staining was employed to evaluate the production of reactive oxygen species (ROS), while flow cytometry and Western blot assays were used to elucidate the molecular mechanism of the synergistic activiy. Results: PEITC and dasatinib combination exhibited a synergistic effect in vitro and in vivo. The combination induced DNA damage and oxidative stress through the production of ROS, which led to the formation of a premature CDK1/Cyclin B1 complex associated with induction of mitotic catastrophe. Furthermore, ROS activated oxeiptosis, a caspase-independent form of programmed cell death. Conclusion: PEITC showed to enhance dasatinib action in treating HCC with increased production of ROS that induced cell cycle arrest followed by mitotic catastrophe, and to induce oxeiptosis. These results highlight the role that ITCs may have in cancer therapy as a complement of clinically approved chemotherapeutic drugs.