Poster Presentation 25th Lorne Cancer Conference 2013

Whole exome sequence analysis of an in vitro model of drug-resistant T-cell acute lymphoblastic leukaemia (#151)

Mark N Cruickshank 1 , Jette Ford 1 , Alex Gout 1 , Richard Francis 1 , Rishi S Kotecha 1 2 3 , Alex H Beesley 1 , Ursula R Kees 1
  1. Telethon Institute for Child Health Research, Subiaco, WA, Australia
  2. Department of Haematology and Oncology, Princess Margaret Hospital for Children, Perth, WA, Australia
  3. School of Paediatrics and Child Health, University of Western Australia, Perth, WA, Australia

The detection of chromosomal abnormalities in childhood leukaemia provides important prognostic markers used to guide treatment. Cytogenetic risk stratification for paediatric B-cell acute lymphoblastic leukaemia (ALL) is well established and is essential for selection of therapy. In contrast, genetic alterations predicting response to chemotherapy in paediatric T-cell ALL (T-ALL) are poorly defined. We are investigating the molecular mechanisms of sensitivity and resistance to a pan-cyclin dependent kinase (CDK) inhibitor, flavopiridol, for the treatment of leukaemia. We have found that flavopiridol is highly cytotoxic in T-ALL cell lines. We generated a series of leukaemia cells resistant to flavopiridol by selection of the T-ALL line PER-255 with pulsed high-dose or continuous low-dose flavopiridol exposure. We isolated clonal populations of flavopiridol-resistant T-ALL cells by limiting dilution and used whole exome sequencing to identify sequence alterations among resistant and sensitive clones. Using Illumina TruSeqTM Exome Enrichment and the HiSeqTM 2000 sequencing platform, we observe 82.3-83.5% of reads aligning to targeted genomic sequence (n=4 resistant PER-255; n=2 sensitive PER-255) with mean coverage range of 59-69. We examined the allele frequencies of single nucleotide variants (SNVs) and predicted functional consequences of structural alterations including SNVs, insertions and deletions associated with resistance to the pan-CDK inhibitor. Our data identifies common variants to the coding genome of independently derived flavopiridol-resistant PER-255 cells and also provides evidence for heterogeneity within a population of resistant PER-255 cells. Genes containing predicted deleterious variants associated with resistance to the CDK inhibitor, include cell-cycle regulators, which may represent novel therapeutic targets for drug-resistant T-ALL. Further studies to explore if these variants arise in T-ALL patients who develop drug-resistance are expected to reveal the clinical relevance of genetic alterations identified in this model system.