Alveolar epithelial cell type 2 populations and single cell gene expressions in non-small cell lung cancer progression and drug resistance

Harry S. Truman Memorial Va Hospital, Columbia MO

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Abstract

Non-small cell lung cancer (NSCLC) is by far the leading cancer killer in Veterans. Major reasons for death after surgical resection are the >50% recurrence rate despite curative intent treatment due to micrometastatic disease at the time of diagnosis. To prevent the development of metastatic recurrence, surgically treated patients receive adjuvant chemotherapy to eradicate undetectable micrometastatic disease. However, this therapy is only associated with a 5% survival benefit due to drug resistances of cancer cells. Identifying personalized treatment strategies to prevent metastases remains an unmet clinical need. Alveolar epithelial type II (AT2) cells are self-renewing epithelial stem cells in the lung that have been identified as the origin for a majority of NSCLCs. However, it is unknown whether and how epithelial cancer cells, such as AT2 cells, contribute to metastatic progression as the major cause of death. Novel data link AT2 cells to the development of NSCLC metastases. A research team led by a surgeon scientist has integrated expertise in human biobanking and translational cancer research at two VA hospitals with collaborators specialized in preclinical cancer models, single cell sequencing, genetics, and bioinformatics. It will be investigated how AT2 and other epithelial cancer cells contribute to NSCLC metastatic progression and drug resistances, as this will allow better molecular targeting to improve cure rates. Micrometastatic cells, such as circulating tumor cells (CTCs), are surrogates of the early metastatic process that can be detected by a blood draw as a liquid biopsy. Currently available preclinical models used to study the spectrum of cancer progression from a localized primary tumor to metastatic disease are limited. To combat this, a library of patient-derived xenograft (PDX) mouse models using primary tumors, micrometastatic CTCs (CDX) and brain metastases was developed. Using single cell RNA sequencing techniques, presence of an additional AT2 population was confirmed in NSCLC patients' metastatic tissues and primary lung tumors that had metastasized already. However, AT2 cells were absent in primary lung tumors of patients that had not metastasized. Importantly, AT2-depleted NSCLC cell lines have reduced tumorigenicity in PDX mouse models. Gene expression and culture data also suggest a strong migratory capacity of AT2-like cancer cells. This indicates an important role of AT2 cells in metastatic progression. For further validation of these critical leads, patient-derived organoid (PDO) pipelines for high-throughput tumor growth and drug response testing were generated. It is innovative to expand on metastasis-associated epithelial cancer cells, such as AT2, using patient-derived models and identify drug targets. Based on the rationale that AT2 populations with an impact on tumorigenicity are identified in metastases, the hypothesis is that AT2 cells promote NSCLC metastatic progression and express cancer- relevant genes. The objective is to identify AT2 lineage-specific phenotypes to detect therapeutic targets in NSCLC. In task 1, AT2-like and other epithelial cancer cell populations will be dissected in NSCLC primary tumors and metastases. AT2 cells will be enriched from a library of human NSCLC cell lines for study of tumorigenicity. Druggable pathways will be identified in NSCLC patient tissues with single cell analyses. In task 2, the impact of epithelial cancer cells, including AT2, on drug responses and patient prognosis will be determined. PDOs from human primary and metastatic NSCLC tumors will be treated with standard-of-care chemotherapy to associate drug responses with gene expressions of epithelial cancer cells. AT2 phenotypes will also be identified in CTCs as liquid biopsies. The impact of AT2-CTCs on survival in surgically treated NSCLC patients will be determined for risk stratification. Upon completion of the proposal, AT2-associated cellular and molecular targets to prevent metastatic progression in NSCLC will be identified. Results have high potential for clinical impact to prevent recurrences in patients undergoing curative surgery for NSCLC. Data will lead to discovery of drug targets at a single cell level that are primed for translation into clinical trials.