Conferences are so packed that it’s often difficult to go through every poster presentation. Now that we’re all back from New Orleans after an amazing AACR conference this year. We’ve summarized the posters we presented at AACR, so you can download and enjoy them from the convenience and comfort of your home or office.
1] A next generation sequencing based sample-to-result pharmacogenomics research solution enables both SNV and CNV detection at once.
The Ion AmpliSeq™ Pharmacogenomics Research Panel covers 40 commonly known targets in genes encoding drug metabolism enzymes and associated transport proteins (Table 1). The panel design was particularly challenging due to the high levels of sequence homology between cytochrome P450 genes. This assay uses Ion AmpliSeq™ technology and contains 119 amplicons in a multiplex single-pool PCR followed by Ion Torrent™ semiconductor sequencing. In addition, the panel includes 9 primer pairs for sample Identification. The assay requires as little as 10 ng of input DNA and is customizable for target addition or removal.
2] Global Gene Expression Profiles from Bladder Tumor FFPE Samples
RNA extracted from formalin fixed paraffin embedded tissue (FFPE) creates considerable additional challenges in acquiring accurate gene expression measurements due to the highly fragmented and compromised integrity of FFPE RNA due to the fixation process. To address the challenges of current sequencing based methods and take advantage of the simplicity of analysis that comes with using technologies such as microarrays; we have tested the Ion AmpliSeq™ Transcriptome Human Gene Expression Kit using RNA isolated from bladder tumor FFPE samples. Research using this targeted RNA sequencing approach allows profiling the global mRNA expression of human RNA in a highly multiplexed fashion using the Ion AmpliSeq™ technology. The Ion AmpliSeq™ Transcriptome Human Gene Expression Kit is a simple research method to measure global gene expression profiles from human RNA samples in a timely, cost effective, and high throughput manner resulting in sensitive and accurate gene expression measurements. The new S5™XL System combined with automated library and template preparation on the Ion Chef™ system enable a simple RNA to gene expression data workflow requiring only 45 minutes of hands on time from 10ng of FFPE RNA.
3] Complete workflow for detection of low frequency somatic mutations from cell-free DNA using Ion Torrent™ platforms
Research detection of somatic mutations in circulating cell-free DNA (cfDNA) using blood samples from subjects previously diagnosed with cancer provides a potential non-invasive approach to monitor cancer status and evaluate cancer evolution in the future. However, most of the existing mutation detection methods show insufficient sensitivity to detect cfDNA mutations since only small amount of mutant gene fragments, derived from tumor cells, is present in a large amount of normal circulating DNA background. Therefore, improvement in the accuracy of measurement systems which includes library preparation method, sequencing, and data analysis, is the key to enable variant detection at sub 1% allelic frequencies. In this study, we developed a highly sensitive and reliable research workflow to detect rare somatic mutations in circulating cfDNA samples. Significant overlapping of mutations discovered in FFPE tumor and cfDNA samples suggests that this workflow may be used to monitor tumor dynamics in NSCLC and potentially other tumors in the future.
4] Clinical research results for a NGS based kit for targeted detection of relevant gene rearrangements in lung tumor samples
Since NGS technologies enable analysis of mutations down to the nucleotide level, it may as a sensitive alternative to current practices used to detect fusion genes from tumour samples. Here, we present the analyses from the testing of one such NGS based technology. This set of kit reagents included for these analyses include an oligonucleotide panel which will simultaneously measure the presence of >75 possible fusion targets containing ALK, RET, ROS-1, and NTRK1 driver genes. Verification of this kit was carried out using the Ion Torrent™ Personal Genome Machine (PGM™) NGS platform and was accomplished through collaboration with three external clinical laboratories located in Portugal, Ireland and the United States.
5] Multiplex TaqMan Assays for Rare Mutation Analysis Using Digital PCR
Detection of rare mutations in tumor tissue and cell free DNA (cfDNA) allows for monitoring of tumor progression and regression for research purposes. cfDNA isolated from plasma combined with a sensitive detection method like digital PCR is non- invasive and enables earlier detection compared to conventional imaging techniques. Building on the TaqMan based Rare Mutation assay set for detection of rare mutations using digital PCR on the QuantStudio™ 3D Digital PCR System, we are now developing multiplex assays for simultaneous detection of several mutations. We selected relevant mutations in the EGFR and KRAS genes for our initial multiplex application: EGFR G719, EGFR exon 19 deletions, and KRAS G12/G13. These mutations may have implications for potential future targeted therapy. undertook a feasibility study multiplexing TaqMan Rare Mutation Assays covering three important mutational loci to accommodate limited amounts of sample frequently encountered in research involving cfDNA or biopsy material. We selected mutations in the EGFR and KRAS genes for our initial multiplex application: EGFR G719, EGFR exon 19 deletions, and KRAS G12/ G13. These mutations may have implications for potential future targeted therapy.
6] Applied Biosystems™ Minor Variant Finder: New Software for Detecting Somatic Mutations at Low Level in Sanger Sequencing Traces
A minor variant is a heterozygous allele in which the proportion of the secondary allele is less than a classical Mendelian germ line heterozygous variant of 50:50. Detecting minor variants has become essential to combating, managing, and advancing our understanding of cancer, inherited and infectious diseases, and the impact of heteroplasmy on mitochondrial diseases. We have developed software that detects and reports 5% minor variants in Sanger Sequencing traces at 95.3% sensitivity and 99.8% specificity. The software calls variants without prior knowledge of location and affords the advantages of Sanger sequencing, of robustness, low error rate, ease of use, human interpretable visual displays of the data, and low cost per sample and target. The software can confirm somatic variants found by NGS.
You can access and download all our posters from AACR 2016 here. We can’t wait to see you again at the next conference!