Abstract 4229: The characterization of drug-tolerant cancer cell populations by Colony Lysate Array (CoLA)

Abstract

Abstract [Background] Increasing evidences indicate that heterogeneity within cancer cell population plays a critical role in response to cytotoxic stresses including drug treatment. We characterized human cancer cell populations that formed colonies in the presence of anticancer drugs at cellular and molecular levels. [Materials and Methods] For colony formation, human cancer cells (HCT-116, HeLa, HT-29, MCF-7, and MKN-45) were sparsely disseminated (10 cells/cm2) in the presence of anticancer drugs (cisplatin, docetaxcell, gefitinib, and sorafenib). To characterize the drug-tolerant individual colonies, we established colony lysate array (CoLA), a tool used for the protein expression analysis of a single-colony, using modified “reverse-phase” protein lysate microarrays (RPAs). [Results] For in vitro modeling of cancer relapse after curative treatment, we consistently observed cell populations that form colonies under 0.1- 10 μM drug concentrations. Interestingly, the 50% colony-inhibition concentration was consistently more than 10-fold higher than that of growth-inhibition for cell-based assays. CoLA assay allows one to use a lysate from a single colony for hierarchical clustering by a panel of cancer stem cell (CSC) markers. Unlike FACS analysis, CoLA is advantageous in detection of full fractions of proteins including those in the nucleus. Thus far, we have found that MKN-45 colonies in the presence of cisplatin expressed CD44, a putative CSC marker in a concentration-dependent manner. Pre-treatment of actinomycin D (AMD), an RNA synthesis inhibitor strongly suppressed MKN-45 colony formation in the presence of cisplatin; however cycloheximide, a protein synthesis inhibitor did not affect the colony formation. [Discussion] Differences in drug concentrations between the colony-inhibition and the growth-inhibition indicate that colony formation and simple cell growth are functionally different. The CD44 expression in MKN-45 colonies in the presence of cisplatin may suggest that these colonies emerged in the presence of drugs acquired a “CSC-like” phenotype. The suppression of colony formation by AMD may also suggest that the colonies require de novo RNA synthesis for the gain of drug resistant phenotype. [Conclusion] The drug-tolerant cell populations exhibit a CSC-like phenotype and require de novo RNA synthesis for the gain of drug resistant phenotype, possibly involved in the heterogeneity of cancer cell population. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4229. doi:1538-7445.AM2012-4229