Development of a novel HAC-based “gain of signal” quantitative assay for measuring chromosome instability (CIN) in cancer cells

• DOI: 10.18632/oncotarget.7854
• Publication Date: 2016-03-03T03:16:30Z
• Authors: Jung-Hyun Kim; Hee-Sheung Lee; Nicholas C. O. Lee; Nikolay V. Goncharov; Vadim Kumeiko; Hiroshi Masumoto; William C. Earnshaw; Natalay Kouprina; Vladimir Larionov
• Keywords: Chromosome instability

// Jung-Hyun Kim 1 , Hee-Sheung Lee Nicholas C. O. Nikolay V. Goncharov 1,2 Vadim Kumeiko 2 Hiroshi Masumoto 3 William Earnshaw 4 Natalay Kouprina and Vladimir Larionov Developmental Therapeutics Branch, National Cancer Institute, NIH, Bethesda, MD, USA School of Biomedicine, Far Eastern Federal University, A. Zhirmunsky Institute Marine Biology, FEB RAS, Vladivostok, Russia Laboratory Cell Engineering, Department Human Genome Research, Kazusa DNA Research Kisarazu, Japan Wellcome Trust Centre for University Edinburgh, Scotland Correspondence to: Larionov, email: Keywords : chromosome instability, CIN, human artificial chromosome, HAC, anticancer drugs Received November 04, 2015 Accepted January 29, 2016 Published March 02, Abstract Accumulating data indicates that instability (CIN) common to cancer cells can be used as a target therapy. At present the rate mis-segregation is quantified by laborious techniques such coupling clonal cell analysis with karyotyping or fluorescence in situ hybridization (FISH). Recently, novel assay was developed based on loss non-essential (HAC) carrying constitutively expressed EGFP transgene ("loss signal" assay). Using this system, easily ranked their effect HAC loss. However, it problematic covert "loss into high-throughput screen identify mutations increase CIN levels. To address point, we re-designed HAC-based assay. In new carries shRNA against integrated genome. Thus, inherit display no green fluorescence, while lacking do. We verified accuracy "gain measuring level induced known antimitotic added list previously inducing compounds, two newly characterized inhibitors centromere-associated protein CENP-E, PF-2771 GSK923295 exhibit highest measured date. The also sensitive enough detect after siRNA depletion genes controlling mitotic progression through distinct mechanisms. Hence utilized future experiments uncover genes, which will provide insight pathogenesis cancer. Also described possible conversion using microplate reader characterize chemical libraries conditions modulate level.