ABSTRACT
The majority of EGFR mutant lung adenocarcinomas respond well to EGFR tyrosine kinase inhibitors (TKI). However, most of these responses are partial, with drug-tolerant residual disease remaining even at the time of maximal response. This residual disease can ultimately lead to relapses, which eventually develop in most patients. To investigate the cellular and molecular properties of residual tumor cells in vivo, we leveraged patient-derived xenograft (PDX) models of EGFR mutant lung cancer. Subcutaneous EGFR mutant PDXs were treated with the third-generation TKI osimertinib until maximal tumor regression. Residual tissue inevitably harbored tumor cells that were transcriptionally distinct from bulk pretreatment tumor. Single-cell transcriptional profiling provided evidence of cells matching the profiles of drug-tolerant cells present in the pretreatment tumor. In one of the PDXs analyzed, osimertinib treatment caused dramatic transcriptomic changes that featured upregulation of the neuroendocrine lineage transcription factor ASCL1. Mechanistically, ASCL1 conferred drug tolerance by initiating an epithelial-to-mesenchymal gene-expression program in permissive cellular contexts. This study reveals fundamental insights into the biology of drug tolerance, the plasticity of cells through TKI treatment, and why specific phenotypes are observed only in certain tumors. SIGNIFICANCE: Analysis of residual disease following tyrosine kinase inhibitor treatment identified heterogeneous and context-specific mechanisms of drug tolerance in lung cancer that could lead to the development of strategies to forestall drug resistance. See related commentary by Rumde and Burns, p. 1188.
Subject(s)
Acrylamides , Carcinoma, Non-Small-Cell Lung , Indoles , Lung Neoplasms , Pyrimidines , Humans , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Lung Neoplasms/pathology , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/genetics , Carcinoma, Non-Small-Cell Lung/pathology , ErbB Receptors/genetics , Drug Resistance, Neoplasm/genetics , Neoplasm Recurrence, Local/drug therapy , Aniline Compounds/pharmacology , Aniline Compounds/therapeutic use , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , Mutation , Basic Helix-Loop-Helix Transcription Factors/geneticsABSTRACT
For over a century, early researchers sought to study biological organisms in a laboratory setting, leading to the generation of both in vitro and in vivo model systems. Patient-derived models of cancer (PDMCs) have more recently come to the forefront of preclinical cancer models and are even finding their way into clinical practice as part of functional precision medicine programs. The PDMC Consortium, supported by the Division of Cancer Biology in the National Cancer Institute of the National Institutes of Health, seeks to understand the biological principles that govern the various PDMC behaviors, particularly in response to perturbagens, such as cancer therapeutics. Based on collective experience from the consortium groups, we provide insight regarding PDMCs established both in vitro and in vivo, with a focus on practical matters related to developing and maintaining key cancer models through a series of vignettes. Although every model has the potential to offer valuable insights, the choice of the right model should be guided by the research question. However, recognizing the inherent constraints in each model is crucial. Our objective here is to delineate the strengths and limitations of each model as established by individual vignettes. Further advances in PDMCs and the development of novel model systems will enable us to better understand human biology and improve the study of human pathology in the lab.
ABSTRACT
Acquired resistance to tyrosine kinase inhibitors (TKI), such as osimertinib used to treat EGFR-mutant lung adenocarcinomas, limits long-term efficacy and is frequently caused by non-genetic mechanisms. Here, we define the chromatin accessibility and gene regulatory signatures of osimertinib sensitive and resistant EGFR-mutant cell and patient-derived models and uncover a role for mammalian SWI/SNF chromatin remodeling complexes in TKI resistance. By profiling mSWI/SNF genome-wide localization, we identify both shared and cancer cell line-specific gene targets underlying the resistant state. Importantly, genetic and pharmacologic disruption of the SMARCA4/SMARCA2 mSWI/SNF ATPases re-sensitizes a subset of resistant models to osimertinib via inhibition of mSWI/SNF-mediated regulation of cellular programs governing cell proliferation, epithelial-to-mesenchymal transition, epithelial cell differentiation, and NRF2 signaling. These data highlight the role of mSWI/SNF complexes in supporting TKI resistance and suggest potential utility of mSWI/SNF inhibitors in TKI-resistant lung cancers.
Subject(s)
Lung Neoplasms , Animals , Humans , Chromatin Assembly and Disassembly , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Lung Neoplasms/pathology , Chromatin , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , ErbB Receptors/genetics , Mutation , Mammals/genetics , DNA Helicases/genetics , Nuclear Proteins/genetics , Transcription Factors/geneticsABSTRACT
The last few years have seen an increasing number of discoveries which collectively demonstrate that histone and DNA modifying enzyme modulate different stages of metastasis. Moreover, epigenomic alterations can now be measured at multiple scales of analysis and are detectable in human tumors or liquid biopsies. Malignant cell clones with a proclivity for relapse in certain organs may arise in the primary tumor as a consequence of epigenomic alterations which cause a loss in lineage integrity. These alterations may occur due to genetic aberrations acquired during tumor progression or concomitant to therapeutic response. Moreover, evolution of the stroma can also alter the epigenome of cancer cells. In this review, we highlight current knowledge with a particular emphasis on leveraging chromatin and DNA modifying mechanisms as biomarkers of disseminated disease and as therapeutic targets to treat metastatic cancers.
Subject(s)
Epigenomics , Neoplasms , Humans , Histones/genetics , Histones/metabolism , Neoplasms/genetics , Neoplasms/therapy , DNA Methylation , DNA , Epigenesis, GeneticABSTRACT
The brain is a major sanctuary site for metastatic cancer cells that evade systemic therapies. Through pre-clinical pharmacological, biological, and molecular studies, we characterize the functional link between drug resistance and central nervous system (CNS) relapse in Epidermal Growth Factor Receptor- (EGFR-) mutant non-small cell lung cancer, which can progress in the brain when treated with the CNS-penetrant EGFR inhibitor osimertinib. Despite widespread osimertinib distribution in vivo, the brain microvascular tumor microenvironment (TME) is associated with the persistence of malignant cell sub-populations, which are poised to proliferate in the brain as osimertinib-resistant lesions over time. Cellular and molecular features of this poised state are regulated through a Ras homolog family member A (RhoA) and Serum Responsive Factor (SRF) gene expression program. RhoA potentiates the outgrowth of disseminated tumor cells on osimertinib treatment, preferentially in response to extracellular laminin and in the brain. Thus, we identify pre-existing and adaptive features of metastatic and drug-resistant cancer cells, which are enhanced by RhoA/SRF signaling and the brain TME during the evolution of osimertinib-resistant disease.
Subject(s)
Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , Humans , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Lung Neoplasms/pathology , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/genetics , Carcinoma, Non-Small-Cell Lung/pathology , rhoA GTP-Binding Protein/genetics , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , Neoplasm Recurrence, Local/drug therapy , ErbB Receptors/genetics , Aniline Compounds/pharmacology , Aniline Compounds/therapeutic use , Brain/pathology , Mutation , Drug Resistance, Neoplasm/genetics , Tumor MicroenvironmentABSTRACT
Metastatic breast cancer remains a major cause of cancer-related deaths in women, and there are few effective therapies against this advanced disease. Emerging evidence suggests that key steps of tumor progression and metastasis are controlled by reversible epigenetic mechanisms. Using an in vivo genetic screen, we identified WDR5 as an actionable epigenetic regulator that is required for metastatic progression in models of triple-negative breast cancer. We found that knockdown of WDR5 in breast cancer cells independently impaired their tumorigenic as well as metastatic capabilities. Mechanistically, WDR5 promotes cell growth by increasing ribosomal gene expression and translation efficiency in a KMT2-independent manner. Consistently, pharmacological inhibition or degradation of WDR5 impedes cellular translation rate and the clonogenic ability of breast cancer cells. Furthermore, a combination of WDR5 targeting with mTOR inhibitors leads to potent suppression of translation and proliferation of breast cancer cells. These results reveal novel therapeutic strategies to treat metastatic breast cancer.
Subject(s)
Breast Neoplasms , Humans , Female , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Histone-Lysine N-Methyltransferase/metabolism , Cell Line, Tumor , Intracellular Signaling Peptides and Proteins/genetics , Cell ProliferationABSTRACT
Brain metastases are the most common brain malignancy. This review discusses the studies presented at the third annual meeting of the Melanoma Research Foundation in the context of other recent reports on the biology and treatment of melanoma brain metastases (MBM). Although symptomatic MBM patients were historically excluded from immunotherapy trials, efforts from clinicians and patient advocates have resulted in more inclusive and even dedicated clinical trials for MBM patients. The results of checkpoint inhibitor trials were discussed in conversation with current standards of care for MBM patients, including steroids, radiotherapy, and targeted therapy. Advances in the basic scientific understanding of MBM, including the role of astrocytes and metabolic adaptations to the brain microenvironment, are exposing new vulnerabilities which could be exploited for therapeutic purposes. Technical advances including single-cell omics and multiplex imaging are expanding our understanding of the MBM ecosystem and its response to therapy. This unprecedented level of spatial and temporal resolution is expected to dramatically advance the field in the coming years and render novel treatment approaches that might improve MBM patient outcomes.
Subject(s)
Brain Neoplasms , Melanoma , Neoplasms, Second Primary , Humans , Ecosystem , Melanoma/pathology , Brain Neoplasms/therapy , Brain Neoplasms/secondary , Immunotherapy/methods , Neoplasms, Second Primary/pathology , Brain , Tumor MicroenvironmentABSTRACT
US medical schools increasingly seek ways to reduce costs and improve productivity. One aspect of this effort has been the development of performance-based incentives for individual faculty. A myriad of such plans exist. Typically, they incentivize clinical revenue generation but vary widely in how teaching, investigation, and administrative contributions are recognized. In Pathology at Yale, we have developed a transparent metrically driven approach that recognizes all missions and allows faculty significant control over their career path. Although some metrics derive from traditional measures such as workload relative value units and one's level of grant support, the key concept underpinning our approach is to define one's contributions not in terms of the revenue generated, but rather on the effort devoted to each of our missions, benchmarked against national or local standards. Full-time faculty are paid a competitive rank-based salary and are expected to contribute at least 100% effort in support of the school's missions: clinical, research, education, administration, and professional service. Metrics define the effort assigned to each activity. Faculty achieving greater than 100% effort receive bonus compensation in proportion to their excess effort. By codifying explicitly how such effort is recognized into a single metric (% effort), we achieve a process that better aligns the professional and personal goals of faculty with the aims of the school. To facilitate its implementation, we have developed a web-based software platform called SWAY (Standardized Workload Analysis at Yale) that enables faculty to monitor their progress and record their activities in real time.
ABSTRACT
Experimental preclinical models have been a cornerstone of lung cancer translational research. Work in these model systems has provided insights into the biology of lung cancer subtypes and their origins, contributed to our understanding of the mechanisms that underlie tumor progression, and revealed new therapeutic vulnerabilities. Initially patient-derived lung cancer cell lines were the main preclinical models available. The landscape is very different now with numerous preclinical models for research each with unique characteristics. These include genetically engineered mouse models (GEMMs), patient-derived xenografts (PDXs) and three-dimensional culture systems ("organoid" cultures). Here we review the development and applications of these models and describe their contributions to lung cancer research.
Subject(s)
Lung Neoplasms , Animals , Disease Models, Animal , Humans , Lung Neoplasms/etiology , Lung Neoplasms/therapy , Mice , Organoids , Translational Research, BiomedicalABSTRACT
PURPOSE: Osimertinib is a potent and selective EGFR tyrosine kinase inhibitor (EGFR-TKI) of both sensitizing and T790M resistance mutations. To treat metastatic brain disease, blood-brain barrier (BBB) permeability is considered desirable for increasing clinical efficacy. EXPERIMENTAL DESIGN: We examined the level of brain penetration for 16 irreversible and reversible EGFR-TKIs using multiple in vitro and in vivo BBB preclinical models. RESULTS: In vitro osimertinib was the weakest substrate for human BBB efflux transporters (efflux ratio 3.2). In vivo rat free brain to free plasma ratios (Kpuu) show osimertinib has the most BBB penetrance (0.21), compared with the other TKIs (Kpuu ≤ 0.12). PET imaging in Cynomolgus macaques demonstrated osimertinib was the only TKI among those tested to achieve significant brain penetrance (C max %ID 1.5, brain/blood Kp 2.6). Desorption electrospray ionization mass spectroscopy images of brains from mouse PC9 macrometastases models showed osimertinib readily distributes across both healthy brain and tumor tissue. Comparison of osimertinib with the poorly BBB penetrant afatinib in a mouse PC9 model of subclinical brain metastases showed only osimertinib has a significant effect on rate of brain tumor growth. CONCLUSIONS: These preclinical studies indicate that osimertinib can achieve significant exposure in the brain compared with the other EGFR-TKIs tested and supports the ongoing clinical evaluation of osimertinib for the treatment of EGFR-mutant brain metastasis. This work also demonstrates the link between low in vitro transporter efflux ratios and increased brain penetrance in vivo supporting the use of in vitro transporter assays as an early screen in drug discovery.
Subject(s)
Acrylamides/pharmacokinetics , Aniline Compounds/pharmacokinetics , Blood-Brain Barrier/metabolism , Brain Neoplasms/drug therapy , Lung Neoplasms/drug therapy , Protein Kinase Inhibitors/pharmacokinetics , Acrylamides/administration & dosage , Aniline Compounds/administration & dosage , Animals , Brain Neoplasms/secondary , Dogs , ErbB Receptors/antagonists & inhibitors , Humans , Lung Neoplasms/pathology , Macaca fascicularis , Madin Darby Canine Kidney Cells , Male , Mice , Permeability , Protein Kinase Inhibitors/administration & dosage , Rats , Tissue Distribution , Xenograft Model Antitumor AssaysABSTRACT
Lineage selective transcription factors (TFs) are important regulators of tumorigenesis, but their biological functions are often context dependent with undefined epigenetic mechanisms of action. In this study, we uncover a conditional role for the endodermal and pulmonary specifying TF GATA6 in lung adenocarcinoma (LUAD) progression. Impairing Gata6 in genetically engineered mouse models reduces the proliferation and increases the differentiation of Kras mutant LUAD tumors. These effects are influenced by the epithelial cell type that is targeted for transformation and genetic context of Kras-mediated tumor initiation. In LUAD cells derived from surfactant protein C expressing progenitors, we identify multiple genomic loci that are bound by GATA6. Moreover, suppression of Gata6 in these cells significantly alters chromatin accessibility, particularly at distal enhancer elements. Analogous to its paradoxical activity in lung development, GATA6 expression fluctuates during different stages of LUAD progression and can epigenetically control diverse transcriptional programs associated with bone morphogenetic protein signaling, alveolar specification, and tumor suppression. These findings reveal how GATA6 can modulate the chromatin landscape of lung cancer cells to control their proliferation and divergent lineage dependencies during tumor progression.
Subject(s)
Adenocarcinoma of Lung/genetics , GATA6 Transcription Factor/genetics , Lung Neoplasms/genetics , Proto-Oncogene Proteins p21(ras)/genetics , Adenocarcinoma of Lung/pathology , Animals , Carcinogenesis/genetics , Cell Differentiation/genetics , Cell Lineage/genetics , Cell Proliferation/genetics , Chromatin/genetics , Disease Models, Animal , Gene Expression Regulation, Neoplastic/genetics , Humans , Lung/metabolism , Lung/pathology , Lung Neoplasms/pathology , MiceABSTRACT
BACKGROUND: Few somatic mutations have been linked to breast cancer metastasis, whereas transcriptomic differences among primary tumors correlate with incidence of metastasis, especially to the lungs and brain. However, the epigenomic alterations and transcription factors (TFs) which underlie these alterations remain unclear. METHODS: To identify these, we performed RNA-seq, Chromatin Immunoprecipitation and sequencing (ChIP-seq) and Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) of the MDA-MB-231 cell line and its brain (BrM2) and lung (LM2) metastatic sub-populations. We incorporated ATAC-seq data from TCGA to assess metastatic open chromatin signatures, and gene expression data from human metastatic datasets to nominate transcription factor biomarkers. RESULTS: Our integrated epigenomic analyses found that lung and brain metastatic cells exhibit both shared and distinctive signatures of active chromatin. Notably, metastatic sub-populations exhibit increased activation of both promoters and enhancers. We also integrated these data with chromosome conformation capture coupled with ChIP-seq (HiChIP) derived enhancer-promoter interactions to predict enhancer-controlled pathway alterations. We found that enhancer changes are associated with endothelial cell migration in LM2, and negative regulation of epithelial cell proliferation in BrM2. Promoter changes are associated with vasculature development in LM2 and homophilic cell adhesion in BrM2. Using ATAC-seq, we identified a metastasis open-chromatin signature that is elevated in basal-like and HER2-enriched breast cancer subtypes and associates with worse prognosis in human samples. We further uncovered TFs associated with the open chromatin landscapes of metastatic cells and whose expression correlates with risk for metastasis. While some of these TFs are associated with primary breast tumor subtypes, others more specifically correlate with lung or brain metastasis. CONCLUSIONS: We identify distinctive epigenomic properties of breast cancer cells that metastasize to the lung and brain. We also demonstrate that signatures of active chromatin sites are partially linked to human breast cancer subtypes with poor prognosis, and that specific TFs can independently distinguish lung and brain relapse.
Subject(s)
Brain Neoplasms , Breast Neoplasms , Chromatin , Lung Neoplasms , Neoplasm Proteins , Transcription Factors , Base Sequence , Brain/metabolism , Brain/pathology , Brain Neoplasms/genetics , Brain Neoplasms/metabolism , Brain Neoplasms/pathology , Brain Neoplasms/secondary , Breast Neoplasms/genetics , Breast Neoplasms/metabolism , Breast Neoplasms/pathology , Cell Line, Tumor , Chromatin/genetics , Chromatin/metabolism , Chromatin/pathology , Chromatin Immunoprecipitation Sequencing , Female , Humans , Lung/metabolism , Lung/pathology , Lung Neoplasms/genetics , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Lung Neoplasms/secondary , Neoplasm Metastasis , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Transcription Factors/genetics , Transcription Factors/metabolismABSTRACT
The integrated stress response (ISR) is a conserved pathway that is activated by cells that are exposed to stress. In lung adenocarcinoma, activation of the ATF4 branch of the ISR by certain oncogenic mutations has been linked to the regulation of amino acid metabolism. In the present study, we provide evidence for ATF4 activation across multiple stages and molecular subtypes of human lung adenocarcinoma. In response to extracellular amino acid limitation, lung adenocarcinoma cells with diverse genotypes commonly induce ATF4 in an eIF2α-dependent manner, which can be blocked pharmacologically using an ISR inhibitor. Although suppressing eIF2α or ATF4 can trigger different biological consequences, adaptive cell-cycle progression and cell migration are particularly sensitive to inhibition of the ISR. These phenotypes require the ATF4 target gene asparagine synthetase (ASNS), which maintains protein translation independently of the mTOR/PI3K pathway. Moreover, NRF2 protein levels and oxidative stress can be modulated by the ISR downstream of ASNS. Finally, we demonstrate that ASNS controls the biosynthesis of select proteins, including the cell-cycle regulator cyclin B1, which are associated with poor lung adenocarcinoma patient outcome. Our findings uncover new regulatory layers of the ISR pathway and its control of proteostasis in lung cancer cells. IMPLICATIONS: We reveal novel regulatory mechanisms by which the ISR controls selective protein translation and is required for cell-cycle progression and migration of lung cancer cells.
Subject(s)
Activating Transcription Factor 4/genetics , Adenocarcinoma of Lung/genetics , Eukaryotic Initiation Factor-2/genetics , Stress, Physiological/genetics , Activating Transcription Factor 4/metabolism , Adenocarcinoma of Lung/pathology , Amino Acids/genetics , Amino Acids/metabolism , Carbon-Nitrogen Ligases with Glutamine as Amide-N-Donor/genetics , Cell Line, Tumor , Cell Proliferation/genetics , Cyclin B1/genetics , Gene Expression Regulation, Neoplastic , Humans , NF-E2-Related Factor 2/genetics , Oxidative Stress/genetics , Phosphatidylinositol 3-Kinases/genetics , Protein Biosynthesis , Proteostasis , Signal Transduction , TOR Serine-Threonine Kinases/geneticsABSTRACT
Mutations associated with tumor development in certain tissues can be nontumorigenic in others, yet the mechanisms underlying these different outcomes remains poorly understood. To address this, we targeted an activating Hras mutation to hair follicle stem cells and discovered that Hras mutant cells outcompete wild-type neighbors yet are integrated into clinically normal skin hair follicles. In contrast, targeting the Hras mutation to the upper noncycling region of the skin epithelium leads to benign outgrowths. Follicular Hras mutant cells autonomously and nonautonomously enhance regeneration, which directs mutant cells into continuous tissue cycling to promote integration rather than aberrancy. This follicular tolerance is maintained under additional challenges that promote tumorigenesis in the epidermis, including aging, injury, and a secondary mutation. Thus, the hair follicle possesses a unique, enhanced capacity to integrate and contain Hras mutant cells within both homeostatic and perturbed tissue, demonstrating that in the skin, multiple, distinct mechanisms exist to suppress oncogenic growth.
Subject(s)
Carcinogenesis , Hair Follicle/metabolism , Neoplasms/metabolism , Neoplasms/pathology , Regeneration , ras Proteins/metabolism , Animals , Mice , Mice, TransgenicABSTRACT
BACKGROUND: Little is known about tumor-associated vasogenic edema in brain metastasis, yet it causes significant morbidity and mortality. Our purpose was to characterize edema in patients treated with anti-PD-1 and to study potential causes of vessel leakage in humans and in pre-clinical models. METHODS: We analyzed tumor and edema volume in 18 non-small cell lung (NSCLC) and 18 melanoma patients with untreated brain metastases treated with pembrolizumab on a phase II clinical trial. Melanoma brain metastases were stained with anti-CD34 to assess vessel density and its association with edema. We employed an in vitro model of the blood-brain barrier using short-term cultures from melanoma brain and extracranial metastases to determine tight junction resistance as a measure of vessel leakiness. RESULTS: Edema volumes are similar in NSCLC and melanoma brain metastases. While larger tumors tended to have more edema, the correlation was weak (R2 = 0.30). Patients responding to pembrolizumab had concurrent shrinkage of edema volume and vice versa (R2 = 0.81). Vessel density was independent of the degree of edema (R2 = 0.037). Melanoma brain metastasis cells in culture caused loss of tight junction resistance in an in vitro blood-brain barrier model system in some cases, whereas extracerebral cell cultures did not. CONCLUSIONS: Edema itself should not preclude using anti-PD-1 with caution, as sensitive tumors have resultant decreases in edema, and anti-PD-1 itself does not exacerbate edema in sensitive tumors. Additional factors aside from tumor mass effect and vessel density cause perilesional edema. Melanoma cells themselves can cause decline in tight junction resistance in a system void of immune cells, suggesting they secrete factors that cause leakiness, which might be harnessed for pharmacologic targeting in patients with significant perilesional edema.
Subject(s)
Antibodies, Monoclonal, Humanized/administration & dosage , Antineoplastic Agents, Immunological/administration & dosage , Brain Edema/diagnostic imaging , Brain Neoplasms/secondary , Carcinoma, Non-Small-Cell Lung/drug therapy , Lung Neoplasms/drug therapy , Melanoma/drug therapy , Administration, Intravenous , Antibodies, Monoclonal, Humanized/pharmacology , Antigens, CD34/metabolism , Antineoplastic Agents, Immunological/pharmacology , Blood-Brain Barrier/immunology , Brain Edema/immunology , Brain Neoplasms/blood supply , Clinical Trials, Phase II as Topic , Drug Administration Schedule , Humans , Retrospective Studies , Tight Junctions/immunology , Treatment Outcome , Tumor Cells, CulturedABSTRACT
The brain is a major site of relapse for several cancers, yet deciphering the mechanisms of brain metastasis remains a challenge because of the complexity of the brain tumor microenvironment (TME). To define the molecular landscape of brain metastasis from intact tissue in vivo, we employ an RNA-sequencing-based approach, which leverages the transcriptome of xenografts and distinguishes tumor cell and stromal gene expression with improved sensitivity and accuracy. Our data reveal shifts in epithelial and neuronal-like lineage programs in malignant cells as they adapt to the brain TME and the reciprocal neuroinflammatory response of the stroma. We identify several transcriptional hallmarks of metastasis that are specific to particular regions of the brain, induced across multiple tumor types, and confirmed in syngeneic models and patient biopsies. These data may serve as a resource for exploring mechanisms of TME co-adaptation within, as well as across, different subtypes of brain metastasis.
Subject(s)
Biomarkers, Tumor/genetics , Brain Neoplasms/secondary , Inflammation/pathology , Neoplasms/pathology , Neuronal Plasticity/genetics , Stromal Cells/pathology , Tumor Microenvironment/genetics , Animals , Brain Neoplasms/genetics , Brain Neoplasms/metabolism , Cell Lineage , Female , High-Throughput Nucleotide Sequencing , Humans , Inflammation/genetics , Inflammation/metabolism , Male , Mice , Mice, Nude , Neoplasms/genetics , Neoplasms/metabolism , Stromal Cells/metabolism , Transcriptome , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
Lung cancer is a deadly treatment refractory disease that is biologically heterogeneous. To understand and effectively treat the full clinical spectrum of thoracic malignancies, additional animal models that can recapitulate diverse human lung cancer subtypes and stages are needed. Allograft or xenograft models are versatile and enable the quantification of tumorigenic capacity in vivo, using malignant cells of either murine or human origin. However, previously described methods of lung cancer cell engraftment have been performed in non-physiological sites, such as the flank of mice, due to the inefficiency of orthotopic transplantation of cells into the lungs. In this study, we describe a method to enhance orthotopic lung cancer cell engraftment by pre-conditioning the airways of mice with the fibrosis inducing agent bleomycin. As a proof-of-concept experiment, we applied this approach to engraft tumor cells of the lung adenocarcinoma subtype, obtained from either mouse or human sources, into various strains of mice. We demonstrate that injuring the airways with bleomycin prior to tumor cell injection increases the engraftment of tumor cells from 0-17% to 71-100%. Significantly, this method enhanced lung tumor incidence and subsequent outgrowth using different models and mouse strains. In addition, engrafted lung cancer cells disseminate from the lungs into relevant distant organs. Thus, we provide a protocol that can be used to establish and maintain new orthotopic models of lung cancer with limiting amounts of cells or biospecimen and to quantitatively assess the tumorigenic capacity of lung cancer cells in physiologically relevant settings.
Subject(s)
Antibiotics, Antineoplastic/therapeutic use , Bleomycin/therapeutic use , Lung Neoplasms/drug therapy , Lung/pathology , Animals , Antibiotics, Antineoplastic/pharmacology , Bleomycin/pharmacology , Disease Models, Animal , Lung Neoplasms/pathology , MiceABSTRACT
Mechanisms underlying the propensity of latent lung adenocarcinoma (LUAD) to relapse are poorly understood. In this study, we show how differential expression of a network of extracellular matrix (ECM) molecules and their interacting proteins contributes to risk of relapse in distinct LUAD subtypes. Overexpression of the hyaluronan receptor HMMR in primary LUAD was associated with an inflammatory molecular signature and poor prognosis. Attenuating HMMR in LUAD cells diminished their ability to initiate lung tumors and distant metastases. HMMR upregulation was not required for dissemination in vivo, but enhanced ECM-mediated signaling, LUAD cell survival, and micrometastasis expansion in hyaluronan-rich microenvironments in the lung and brain metastatic niches. Our findings reveal an important mechanism by which disseminated cancer cells can coopt the inflammatory ECM to persist, leading to brain metastatic outgrowths. Cancer Res; 77(8); 1905-17. ©2017 AACR.
Subject(s)
Adenocarcinoma/metabolism , Adenocarcinoma/pathology , Extracellular Matrix Proteins/biosynthesis , Hyaluronan Receptors/biosynthesis , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Adenocarcinoma/genetics , Adenocarcinoma of Lung , Animals , Brain Neoplasms/metabolism , Brain Neoplasms/secondary , Cell Line, Tumor , Extracellular Matrix/genetics , Extracellular Matrix/metabolism , Extracellular Matrix/pathology , Extracellular Matrix Proteins/genetics , Gene Expression , Humans , Hyaluronan Receptors/genetics , Lung Neoplasms/genetics , Male , Mice , Mice, Nude , Neoplasm Micrometastasis , Transcriptome , Tumor MicroenvironmentABSTRACT
Glycosylation is one of the most predominant forms of cell-surface protein modifications, yet its deregulation in cancer and contribution to tumor microenvironment interactions remain poorly understood. In this issue of Cancer Discovery, Reticker-Flynn and Bhatia characterize an enzymatic switch in lung cancer cells that triggers aberrant surface protein glycosylation patterns, adhesion to lectins on the surface of inflammatory cells, and subsequent metastatic colonization of the liver.