Abstract 4868: A high-throughput allelic copy-number and somatic mutation platform utilizing a 75-ng DNA input in a 330,000 Molecular Inversion Probes (MIP) assay with FFPE samples

Abstract

Abstract Background: Copy number (CN) and somatic mutation studies of cancer hold great promise for the discovery of reliable biomarkers that can predict clinical outcomes. One of the challenges in such study is the majority of banked samples are formalin-fixed paraffin embedded (FFPE). Due to DNA degradation, FFPE samples generally perform poorly with CN technologies. However, MIP technology works well on FFPE samples, requiring only small stretches of intact genomic DNA (∼40 bp) in a 75-ng input. We have applied this 330,000-plex platform on thousands of FFPE samples from various tissues, e.g. breast, colon and brain, with 90% overall pass rate. Experimental Design: MIP probes were synthesized and screened for the performance. The selection criteria were: (1) good quantitative performance in reproducibility and dynamic range; (2) non-redundant whole genome coverage. Our platform employs a measurement comparing adjacent markers across the genome: this median of absolute pairwise distribution, or MAPD, is a reliable metric for assessing sample quality1. For array design, the best sequence was chosen based on empirical screening data. Results: Using only 75 ng of genomic DNA input, we have obtained both good genotyping and CN data, which is offered as a service-only product under the name “Oncoscan FFPE Express” by Affymetrix. For FFPE samples, good concordance between normal and tumor sample pairs is also observed1. Using an arbitrary cutoff of MAPD ≤ 0.6, a 2000-FFPE sample project has an overall pass rate > 92%. In addition to CN information, data for 400 frequent occurred somatic mutations in cancer listed in the Sanger COSMIC database are also generated in the same assay. They range from single nucleotide change to 10 bp insertion-deletion. Preliminary validation data by different platforms show 10% sensitivity (10% tumor with 90% normal) from a few key mutations (unpublished). While more validation is needed for every mutation interrogated, the current data show great promise of a complete solution for cancer mutation spectrum by a single assay. Conclusion: We have developed a powerful 330K MIP-CCN platform that works well on both frozen and archival FFPE samples. All three types of mutation (CN, SNP and somatic) can be interrogated by the same assay, offering unprecedented opportunity for retrospective study where only FFPE samples are available, and ongoing clinical trials where only small quantity of biopsy samples are accessible.