Abstract P6-12-09: Targeting deregulated cyclin E in inflammatory breast cancer via sequential combination therapy
Authors: Alexander A , Karakas C , Gong Y , Alvarez R , Ueno NT , Keyomarsi K 
Published: August 2014

Inflammatory breast cancer (IBC) is a rare, highly aggressive form of breast cancer, accounting for 2-5% of all breast cancers in the US and 10% of breast cancer deaths. Unfortunately, until recently this disease has not been studied extensively at the molecular/cellular level despite the compelling unmet need to develop more effective therapies. In spite of receiving aggressive multidisciplinary therapy (anthracycline-taxane-based neoadjuvant chemotherapy, modified-radical-mastectomy and adjuvant radiation therapy), only about 40% of IBC patients survive 5 years. The goal of this project is to identify new targets that can be exploited pharmacologically either in combination with current therapies or with other novel agents.

Previous work has identified tumor-specific low molecular weight forms of cyclin E, termed LMW-E, that are preferentially found in triple negative breast cancers (TNBC) and serve as a poor prognostic biomarker. As a result of cleavage, LMW-E preferentially accumulates in the cytoplasm where it exerts its pleiotropic oncogenic functions including deregulating the cell cycle. Furthermore, we have shown that TNBC cells harboring LMW-E are sensitive to CDK inhibitors.

To establish the clinical relevance of targeting the cyclin E/CDK2 axis in IBC, a pilot study was performed to determine the incidence of LMW-E. We performed IHC on 11 post-chemotherapy surgical specimens from MDACC. All tumors expressed cyclin E, 8 of which had predominantly cytoplasmic staining, indicative of LMW-E expression. These results are now being validated in a larger cohort of samples. These promising preliminary results led us to perform in vitro experiments to determine sensitivity to CDK2 inhibitors and characterize cell cycle profiles.

Similar to non-IBC breast cancers, IBC tumors can be ER-positive, TNBC or HER2-overexpressing; however, there are only TN-IBC or HER2+ cell lines. We utilized 2 cell lines for these studies, one of which is TN-IBC, the other of which is ER-negative and HER2-overexpressing. These cell lines were treated with a panel of CDK inhibitors to evaluate their sensitivity in both short-term and long-term assays. The long-term assay we developed called the high-throughput survival assay was used to examine combinations of agents for potential synergies using isobologram analysis. We have demonstrated chemotherapy-specific synergistic cell death when cells are first treated with CDK inhibitor in IBC and non-IBC cell lines.

Using flow cytometry, we have demonstrated that CDK inhibitors induce a G2/M arrest, which is further increased by sequential treatment with chemotherapy. Furthermore, combination treatment leads to cell death, as measured by sub-G1 DNA content.

Taken together, these studies provide solid evidence that targeting CDK2 is a promising strategy in treating IBC tumors that express cyclin E. Beyond these early pre-clinical advances, however, we demonstrated clear sequence-specificity for certain pairs of drugs, implying that future clinical studies should be designed taking into account the mechanism of action of cell cycle targeting drugs.

Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P6-12-09.