Poster Presentation 26th Lorne Cancer Conference 2014

Molecular Analysis of Circulating Tumour Cells (CTCs) from the Blood of Patients with Colorectal Cancer (CRC) detected with the CellSearch System (#151)

Christin Gasch 1 2 , Thomas Bauernhofer 3 , Ellen Heitzer 4 , Martina Auer 4 , Prue Plummer 2 5 , Lidija Jovanovic 6 , Oliver Mauermann 1 , Jochen Geigl 4 , Michael Speicher 4 , Colleen Nelson 6 , Albert Mellick 2 5 , Klaus Pantel 1 , Sabine Riethdorf 1
  1. Department of Tumour Biology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
  2. School of Medicine, Deakin University, Geelong Waurn Ponds Campus, Geelong, VIC, Australia
  3. Department of Clinical Oncology, Medical University of Graz, Graz, Austria
  4. Institute of Human Genetics, Medical University of Graz, Graz, Austria
  5. Host Response to Cancer Laboratory, School of Medical Science, Griffith University, Gold Coast, QLD, Australia
  6. Australian Prostate Cancer Research Centre, Queensland University of Technology, Brisbane, QLD, Australia

Clinical data over the last 15 years has increasingly shown that there is a correlation between the presence of tumour cells circulating in the peripheral blood, and a poor prognosis for colorectal cancer (CRC) patients. However, further characterization of circulating tumour cells (CTCs) remains pivotal: not only to confirm their malignant origin but also to help identify diagnostically and therapeutically relevant targets for therapy; as well as to assist in stratifying cancer patients for individual therapies. Furthermore, while only a small subpopulation of CTCs might lead to a poor patient outcome, the genetic and phenotypic heterogeneity of CTCs and link with outcome remains poorly understood. To analyse clinically relevant genetic markers, we firstly isolated 141 CTCs (using micromanipulation following selection via the CellSearch System, Veridex LLC) from 13 CRC patients. We then applied techniques in whole genome amplification (WGA), on single CTCs individually, and analysed the material by array comparative genomic hybridisation (CGH: 19 CTC/5 patients), quantitative real time PCR (Q-PCR: 29 CTC/8 patients) and Sanger sequencing (93 CTC/12 patients). Following analysis we identified gene amplifications in key markers including EGFR, LGR5, AURKA and c-MYC; as well as mutations in p53, KRAS, BRAF and PIK3CA gene. Taken as a whole the data obtained in this work revealed an unexpected degree of genetic heterogeneity, even among CTCs from the same patient (Gasch et al. 2013). To follow up this study we have begun work to establish a protocol for RNA-in situ hybridisation (RNA-ISH) analysis to analyse CTC phenotype directly (esp. miRNA expression); and show that this can be performed directly in the CellSearch cartridge itself.
In conclusion, determining the genetic and phenotypic heterogeneity of CTCs, and link to patient outcome will be key to the development and rational application of future effective anticancer therapies.