Mutation Enriched Targeted Re-Sequencing

Information

  • Research Project
  • 9042780
  • ApplicationId
    9042780
  • Core Project Number
    R42CA180389
  • Full Project Number
    2R42CA180389-02A1
  • Serial Number
    180389
  • FOA Number
    PA-14-072
  • Sub Project Id
  • Project Start Date
    8/1/2013 - 11 years ago
  • Project End Date
    12/31/2017 - 6 years ago
  • Program Officer Name
    LOU, XING-JIAN
  • Budget Start Date
    1/1/2016 - 8 years ago
  • Budget End Date
    12/31/2016 - 7 years ago
  • Fiscal Year
    2016
  • Support Year
    02
  • Suffix
    A1
  • Award Notice Date
    12/15/2015 - 9 years ago
Organizations

Mutation Enriched Targeted Re-Sequencing

? DESCRIPTION (provided by applicant): Low-level, tumor-associated somatic DNA mutations can have profound implications for development of metastasis, prognosis, choice of treatment, follow-up or early cancer detection. Unless they are effectively detected, these low-level mutations can misinform patient management decisions or become missed opportunities for personalized medicine. Widely-used technologies such as sequencing are not sensitive enough to detect these mutations when they are at very low percentages compared to normal DNA. Likewise the next generation sequencing technologies (NGS) are promising technology advances that can effectively detect prevalent somatic mutations in targeted gene panels; however due to the limited quantity of DNA in most patient samples and the abundance of normal DNA when analyzing blood, NGS 'loses steam' and its integration with clinical practice is problematic. For mutations at an abundance of ~2-5% or below, NGS generates false positives (`noise') independent of sequencing depth; yet these are often the clinically relevant mutations causing resistance to drug treatments. Commercial sample preparation kits for targeted re-sequencing of cancer gene panels have emerged, however they are uniformly unable to detect mutations below a 2% abundance level. Thus, while targeted re-sequencing provides an opportunity for integration of NGS with clinical oncology, the technology is ineffective in detecting DNA mutations in circulating DNA, urine, or heterogeneous cancers. We intend to use COLD-PCR, a recently developed method that enriches unknown mutation-containing sequences over wild-type, normal alleles during PCR amplification. In previous work we showed COLD-PCR- NGS-based sequencing for mutations down to 0.02% abundance. However, COLD-PCR was only applicable with a single amplicon per reaction, limiting its efficient combination with NGS. This STTR proposes a simple and powerful modification that enables COLD-PCR to be applied to hundreds or thousands of DNA targets in a single reaction, thus enabling mutation enrichment in disease- specific gene panels prior to NGS. The new approach, temperature-tolerant-COLD-PCR (TT-COLD-PCR) converts the rare mutations to high abundance mutations, overcoming the `noise' and avoiding the costly need for repeated sequence reads during NGS. In Phase I we obtained proof of principle for TT-COLD-PCR. In Phase II, TT-COLD-PCR will be developed into kits for cancer-specific gene panels, to magnify rare mutations in multiple DNA targets thus enabling expanded application of targeted re-sequencing for heterogeneous cancers or circulating DNA. This project meets one of the aims of the NCI to support the development of new methods of diagnosis for the detection, discovery and validation of biomarkers for cancer detection, diagnosis and prognosis.

IC Name
NATIONAL CANCER INSTITUTE
  • Activity
    R42
  • Administering IC
    CA
  • Application Type
    2
  • Direct Cost Amount
  • Indirect Cost Amount
  • Total Cost
    773511
  • Sub Project Total Cost
  • ARRA Funded
    False
  • CFDA Code
    394
  • Ed Inst. Type
  • Funding ICs
    NCI:773511\
  • Funding Mechanism
    SBIR-STTR RPGs
  • Study Section
    ZRG1
  • Study Section Name
    Special Emphasis Panel
  • Organization Name
    TRANSGENOMIC, INC.
  • Organization Department
  • Organization DUNS
    969419258
  • Organization City
    Omaha
  • Organization State
    NE
  • Organization Country
    UNITED STATES
  • Organization Zip Code
    681644268
  • Organization District
    UNITED STATES