Prevalence and Therapeutic Targeting of High-Level ERBB2 Amplification in NSCLC.

INTRODUCTION: ERBB2 amplification in lung cancer remains poorly characterized. HER2 (encoded by ERBB2) is a transmembrane tyrosine kinase capable of ligand-independent dimerization and signaling when overexpressed, and a common cause of HER2 overexpression is ERBB2 amplification. Here, we evaluated the clinicopathologic and genomic characteristics of ERBB2-amplified NSCLC and explored a HER2 antibody-drug conjugate (ADC) therapeutic strategy. METHODS: Our institutional next-generation DNA sequencing data (OncoPanel) from 5769 NSCLC samples (5075 patients) were queried for cases having high-level ERBB2 amplification (≥6 copies). Clinical and demographic characteristics were extracted from the electronic medical records. Efficacy of the pan-ERBB inhibitor afatinib or HER2 ADCs (trastuzumab deruxtecan and trastuzumab emtansine) was evaluated in NSCLC preclinical models and patients with ERBB2 amplification. RESULTS: High-level ERBB2 amplification was identified in 0.9% of lung adenocarcinomas and reliably predicted overexpression of HER2. ERBB2 amplification events are detected in two distinct clinicopathologic and genomic subsets of NSCLC: as the sole mitogenic driver in tumors arising in patients with a smoking history or as a concomitant alteration with other mitogenic drivers in patients with a light or never smoking history. We further reveal that trastuzumab deruxtecan is effective therapy in in vitro and in vivo preclinical models of NSCLC harboring ERBB2 amplification and report two cases of clinical activity of an anti-HER2 ADC in patients who acquired ERBB2 amplification after previous targeted therapy. CONCLUSIONS: High-level ERBB2 amplification reliably predicts HER2 overexpression in patients with NSCLC, and HER2 ADC is effective therapy in this population.

Delineating spatial cell-cell interactions in the solid tumour microenvironment through the lens of highly multiplexed imaging.

The growth and metastasis of solid tumours is known to be facilitated by the tumour microenvironment (TME), which is composed of a highly diverse collection of cell types that interact and communicate with one another extensively. Many of these interactions involve the immune cell population within the TME, referred to as the tumour immune microenvironment (TIME). These non-cell autonomous interactions exert substantial influence over cell behaviour and contribute to the reprogramming of immune and stromal cells into numerous pro-tumourigenic phenotypes. The study of some of these interactions, such as the PD-1/PD-L1 axis that induces CD8+ T cell exhaustion, has led to the development of breakthrough therapeutic advances. Yet many common analyses of the TME either do not retain the spatial data necessary to assess cell-cell interactions, or interrogate few (<10) markers, limiting the capacity for cell phenotyping. Recently developed digital pathology technologies, together with sophisticated bioimage analysis programs, now enable the high-resolution, highly-multiplexed analysis of diverse immune and stromal cell markers within the TME of clinical specimens. In this article, we review the tumour-promoting non-cell autonomous interactions in the TME and their impact on tumour behaviour. We additionally survey commonly used image analysis programs and highly-multiplexed spatial imaging technologies, and we discuss their relative advantages and limitations. The spatial organization of the TME varies enormously between patients, and so leveraging these technologies in future studies to further characterize how non-cell autonomous interactions impact tumour behaviour may inform the personalization of cancer treatment.​.

Loss of ATG4B and ATG4A results in two-stage cell cycle defects in pancreatic ductal adenocarcinoma cells.

Pancreatic ductal adenocarcinoma (PDAC) exhibits elevated levels of autophagy, which promote tumor progression and treatment resistance. ATG4B is an autophagy-related cysteine protease under consideration as a potential therapeutic target, but it is largely unexplored in PDAC. Here, we investigated the clinical and functional relevance of ATG4B expression in PDAC. Using two PDAC patient cohorts, we found that low ATG4B mRNA or protein expression is associated with worse patient survival outcomes, poorly differentiated PDAC tumors and a lack of survival benefit from adjuvant chemotherapy. In PDAC cell lines, ATG4B knockout reduced proliferation, abolished processing of LC3B (also known as MAP1LC3B), and reduced GABARAP and GABARAPL1 levels, but increased ATG4A levels. ATG4B and ATG4A double knockout lines displayed a further reduction in proliferation, characterized by delays in G1-S phase transition and mitosis. Pro-LC3B accumulated aberrantly at the centrosome with a concomitant increase in centrosomal proteins PCM1 and CEP131, which was rescued by exogenous ATG4B. The two-stage cell cycle defects following ATG4B and ATG4A loss have important therapeutic implications for PDAC.

New insights into the biology and development of lung cancer in never smokers-implications for early detection and treatment.

Lung cancer is the leading cause of cancer deaths worldwide. Despite never smokers comprising between 10 and 25% of all cases, lung cancer in never smokers (LCNS) is relatively under characterized from an etiological and biological perspective. The application of multi-omics techniques on large patient cohorts has significantly advanced the current understanding of LCNS tumor biology. By synthesizing the findings of multi-omics studies on LCNS from a clinical perspective, we can directly translate knowledge regarding tumor biology into implications for patient care. Primarily focused on never smokers with lung adenocarcinoma, this review details the predominance of driver mutations, particularly in East Asian patients, as well as the frequency and importance of germline variants in LCNS. The mutational patterns present in LCNS tumors are thoroughly explored, highlighting the high abundance of the APOBEC signature. Moreover, this review recognizes the spectrum of immune profiles present in LCNS tumors and posits how it can be translated to treatment selection. The recurring and novel insights from multi-omics studies on LCNS tumor biology have a wide range of clinical implications. Risk factors such as exposure to outdoor air pollution, second hand smoke, and potentially diet have a genomic imprint in LCNS at varying degrees, and although they do not encompass all LCNS cases, they can be leveraged to stratify risk. Germline variants similarly contribute to a notable proportion of LCNS, which warrants detailed documentation of family history of lung cancer among never smokers and demonstrates value in developing testing for pathogenic variants in never smokers for early detection in the future. Molecular driver subtypes and specific co-mutations and mutational signatures have prognostic value in LCNS and can guide treatment selection. LCNS tumors with no known driver alterations tend to be stem-like and genes contributing to this state may serve as potential therapeutic targets. Overall, the comprehensive findings of multi-omics studies exert a wide influence on clinical management and future research directions in the realm of LCNS.

Methionine-producing tumor micro(be) environment fuels growth of solid tumors.

BACKGROUND: Recent studies have uncovered the near-ubiquitous presence of microbes in solid tumors of diverse origins. Previous literature has shown the impact of specific bacterial species on the progression of cancer. We propose that local microbial dysbiosis enables certain cancer phenotypes through provisioning of essential metabolites directly to tumor cells. METHODS: 16S rDNA sequencing of 75 patient lung samples revealed the lung tumor microbiome specifically enriched for bacteria capable of producing methionine. Wild-type (WT) and methionine auxotrophic (metA mutant) E. coli cells were used to condition cell culture media and the proliferation of lung adenocarcinoma (LUAD) cells were measured using SYTO60 staining. Further, colony forming assay, Annexin V Staining, BrdU, AlamarBlue, western blot, qPCR, LINE microarray and subcutaneous injection with methionine modulated feed were used to analyze cellular proliferation, cell-cycle, cell death, methylation potential, and xenograft formation under methionine restriction. Moreover, C14-labeled glucose was used to illustrate the interplay between tumor cells and bacteria. RESULTS/DISCUSSION: Our results show bacteria found locally within the tumor microenvironment are enriched for methionine synthetic pathways, while having reduced S-adenosylmethionine metabolizing pathways. As methionine is one of nine essential amino acids that mammals are unable to synthesize de novo, we investigated a potentially novel function for the microbiome, supplying essential nutrients, such as methionine, to cancer cells. We demonstrate that LUAD cells can utilize methionine generated by bacteria to rescue phenotypes that would otherwise be inhibited due to nutrient restriction. In addition to this, with WT and metA mutant E. coli, we saw a selective advantage for bacteria with an intact methionine synthetic pathway to survive under the conditions induced by LUAD cells. These results would suggest that there is a potential bi-directional cross-talk between the local microbiome and adjacent tumor cells. In this study, we focused on methionine as one of the critical molecules, but we also hypothesize that additional bacterial metabolites may also be utilized by LUAD. Indeed, our radiolabeling data suggest that other biomolecules are shared between cancer cells and bacteria. Thus, modulating the local microbiome may have an indirect effect on tumor development, progression, and metastasis.

Combination Therapy With MDM2 and MEK Inhibitors Is Effective in Patient-Derived Models of Lung Adenocarcinoma With Concurrent Oncogenic Drivers and MDM2 Amplification.

INTRODUCTION: Although targeted therapies have revolutionized the therapeutic landscape of lung adenocarcinomas (LUADs), disease progression on single-agent targeted therapy against known oncogenic drivers is common, and therapeutic options after disease progression are limited. In patients with MDM2 amplification (MDM2amp) and a concurrent oncogenic driver alteration, we hypothesized that targeting of the tumor-suppressor pathway (by means of restoration of p53 using MDM2 inhibition) and simultaneous targeting of co-occurring MAPK oncogenic pathway might represent a more durably effective therapeutic strategy. METHODS: We evaluated genomic next-generation sequencing data using the Memorial Sloan Kettering Cancer Center-Integrated Mutation Profiling of Actionable Cancer Targets platform to nominate potential targets for combination therapy in LUAD. We investigated the small molecule MDM2 inhibitor milademetan in cell lines and patient-derived xenografts of LUAD with a known driver alteration and MDM2amp. RESULTS: Of 10,587 patient samples from 7121 patients with LUAD profiled by next-generation sequencing, 6% (410 of 7121) harbored MDM2amp. MDM2amp was significantly enriched among tumors with driver alterations in METex14 (36%, p < 0.001), EGFR (8%, p < 0.001), RET (12%, p < 0.01), and ALK (10%, p < 0.01). The combination of milademetan and the MEK inhibitor trametinib was synergistic in growth inhibition of ECLC5-GLx (TRIM33-RET/MDM2amp), LUAD12c (METex14/KRASG12S/MDM2amp), SW1573 (KRASG12C, TP53 wild type), and A549 (KRASG12S) cells and in increasing expression of proapoptotic proteins PUMA and BIM. Treatment of ECLC5-GLx and LUAD12c with single-agent milademetan increased ERK phosphorylation, consistent with previous data on ERK activation with MDM2 inhibition. This ERK activation was effectively suppressed by concomitant administration of trametinib. In contrast, ERK phosphorylation induced by milademetan was not suppressed by concurrent RET inhibition using selpercatinib (in ECLC5-GLx) or MET inhibition using capmatinib (in LUAD12c). In vivo, combination milademetan and trametinib was more effective than either agent alone in ECLC5-GLx, LX-285 (EGFRex19del/MDM2amp), L13BS1 (METex14/MDM2amp), and A549 (KRASG12S, TP53 wild type). CONCLUSIONS: Combined MDM2/MEK inhibition was found to have efficacy across multiple patient-derived LUAD models harboring MDM2amp and concurrent oncogenic drivers. This combination, potentially applicable to LUADs with a wide variety of oncogenic driver mutations and kinase fusions activating the MAPK pathway, has evident clinical implications and will be investigated as part of a planned phase 1/2 clinical trial.

Tandem mass tag-based thermal proteome profiling for the discovery of drug-protein interactions in cancer cells.

Identification of effector targets is imperative to the characterization of the mechanisms of action of novel small molecules. Here, we describe steps to identify effector drug-protein interactions in lysates derived from cancer cell lines using a thermal proteome profiling (TPP) protocol. Building on existing TTP approaches, we detail the use of an in-solution trypsin digestion technique to streamline sample preparation, a nonparametric analysis to rank proteins for prioritization, and a follow-up strategy for identifying effector interactors. For complete details on the use and execution of this protocol, please refer to Johnson et al. (2022).1.

Characterizing the secretome of EGFR mutant lung adenocarcinoma.

Background: Lung cancer is the leading cause of cancer related death worldwide, mainly due to the late stage of disease at the time of diagnosis. Non-invasive biomarkers are needed to supplement existing screening methods to enable earlier detection and increased patient survival. This is critical to EGFR-driven lung adenocarcinoma as it commonly occurs in individuals who have never smoked and do not qualify for current screening protocols. Methods: In this study, we performed mass spectrometry analysis of the secretome of cultured lung cells representing different stages of mutant EGFR driven transformation, from normal to fully malignant. Identified secreted proteins specific to the malignant state were validated using orthogonal methods and their clinical activity assessed in lung adenocarcinoma patient cohorts. Results: We quantified 1020 secreted proteins, which were compared for differential expression between stages of transformation. We validated differentially expressed proteins at the transcriptional level in clinical tumor specimens, association with patient survival, and absolute concentration to yield three biomarker candidates: MDK, GDF15, and SPINT2. These candidates were validated using ELISA and increased levels were associated with poor patient survival specifically in EGFR mutant lung adenocarcinoma patients. Conclusions: Our study provides insight into changes in secreted proteins during EGFR driven lung adenocarcinoma transformation that may play a role in the processes that promote tumor progression. The specific candidates identified can harnessed for biomarker use to identify high risk individuals for early detection screening programs and disease management for this molecular subgroup of lung adenocarcinoma patients.

MEK inhibitor resistance in lung adenocarcinoma is associated with addiction to sustained ERK suppression.

MEK inhibitors (MEKi) have limited efficacy in KRAS mutant lung adenocarcinoma (LUAD) patients, and this is attributed to both intrinsic and adaptive mechanisms of drug resistance. While many studies have focused on the former, there remains a dearth of data regarding acquired resistance to MEKi in LUAD. We established trametinib-resistant KRAS mutant LUAD cells through dose escalation and performed targeted MSK-IMPACT sequencing to identify drivers of MEKi resistance. Comparing resistant cells to their sensitive counterparts revealed alteration of genes associated with trametinib response. We describe a state of "drug addiction" in resistant cases where cells are dependent on continuous culture in trametinib for survival. We show that dependence on ERK2 suppression underlies this phenomenon and that trametinib removal hyperactivates ERK, resulting in ER stress and apoptosis. Amplification of KRASG12C occurs in drug-addicted cells and blocking mutant-specific activity with AMG 510 rescues the lethality associated with trametinib withdrawal. Furthermore, we show that increased KRASG12C expression is lethal to other KRAS mutant LUAD cells, consequential to ERK hyperactivation. Our study determines the drug-addicted phenotype in lung cancer is associated with KRAS amplification and demonstrates that toxic acquired genetic changes can develop de novo in the background of MAPK suppression with MEK inhibitors. We suggest that the presence of mutant KRAS amplification in patients may identify those that may benefit from a "drug holiday" to circumvent drug resistance. These findings demonstrate the toxic potential of hyperactive ERK signaling and highlight potential therapeutic opportunities in patients bearing KRAS mutations.

CCL5 production in lung cancer cells leads to an altered immune microenvironment and promotes tumor development.

Current immunotherapies for lung cancer are only effective in a subset of patients. Identifying tumor-derived factors that facilitate immunosuppression offers the opportunity to develop novel strategies to supplement and improve current therapeutics. We sought to determine whether expression of driver oncogenes in lung cancer cells affects cytokine secretion, alters the local immune environment, and influences lung tumor progression. We demonstrate that oncogenic EGFR and KRAS mutations, which are early events in lung tumourigenesis, can drive cytokine and chemokine production by cancer cells. One of the most prominent changes was in CCL5, which was rapidly induced by KRASG12V or EGFRL858R expression, through MAPK activation. Immunocompetent mice implanted with syngeneic KRAS-mutant lung cancer cells deficient in CCL5 have decreased regulatory T cells (Tregs), evidence of T cell exhaustion, and reduced lung tumor burden, indicating tumor-cell CCL5 production contributes to an immune suppressive environment in the lungs. Furthermore, high CCL5 expression correlates with poor prognosis, immunosuppressive regulatory T cells, and alteration to CD8 effector function in lung adenocarcinoma patients. Our data support targeting CCL5 or CCL5 receptors on immune suppressive cells to prevent formation of an immune suppressive tumor microenvironment that promotes lung cancer progression and immunotherapy insensitivity.

MET Exon 14 Splice-Site Mutations Preferentially Activate KRAS Signaling to Drive Tumourigenesis.

Targeted therapies for MET exon 14-skipping (METΔex14)-driven lung cancers have generated some promising results but response rates remain below that seen for other kinase-driven cancers. One strategy for improving treatment outcomes is to employ rational combination therapies to enhance the suppression of tumour growth and delay or prevent the emergence of resistance. To this end, we profiled the transcriptomes of MET-addicted lung tumours and cell lines and identified the RAS-mitogen-activated protein kinase (MAPK) pathway as a critical effector required for METΔex14-dependent growth. Ectopic expression of MET in an isogenic cell line model showed that overexpression of the mutant MET receptor led to higher levels of MAPK phosphorylation and nuclear import, resulting in increased expression and phosphorylation of nuclear MAPK targets. In comparison, other known MET effectors were unaffected. Inhibition of this pathway by KRAS knockdown in MET-addicted cells in vitro led to decreased viability in only the METΔex14-mutant cells. Conversely, decoupling RAS-MAPK axis, but not other effector pathways, from MET activity via the introduction of constitutively active mutants conferred resistance to MET inhibitors in vitro. Our results suggest that aberrant hyperactivity of the MET receptor caused by the exon 14-skipping mutation does not uniformly upregulate all known downstream effectors, rather gaining a predilection for aberrantly activating and subsequently relying on the RAS-MAPK pathway. These findings provide a rationale for the co-targeting of the RAS-MAPK pathway alongside MET to prolong therapeutic response and circumvent resistance to improve patient survival.

Characterization of a small molecule inhibitor of disulfide reductases that induces oxidative stress and lethality in lung cancer cells.

Phenotype-based screening can identify small molecules that elicit a desired cellular response, but additional approaches are required to characterize their targets and mechanisms of action. Here, we show that a compound termed LCS3, which selectively impairs the growth of human lung adenocarcinoma (LUAD) cells, induces oxidative stress. To identify the target that mediates this effect, we use thermal proteome profiling (TPP) and uncover the disulfide reductases GSR and TXNRD1 as targets. We confirm through enzymatic assays that LCS3 inhibits disulfide reductase activity through a reversible, uncompetitive mechanism. Further, we demonstrate that LCS3-sensitive LUAD cells are sensitive to the synergistic inhibition of glutathione and thioredoxin pathways. Lastly, a genome-wide CRISPR knockout screen identifies NQO1 loss as a mechanism of LCS3 resistance. This work highlights the ability of TPP to uncover targets of small molecules identified by high-throughput screens and demonstrates the potential therapeutic utility of inhibiting disulfide reductases in LUAD.

Emerging Molecular Dependencies of Mutant EGFR-Driven Non-Small Cell Lung Cancer.

Epidermal growth factor receptor (EGFR) mutations are the molecular driver of a subset of non-small cell lung cancers (NSCLC); tumors that harbor these mutations are often dependent on sustained oncogene signaling for survival, a concept known as "oncogene addiction". Inhibiting EGFR with tyrosine kinase inhibitors has improved clinical outcomes for patients; however, successive generations of inhibitors have failed to prevent the eventual emergence of resistance to targeted agents. Although these tumors have a well-established dependency on EGFR signaling, there remain questions about the underlying genetic mechanisms necessary for EGFR-driven oncogenesis and the factors that allow tumor cells to escape EGFR dependence. In this review, we highlight the latest findings on mutant EGFR dependencies, co-operative drivers, and molecular mechanisms that underlie sensitivity to EGFR inhibitors. Additionally, we offer perspective on how these discoveries may inform novel combination therapies tailored to EGFR mutant NSCLC.

Extracellular signal-regulated kinase mediates chromatin rewiring and lineage transformation in lung cancer.

Lineage transformation between lung cancer subtypes is a poorly understood phenomenon associated with resistance to treatment and poor patient outcomes. Here, we aimed to model this transition to define underlying biological mechanisms and identify potential avenues for therapeutic intervention. Small cell lung cancer (SCLC) is neuroendocrine in identity and, in contrast to non-SCLC (NSCLC), rarely contains mutations that drive the MAPK pathway. Likewise, NSCLCs that transform to SCLC concomitantly with development of therapy resistance downregulate MAPK signaling, suggesting an inverse relationship between pathway activation and lineage state. To test this, we activated MAPK in SCLC through conditional expression of mutant KRAS or EGFR, which revealed suppression of the neuroendocrine differentiation program via ERK. We found that ERK induces the expression of ETS factors that mediate transformation into a NSCLC-like state. ATAC-seq demonstrated ERK-driven changes in chromatin accessibility at putative regulatory regions and global chromatin rewiring at neuroendocrine and ETS transcriptional targets. Further, ERK-mediated induction of ETS factors as well as suppression of neuroendocrine differentiation were dependent on histone acetyltransferase activities of CBP/p300. Overall, we describe how the ERK-CBP/p300-ETS axis promotes a lineage shift between neuroendocrine and non-neuroendocrine lung cancer phenotypes and provide rationale for the disruption of this program during transformation-driven resistance to targeted therapy.

Resistance to BET inhibitors in lung adenocarcinoma is mediated by casein kinase phosphorylation of BRD4.

Targeting the epigenome to modulate gene expression programs driving cancer development has emerged as an exciting avenue for therapeutic intervention. Pharmacological inhibition of the bromodomain and extraterminal (BET) family of chromatin adapter proteins has proven effective in this regard, suppressing growth of diverse cancer types mainly through downregulation of the c-MYC oncogene, and its downstream transcriptional program. While initially effective, resistance to BET inhibitors (BETi) typically occurs through mechanisms that reactivate MYC expression. We have previously shown that lung adenocarcinoma (LAC) is inhibited by JQ1 through suppression of FOSL1, suggesting that the epigenetic landscape of tumor cells from different origins and differentiation states influences BETi response. Here, we assessed how these differences affect mechanisms of BETi resistance through the establishment of isogenic pairs of JQ1 sensitive and resistant LAC cell lines. We found that resistance to JQ1 in LAC occurs independent of FOSL1 while MYC levels remain unchanged between resistant cells and their JQ1-treated parental counterparts. Furthermore, while epithelial-mesenchymal transition (EMT) is observed upon resistance, TGF-ß induced EMT did not confer resistance in JQ1 sensitive LAC lines, suggesting this is a consequence, rather than a driver of BETi resistance in our model systems. Importantly, siRNA knockdown demonstrated that JQ1 resistant cell lines are still dependent on BRD4 expression for survival and we found that phosphorylation of BRD4 is elevated in resistant LACs, identifying casein kinase 2 (CK2) as a candidate protein mediating this effect. Inhibition of CK2, as well as downstream transcriptional targets of phosphorylated BRD4-including AXL and activators of the PI3K pathway-synergize with JQ1 to inhibit BETi resistant LAC. Overall, this demonstrates that the mechanism of resistance to BETi varies depending on cancer type, with LAC cells developing JQ1 resistance independent of MYC regulation, and identifying CK2 phosphorylation of BRD4 as a potential target to overcome resistance in this cancer.

Erratum: MicroRNA Modification of Coxsackievirus B3 Decreases Its Toxicity, while Retaining Oncolytic Potency against Lung Cancer.

[This corrects the article DOI: 10.1016/j.omto.2020.01.002.].

An ErbB2 splice variant lacking exon 16 drives lung carcinoma.

Lung cancer causes more deaths annually than any other malignancy. A subset of non-small cell lung cancer (NSCLC) is driven by amplification and overexpression or activating mutation of the receptor tyrosine kinase (RTK) ERBB2 In some contexts, notably breast cancer, alternative splicing of ERBB2 causes skipping of exon 16, leading to the expression of an oncogenic ERBB2 isoform (ERBB2ΔEx16) that forms constitutively active homodimers. However, the broader implications of ERBB2 alternative splicing in human cancers have not been explored. Here, we have used genomic and transcriptomic analysis to identify elevated ERBB2ΔEx16 expression in a subset of NSCLC cases, as well as splicing site mutations facilitating exon 16 skipping and deletions of exon 16 in a subset of these lung tumors and in a number of other carcinomas. Supporting the potential of ERBB2ΔEx16 as a lung cancer driver, its expression transformed immortalized lung epithelial cells while a transgenic model featuring inducible ERBB2ΔEx16 specifically in the lung epithelium rapidly developed lung adenocarcinomas following transgene induction. Collectively, these observations indicate that ERBB2ΔEx16 is a lung cancer oncogene with potential clinical importance for a proportion of patients.

Drug Sensitivity and Allele Specificity of First-Line Osimertinib Resistance EGFR Mutations.

Osimertinib, a mutant-specific third-generation EGFR tyrosine kinase inhibitor, is emerging as the preferred first-line therapy for EGFR-mutant lung cancer, yet resistance inevitably develops in patients. We modeled acquired resistance to osimertinib in transgenic mouse models of EGFRL858R -induced lung adenocarcinoma and found that it is mediated largely through secondary mutations in EGFR-either C797S or L718V/Q. Analysis of circulating free DNA data from patients revealed that L718Q/V mutations almost always occur in the context of an L858R driver mutation. Therapeutic testing in mice revealed that both erlotinib and afatinib caused regression of osimertinib-resistant C797S-containing tumors, whereas only afatinib was effective on L718Q mutant tumors. Combination first-line osimertinib plus erlotinib treatment prevented the emergence of secondary mutations in EGFR. These findings highlight how knowledge of the specific characteristics of resistance mutations is important for determining potential subsequent treatment approaches and suggest strategies to overcome or prevent osimertinib resistance in vivo. SIGNIFICANCE: This study provides insight into the biological and molecular properties of osimertinib resistance EGFR mutations and evaluates therapeutic strategies to overcome resistance. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/10/2017/F1.large.jpg.

MicroRNA Modification of Coxsackievirus B3 Decreases Its Toxicity, while Retaining Oncolytic Potency against Lung Cancer.

We recently discovered that coxsackievirus B3 (CVB3) is a potent oncolytic virus against KRAS mutant lung adenocarcinoma. Nevertheless, the evident toxicity restricts the use of wild-type (WT)-CVB3 for cancer therapy. The current study aims to engineer the CVB3 to decrease its toxicity and to extend our previous research to determine its safety and efficacy in treating TP53/RB1 mutant small-cell lung cancer (SCLC). A microRNA-modified CVB3 (miR-CVB3) was generated via inserting multiple copies of tumor-suppressive miR-145/miR-143 target sequences into the viral genome. In vitro experiments revealed that miR-CVB3 retained the ability to infect and lyse KRAS mutant lung adenocarcinoma and TP53/RB1-mutant SCLC cells, but with a markedly reduced cytotoxicity toward cardiomyocytes. In vivo study using a TP53/RB1-mutant SCLC xenograft model demonstrated that a single dose of miR-CVB3 via systemic administration resulted in a significant tumor regression. Most strikingly, mice treated with miR-CVB3 exhibited greatly attenuated cardiotoxicities and decreased viral titers compared to WT-CVB3-treated mice. Collectively, we generated a recombinant CVB3 that is powerful in destroying both KRAS mutant lung adenocarcinoma and TP53/RB1-mutant SCLC, with a negligible toxicity toward normal tissues. Future investigation is needed to address the issue of genome instability of miR-CVB3, which was observed in ~40% of mice after a prolonged treatment.

Epithelial tumor suppressor ELF3 is a lineage-specific amplified oncogene in lung adenocarcinoma.

Gene function in cancer is often cell type-specific. The epithelial cell-specific transcription factor ELF3 is a documented tumor suppressor in many epithelial tumors yet displays oncogenic properties in others. Here, we show that ELF3 is an oncogene in the adenocarcinoma subtype of lung cancer (LUAD), providing genetic, functional, and clinical evidence of subtype specificity. We discover a region of focal amplification at chromosome 1q32.1 encompassing the ELF3 locus in LUAD which is absent in the squamous subtype. Gene dosage and promoter hypomethylation affect the locus in up to 80% of LUAD analyzed. ELF3 expression was required for tumor growth and a pan-cancer expression network analysis supports its subtype and tissue specificity. We further show that ELF3 displays strong prognostic value in LUAD but not LUSC. We conclude that, contrary to many other tumors of epithelial origin, ELF3 is an oncogene and putative therapeutic target in LUAD.