Mutation profile

Prognosis for non-small cell lung cancer (NSCLC) is substantially better in early-stage as opposed to late-stage disease. If high-risk individuals could be effectively identified, they could significantly benefit from intensive surveillance, early detection, and prevention strategies. While smoking is a primary risk factor, only about 15% of smokers develop the disease. Consistent with a genetic predisposition, some patients have a family history of the disease and are affected at a young age, although they have never smoked. Unfortunately, NSCLC predisposing gene mutations are poorly characterized. We aim to identify novel NSCLC germline risk gene mutations. Our team members have previously identified novel risk gene mutations for several familial cancers. We have assembled a large clinically and molecularly well-characterized NSCLC case-control cohort of never-smokers, performed whole-exome sequencing (WES) on familial and early onset NSCLC probands, identified promising candidate risk genes, and externally validated selected samples. We follow a resource-conscious approach by focusing on a founder population, Ashkenazi Jews (AJ), to decrease genetic heterogeneity and to increase the allele frequencies of candidate recurrent risk mutations. We will perform WES on additional familial and early onset NSCLC probands to identify recurrently mutated NSCLC genes and cross-validate the genes using whole-genome sequencing on matching tumors. The gene candidates identified will be validated using a targeted sequencing approach in a larger non-AJ cohort. These are anticipated to provide new biological insights, serve as future chemoprevention targets, and be used in the development of a genetic diagnostic test to identify high-risk individuals.

Approximately 30% of patients with stage 1 non-small cell lung cancer (NSCLC) will die from recurrent disease despite surgical resection . Currently, there is a lack of reliable molecular predictors being routinely used in the clinic for identifying patients at high risk for developing recurrent disease and therefore may benefit from more aggressive therapies. To date several groups have applied microarray-based methods for global gene expression profiling of frozen tumor tissues in an attempt to identify prognostic ‘signatures’ in patients with early stage NSCLC.  Building upon this, through a meta-analysis of seven independent microarray studies Govindan et al. were able to identify a 64-gene signature that is highly predictive of survival in patients with stage I NSCLC. However, recent advances in high-throughput sequencing have enabled an unbiased view of the transcriptome including novel transcriptional events such as gene fusions, splicing variants, and novel transcripts. This offers an opportunity to incorporate additional markers, which may have eluded previous technologies, but could serve for improving a predictive signature of survival in non-small cell lung cancer (NSCLC). However, despite the potential of RNA-Seq to improve existing predictive signatures, we are still faced with the challenge of translating these findings into a platform where they can be reliably applied to formalin fixed paraffin embedded (FFPE) clinical tissue specimens routinely collected in tertiary referral centers. Therefore, this proposal focuses on refining the 64-gene predictive signature using transcriptome sequencing and developing a NanoString nCounter assay to reliably predict survival of patients with resected stage I NSCLC, using RNA derived from FFPE tumor blocks. To accomplish this we have proposed the following specific aims: (1) use transcriptome sequencing to detect molecular predictors of outcome in patients with stage I NSCLC, (2) validate the technical performance of the NanoString nCounter using an RNA-Seq predictive signature across FFPE tumor blocks of resected stage I NSCLC, and (3) clinically validate the prognostic power of the predictive signature across an independent sample set. Overall, if the proposed aims are met, this research could lead to an improved assay that could be translated into future prospective studies for risk-stratifying stage I NSCLC patients.

Recently, the National Lung Screening Trial reported a 20% reduction in lung cancer mortality and lung cancer chemoprevention trials targeting the arachidonic acid pathway have demonstrated decreases in lung cancer associate markers .These studies highlight possibilities for future reductions in lung cancer mortality through early detection and chemoprevention. Our group has previously identified smoking- and lung cancer-specific gene expression alterations in cytologically normal airway epithelial cells that can serve s a clinically relevant biomarker for the early detection of lung cancer. We propose to extend our studies of the field of injury by profiling  these cells from high-risk smokers with preinvasive lesions (dysplasia) and matched controls without lesions by RNA-Seq. We will characterize the molecular alterations (coding, noncoding, and novel RNA expression and alterative splicing) associated with the field of injury in premalignancy and identify whether these differences are similar in both current and former smokers. Next, we will develop molecular signatures that predict subsequent progression or regression of premalignant lesions as well as changes that occur as a result of progression or regression. Collectively, these signatures will aid in the stratification of high-risk smokers for chemoprevention trials, serve as intermediate markers of efficacy in these trials, and identify new targets for chemoprevention. Finally, we will explore whether these molecular signatures re associated with occult lung cancer or future lung cancer development in independent cohorts as common alterations may indicate early events in lung carcinogenesis and have additional utility as biomarkers in the early detection of lung cancer.