K-RAS Mutation Test Kit

Mutations in the K-RAS  oncogene are frequently found in human cancers.  They are common in pancreatic cancer, colorectal cancer, lung adenocarcinoma, gall bladder cancer, bile duct cancer and thyroid cancer.  These mutations may indicate prognosis and drug response, and many new cancer therapies are being targeted to the K-RAS pathway. Recent publications (1-5) suggest that in lung cancer patients the K-RAS mutation status is strong predictor of resistance to therapy with tyrosine kinase inhibitors such as Iressa™ (AstraZeneca) or Tarceva™ (Roche /OSI Pharmaceuticals). This kit will detect 7 K-RAS mutations in codons 12 and 13, allowing determination of whether there is a correlation between K-RAS mutation status and drug response, the kit can also be used to provide valuable biomarker information to drug companies with novel therapies in development.

  1. Gly12Asp (GGT>GAT)
  2. Gly12Ala (GGT>GCT)
  3. Gly12Val (GGT>GTT)
  4. Gly12Ser (GGT>AGT)
  5. Gly12Arg (GGT>CGT)
  6. Gly12Cys (GGT>TGT)
  7. Gly13Asp (GGC>GAC)

The methods used in this kit are highly selective and, depending on the total amount of DNA present, can detect approximately 1% of mutant in a background of wild type genomic DNA.  The assays have limits of detection of between 5 and 10 copies.  These selectivity and detection limits are superior to technologies such as dye terminator sequencing.

 

Intended Use

K-RAS mutation kitThe K-RAS Mutation Test Kit is intended for the detection of 7 somatic mutations in the K-RAS oncogene.  The kit is for use on DNA samples and will provide a qualitative assessment of mutation status. 

The DxS K-RAS Mutation Kit is for research use only.


Technological Principles

DxS has combined ARMS™ (allele specific PCR, 6) with the Scorpions™ real-time PCR technology (7,8) to develop sensitive and robust tests for tumour-borne K-RAS mutations. 
 

Protocol

After DNA extraction, real time PCR assays are performed to detect the target molecule.  By comparing control and mutant sample reactions users can detect and estimate low levels of mutation. No further sample processing is necessary and the time to result is <3 hours.
 

Key Features:

  • The kit is highly selective and robust, detecting seven key mutations in the K-RAS gene.
  • The un-paralleled sensitivity of the kit means that the assay detects mutations frequently missed by sequencing methods.
  • The assay detects mutations frequently missed by sequencing methods
  • Limit of detection of 10 copies of below
  • Detects very low, <1% , levels of mutant in a background of normal DNA
  • Qualitative real-time PCR assay combining Scorpions® and ARMS® (allele-specific PCR) technologies
  • Compatible with most real-time PCR machines
  • Sample types include DNA from fresh or frozen tumour tissue and PET samples

 

References

  1. E. Massarelli, M. Varella-Garcia, X. Tang, A. C. Xavier et al. (2007). KRAS Mutation Is an Important Predictor of Resistance to Therapy with Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitors in Non-Small Cell Lung Cancer. Clin Cancer Res 2007; 13 (10).

  2. William Pao, Theresa Y. Wang, Gregory J. Riely, Vincent A. Miller, Qiulu Pan, Marc Ladanyi et al. (2005). KRAS mutations and Primary Resistance of Lung Adenocarcinomas to Gefitinib or Erlotinib.  PloS Medicine 2(1): 57-61.

  3. D. A. Eberhard, B. E. Johnson, L.C. Amler, A. D. Goddard et al (2005). Mutations in the EGFR and in K-RAS are predictive and prognostic indicators in patients with NSCLC treated with chemotherapy alone and in combination with erlotinib. J Clin Oncol 23: 5900-5909.

  4. L. Toschi & F. Cappuzzo. (2007) Understanding the new genetics of responsiveness to EGFR tyrosine kinase inhibitors. Oncologist 12; 211-220.

  5. Sae-Won Han, Tae-You Kim, Yoon Kyung Jeon, Pil Gyu Hwang et al. (2006). Optimization of Patient Selection for Gefitinib in Non-Small Cell Lung Cancer by combined analysis of Epidermal Growth Factor Receptor Mutation, K-RAS Mutation, and AKT Phosphorylation.  Clin Cancer Res 12(8):2538-2544.

  6. Newton CR, Graham A, Heptinstall LE, Powell SJ, Summers C et al. (1989). Analysis of any point mutation in DNA. The amplification refractory mutation system (ARMS) Nucleic Acids Res. 17 (7): 2503-16.

 
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