The synthetic compound, T0901317, is an LXR agonist commonly used to test LXR function. The influence of T0901317 and LXR in mediating cancer cell proliferation has been tested in many cell lines, including ovarian cancer cells [26
]. We aimed to test how T0901317 may enhance cisplatin efficacy as a combination treatment. However, unlike previous reports [26
], we observe that T0901317 has very little influence on cell growth when used alone, but significantly increases the IC50
of both cisplatin and paclitaxel it is added in combination. In the A2780-cp cell line, selected for drug resistance, the effect of T0901317 is abrogated, suggesting that T0901317-regulated pathways are important contributors to cisplatin sensitivity in the parental A2780 cell line. Together, these data suggest that T0901317 is not an effective anti-tumor therapeutic for some ovarian cancer cells, but may instead be harmful, as it appears to actually reduce the efficacy of cisplatin and paclitaxel.
To investigate the mechanisms of T0901317's influence on the cell cycle in ovarian cancer cell lines, we probed the protein levels of cell cycle and survival regulators. These data suggest an apparent paradox of T0901317 activity. CDK4, cyclin D1, and cyclin D3 are all significantly increased upon T0901317 treatment, consistent with T0901317 increasing the number of viable cells. At the same time, cell cycle inhibitors such as p27 and p21, which often are up-regulated from toxic compounds such as cisplatin [24
], are significantly increased. When co-treated with cisplatin, T0901317 significantly increases the expression of CDK4, cyclin D1, and cyclin D3, likely promoting the observed survival and reduced apoptosis. These data suggest that T0901317 is activating both pro-growth and pro-apoptotic signals, but that its action leads to a net decrease in cisplatin efficacy on some ovarian cancer cells.
The observation that myriad LXR ligands do not affect ovarian cancer cells in the same fashion as T0901317 suggests that T0901317 is not working through LXR. LXR is expressed in these cell lines and T0901317 stimulates the up-regulation of known LXR targets, suggesting LXR signaling is activated. However, none of the other LXR ligands showed similar significant effects on cell viability and drug response, suggesting that T0901317 may not be reducing chemotherapy efficacy through an LXR mediated mechanism. PXR and FXR agonists also showed no significant effects, suggesting that T0901317 affects drug response in ovarian cell lines by an LXR, PXR, and FXR independent mechanism. SiRNA knock-down of LXRα and LXRβ did not affect the number of viable OVCAR-8 cells (data not shown), further suggesting that LXR activity does not have significant effects on cell proliferation in OVCAR-8. The concentration of T0901317 required to yield the effect on cisplatin efficacy is also indicative of LXR independence. Typically, T0901317 induces activation of LXR at concentrations <1 μM. We have seen no significant effects of T0901317 at concentrations this low (data not shown), suggesting that the effects presented here, seen at treatments between 1 and 10 μM, are likely independent of LXR.
Together, these data suggest that T0901317 increases ovarian cancer cell survival and drug resistance, likely in an LXR independent manner. These data differ from previous reports of anti-proliferative effects of T0901317 where very high concentrations were used [12
]. At concentrations above 20 μM, we also observe T0901317 anti-proliferative effects. We observe T0901317 protective effects in the 1-10 μM range. We have performed these experiments using multiple lots of T0901317 and have observed very consistent data from both relatively early and late passage cells. Because we see T0901317 killing MCF-7 cells, we believe that the T0901317 we are using is effective to reduce the number of viable cells for some cell lines, as previously reported [13
]. We also observe T0901317 stimulating p27 and p21 expression, similar to previous reports [26
], but these changes do not seem to manifest as changes in the number of viable cells in the 1-10 μM range. These earlier studies of T0901317's effect on ovarian cancer cells did not address its effect in combination with cisplatin, where we observe a significant reduction in efficacy, suggesting that T0901317 has different activities at low and high concentrations.
We have identified new effects of T0901317 in mediating cytotoxic drugs in ovarian cancer cells. Future efforts to investigate these concentration dependent effects may uncover LXR dependent and independent pathways mediating ovarian cancer cell growth and apoptosis, and may lead to new mechanisms controlling chemotherapy. Identifying these key factors and mechanisms mediating T0901037 and LXR phenotypes will be critical to translating these observations into advances in the clinical treatment of ovarian cancer.