Thioformin

Aim: Fluopsin C, an antibiotic isolated from Pseudomonas jinanesis, has shown antitumor effects on several cancer cell lines. In the current study, the oncotic cell death induced by fluopsin C was investigated in human breast adenocarcinoma cells in vitro. Methods: Human breast adenocarcinoma cell lines MCF-7 and MD-MBA-231 were used. The cytotoxicity was evaluated using MTT assay. Time-lapse microscopy and transmission electron microscopy were used to observe the morphological changes. Cell membrane integrity was assessed with propidium iodide (PI) uptake and lactate dehydrogenase (LDH) assay. Flow cytometry was used to measure reactive oxygen species (ROS) level and mitochondrial membrane potential (Δψm). A multimode microplate reader was used to analyze the intracellular ATP level. The changes in cytoskeletal system were investigated with Western blotting and immunostaining. Results: Fluopsin C (0.5-8 μmol/L) reduced the cell viability in dose- and time-dependent manners. Its IC50 values in MCF-7 and MD-MBA-231 cells at 24 h were 0.9 and 1.03 μmol/L, respectively. Fluopsin C (2 μmol/L) induced oncosis in both the breast adenocarcinoma cells characterized by membrane blebbing and swelling, which was blocked by pretreatment with the pan-caspase inhibitor Z-VAD-fmk. In MCF-7 cells, fluopsin C caused PI uptake into the cells, significantly increased LDH release, induced cytoskeletal system degradation and ROS accumulation, decreased the intracellular ATP level and Δψm. Noticeably, fluopsin C exerted comparable cytotoxicity against the normal human hepatocytes (HL7702) and human mammary epithelial cells with the IC50 values at 24 h of 2.7 and 2.4 μmol/L, respectively. Conclusion: Oncotic cell death was involved in the anticancer effects of fluopsin C on human breast adenocarcinoma cells in vitro. The hepatoxicity of fluopsin C should not be ignored.

Fluopsin C is an antibiotic compound derived from secondary metabolism of different microorganisms, which possesses antitumor, antibacterial, and antifungal activity. Related to fluopsin C antiproliferative activity, the aim of this study was to examine the following parameters: cytotoxicity, genotoxicity, cell cycle arrest, cell death induction (apoptosis), mitochondrial membrane potential (MMP), colony formation, and mRNA expression of genes involved in adaptive stress responses and cellular death utilizing a monolayer. In addition, a three-dimensional cell culture was used to evaluate the effects on growth of tumor spheroids. Fluopsin C was cytotoxic (1) producing cell division arrest in the G1 phase, (2) elevating expression of mRNA of the CDKN1A gene and (3) decrease in expression of mRNA H2AFX gene. Further, fluopsin C enhanced DNA damage as evidenced by increased expression of mRNA of GADD45A and GPX1 genes, indicating that reactive oxygen species (ROS) may be involved in the observed genotoxic response. Reticulum stress was also detected as noted from activation of the ribonuclease inositol-requiring protein 1 (IRE1) pathway, since a rise in mRNA expression of the ERN1 and TRAF2 genes was observed. During the cell death process, an increase in mRNA expression of the BBC3 gene was noted, indicating participation of this antibiotic in oncotic (ischemic) cell death. Data thus demonstrated for the first time that fluopsin C interferes with the volume of tumor spheroids, in order to attenuate their growth. Our findings show that fluopsin C modulates essential molecular processes in response to stress and cell death.