RESUMO
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.
Assuntos
Antineoplásicos , Neoplasias do Colo , Neoplasias Pancreáticas , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/genética , Combinação de Medicamentos , Sinergismo Farmacológico , Humanos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas p21(ras)/genéticaRESUMO
Functional genomics approaches can overcome limitations-such as the lack of identification of robust targets and poor clinical efficacy-that hamper cancer drug development. Here we performed genome-scale CRISPR-Cas9 screens in 324 human cancer cell lines from 30 cancer types and developed a data-driven framework to prioritize candidates for cancer therapeutics. We integrated cell fitness effects with genomic biomarkers and target tractability for drug development to systematically prioritize new targets in defined tissues and genotypes. We verified one of our most promising dependencies, the Werner syndrome ATP-dependent helicase, as a synthetic lethal target in tumours from multiple cancer types with microsatellite instability. Our analysis provides a resource of cancer dependencies, generates a framework to prioritize cancer drug targets and suggests specific new targets. The principles described in this study can inform the initial stages of drug development by contributing to a new, diverse and more effective portfolio of cancer drug targets.
Assuntos
Sistemas CRISPR-Cas/genética , Descoberta de Drogas/métodos , Edição de Genes , Terapia de Alvo Molecular/métodos , Neoplasias/genética , Neoplasias/terapia , Animais , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Feminino , Genoma Humano/genética , Humanos , Camundongos , Instabilidade de Microssatélites , Transplante de Neoplasias , Neoplasias/classificação , Neoplasias/patologia , Especificidade de Órgãos , Reprodutibilidade dos Testes , Mutações Sintéticas Letais/genética , Síndrome de Werner/genética , Helicase da Síndrome de Werner/genéticaRESUMO
HER2 amplification occurs in approximately 5% of colorectal cancer (CRC) cases and is associated only partially with clinical response to combined human epidermal growth factor receptor 2 (HER2)/epidermal growth factor receptor (EGFR)-targeted treatment. An alternative approach based on adoptive cell therapy using T cells engineered with anti-HER2 chimeric antigen receptor (CAR) proved to be toxic due to on-target/off-tumor activity. Here we describe a combinatorial strategy to safely target HER2 amplification and carcinoembryonic antigen (CEA) expression in CRC using a synNotch-CAR-based artificial regulatory network. The natural killer (NK) cell line NK-92 was engineered with an anti-HER2 synNotch receptor driving the expression of a CAR against CEA only when engaged. After being transduced and sorted for HER2-driven CAR expression, cells were cloned. The clone with optimal performances in terms of specificity and amplitude of CAR induction demonstrated significant activity in vitro and in vivo specifically against HER2-amplified (HER2amp)/CEA+ CRC models, with no effects on cells with physiological HER2 levels. The HER2-synNotch/CEA-CAR-NK system provides an innovative, scalable, and safe off-the-shelf cell therapy approach with potential against HER2amp CRC resistant or partially responsive to HER2/EGFR blockade.
Assuntos
Neoplasias Colorretais , Receptor ErbB-2 , Receptores de Antígenos Quiméricos , Ensaios Antitumorais Modelo de Xenoenxerto , Neoplasias Colorretais/terapia , Neoplasias Colorretais/genética , Neoplasias Colorretais/imunologia , Humanos , Receptor ErbB-2/metabolismo , Receptor ErbB-2/genética , Animais , Receptores de Antígenos Quiméricos/imunologia , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos Quiméricos/metabolismo , Camundongos , Linhagem Celular Tumoral , Antígeno Carcinoembrionário/imunologia , Antígeno Carcinoembrionário/genética , Amplificação de Genes , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Imunoterapia/métodos , Imunoterapia Adotiva/métodos , Modelos Animais de Doenças , FemininoRESUMO
MOTIVATION: The transition from evaluating a single time point to examining the entire dynamic evolution of a system is possible only in the presence of the proper framework. The strong variability of dynamic evolution makes the definition of an explanatory procedure for data fitting and clustering challenging. RESULTS: We developed CONNECTOR, a data-driven framework able to analyze and inspect longitudinal data in a straightforward and revealing way. When used to analyze tumor growth kinetics over time in 1599 patient-derived xenograft growth curves from ovarian and colorectal cancers, CONNECTOR allowed the aggregation of time-series data through an unsupervised approach in informative clusters. We give a new perspective of mechanism interpretation, specifically, we define novel model aggregations and we identify unanticipated molecular associations with response to clinically approved therapies. AVAILABILITY AND IMPLEMENTATION: CONNECTOR is freely available under GNU GPL license at https://qbioturin.github.io/connector and https://doi.org/10.17504/protocols.io.8epv56e74g1b/v1.
Assuntos
Software , Humanos , Animais , Análise por Conglomerados , Fatores de Tempo , Modelos Animais de Doenças , Medição de RiscoRESUMO
The MET tyrosine kinase receptor (also known as the HGF receptor) promotes tissue remodelling, which underlies developmental morphogenesis, wound repair, organ homeostasis and cancer metastasis, by integrating growth, survival and migration cues in response to environmental stimuli or cell-autonomous perturbations. The versatility of MET-mediated biological responses is sustained by qualitative and quantitative signal modulation. Qualitative mechanisms include the engagement of dedicated signal transducers and the subcellular compartmentalization of MET signalling pathways, whereas quantitative regulation involves MET partnering with adaptor amplifiers or being degraded through the shedding of its extracellular domain or through intracellular ubiquitylation. Controlled activation of MET signalling can be exploited in regenerative medicine, whereas MET inhibition might slow down tumour progression.
Assuntos
Estruturas Animais/fisiologia , Crescimento e Desenvolvimento/genética , Neoplasias/genética , Proteínas Proto-Oncogênicas c-met/fisiologia , Regeneração/genética , Estruturas Animais/metabolismo , Animais , Crescimento e Desenvolvimento/fisiologia , Humanos , Modelos Biológicos , Modelos Moleculares , Neoplasias/etiologia , Proteínas Proto-Oncogênicas c-met/química , Proteínas Proto-Oncogênicas c-met/genética , Proteínas Proto-Oncogênicas c-met/metabolismo , Regeneração/fisiologia , Transdução de Sinais/genética , Transdução de Sinais/fisiologiaRESUMO
Colorectal cancer is the third most common cancer worldwide, with 1.2 million patients diagnosed annually. In late-stage colorectal cancer, the most commonly used targeted therapies are the monoclonal antibodies cetuximab and panitumumab, which prevent epidermal growth factor receptor (EGFR) activation. Recent studies have identified alterations in KRAS and other genes as likely mechanisms of primary and secondary resistance to anti-EGFR antibody therapy. Despite these efforts, additional mechanisms of resistance to EGFR blockade are thought to be present in colorectal cancer and little is known about determinants of sensitivity to this therapy. To examine the effect of somatic genetic changes in colorectal cancer on response to anti-EGFR antibody therapy, here we perform complete exome sequence and copy number analyses of 129 patient-derived tumour grafts and targeted genomic analyses of 55 patient tumours, all of which were KRAS wild-type. We analysed the response of tumours to anti-EGFR antibody blockade in tumour graft models and in clinical settings and functionally linked therapeutic responses to mutational data. In addition to previously identified genes, we detected mutations in ERBB2, EGFR, FGFR1, PDGFRA, and MAP2K1 as potential mechanisms of primary resistance to this therapy. Novel alterations in the ectodomain of EGFR were identified in patients with acquired resistance to EGFR blockade. Amplifications and sequence changes in the tyrosine kinase receptor adaptor gene IRS2 were identified in tumours with increased sensitivity to anti-EGFR therapy. Therapeutic resistance to EGFR blockade could be overcome in tumour graft models through combinatorial therapies targeting actionable genes. These analyses provide a systematic approach to evaluating response to targeted therapies in human cancer, highlight new mechanisms of responsiveness to anti-EGFR therapies, and delineate new avenues for intervention in managing colorectal cancer.
Assuntos
Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Receptores ErbB/antagonistas & inibidores , Genoma Humano/genética , Genômica , Animais , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Cetuximab/farmacologia , Cetuximab/uso terapêutico , Neoplasias Colorretais/metabolismo , Variações do Número de Cópias de DNA/genética , Receptores ErbB/química , Receptores ErbB/genética , Exoma/genética , Feminino , Humanos , Proteínas Substratos do Receptor de Insulina/genética , MAP Quinase Quinase 1/genética , Camundongos , Terapia de Alvo Molecular , Mutação/genética , Panitumumabe , Proteínas Proto-Oncogênicas p21(ras)/genética , Receptor ErbB-2/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Antibodies targeting the human epidermal growth factor receptor (EGFR) are used for the treatment of RAS wild-type metastatic colorectal cancer. A significant proportion of patients remains unresponsive to this therapy. Here, we performed a reverse-phase protein array-based (phospho)protein analysis of 63 KRAS, NRAS, BRAF and PIK3CA wild-type metastatic CRC tumours. Responses of tumours to anti-EGFR therapy with cetuximab were recorded in patient-derived xenograft (PDX) models. Unsupervised hierarchical clustering of pretreatment tumour tissue identified three clusters, of which Cluster C3 was exclusively composed of responders. Clusters C1 and C2 exhibited mixed responses. None of the three protein clusters exhibited a significant correlation with transcriptome-based subtypes. Analysis of protein signatures across all PDXs identified 14 markers that discriminated cetuximab-sensitive and cetuximab-resistant tumours: PDK1 (S241), caspase-8, Shc (Y317), Stat3 (Y705), p27, GSK-3ß (S9), HER3, PKC-α (S657), EGFR (Y1068), Akt (S473), S6 ribosomal protein (S240/244), HER3 (Y1289), NF-κB-p65 (S536) and Gab-1 (Y627). Least absolute shrinkage and selection operator and binominal logistic regression analysis delivered refined protein signatures for predicting response to cetuximab. (Phospo-)protein analysis of matched pretreated and posttreated models furthermore showed significant reduction of Gab-1 (Y627) and GSK-3ß (S9) exclusively in responding models, suggesting novel targets for treatment.
Assuntos
Cetuximab/administração & dosagem , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/secundário , Fosfoproteínas/metabolismo , Proteômica/métodos , Animais , Proliferação de Células/efeitos dos fármacos , Cetuximab/farmacologia , Classe I de Fosfatidilinositol 3-Quinases/genética , Análise por Conglomerados , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Feminino , GTP Fosfo-Hidrolases/genética , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Masculino , Proteínas de Membrana/genética , Camundongos , Fosfoproteínas/efeitos dos fármacos , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Aprendizado de Máquina não Supervisionado , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Identifying hyperactive kinases in cancer is crucial for individualized treatment with specific inhibitors. Kinase activity can be discerned from global protein phosphorylation profiles obtained with mass spectrometry-based phosphoproteomics. A major challenge is to relate such profiles to specific hyperactive kinases fueling growth/progression of individual tumors. Hitherto, the focus has been on phosphorylation of either kinases or their substrates. Here, we combined label-free kinase-centric and substrate-centric information in an Integrative Inferred Kinase Activity (INKA) analysis. This multipronged, stringent analysis enables ranking of kinase activity and visualization of kinase-substrate networks in a single biological sample. To demonstrate utility, we analyzed (i) cancer cell lines with known oncogenes, (ii) cell lines in a differential setting (wild-type versus mutant, +/- drug), (iii) pre- and on-treatment tumor needle biopsies, (iv) cancer cell panel with available drug sensitivity data, and (v) patient-derived tumor xenografts with INKA-guided drug selection and testing. These analyses show superior performance of INKA over its components and substrate-based single-sample tool KARP, and underscore target potential of high-ranking kinases, encouraging further exploration of INKA's functional and clinical value.
Assuntos
Neoplasias/enzimologia , Fosfotransferases/análise , Proteômica/métodos , Biologia de Sistemas/métodos , Linhagem Celular Tumoral , Ativação Enzimática , Humanos , Células K562 , Espectrometria de Massas , Fosfoproteínas/análiseRESUMO
BACKGROUND: HER2 amplification is detected in 3% of patients with colorectal cancer (CRC), making tumors in the metastatic setting vulnerable to double pharmacological HER2 blockade. Preclinical findings show that it also might impair response to anti-epidermal growth factor receptor (EGFR) treatment. SUBJECTS AND METHODS: Patients with KRAS exon 2 wild-type metastatic CRC underwent molecular screening of HER2 positivity by HERACLES criteria (immunohistochemistry 3+ or 2+ in ≥50% of cells, confirmed by fluorescence in situ hybridization). A sample of consecutive HER2-negative patients was selected as control. A regression modeling strategy was applied to identify predictors explaining the bulk of HER2 positivity and the association with response to previous anti-EGFR treatment. RESULTS: From August 2012 to April 2018, a total of 100 HER2-positive metastatic CRC tumors were detected out of 1,485 KRAS exon 2 wild-type screened patients (6.7%). HER2-positive patients show more frequently lung metastases (odds ratio [OR], 2.04; 95% confidence interval [CI], 1.15-3.61; p = .014) and higher tumor burden (OR, 1.48; 95% CI, 1.10-2.01; p = .011), and tumors were more likely to be left sided (OR, 0.50; 95% CI, 0.22-1.11; p = .088). HER2-positive patients who received treatment with anti-EGFR agents (n = 79) showed poorer outcome (objective response rate, 31.2% vs. 46.9%, p = .031; progression-free survival, 5.7 months vs. 7 months, p = .087). CONCLUSION: Testing for HER2 should be offered to all patients with metastatic CRC because the occurrence of this biomarker is unlikely to be predicted based on main clinicopathological features. Patients with HER2-amplified metastatic CRC are less likely to respond to anti-EGFR therapy. IMPLICATIONS FOR PRACTICE: Patients with HER2-amplified/overexpressed metastatic colorectal cancer (mCRC) harbor a driver actionable molecular alteration that has been shown in preclinical models to hamper efficacy of the anti-epidermal growth factor receptor (EGFR) targeted therapies. The present study confirmed that this molecular feature was associated with worse objective tumor response and shorter progression-free survival in response to previous anti-EGFR therapies. Moreover, it was found that the occurrence of this biomarker is unlikely to be predicted based on main clinicopathological features. Therefore, HER2 status assessment should be included in the molecular diagnostic workup of all mCRC for speedy referral to clinical trials encompassing HER2-targeted double blockade independently of previous anti-EGFR treatment.
Assuntos
Neoplasias Colorretais/tratamento farmacológico , Receptor ErbB-2/metabolismo , Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Receptores ErbB/metabolismo , Feminino , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Terapia de Alvo Molecular , Metástase Neoplásica , Intervalo Livre de Progressão , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-2/genética , Estudos RetrospectivosRESUMO
BACKGROUND: Tankyrases are poly(adenosine diphosphate)-ribose polymerases that contribute to biological processes as diverse as modulation of Wnt signaling, telomere maintenance, vesicle trafficking, and microtubule-dependent spindle pole assembly during mitosis. At interphase, polarized reshaping of the microtubule network fosters oriented cell migration. This is attained by association of adenomatous polyposis coli with the plus end of microtubules at the cortex of cell membrane protrusions and microtubule-based centrosome reorientation towards the migrating front. RESULTS: Here we report a new function for tankyrases, namely, regulation of directional cell locomotion. Using a panel of lung cancer cell lines as a model system, we found that abrogation of tankyrase activity by two different, structurally unrelated small-molecule inhibitors (one introduced and characterized here for the first time) or by RNA interference-based genetic silencing weakened cell migration, invasion, and directional movement induced by the motogenic cytokine hepatocyte growth factor. Mechanistically, the anti-invasive outcome of tankyrase inhibition could be ascribed to sequential deterioration of the distinct events that govern cell directional sensing. In particular, tankyrase blockade negatively impacted (1) microtubule dynamic instability; (2) adenomatous polyposis coli plasma membrane targeting; and (3) centrosome reorientation. CONCLUSIONS: Collectively, these findings uncover an unanticipated role for tankyrases in influencing at multiple levels the interphase dynamics of the microtubule network and the subcellular distribution of related polarity signals. These results encourage the further exploration of tankyrase inhibitors as therapeutic tools to oppose dissemination and metastasis of cancer cells.
Assuntos
Movimento Celular/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Pulmão/efeitos dos fármacos , Tanquirases/antagonistas & inibidores , Linhagem Celular Tumoral , Inibidores Enzimáticos/química , Humanos , Pulmão/metabolismo , Pulmão/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Microtúbulos/metabolismo , Microtúbulos/patologia , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Invasividade Neoplásica/prevenção & controle , Interferência de RNA , Tanquirases/genética , Tanquirases/metabolismoRESUMO
BACKGROUND: We previously found that dual HER2 blockade with trastuzumab and lapatinib led to inhibition of tumour growth in patient-derived xenografts of HER2-amplified metastatic colorectal cancer. In this study, we aimed to assess the antitumour activity of trastuzumab and lapatinib in patients with HER2-positive colorectal cancer. METHODS: HERACLES was a proof-of-concept, multicentre, open-label, phase 2 trial done at four Italian academic cancer centres. We enrolled adult patients with KRAS exon 2 (codons 12 and 13) wild-type and HER2-positive metastatic colorectal cancer refractory to standard of care (including cetuximab or panitumumab), an Eastern Cooperative Oncology Group performance status of 0 or 1, and at least one measurable lesion. We defined HER2 positivity in tumour samples by use of immunohistochemistry and fluorescence in-situ hybridisation in accordance with our previously validated colorectal cancer-specific diagnostic criteria. Eligible patients received intravenous trastuzumab at 4 mg/kg loading dose followed by 2 mg/kg once per week, and oral lapatinib at 1000 mg per day until evidence of disease progression. The primary endpoint was the proportion of patients achieving an objective response (defined as complete response or partial response), which was assessed by independent central review in the intention-to-treat population. This trial is registered with EudraCT, number 2012-002128-33. FINDINGS: Between Aug 27, 2012, and May 15, 2015, we screened 914 patients with KRAS exon 2 (codons 12 and 13) wild-type metastatic colorectal cancer and identified 48 (5%) patients with HER2-positive tumours, although two died before enrolment. Of these patients, 27 were eligible for the trial. All were evaluable for response. At the time of data cutoff on Oct 15, 2015, with a median follow-up of 94 weeks (IQR 51-127), eight (30%, 95% CI 14-50) of 27 patients had achieved an objective response, with one patient (4%, 95% CI -3 to 11) achieving a complete response, and seven (26%, 95% CI 9-43) achieving partial responses; 12 (44%, 95% CI 25-63) patients had stable disease. Six (22%) of 27 patients had grade 3 adverse events, which consisted of fatigue in four patients, skin rash in one patient, and increased bilirubin concentration in one patient. No grade 4 or 5 adverse events were reported. We detected no drug-related serious adverse events. INTERPRETATION: The combination of trastuzumab and lapatinib is active and well tolerated in treatment-refractory patients with HER2-positive metastatic colorectal cancer. FUNDING: Associazione Italiana Ricerca Cancro (AIRC), Fondazione Oncologia Niguarda Onlus, and Roche.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Terapia de Alvo Molecular , Recidiva Local de Neoplasia/tratamento farmacológico , Proteínas Proto-Oncogênicas p21(ras)/genética , Receptor ErbB-2/antagonistas & inibidores , Adulto , Idoso , Biomarcadores Tumorais , Códon/genética , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Feminino , Seguimentos , Humanos , Técnicas Imunoenzimáticas , Lapatinib , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/metabolismo , Recidiva Local de Neoplasia/patologia , Estadiamento de Neoplasias , Prognóstico , Quinazolinas/administração & dosagem , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Terapia de Salvação , Taxa de Sobrevida , Trastuzumab/administração & dosagemRESUMO
Inhibitors of poly(ADP-ribose) polymerases actualized the biological concept of synthetic lethality in the clinical practice, yielding a paradigmatic example of translational medicine. The profound sensitivity of tumors with germline BRCA mutations to PARP1/2 blockade owes to inherent defects of the BRCA-dependent homologous recombination machinery, which are unleashed by interruption of PARP DNA repair activity and lead to DNA damage overload and cell death. Conversely, aspirant BRCA-like tumors harboring somatic DNA repair dysfunctions (a vast entity of genetic and epigenetic defects known as "BRCAness") not always align with the familial counterpart and appear not to be equally sensitive to PARP inhibition. The acquisition of secondary resistance in initially responsive patients and the lack of standardized biomarkers to identify "BRCAness" pose serious threats to the clinical advance of PARP inhibitors; a feeling is also emerging that a BRCA-centered perspective might have missed the influence of additional, not negligible and DNA repair-independent PARP contributions onto therapy outcome. While regulatory approval for PARP1/2 inhibitors is still pending, novel therapeutic opportunities are sprouting from different branches of the PARP family, although they remain immature for clinical extrapolation. This review is an endeavor to provide a comprehensive appraisal of the multifaceted biology of PARPs and their evolving impact on cancer therapeutics.
Assuntos
Antineoplásicos/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Neoplasias/tratamento farmacológico , Inibidores de Poli(ADP-Ribose) Polimerases , Adenosina Difosfato Ribose/metabolismo , Animais , Dano ao DNA/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Poli(ADP-Ribose) Polimerases/metabolismoRESUMO
MET is a master gene controlling a genetic program driving proliferation, apoptosis protection and invasion. The ROR1 pseudokinase acts as a MET substrate. However, its contribution to MET signaling and MET-dependent biological outcomes remains to be elucidated. By structure-function analysis of ROR1 mutants, we show that ROR1 encompasses two major substrate regions: one is located in the proline-rich domain and is directly phosphorylated by MET; the other resides in the pseudokinase domain and is phosphorylated through intermediate activation of SRC. Differential phosphorylation of these two regions dictates the execution of specific responses: phosphorylation of the ROR1 proline-rich domain by MET-but not phosphorylation of the pseudokinase domain by SRC-is necessary and sufficient to control MET-driven proliferation and protection from apoptosis. Differently, both the proline-rich and the pseudokinase domains mediate cell invasion. Consistent with the role of ROR1 in specifying the functional consequences of MET-dependent signals, ROR1 silencing leads to selective attenuation of only some of the signal transduction pathways sustained by MET. These data enlighten the so far elusive function(s) of pseudokinases and identify a mechanism of biological diversification, based on substrate specificity of oncogenic kinases.
Assuntos
Proteínas Proto-Oncogênicas c-met/metabolismo , Pseudogenes/fisiologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Transdução de Sinais , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Animais , Apoptose , Western Blotting , Células COS , Movimento Celular , Proliferação de Células , Células Cultivadas , Chlorocebus aethiops , Humanos , Imunoprecipitação , Lentivirus/genética , Fosforilação , Plasmídeos/genética , Proteínas Proto-Oncogênicas c-met/genética , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genéticaRESUMO
Abnormal activation of MET/HGF (Hepatocyte Growth Factor) pathway has been described in several tumours and increased HGF plasmatic levels have been detected in patients with aggressive multiple myeloma (MM). MET and HGF mRNA expression was investigated in 105 samples of purified plasma cells derived from newly diagnosed MM patients treated with bortezomib-based induction therapy. Gene expression was compared with response to therapy and clinical outcome. MET gene copy number was also evaluated. MET mRNA expression was higher in CD138(+) than in CD138(-) cells (median 76·90 vs. 11·24; P = 0·0009). Low MET mRNA expression characterized patients with better response (complete response or very good partial response) compared to other patients (median 56·10 vs. 134·83; P = 0·0006). After a median follow-up of 50 months, patients with high MET mRNA expression displayed a worse progression-free survival (PFS; P = 0·0029) and overall survival (OS; P = 0·0023) compared to those with low MET mRNA levels. Patients with both high MET mRNA expression and high ß2-microglobulin level (>5·5 mg/l) had further worse median PFS (P < 0·0001) and OS (P < 0·0001). Patients carrying 4 MET gene copies (8 out of 82, 9·8%) also had a short PFS. High MET mRNA expression identifies patients with dismal PFS and OS and the combination with high ß2-microglobulin further characterizes patients with worse outcome.
Assuntos
Fator de Crescimento de Hepatócito/metabolismo , Mieloma Múltiplo/metabolismo , Mieloma Múltiplo/patologia , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/metabolismo , Feminino , Fator de Crescimento de Hepatócito/biossíntese , Fator de Crescimento de Hepatócito/sangue , Fator de Crescimento de Hepatócito/genética , Humanos , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/genética , Mieloma Múltiplo/terapia , Prognóstico , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
BACKGROUND: Standard chemotherapy in unresectable biliary tract carcinoma (BTC) patients is based on gemcitabine combined with platinum derivatives. However, primary or acquired resistance is inevitable and no second-line chemotherapy is demonstrated to be effective. Thus, there is an urgent need to identify new alternative (chemo)therapy approaches. METHODS: We evaluated the mechanism of action of ET-743 in preclinical models of BTC. Six BTC cell lines (TFK-1, EGI-1, TGBC1, WITT, KMCH, HuH28), two primary cell cultures derived from BTC patients, the EGI-1 and a new established BTC patient-derived xenografts, were used as preclinical models to investigate the anti-tumor activity of ET-743 in vitro and in vivo. Gene expression profiling was also analyzed upon ET-743 treatment in in vivo models. RESULTS: We found that ET-743 inhibited cell growth of BTC cell lines and primary cultures (IC50 ranging from 0.37 to 3.08 nM) preferentially inducing apoptosis and activation of the complex DNA damage-repair proteins (p-ATM, p-p53 and p-Histone H2A.x) in vitro. In EGI-1 and patient-derived xenografts, ET-743 induced tumor growth delay and reduction of vasculogenesis. In vivo ET-743 induced a deregulation of genes involved in cell adhesion, stress-related response, and in pathways involved in cholangiocarcinogenesis, such as the IL-6, Sonic Hedgehog and Wnt signaling pathways. CONCLUSIONS: These results suggest that ET-743 could represent an alternative chemotherapy for BTC treatment and encourage the development of clinical trials in BTC patients resistant to standard chemotherapy.
Assuntos
Antineoplásicos Alquilantes/farmacologia , Neoplasias do Sistema Biliar/tratamento farmacológico , Dioxóis/farmacologia , Tetra-Hidroisoquinolinas/farmacologia , Animais , Apoptose/efeitos dos fármacos , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Neoplasias do Sistema Biliar/irrigação sanguínea , Neoplasias do Sistema Biliar/genética , Adesão Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Transformação Celular Neoplásica/genética , Reparo do DNA/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Histonas/metabolismo , Humanos , Interleucina-6/genética , Camundongos , Camundongos Endogâmicos NOD , Neovascularização Patológica/tratamento farmacológico , Fosforilação , Trabectedina , Proteína Supressora de Tumor p53/metabolismo , Via de Sinalização Wnt/efeitos dos fármacosAssuntos
Experimentação Animal/legislação & jurisprudência , Bem-Estar do Animal/legislação & jurisprudência , Animais de Laboratório , Pesquisa Biomédica/métodos , Bem-Estar do Animal/normas , Animais , União Europeia , Humanos , Itália , Camundongos , Transtornos Relacionados ao Uso de Substâncias , Transplante HeterólogoRESUMO
The bacterial genotoxin colibactin promotes colorectal cancer (CRC) tumorigenesis, but systematic assessment of its impact on DNA repair is lacking, and its effect on response to DNA-damaging chemotherapeutics is unknown. We find that CRC cell lines display differential response to colibactin on the basis of homologous recombination (HR) proficiency. Sensitivity to colibactin is induced by inhibition of ATM, which regulates DNA double-strand break repair, and blunted by HR reconstitution. Conversely, CRC cells chronically infected with colibactin develop a tolerant phenotype characterized by restored HR activity. Notably, sensitivity to colibactin correlates with response to irinotecan active metabolite SN38, in both cell lines and patient-derived organoids. Moreover, CRC cells that acquire colibactin tolerance develop cross-resistance to SN38, and a trend toward poorer response to irinotecan is observed in a retrospective cohort of CRCs harboring colibactin genomic island. Our results shed insight into colibactin activity and provide translational evidence on its chemoresistance-promoting role in CRC.
Assuntos
Neoplasias Colorretais , Escherichia coli , Peptídeos , Policetídeos , Humanos , Irinotecano/farmacologia , Escherichia coli/genética , Escherichia coli/metabolismo , Estudos Retrospectivos , DNA/metabolismo , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Neoplasias Colorretais/microbiologiaRESUMO
Molecular stratification using gene-level transcriptional data has identified subtypes with distinctive genotypic and phenotypic traits, as exemplified by the consensus molecular subtypes (CMS) in colorectal cancer (CRC). Here, rather than gene-level data, we make use of gene ontology and biological activation state information for initial molecular class discovery. In doing so, we defined three pathway-derived subtypes (PDS) in CRC: PDS1 tumors, which are canonical/LGR5+ stem-rich, highly proliferative and display good prognosis; PDS2 tumors, which are regenerative/ANXA1+ stem-rich, with elevated stromal and immune tumor microenvironmental lineages; and PDS3 tumors, which represent a previously overlooked slow-cycling subset of tumors within CMS2 with reduced stem populations and increased differentiated lineages, particularly enterocytes and enteroendocrine cells, yet display the worst prognosis in locally advanced disease. These PDS3 phenotypic traits are evident across numerous bulk and single-cell datasets, and demark a series of subtle biological states that are currently under-represented in pre-clinical models and are not identified using existing subtyping classifiers.
Assuntos
Neoplasias Colorretais , Humanos , Neoplasias Colorretais/patologia , Prognóstico , Diferenciação Celular/genética , Fenótipo , Biomarcadores Tumorais/genética , Perfilação da Expressão GênicaRESUMO
PURPOSE: ERBB2-amplified colorectal cancer is a distinct molecular subtype with expanding treatments. Implications of concurrent oncogenic RAS/RAF alterations are not known. EXPERIMENTAL DESIGN: Dana-Farber and Foundation Medicine Inc. Colorectal cancer cohorts with genomic profiling were used to identify ERBB2-amplified cases [Dana-Farber, n = 47/2,729 (1.7%); FMI, n = 1857/49,839 (3.7%)]. Outcomes of patients receiving HER2-directed therapies are reported (Dana-Farber, n = 9; Flatiron Health-Foundation Medicine clinicogenomic database, FH-FMI CGDB, n = 38). Multisite HER2 IHC and genomic profiling were performed to understand HER2 intratumoral and interlesional heterogeneity. The impact of concurrent RAS comutations on the effectiveness of HER2-directed therapies were studied in isogenic colorectal cancer cell lines and xenografts. RESULTS: ERBB2 amplifications are enriched in left-sided colorectal cancer. Twenty percent of ERBB2-amplified colorectal cancers have co-occurring oncogenic RAS/RAF alterations. While RAS/RAF WT colorectal cancers typically have clonal ERBB2 amplification, colorectal cancers with co-occurring RAS/RAF alterations have lower level ERRB2 amplification, higher intratumoral heterogeneity, and interlesional ERBB2 discordance. These distinct genomic patterns lead to differential responsiveness and patterns of resistance to HER2-directed therapy. ERBB2-amplified colorectal cancer with RAS/RAF alterations are resistant to trastuzumab-based combinations, such as trastuzumab/tucatinib, but retain sensitivity to trastuzumab deruxtecan in in vitro and murine models. Trastuzumab deruxtecan shows clinical efficacy in cases with high-level ERBB2-amplified RAS/RAF coaltered colorectal cancer. CONCLUSIONS: Co-occurring RAS/RAF alterations define a unique subtype of ERBB2-amplified colorectal cancer that has increased intratumoral heterogeneity, interlesional discordance, and resistance to trastuzumab-based combinations. Further examination of trastuzumab deruxtecan in this previously understudied cohort of ERBB2-amplified colorectal cancer is warranted.