benzyloxycarbonylvalyl-alanyl-aspartyl-fluoromethyl-ketone and phenethyl-isothiocyanate

The treatment of pancreatic cancer, one of the most aggressive gastrointestinal tract malignancies, with current chemotherapeutic drugs has had limited success due to its chemoresistance and poor prognosis. Therefore, the development of new drugs or effective combination therapies is urgently needed. Cotylenin A (CN-A) (a plant growth regulator) is a potent inducer of differentiation in myeloid leukemia cells and exhibits potent antitumor activities in several cancer cell lines. In the present study, we demonstrated that CN-A and phenethyl isothiocyanate (PEITC), an inducer of reactive oxygen species (ROS) and a dietary anticarcinogenic compound, synergistically inhibited the proliferation of MIAPaCa-2, PANC-1 and gemcitabine-resistant PANC-1 cells. A combined treatment with CN-A and PEITC also effectively inhibited the anchorage-independent growth of these cancer cells. The combined treatment with CN-A and PEITC strongly induced cell death within 1 day at concentrations at which CN-A or PEITC alone did not affect cell viability. A combined treatment with synthetic CN-A derivatives (ISIR-005 and ISIR-042) or fusicoccin J (CN-A-related natural product) and PEITC did not have synergistic effects on cell death. The combined treatment with CN-A and PEITC synergistically induced the generation of ROS. Antioxidants (N-acetylcysteine and trolox), ferroptosis inhibitors (ferrostatin-1 and liproxstatin), and the lysosomal iron chelator deferoxamine canceled the synergistic cell death. Apoptosis inhibitors (Z-VAD-FMK and Q-VD-OPH) and the necrosis inhibitor necrostatin-1s did not inhibit synergistic cell death. Autophagy inhibitors (3-metyladenine and chloroquine) partially prevented cell death. These results show that synergistic cell death induced by the combined treatment with CN-A and PEITC is mainly due to the induction of ferroptosis. Therefore, the combination of CN-A and PEITC has potential as a novel therapeutic strategy against pancreatic cancer.

Topics: Acetylcysteine; Amino Acid Chloromethyl Ketones; Antineoplastic Agents; Antioxidants; Apoptosis; Cell Death; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cyclohexylamines; Diterpenes; Glycosides; Humans; Isothiocyanates; Pancreatic Neoplasms; Phenylenediamines; Quinolines; Reactive Oxygen Species

The present study was undertaken to gain insights into the molecular mechanism of apoptosis induction by phenethyl isothiocyanate (PEITC) using prostate cancer cell lines derived from transgenic adenocarcinoma mouse prostate (TRAMP) mice (TRAMP-C1 and TRAMP-C2).. The viability of TRAMP-C1 and TRAMP-C2 cells was reduced significantly in the presence of PEITC in a concentration-dependent manner as determined by sulforhodamine B and trypan blue dye exclusion assays. Treatment of TRAMP-derived cells with PEITC revealed features characteristic of apoptosis induction, including appearance of subdiploid cells (determined by flow cytometry), cytoplasmic histone-associated DNA fragmentation (determined by an ELISA assay), and cleavage of caspase-3 (determined by immunoblotting). The PEITC-induced apoptosis in TRAMP-derived cells was associated with a marked increase in the level of proapoptotic protein Bak and/or a decrease in the levels of antiapoptotic protein Mcl-1 or Bcl-xL and disruption of mitochondrial membrane potential. The SV40 immortalized mouse embryonic fibroblasts derived from Bak and Bax double knockout mice were significantly more resistant to PEITC-induced DNA fragmentation compared with wild-type or Bak-/- mouse embryonic fibroblasts. The PEITC-induced apoptosis in both cell lines was significantly attenuated in the presence of caspase inhibitors zVAD-fmk, zLEHD-fmk, and zIETD-fmk. Oral administration of PEITC (9 or 12 micromol PEITC/d, Monday-Friday) significantly retarded growth of TRAMP-C1 xenografts in nude mice without causing weight loss or any other side effects.. The results of the present study indicate that caspase-dependent apoptosis by PEITC is mediated by Bak and Bax proteins.

Topics: Adenocarcinoma; Amino Acid Chloromethyl Ketones; Animals; Apoptosis; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein; Caspase Inhibitors; Caspases; Cell Line, Tumor; Cell Survival; Cysteine Proteinase Inhibitors; Dose-Response Relationship, Drug; Immunoblotting; Intracellular Membranes; Isothiocyanates; Male; Membrane Potentials; Membrane Proteins; Mice; Mice, Knockout; Mice, Transgenic; Mitochondria; Neoplasms; Plant Preparations; Prostatic Neoplasms; Proto-Oncogene Proteins c-bcl-2; Vegetables

Phenethyl isothiocyanate and allyl isothiocyanate induce apoptosis of human leukaemia HL60 cells in vitro. Apoptosis was associated with cleavage of p22 BID protein to p15, p13 and p11 fragments and activation of JNK and tyrosine phosphorylation (18 kDa and 45 kDa proteins). All these effects and apoptosis were prevented by exogenous glutathione (15 mM). Protein tyrosine phosphatase activity was unchanged. The general caspase inhibitor Z-VAD-fmk prevented apoptosis but not JNK activation - excluding a role for caspases in JNK activation, whereas curcumin prevented JNK activation but only delayed apoptosis. This suggests that in isothiocyanate-induced apoptosis, the caspase pathway has an essential role, the JNK pathway a supporting role, and inhibition of protein tyrosine phosphatases is not involved.

Topics: Amino Acid Chloromethyl Ketones; Anticarcinogenic Agents; Apoptosis; BH3 Interacting Domain Death Agonist Protein; Carrier Proteins; Cysteine Proteinase Inhibitors; Enzyme Activation; Enzyme Inhibitors; HL-60 Cells; Humans; Isothiocyanates; JNK Mitogen-Activated Protein Kinases; Mitogen-Activated Protein Kinases; Neoplasms; Phosphorylation; Phosphotyrosine; Signal Transduction