Abstract 2256: Establishment and characterization of circulating tumor cell-derived xenografts in non-small cell lung cancer
Authors: Faugeroux Vincent , Deas Olivier , Michels Judith , Judde Jean Gabriel , Cairo Stefano , Vielh Philippe , Marty Virginie , Billiot Fanny , Ngocamus Maud , Besse Benjamin , Kannouche Patricia , Farace Françoise 
Published: July 2016

Low numbers of circulating tumor cells (CTCs) have so far limited the establishment of CTC-derived xenografts (CDXs) to improve our understanding of tumor progression, drug resistance mechanisms, and their biological properties. We report the establishment and the phenotypic and molecular characterization of one NSCLC CDX.

Blood samples (30 ml) were drawn from 49 NSCLC patients with advanced metastatic disease. CTCs were enriched by RosetteSep, embedded in matrigel, and implanted in the interscapular aspect of NSG mouse (Nod/Scid-IL2Rγ-/-). Mice were followed-up for one year according to ethical regulations. CDX tumours and CDX derived cell lines were phenotypically and molecularly characterized by immunofluorescence, immunohistochemistry, CGHarray, exome sequencing and transcriptome gene expression.

CTCs from one NSCLC patient with 750 CTCs detected by CellSearch gave rise to a tumor 5 months after initial murine injection. Histological analysis confirmed the human origin of the tumor and the presence of a poorly differentiated adenocarcinoma consistent with the patient's biopsy. Tumor was positive for EpCAM, EMA, CK8;18, and Ki67, and negative for vimentin. A fraction of cells (25%) from freshly dissociated tumors exhibited ALDH activity. CGH from CDX tumors at passage 1 and 2 shows multiple gene rearrangement, revealing a high degree of genomic instability. Transcriptome analysis of ALDH positive and negative cells is ongoing and should help of identifying a cancer stem cell gene expression signature. Whole-exome sequencing of CDX tumor is ongoing and will be compared to data obtained from single CTCs from the patient.

A cell line established in vitro from the CDX model grows in 3D clusters and is tumorigenic in mice. Interestingly, this cell line is positive for cytokeratins, EpCAM, E-cadherin, N-cadherin, vimentin, and expresses multiple cancer stem cell markers including CD166, CD24, CD133 and, ALDH activity. The cell line is hypotetraploid (about 70 chromosomes) and its CGH profile was similar to that of the CDX tumour, revealing a high level of genome instability. By investigating DNA replication process in this cell line, we found that it exhibits a spontaneous enhanced DNA damage signaling associated to an accumulation of DNA double strand breaks mainly in S phase strongly suggesting that the CDX-derived cell line displays hallmarks on replication stress that could explain, at least partially, the genomic instability in the cells.

We report a low success rate in the establishment of NSCLC CDX (2%). However one NSCLC CDX model harboring cancer stem cell properties and deficiency of DNA replication maintenance was established. Ongoing work to identify a cancer stem cell signature and characteristics replication stress markers in this CDX model will be presented. This NSCLC CDX model will be useful to test drugs targeting these alterations in vivo and improve our knowledge of drug resistance.

Citation Format: Vincent Faugeroux, Olivier Deas, Judith Michels, Jean Gabriel Judde, Stefano Cairo, Philippe Vielh, Virginie Marty, Fanny Billiot, Maud Ngocamus, Benjamin Besse, Patricia Kannouche, Françoise Farace. Establishment and characterization of circulating tumor cell-derived xenografts in non-small cell lung cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2256.