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1.
Brief Bioinform ; 25(4)2024 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-38783705

RESUMO

Tumor mutational signatures have gained prominence in cancer research, yet the lack of standardized methods hinders reproducibility and robustness. Leveraging colorectal cancer (CRC) as a model, we explored the influence of computational parameters on mutational signature analyses across 230 CRC cell lines and 152 CRC patients. Results were validated in three independent datasets: 483 endometrial cancer patients stratified by mismatch repair (MMR) status, 35 lung cancer patients by smoking status and 12 patient-derived organoids (PDOs) annotated for colibactin exposure. Assessing various bioinformatic tools, reference datasets and input data sizes including whole genome sequencing, whole exome sequencing and a pan-cancer gene panel, we demonstrated significant variability in the results. We report that the use of distinct algorithms and references led to statistically different results, highlighting how arbitrary choices may induce variability in the mutational signature contributions. Furthermore, we found a differential contribution of mutational signatures between coding and intergenic regions and defined the minimum number of somatic variants required for reliable mutational signature assignment. To facilitate the identification of the most suitable workflows, we developed Comparative Mutational Signature analysis on Coding and Extragenic Regions (CoMSCER), a bioinformatic tool which allows researchers to easily perform comparative mutational signature analysis by coupling the results from several tools and public reference datasets and to assess mutational signature contributions in coding and non-coding genomic regions. In conclusion, our study provides a comparative framework to elucidate the impact of distinct computational workflows on mutational signatures.


Assuntos
Neoplasias Colorretais , Biologia Computacional , Mutação , Humanos , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Biologia Computacional/métodos , Fluxo de Trabalho , Linhagem Celular Tumoral , Sequenciamento do Exoma/métodos , Feminino , Algoritmos
2.
Mol Oncol ; 18(6): 1460-1485, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38468448

RESUMO

Multiple strategies are continuously being explored to expand the drug target repertoire in solid tumors. We devised a novel computational workflow for transcriptome-wide gene expression outlier analysis that allows the systematic identification of both overexpression and underexpression events in cancer cells. Here, it was applied to expression values obtained through RNA sequencing in 226 colorectal cancer (CRC) cell lines that were also characterized by whole-exome sequencing and microarray-based DNA methylation profiling. We found cell models displaying an abnormally high or low expression level for 3533 and 965 genes, respectively. Gene expression abnormalities that have been previously associated with clinically relevant features of CRC cell lines were confirmed. Moreover, by integrating multi-omics data, we identified both genetic and epigenetic alternations underlying outlier expression values. Importantly, our atlas of CRC gene expression outliers can guide the discovery of novel drug targets and biomarkers. As a proof of concept, we found that CRC cell lines lacking expression of the MTAP gene are sensitive to treatment with a PRMT5-MTA inhibitor (MRTX1719). Finally, other tumor types may also benefit from this approach.


Assuntos
Neoplasias Colorretais , Regulação Neoplásica da Expressão Gênica , Transcriptoma , Humanos , Neoplasias Colorretais/genética , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/patologia , Neoplasias Colorretais/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Linhagem Celular Tumoral , Transcriptoma/genética , Perfilação da Expressão Gênica , Metilação de DNA/genética
3.
Mol Oncol ; 18(2): 241-244, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38308461

RESUMO

Drug resistance represents a major limitation to the long-term efficacy of anti-cancer treatments. The commonly accepted view is that the selection of inheritable genetic mechanisms governs the development of secondary resistance. However, compelling evidence suggests an important role for adaptive cell plasticity and non-genetic mechanisms in the development of therapy resistance. The two phenomena are not mutually exclusive and the interplay between genetic and non-genetic mechanisms may affect tumor evolution during treatment. A broader characterization of the genetic and non-genetic mechanisms of drug resistance may pave the way for more precise and effective therapeutic strategies to overcome resistance.


Assuntos
Neoplasias , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/genética , Resistência a Medicamentos , Epigênese Genética , Resistencia a Medicamentos Antineoplásicos/genética
4.
Cell Rep Med ; 5(2): 101376, 2024 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-38228147

RESUMO

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/microbiologia
5.
J Exp Clin Cancer Res ; 42(1): 120, 2023 May 12.
Artigo em Inglês | MEDLINE | ID: mdl-37170152

RESUMO

BACKGROUND: MET-driven acquired resistance is emerging with unanticipated frequency in patients relapsing upon molecular therapy treatments. However, the determination of MET amplification remains challenging using both standard and next-generation sequencing-based methodologies. Liquid biopsy is an effective, non-invasive approach to define cancer genomic profiles, track tumor evolution over time, monitor treatment response and detect molecular resistance in advance. Circular RNAs (circRNAs), a family of RNA molecules that originate from a process of back-splicing, are attracting growing interest as potential novel biomarkers for their stability in body fluids. METHODS: We identified a circRNA encoded by the MET gene (circMET) and exploited blood-derived cell-free RNA (cfRNA) and matched tumor tissues to identify, stratify and monitor advanced cancer patients molecularly characterized by high MET activity, generally associated with genomic amplification. RESULTS: Using publicly available bioinformatic tools, we discovered that the MET locus transcribes several circRNA molecules, but only one candidate, circMET, was particularly abundant. Deeper molecular analysis revealed that circMET levels positively correlated with MET expression and activity, especially in MET-amplified cells. We developed a circMET-detection strategy and, in parallel, we performed standard FISH and IHC analyses in the same specimens to assess whether circMET quantification could identify patients displaying high MET activity. Longitudinal monitoring of circMET levels in the plasma of selected patients revealed the early emergence of MET amplification as a mechanism of acquired resistance to molecular therapies. CONCLUSIONS: We found that measurement of circMET levels allows identification and tracking of patients characterized by high MET activity. Circulating circMET (ccMET) detection and analysis could be a simple, cost-effective, non-invasive approach to better implement patient stratification based on MET expression, as well as to dynamically monitor over time both therapy response and clonal evolution during treatment.


Assuntos
Neoplasias , RNA Circular , Humanos , Biomarcadores , Biologia Computacional , Neoplasias/genética , RNA/genética , RNA/metabolismo , RNA Circular/genética
6.
Nat Genet ; 54(7): 976-984, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35817983

RESUMO

Compelling evidence shows that cancer persister cells represent a major limit to the long-term efficacy of targeted therapies. However, the phenotype and population dynamics of cancer persister cells remain unclear. We developed a quantitative framework to study persisters by combining experimental characterization and mathematical modeling. We found that, in colorectal cancer, a fraction of persisters slowly replicates. Clinically approved targeted therapies induce a switch to drug-tolerant persisters and a temporary 7- to 50-fold increase of their mutation rate, thus increasing the number of persister-derived resistant cells. These findings reveal that treatment may influence persistence and mutability in cancer cells and pinpoint inhibition of error-prone DNA polymerases as a strategy to restrict tumor recurrence.


Assuntos
Neoplasias Colorretais , Taxa de Mutação , Antibacterianos/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Humanos , Dinâmica Populacional
7.
Cancer Discov ; 11(8): 1886-1895, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33952585

RESUMO

Cancer is characterized by loss of the regulatory mechanisms that preserve homeostasis in multicellular organisms, such as controlled proliferation, cell-cell adhesion, and tissue differentiation. The breakdown of multicellularity rules is accompanied by activation of "selfish," unicellular-like life features, which are linked to the increased adaptability to environmental changes displayed by cancer cells. Mechanisms of stress response, resembling those observed in unicellular organisms, are actively exploited by mammalian cancer cells to boost genetic diversity and increase chances of survival under unfavorable conditions, such as lack of oxygen/nutrients or exposure to drugs. Unicellular organisms under stressful conditions (e.g., antibiotic treatment) stop replicating or slowly divide and transiently increase their mutation rates to foster diversity, a process known as adaptive mutability. Analogously, tumor cells exposed to drugs enter a persister phenotype and can reduce DNA replication fidelity, which in turn fosters genetic diversity. The implications of adaptive evolution are of relevance to understand resistance to anticancer therapies.


Assuntos
Bactérias , Evolução Biológica , Homeostase , Neoplasias , Humanos
8.
Clin Cancer Res ; 26(6): 1372-1384, 2020 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-31831554

RESUMO

PURPOSE: Defects in the homologous recombination (HR) repair pathway are of clinical interest due to sensitivity of HR-deficient cells to PARP inhibitors. We were interested in defining PARP vulnerability in patients with metastatic colorectal cancer (mCRC) carrying KRAS and BRAF mutations who display poor prognosis, have limited therapeutic options, and represent an unmet clinical need. EXPERIMENTAL DESIGN: We tested colorectal cancer cell lines, patient-derived organoids (PDO), and patient-derived xenografts (PDX) enriched for KRAS and BRAF mutations for sensitivity to the PARP inhibitor olaparib, and the chemotherapeutic agents oxaliplatin and 5-fluorouracil (5-FU). Genomic profiles and DNA repair proficiency of colorectal cancer models were compared with pharmacologic response. RESULTS: Thirteen of 99 (around 13%) colorectal cancer cell lines were highly sensitive to clinically active concentrations of olaparib and displayed functional deficiency in HR. Response to PARP blockade was positively correlated with sensitivity to oxaliplatin in colorectal cancer cell lines as well as patient-derived organoids. Treatment of PDXs with olaparib impaired tumor growth and maintenance therapy with PARP blockade after initial oxaliplatin response delayed disease progression in mice. CONCLUSIONS: These results indicate that a colorectal cancer subset characterized by poor prognosis and limited therapeutic options is vulnerable to PARP inhibition and suggest that PDO-based drug-screening assays can be used to identify patients with colorectal cancer likely to benefit from olaparib. As patients with mCRC almost invariably receive therapies based on oxaliplatin, "maintenance" treatment with PARP inhibitors warrants further clinical investigation.


Assuntos
Neoplasias Colorretais/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica , Oxaliplatina/farmacologia , Ftalazinas/farmacologia , Piperazinas/farmacologia , Reparo de DNA por Recombinação , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Mutação , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Science ; 366(6472): 1473-1480, 2019 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-31699882

RESUMO

The emergence of drug resistance limits the efficacy of targeted therapies in human tumors. The prevalent view is that resistance is a fait accompli: when treatment is initiated, cancers already contain drug-resistant mutant cells. Bacteria exposed to antibiotics transiently increase their mutation rates (adaptive mutability), thus improving the likelihood of survival. We investigated whether human colorectal cancer (CRC) cells likewise exploit adaptive mutability to evade therapeutic pressure. We found that epidermal growth factor receptor (EGFR)/BRAF inhibition down-regulates mismatch repair (MMR) and homologous recombination DNA-repair genes and concomitantly up-regulates error-prone polymerases in drug-tolerant (persister) cells. MMR proteins were also down-regulated in patient-derived xenografts and tumor specimens during therapy. EGFR/BRAF inhibition induced DNA damage, increased mutability, and triggered microsatellite instability. Thus, like unicellular organisms, tumor cells evade therapeutic pressures by enhancing mutability.


Assuntos
Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Reparo de Erro de Pareamento de DNA/genética , Resistencia a Medicamentos Antineoplásicos/genética , Receptores ErbB/antagonistas & inibidores , Terapia de Alvo Molecular , Mutagênese , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Adaptação Biológica/genética , Regulação para Baixo , Humanos , Seleção Genética
10.
Cancer Cell ; 34(1): 148-162.e7, 2018 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-29990497

RESUMO

Targeting HER2 is effective in 24% of ERBB2 amplified metastatic colorectal cancer; however, secondary resistance occurs in most of the cases. We studied the evolution of individual metastases during treatment to discover spatially resolved determinants of resistance. Circulating tumor DNA (ctDNA) analysis identified alterations associated with resistance in the majority of refractory patients. ctDNA profiles and lesion-specific radiographic reports revealed organ- or metastasis-private evolutionary patterns. When radiologic assessments documented progressive disease in target lesions, response to HER2 blockade was retained in other metastases. Genomic and functional analyses on samples and cell models from eight metastases of a patient co-recruited to a postmortem study unveiled lesion-specific evolutionary trees and pharmacologic vulnerabilities. Lesion size and contribution of distinct metastases to plasma ctDNA were correlated.


Assuntos
Adenocarcinoma/tratamento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Lapatinib/administração & dosagem , Neoplasias Hepáticas/tratamento farmacológico , Inibidores de Proteínas Quinases/administração & dosagem , Receptor ErbB-2/antagonistas & inibidores , Tomografia Computadorizada por Raios X , Trastuzumab/administração & dosagem , Adenocarcinoma/diagnóstico por imagem , Adenocarcinoma/genética , Adenocarcinoma/secundário , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Classe I de Fosfatidilinositol 3-Quinases/genética , Tomada de Decisão Clínica , Neoplasias Colorretais/diagnóstico por imagem , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Análise Mutacional de DNA , Progressão da Doença , Feminino , Amplificação de Genes , Humanos , Itália , Lapatinib/efeitos adversos , Biópsia Líquida , Neoplasias Hepáticas/diagnóstico por imagem , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/secundário , Imageamento por Ressonância Magnética , Masculino , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Valor Preditivo dos Testes , Intervalo Livre de Progressão , Inibidores de Proteínas Quinases/efeitos adversos , Receptor ErbB-2/genética , Fatores de Risco , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Trastuzumab/efeitos adversos , Resultado do Tratamento , Células Tumorais Cultivadas , Proteínas ras/genética
11.
Nat Commun ; 9(1): 2287, 2018 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-29895949

RESUMO

Attempts at eradicating metastatic cancers with targeted therapies are limited by the emergence of resistant subclones bearing heterogeneous (epi)genetic changes. We used colorectal cancer (CRC) to test the hypothesis that interfering with an ancestral oncogenic event shared by all the malignant cells (such as WNT pathway alterations) could override heterogeneous mechanisms of acquired drug resistance. Here, we report that in CRC-resistant cell populations, phylogenetic analysis uncovers a complex subclonal architecture, indicating parallel evolution of multiple independent cellular lineages. Functional and pharmacological modulation of WNT signalling induces cell death in CRC preclinical models from patients that relapsed during the treatment, regardless of the drug type or resistance mechanisms. Concomitant blockade of WNT and MAPK signalling restrains the emergence of drug-resistant clones. Reliance upon the WNT-APC pathway is preserved throughout the branched genomic drift associated with emergence of treatment relapse, thus offering the possibility of a common therapeutic strategy to overcome secondary drug resistance.


Assuntos
Neoplasias Colorretais/genética , Deriva Genética , Terapia de Alvo Molecular , Mutação , Animais , Biópsia , Técnicas de Cultura de Células , Linhagem da Célula , Proliferação de Células , Neoplasias Colorretais/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Sistema de Sinalização das MAP Quinases , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Metástase Neoplásica , Recidiva Local de Neoplasia , Transplante de Neoplasias , Oncogenes , Filogenia , Transdução de Sinais , Via de Sinalização Wnt
12.
Nature ; 552(7683): 116-120, 2017 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-29186113

RESUMO

Molecular alterations in genes involved in DNA mismatch repair (MMR) promote cancer initiation and foster tumour progression. Cancers deficient in MMR frequently show favourable prognosis and indolent progression. The functional basis of the clinical outcome of patients with tumours that are deficient in MMR is not clear. Here we genetically inactivate MutL homologue 1 (MLH1) in colorectal, breast and pancreatic mouse cancer cells. The growth of MMR-deficient cells was comparable to their proficient counterparts in vitro and on transplantation in immunocompromised mice. By contrast, MMR-deficient cancer cells grew poorly when transplanted in syngeneic mice. The inactivation of MMR increased the mutational burden and led to dynamic mutational profiles, which resulted in the persistent renewal of neoantigens in vitro and in vivo, whereas MMR-proficient cells exhibited stable mutational load and neoantigen profiles over time. Immune surveillance improved when cancer cells, in which MLH1 had been inactivated, accumulated neoantigens for several generations. When restricted to a clonal population, the dynamic generation of neoantigens driven by MMR further increased immune surveillance. Inactivation of MMR, driven by acquired resistance to the clinical agent temozolomide, increased mutational load, promoted continuous renewal of neoantigens in human colorectal cancers and triggered immune surveillance in mouse models. These results suggest that targeting DNA repair processes can increase the burden of neoantigens in tumour cells; this has the potential to be exploited in therapeutic approaches.


Assuntos
Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Reparo de Erro de Pareamento de DNA/genética , Imunoterapia/métodos , Neoplasias/imunologia , Neoplasias/patologia , Animais , Anticorpos Antineoplásicos/imunologia , Anticorpos Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Proliferação de Células/genética , Feminino , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos SCID , Proteína 1 Homóloga a MutL/deficiência , Proteína 1 Homóloga a MutL/genética , Neoplasias/genética , Neoplasias/terapia , Receptores de Antígenos de Linfócitos T/imunologia , Evasão Tumoral/genética , Evasão Tumoral/imunologia
13.
Artigo em Inglês | MEDLINE | ID: mdl-28003276

RESUMO

Precision oncology relies on targeted drugs, such as kinase inhibitors, that are presently administered based on molecular profiles obtained from surgical or bioptic tissue samples. The inherent ability of human tumors to molecularly evolve in response to drug pressures represents a daunting diagnostic challenge. Circulating free DNA (cfDNA) released from primary and metastatic lesions can be used to draw molecular maps that can be continuously updated to match each tumor's evolution. We will present evidence that liquid biopsies can effectively interrogate how targeted therapies drive lesion-specific drug-resistance mechanisms. The impact of drug-induced molecular heterogeneity on subsequent lines of treatment will also be discussed.


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias/tratamento farmacológico , Neoplasias/patologia , DNA de Neoplasias/genética , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Biópsia Líquida/métodos , Mutação
14.
Cell ; 165(2): 317-30, 2016 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-27058664

RESUMO

BRAF(V600E) mutant colon cancers (CCs) have a characteristic gene expression signature that is also found in some tumors lacking this mutation. Collectively, they are referred to as "BRAF-like" tumors and represent some 20% of CCs. We used a shRNA-based genetic screen focused on genes upregulated in BRAF(V600E) CCs to identify vulnerabilities of this tumor subtype that might be exploited therapeutically. Here, we identify RANBP2 (also known as NUP358) as essential for survival of BRAF-like, but not for non-BRAF-like, CC cells. Suppression of RANBP2 results in mitotic defects only in BRAF-like CC cells, leading to cell death. Mechanistically, RANBP2 silencing reduces microtubule outgrowth from the kinetochores, thereby inducing spindle perturbations, providing an explanation for the observed mitotic defects. We find that BRAF-like CCs display far greater sensitivity to the microtubule poison vinorelbine both in vitro and in vivo, suggesting that vinorelbine is a potential tailored treatment for BRAF-like CCs.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Vimblastina/análogos & derivados , Animais , Antineoplásicos Fitogênicos/administração & dosagem , Células Cultivadas , Neoplasias do Colo/classificação , Neoplasias do Colo/tratamento farmacológico , Xenoenxertos , Humanos , Camundongos , Camundongos Nus , Microtúbulos/efeitos dos fármacos , Microtúbulos/metabolismo , Chaperonas Moleculares/genética , Transplante de Neoplasias , Complexo de Proteínas Formadoras de Poros Nucleares/genética , Proteínas Proto-Oncogênicas B-raf/genética , Vimblastina/administração & dosagem , Vimblastina/farmacologia , Vinorelbina
15.
Cancer Discov ; 6(1): 36-44, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26546295

RESUMO

UNLABELLED: Entrectinib is a first-in-class pan-TRK kinase inhibitor currently undergoing clinical testing in colorectal cancer and other tumor types. A patient with metastatic colorectal cancer harboring an LMNA-NTRK1 rearrangement displayed a remarkable response to treatment with entrectinib, which was followed by the emergence of resistance. To characterize the molecular bases of the patient's relapse, circulating tumor DNA (ctDNA) was collected longitudinally during treatment, and a tissue biopsy, obtained before entrectinib treatment, was transplanted in mice (xenopatient), which then received the same entrectinib regimen until resistance developed. Genetic profiling of ctDNA and xenopatient samples showed acquisition of two point mutations in the catalytic domain of NTRK1, p.G595R and p.G667C. Biochemical and pharmacologic analysis in multiple preclinical models confirmed that either mutation renders the TRKA kinase insensitive to entrectinib. These findings can be immediately exploited to design next-generation TRKA inhibitors. SIGNIFICANCE: We provide proof of principle that analyses of xenopatients (avatar) and liquid biopsies allow the identification of drug resistance mechanisms in parallel with clinical treatment of an individual patient. We describe for the first time that p.G595R and p.G667C TRKA mutations drive acquired resistance to entrectinib in colorectal cancers carrying NTRK1 rearrangements.


Assuntos
Benzamidas/administração & dosagem , Neoplasias Colorretais/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Indazóis/administração & dosagem , Inibidores de Proteínas Quinases/administração & dosagem , Receptor trkA/genética , Animais , Domínio Catalítico , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Rearranjo Gênico , Humanos , Camundongos , Mutação , Transplante de Neoplasias , Células Neoplásicas Circulantes/patologia , Receptor trkA/química
16.
Cancer Discov ; 6(2): 147-153, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26644315

RESUMO

UNLABELLED: How genomic heterogeneity associated with acquired resistance to targeted agents affects response to subsequent therapy is unknown. We studied EGFR blockade in colorectal cancer to assess whether tissue and liquid biopsies can be integrated with radiologic imaging to monitor the impact of individual oncogenic alterations on lesion-specific responses. Biopsy of a patient's progressing liver metastasis following prolonged response to cetuximab revealed a MEK1(K57T) mutation as a novel mechanism of acquired resistance. This lesion regressed upon treatment with panitumumab and the MEK inhibitor trametinib. In circulating tumor DNA (ctDNA), mutant MEK1 levels declined with treatment, but a previously unrecognized KRAS(Q61H) mutation was also identified that increased despite therapy. This same KRAS mutation was later found in a separate nonresponding metastasis. In summary, parallel analyses of tumor biopsies and serial ctDNA monitoring show that lesion-specific radiographic responses to subsequent targeted therapies can be driven by distinct resistance mechanisms arising within separate tumor lesions in the same patient. SIGNIFICANCE: Molecular heterogeneity ensuing from acquired resistance drives lesion-specific responses to subsequent targeted therapies. Analysis of a single-lesion biopsy is inadequate to guide selection of subsequent targeted therapies. ctDNA profiles allow the detection of concomitant resistance mechanisms residing in separate metastases and assessment of the effect of therapies designed to overcome resistance.


Assuntos
Neoplasias Colorretais/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Neoplasias Hepáticas/tratamento farmacológico , MAP Quinase Quinase 1/genética , Mutação , Proteínas Proto-Oncogênicas p21(ras)/genética , Anticorpos Monoclonais/uso terapêutico , Linhagem Celular Tumoral , Cetuximab/uso terapêutico , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , DNA de Neoplasias/sangue , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/secundário , Terapia de Alvo Molecular , Panitumumabe , Medicina de Precisão , Resultado do Tratamento
17.
Nat Commun ; 6: 7002, 2015 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-25926053

RESUMO

The development of molecularly targeted anticancer agents relies on large panels of tumour-specific preclinical models closely recapitulating the molecular heterogeneity observed in patients. Here we describe the mutational and gene expression analyses of 151 colorectal cancer (CRC) cell lines. We find that the whole spectrum of CRC molecular and transcriptional subtypes, previously defined in patients, is represented in this cell line compendium. Transcriptional outlier analysis identifies RAS/BRAF wild-type cells, resistant to EGFR blockade, functionally and pharmacologically addicted to kinase genes including ALK, FGFR2, NTRK1/2 and RET. The same genes are present as expression outliers in CRC patient samples. Genomic rearrangements (translocations) involving the ALK and NTRK1 genes are associated with the overexpression of the corresponding proteins in CRC specimens. The approach described here can be used to pinpoint CRCs with exquisite dependencies to individual kinases for which clinically approved drugs are already available.


Assuntos
Neoplasias Colorretais/enzimologia , Receptores ErbB/antagonistas & inibidores , Receptores Proteína Tirosina Quinases/metabolismo , Quinase do Linfoma Anaplásico , Linhagem Celular Tumoral , Cetuximab , Neoplasias Colorretais/genética , Genes erbB-1 , Heterogeneidade Genética , Humanos , Terapia de Alvo Molecular , Proteínas Proto-Oncogênicas c-ret/metabolismo , Receptores Proteína Tirosina Quinases/genética , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo
18.
Clin Cancer Res ; 21(9): 2157-66, 2015 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-25623215

RESUMO

PURPOSE: Patients with colorectal cancer who respond to the anti-EGFR antibody cetuximab often develop resistance within several months of initiating therapy. To design new lines of treatment, the molecular landscape of resistant tumors must be ascertained. We investigated the role of mutations in the EGFR signaling axis on the acquisition of resistance to cetuximab in patients and cellular models. EXPERIMENTAL DESIGN: Tissue samples were obtained from 37 patients with colorectal cancer who became refractory to cetuximab. Colorectal cancer cells sensitive to cetuximab were treated until resistant derivatives emerged. Mutational profiling of biopsies and cell lines was performed. Structural modeling and functional analyses were performed to causally associate the alleles to resistance. RESULTS: The genetic profile of tumor specimens obtained after cetuximab treatment revealed the emergence of a complex pattern of mutations in EGFR, KRAS, NRAS, BRAF, and PIK3CA genes, including two novel EGFR ectodomain mutations (R451C and K467T). Mutational profiling of cetuximab-resistant cells recapitulated the molecular landscape observed in clinical samples and revealed three additional EGFR alleles: S464L, G465R, and I491M. Structurally, these mutations are located in the cetuximab-binding region, except for the R451C mutant. Functionally, EGFR ectodomain mutations prevent binding to cetuximab but a subset is permissive for interaction with panitumumab. CONCLUSIONS: Colorectal tumors evade EGFR blockade by constitutive activation of downstream signaling effectors and through mutations affecting receptor-antibody binding. Both mechanisms of resistance may occur concomitantly. Our data have implications for designing additional lines of therapy for patients with colorectal cancer who relapse upon treatment with anti-EGFR antibodies.


Assuntos
Neoplasias Colorretais/genética , Resistencia a Medicamentos Antineoplásicos/genética , Genes erbB-1/genética , Mutação , Antineoplásicos/uso terapêutico , Western Blotting , Linhagem Celular Tumoral , Cetuximab/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Análise Mutacional de DNA , Espaço Extracelular/genética , Citometria de Fluxo , Humanos , Reação em Cadeia da Polimerase em Tempo Real
19.
Cell Rep ; 8(5): 1475-83, 2014 Sep 11.
Artigo em Inglês | MEDLINE | ID: mdl-25199829

RESUMO

KRAS is the most frequently mutated oncogene in human cancer, yet no therapies are available to treat KRAS mutant cancers. We used two independent reverse genetic approaches to identify components of the RAS-signaling pathways required for growth of KRAS mutant tumors. Small interfering RNA (siRNA) screening of 37 KRAS mutant colorectal cancer cell lines showed that RAF1 suppression was synthetic lethal with MEK inhibition. An unbiased kinome short hairpin RNA (shRNA)-based screen confirmed this synthetic lethal interaction in colorectal as well as in lung cancer cells bearing KRAS mutations. Compounds targeting RAF kinases can reverse resistance to the MEK inhibitor selumetinib. MEK inhibition induces RAS activation and BRAF-RAF1 dimerization and sustains MEK-ERK signaling, which is responsible for intrinsic resistance to selumetinib. Prolonged dual blockade of RAF and MEK leads to persistent ERK suppression and efficiently induces apoptosis. Our data underlie the relevance of developing combinatorial regimens of drugs targeting the RAF-MEK pathway in KRAS mutant tumors.


Assuntos
Neoplasias Colorretais/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Mutação , Proteínas Proto-Oncogênicas/genética , Quinases raf/metabolismo , Proteínas ras/genética , Antineoplásicos/farmacologia , Apoptose , Benzimidazóis/farmacologia , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Humanos , Sistema de Sinalização das MAP Quinases , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Inibidores de Proteínas Quinases/farmacologia , Multimerização Proteica , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas p21(ras) , Quinases raf/antagonistas & inibidores , Quinases raf/genética , Proteínas ras/metabolismo
20.
Sci Transl Med ; 6(224): 224ra26, 2014 Feb 19.
Artigo em Inglês | MEDLINE | ID: mdl-24553387

RESUMO

Colorectal cancers (CRCs) that are sensitive to the anti-epidermal growth factor receptor (EGFR) antibodies cetuximab or panitumumab almost always develop resistance within several months of initiating therapy. We report the emergence of polyclonal KRAS, NRAS, and BRAF mutations in CRC cells with acquired resistance to EGFR blockade. Regardless of the genetic alterations, resistant cells consistently displayed mitogen-activated protein kinase kinase (MEK) and extracellular signal-regulated kinase (ERK) activation, which persisted after EGFR blockade. Inhibition of MEK1/2 alone failed to impair the growth of resistant cells in vitro and in vivo. An RNA interference screen demonstrated that suppression of EGFR, together with silencing of MEK1/2, was required to hamper the proliferation of resistant cells. Indeed, concomitant pharmacological blockade of MEK and EGFR induced prolonged ERK inhibition and severely impaired the growth of resistant tumor cells. Heterogeneous and concomitant mutations in KRAS and NRAS were also detected in plasma samples from patients who developed resistance to anti-EGFR antibodies. A mouse xenotransplant from a CRC patient who responded and subsequently relapsed upon EGFR therapy showed exquisite sensitivity to combinatorial treatment with MEK and EGFR inhibitors. Collectively, these results identify genetically distinct mechanisms that mediate secondary resistance to anti-EGFR therapies, all of which reactivate ERK signaling. These observations provide a rational strategy to overcome the multifaceted clonal heterogeneity that emerges when tumors are treated with targeted agents. We propose that MEK inhibitors, in combination with cetuximab or panitumumab, should be tested in CRC patients who become refractory to anti-EGFR therapies.


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Receptores ErbB/antagonistas & inibidores , MAP Quinase Quinase Quinases/antagonistas & inibidores , Resistencia a Medicamentos Antineoplásicos , Receptores ErbB/genética , Receptores ErbB/metabolismo , Inativação Gênica , Humanos , MAP Quinase Quinase Quinases/genética , Transdução de Sinais
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