In vitro studies of Psorinum 6x on several human cancer cell lines reveal its anti-cancer potential
Keywords: Psorinum therapy, reactive oxygen species, anticancer potential, apoptosis, drug-DNA interaction
AbstractObjective: Psorinum therapy is claimed to combat/ameliorate a variety of human cancers, but for obvious reasons these studies are devoid of any untreated/placebo-treated controls. Therefore, if Psorinum 6x administration to cancer cells of different origin can show visible anti-cancer effects in a controlled in vitro study has been examined. Materials & Methods: Psorinum 6x was provided by Hahnemann Publishing Company (HAPCO), 165 BB Ganguly Street, Kolkata for our research. It was prepared by HAPCO by following standard homoeopathic guidelines from authentic pus cells obtained from an eczema patient being treated at National Institute of Homeopathy, Salt Lake, Kolkata. MTT assay was initially done on several cancer cell lines like A549 (lung cancer), HeLa (cervix cancer), HepG2 (liver cancer) and MCF7 (breast cancer). Psorinum 6x showed strongest anticancer effect against A549 though it also had lesser effect against other cell lines tested. Therefore, A549 was chosen as the model cell line for further study using several relevant protocols. Effects of Psorinum 6x were compared with that of "Placebo 6x" control made of the same stock of "vehicle" used for preparation of Psorinum 6x. Protocols like analysis of cell cycle progression, generation of reactive oxygen species (ROS), change in mitochondrial membrane potential (MMP) and actual cell death (apoptosis), if any, were analysed flow-cytometrically. Whether Psorinum 6x could damage DNA and induce morphological changes were also determined microscopically. Expression of different signal proteins related to cell death (apoptosis) and survival were critically studied by western blot analysis and confocal microscopy. Further, to determine if Psorinum 6x could interact directly with DNA to induce any conformational changes was also determined by circular dichroism (CD) spectroscopy. Results: Psorinum 6x treatment reduced cell viability and inhibited cell proliferation at 24h after treatment, arresting cell cycle at sub-G stage. Upon Psorinum treatment, there were increase of ROS and MMP depolarization, morphological changes and DNA damage typical of apoptosis in A549 cells, along with externalization of phosphatidyl serine. Further, an increase in p53 level, Bax translocation into mitochondria, cytochrome c release into cytosol along with reduction of Bcl2 level and caspase 3 activation were noted which eventually drove A549 cells towards apoptosis; the apoptotic signalling was found to be through mitochondria-mediated caspase 3 dependent pathway. Evidence of direct interaction of Psorinum with cellular DNA was revealed from CD-spectroscopy. Conclusion: Thus, Psorinum 6 x had positive anti-cancer effects against a number of cancer cells, of which it appeared to have strongest effect against the lung cancer cell, A549.