Tumours have a much greater dependence than normal tissues on anaerobic glycolysis for energy generation. We have studied the effects of glycolysis inhibition on tumour cells in vitro. Cellular ATP fell during exposure of cells in air to 2-deoxy glucose or to the lactate dehydrogenase inhibitor oxamate. Glycolysis inhibition alone did not alter clonogenic cell survival over 6 h, but a 6-h exposure to oxamate combined with doxorubicin (Dox) gave greater than additive cell killing. This effect was greatest when oxamate was dosed after Dox, suggesting that oxamate inhibited repair of Dox-induced damage. Oxamate also gave greater than additive cell killing in multicellular spheroids when combined with Dox and there was a greater than additive growth delay in spheroids dosed with Dox plus oxamate. These data demonstrate that inhibition of anaerobic glycolysis might be used to obtain a significant therapeutic gain in combination treatments with cytotoxic drugs or radiotherapy.