Prodigiosins are hydrophobic molecules, very unstable in water solutions, and might diffuse freely through membranes and interact with the DNA with a preference for AT sites from the minor-groove promoting dsDNA cleavage event. Cells respond to DNA damage by activating cell cycle arrest, DNA repair, and in some circumstances, the triggering of apoptosis (Route 1). Prodigiosins also might be incorporated into the lipids bilayer of the plasmatic membrane, where by endocytosis it reaches the endosome compartment, uncouples vacuolar H+-ATPase (V-ATPase) through promotion of the H+/Cl- symporter, and induces neutralisation of the acid compartment of cells, inducing intracellular acidification and eventually apoptosis (Route 2). Apoptosis also might occur through the activation of an unidentified prodigiosin receptor or by the activation of a known death receptor, inducing caspase 8 activation and consequently, apoptosis (Route 3). Finally, it might diffuse freely through membranes and interact with the mitochondrial outer membrane, uncoupling Fo-F1-ATPase and therefore, inducing apoptosis (Route 4). In conclusion, the pathway followed would depend very much on the cell type studied, the drug concentration inside the cell, the hierarchy of the prodigiosin targets and the interaction of distinct pathways mentioned above.
DNA-binding molecules regulate mechanisms central to cellular function, including DNA replication and gene expression. The planar prodigiosin nucleus binds DNA by intercalation, while the methoxy group and ring nitrogens provide hydrogen-bonding sites to facilitate DNA binding. The cationic nature at neutral pH also provides electrostatic interaction with the negatively phosphate groups of the DNA helix. Prodigiosin is a DNA interacting agent, with a preference for AT sites from the minor-groove. In addition, it facilitates copper-promoted oxidative double strand (ds) DNA cleavage through reductive activation of Cu(II), by oxidation of the electron-rich prodigiosin molecule. Copper is an essential trace element distributed in all cellular organelles including nucleus, its levels are usually high in cancer. In dry non-cancerous breast tissue, the mean concentration of copper is 1.47 ppm, whereas the mean concentration increases to 5.12 ppm in cancerous tissue.
|FIG.14||A recent crystallographic study8 confirmed this scenario: Manderville and co-workers succeeded in obtaining crystals from a 1:1 mixture of prodigiosin and Cu(II), which show several remarkable features. As expected, all three nitrogen atoms of the ligand in complex are coordinated to the metal cation in a distorted square-planar arrangement; remarkably, however, the pyrrole ring C has been oxidized and attacked by an external nucleophile (water or MeOH from the medium).(FIG.14) The analogous Zn(II) complex, in contrast, contains two molecules of prodigiosin in the ligand sphere of the metal. Neither of the ligands has been oxidized and only two nitrogen atoms of each ligand are bound to the metal. (FIG.15) These results shed light on the specific action of copper and corroborate the previous assumptions as to the elementary steps responsible for the nucleaselike activity of certain metal–prodigiosin combinations in the oxidative degradation of DNA.||FIG. 15|
By far the most attractive biological response of mammals to pyrrole alkaloids of this type, however, lies in their immunosuppressive activity at doses that are not cytotoxic. Their efficacy in vivo has been demonstrated by several independent studies on animal models. Although a recent investigation indicates a rather steep dose-response curve of prodigiosin on lymphocytes, inhibition of proliferation for such cells is reached at nanomolar concentrations (IC50 3–8 ngmL-1). In mice, the ED50 was reported to be 1.5 mgkg-1 for 6 days, whereas acute systemic toxicity is observed at doses = 4 mgkg-1. Although this therapeutic window may be too narrow for direct clinical applications, these alkaloids constitute new leads in the search for supplementary immunosuppressive drugs. Most encouraging is the fact that the mechanism of action of prodigiosin and its congeners is distinctly different from that of the clinically used cyclosporin or FK 506, which act as calcineurine inhibitors and block the proliferation of T cells in the early G1 stages of the cell cycle. As a consequence, a synergistic effect on the immune system is observed if, for example, prodigiosin is administered simultaneously with either cyclosporin or FK 506, which presently dominate clinical immunosuppressive regimens for the treatment of patients after organ transplantation. The in vitro 60 human tumour cell panel of the National Cancer Institute Drug Discovery Program (NCI, Bethesda, MD) provides an interesting tool that is available on Internet at www.dtp.nci.nih.gov with the NSC Number: 47147-F. Prodigiosin has been screened with an average IC50 of 2.1 mM. Recently, an extensive chemical research programme was undertaken by D’Alessio and co-workers from Pharmacia & Upjohn, in order to obtain synthetic derivatives of prodigiosin and identify more active and less toxic drugs than natural prodigiosin compounds.