Abstract 3048: microRNA biomarkers associated with sunitinib-resistance in non-small cell lung cancer

Abstract

Abstract Background: Tyrosine kinase inhibitors (TKIs) are anticancer agents capable of specifically targeting tyrosine kinase receptors (TKR) often up-regulated in certain cancers. Sunitinib is a TKI acting on multiple TKRs including VEGFR, PDGFRβ, and FLT3. Sunitinib is approved for the treatment of advanced kidney and GIST cancers and is currently in clinical trials in other solid tumors, including NSCLC. We sought to identify in vitro microRNA (miRNA) biomarkers associated with sunitinib resistance (SR) in NSCLC cell lines. Methods: Sunitinib sensitivity was determined in seven NSCLC cell lines. Drug IC50s were determined using Alamar Blue proliferation assays. The sensitive NSCLC cell line H1703 was used to create SR cells by exposing three independent sets of cells to half the IC50 dose of sunitinib over several weeks. miRNA microarray profiling was performed on the three H1703 SR cell lines and the parent H1703 cells using the GenoExplorer miRNA Expression System (Sanger miRNA Registry version 14). Differences between the parent cells and the SR cells were identified using a t test with Benjamini-Hochberg correction and validated by qPCR. qPCR data were normalized to RNU6, and fold changes (FC) were computed against the parental H1703 line. A two-sided z-test was used to compute the p-values for the FC. To determine whether a miRNA can determine resistance across cell lines, a score was calculated using the following method. First, the FC were ternarized: significant increase (+1; FC >1 AND p-value < 0.05), significant decrease (−1; FC < 1 AND p-value < 0.05), non-significant FC(0). These values were then compared with the sunitinib IC50 of the cell lines: +1 on a resistant cell line or −1 on a sensitive cell line will be counted as one plus point, and otherwise as one minus point. A non-significant FC was not counted. Finally the score was computed as the total points divided by the number of cell lines. The “TKI resistance score” near “1” or “-1” suggests a good miRNA resistance indicator for the cell lines tested, Results: Three H1703 SR cell lines were successfully created with 2-7 fold increases in resistance as compared to the parental H1703 line. miRNA profiling identified up-regulation of miR-21, miR-23a, miR-23b, and miR-29 in the H1703 SR cells. TKI resistance scores based on qPCR data revealed miR-23b to be the best indicator of SR having a score of 1 when analyzing only the H1703 SR lines and a score of 0.67 when analyzing the H1703 SR lines together with the other six NSCLC lines. Conclusions: miRNA markers for SR exist in NSCLC cell lines as well as in H1703 cells with acquired SR. miR-23b appears to be the best potential biomarker for SR in NSCLC and if validated in clinical samples, may assist in enriching stratification for enrollment on NSCLC treatment clinical trials. Evaluation of miR-23b's mechanistic role in SR is ongoing. Supported by TGen Foundation, SHC Foundation, and the Flinn Foundation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3048.