Abstract 3947: Intratumor heterogeneity impacts treatment response in rectal carcinoma
Authors: Meyer Rüdiger , McNeil Nicole E. , Wangsa Darawalee , Anthuber Lena , Auslander Noam , Wangsa Danny , Zhang Zhongqiu , Rosenberg Daniel , Ruppin Eytan , Habermann Jens K. , Ried Thomas 
Published: July 2017

Abstract
Individual response to radiochemotherapy (RCT) in rectal cancer patients is highly variable and the underlying mechanisms of treatment resistance of cancer cells are poorly understood. Recent studies revealed a considerable degree of genomic tumor heterogeneity. We hypothesize that this heterogeneity has a direct impact on treatment response as subpopulations of cancer cells are resistant to currently used RCT and facilitate tumor growth under treatment. To address this highly relevant clinical issue, patient-derived rectal cell lines were established. The tissue was derived from a 59-year-old male presenting with an adenocarcinoma of lower rectum (T3,N1,M0), who was treated by neoadjuvant RCT (50.4 Gy plus 5-FU) and low anterior resection. The neoadjuvant treatment induced a shrinkage of the tumor (staging of the surgical specimen: ypT3,N0), suggesting a partial response to RCT. A biopsy of the treatment naïve tumor was implanted heterotopically into an immunodeficient nude mouse. After in vivo growth, the tumor was dissociated and introduced to an in vitro culture where murine stromal cells were depleted using antibody-based columns. The cell line proved to be of human origin by immunofluorescence for human Keratin 20 as well as genotyping using short tandem repeat profiling. The characterization of the genome by array CGH and spectral karyotyping (SKY) revealed a highly complex near-triploid karyotype with numerous chromosomal imbalances, which are specific for colorectal cancers, including gains in chromosomes 7, 13 and 20. We also observed other structural abnormalities including an isochromosome 1q. Multiplex-FISH by consecutive hybridization of probes for 15 gene loci, which are known to be relevant in colorectal cancer genesis, revealed a significant degree of clonal heterogeneity of the cell line. Single cell sorting was then performed to establish single cell derived cell lines from the genomically well characterized parental cell line. Exposure of the single cell derived cell lines to irradiation in combination with 3 uM 5-FU revealed substantial differences in treatment response. Analyses of the genome by array CGH and transcriptome by RNA-Seq of the respective single cell derived cell lines are currently underway. This will allow the comparison of the respective sensitivities to RCT to the identified aberration profiles. These data will facilitate the understanding of therapy resistance and potentially allow a reliable prediction of the patient’s response. A tailored therapy is an important step towards individualized treatment in colorectal cancer patients to avoid therapy resistance.
Citation Format: Rüdiger Meyer, Nicole E. McNeil, Darawalee Wangsa, Lena Anthuber, Noam Auslander, Danny Wangsa, Zhongqiu Zhang, Daniel Rosenberg, Eytan Ruppin, Jens K. Habermann, Thomas Ried. Intratumor heterogeneity impacts treatment response in rectal carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3947. doi:10.1158/1538-7445.AM2017-3947