PROJECT SUMMARY/ABSTRACT A tumor biopsy is traditionally performed for lung cancer diagnosis using either bronchoscopy through the airway or a needle aspiration through the chest wall. Advances in targeted and immune therapies now often require more tissue for molecular and immune profiling to optimally manage lung cancer. The yield for cancer diagnosis using modern bronchoscopic tools approaches only 50% across the spectrum of lung tumors biopsied, and the additional requirement for molecular and immune profiling erodes this yield further. This leads to delayed and suboptimal care, increased healthcare costs, and increased patient morbidity since patients can often require multiple procedures to obtain the correct information for treatment. Our group has recently demonstrated that targeted Bronchoalveolar Lavage (BAL) – or a washing of the lung cancer performed during a procedure – is a reservoir of genomic and cellular biomarkers in the lung tumor macroenvironment (TMaE). Further, we have demonstrated that molecular analyses of BAL from the lung TMaE recapitulate cancer biology in the lung tumor microenvironment (TMiE). While BAL is very safe and routinely performed during bronchoscopy, to date, it is a pauci-cellular fluid that is of limited clinical utility for cancer diagnosis. Beyond cytology that is low yield, there are no molecular or cellular assays that are used in the clinic to fully inform providers who treat lung cancer. Because of this, rigorous attention to how methods of collection, patient host factors and processing of BAL will alter genomic and high dimensional cell based assays is lacking. Our central hypothesis is: BAL globally samples the tumor microenvironment (TMiE) to overcome limitations of tumor heterogeneity and is more sensitive than blood for immunogenomic profiling due to increased quantities of tumor specific biomarkers. To realize our goal and prove our hypothesis, in depth analysis of the conditions affecting BAL for high dimensional genome and cell assays is required. Here, we propose studying how basic conditions in the lung, variations in acquisition of BAL, and storage and processing of BAL affect its utility for comprehensive genome profiling and analysis of the T cell repertoire. Following identification of key pre-analytic variables, we propose a standard operating procedure for implementation in observational biomarker and first-in-man clinical trials to demonstrate the clinical utility of our approach. Proposal success will facilitate the introduction of novel molecular assays into the clinic that augment extant and developing blood and tumor assays. This approach will be particularly relevant as we move into the era of precision guided therapies for lung cancer treatment, which have begun to reduce mortality in even the most advanced stages, over the coming years and decades.