Abstract B56: High sensitivity characterization of KRAS mutant subpopulations in normal lung tissue and adenocarcinomas of the lung
Authors: Myers Meagan B. , McKim Karen L. , Meng Fanxue , Parsons Barbara L. 
Published: May 2015

Abstract
KRAS tumor mutational status predicts response to therapies that target the epidermal growth factor receptor (EGFR) and is being used to make treatment decisions for individual colon cancer patients. There is evidence showing that undetected KRAS mutant tumor subpopulations can subvert durable responses, lead to drug resistance, and drive relapse in colon cancer patients treated with EGFR-targeted therapies. Patients with adenocarcinomas of the lung are treated with EGFR thymidine kinase inhibitors and, theoretically, the efficacy of these treatments could be influenced by KRAS mutational status, even though KRAS mutational status is not currently evaluated prior to treatment of lung cancer patients. In order to assess the potential for undetected KRAS mutant subpopulations to impact response in lung adenocarcinoma patients treated with EGFR thymidine kinase inhibitors, the sensitive allele-specific competitive blocker polymerase chain reaction (ACB-PCR) method was used to quantify KRAS codon 12 GGT to GAT and GGT to GTT mutant fraction (MF) in 19 normal lung samples and 21 primary lung adenocarcinomas. ACB-PCR is a type of allele-specific amplification that can measure 3 mutant alleles in a background of 300,000 normal alleles and gives a quantitative readout, rather than reporting the presence/absence of mutation. ACB-PCR was used to quantify KRAS mutation in normal lung and lung tumor tissue in order to empirically define pathological levels of KRAS mutation as those above the upper 95% confidence interval of that present in normal lung. Measurable levels (≥10-5) of KRAS codon 12 GTT and/or GAT mutations were observed in 17/19 (89%) of normal lung samples. The KRAS codon 12 GAT and GTT geometric mean MFs in normal lung were 3.25 x 10-5 and 3.15 x 10-5, respectively. Normal colonic mucosa and normal lung tissue contain similar levels of KRAS codon 12 GAT mutation, however normal lung tissue has a significantly higher level of the KRAS codon 12 GTT mutation than colonic mucosa. All lung adenocarcinomas had measurable levels of both mutations. Adenocarcinomas from male smokers had significantly greater KRAS codon 12 GTT MFs than adenocarcinomas from female smokers. The highest observed KRAS codon 12 GAT MFs were associated with tumors characterized as having relatively low necrosis and tumor cell content. Importantly, 47.6% and 57.1% of lung adenocarcinomas had KRAS codon 12 GAT and GTT MFs greater than the upper 95% confidence interval for MF in normal lung (1.02 x 10-4 and 9.16 x 10-5, respectively). In addition, 28% of lung adenocarcinomas had abnormally-high levels of both mutations. Extrapolation of these results to include additional KRAS hotspot point mutations leads to the conclusion that most, if not all, lung adenocarcinomas possess greater KRAS mutant cell content than normal lung tissue. These results suggest there is a strong possibility that treatment of lung adenocarcinoma patients with therapies that target EGFR may select for the outgrowth of KRAS mutant subpopulations and contribute to the development of drug resistance and relapse.
Citation Format: Meagan B. Myers, Karen L. McKim, Fanxue Meng, Barbara L. Parsons. High sensitivity characterization of KRAS mutant subpopulations in normal lung tissue and adenocarcinomas of the lung. [abstract]. In: Proceedings of the AACR Special Conference on RAS Oncogenes: From Biology to Therapy; Feb 24-27, 2014; Lake Buena Vista, FL. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(12 Suppl):Abstract nr B56. doi: 10.1158/1557-3125.RASONC14-B56