Compared to vector controls, cells with reduced NVP-BKM120 cost Metnase levels showed a 17-fold higher frequency of apoptosis after adriamycin exposure. This finding suggests that Metnase suppresses adriamycin-induced apoptosis, contributing to the increased resistance of breast cancer cells to this drug. We examined the effect of Metnase on adriamycin inhibition of Topo IIa-mediated decatention using a Temozolomide kinetoplast DNA in vitro decatenation assay. Topo IIa decatenates kDNA and adriamycin completely inhibits this activity. As shown previously, purified Metnase does not decatenate kDNA on its own, but enhances Topo IIa-dependent kDNA decatenation by 4-fold. Importantly, when Metnase is present, it overcomes the inhibition of Topo IIa by adriamycin, and this is true whether Metnase is added to the reaction before or after adriamycin. Note also that in the presence of Metnase, there is a greater level of decatentation in the presence of adriamycin than with Topo IIa alone 
in the absence of adriamycin. Metnase is a known component of the DSB repair pathway, and may enhance resistance to Topo IIa inhibitors by two mechanisms, enhancing DSB repair or enhancing Topo IIa function. The data presented here suggest that the ability of Metnase to interact with Topo IIa, and enhance Topo IIa dependent decatenation in vivo and in vitro may be at least as important as its ability to promote DSB repair in surviving exposure to clinical Topo IIa inhibitors. It is possible that Metnase could bind Topo IIa and physically block binding by adriamycin. In this model, Metnase would be bound to Topo IIa on DNA, and prevent adriamycin from stabilizing the Topo IIa/DNA cleavage complex, allowing Topo IIa to complete re-ligation. Alternatively, Metnase may function as a co-factor or chaperone to increase Topo IIa reaction kinetics. Here Metnase would bind transiently to Topo IIa and increase its reaction rate regardless of adriamycin binding. The mechanism may also be a functional combination of these two mechanisms where Metnase increases Topo IIa kinetics while also blocking further binding of the drug. Our interpretation of these data is that Metnase increases the intrinsic function of Topo IIa via one of the above mentioned molecular mechanisms, and that this will result in fewer DSBs, not necessarily from enhanced DNA repair, but from Topo IIa directly resisting adriamycin inhibition and thus inhibiting the production of DSBs. This model is supported by our findings that Metnase significantly blocks breast cancer cell metaphase arrest induced by ICRF-193, and that cellular resistance to Topo IIa inhibitors is directly proportional to the Metnase expression level. Our data reveal a novel mechanism for adriamycin resistance in breast cancer cells that may have important clinical implications. Metnase may be a critical biomarker for predicting tumor response to Topo IIa inhibitors. By monitoring Metnase levels, treatments with Topo IIa inhibitors may be tailored to improve efficacy. In addition, since reduced Metnase levels increase sensitivity to clinical Topo IIa inhibitors, inhibiting Metnase with a small molecule could improve response in combination therapies. Metnase inhibition may be especially important in a recurrent breast tumor that was previously exposed to Topo IIa inhibitors, since resistance to these agents may be due to upregulation of Metnase and/or Topo IIa. In summary, Metnase mediates the ability of Topo IIa to resist clinically relevant inhibitors, and may itself prove clinically useful in the treatment of breast cancer. Translationally controlled tumor protein is expressed in almost all mammalian tissues. Intracellular TCTP levels respond to various extracellular signals and agents such as growth factors, cytokines, and stress conditions. Extracellular TCTP has also been reported to be present in the supernatants of human U937 macrophage cell cultures, outside of mononuclear cells and platelets, in nasal washings, skin blister fluids, and bronchoalveolar lavage fluids during late allergic reactions.