IngredientsPhenethyl isothiocyanate (PEITC) is a naturally occurring isothiocyanate whose precursor, gluconasturtiin, is found in some cruciferous vegetables, especially watercress. Its systematic IUPAC name is 2-isothiocyanatoethylbenzene, and it is also referred to in the scientific literature as 2-phenethyl isothiocyanate, β-phenethyl isothiocyanate, phenyl ethyl ITC, or 2-phenyl ethyl ITC. Structurally, ChEBI designates it as an isothiocyanate having a phenethyl group attached to the nitrogen. Its CAS registry number is 2257-09-2. PEITC is a low molecular weight compound (MW = 163.2 g/mol) and is fairly lipophilic (logP = 3.47).
Glucosinolates (β-thioglucoside N-hydroxysulfates) are naturally occurring compounds that are precursors for isothiocyanates. These phytochemicals are of widespread occurrence in the cruciferous vegetables of the Cruciferae or Brassicaceae family, containing hundreds of genera and thousands of species. These compounds come from cruciferous plants (Brassicaceae), such as watercress, cabbage, cauliflower, turnip, horseradish, broccoli, and Brussels sprouts.
PEITC occurs as its glucosinolate precursor gluconasturtiin in the cruciferous vegetable watercress (Nasturtium officinale). Watercress also contains smaller quantities of 7-methylsulfinylheptyl GS and 8-methylsulfinyloctyl GS. The seeds of the cruciferous Barbarea verna (land cress) are enriched with only one glucosinolate, gluconasturtiin, containing approximately 3% by weight. PEITC originates from gluconasturtiin and has been previously shown to be highly accumulated in watercress flowers, while it is less abundant in leaves and stems. Horseradish (Armoracia rusticana) powder contains mainly PEITC.
In the intact plant, myrosinase is stored separately from glucosinolates; when the plant tissue is damaged by chopping or chewing, the enzyme contacts with glucosinolates and catalyzes the lysis, and the reaction produces indoles, nitriles, thiocyanates, or isothiocyanates. PEITC is thus prepared by hydrolysis of gluconasturtiin, an abundant natural product present in cruciferous vegetables. Glucosinolates are water-soluble and stable compounds, but isothiocyanates are hydrophobic and very reactive compounds.
PEITC is a bioactive compound involved in several biological mechanisms that are naturally related to protecting the plant against external factors. The plant produces PEITC in response to specific stress situations, since it presents biocidal activity against various pathogens, such as bacteria, fungi, insects, and other biotic stressors.
PEITC is encountered in research and commercial settings in several forms:
PEITC as a discrete chemical entity was not known to pre-modern practitioners; rather, its history is inseparable from the historical use of the plants in which it occurs—principally watercress (Nasturtium officinale), horseradish (Armoracia rusticana), mustard greens, and related Brassicaceae species. Watercress leaf is rich in vitamins, minerals, tannins, phenolic acids, and terpenes. Like other members of the Brassicaceae family, watercress contains glucosinolates, sulfur-containing compounds that are converted to bioactive isothiocyanates when cell walls are breached through chopping or chewing. The most abundant glucosinolate in watercress is gluconasturtiin, which is converted to phenethyl isothiocyanate (PEITC).
Hippocrates, a Greek physician who lived in the 4th century BC, was particularly convinced of the health benefits of watercress, a plant he often referred to as the "cure of cures." Watercress contains phenethyl isothiocyanate (PEITC), a special mustard oil known to have significant anti-cancer properties. Numerous studies have documented that PEITC can be utilized therapeutically in various human diseases including hyperglycemia, hypertension, hypercholesterolemia, bronchitis, arthritis and scurvy. These indications reflect the broader traditional medicinal use of watercress-containing preparations.
Ethnobotanists have noted that in Europe, watercress soups date back to medieval times; in parts of India, mustard oil and greens have long been part of winter and early spring cuisines. Interest in natural products as an alternative to synthetic drugs for cancer treatment is largely due to low cost, established historical use in traditional medicinal systems, easy availability and minimal or no toxicity.
Phenethyl isothiocyanate was first isolated in the early 1980s by phytochemists intrigued by the cancer-preventive reputation of cruciferous vegetables. Through the 1990s and 2000s, dozens of laboratory and animal studies charted its ability to induce detoxification enzymes, particularly glutathione S-transferase (GST) and NAD(P)H:quinone oxidoreductase 1 (NQO1). It has been approximately 30 years since Chung and colleagues first showed that PEITC, which is found in substantial quantities as the conjugate gluconasturtiin in watercress and several other cruciferous vegetables, inhibited an obligatory step in the metabolic activation of the tobacco-specific lung carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK).
The epidemiological evidence suggests a strong inverse relationship between dietary intake of cruciferous vegetables and the incidence of cancer. Among other constituents of cruciferous vegetables, isothiocyanates are the main bioactive chemicals present. Isothiocyanates are the most common products that result from the enzymatic breakdown of glucosinolates by the myrosinase (β-thioglucosidase) enzyme.
Of all the isothiocyanates, phenethyl isothiocyanate (PEITC) is the only one that has reached the clinical stage of testing. Within watercress, PEITC is the dominant bioactive phytochemical of interest, accompanied by other glucosinolate-derived compounds and polyphenols. Watercress (Nasturtium officinale) belongs to the Brassicaceae family and is a well-known aquatic plant due to its high content of phytochemicals with high nutritional impact including polyphenols (quercetin-3-O-rutinoside, kaempferol-3-O-rutinoside, isorhamnetin, protocatechuic acid and chlorogenic acid), minerals (Ca, K, Zn, Fe), vitamins (A, B, C, D and K), soluble sugars and proteins.
Underlying PEITC's biological activities is the sulfhydryl reactivity of PEITC with cysteine residues in its protein targets. PEITC is a naturally occurring electrophilic compound that readily undergoes thiocarbamoylation reactions with cellular thiols. Following uptake into cells, the predominant initial reaction of PEITC is with glutathione (GSH), the major intracellular antioxidant. PEITC conjugates are then effluxed from the cell, but breakdown of extracellular PEITC conjugates results in liberation of PEITC which is free to re-enter the cell.
The inhibitory effects of isothiocyanates have been attributed to their ability to regulate multiple molecular mechanisms including inhibition of Phase I drug-metabolizing enzymes (e.g., cytochrome P-450), induction of Phase II detoxifying enzymes (e.g., glutathione-S-transferases), induction of cell cycle arrest and apoptosis, and inhibition of histone deacetylases.
PEITC modulates the activity and protein expression of a wide range of Phase I oxidative and Phase II conjugating drug-metabolizing enzymes, inhibiting the formation of carcinogens and increasing their detoxification. Specifically, nearly all of the carcinogen-inhibiting effects of PEITC can be traced to its inhibitory effects on cytochrome P450 enzymes including human P450s 2A13, 2A6, 1A2, and 2B6, which are catalysts of NNK bioactivation.
PEITC has multiple biological effects, including activation of cytoprotective pathways, such as those mediated by the transcription factor nuclear factor erythroid 2 p45-related factor 2 (NRF2) and the transcription factor heat shock factor 1 (HSF1), and can cause changes in the epigenome. This chemical reactivity highlights the critical importance of the dose of PEITC for achieving on-target selectivity, which should be carefully considered in the design of future clinical trials.
The conjugation of PEITC with intracellular glutathione and the subsequent removal of the conjugate result in depletion of glutathione and alteration in redox homeostasis leading to oxidative stress. PEITC binds to GSH and causes its depletion in cancer cells, leading to ROS-induced cell damage. Interestingly, the sensitivity towards PEITC correlates with constitutive GSH levels present in the cells. The cells with higher levels of GSH were relatively more sensitive to PEITC, which may also explain the selectivity toward cancer cells compared with normal cells, since cancer cells have higher levels of GSH.
At high concentrations, PEITC leads to accumulation of reactive oxygen species and cytoskeletal changes, resulting in cytotoxicity. Based on the available data, it is well-established that PEITC acts as a pro-oxidant to initiate ROS generation, leading to apoptosis in cell culture models.
PEITC treatment induces G2/M phase cell cycle arrest, down-regulates the anti-apoptotic proteins Bcl-2 and Bcl-XL, up-regulates the pro-apoptotic protein Bak, and suppresses Notch 1 and 2 levels. Depending on the cancer cell type, arrest can occur at different checkpoints. In human oral cancer HSC-3 cells, PEITC induced cell death through the G0/G1 phase arrest and induction of apoptosis.
PEITC's anticancer activities are mediated through several mechanisms, including the induction of apoptosis, inhibition of cell proliferation, suppression of angiogenesis, and reduction of metastasis. PEITC targets crucial cellular signaling pathways involved in cancer progression, notably the NF-κB, Akt, and MAPK pathways. Multiple cell death modalities, including ferroptosis, apoptosis, and autophagy, can be triggered in cancer cells.
Mechanism studies in human prostate cancer cells revealed that PEITC is a dual inhibitor of aberrant DNA hypermethylation and histone deacetylases, reactivating silenced genes and regulating the androgen-mediated growth of tumor cells. In a study examining human prostate cancer LNCaP cells, treatment with PEITC resulted in demethylation of the GSTP1 gene promoter, inhibited the activity of HDACs, and induced selective histone acetylation and methylation.
Some isothiocyanates, including PEITC, bind covalently to cysteines in tubulin as a molecular target, which may mediate microtubule disruption and blocking of mitosis.
PEITC alters iron metabolism related to the processes of iron import, storage, and export, resulting in increased labile iron. PEITC causes oxidative stress as a consequence of GSH depletion, inducing ROS generation and lipid peroxidation. Multiple cell death modalities, including ferroptosis, apoptosis, and autophagy, are triggered in human cancer cells.
Inhibition of angiogenesis is one of the recently reported mechanisms of breast cancer prevention by PEITC.
The relationship between the consumption of cruciferous vegetables and chemoprotection has been widely documented in epidemiological studies. Several epidemiological studies from different parts of the world provide evidence for the reduced risk of cancer with higher total isothiocyanate intake in the form of cruciferous vegetables, covering cancers of the lung, breast, stomach, bladder, colorectum, pancreas, prostate, and kidney. These epidemiological associations form the backdrop against which laboratory and clinical investigation of PEITC has proceeded.
PEITC is known to not only prevent the initiation phase of the carcinogenesis process but also to inhibit the progression of tumorigenesis. PEITC has been found in research to be relevant against various cancer types, such as breast, prostate, lung, colon, and pancreatic cancers.
2-Phenethyl isothiocyanate (PEITC) is an inhibitor of the metabolic activation and lung carcinogenicity of the tobacco carcinogen NNK in F344 rats and A/J mice. In human lung cancer L9981 cells, both BITC and PEITC inhibited cell growth in a dose-dependent manner; cell migrations were substantially reduced, as were cell invasions. Metastasis-related genes MMP-2, Twist, and β-catenin were also modulated.
A clinical trial was carried out to determine whether PEITC inhibits the metabolic activation of NNK in smokers. Cigarette smokers were recruited and asked to smoke cigarettes containing deuterium-labelled NNK for an acclimation period. Subjects were then randomly assigned to one of two arms: PEITC followed by placebo, or placebo followed by PEITC. During the one-week treatment period, each subject took PEITC (10 mg in 1 mL of olive oil, 4 times per day). Overall, intake of PEITC 40 mg/day for 5 days significantly inhibited NNK metabolic activation by 8% (P = 0.023), and increased urinary detoxification mercapturic acids formed from benzene by 25% (P = 0.002) and acrolein by 15% (P = 0.005), in 82 current smokers in the US.
Four clinical studies for human testing of PEITC were initiated, out of which one study (NCT00968461) was withdrawn for unexplained reasons. Of the remaining three, one study (NCT00005883) was completed with a goal of multi-dose testing of PEITC in lung cancer patients, although the results of this phase I study had not been published at the time of a 2014 review. Another Phase I clinical study (NCT01790204) was planned at Georgetown University, with the goal of testing the beneficial effects of PEITC in oral carcinoma with mutant p53.
Evidence strength: For lung cancer chemoprevention in smokers, there is one completed small randomized crossover clinical trial demonstrating modulation of carcinogen metabolism, and at least one additional Phase I dose-finding trial. The biomarker data from the clinical study are statistically significant but the trial was small, short-term, and focused on pharmacodynamic markers rather than cancer incidence endpoints. The overall body of human clinical evidence remains preliminary.
In vivo chemopreventive efficacy of PEITC against prostate cancer has been established in a transgenic mouse model (TRAMP). Feeding of 3 µmol PEITC/g diet significantly decreased incidence as well as burden of poorly-differentiated cancer in the dorsolateral prostate of TRAMP mice. In studies of prostate cancer, PEITC has been demonstrated as a dual inhibitor of DNA hypermethylation and histone deacetylases, reactivating silenced genes including the π-class glutathione S-transferase which is inactivated by aberrant DNA methylation in the vast majority of clinical prostate tumors. PEITC and its major metabolite have been demonstrated to inhibit post-initiation progression of carcinogenesis, including the induction of autophagy, apoptosis, and down-regulation of the androgen receptor, thus reducing its growth stimulation on prostate tumor cells.
Unique to prostate cancer is that PEITC downregulates the transcriptional factor Sp1, a regulator of androgen receptor expression.
Evidence for the protective effect of cruciferous vegetables and their components, including PEITC, against prostate cancer derives from population-based observational studies as well as laboratory investigations. For example, a population-based case-control study suggested an inverse association between intake of cruciferous vegetables and the risk of prostate cancer. PEITC has undergone small human clinical trials against cancers, such as breast cancer, lung cancer, and prostate cancer, either within cruciferous food or as a single agent.
Evidence strength: Strong preclinical evidence in animal models and cell lines; epidemiological evidence is observational and does not isolate PEITC from other cruciferous vegetable constituents. Human clinical evidence is limited to small Phase I trials assessing safety and bioavailability rather than efficacy endpoints.
Exposure to PEITC inhibited pancreatic cancer cell growth in a dose-dependent manner, with an IC50 of approximately 7 µmol/L. PEITC treatment induced G2/M phase cell cycle arrest, down-regulated anti-apoptotic proteins Bcl-2 and Bcl-XL, up-regulated pro-apoptotic protein Bak, and suppressed Notch 1 and 2 levels. In addition, treatment with PEITC induced cleavage of poly-(ADP-ribose) polymerase and led to increased cytoplasmic histone-associated DNA fragmentation. Oral administration of PEITC suppressed the growth of pancreatic cancer cells in a MIAPaca2 xenograft animal model.
Evidence strength: Preclinical only (in vitro and xenograft animal models). No human clinical trial data were identified specifically for pancreatic cancer.
Several in vitro and in vivo studies have elucidated molecular mechanisms underlying the pharmacodynamics of PEITC in breast cancer, including cancer cell apoptosis by upregulation of apoptotic genes, cell cycle arrest at G2/M phase by generation of reactive oxygen species and depletion of intracellular glutathione, downregulation of the estrogen receptor, decrease in sensitivity to estrogen, and inhibition of tumor metastasis. PEITC readily induced apoptosis in MDA-MB-231 cells, associated with rapid activation of caspases 9 and 3, and decreased expression of BAX. MCF7 cells were relatively resistant to the apoptosis-promoting effects of PEITC, and this relative resistance was associated with high basal expression of NRF2 and raised intracellular levels of GSH.
Evidence strength: Predominantly in vitro and animal model data; the differential sensitivity of cancer cell lines based on NRF2 and GSH status is a noteworthy mechanistic finding. Human clinical trial data for breast cancer specifically were not identified in the sources reviewed.
Both Brassicaceae extracts and isothiocyanates exert antiproliferative effects in HT29 colorectal cancer spheroids, arresting cell cycle at G2/M, possibly due to ITC-induced DNA damage. Colony formation and expression of LGR5 and CD133 cancer stemness markers were significantly reduced. Only watercress extract and PEITC decreased ALDH1 activity in a dose-dependent manner, as well as β-catenin expression.
Evidence strength: In vitro and animal model evidence only. No completed human clinical trials for colorectal cancer chemoprevention with PEITC as a single agent were identified in the sources reviewed.
Research indicates that PEITC-enriched watercress flower extract, in a cell-based system, can modulate the activity of key enzymes involved in cellular antioxidant defense mechanisms. In addition, PEITC-enriched extract induces lipid and protein oxidation in a concentration-dependent manner, and its cytotoxicity is not only dependent on PEITC itself but also on its N-acetylated cysteine conjugated form.
Evidence strength: In vitro only. No human clinical trials for melanoma were identified in the sources reviewed.
Studies investigated the molecular mechanism and anticancer potential of PEITC in oral squamous cell carcinoma (OSCC) cells with various p53 statuses. PEITC inhibited the growth of multiple oral cancer cell lines (OC2, SCC4, and SCC25) in a dose-dependent manner with low toxicity to normal cells. Treatment with PEITC induced reactive oxygen species production, nitric oxide generation, and GSH depletion and triggered DNA damage response. PEITC is in one clinical trial evaluating its safety and efficacy profile in head and neck cancer patients (NCT03034603).
Evidence strength: Preclinical evidence is robust across multiple OSCC cell lines; one registered clinical trial was noted as of publication of the 2018 review.
PEITC has antioxidant, anti-inflammatory, bactericidal, and anticarcinogenic properties. One published review summarized the current knowledge on the role of PEITC as a potential natural nutraceutical or adjuvant against oxidative/inflammatory-related disorders in the gastrointestinal tract.
Evidence strength: In vitro and animal model evidence; no human clinical trials focused specifically on GI health were identified in sources reviewed.
PEITC increased the expression of cytoprotective proteins and protected human skin tissues from UVR-induced skin photodamage. Some members of the isothiocyanate class, including PEITC, have undergone human clinical trials against skin disorders.
Evidence strength: Limited; mechanistic in vitro data and mention of some clinical trial activity, but details of completed human trials for skin photoprotection were not available in the sources reviewed.
PEITC is a hydrophobic compound and is rapidly absorbed, with a bioavailability of 70% observed after oral administration. Intake of 100 g of watercress, providing about 25 mg of PEITC, showed a Tmax of about 2.6 h, a Cmax of 1.0 µM, and a half-life of approximately 5 h.
The half-life of PEITC was 2.4 hours in one phase I clinical study. A Phase I study (NCI CN-55120) confirmed that after oral intake of 200 mg PEITC by human volunteers, 10 µM PEITC can be achieved in plasma. Interestingly, many studies have shown the anti-cancer efficacy of PEITC at concentrations less than 10 µM.
PEITC demonstrates dose-dependent pharmacokinetics with first-order absorption, nonlinear tissue distribution due to interactions with ABC transporters and intracellular glutathione depletion, and capacity-limited elimination due to saturation of glutathione-S-transferases in the liver.
After individuals ate watercress as part of a breakfast meal and 24-hour urine samples were collected, a urinary metabolite was found and its identity was confirmed as the N-acetylcysteine conjugate of PEITC by comparison with the synthetic standard using nuclear magnetic resonance and mass spectrometry. This PEITC-NAC conjugate serves as the primary urinary biomarker of PEITC exposure. Another aspect of chemoprevention by isothiocyanates involves the glutathione-S-transferase enzymes (GSTs) which catalyze the conjugation of isothiocyanates such as PEITC with glutathione, leading to the excretion in urine of PEITC-NAC.
PEITC has unique biological features characterized by high oral bioavailability, low clearance, and high protein binding.
The following dosages are reported directly from published research and should not be interpreted as recommendations:
The investigators of the smoker clinical trial noted that the dosing form of PEITC used was not optimal, and the dose may have been too low. Further research is necessary to determine the optimal dosage, understand its bioavailability, and assess potential side effects for clinical use.
At high concentrations, PEITC leads to accumulation of reactive oxygen species and cytoskeletal changes, resulting in cytotoxicity. Underlying these activities is the sulfhydryl reactivity of PEITC with cysteine residues in its protein targets. This chemical reactivity highlights the critical importance of the dose of PEITC for achieving on-target selectivity, which should be carefully considered in the design of future clinical trials.
PEITC is a naturally occurring electrophile which depletes intracellular glutathione (GSH) levels and triggers accumulation of reactive oxygen species (ROS). Due to isothiocyanates' inherently high level of thiol reactivity, elevated intracellular levels of PEITC have the potential to deplete intracellular glutathione (GSH) reserves. Since GSH is a primary intracellular antioxidant and cytoprotective enzyme cofactor, preservation of intracellular GSH status is crucial for cytoprotection.
PEITC demonstrates capacity-limited elimination due to saturation of glutathione-S-transferases in the liver, which has implications for dosing and pharmacokinetics at higher intakes.
PEITC exerts inhibitory effects on cytochrome P450 enzymes including human P450s 2A13, 2A6, 1A2, and 2B6. This inhibitory activity, while of interest for carcinogen detoxification, carries the potential to alter the metabolism of pharmaceutical drugs that are substrates of these enzymes. PEITC has been specifically characterized as a phytochemical inhibitor of CYP2E1 in animal models, which is relevant given that CYP2E1 participates in the metabolism of a range of xenobiotics, solvents, and drugs.
The relative resistance of some cancer cell lines to PEITC-induced apoptosis was associated with high basal expression of NRF2 and raised intracellular levels of GSH. Differences in the basal expression of NRF2 and resultant changes in GSH levels may be an important determinant of sensitivity to PEITC-induced apoptosis. This suggests that individual variation in NRF2 pathway activity may significantly influence both therapeutic responses and safety profiles.
In the smoker clinical trial, the mean level of the 24-hour urinary excretion of PEITC-NAC was approximately 80 micromoles. In the phase I clinical study, the half-life of PEITC was 2.4 hours. The urinary excretion of PEITC equivalents averaged 13.5 mg in 24 hours in participants taking supplemental PEITC, which was approximately only one-third the amount excreted by watercress consumers achieving the same nominal intake, suggesting that the food matrix significantly affects bioavailability.
Pre-clinical evidence suggests that the combination of PEITC with conventional anti-cancer agents is highly effective in improving overall efficacy. PEITC, at 5 µM, was found to potentiate the cytotoxic effects of imatinib in K562 chronic myeloid leukemia cells and enhanced cell death by modulating the BCR-ABL and STAT5 signaling pathways. The potential for synergistic interactions with chemotherapy agents warrants attention in settings where PEITC supplementation may occur concurrently with pharmaceutical cancer treatment.
It is well-established that PEITC acts as a pro-oxidant to initiate ROS generation, leading to apoptosis in cell culture models. However, resultant modulation of ROS from an in vitro study can vary significantly from an in vivo study with the same agent. Thus, outcomes based on the in vitro studies may be debatable. The complex pharmacokinetics and pharmacodynamics of PEITC necessitate a systems-biology approach in parallel with PK/PD modeling to develop PEITC as a therapeutic agent for treating cancers.
Based on accumulating evidence, PEITC appears to be a promising agent for cancer therapy and has been under clinical trials for leukemia and lung cancer. Of all the isothiocyanates, PEITC is the only one that has reached the clinical stage of testing. PEITC has been registered in clinical trials for evaluating its safety and efficacy profile in head and neck cancer patients (NCT03034603), and for its long-term effects in cancer patients' outcomes (NCT02468882).
PEITC has unique biological features characterized by high oral bioavailability, low clearance, and high protein binding. Current evidence and outcomes suggest that it holds potential for future clinical development and application in cancer therapy.
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.
Health conditions that PEITC (phenethyl isothiocyanate) may help support.
Body systems that PEITC (phenethyl isothiocyanate) may help support.
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