Abstract LB-286: The patient-derived xenograft mouse model dilemma; a new solution for an old problem
Authors: Schneeberger Valentina , Poirier John , Rudin Charles 
Published: August 2015

Patient derived xenograft (PDX) mouse models have become an essential tool for cancer research. Our laboratory was one of the first to develop patient derived xenografts of multiple lung cancer types and has used these models extensively for therapeutic analysis in vivo. We previously demonstrated that lung PDXs more closely recapitulate patterns of gene expression found in primary tumors while revealing that traditional cell line xenografts are genetically divergent from the tumor of origin, and thus may not accurately model human disease. One disadvantage of PDX mouse models is the admixture of human tumor cells with murine stroma during growth in vivo. This phenomenon has important implications on genomic sequencing including wasteful sequencing of mouse DNA and false positive mutation calls in reads that map to both the human and mouse reference genomes. Bioinformatic approaches to mapping human reads obtained in the context of admixed mouse DNA can be helpful, but insufficient in the context of xenograft tissue with significant stromal content. We therefore evaluated immunomagnetic approaches to tumor cell purification prior to next generation sequencing.

We tested the Miltenyi mouse cell depletion kit (MCD), the STEMCELL technologies EasySep EpCam positive selection kit and the Miltenyi EpCam positive selection kit for their ability to purify human tumor cells from mouse stroma using a quantitative PCR and fragment analysis based approach, using fluorescence activated cell sorting (FACS) as a gold standard for purity. In our hands, the Miltenyi MCD kit yielded consistently higher purity across a range of stromal contamination levels. Prior to purification, samples ranged from 20% to 95% human tissue composition and were consistently purified to greater than 93% purity after MCD.

DNA from samples representing germline DNA, input, FACS sorted, or MCD depleted cell suspensions was then subjected to next generation sequencing by MSK-IMPACT, a targeted sequencing approach that enriches for exonic regions in 341 genes. All samples were sequenced to &gt;800x average depth. Immunomagnetic depletion of mouse stromal cells resulted in identical mutation and copy number variation calls as FACS purified samples, while eliminating spurious false positive mutations caused by contaminating mouse DNA, demonstrating the effectiveness of this technique.

The increasing use of PDX mouse models for the study of cancer calls for the development of tools that will ensure the data produced by these samples is of the best quality possible. This project describes a novel technique with far reaching applications.

Citation Format: Valentina Schneeberger, John Poirier, Charles Rudin. The patient-derived xenograft mouse model dilemma; a new solution for an old problem. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-286. doi:10.1158/1538-7445.AM2015-LB-286