Cytotoxicity of spermine oxidation products to multidrug resistant melanoma M14 ADR2 cells: Sensitization by the MDL 72527 lysosomotropic compound

It has been confirmed that multidrug resistant (MDR) human melanoma cells are more sensitive than their wild-type counterparts to H(2)O(2) and aldehydes, the products of bovine serum amine oxidase (BSAO)-catalyzed oxidation of spermine. The metabolites formed by BSAO and spermine are more toxic than exogenous H(2)O(2) and acrolein, even thou-h their concentration is lower during the initial phase of incubation due to their more gradual release than the exogenous products. Both wild-type and MDR cells, after pre-treatment with MDL 72527, an inactivator of polyamine oxidase and a lysosomotropic compound, show to be sensitized to subsequent exposure to BSAO/spermine. Evidence of ultrastructural aberrations and acridine orange release from lysosomes is presented in this work that is in favor of the permeabilization of the lysosomal membrane as the major cause of sensitization by MDL 72527. Owing to its lysosomotropic effect, pre-treatment with MDL 72527 amplifies the ability of the metabolites formed from spermine by oxidative deamination to induce cell death. Since it is conceivable that combined treatment with a lysosomotropic compound and BSAO/spermine would be effective against tumor cells, it is of interest to search for such novel compounds, which might be promising for application in a therapeutic setting

Anticancer drugs and hyperthermia enhance cytotoxicity induced by polyamine enzymatic oxidation products

A correlation between regulation of cell proliferation and polyamine metabolism is described. The latter can enter protein synthesis through the modification of eukaryotic initiation factor 5A (eIF5A) and the formation of the peculiar amino acid hypusine. Specific inhibitors of hypusine formation induce apoptosis that can be potentiated by the combination with cytokines such as interferon alpha (IFN alpha) that itself decreases hypusine synthesis. We have also demonstrated that the concomitant treatment of cancer cells with IFNa and the protein synthesis inhibitor fusion protein TGF alpha/Pseudomonas Aeruginosa toxin synergize in inducing cancer cell growth inhibition. Another way used by polyamines to induce apoptosis is the generation of intracellular oxidative stress through the interaction with bovine serum amine oxidase (BSAO). This enzyme used simultaneously to spermine induces apoptosis, necrosis, inhibition of cell proliferation and inhibition of DNA and protein synthesis in several cell types. The enzymatic oxidation products of polyamine, H2O2 and aldehyde(s) cause these effects. We have recently found that the cytotoxicity of anti-cancer agents, either etoposide or docetaxel, in cancer cells is potentiated in the presence of BSAO/Spermine. In conclusion, polyamine metabolites could be useful in the design of new therapeutic strategies

Bidirectional fluxes of spermine across the mitochondrial membrane.

The polyamine spermine is transported into the mitochondrial matrix by an electrophoretic mechanism having as driving force the negative electrical membrane potential (DW). The presence of phosphate increases spermine uptake by reducingDpH and enhancingDW. The transport system is a specific uniporter constituted by a protein channel exhibiting two asymmetric energy barriers with the spermine binding site located in the energy well between the two barriers. Although spermine transport is electrophoretic in origin, its accumulation does not follow the Nernst equation for the presence of an efflux pathway. Spermine efflux may be induced by different agents, such as FCCP, antimycin A and mersalyl, able to completely or partially reduce theDWvalue and, consequently, suppress or weaken the force necessary to maintain spermine in the matrix. However this efflux may also take place in normal conditions when the electrophoretic accumulation of the polycationic polyamine induces a sufficient drop inDWable to trigger the efflux pathway. The release of the polyamine is most probably electroneutral in origin and can take place in exchange with protons or in symport with phosphate anion. The activity of both the uptake and efflux pathways induces a continuous cycling of spermine across the mitochondrial membrane, the rate of which may be prominent in imposing the concentrations of spermine in the inner and outer compartment. Thus, this event has a significant role on mitochondrial permeability transition modulation and consequently on the triggering of intrinsic apoptosis

A combined structure and ligand-based application on BSAO as an encouraging approach in anti-cancer therapy.

Bovine serum amine oxidase (BSAO) is a copper-containing enzyme that catalyzes the oxidation of primary amines, such natural polyamines spermine and spermidine, into their corresponding aldehydes, to release the cytotoxic hydrogen peroxide and ammonia. Since these products were recognized to induce stress-activated signal transduction pathways, determining necrosis or apoptosis in tumor-cultured cell lines,1 new strategies are emerging2 for anti-cancer treatments with BSAO and the development of new polyamine analogs is mandatory. To improve enzyme-substrate affinity, structure based (SB) and ligand based (LB) studies were performed on 25 among natural polyamines and newly synthesized and biologically assayed polyamine analogs. The connubium3 between the AutoDock suite,4 3-D QSAutogrid/R5 and COMBINEr6, 7 methodologies, allowed to define the most influent ligand-residues interactions and derive a predictive 3-D QSAR pharmacophoric model as a promising paradigm to predict and design novel BSAO substrates, endowed with both high potency and selectivity. As future perspective, these molecules will be biologically tested as BSAO substrates to evaluate their cytotoxic effects and eligibility to be used in this new anti-cancer therapy. Details and methodologies will be reported