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Ion Torrent Oncomine cell-free nucleic acids assays are multi-biomarker next-generation sequencing (NGS) assays that enable reproducible detection of somatic mutations in plasma, down to a level of 0.1% in genes relevant to solid tumors. When used as part of a complete liquid biopsy NGS workflow, the road from research blood sample to variant data only takes two days.
Each assay contains the reagents for library construction and a single pool of primers used to perform multiplex PCR for preparation of amplicon libraries from cell-free DNA (cfDNA) and cell-free RNA (cfRNA) found in the plasma fraction of whole blood, and each is part of a complete research solution to detect tumor-derived DNA and RNA from cell-free nucleic acids.
Using a single tube of blood, our assays are designed for analysis of single nucleotide variants, short indels, copy number variations, and fusions that are frequently mutated in research cancer samples. The assays enable a limit of detection (LOD) down to 0.1%.
Learn how the assay performs against commercially available pre-characterized reference materials and clinical plasma samples.
Liquid biopsies offer several advantages over conventional solid tumor samples:
We now know that cancer is a molecular disease. To better advance cancer research in the future, we need a clearer understanding of how those oncogenes and oncoproteins change in both time and space. Liquid biopsy applications enable clinical researchers to investigate tumors in ways not previously possible.
In the past, clinical cancer research on cfDNA focused on digital PCR or droplet digital PCR applications to analyze samples with low-frequency mutations. These approaches, while sensitive, limit the number of target genes, thereby limiting the field of vision of the researcher.
The Oncomine Cell-Free Research Assays and the Ion GeneStudio S5 Systems enable tumor heterogeneity research, therapy selection research, therapy monitoring research, and reoccurrence research studies from as little as 1ng of sample input.
“Using the Oncomine Lung cfDNA Assay, we were able to amplify more difficult samples. The assay allowed us the detection of mutations like EGFR T790M down to 0.14% allelic fraction.”
José Luis Costa, PhD
Senior Researcher
Institute of Molecular Pathology and Immunology of the University of Porto, Portugal
Oncomine cell-free research assays include targets identified by the Oncomine Reporter, a cancer genomics data resource, and reviewed by clinical researchers.
Assay | DNA/RNA | Gene | Selected SNV hotspots | CNVs | Fusions | Extra | |
DNA & RNA | AKT1 ALK APC AR ARAF BRAF CCND1 CCND2 CCND3 CDK4 CDK6 CHEK2 CTNNB1 DDR2 EGFR ERBB2 ERBB3 ERG ESR1 ETV1 FBXW7 FGFR1 FGFR2 FGFR3 FGFR4 FLT3 | GNA11 GNAQ GNAS HRAS IDH1 IDH2 KIT KRAS MAP2K1 MAP2K2 MET MTOR MYC NRAS NTRK1 NTRK3 PDGFRA PIK3CA PTEN RAF1 RET ROS1 SF3B1 SMAD4 SMO TP53 | >900 hotspots including: BRAF
| CCND1 CCND2 CCND3 CDK4 CDK6 EGFR ERBB2 FGFR1 FGFR2 FGFR3 MET MYC | ALK BRAF ERG ETV1 FGFR1 FGFR2 FGFR3 MET NTRK1 NTRK3 RET ROS1 | MET exon Tumor | |
Oncomine Lung cfTNA Assay | DNA & RNA | ALK BRAF EGFR ERBB2 KRAS MAP2K1 MET NRAS PIK3CA RET ROS1 TP53 | >150 hotspots including: EGFR: T790M, C797S, | MET | ALK, RET,ROS1 | MET exon 14 skipping | |
Oncomine Lung cfDNA Assay | DNA | ALK BRAF EGFR ERBB2 KRAS MAP2K1 MET NRAS PIK3CA ROS1 TP53 | >150 hotspots including: EGFR: T790M, C797S, L858R, Exon 19 del KRAS: G12X, G13X, Q61X | --- | --- | --- | |
Oncomine Breast cfDNA Assay v2 | DNA | AKT1 CCND1 EGFR ERBB2 ERBB3 ESR1 FBXW7 FGFR1 KRAS PIK3CA SF3B1 TP53 | >150 hotspots including: ESR1: mutations associated with anti- estrogen resistance TP53: mutations associated with loss of function ERBB2: mutations associated with sensitivity to anti-ERBB2 therapies | CCND1, ERBB2, | --- | Expanded coverage of TP53 | |
Oncomine Breast cfDNA Assay | DNA | AKT1 EGFR ERBB2 ERBB3 ESR1 FBXW7 KRAS PIK3CA SF3B1 TP53 | >150 hotspots including: PIK3CA: E545K, H1047R AKT1: E17K ESR1: mutations associated with anti- estrogen resistance TP53: mutations associated with loss of function ERBB2: mutations associated with sensitivity to anti-ERBB2 therapies | --- | --- | --- | |
Oncomine Colon cfDNA Assay | DNA | AKT1 APC BRAF CTNNB1 EGFR ERBB2 FBXW7 GNAS KRAS MAP2K1 NRAS PIK3CA SMAD4 TP53 | >240 hotspots including: KRAS/NRAS: G12/G13/Q61 BRAF: V600E PIK3CA: E545K, H1047R TP53: R175H R273H/C/L Recurrent deleterious APC mutations (including p.R876*, p.R1114*, p.Q1378*, p.R1450*) SMAD4: R361C/H CTNNB1: S45F, T41A | --- | --- | --- |
Hotspot Genes | Tumor Suppressor Genes | Copy Number Genes | Gene Fusions | |
AKT1 ALK AR ARAF BRAF CHEK2 CTNNB1 DDR2 EGFR ERBB2 ERBB3 ESR1 FGFR1 FGFR2 FGFR3 FGFR4 FLT3 GNA11 GNAQ GNAS | HRAS | APC | CCND1 | ALK |
Since cell-free nucleic acids can vary from sample to sample, flexibility in supporting varying amounts of input DNA, especially small amounts, is critical. Oncomine cell-free assays have the ability to support varying input amounts (Figure 1). Getting data from as many samples as possible is a crucial step in advancing liquid biopsy research.
In verification studies, the assays demonstrate high correlation between the variants called in formalin-fixed, paraffin-embedded (FFPE) samples and those called in plasma (Table 1). As expected, there is a higher mutant allelic fraction in the FFPE tumor sample compared to that measured in plasma, and germline variants are seen at expected levels of ~50% in both FFPE and plasma.
Sample | Variant | FFPE | Plasma |
---|---|---|---|
1 | EGFR-L858R | 71.42% | 2.62% |
2 | TP53-R158L | 51.89% | 4.32% |
3 | MET-T1010I | 43.87% | 51.57% |
4 | N/A | No detection | No detection |
5 | EGFR-L858R | 58.44% | 7.28% |
6 | TP53-R158L | 10.19% | 1.26% |
Table 1. Correlation of FFPE and matched plasma sample results (late-stage lung cancer samples). Oncomine cell-free assays demonstrate high correlation between variants called in FFPE samples and those called in matched plasma samples. The above data was obtained using the Oncomine Lung cfDNA Assay. Bolded values indicate somatic mutations. Non-bolded values indicate germline mutations.
Overall, Oncomine cell-free assays enable results from more samples, with demonstrated, repeatable results using clinical research samples at 0.1% limit of detection with 90% sensitivity and >98% specificity across our whole portfolio of Oncomine cell-free assays.
Sample | EGFR | EGFR | EGFR | EGFR | KRAS | NRAS | NRAS | PIK3CA |
---|---|---|---|---|---|---|---|---|
0.1% HDX | 0.06 | 0.17 | 0.06 | 0.10 | 0.22 | 0.17 | 0.15 | 0.10 |
1% HDX | 0.72 | 1.07 | 0.75 | 0.74 | 1.14 | 1.15 | 1.15 | 2.29 |
5% HDX | 4.52 | 4.86 | 6.32 | 3.97 | 6.34 | 6.11 | 6.94 | 5.29 |
100% WT | 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 |
Table 2. Variants called from Horizon cfDNA Multiplex Reference Set. All 8 mutant hotspots were called at 0.1%. Data obtained using the Oncomine Lung cfDNA Assay.
Running up to 8 samples on an Ion 530 chip or up to 32 samples on an Ion 540 chip, the Oncomine cell-free assays enable efficiently multiplexed PCR analysis of single nucleotide variants (SNVs) , short indels, copy number variations, and fusions across our whole portfolio of Oncomine cell-free assays. frequently mutated in common tumor types, tolerating sample input variability and accommodating flexible input amounts as low as 1 ng.
The entire workflow (Figure 2), from isolation of cell-free nucleic acids using the appropriate Applied Biosystems MagMAX Cell-Free Isolation Kit to analysis of samples, can be accomplished in just 2 days on the Ion GeneStudio S5 Prime system.
Oncomine cell-free assays are part of a complete workflow designed for liquid biopsy clinical research, which includes:
A kit containing reagents for library construction and a single pool of multiplex PCR primers to prepare amplicon libraries from cell-free nucleic acids found in the plasma fraction of whole blood.
A set of unique barcode adapters specifically designed for optimal performance with our technology. The unique barcode adapters are compatible with the Ion S5, Ion PGM, and Ion Proton systems. When used in combination with kits, this set enables you to pool up to 24 amplicon libraries and conduct multiplex sequencing analysis, which helps reduce the sequencing cost per sample.
The Ion GeneStudio S5 Series supports five different chip types, so you can run multiple applications on a single sequencer. This flexibility eliminates the need to batch samples in order to achieve the optimum cost efficiency. Just choose the chip type that matches your specific throughput or application needs.
Learn more in this featured poster from AACR 2019.
** Oncomine Reporter is enabled in Ion Reporter™ Software version 5.2 or later.
For Research Use Only. Not for use in diagnostic procedures.