Purpose Promising results in the treatment of NSCLC have been seen with agents targeting immune checkpoints, such as PD-1 or PD-L1. However, only a select group of patients respond to these interventions. The identification of biomarkers that predict clinical benefit to immune checkpoint blockade is critical to successful clinical translation of these agents. Methods We conducted an integrated analysis of three independent large datasets, including The Cancer Genome Atlas (TCGA) of lung adenocarcinoma and two datasets from MD Anderson Cancer Center, Profiling of Resistance patterns and Oncogenic Signaling Pathways in Evaluation of Cancers of the Thorax (named PROSPECT) and Biomarker-integrated Approaches of Targeted Therapy for Lung Cancer Elimination (named BATTLE-1). Comprehensive analysis of mRNA gene expression, reverse phase protein array (RPPA), immunohistochemistry and correlation with clinical data were performed. Results Epithelial-mesenchymal transition (EMT) is highly associated with an inflammatory tumor microenvironment in lung adenocarcinoma, independent of tumor mutational burden. We found immune activation co-existent with elevation of multiple targetable immune checkpoint molecules, including PD-L1, PD-L2, PD-1, TIM-3, B7-H3, BTLA and CTLA-4, along with increases in tumor infiltration by CD4+Foxp3+ regulatory T cells in lung adenocarcinomas that displayed an EMT phenotype. Furthermore, we identify B7-H3 as a prognostic marker for NSCLC. Conclusions The strong association between EMT status and an inflammatory tumor microenvironment with elevation of multiple targetable immune checkpoint molecules warrants further investigation of using EMT as a predictive biomarker for immune checkpoint blockade agents and other immunotherapies in NSCLC and possibly a broad range of other cancers.

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Seven patients with typical or atypical pulmonary carcinoid tumors overdiagnosed as small-cell carcinoma on bronchoscopic biopsies are described. Bronchial biopsies from 9 consecutive small-cell lung carcinoma patients were used as control group for histologic and immunohistochemical studies (cytokeratins, chromogranin A, synaptophysin, Ki-67 [MIB-1], and TTF-1). The carcinoid tumors presented as either central or peripheral lesions composed of tumor cells with granular, sometimes coarse chromatin pattern, high levels of chromogranin A/synaptophysin immunoreactivity, and low (<20%) Ki-67 (MIB-1) labeling index. The tumor stroma contained thin-walled blood vessels. Small-cell carcinomas always showed central tumor location, finely dispersed nuclear chromatin, lower levels of chromogranin A/synaptophysin, and high (>50%) Ki-67 (MIB-1) labeling index. The stroma contained thick-walled blood vessels with glomeruloid configuration. Judging from this study, overdiagnosis of carcinoid tumor as small-cell carcinoma in small crushed bronchial biopsies remains a significant potential problem in a worldwide sample of hospital settings. Careful evaluation of hematoxylin and eosin sections remains the most important tool for the differential diagnosis, with evaluation of tumor cell proliferation by Ki-67 (MIB-1) labeling index emerging from our review as the most useful ancillary technique for the distinction.

Non-Small Cell Lung Cancer (NSCLC) demonstrates remarkable molecular diversity. With the completion of The Cancer Genome Atlas (TCGA), there is opportunity for systematic analyses of the entire TCGA NSCLC cohort, including comparisons and contrasts between different disease subsets. On the basis of multidimensional and comprehensive molecular characterization (including DNA methylation and copy, and RNA and protein expression), 1023 NSCLC cases-519 from TCGA adenocarcinoma (AD) project and 504 from TCGA squamous cell (SQCC) project-were classified using a "cluster-of-clusters" analytic approach. Patterns from TCGA NSCLC subsets were examined in independent external databases, including the PROSPECT (Profiling of Resistance patterns and Oncogenic Signaling Pathways in Evaluation of Cancers of the Thorax) NSCLC dataset. Nine genomic subtypes of NSCLC were identified, three within SQCC and six within AD. SQCC subtypes were associated with transcriptional targets of SOX2 or p63. One predominately AD subtype (with a large proportion of SQCC) shared molecular features Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use: http://www.nature.com/authors/editorial_policies/license.html#terms HHS Public Access Author Manuscript Author ManuscriptAuthor ManuscriptAuthor Manuscript with neuroendocrine tumors. Two AD subtypes manifested a CpG island methylator phenotype (CIMP). Three AD subtypes showed high p38 and mTOR pathway activation. AD subtypes associated with low differentiation showed relatively worse prognosis. SQCC subtypes and two of the AD subtypes expressed cancer-testis antigen genes, while three AD subtypes expressed several immune checkpoint genes including PDL1 and PDL2, corresponding with patterns of greater immune cell infiltration. Subtype associations for several immune-related markers-including PD1, PDL1, CD3, and CD8-were confirmed in the PROSPECT cohort using immunohistochemistry. NSCLC molecular subtypes have therapeutic implications and lend support to a personalized approach to NSCLC management based on molecular characterization.

Epithelial to mesenchymal transition (EMT) is a key process associated with tumor progression and metastasis. To define molecular features associated with EMT states, we undertook an integrative approach combining mRNA, microRNA, DNA methylation and proteomic profiles of 38 cell populations representative of the genomic heterogeneity in lung adenocarcinoma were integrated with functional profiles consisting of cell invasiveness, adhesion and motility. A subset of cell lines that were readily defined as epithelial or mesenchymal based on their morphology and E-cadherin and vimentin expression elicited distinctive molecular signatures. However, most cell populations displayed intermediate/hybrid states of EMT, with mixed epithelial and mesenchymal characteristics. A dominant proteomic feature of aggressive hybrid cell lines was upregulation of cytoskeletal and actin binding proteins, a signature shared with mesenchymal cell lines. Cytoskeletal reorganization preceded loss of E-cadherin in epithelial cells in which EMT was induced by TGFβ. A set of transcripts corresponding to the mesenchymal protein signature enriched in cytoskeletal proteins was found to be predictive of survival in independent datasets of lung adenocarcinomas. Our findings point to an association between cytoskeletal and actin-binding proteins, a mesenchymal or hybrid EMT phenotype and invasive properties of lung adenocarcinomas.