Differential sensitivity of double minute chromosomes to hydroxyurea treatment in cultured methotrexate-resistant...

Treating mammalian cells with continuous sub-lethal doses of Hydroxyurea (HU) causes the loss of double minute chromosomes (DMs) containing amplified oncogenes in culture. Recently, we have shown that treating glioblastoma multiforme cells in culture with low doses of HU causes the loss of DMs containing epidermal growth factor receptor genes. Loss of amplified EGFR genes was accompanied by cessation of growth, and greatly decreased tumorigenicity. To further study HU-induced elimination of DMs we have now followed the fate of dihydrofolate reductase gene (DHFR) amplifying DMs in methotrexate-resistant mouse cells during simultaneous treatment with both MTX and HU. We report that in the presence of both HU and MTX, the amplified genes decreased to 25% of starting levels in the first week of treatment, but that ultimately the cells become resistant to HU and reamplify the DHFR gene. We also report that some DHFR amplifying DMs are much more sensitive to HU than others. This study demonstrates that HU does not simply increase the rate of passive loss of DMs.