Effects of PI3K inhibitor NVP-BKM120 on overcoming drug resistance and eliminating cancer stem cells in human breast cancer cells

The multidrug resistance (MDR) phenotype often accompanies activation of the phosphatidylinositol 3-kinase (PI3K)/AKT pathway, which renders a survival signal to withstand cytotoxic anticancer drugs and enhances cancer stem cell (CSC) characteristics. As a result, PI3K/AKT-blocking approaches have been proposed as antineoplastic strategies, and inhibitors of PI3K/AKT are currently being trailed clinically in breast cancer patients. However, the effects of PI3K inhibitors on MDR breast cancers have not yet been elucidated. In the present study, the tumorigenic properties of three MDR breast cancer cell lines to a selective inhibitor of PI3K, NVP-BKM120 (BKM120), were assessed. We found that BKM120 showed a significant cytotoxic activity on MDR breast cancer cells both in vitro and in vivo. When doxorubicin (DOX) was combined with BKM120, strong synergistic antiproliferative effect was observed. BKM120 activity induced the blockage of PI3K/AKT signaling and NF-κB expression, which in turn led to activate caspase-3/7 and caspase-9 and changed the expression of several apoptosis-related gene expression. Furthermore, BKM120 effectively eliminated CSC subpopulation and reduced sphere formation of these drug-resistant cells. Our findings indicate that BKM120 partially overcomes the MDR phenotype in chemoresistant breast cancer through cell apoptosis induction and CSC abolishing, which appears to be mediated by the inhibition of the PI3K/AKT/NF-κB axis. This offers a strong rationale to explore the therapeutic strategy of using BKM120 alone or in combination for chemotherapy-nonresponsive breast cancer patients.Multidrug resistance (MDR) remains a major cause for failure of chemotherapy-based treatment of breast cancer, in which cells become refractory to many structurally and functionally unrelated chemotherapeutic drugs.¹ Overexpression of P-glycoprotein (P-gp), a member of the ATP-binding cassette (ABC) transporter family encoded by the mdr-1 gene, represents one of the principal mechanisms that contribute to the MDR phenotype.² However, many other mechanisms contribute simultaneously to the MDR phenotype, which may affect drug absorption, distribution and metabolism, thus modulating the efficacy of chemotherapeutic agents consequently.^{3, 4} Growing evidence supports the notion that a subset of cancer cells, with self-renewal and differentiation features, are the cancer stem cells (CSCs) thought to be responsible for resistance to chemotherapy.⁵ CSCs seem to be protected against chemotherapeutic agents by means of different mechanisms, such as robust proficiency of DNA damage repair, overexpression of ABC transporters, abnormal activation of numerous signaling pathways, including phosphatidylinositol 3-kinase (PI3K)/AKT, Notch, Hedgehog and Wnt pathways.^{6, 7, 8} On the other hand, the CSC fraction is probably enriched after chemotherapy, as demonstrated by the increased expression of stemness markers in patients who are receiving primary systematic therapy.⁹The activation of the PI3K/AKT pathway is frequently implicated in resistance to anticancer therapies. Once activated, AKT can phosphorylate multiple substrates and downstream effectors, such as mTOR family, caspase family, cell cycle protein family and nuclear factor-κB (NF-κB), which contribute collectively to promote cell proliferation, survival, metastasis and chemoresistance.^{10, 11, 12} As this signaling cascade has a central role in human breast cancer, development of novel strategies to overcome resistance and eliminate CSC by targeting the PI3K/AKT pathway is apparently warranted.¹³NVP-BKM120 (referred hereafter as BKM120) is a potent and highly selective pan-class I PI3K inhibitor, which belongs to the 2,6-dimorpholino pyrimidine derivatives.¹⁴ It selectively inhibits wild type and mutant PI3K p110 α, β, δ, γ isoforms and exerts a strong antiproliferative effect to induce apoptosis in several cancers by specifically inhibiting the PI3K/AKT signaling pathway.^{15, 16, 17} Phase I clinical trials show that overall BKM120 is well tolerated in several solid tumors, and Phase II clinical trials are ongoing.¹⁷ Several recent reports also emphasized the enhanced antitumor effects in mouse models when BKM120 was co-treated with inhibitors of other signaling pathways.^{18, 19, 20}In this study, we analyzed, for the first time, the efficacy of BKM120 in several MDR breast cancer cell lines with which the MDR phenotype is induced by different molecular mechanisms. BKM120 exerted potent efficacy of apoptosis promoting as well as CSCs eliminating through inhibiting the PI3K/AKT/NF-κB cascade in vitro and in vivo. In addition, BKM120 synergized with DOX, a common chemotherapeutic agent of breast cancer. Here we demonstrate the potential of BKM120 in overcoming chemotherapy resistance in breast cancer.