A Screen for Selective Killing of Cells with Chromosomal Instability Induced by a Spindle Checkpoint Defect Zeeshan Shaukat1, Heidi W. S. Wong2, Shannon Nicolson1, Robert B. Saint2, Stephen L. Gregory1* 1 School of Molecular and Biomedical Sciences, University of Adelaide, Adelaide, South Australia, Australia, 2 Department of Genetics, University of Melbourne, Melbourne, Victoria, Australia Abstract Background: The spindle assembly checkpoint is crucial for the maintenance of a stable chromosome number. Defects in the checkpoint lead to Chromosomal INstability (CIN), which is linked to the progression of tumors with poor clinical outcomes such as drug resistance and metastasis. As CIN is not found in normal cells, it offers a cancer-specific target for therapy, which may be particularly valuable because CIN is common in advanced tumours that are resistant to conventional therapy. Principal Findings: Here we identify genes that are required for the viability of cells with a CIN phenotype. We have used RNAi knockdown of the spindle assembly checkpoint to induce CIN in Drosophila and then screened the set of kinase and phosphatase genes by RNAi knockdown to identify those that induce apoptosis only in the CIN cells. Genes identified include those involved in JNK signaling pathways and mitotic cytoskeletal regulation. Conclusions/Significance: The screen demonstrates that it is feasible to selectively kill cells with CIN induced by spindle checkpoint defects. It has identified candidates that are currently being pursued as cancer therapy targets (e.g. Nek2: NIMA related kinase 2), confirming that the screen is able to identify promising drug targets of clinical significance. In addition, several other candidates were identified that have no previous connection with mitosis or apoptosis. Further screening and detailed characterization of the candidates could potentially lead to the therapies that specifically target advanced cancers that exhibit CIN. Citation: Shaukat Z, Wong HWS, Nicolson S, Saint RB, Gregory SL (2012) A Screen for Selective Killing of Cells with Chromosomal Instability Induced by a Spindle Checkpoint Defect. PLoS ONE 7(10): e47447. doi:10.1371/journal.pone.0047447 Editor: Barbara Jennings, University College London, United Kingdom Received June 20, 2012; Accepted September 17, 2012; Published October 15, 2012 Copyright: 2012 Shaukat et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was funded in part by National Health and Medical Research Council grants 525477 and 1027878 (www.nhmrc.gov.au). No additional external funding was received for this study. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: stephen.gregory@adelaide.edu.au Introduction Chromosomal INstability (CIN) is a common feature of nearly all solid tumors [1]. CIN results in ongoing numerical and structural aberrations of chromosomes as tumors proliferate, and is associated with poor clinical outcomes such as tumour metastasis and adaptability to environmental and chemical stresses [2,3]. Drug resistance and relapse is common in cancers with CIN as they evolve rapidly, making them difficult to target with regular therapies [4,5]. The most common errors seen in CIN cancer cells are lagging chromosomes and chromosomal bridges. The mechanisms pro- posed to be responsible for these errors include defects in: sister chromatid cohesion [6], kinetochore–spindle attachment [7], cytokinesis [8], and centrosome duplication [9]. Perhaps the best-characterized cause of chromosomal instability is weakening of the Spindle Assembly Checkpoint (SAC) [10,11]. The SAC is the only mechanism by which cells can detect aberrant kinetochore attachments in metaphase and delay the entry to anaphase until the problem is resolved, or otherwise induce apoptosis [12,13]. This mechanism is not perfect, so mutations that cause high rates of segregation defects or shorten the duration of the metaphase error checking can cause CIN and lead to tumorigenesis [10,11]. The list of mutations known to have effects on chromosomal segregation is long, and includes over- and under-expression of spindle checkpoint proteins and clinically relevant cancer mutations such as loss of Rb in retinoblastoma or Adenomatous Polyposis Coli (APC) in colorectal cancer [11,14]. Reduction of the SAC protein Mad2 (mitotic arrest deficient 2) or its partner proteins (e.g. BubR1: Budding uninhibited by benzimidazoles Related 1) have been shown to shorten metaphase, causing CIN, aneuploidy and tumour susceptibility in humans and mouse models [15–17]. However, even in those CIN cancers that retain a SAC capable of detecting gross spindle abnormalities [18], the checkpoint is not able to respond to the merotelic kinetochore attachments that cause instability [19]. Loss of function of p53, which is common in cancers [20], increases the tolerance level for such missegregation, allowing the continual reassortment of the genome seen in CIN tumours [21,22]. CIN levels are higher in malignant tumors than in benign ones [23,24] and CIN is not found in normal cells, so it offers an attractive target for a cancer- specific therapy. CIN is particularly common in tumour types that are most in need of new drugs (e.g. colorectal cancers). Targeting CIN could also help to limit the ability of cancer cells to evolve PLOS ONE | www.plosone.org 1 October 2012 | Volume 7 | Issue 10 | e47447