aphidicolin and Skin-Neoplasms

Recently, the three-dimensional structure of the active site of human DNA polymerase alpha (pol alpha) was proposed based on the application of molecular modeling methods and molecular dynamic simulations. The modeled structure of the enzyme was used for docking selective inhibitors (nucleotide analogs and the non-nucleoside inhibitor aphidicolin) in its active site in order to design new drugs for actinic keratosis and squamous cell carcinoma (SCC). The resulting complexes explained the geometrical and physicochemical interactions of the inhibitors with the amino acid residues involved in binding to the catalytic site, and offered insight into the experimentally derived binding data. The proposed structures were synthesized and tested in vitro for their influence on human keratinocytes and relevant tumor cell lines. Effects were compared to aphidicolin which inhibits pol alpha in a non-competitive manner, as well as to diclofenac and 5-fluorouracil, both approved for therapy of actinic keratosis. Here we describe three new nucleoside analogs inhibiting keratinocyte proliferation by inhibiting DNA synthesis and inducing apoptosis and necrosis. Thus, the combination of modeling studies and in vitro tests should allow the derivation of new drug candidates for the therapy of skin tumors, given that the agents are not relevant substrates of nucleotide transporters expressed by skin cancer cells. Kinases for nucleoside activation were detected, too, corresponding with the observed effects of nucleoside analogs.

Topics: Aphidicolin; Apoptosis; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; DNA Polymerase I; Humans; Keratinocytes; Keratosis, Actinic; Models, Chemical; Models, Molecular; Necrosis; Nucleic Acid Synthesis Inhibitors; Nucleotide Transport Proteins; Protein Binding; Purines; Skin Neoplasms; Thymidine

The induction of micronuclei (MN) by incubation with different mutagenic agents was tested in diploid fibroblast cultures obtained from 15 probands with constitutively high MN rates (15.75-77.25 MN/500 cells; average 36.27 +/- 17.60 MN) and 15 probands (controls) with low MN rates (4-13.75 MN/500 cells; average 8.97 +/- 2.73 MN). In order to find out whether fibroblast cultures of individuals with increased spontaneous MN levels exhibit an increased sensitivity to various agents with different genotoxic mechanisms, we studied the induction of MN in these cell cultures by ultraviolet (UV) irradiation, mitomycin C (MMC), N-methyl-N-nitro-N-nitrosoguanidine (MNNG) and benzo-(a)pyrene-diol-expoxid (BPDE). In addition, we tested aphidicolin (APC), a polymerase alpha inhibitor, which is a potent inducer of common fragile sites. Probands with spontaneously high MN showed an significantly increased sensitivity to UV (p less than or equal to 0.005), MMC (p less than or equal to 0.005), and BPDE (p less than or equal to 0.005). No significant differences were found for MNNG and APC as compared to controls.

Topics: 7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; Aphidicolin; Cells, Cultured; Fibroblasts; Humans; Melanoma; Methylnitronitrosoguanidine; Mitomycin; Mutagens; Reference Values; Skin; Skin Neoplasms; Ultraviolet Rays