Radiosensitization of tumour cell lines by the polyphenol Gossypol results from depressed double-strand break repair and not from enhanced apoptosis.
OBJECTIVE
New drugs are needed to increase the efficiency of radiotherapy in order to improve the therapeutic outcome of tumour patients. In this respect, the polyphenol Gossypol might be of interest, because of its effect on apoptosis and DNA repair, which is either mediated directly or indirectly via the inositol phosphate metabolism. It was investigated, whether these effects result in enhanced radiosensitivity of tumour cells.


METHODS
Tumour cell lines investigated: A549, FaDu, H1299, MCF7 and Du145. Cell cycle distribution was determined by FACS analysis, apoptosis was measured by DAPI staining and caspase3/7 activity. Double-strand breaks (DSB) were investigated via gammaH2AX-foci and cell survival by colony formation assay. The level of inositol phosphates was determined by HPLC, protein expression by Western blot.


RESULTS
In A549 cells, Gossypol at concentrations 1microM strongly affects proliferation with only a modest arrest in the G1-phase, but with no increase in the fraction of apoptotic cells or the number of additional DSB. Additional DSB were only seen in FaDu cells, where Gossypol (2microM) was extremely toxic with a plating efficiency <0.002. When combined with irradiation, incubation with Gossypol (1-2microM) was found to result in an enhanced radiosensitivity with, however, a substantial variation. While there was a strong radiosensitization for FaDu and Du145 cells, there was an intermediate response for A549 cells, but almost no effect for H1299 and MCF7 cells. This sensitization was not caused from an elevated rate of apoptosis, but primarily resulted from reduced DSB repair capacity. The reduction in DSB repair could be ascribed neither to changes in the level of repair proteins relevant for non-homologous end-joining (Ku70, Ku80, DNA-PKcs) nor to changes in the level of higher phosphorylated inositols, whereby the latter were even found to be enhanced by Gossypol.


CONCLUSIONS
For some tumour cell lines treatment with low concentrations of Gossypol can be used to inhibit DSB repair capacity and with that to increase the cellular radiosensitivity.
Kasten-Pisula
Ulla
U
Laboratory of Radiobiology & Experimental Radiooncology, University Medical Center Hamburg-Eppendorf, Germany.
Windhorst
Sabine
S
Dahm-Daphi
Jochen
J
Mayr
Georg
G
Dikomey
Ekkehard
E
D017209:Apoptosis / Q000187:drug effects* / Q000528:radiation effects; D045744:Cell Line, Tumor; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects; D004260:DNA Repair / Q000187:drug effects; D005419:Flavonoids / Q000494:pharmacology*; D006072:Gossypol / Q000494:pharmacology*; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism / Q000528:radiation effects; D009369:Neoplasms / Q000188:drug therapy* / Q000235:genetics; D010636:Phenols / Q000494:pharmacology*; D059808:Polyphenols; D011836:Radiation Tolerance / Q000187:drug effects*; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*
D005419:Flavonoids; D007295:Inositol Phosphates; D010636:Phenols; D059808:Polyphenols; D011838:Radiation-Sensitizing Agents; D006072:Gossypol