RESUMEN
Small blue round cell sarcomas (SBRCSs) are a heterogeneous group of tumors with overlapping morphologic features but markedly varying prognosis. They are characterized by distinct chromosomal alterations, particularly rearrangements leading to gene fusions, whose detection currently represents the most reliable diagnostic marker. Ewing sarcomas are the most common SBRCSs, defined by gene fusions involving EWSR1 and transcription factors of the ETS family, and the most frequent non-EWSR1-rearranged SBRCSs harbor a CIC rearrangement. Unfortunately, currently the identification of CIC::DUX4 translocation events, the most common CIC rearrangement, is challenging. Here, we present a machine-learning approach to support SBRCS diagnosis that relies on gene expression profiles measured via targeted sequencing. The analyses on a curated cohort of 69 soft-tissue tumors showed markedly distinct expression patterns for SBRCS subgroups. A random forest classifier trained on Ewing sarcoma and CIC-rearranged cases predicted probabilities of being CIC-rearranged >0.9 for CIC-rearranged-like sarcomas and <0.6 for other SBRCSs. Testing on a retrospective cohort of 1335 routine diagnostic cases identified 15 candidate CIC-rearranged tumors with a probability >0.75, all of which were supported by expert histopathologic reassessment. Furthermore, the multigene random forest classifier appeared advantageous over using high ETV4 expression alone, previously proposed as a surrogate to identify CIC rearrangement. Taken together, the expression-based classifier can offer valuable support for SBRCS pathologic diagnosis.
Asunto(s)
Sarcoma de Células Pequeñas , Sarcoma , Neoplasias de los Tejidos Blandos , Humanos , Estudios Retrospectivos , Sarcoma de Células Pequeñas/diagnóstico , Sarcoma de Células Pequeñas/genética , Sarcoma de Células Pequeñas/patología , Factores de Transcripción/genética , Sarcoma/genética , Neoplasias de los Tejidos Blandos/genética , Análisis de Secuencia de ARN , Proteínas de Fusión Oncogénica/genética , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/análisisRESUMEN
Trastuzumab emtansine (T-DM1) was the first and one of the most successful antibody-drug conjugates (ADC) approved for treating refractory HER2-positive breast cancer. Despite its initial clinical efficacy, resistance is unfortunately common, necessitating approaches to improve response. Here, we found that in sensitive cells, T-DM1 induced spindle assembly checkpoint (SAC)-dependent immunogenic cell death (ICD), an immune-priming form of cell death. The payload of T-DM1 mediated ICD by inducing eIF2α phosphorylation, surface exposure of calreticulin, ATP and HMGB1 release, and secretion of ICD-related cytokines, all of which were lost in resistance. Accordingly, ICD-related gene signatures in pretreatment samples correlated with clinical response to T-DM1-containing therapy, and increased infiltration of antitumor CD8+ T cells in posttreatment samples was correlated with better T-DM1 response. Transforming acidic coiled-coil containing 3 (TACC3) was overexpressed in T-DM1-resistant cells, and T-DM1 responsive patients had reduced TACC3 protein expression whereas nonresponders exhibited increased TACC3 expression during T-DM1 treatment. Notably, genetic or pharmacologic inhibition of TACC3 restored T-DM1-induced SAC activation and induction of ICD markers in vitro. Finally, TACC3 inhibition in vivo elicited ICD in a vaccination assay and potentiated the antitumor efficacy of T-DM1 by inducing dendritic cell maturation and enhancing intratumoral infiltration of cytotoxic T cells. Together, these results illustrate that ICD is a key mechanism of action of T-DM1 that is lost in resistance and that targeting TACC3 can restore T-DM1-mediated ICD and overcome resistance. SIGNIFICANCE: Loss of induction of immunogenic cell death in response to T-DM1 leads to resistance that can be overcome by targeting TACC3, providing an attractive strategy to improve the efficacy of T-DM1.
Asunto(s)
Ado-Trastuzumab Emtansina , Neoplasias de la Mama , Muerte Celular Inmunogénica , Proteínas Asociadas a Microtúbulos , Receptor ErbB-2 , Humanos , Femenino , Neoplasias de la Mama/inmunología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/genética , Muerte Celular Inmunogénica/efectos de los fármacos , Receptor ErbB-2/metabolismo , Ado-Trastuzumab Emtansina/farmacología , Ado-Trastuzumab Emtansina/uso terapéutico , Animales , Ratones , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/inmunología , Ensayos Antitumor por Modelo de Xenoinjerto , Línea Celular Tumoral , Antineoplásicos Inmunológicos/farmacología , Antineoplásicos Inmunológicos/uso terapéutico , Resistencia a Antineoplásicos/inmunología , Resistencia a Antineoplásicos/efectos de los fármacos , Antígenos de Neoplasias/inmunología , Antígenos de Neoplasias/genética , Trastuzumab/farmacología , Trastuzumab/uso terapéutico , Linfocitos T CD8-positivos/inmunologíaRESUMEN
GRB2 is an adaptor protein of HER2 (and several other tyrosine kinases), which we identified as a novel BECN1 (Beclin 1) interacting partner. GRB2 co-immunoprecipitated with BECN1 in several breast cancer cell lines and regulates autophagy through a mechanism involving the modulation of the class III PI3Kinase VPS34 activity. In ovo studies in a CAM (Chicken Chorioallantoic Membrane) model indicated that GRB2 knockdown, as well as overexpression of GRB2 loss-of-function mutants (Y52A and S86A-R88A) compromised tumor growth. These differences in tumor growth correlated with differential autophagy activity, indicating that autophagy effects might be related to the effects on tumorigenesis. Our data highlight a novel function of GRB2 as a BECN1 binding protein and a regulator of autophagy.
Asunto(s)
Autofagia , Beclina-1 , Proteína Adaptadora GRB2 , Animales , Proteínas Adaptadoras Transductoras de Señales , Beclina-1/metabolismo , Carcinogénesis , Transformación Celular Neoplásica , Humanos , Proteína Adaptadora GRB2/metabolismoRESUMEN
Intratumoral heterogeneity impacts the success or failure of anti-cancer therapies. Here, we investigated the evolution and mechanistic heterogeneity in clonal populations of cell models for estrogen receptor positive breast cancer. To this end, we established barcoded models of luminal breast cancer and rendered them resistant to commonly applied first line endocrine therapies. By isolating single clones from the resistant cell pools and characterizing replicates of individual clones we observed inter- (between cell lines) and intra-tumor (between different clones from the same cell line) heterogeneity. Molecular characterization at RNA and phospho-proteomic levels revealed private clonal activation of the unfolded protein response and respective sensitivity to inhibition of the proteasome, and potentially shared sensitivities for repression of protein kinase C. Our in vitro findings are consistent with tumor-heterogeneity that is observed in breast cancer patients thus highlighting the need to uncover heterogeneity at an individual patient level and to adjust therapies accordingly.
RESUMEN
Immunogenic cell death (ICD), an immune-priming form of cell death, has been shown to be induced by several different anti-cancer therapies. Despite being the first and one of the most successful antibody-drug conjugates (ADCs) approved for refractory HER2-positive breast cancer, little is known if response and resistance to trastuzumab emtansine (T-DM1) involves ICD modulation that can be leveraged to enhance T-DM1 response. Here, we report that T-DM1 induces spindle assembly checkpoint (SAC)-dependent ICD in sensitive cells by inducing eIF2α phosphorylation, surface exposure of calreticulin, ATP and HMGB1 release, and secretion of ICD-related cytokines, all of which are lost in resistance. Accordingly, an ICD-related gene signature correlates with clinical response to T-DM1-containing therapy. We found that transforming acidic coiled-coil containing 3 (TACC3) is overexpressed in T-DM1 resistant cells, and that T-DM1 responsive patients have reduced TACC3 protein while the non-responders exhibited increased TACC3 expression during T-DM1 treatment. Notably, genetic or pharmacological inhibition of TACC3 revives T-DM1-induced SAC activation and induction of ICD markers in vitro. Finally, TACC3 inhibition elicits ICD in vivo shown by vaccination assay, and it potentiates T-DM1 by inducing dendritic cell (DC) maturation and enhancing infiltration of cytotoxic T cells in the human HER2-overexpressing MMTV.f.huHER2#5 (Fo5) transgenic model. Together, our results show that ICD is a key mechanism of action of T-DM1 which is lost in resistance, and that targeting TACC3 restores T-DM1-mediated ICD and overcomes resistance.
RESUMEN
Anticancer therapies have been limited by the emergence of mutations and other adaptations. In bacteria, antibiotics activate the SOS response, which mobilizes error-prone factors that allow for continuous replication at the cost of mutagenesis. We investigated whether the treatment of lung cancer with EGFR inhibitors (EGFRi) similarly engages hypermutators. In cycling drug-tolerant persister (DTP) cells and in EGFRi-treated patients presenting residual disease, we observed upregulation of GAS6, whereas ablation of GAS6's receptor, AXL, eradicated resistance. Reciprocally, AXL overexpression enhanced DTP survival and accelerated the emergence of T790M, an EGFR mutation typical to resistant cells. Mechanistically, AXL induces low-fidelity DNA polymerases and activates their organizer, RAD18, by promoting neddylation. Metabolomics uncovered another hypermutator, AXL-driven activation of MYC, and increased purine synthesis that is unbalanced by pyrimidines. Aligning anti-AXL combination treatments with the transition from DTPs to resistant cells cured patient-derived xenografts. Hence, similar to bacteria, tumors tolerate therapy by engaging pharmacologically targetable endogenous mutators. SIGNIFICANCE: EGFR-mutant lung cancers treated with kinase inhibitors often evolve resistance due to secondary mutations. We report that in similarity to the bacterial SOS response stimulated by antibiotics, endogenous mutators are activated in drug-treated cells, and this heralds tolerance. Blocking the process prevented resistance in xenograft models, which offers new treatment strategies. This article is highlighted in the In This Issue feature, p. 2483.
Asunto(s)
Resistencia a Antineoplásicos , Neoplasias Pulmonares , Proteínas Proto-Oncogénicas , Proteínas Tirosina Quinasas Receptoras , Humanos , Línea Celular Tumoral , Replicación del ADN , Proteínas de Unión al ADN/genética , Resistencia a Antineoplásicos/genética , Receptores ErbB/genética , Neoplasias Pulmonares/genética , Mutación , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas/genética , Proteínas Tirosina Quinasas Receptoras/genética , Ubiquitina-Proteína Ligasas/genética , Animales , Tirosina Quinasa del Receptor AxlRESUMEN
BACKGROUND: MicroRNAs (miRNAs) and isomiRs play important roles in tumorigenesis as essential regulators of gene expression. 5'isomiRs exhibit a shifted seed sequence compared to the canonical miRNA, resulting in different target spectra and thereby extending the phenotypic impact of the respective common pre-miRNA. However, for most miRNAs, expression and function of 5'isomiRs have not been studied in detail yet. Therefore, this study aims to investigate the functions of miRNAs and their 5'isomiRs. METHODS: The expression of 5'isomiRs was assessed in The Cancer Genome Atlas (TCGA) breast cancer patient dataset. Phenotypic effects of miR-183 overexpression in triple-negative breast cancer (TNBC) cell lines were investigated in vitro and in vivo by quantifying migration, proliferation, tumor growth and metastasis. Direct targeting of E2F1 by miR-183-5p|+2 was validated with a 3'UTR luciferase assay and linked to the phenotypes of isomiR overexpression. RESULTS: TCGA breast cancer patient data indicated that three variants of miR-183-5p are highly expressed and upregulated, namely miR-183-5p|0, miR-183-5p|+1 and miR-183-5p|+2. However, TNBC cell lines displayed reduced proliferation and invasion upon overexpression of pre-miR-183. While invasion was reduced individually by all three isomiRs, proliferation and cell cycle progression were specifically inhibited by overexpression of miR-183-5p|+2. Proteomic analysis revealed reduced expression of E2F target genes upon overexpression of this isomiR, which could be attributed to direct targeting of E2F1, specifically by miR-183-5p|+2. Knockdown of E2F1 partially phenocopied the effect of miR-183-5p|+2 overexpression on cell proliferation and cell cycle. Gene set enrichment analysis of TCGA and METABRIC patient data indicated that the activity of E2F strongly correlated with the expression of miR-183-5p, suggesting transcriptional regulation of the miRNA by a factor of the E2F family. Indeed, in vitro, expression of miR-183-5p was regulated by E2F1. Hence, miR-183-5p|+2 directly targeting E2F1 appears to be part of a negative feedback loop potentially fine-tuning its activity. CONCLUSIONS: This study demonstrates that 5'isomiRs originating from the same arm of the same pre-miRNA (i.e. pre-miR-183-5p) may exhibit different functions and thereby collectively contribute to the same phenotype. Here, one of three isomiRs was shown to counteract expression of the pre-miRNA by negatively regulating a transcriptional activator (i.e. E2F1). We speculate that this might be part of a regulatory mechanism to prevent uncontrolled cell proliferation, which is disabled during cancer progression.
Asunto(s)
MicroARNs , Neoplasias de la Mama Triple Negativas , Línea Celular Tumoral , Factor de Transcripción E2F1/genética , Factor de Transcripción E2F1/metabolismo , Retroalimentación , Humanos , MicroARNs/metabolismo , Proteómica , Neoplasias de la Mama Triple Negativas/metabolismoRESUMEN
Targeted therapies have shown striking success in the treatment of cancer over the last years. However, their specific effects on an individual tumor appear to be varying and difficult to predict. Using an integrative modeling approach that combines mechanistic and regression modeling, we gained insights into the response mechanisms of breast cancer cells due to different ligand-drug combinations. The multi-pathway model, capturing ERBB receptor signaling as well as downstream MAPK and PI3K pathways was calibrated on time-resolved data of the luminal breast cancer cell lines MCF7 and T47D across an array of four ligands and five drugs. The same model was then successfully applied to triple negative and HER2-positive breast cancer cell lines, requiring adjustments mostly for the respective receptor compositions within these cell lines. The additional relevance of cell-line-specific mutations in the MAPK and PI3K pathway components was identified via L1 regularization, where the impact of these mutations on pathway activation was uncovered. Finally, we predicted and experimentally validated the proliferation response of cells to drug co-treatments. We developed a unified mathematical model that can describe the ERBB receptor and downstream signaling in response to therapeutic drugs targeting this clinically relevant signaling network in cell line that represent three major subtypes of breast cancer. Our data and model suggest that alterations in this network could render anti-HER therapies relevant beyond the HER2-positive subtype.
RESUMEN
Primary lymphomas of the central nervous system (PCNSL) are mainly diffuse large B-cell lymphomas (DLBCLs) confined to the central nervous system (CNS). Molecular drivers of PCNSL have not been fully elucidated. Here, we profile and compare the whole-genome and transcriptome landscape of 51 CNS lymphomas (CNSL) to 39 follicular lymphoma and 36 DLBCL cases outside the CNS. We find recurrent mutations in JAK-STAT, NFkB, and B-cell receptor signaling pathways, including hallmark mutations in MYD88 L265P (67%) and CD79B (63%), and CDKN2A deletions (83%). PCNSLs exhibit significantly more focal deletions of HLA-D (6p21) locus as a potential mechanism of immune evasion. Mutational signatures correlating with DNA replication and mitosis are significantly enriched in PCNSL. TERT gene expression is significantly higher in PCNSL compared to activated B-cell (ABC)-DLBCL. Transcriptome analysis clearly distinguishes PCNSL and systemic DLBCL into distinct molecular subtypes. Epstein-Barr virus (EBV)+ CNSL cases lack recurrent mutational hotspots apart from IG and HLA-DRB loci. We show that PCNSL can be clearly distinguished from DLBCL, having distinct expression profiles, IG expression and translocation patterns, as well as specific combinations of genetic alterations.
Asunto(s)
Neoplasias del Sistema Nervioso Central , Infecciones por Virus de Epstein-Barr , Linfoma de Células B Grandes Difuso , Sistema Nervioso Central/metabolismo , Neoplasias del Sistema Nervioso Central/genética , Neoplasias del Sistema Nervioso Central/patología , Genómica , Herpesvirus Humano 4 , Humanos , Linfoma de Células B Grandes Difuso/metabolismoRESUMEN
Seventy percent of patients with colorectal cancer develop liver metastases (CRLM), which are a decisive factor in cancer progression. Therapy outcome is largely influenced by tumor heterogeneity, but the intra- and inter-patient heterogeneity of CRLM has been poorly studied. In particular, the contribution of the WNT and EGFR pathways, which are both frequently deregulated in colorectal cancer, has not yet been addressed in this context. To this end, we comprehensively characterized normal liver tissue and eight CRLM from two patients by standardized histopathological, molecular, and proteomic subtyping. Suitable fresh-frozen tissue samples were profiled by transcriptome sequencing (RNA-Seq) and proteomic profiling with reverse phase protein arrays (RPPA) combined with bioinformatic analyses to assess tumor heterogeneity and identify WNT- and EGFR-related master regulators and metastatic effectors. A standardized data analysis pipeline for integrating RNA-Seq with clinical, proteomic, and genetic data was established. Dimensionality reduction of the transcriptome data revealed a distinct signature for CRLM differing from normal liver tissue and indicated a high degree of tumor heterogeneity. WNT and EGFR signaling were highly active in CRLM and the genes of both pathways were heterogeneously expressed between the two patients as well as between the synchronous metastases of a single patient. An analysis of the master regulators and metastatic effectors implicated in the regulation of these genes revealed a set of four genes (SFN, IGF2BP1, STAT1, PIK3CG) that were differentially expressed in CRLM and were associated with clinical outcome in a large cohort of colorectal cancer patients as well as CRLM samples. In conclusion, high-throughput profiling enabled us to define a CRLM-specific signature and revealed the genes of the WNT and EGFR pathways associated with inter- and intra-patient heterogeneity, which were validated as prognostic biomarkers in CRC primary tumors as well as liver metastases.
RESUMEN
Autophagy is an intracellular degradation process that maintains the cellular homeostasis and it is regulated in multiple ways, both in health and disease. Assessment of autophagic flux in cells is an important approach for understanding the function of autophagy in biological contexts. Here, we describe a new tool for the qualitative and quantitative determination of autophagic flux using a dual lentiviral reporter system that generates a fusion HiBiT-GFP-LC3B protein suitable for generating stable cell lines.
Asunto(s)
Autofagia , Proteínas Asociadas a Microtúbulos , Autofagia/genética , Línea Celular , Proteínas Asociadas a Microtúbulos/metabolismoRESUMEN
BACKGROUND: Triple negative breast cancer (TNBC) is the most aggressive subtype of breast cancer (BC). Treatment options for TNBC patients are limited and further insights into disease aetiology are needed to develop better therapeutic approaches. microRNAs' ability to regulate multiple targets could hold a promising discovery approach to pathways relevant for TNBC aggressiveness. Thus, we address the role of miRNAs in controlling three signalling pathways relevant to the biology of TNBC, and their downstream phenotypes. METHODS: To identify miRNAs regulating WNT/ß-catenin, c-Met, and integrin signalling pathways, we performed a high-throughput targeted proteomic approach, investigating the effect of 800 miRNAs on the expression of 62 proteins in the MDA-MB-231 TNBC cell line. We then developed a novel network analysis, Pathway Coregulatory (PC) score, to detect miRNAs regulating these three pathways. Using in vitro assays for cell growth, migration, apoptosis, and stem-cell content, we validated the function of candidate miRNAs. Bioinformatic analyses using BC patients' datasets were employed to assess expression of miRNAs as well as their pathological relevance in TNBC patients. RESULTS: We identified six candidate miRNAs coordinately regulating the three signalling pathways. Quantifying cell growth of three TNBC cell lines upon miRNA gain-of-function experiments, we characterised miR-193b as a strong and consistent repressor of proliferation. Importantly, the effects of miR-193b were stronger than chemical inhibition of the individual pathways. We further demonstrated that miR-193b induced apoptosis, repressed migration, and regulated stem-cell markers in MDA-MB-231 cells. Furthermore, miR-193b expression was the lowest in patients classified as TNBC or Basal compared to other subtypes. Gene Set Enrichment Analysis showed that miR-193b expression was significantly associated with reduced activity of WNT/ß-catenin and c-Met signalling pathways in TNBC patients. CONCLUSIONS: Integrating miRNA-mediated effects and protein functions on networks, we show that miRNAs predominantly act in a coordinated fashion to activate or repress connected signalling pathways responsible for metastatic traits in TNBC. We further demonstrate that our top candidate, miR-193b, regulates these phenotypes to an extent stronger than individual pathway inhibition, thus emphasizing that its effect on TNBC aggressiveness is mediated by the coordinated repression of these functionally interconnected pathways.
Asunto(s)
MicroARNs/metabolismo , Proteínas Proto-Oncogénicas c-met/metabolismo , Neoplasias de la Mama Triple Negativas/genética , Vía de Señalización Wnt/genética , beta Catenina/metabolismo , Línea Celular Tumoral , Proliferación Celular , Humanos , Metástasis de la Neoplasia , TransfecciónRESUMEN
BACKGROUND: Breast cancer has been associated with activation of the WNT signaling pathway, although no driver mutations in WNT genes have been found yet. Instead, a high expression of the alternative WNT receptor ROR2 was observed, in particular in breast cancer brain metastases. However, its respective ligand and downstream signaling in this context remained unknown. METHODS: We modulated the expression of ROR2 in human breast cancer cells and characterized their gene and protein expression by RNA-Seq, qRT-PCR, immunoblots and reverse phase protein array (RPPA) combined with network analyses to understand the molecular basis of ROR2 signaling in breast cancer. Using co-immunoprecipitations, we verified the interaction of ROR2 with the identified ligand, WNT11. The functional consequences of WNT11/ROR2 signaling for tumor cell aggressiveness were assessed by microscopy, impedance sensing as well as viability and invasion assays. To evaluate the translational significance of our findings, we performed gene set enrichment, expression and survival analyses on human breast cancer brain metastases. RESULTS: We found ROR2 to be highly expressed in aggressive breast tumors and associated with worse metastasis-free survival. ROR2 overexpression induced a BRCAness-like phenotype in a cell-context specific manner and rendered cells resistant to PARP inhibition. High levels of ROR2 were furthermore associated with defects in cell morphology and cell-cell-contacts leading to increased tumor invasiveness. On a molecular level, ROR2 overexpression upregulated several non-canonical WNT ligands, in particular WNT11. Co-immunoprecipitation confirmed that WNT11 indeed interacts with the cysteine-rich domain of ROR2 and triggers its invasion-promoting signaling via RHO/ROCK. Knockdown of WNT11 reversed the pro-invasive phenotype and the cellular changes in ROR2-overexpressing cells. CONCLUSIONS: Taken together, our study revealed a novel auto-stimulatory loop in which ROR2 triggers the expression of its own ligand, WNT11, resulting in enhanced tumor invasion associated with breast cancer metastasis.
Asunto(s)
Neoplasias Encefálicas/genética , Vía de Señalización Wnt/genética , Neoplasias Encefálicas/mortalidad , Humanos , Invasividad Neoplásica , Metástasis de la Neoplasia , Análisis de Supervivencia , TransfecciónRESUMEN
Tumour cells do not exist as an isolated entity. Instead, they are surrounded by and closely interact with cells of the environment they are emerged in. The tumour microenvironment (TME) is not static and several factors, including cancer cells and therapies, have been described to modulate several of its components. Fibroblasts are key elements of the TME with the capacity to influence tumour progression, invasion and response to therapy, which makes them attractive targets in cancer treatment. In this review, we focus on fibroblasts and their numerous roles in the TME with a special attention to recent findings describing their heterogeneity and role in therapy response. Furthermore, we explore how different therapies can impact these cells and their communication with cancer cells. Finally, we highlight potential strategies targeting this cell type that can be employed for improving patient outcome.
RESUMEN
Lung cancers driven by mutant forms of EGFR invariably develop resistance to kinase inhibitors, often due to secondary mutations. Here we describe an unconventional mechanism of resistance to dacomitinib, a newly approved covalent EGFR kinase inhibitor, and uncover a previously unknown step of resistance acquisition. Dacomitinib-resistant (DR) derivatives of lung cancer cells were established by means of gradually increasing dacomitinib concentrations. These DR cells acquired no secondary mutations in the kinase or other domains of EGFR. Along with resistance to other EGFR inhibitors, DR cells acquired features characteristic to epithelial-mesenchymal transition, including an expanded population of aldehyde dehydrogenase-positive cells and upregulation of AXL, a receptor previously implicated in drug resistance. Unexpectedly, when implanted in animals, DR cells reverted to a dacomitinib-sensitive state. Nevertheless, cell lines derived from regressing tumors displayed renewed resistance when cultured in vitro. Three-dimensional and cocultures along with additional analyses indicated lack of involvement of hypoxia, fibroblasts, and immune cells in phenotype reversal, implying that other host-dependent mechanisms might nullify nonmutational modes of resistance. Thus, similar to the phenotypic resistance of bacteria treated with antibiotics, the reversible resisters described here likely evolve from drug-tolerant persisters and give rise to the irreversible, secondary mutation-driven nonreversible resister state. SIGNIFICANCE: This study reports that stepwise acquisition of kinase inhibitor resistance in lung cancers driven by mutant EGFR comprises a nonmutational, reversible resister state. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/14/3862/F1.large.jpg.
Asunto(s)
Inhibidores de Proteínas Quinasas/uso terapéutico , Animales , Femenino , Humanos , Ratones , Ratones Desnudos , Fenotipo , Inhibidores de Proteínas Quinasas/farmacologíaRESUMEN
Solitary fibrous tumors (SFTs) harbor activating NAB2-STAT6 gene fusions. Different variants of the NAB2-STAT6 gene fusion have been associated with distinct clinicopathologic features. Lipomatous SFTs are a morphologic variant of SFTs, characterized by a fat-forming tumor component. Our aim was to evaluate NAB2-STAT6 fusion variants and to further study the molecular genetic features in a cohort of lipomatous SFTs. A hybrid-capture-based next-generation sequencing panel was employed to detect NAB2-STAT6 gene fusions at the RNA level. In addition, the RNA expression levels of 507 genes were evaluated using this panel, and were compared with a control cohort of nonlipomatous SFTs. Notably, 5 of 11 (45%) of lipomatous SFTs in the current series harbored the uncommon NAB2 exon 4-STAT6 exon 4 gene fusion variant, which is observed in only 0.9% to 1.4% of nonlipomatous SFTs. Furthermore, lipomatous SFTs displayed significant differences in gene expression compared with their nonlipomatous counterparts, including up-regulation of the gene peroxisome proliferator activated receptor-γ (PPARG). Peroxisome proliferator activated receptor-γ is a nuclear receptor regulating adipocyte differentiation, providing a possible explanation for the fat-forming component in lipomatous SFTs. In summary, the current study provides a possible molecular genetic basis for the distinct morphologic features of lipomatous SFTs.
Asunto(s)
Adipocitos/patología , PPAR gamma/genética , Proteínas Represoras/genética , Factor de Transcripción STAT6/genética , Tumores Fibrosos Solitarios/genética , Adulto , Anciano , Diferenciación Celular/genética , Femenino , Humanos , Masculino , Persona de Mediana Edad , Fusión de Oncogenes , Tumores Fibrosos Solitarios/patología , Regulación hacia ArribaRESUMEN
HER3 is highly expressed in luminal breast cancer subtypes. Its activation by NRG1 promotes activation of AKT and ERK1/2, contributing to tumour progression and therapy resistance. HER3-targeting agents that block this activation, are currently under phase 1/2 clinical studies, and although they have shown favorable tolerability, their activity as a single agent has proven to be limited. Here we show that phosphorylation and activation of HER3 in luminal breast cancer cells occurs in a paracrine manner and is mediated by NRG1 expressed by cancer-associated fibroblasts (CAFs). Moreover, we uncover a HER3-independent NRG1 signaling in CAFs that results in the induction of a strong migratory and pro-fibrotic phenotype, describing a subtype of CAFs with elevated expression of NRG1 and an associated transcriptomic profile that determines their functional properties. Finally, we identified Hyaluronan Synthase 2 (HAS2), a targetable molecule strongly correlated with NRG1, as an attractive player supporting NRG1 signaling in CAFs.
Asunto(s)
Neoplasias de la Mama/genética , Fibroblastos Asociados al Cáncer/metabolismo , Neurregulina-1/metabolismo , Proteómica/métodos , Femenino , Humanos , Microambiente TumoralRESUMEN
Solitary fibrous tumors (SFTs) harbor recurrent NAB2-STAT6 gene fusions, promoting constitutional up-regulation of oncogenic early growth response 1 (EGR1)-dependent gene expression. SFTs with the most common canonical NAB2 exon 4-STAT6 exon 2 fusion variant are often located in the thorax (pleuropulmonary) and are less cellular with abundant collagen. In contrast, SFTs with NAB2 exon 6-STAT6 exon 16/17 fusion variants typically display a cellular round to ovoid cell morphology and are often located in the deep soft tissue of the retroperitoneum and intra-abdominal pelvic region or in the meninges. Here, we employed next-generation sequencing-based gene expression profiling to identify significant differences in gene expression associated with anatomic localization and NAB2-STAT6 gene fusion variants. SFTs with the NAB2 exon 4-STAT6 exon 2 fusion variant showed a transcriptional signature enriched for genes involved in DNA binding, gene transcription, and nuclear localization, whereas SFTs with the NAB2 exon 6-STAT6 exon 16/17 fusion variants were enriched for genes involved in tyrosine kinase signaling, cell proliferation, and cytoplasmic localization. Specific transcription factor binding motifs were enriched among differentially expressed genes in SFTs with different fusion variants, implicating co-transcription factors in the modification of chimeric NGFI-A binding protein 2 (NAB2)-STAT6-dependent deregulation of EGR1-dependent gene expression. In summary, this study establishes a potential molecular biologic basis for clinicopathologic differences in SFTs with distinct NAB2-STAT6 gene fusion variants.
Asunto(s)
Biomarcadores de Tumor/genética , Proteínas Represoras/genética , Factor de Transcripción STAT6/genética , Tumores Fibrosos Solitarios/genética , Exones/genética , Femenino , Expresión Génica/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Masculino , Proteínas de Fusión Oncogénica/genética , Proteínas Represoras/metabolismo , Tumores Fibrosos Solitarios/patologíaRESUMEN
Chemotherapy (CTX) remains the standard of care for most aggressive tumours, including breast cancer (BC). In BC chemotherapeutic regimens, the maximum tolerated dose of cytotoxic drugs is administered at regular intervals, and cancer cells can re-grow or adapt during the resting periods between cycles. The impact of the tumour microenvironment on the fate of cancer cells after CTX remains poorly understood. Here, we show that paracrine signalling from CTX-treated cancer cells to stromal fibroblasts can drive cancer cell recovery after cytotoxic drug withdrawal. Interferon ß1 (IFNß1) secreted by cancer cells following treatment with high doses of CTX instigates the acquisition of an anti-viral state in stromal fibroblasts. This state is associated with an expression pattern here referred to as interferon signature (IFNS), which encompasses several interferon-stimulated genes (ISGs), including numerous pro-inflammatory cytokine genes. This crosstalk is an important driver of the expansion of BC cells after CTX, and IFNß1 blockade in tumour cells abrogated their fibroblast-dependent recovery potential. Analysis of human breast carcinomas supported a link between CTX-induced IFNS in tumour stroma and poor response to CTX treatment. First, IFNß1 expression in human breast carcinomas was found to inversely correlate with recurrence free survival (RFS). Second, using laser capture microdissection data sets, we show a higher expression of IFNS in the stromal tumour compartment compared to the epithelial one and this signature was found to be more prominent in more aggressive subtypes of BC (basal-like), pointing to a pro-tumorigenic role of this signature. Moreover, IFNS was associated with higher recurrence rates and a worse outcome in BC patients. Our study unravels a novel form of paracrine communication between cancer cells and fibroblasts that ultimately results in CTX resistance. Targeting this axis has the potential to improve CTX outcomes in patients with BC.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Fibroblastos/efectos de los fármacos , Interferón beta/metabolismo , Interferón beta/farmacología , Recurrencia Local de Neoplasia/etiología , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Técnicas de Cocultivo , Femenino , Fibroblastos/metabolismo , Fibroblastos/patología , Células HEK293 , Humanos , Células MCF-7 , Recurrencia Local de Neoplasia/inducido químicamente , Recurrencia Local de Neoplasia/metabolismo , Comunicación Paracrina/efectos de los fármacos , Comunicación Paracrina/fisiología , Células del Estroma/efectos de los fármacos , Células del Estroma/metabolismo , Células del Estroma/patología , Microambiente Tumoral/efectos de los fármacosRESUMEN
DNA sequencing and RNA sequencing are increasingly applied in precision oncology, where molecular tumor boards evaluate the actionability of genetic events in individual tumors to guide targeted treatment. To work toward an additional level of patient characterization, we assessed the abundance and activity of 27 proteins in 134 patients whose tumors had previously undergone whole-exome and RNA sequencing within the Molecularly Aided Stratification for Tumor Eradication Research (MASTER) program of National Center for Tumor Diseases, Heidelberg. Proteomic and phosphoproteomic targets were selected to reflect the most relevant therapeutic baskets in MASTER. Among six different therapeutic baskets, the proteomic data supported treatment recommendations that were based on DNA and RNA analyses in 10% to 57% and frequently suggested alternative treatment options. In several cases, protein activities explained the patients' clinical course and provided potential explanations for treatment failure. Our study indicates that the integrative analysis of DNA, RNA and protein data may refine therapeutic stratification of individual patients and, thus, holds potential to increase the success rate of precision cancer therapy. Prospective validation studies are needed to advance the integration of proteomic analysis into precision oncology.