Abstract A256: Rational combination of the IGF-1R/IR tyrosine kinase inhibitor (TKI), OSI-906, with the MEK inhibitor, U0126, results in synergistic and apoptotic effects in human colorectal cancer (CRC) cell lines
Authors: Flanigan Sara A. , Pitts Todd M. , Kulikowski Gillian N. , Tan Aik Choon , Nallapareddy Sujatha , Tentler John J. , Eckhardt S. Gail , Leong Stephen 
Published: June 2012

Background: Signaling by the insulin-like growth factor 1 receptor (IGF-1R) promotes cell growth, migration and survival in several human tumor types. IGF-1R is overexpressed in CRC and is associated with a poor prognosis and resistance to chemotherapy. Our prior transcriptional profiling analysis of CRC cell lines treated with the small molecule IGF-1R/IR TKI, OSI-906, indicated that overexpression of the mitogen-activated protein kinase (MAPK) pathway conferred resistance to OSI-906. The purpose of this study was to evaluate the rational combination of the MEK1/2 inhibitor, U0126, and OSI-906 against human CRC cell lines.

Methods and Results: The antiproliferative effects of OSI-906 and U0126 were assessed as single agents and in combination using the Sulforhodamine B (SRB) cell viability assay. Twenty-eight CRC cell lines were exposed to either OSI-906 (0–5µM), or U0126 (0–20µM). In reference to OSI-906, cell lines with IC50 ≤ 1µM were considered sensitive (S) and cell lines with IC50 ≥ 5µM were deemed resistant (R). Likewise, cell lines with IC50 &lt; 5µM were regarded as S to U0126 and cell lines with IC50 &gt; 10µM were considered R. We selected 12 cell lines for combination studies according to the following conditions: 1) S to both drugs, 2) R to both drugs, 3) S to OSI-906 but R to U0126, and 4) R to OSI-906 but S to U0126, and treated them with varying doses of the two drugs as single agents and in combination. Combination effects between OSI-906 and U0126 were evaluated using the Chou and Talalay method. Robust synergy was observed in most CRC cell lines, including those that were resistant to OSI-906 or U0126. Apoptosis was then analyzed using bioluminescent caspase 3/7 detection and validated through analysis of PARP cleavage by immunoblotting. Surprisingly, some of the cell lines demonstrated induction of apoptosis when exposed to the combination but not with either agent alone. The CRC cell lines with the greatest apoptotic induction were inherently sensitive to OSI-906 in the SRB assay.

Conclusion: Our prior transcriptional profiling data revealed elevated expression of genes in the MAPK pathway in cell lines that were less sensitive to OSI-906, providing the basis for rational combination therapy with U0126. Interestingly, the combination of OSI-906 and U0126 displayed synergy in nearly all cell lines tested, regardless of sensitivity to either compound. Similarly, the apoptosis exhibited by some CRC cell lines could not be predicted by the sensitivity profile, or RAS/RAF mutational status. However, CRC cell lines with the greatest synergistic induction of apoptosis were sensitive to OSI-906. Further pathway and transcriptional profiling analysis is ongoing to reveal the underlying mechanism(s) of the observed synergy. These results, if further validated in vivo, will support the rational combination of OSI-906 and a MEK inhibitor in patients with CRC.

Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A256.