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Background Despite the importance of tumor-infiltrating T lymphocytes (TILs) in cancer biology, the relationship between TIL phenotypes and their prognostic relevance for localized non-small-cell lung cancer (NSCLC) has not been well established. Patients and methods Fresh tumor and normal adjacent tissue was prospectively collected from 150 patients with localized NSCLC. Tissue was comprehensively characterized by high-dimensional flow cytometry of TILs integrated with immunogenomic data from multiplex immunofluorescence, T-cell receptor sequencing, exome sequencing, RNA sequencing, targeted proteomics, and clinicopathologic features. Results While neither the magnitude of TIL infiltration nor specific TIL subsets were significantly prognostic alone, the integration of high-dimensional flow cytometry data identified two major immunotypes (IM1 and IM2) that were predictive of recurrence-free survival independent of clinical characteristics. IM2 was associated with poor prognosis and characterized by the presence of proliferating TILs expressing cluster of differentiation 103, programmed cell death protein 1, T-cell immunoglobulin and mucin-domain containing protein 3, and inducible T-cell costimulator. Conversely, IM1 was associated with good prognosis and differentiated by an abundance of CD8+ T cells expressing cytolytic enzymes, CD4+ T cells lacking the expression of inhibitory receptors, and increased levels of B-cell infiltrates and tertiary lymphoid structures. While increased B-cell infiltration was associated with good prognosis, the best prognosis was observed in patients with tumors exhibiting high levels of both B cells and T cells. These findings were validated in patient tumors from The Cancer Genome Atlas. Conclusions Our study suggests that although the number of infiltrating T cells is not associated with patient survival, the nature of the infiltrating T cells, resolved in distinct TIL immunotypes, is prognostically relevant in NSCLC and may inform therapeutic approaches to clinical care.

Background: Patients with thoracic cancers affected by the coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), appear to have poor clinical outcomes AXL, a TAM (Tyro3, AXL, Mer) family receptor tyrosine kinase, is a known mediator of epithelial to mesenchymal transition (EMT) and therapeutic resistance in non-small cell lung cancer (NSCLC) and other cancers Additionally, AXL plays a role in efficient Ebola and Zika viral entry and infection and AXL inhibition has demonstrated antiviral activities Recently, bemcentinib, a highly selective and potent AXL inhibitor with antiviral activity, has been fast-tracked as the first potential treatment for assessment in the United Kingdom’s ACcelerating COVID-19 Research & Development (ACCORD) multicenter, randomized phase II trial Methods: We analyzed mRNA expression of AXL and other TAM family members as well as angiotensin-converting enzyme 2 (ACE2), the SARS-CoV-2 receptor, in treatment-naive (n=1016) and previously treated (n=239) NSCLC tumors and in a panel of NSCLC cell lines (n=70) We also analyzed AXL mRNA levels in NSCLC cell lines (n=3) infected with SARS-CoV-2 Results: In treatment-naive and previously-treated NSCLC tumors, AXL mRNA expression was higher in mesenchymal tumors, as expected, and inversely correlated with ACE2 Similarly, in NSCLC cell lines, high ACE2 expression was associated with low AXL mRNA and protein expression Notably, expression of ACE2 was downregulated while that of AXL and ZEB1, an EMT transcription factor, were upregulated in NSCLC cells infected with SARS-CoV-2 as compared to mock infected cells, suggesting a shift to a more mesenchymal phenotype Treatment with bemcentinib for 24h downregulated ZEB1 expression in mesenchymal cell lines, reversing EMT Conclusions: These data, in the context of ACE2’s role in preventing acute respiratory distress syndrome, suggest a shift from ACE2-expressing epithelial cells to a more mesenchymal phenotype characterized by low ACE2 and high AXL expression, upon infection of NSCLC cells with SARS-CoV-2 In addition to bemcentinib’s antiviral activity, it can also reverse EMT, further supporting AXL and EMT as novel therapeutic targets for COVID-19 treatment Legal entity responsible for the study: Lauren A Byers Funding: NIH/NCI CCSG P30-CA016672 (Bioinformatics Shared Resource), NIH/NCI T32 CA009666, ASCO Young Investigator Award (C M G );University of Texas SPORE in Lung Cancer P5-CA070907 (L A B C M G ), NIH/NCI R01-CA207295, NIH/NCI U01-CA213273, the Department of Defense (LC170171) (L A B ), The LUNGevity foundation (D G , L A B ), through generous philanthropic contributions to The University of Texas MD Anderson Lung Cancer Moon Shot Program (J V H , J W , L A B );an Andrew Sabin Family Fellowship (L A B ), and The Rexanna Foundation for Fighting Lung Cancer (J V H , L A B ) Disclosure: C Gay: Research grant/Funding (self): AstraZeneca D Gibbons: Advisory/Consultancy, Research grant/Funding (self): AstraZeneca;Advisory/Consultancy: GlaxoSmithKline;Advisory/Consultancy: Sanofi;Advisory/Consultancy, Research grant/Funding (self): Janssen;Research grant/Funding (self): Takeda;Research grant/Funding (self): Ribon Therapeutics;Research grant/Funding (self): Astellas J V Heymach: Advisory/Consultancy, Research grant/Funding (self): AstraZeneca;Advisory/Consultancy: Boehringer Ingelheim;Advisory/Consultancy: Exelixis;Advisory/Consultancy: Genentech;Advisory/Consultancy, Research grant/Funding (self): GlaxoSmithKline;Advisory/Consultancy: Guardant Health;Advisory/Consultancy: Hengrui;Advisory/Consultancy: Lilly;Advisory/Consultancy: Novartis;Advisory/Consultancy, Research grant/Funding (self), Licensing/Royalties: Spectrum;Advisory/Consultancy: EMD Serono;Advisory/Consultancy: Synta;Research grant/Funding (self): Bayer L A Byers: Advisory/Consultancy, Research grant/Funding (self): AstraZeneca;AbbVie;GenMab;Sierra Oncology;Advisory/Consultancy: BergenBio;Pharma Mar SA;Merck;Bristol Myers Squibb;G nentech;Pfizer;Research grant/Funding (self): Tolero Pharmaceuticals All other authors have declared no conflicts of interest

Background: The novel coronavirus SARS-CoV-2 is the cause of the respiratory illness COVID-19—a global pandemic affecting over 4 million individuals worldwide Viruses efficiently replicate by hijacking host cell machinery to obtain macromolecules and energy by similar mechanisms as cancer cells Since viral infection is known to alter cellular nutrient requirements, this study explores the metabolites and metabolic pathways associated with SARS-CoV-2 infection Methods: Bulk and single-cell seequencing data from cell lines and tumor samples were retrieved from publically available datasets Transcriptional data were retrieved from publically available datasets of gefitinib- and erlotinib-resistant EGFR-mutant cell lines and Calu3 and A549 cells mock treated or infected with SARS-CoV-2 Single-cell RNAseq datasets of EGFR-mutant PC-9 mock and osimertinib treated were downloaded from GEO 225 metabolites were profiled in CCLE cell lines using LC-MS Results: To identify metabolic features of cells able to be infected by SARS-CoV-2 via the ACE2 receptor, metabolites associated with ACE2 expression were investigated ACE2 expression positively correlates with glutamine in upper aerodigestive tract cell lines Consistent with this, ACE2 expression was examined against a list of 253 metabolism-associated genes and GLUL, which encodes an enzyme (glutamine synthetase) responsible for conversion of glutamate to glutamine, was significantly positively correlated in NSCLC, HNSCC, and SCLC cell lines and confirmed in human tumor datasets Additionally, GLS, which encodes the enzyme (glutaminase) that catalyzes the opposing reaction, is negatively correlated with ACE2 expression Further, we analyzed RNA sequencing data from NSCLC cell lines infected with SARS-CoV-2 for 24 hours and revealed that upon infection there is a down regulation of GLUL signifying a metabolic-shift away from glutamine as the cells undergo EMT Conclusions: We show that SARS-CoV-2 targeting of ACE-2 expressing, metabolically-primed epithelial cells is advantageous to exploit the abundance of glutamine to synthesize nucleotides for rapid replication and viral spread Legal entity responsible for the study: Lauren A Byers Funding: NIH/NCI R01-CA207295 (L A B ), NIH/NCI U01-CA213273 (L A B , J V H ), CCSG P30-CA01667 (L A B ), University of Texas SPORE in Lung Cancer P5-CA070907 (L A B , D L G , J V H , C M G ), the Department of Defense (LC170171;L A B ), Khalifa Bin Zayed Al Nahyan Foundation(C M G ), The University of Texas MD Anderson Cancer Center-Oropharynx Cancer Program generously supported by Mr and Mrs Charles W Stiefel (F M J ), through generous philanthropic contributions to The University of Texas MD Anderson Lung Cancer Moon Shot Program and Andrew Sabin Family Fellowship, and The Rexanna Foundation for Fighting Lung Cancer Disclosure: C Gay: Research grant/Funding (self): AstraZeneca S Heeke: Honoraria (self): Qiagen;Boehringer Ingelheim;Travel/Accommodation/Expenses: Roche D Gibbons: Advisory/Consultancy, Research grant/Funding (self): AstraZeneca;Janssen;Advisory/Consultancy: GlaxoSmithKline;Sanofi;Research grant/Funding (self): Takeda;Ribon Therapeutics;Astellas J V Heymach: Advisory/Consultancy, Research grant/Funding (self): AstraZeneca;Advisory/Consultancy: Boehringer Ingelheim;EMD Serono;Novartis;Lilly;Hengrui;Guardant Health;GSK;Genentech;Exelixis;Synta;Advisory/Consultancy, Research grant/Funding (self), Licensing/Royalties: Spectrum;Research grant/Funding (self): Bayer;GlaxoSmithKline L A Byers: Advisory/Consultancy: Pfizer;Genentech;: Bristol Myers Squibb;Merck;Pharma Mar SA;BergenBio;Advisory/Consultancy, Research grant/Funding (self): Sierra Oncology;GenMab;AstraZeneca;AbbVie;Research grant/Funding (self): Tolero Pharmaceuticals All other authors have declared no conflicts of interest

Background Aurora Kinases (AURK) regulate mitosis and are often upregulated in cancer. SCLC is characterized by TP53 and RB1 loss and frequent MYC amplification. Our group, and others, have proved that MYC-driven SCLC are vulnerable to AURKA-inhibitors (AURKAi), but their use is limited by toxicity due to high AURKA expression. AURKB levels are variable in SCLC, making AURKB an attractive targeted therapeutic approach. A novel AURKBi, AZD2811NP (nanoparticle), is now being investigated in relapsed SCLC patients (NCT02579226). We hypothesize that molecularly defined subsets may be sensitive to AZD2811. Methods We tested 50 human-derived SCLC cell lines with AZD2811 in 96 hour proliferation assays. To identify translatable biomarkers of response, we correlated IC50 values with genomic (WES), transcriptomic (RNASeq), and proteomic profiling (RPPA). Results AZD2811 is active in a subset of SCLC cells: 13/30 (26%) have high sensitivity (IC50 Conclusions Our results show encouraging single agent in vitro activity of AZD2811 in SCLC cells, and suggest a novel biomarker-driven approach for combinations. Candidate biomarkers will be tested in samples from the ongoing clinical trial. The high BCL2 levels observed in resistant cells provide a rationale for exploring AURKB/BCL2-i combinations. Clinical trial identification NCT02579226. Legal entity responsible for the study The authors. Funding AstraZeneca. Disclosure C.A. Adelman: Full / Part-time employment: AstraZeneca. J. Travers: Full / Part-time employment: AstraZeneca. J. Heymach: Advisory / Consultancy, Research grant / Funding (self): AstraZeneca; Advisory / Consultancy, Licensing / Royalties: BioTree; Advisory / Consultancy: Boehringer Ingelheim; Advisory / Consultancy: Exelixis; Advisory / Consultancy: Genentech; Advisory / Consultancy: GSK; Advisory / Consultancy: Guardant Health; Advisory / Consultancy: Hengrui; Advisory / Consultancy: Lilly; Advisory / Consultancy: Novartis; Advisory / Consultancy, Research grant / Funding (self), Licensing / Royalties: Spectrum; Advisory / Consultancy: EMD Serono; Advisory / Consultancy: Synta; Research grant / Funding (self): Bayer; Research grant / Funding (self): GlaxoSmithKline. L.A. Byers: Advisory / Consultancy, Research grant / Funding (self): AstraZeneca; Advisory / Consultancy, Research grant / Funding (self): AbbVie; Advisory / Consultancy, Research grant / Funding (self): GenMab; Advisory / Consultancy: BergenBio; Advisory / Consultancy: Pharma Mar; Advisory / Consultancy: SA; Advisory / Consultancy, Research grant / Funding (self): Sierra Oncology; Research grant / Funding (self): Tolero Pharmaceuticals. All other authors have declared no conflicts of interest.