Effect of Transient Hypoxia on Sensitivity to Doxorubicin in Human and Murine Cell Lines

Abstract

Overreplication of DNA associated with gene amplification and drug resistance has been reported to occur after transient hypoxia of rodent cells. Because oxygen levels fluctuate in solid tumors, clinical drug resistance might be stimulated by this mechanism. We have therefore studied the effect of transient hypoxia on sensitivity to doxorubicin in human and murine cell lines. Exposure to hypoxia led to a decreased rate of cell proliferation, and most of the observed changes in sensitivity to doxorubicin were consistent with cell cycle-dependent cytotoxicity of this drug. After transient hypoxia, about 10% of the murine cells (EMT6/Ro and KHT-LP1) contained greater than four times the haploid DNA content (greater than 4C DNA), but only 0%-5% of the human cells (MGH-U1, A549, and Hey) had greater than 4C DNA content. Murine cells that had been exposed to hypoxia and reoxygenation, and which had greater than 4C DNA content, were separated by flow cytometry. For KHT-LP1 cells, but not for EMT6/Ro cells, this subpopulation was found to be more resistant to doxorubicin than the subpopulation with less than 4C DNA content and the aerobic control. When resistant KHT-LP1 clones were expanded in the presence of doxorubicin, six of six clones showed amplification of the P-glycoprotein gene family. The ability and efficiency of hypoxia to induce DNA overreplication, gene amplification, and drug resistance appears to be cell-line dependent.

Induction of multiple double-strand breaks within an hsr by meganucleaseI-SceI expression or fragile site activation leads to formation of double minutes and other chromosomal rearrangements

“…We previously demonstrated that oxygen privation (hypoxia), an important parameter of the tumor microenvironment (Brizel et al, 1996;Graeber et al, 1996;Kinzler and Vogelstein, 1996), which enhances gene amplification frequency (Luk et al, 1990;Rice et al, 1986), induces breaks at fragile sites . Here we show that activation of fragile sites, by drugs such as aphidicolin or by hypoxia, induces hsr breakdown as do I-SceI induced DSBs.…”

Carbonic anhydrase-9 expression levels and prognosis in human breast cancer: association with treatment outcome

“…Hypoxic cells are particularly radioresistant (Höckel et al, 1996b), but hypoxia has also been associated with resistance to chemotherapy (Luk et al, 1990). Not only does tumour hypoxia directly attenuate therapy success, but it also leads to clonal selection of more aggressive cells leading to poor prognosis, independent of treatment modality (Höckel et al, 1996a;Rofstad, 2000).…”

Hypoxia-induced angiogenesis and vascular endothelial growth factor secretion in human melanoma

“…Hypoxia-response elements governing the increased gene expression in response to hypoxia have been discovered in the vicinity of most of these genes (Dachs and Stratford, 1996). Moreover, tumour cells subjected to transient hypoxia in vitro can show increased metastatic potential in vivo (Young and Hill, 1988) and increased resistance to some chemotherapeutic agents (Rice et al, 1987;Luk et al, 1990;Sanna and Rofstad, 1994). Exposure of tumour cells to transient hypoxia in vitro can also induce cell subpopulations showing a slightly increased DNA content (Rice et al, 1986;Wilson et al, 1989) or a doubling of the number of chromosomes (Rofstad et al, 1996).…”