RESUMEN
Biliary tract cancers (BTCs) are a group of deadly malignancies encompassing intrahepatic and extrahepatic cholangiocarcinoma, gallbladder carcinoma, and ampullary carcinoma. Here, we present the integrative analysis of 63 BTC cell lines via multi-omics clustering and genome- scale CRISPR screens, providing a platform to illuminate BTC biology and inform therapeutic development. We identify dependencies broadly enriched in BTC compared to other cancers as well as dependencies selective to the anatomic subtypes. Notably, cholangiocarcinoma cell lines are stratified into distinct lineage subtypes based on biliary or dual biliary/hepatocyte marker signatures, associated with dependency on specific lineage survival factors. Transcriptional analysis of patient specimens demonstrates the prognostic significance of these lineage subtypes. Additionally, we delineate strategies to enhance targeted therapies or to overcome resistance in cell lines with key driver gene mutations. Furthermore, clustering based on dependencies and proteomics data elucidates unexpected functional relationships, including a BTC subgroup with partial squamous differentiation. Thus, this cell line atlas reveals potential therapeutic targets in molecularly defined BTCs, unveils biologically distinct disease subtypes, and offers a vital resource for BTC research.
RESUMEN
[This corrects the article DOI: 10.1371/journal.pone.0140310.].
RESUMEN
Tailoring optimal treatment for individual cancer patients remains a significant challenge. To address this issue, we developed PERCEPTION (PERsonalized Single-Cell Expression-Based Planning for Treatments In ONcology), a precision oncology computational pipeline. Our approach uses publicly available matched bulk and single-cell (sc) expression profiles from large-scale cell-line drug screens. These profiles help build treatment response models based on patients' sc-tumor transcriptomics. PERCEPTION demonstrates success in predicting responses to targeted therapies in cultured and patient-tumor-derived primary cells, as well as in two clinical trials for multiple myeloma and breast cancer. It also captures the resistance development in patients with lung cancer treated with tyrosine kinase inhibitors. PERCEPTION outperforms published state-of-the-art sc-based and bulk-based predictors in all clinical cohorts. PERCEPTION is accessible at https://github.com/ruppinlab/PERCEPTION . Our work, showcasing patient stratification using sc-expression profiles of their tumors, will encourage the adoption of sc-omics profiling in clinical settings, enhancing precision oncology tools based on sc-omics.
Asunto(s)
Resistencia a Antineoplásicos , Medicina de Precisión , Análisis de la Célula Individual , Transcriptoma , Humanos , Análisis de la Célula Individual/métodos , Medicina de Precisión/métodos , Resistencia a Antineoplásicos/genética , Neoplasias/genética , Neoplasias/tratamiento farmacológico , Perfilación de la Expresión Génica/métodos , Femenino , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/tratamiento farmacológico , Regulación Neoplásica de la Expresión Génica , Línea Celular Tumoral , Biología Computacional/métodosRESUMEN
Combination of anti-cancer drugs is broadly seen as way to overcome the often-limited efficacy of single agents. The design and testing of combinations are however very challenging. Here we present a uniquely large dataset screening over 5000 targeted agent combinations across 81 non-small cell lung cancer cell lines. Our analysis reveals a profound heterogeneity of response across the tumor models. Notably, combinations very rarely result in a strong gain in efficacy over the range of response observable with single agents. Importantly, gain of activity over single agents is more often seen when co-targeting functionally proximal genes, offering a strategy for designing more efficient combinations. Because combinatorial effect is strongly context specific, tumor specificity should be achievable. The resource provided, together with an additional validation screen sheds light on major challenges and opportunities in building efficacious combinations against cancer and provides an opportunity for training computational models for synergy prediction.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Combinación de MedicamentosRESUMEN
Adult liver malignancies, including intrahepatic cholangiocarcinoma and hepatocellular carcinoma, are the second leading cause of cancer-related deaths worldwide. Most individuals are treated with either combination chemotherapy or immunotherapy, respectively, without specific biomarkers for selection. Here using high-throughput screens, proteomics and in vitro resistance models, we identify the small molecule YC-1 as selectively active against a defined subset of cell lines derived from both liver cancer types. We demonstrate that selectivity is determined by expression of the liver-resident cytosolic sulfotransferase enzyme SULT1A1, which sulfonates YC-1. Sulfonation stimulates covalent binding of YC-1 to lysine residues in protein targets, enriching for RNA-binding factors. Computational analysis defined a wider group of structurally related SULT1A1-activated small molecules with distinct target profiles, which together constitute an untapped small-molecule class. These studies provide a foundation for preclinical development of these agents and point to the broader potential of exploiting SULT1A1 activity for selective targeting strategies.
Asunto(s)
Alquilantes , Neoplasias Hepáticas , Humanos , Sulfotransferasas , Neoplasias Hepáticas/tratamiento farmacológico , ArilsulfotransferasaRESUMEN
Preclinical and clinical studies have evidenced that effective targeted therapy treatment against receptor tyrosine kinases (RTKs) in different solid tumor paradigms is predicated on simultaneous inhibition of both the PI3K and MEK intracellular signaling pathways. Indeed, re-activation of either pathway results in resistance to these therapies. Recently, oncogenic phosphatase SHP2 inhibitors have been developed with some now reaching clinical trials. To expand on possible indications for SHP099, we screened over 800 cancer cell lines covering over 25 subsets of cancer. We found HNSCC was the most sensitive adult subtype of cancer to SHP099. We found that, in addition to the MEK pathway, SHP2 inhibition blocks the PI3K pathway in sensitive HNSCC, resulting in downregulation of mTORC signaling and anti-tumor effects across several HNSCC mouse models, including an HPV+ patient-derived xenograft (PDX). Importantly, we found low levels of the RTK ligand epiregulin identified HNSCCs that were sensitive to SHP2 inhibitor, and, adding exogenous epiregulin mitigated SHP099 efficacy. Mechanistically, epiregulin maintained SHP2-GAB1 complexes in the presence of SHP2 inhibition, preventing downregulation of the MEK and PI3K pathways. We demonstrate HNSCCs were highly dependent on GAB1 for their survival and knockdown of GAB1 is sufficient to block the ability of epiregulin to rescue MEK and PI3K signaling. These data connect the sensitivity of HNSCC to SHP2 inhibitors and to a broad reliance on GAB1-SHP2, revealing an important and druggable signaling axis. Overall, SHP2 inhibitors are being heavily developed and may have activity in HNSCCs, and in particular those with low levels of epiregulin.
Asunto(s)
Neoplasias de Cabeza y Cuello , Fosfatidilinositol 3-Quinasas , Animales , Ratones , Humanos , Fosfatidilinositol 3-Quinasas/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/tratamiento farmacológico , Epirregulina/metabolismo , Inhibidores Enzimáticos/farmacología , Neoplasias de Cabeza y Cuello/tratamiento farmacológico , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proteínas Adaptadoras Transductoras de Señales/metabolismoRESUMEN
The search for cell-permeable drugs has conventionally focused on low-molecular weight (MW), nonpolar, rigid chemical structures. However, emerging therapeutic strategies break traditional drug design rules by employing flexibly linked chemical entities composed of more than one ligand. Using complementary genome-scale chemical-genetic approaches we identified an endogenous chemical uptake pathway involving interferon-induced transmembrane proteins (IFITMs) that modulates the cell permeability of a prototypical biopic inhibitor of MTOR (RapaLink-1, MW: 1784 g/mol). We devised additional linked inhibitors targeting BCR-ABL1 (DasatiLink-1, MW: 1518 g/mol) and EIF4A1 (BisRoc-1, MW: 1466 g/mol), uptake of which was facilitated by IFITMs. We also found that IFITMs moderately assisted some proteolysis-targeting chimeras and examined the physicochemical requirements for involvement of this uptake pathway.
RESUMEN
Targeting TEAD autopalmitoylation has been proposed as a therapeutic approach for YAP-dependent cancers. Here we show that TEAD palmitoylation inhibitor MGH-CP1 and analogues block cancer cell "stemness", organ overgrowth and tumor initiation in vitro and in vivo. MGH-CP1 sensitivity correlates significantly with YAP-dependency in a large panel of cancer cell lines. However, TEAD inhibition or YAP/TAZ knockdown leads to transient inhibition of cell cycle progression without inducing cell death, undermining their potential therapeutic utilities. We further reveal that TEAD inhibition or YAP/TAZ silencing leads to VGLL3-mediated transcriptional activation of SOX4/PI3K/AKT signaling axis, which contributes to cancer cell survival and confers therapeutic resistance to TEAD inhibitors. Consistently, combination of TEAD and AKT inhibitors exhibits strong synergy in inducing cancer cell death. Our work characterizes the therapeutic opportunities and limitations of TEAD palmitoylation inhibitors in cancers, and uncovers an intrinsic molecular mechanism, which confers potential therapeutic resistance.
Asunto(s)
Neoplasias , Fosfatidilinositol 3-Quinasas , Humanos , Lipoilación , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal , Factores de Transcripción SOXC/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Factores de Transcripción de Dominio TEA/metabolismoRESUMEN
Reoccurring/high-risk neuroblastoma (NB) tumors have the enrichment of non-RAS/RAF mutations along the mitogen-activated protein kinase (MAPK) signaling pathway, suggesting that activation of MEK/ERK is critical for their survival. However, based on preclinical data, MEK inhibitors are unlikely to be active in NB and have demonstrated dose-limiting toxicities that limit their use. Here, we explore an alternative way to target the MAPK pathway in high-risk NB. We find that NB models are among the most sensitive among over 900 tumor-derived cell lines to the allosteric SHP2 inhibitor SHP099. Sensitivity to SHP099 in NB is greater in models with loss or low expression of the RAS GTPase activation protein (GAP) neurofibromin 1 (NF1). Furthermore, NF1 is lower in advanced and relapsed NB and NF1 loss is enriched in high-risk NB tumors regardless of MYCN status. SHP2 inhibition consistently blocks tumor growth in high-risk NB mouse models, revealing a new drug target in relapsed NB.
Asunto(s)
Neuroblastoma , Neurofibromina 1 , Proteína Tirosina Fosfatasa no Receptora Tipo 11/metabolismo , Animales , Línea Celular Tumoral , Ratones , Quinasas de Proteína Quinasa Activadas por Mitógenos , Proteínas Quinasas Activadas por Mitógenos , Recurrencia Local de Neoplasia , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/genética , Neuroblastoma/patología , Neurofibromina 1/genética , Neurofibromina 1/metabolismo , Inhibidores de Proteínas Quinasas/farmacologíaRESUMEN
FGFR inhibitors are approved for the treatment of advanced cholangiocarcinoma harboring FGFR2 fusions. However, the response rate is moderate, and resistance emerges rapidly due to acquired secondary FGFR2 mutations or due to other less-defined mechanisms. Here, we conducted high-throughput combination drug screens, biochemical analysis, and therapeutic studies using patient-derived models of FGFR2 fusion-positive cholangiocarcinoma to gain insight into these clinical profiles and uncover improved treatment strategies. We found that feedback activation of EGFR signaling limits FGFR inhibitor efficacy, restricting cell death induction in sensitive models and causing resistance in insensitive models lacking secondary FGFR2 mutations. Inhibition of wild-type EGFR potentiated responses to FGFR inhibitors in both contexts, durably suppressing MEK/ERK and mTOR signaling, increasing apoptosis, and causing marked tumor regressions in vivo. Our findings reveal EGFR-dependent adaptive signaling as an important mechanism limiting FGFR inhibitor efficacy and driving resistance and support clinical testing of FGFR/EGFR inhibitor therapy for FGFR2 fusion-positive cholangiocarcinoma. SIGNIFICANCE: We demonstrate that feedback activation of EGFR signaling limits the effectiveness of FGFR inhibitor therapy and drives adaptive resistance in patient-derived models of FGFR2 fusion-positive cholangiocarcinoma. These studies support the potential of combination treatment with FGFR and EGFR inhibitors as an improved treatment for patients with FGFR2-driven cholangiocarcinoma. This article is highlighted in the In This Issue feature, p. 1171.
Asunto(s)
Neoplasias de los Conductos Biliares , Colangiocarcinoma , Neoplasias de los Conductos Biliares/tratamiento farmacológico , Neoplasias de los Conductos Biliares/genética , Neoplasias de los Conductos Biliares/metabolismo , Conductos Biliares Intrahepáticos/metabolismo , Conductos Biliares Intrahepáticos/patología , Colangiocarcinoma/tratamiento farmacológico , Colangiocarcinoma/genética , Colangiocarcinoma/metabolismo , Receptores ErbB/genética , Humanos , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Pirimidinas/farmacología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismoRESUMEN
Combinations of anti-cancer drugs can overcome resistance and provide new treatments1,2. The number of possible drug combinations vastly exceeds what could be tested clinically. Efforts to systematically identify active combinations and the tissues and molecular contexts in which they are most effective could accelerate the development of combination treatments. Here we evaluate the potency and efficacy of 2,025 clinically relevant two-drug combinations, generating a dataset encompassing 125 molecularly characterized breast, colorectal and pancreatic cancer cell lines. We show that synergy between drugs is rare and highly context-dependent, and that combinations of targeted agents are most likely to be synergistic. We incorporate multi-omic molecular features to identify combination biomarkers and specify synergistic drug combinations and their active contexts, including in basal-like breast cancer, and microsatellite-stable or KRAS-mutant colon cancer. Our results show that irinotecan and CHEK1 inhibition have synergistic effects in microsatellite-stable or KRAS-TP53 double-mutant colon cancer cells, leading to apoptosis and suppression of tumour xenograft growth. This study identifies clinically relevant effective drug combinations in distinct molecular subpopulations and is a resource to guide rational efforts to develop combinatorial drug treatments.
Asunto(s)
Antineoplásicos , Neoplasias del Colon , Neoplasias Pancreáticas , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Línea Celular Tumoral , Proliferación Celular , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/genética , Combinación de Medicamentos , Sinergismo Farmacológico , Humanos , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogénicas p21(ras)/genéticaRESUMEN
Triple negative breast cancer (TNBC) accounts for over 30% of all breast cancer (BC)-related deaths, despite accounting for only 10% to 15% of total BC cases. Targeted therapy development has largely stalled in TNBC, underlined by a lack of traditionally druggable addictions like receptor tyrosine kinases (RTKs). Here, through full genome CRISPR/Cas9 screening of TNBC models, we have uncovered the sensitivity of TNBCs to the depletion of the ubiquitin-like modifier activating enzyme 1 (UBA1). Targeting UBA1 with the first-in-class UBA1 inhibitor TAK-243 induced unresolvable endoplasmic reticulum (ER)-stress and activating transcription factor 4 (ATF4)-mediated upregulation of proapoptotic NOXA, leading to cell death. c-MYC expression correlates with TAK-243 sensitivity and cooperates with TAK-243 to induce a stress response and cell death. Importantly, there was an order of magnitude greater sensitivity of TNBC lines to TAK-243 compared to normal tissue-derived cells. In five patient derived xenograft models (PDXs) of TNBC, TAK-243 therapy led to tumor inhibition or frank tumor regression. Moreover, in an intracardiac metastatic model of TNBC, TAK-243 markedly reduced metastatic burden, indicating UBA1 is a potential new target in TNBC expressing high levels of c-MYC.
RESUMEN
Cancer-associated fibroblasts (CAFs) are highly heterogeneous. With the lack of a comprehensive understanding of CAFs' functional distinctions, it remains unclear how cancer treatments could be personalized based on CAFs in a patient's tumor. We have established a living biobank of CAFs derived from biopsies of patients' non-small lung cancer (NSCLC) that encompasses a broad molecular spectrum of CAFs in clinical NSCLC. By functionally interrogating CAF heterogeneity using the same therapeutics received by patients, we identify three functional subtypes: (1) robustly protective of cancers and highly expressing HGF and FGF7; (2) moderately protective of cancers and highly expressing FGF7; and (3) those providing minimal protection. These functional differences among CAFs are governed by their intrinsic TGF-ß signaling, which suppresses HGF and FGF7 expression. This CAF functional classification correlates with patients' clinical response to targeted therapies and also associates with the tumor immune microenvironment, therefore providing an avenue to guide personalized treatment.
Asunto(s)
Fibroblastos Asociados al Cáncer/patología , Carcinoma de Pulmón de Células no Pequeñas/patología , Factor 7 de Crecimiento de Fibroblastos/genética , Factor de Crecimiento de Hepatocito/genética , Neoplasias Pulmonares/patología , Biopsia , Fibroblastos Asociados al Cáncer/química , Carcinoma de Pulmón de Células no Pequeñas/genética , Resistencia a Antineoplásicos , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Pulmonares/genética , Medicina de Precisión , Transducción de Señal , Factor de Crecimiento Transformador beta/metabolismo , Microambiente Tumoral , Regulación hacia ArribaRESUMEN
The development of molecular targeted drugs with radiation and chemotherapy is critically important for improving the outcomes of patients with hard-to-treat, potentially curable cancers. However, too many preclinical studies have not translated into successful radiation oncology trials. Major contributing factors to this insufficiency include poor reproducibility of preclinical data, inadequate preclinical modeling of intertumoral genomic heterogeneity that influences treatment sensitivity in the clinic, and a reliance on tumor growth delay instead of local control (TCD50) endpoints. There exists an urgent need to overcome these barriers to facilitate successful clinical translation of targeted radiosensitizers. To this end, we have used 3-dimensional (3D) cell culture assays to better model tumor behavior in vivo. Examples of successful prediction of in vivo effects with these 3D assays include radiosensitization of head and neck cancers by inhibiting epidermal growth factor receptor or focal adhesion kinase signaling, and radioresistance associated with oncogenic mutation of KRAS. To address the issue of tumor heterogeneity, we leveraged institutional resources that allow high-throughput 3D screening of radiation combinations with small-molecule inhibitors across genomically characterized cell lines from lung, head and neck, and pancreatic cancers. This high-throughput screen is expected to uncover genomic biomarkers that will inform the successful clinical translation of targeted agents from the National Cancer Institute Cancer Therapy Evaluation Program portfolio and other sources. Screening "hits" need to be subjected to refinement studies that include clonogenic assays, addition of disease-specific chemotherapeutics, target/biomarker validation, and integration of patient-derived tumor models. The chemoradiosensitizing activities of the most promising drugs should be confirmed in TCD50 assays in xenograft models with or without relevant biomarker and using clinically relevant radiation fractionation. We predict that appropriately validated and biomarker-directed targeted therapies will have a higher likelihood than past efforts of being successfully incorporated into the standard management of hard-to-treat tumors.
Asunto(s)
Terapia Molecular Dirigida , Biomarcadores de Tumor , Humanos , Neoplasias , Preparaciones Farmacéuticas , Fármacos Sensibilizantes a Radiaciones/uso terapéutico , Reproducibilidad de los ResultadosRESUMEN
The EWSR1-FLI1 t(11;22)(q24;q12) translocation is the hallmark genomic alteration of Ewing sarcoma, a malignancy of the bone and surrounding tissue, predominantly affecting children and adolescents. Although significant progress has been made for the treatment of localized disease, patients with metastasis or who relapse after chemotherapy have less than a 30% five-year survival rate. EWS-FLI1 is currently not clinically druggable, driving the need for more effective targeted therapies. Treatment with the H3K27 demethylase inhibitor, GSK-J4, leads to an increase in H3K27me and a decrease in H3K27ac, a significant event in Ewing sarcoma because H3K27ac associates strongly with EWS-FLI1 binding at enhancers and promoters and subsequent activity of EWS-FLI1 target genes. We were able to identify targets of EWS-FLI1 tumorigenesis directly inhibited by GSK-J4. GSK-J4 disruption of EWS-FLI1-driven transcription was toxic to Ewing sarcoma cells and slowed tumor growth in patient-derived xenografts (PDX) of Ewing sarcoma. Responses were markedly exacerbated by cotreatment with a disruptor of RNA polymerase II activity, the CDK7 inhibitor THZ1. This combination together suppressed EWS-FLI1 target genes and viability of ex vivo PDX Ewing sarcoma cells in a synergistic manner. In PDX models of Ewing Sarcoma, the combination shrank tumors. We present a new therapeutic strategy to treat Ewing sarcoma by decreasing H3K27ac at EWS-FLI1-driven transcripts, exacerbated by blocking phosphorylation of the C-terminal domain of RNA polymerase II to further hinder the EWS-FLI1-driven transcriptome.
Asunto(s)
Benzazepinas/farmacología , ARN Polimerasas Dirigidas por ADN/antagonistas & inhibidores , Histonas/antagonistas & inhibidores , Proteínas de Fusión Oncogénica/antagonistas & inhibidores , Fenilendiaminas/farmacología , Proteína Proto-Oncogénica c-fli-1/antagonistas & inhibidores , Pirimidinas/farmacología , Proteína EWS de Unión a ARN/antagonistas & inhibidores , Sarcoma de Ewing/tratamiento farmacológico , Transcriptoma , Animales , Apoptosis , Neoplasias Óseas/tratamiento farmacológico , Neoplasias Óseas/genética , Neoplasias Óseas/patología , Proliferación Celular , Regulación Neoplásica de la Expresión Génica , Humanos , Ratones , Ratones Endogámicos NOD , Ratones SCID , Sarcoma de Ewing/genética , Sarcoma de Ewing/patología , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Venetoclax is a small molecule inhibitor of the prosurvival protein BCL-2 that has gained market approval in BCL-2-dependent hematologic cancers including chronic lymphocytic leukemia and acute myeloid leukemia. Neuroblastoma is a heterogenous pediatric cancer with a 5-year survival rate of less than 50% for high-risk patients, which includes nearly all cases with amplified MYCN We previously demonstrated that venetoclax is active in MYCN-amplified neuroblastoma but has limited single-agent activity in most models, presumably the result of other pro-survival BCL-2 family protein expression or insufficient prodeath protein mobilization. As the relative tolerability of venetoclax makes it amenable to combining with other therapies, we evaluated the sensitivity of MYCN-amplified neuroblastoma models to rational combinations of venetoclax with agents that have both mechanistic complementarity and active clinical programs. First, the MDM2 inhibitor NVP-CGM097 increases the prodeath BH3-only protein NOXA to sensitize p53-wild-type, MYCN-amplified neuroblastomas to venetoclax. Second, the MCL-1 inhibitor S63845 sensitizes MYCN-amplified neuroblastoma through neutralization of MCL-1, inducing synergistic cell killing when combined with venetoclax. Finally, the standard-of-care drug cocktail cyclophosphamide and topotecan reduces the apoptotic threshold of neuroblastoma, thus setting the stage for robust combination efficacy with venetoclax. In all cases, these rational combinations translated to in vivo tumor regressions in MYCN-amplified patient-derived xenograft models. Venetoclax is currently being evaluated in pediatric patients in the clinic, including neuroblastoma (NCT03236857). Although establishment of safety is still ongoing, the data disclosed herein indicate rational and clinically actionable combination strategies that could potentiate the activity of venetoclax in patients with amplified MYCN with neuroblastoma.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Amplificación de Genes , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Proteína Proto-Oncogénica N-Myc/genética , Neuroblastoma/tratamiento farmacológico , Animales , Apoptosis , Compuestos Bicíclicos Heterocíclicos con Puentes/administración & dosificación , Proliferación Celular , Ciclofosfamida/administración & dosificación , Femenino , Humanos , Ratones , Ratones Endogámicos NOD , Ratones SCID , Neuroblastoma/genética , Neuroblastoma/metabolismo , Neuroblastoma/patología , Sulfonamidas/administración & dosificación , Topotecan/administración & dosificación , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Two-dimensional (2D) culture of tumor cells fails to recapitulate some important aspects of cellular organization seen in in vivo experiments. In addition, cell cultures traditionally use non-physiological concentration of nutrients. Here, we describe a protocol for a facile three-dimensional (3D) culture format for cancer cells. This 3D platform helps overcome the 2D culture limitations. In addition, it allows for longitudinal modeling of responses to cancer therapeutics. For complete details on the use and execution of this protocol, please refer to Lhuissier et al. (2017), Lehmann et al. (2016), Liu et al. (2016), and Duval et al. (2011).
Asunto(s)
Alginatos/química , Antineoplásicos/farmacología , Técnicas de Cultivo Tridimensional de Células/métodos , Cloruro de Calcio , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Humanos , Modelos BiológicosRESUMEN
MYCN-amplified neuroblastoma is a lethal subset of pediatric cancer. MYCN drives numerous effects in the cell, including metabolic changes that are critical for oncogenesis. The understanding that both compensatory pathways and intrinsic redundancy in cell systems exists implies that the use of combination therapies for effective and durable responses is necessary. Additionally, the most effective targeted therapies exploit an "Achilles' heel" and are tailored to the genetics of the cancer under study. We performed an unbiased screen on select metabolic targeted therapy combinations and correlated sensitivity with over 20 subsets of cancer. We found that MYCN-amplified neuroblastoma is hypersensitive to the combination of an inhibitor of the lactate transporter MCT1, AZD3965, and complex I of the mitochondrion, phenformin. Our data demonstrate that MCT4 is highly correlated with resistance to the combination in the screen and lowly expressed in MYCN-amplified neuroblastoma. Low MCT4 combines with high expression of the MCT2 and MCT1 chaperone CD147 in MYCN-amplified neuroblastoma, altogether conferring sensitivity to the AZD3965 and phenformin combination. The result is simultaneous disruption of glycolysis and oxidative phosphorylation, resulting in dramatic disruption of adenosine triphosphate (ATP) production, endoplasmic reticulum stress, and cell death. In mouse models of MYCN-amplified neuroblastoma, the combination was tolerable at concentrations where it shrank tumors and did not increase white-blood-cell toxicity compared to single drugs. Therefore, we demonstrate that a metabolic combination screen can identify vulnerabilities in subsets of cancer and put forth a metabolic combination therapy tailored for MYCN-amplified neuroblastoma that demonstrates efficacy and tolerability in vivo.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Complejo I de Transporte de Electrón/antagonistas & inhibidores , Transportadores de Ácidos Monocarboxílicos/antagonistas & inhibidores , Proteína Proto-Oncogénica N-Myc/genética , Neuroblastoma/tratamiento farmacológico , Simportadores/antagonistas & inhibidores , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Apoptosis/efectos de los fármacos , Basigina/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Complejo I de Transporte de Electrón/metabolismo , Amplificación de Genes , Humanos , Ratones , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Transportadores de Ácidos Monocarboxílicos/metabolismo , Neuroblastoma/genética , Neuroblastoma/patología , Fenformina/farmacología , Fenformina/uso terapéutico , Pirimidinonas/farmacología , Pirimidinonas/uso terapéutico , Simportadores/metabolismo , Tiofenos/farmacología , Tiofenos/uso terapéutico , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Identifying robust, patient-specific, and predictive biomarkers presents a major obstacle in precision oncology. To optimize patient-specific therapeutic strategies, here we couple pathway knowledge with large-scale drug sensitivity, RNAi, and CRISPR-Cas9 screening data from 460 cell lines. Pathway activity levels are found to be strong predictive biomarkers for the essentiality of 15 proteins, including the essentiality of MAD2L1 in breast cancer patients with high BRCA-pathway activity. We also find strong predictive biomarkers for the sensitivity to 31 compounds, including BCL2 and microtubule inhibitors (MTIs). Lastly, we show that Bcl-xL inhibition can modulate the activity of a predictive biomarker pathway and re-sensitize lung cancer cells and tumors to MTI therapy. Overall, our results support the use of pathways in helping to achieve the goal of precision medicine by uncovering dozens of predictive biomarkers.
Asunto(s)
Biomarcadores de Tumor/genética , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Neoplasias/genética , Transducción de Señal/genética , Animales , Antineoplásicos/farmacología , Proteína BRCA1/genética , Proteína BRCA1/metabolismo , Proteína BRCA2/genética , Proteína BRCA2/metabolismo , Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Femenino , Redes Reguladoras de Genes , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Ratones , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Medicina de Precisión/métodos , Interferencia de ARN , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Multidrug resistance (MDR) in cancer remains a major challenge for the success of chemotherapy. Natural products have been a rich source for the discovery of drugs against MDR cancers. Here, we applied high-throughput cytotoxicity screening of an in-house natural product library against MDR SGC7901/VCR cells and identified that the cyclodepsipeptide verucopeptin demonstrated notable antitumor potency. Cytological profiling combined with click chemistry-based proteomics revealed that ATP6V1G directly interacted with verucopeptin. ATP6V1G, a subunit of the vacuolar H+-ATPase (v-ATPase) that has not been previously targeted, was essential for SGC7901/VCR cell growth. Verucopeptin exhibited strong inhibition of both v-ATPase activity and mTORC1 signaling, leading to substantial pharmacological efficacy against SGC7901/VCR cell proliferation and tumor growth in vivo. Our results demonstrate that targeting v-ATPase via its V1G subunit constitutes a unique approach for modulating v-ATPase and mTORC1 signaling with great potential for the development of therapeutics against MDR cancers.