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1.
Cell ; 157(2): 382-394, 2014 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-24725405

RESUMO

Missense mutations in the p53 tumor suppressor inactivate its antiproliferative properties but can also promote metastasis through a gain-of-function activity. We show that sustained expression of mutant p53 is required to maintain the prometastatic phenotype of a murine model of pancreatic cancer, a highly metastatic disease that frequently displays p53 mutations. Transcriptional profiling and functional screening identified the platelet-derived growth factor receptor b (PDGFRb) as both necessary and sufficient to mediate these effects. Mutant p53 induced PDGFRb through a cell-autonomous mechanism involving inhibition of a p73/NF-Y complex that represses PDGFRb expression in p53-deficient, noninvasive cells. Blocking PDGFRb signaling by RNA interference or by small molecule inhibitors prevented pancreatic cancer cell invasion in vitro and metastasis formation in vivo. Finally, high PDGFRb expression correlates with poor disease-free survival in pancreatic, colon, and ovarian cancer patients, implicating PDGFRb as a prognostic marker and possible target for attenuating metastasis in p53 mutant tumors.


Assuntos
Carcinoma Ductal Pancreático/metabolismo , Metástase Neoplásica , Neoplasias Pancreáticas/metabolismo , Receptor beta de Fator de Crescimento Derivado de Plaquetas/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Carcinoma Ductal Pancreático/patologia , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Humanos , Camundongos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Proteína Supressora de Tumor p53/genética
2.
Cell ; 144(4): 465-6, 2011 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-21335229

RESUMO

Tumor cells frequently display an abnormal number of chromosomes, a phenomenon known as aneuploidy. Tang et al. (2011) now show that aneuploid cells are particularly sensitive to compounds that induce proteotoxic and energy stress. Could this vulnerability lead to new cancer therapies?

3.
Chembiochem ; : e202400491, 2024 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-39076125

RESUMO

The natural product FR900359 (FR) has generated significant attention lately, due to its characteristics as potent and selective inhibitor of Gq/11 mediated signal transduction of associated G protein-coupled receptors (GPCRs). This makes FR both a widely used pharmacological tool compound and a lead molecule for targeted cancer therapy. The exploration of structure-activity-relationship (SAR) of the scaffold by total synthesis has been complicated by its structural complexity and its incompatibility with standard approaches of solid-phase peptide synthesis. Options for late-stage functionalization of FR are limited due to a lack of tractable functional groups. Here we present a mixed approach combining i) genetic engineering of the FR-assembly line in Chromobacterium vaccinii, to obtain a novel FR analog featuring a primary amine, with ii) its subsequent synthetic modification and biological profiling for further SAR exploration of the FR scaffold.

4.
Biochemistry ; 62(7): 1321-1329, 2023 04 04.
Artigo em Inglês | MEDLINE | ID: mdl-36883372

RESUMO

The Myb transcription factor is involved in the proliferation of hematopoietic cells, and deregulation of its expression can lead to cancers such as leukemia. Myb interacts with various proteins, including the histone acetyltransferases p300 and CBP. Myb binds to a small domain of p300, the KIX domain (p300KIX), and inhibiting this interaction is a potential new drug discovery strategy in oncology. The available structures show that Myb binds to a very shallow pocket of the KIX domain, indicating that it might be challenging to identify inhibitors of this interaction. Here, we report the design of Myb-derived peptides which interact with p300KIX. We show that by mutating only two Myb residues that bind in or near a hotspot at the surface of p300KIX, it is possible to obtain single-digit nanomolar peptidic inhibitors of the Myb/p300KIX interaction that bind 400-fold tighter to p300KIX than wildtype Myb. These findings suggest that it might also be possible to design potent low molecular-weight compounds to disrupt the Myb/p300KIX interaction.


Assuntos
Proteína p300 Associada a E1A , Peptídeos , Proteínas Proto-Oncogênicas c-myb , Peptídeos/farmacologia , Ligação Proteica , Proteínas Proto-Oncogênicas c-myb/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-myb/química , Proteína p300 Associada a E1A/antagonistas & inibidores , Proteína p300 Associada a E1A/química
5.
Chemistry ; 28(8): e202103888, 2022 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-34878202

RESUMO

Access to the cyclic depsipeptide FR900359 (FR), a selective Gq/11 protein inhibitor of high pharmacological interest and a potential lead molecule for targeted therapy of cancers with oncogenic GNAQ or GNA11 mutations (encoding Gq and G11 respectively), has been challenging ever since its initial discovery more than three decades ago. The recent discovery of Chromobacterium vaccinii as a cultivable FR producer enables the development of approaches leading to a high-yielding, scalable and sustainable biotechnological process for production of FR, thereby removing this bottleneck. Here we characterize different promoters in exchange of the native promoter of the FR assembly line, resulting in an overexpression mutant with significantly increased production of FR. Thereby, the isolation and structure elucidation of novel FR analogs of low abundance is enabled. Further, we explore the antiproliferative activities of fifteen chromodepsins against uveal melanoma cell lines harboring Gq/11 mutations and characterize the major metabolite of FR formed in plasma.


Assuntos
Chromobacterium , Depsipeptídeos , Linhagem Celular Tumoral , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/antagonistas & inibidores , Humanos , Mutação , Regiões Promotoras Genéticas , Neoplasias Uveais
6.
Nature ; 534(7609): 647-51, 2016 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-27338794

RESUMO

Therapeutic targeting of KRAS-mutant lung adenocarcinoma represents a major goal of clinical oncology. KRAS itself has proved difficult to inhibit, and the effectiveness of agents that target key KRAS effectors has been thwarted by activation of compensatory or parallel pathways that limit their efficacy as single agents. Here we take a systematic approach towards identifying combination targets for trametinib, a MEK inhibitor approved by the US Food and Drug Administration, which acts downstream of KRAS to suppress signalling through the mitogen-activated protein kinase (MAPK) cascade. Informed by a short-hairpin RNA screen, we show that trametinib provokes a compensatory response involving the fibroblast growth factor receptor 1 (FGFR1) that leads to signalling rebound and adaptive drug resistance. As a consequence, genetic or pharmacological inhibition of FGFR1 in combination with trametinib enhances tumour cell death in vitro and in vivo. This compensatory response shows distinct specificities: it is dominated by FGFR1 in KRAS-mutant lung and pancreatic cancer cells, but is not activated or involves other mechanisms in KRAS wild-type lung and KRAS-mutant colon cancer cells. Importantly, KRAS-mutant lung cancer cells and patients' tumours treated with trametinib show an increase in FRS2 phosphorylation, a biomarker of FGFR activation; this increase is abolished by FGFR1 inhibition and correlates with sensitivity to trametinib and FGFR inhibitor combinations. These results demonstrate that FGFR1 can mediate adaptive resistance to trametinib and validate a combinatorial approach for treating KRAS-mutant lung cancer.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica , Imidazóis/uso terapêutico , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/genética , Piridazinas/uso terapêutico , Piridonas/uso terapêutico , Pirimidinonas/uso terapêutico , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/patologia , Adenocarcinoma de Pulmão , Animais , Morte Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos , Ensaios de Seleção de Medicamentos Antitumorais , Ativação Enzimática/efeitos dos fármacos , Retroalimentação Fisiológica , Feminino , Humanos , Imidazóis/farmacologia , Neoplasias Pulmonares/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Proteínas Mutantes/genética , Mutação , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Fosforilação/efeitos dos fármacos , Piridazinas/farmacologia , Piridonas/farmacologia , Pirimidinonas/farmacologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
7.
Nature ; 516(7531): 423-7, 2014 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-25337876

RESUMO

Chromosomal rearrangements have a central role in the pathogenesis of human cancers and often result in the expression of therapeutically actionable gene fusions. A recently discovered example is a fusion between the genes echinoderm microtubule-associated protein like 4 (EML4) and anaplastic lymphoma kinase (ALK), generated by an inversion on the short arm of chromosome 2: inv(2)(p21p23). The EML4-ALK oncogene is detected in a subset of human non-small cell lung cancers (NSCLC) and is clinically relevant because it confers sensitivity to ALK inhibitors. Despite their importance, modelling such genetic events in mice has proven challenging and requires complex manipulation of the germ line. Here we describe an efficient method to induce specific chromosomal rearrangements in vivo using viral-mediated delivery of the CRISPR/Cas9 system to somatic cells of adult animals. We apply it to generate a mouse model of Eml4-Alk-driven lung cancer. The resulting tumours invariably harbour the Eml4-Alk inversion, express the Eml4-Alk fusion gene, display histopathological and molecular features typical of ALK(+) human NSCLCs, and respond to treatment with ALK inhibitors. The general strategy described here substantially expands our ability to model human cancers in mice and potentially in other organisms.


Assuntos
Proteínas Associadas a CRISPR/genética , Sistemas CRISPR-Cas , Engenharia Genética/métodos , Translocação Genética/genética , Quinase do Linfoma Anaplásico , Animais , Antineoplásicos/uso terapêutico , Células Cultivadas , Inversão Cromossômica/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Crizotinibe , Modelos Animais de Doenças , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/patologia , Camundongos , Células NIH 3T3 , Inibidores de Proteínas Quinases/uso terapêutico , Pirazóis/uso terapêutico , Piridinas/uso terapêutico , Receptores Proteína Tirosina Quinases/metabolismo
8.
J Biol Chem ; 292(10): 4164-4175, 2017 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-28154181

RESUMO

KRAS is the most frequently mutated oncogene in human cancer, but its therapeutic targeting remains challenging. Here, we report a synthetic lethal screen with a library of deubiquitinases and identify USP39, which encodes an essential splicing factor, as a critical gene for the viability of KRAS-dependent cells. We show that splicing fidelity inhibitors decrease preferentially the proliferation rate of KRAS-active cells. Moreover, depletion of DHX38, encoding an USP39-interacting splicing factor, also reduces the viability of these cells. In agreement with these results, USP39 depletion caused a significant reduction in pre-mRNA splicing efficiency, as demonstrated through RNA-seq experiments. Furthermore, we show that USP39 is up-regulated in lung and colon carcinomas and its expression correlates with KRAS levels and poor clinical outcome. Accordingly, our work provides critical information for the development of splicing-directed antitumor treatments and supports the potential of USP39-targeting strategies as the basis of new anticancer therapies.


Assuntos
Neoplasias do Colo/patologia , Neoplasias Pulmonares/patologia , Mutação/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteases Específicas de Ubiquitina/metabolismo , Animais , Apoptose , Western Blotting , Proliferação de Células , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Nus , Prognóstico , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Taxa de Sobrevida , Células Tumorais Cultivadas , Proteases Específicas de Ubiquitina/genética , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Cancer Discov ; 14(2): 308-325, 2024 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-37931288

RESUMO

Lung adenocarcinoma (LUAD), commonly driven by KRAS mutations, is responsible for 7% of all cancer mortality. The first allele-specific KRAS inhibitors were recently approved in LUAD, but the clinical benefit is limited by intrinsic and acquired resistance. LUAD predominantly arises from alveolar type 2 (AT2) cells, which function as facultative alveolar stem cells by self-renewing and replacing alveolar type 1 (AT1) cells. Using genetically engineered mouse models, patient-derived xenografts, and patient samples, we found inhibition of KRAS promotes transition to a quiescent AT1-like cancer cell state in LUAD tumors. Similarly, suppressing Kras induced AT1 differentiation of wild-type AT2 cells upon lung injury. The AT1-like LUAD cells exhibited high growth and differentiation potential upon treatment cessation, whereas ablation of the AT1-like cells robustly improved treatment response to KRAS inhibitors. Our results uncover an unexpected role for KRAS in promoting intratumoral heterogeneity and suggest that targeting alveolar differentiation may augment KRAS-targeted therapies in LUAD. SIGNIFICANCE: Treatment resistance limits response to KRAS inhibitors in LUAD patients. We find LUAD residual disease following KRAS targeting is composed of AT1-like cancer cells with the capacity to reignite tumorigenesis. Targeting the AT1-like cells augments responses to KRAS inhibition, elucidating a therapeutic strategy to overcome resistance to KRAS-targeted therapy. This article is featured in Selected Articles from This Issue, p. 201.


Assuntos
Adenocarcinoma de Pulmão , Neoplasias Pulmonares , Camundongos , Animais , Humanos , Proteínas Proto-Oncogênicas p21(ras)/genética , Adenocarcinoma de Pulmão/tratamento farmacológico , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/patologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Diferenciação Celular , Células Epiteliais Alveolares/patologia
11.
Nat Cancer ; 5(3): 481-499, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38233483

RESUMO

Activating mutations in GNAQ/GNA11 occur in over 90% of uveal melanomas (UMs), the most lethal melanoma subtype; however, targeting these oncogenes has proven challenging and inhibiting their downstream effectors show limited clinical efficacy. Here, we performed genome-scale CRISPR screens along with computational analyses of cancer dependency and gene expression datasets to identify the inositol-metabolizing phosphatase INPP5A as a selective dependency in GNAQ/11-mutant UM cells in vitro and in vivo. Mutant cells intrinsically produce high levels of the second messenger inositol 1,4,5 trisphosphate (IP3) that accumulate upon suppression of INPP5A, resulting in hyperactivation of IP3-receptor signaling, increased cytosolic calcium and p53-dependent apoptosis. Finally, we show that GNAQ/11-mutant UM cells and patients' tumors exhibit elevated levels of IP4, a biomarker of enhanced IP3 production; these high levels are abolished by GNAQ/11 inhibition and correlate with sensitivity to INPP5A depletion. Our findings uncover INPP5A as a synthetic lethal vulnerability and a potential therapeutic target for GNAQ/11-mutant-driven cancers.


Assuntos
Melanoma , Humanos , Melanoma/tratamento farmacológico , Subunidades alfa de Proteínas de Ligação ao GTP/genética , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/genética , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/uso terapêutico , Mutação , Transdução de Sinais , Inositol Polifosfato 5-Fosfatases/genética
12.
Semin Cell Dev Biol ; 22(6): 572-8, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21439391

RESUMO

The Anaphase-Promoting Complex or Cyclosome (APC/C) is an E3 ubiquitin ligase whose activation requires the binding of a cofactor, either Cdc20 or Cdh1. While APC/C-Cdc20 is a major player during mitotic exit, APC/C-Cdh1 plays a central role in maintaining quiescence and controlling the onset of DNA replication. In addition, APC/C-Cdh1 is essential for endoreduplication, a process in which several rounds of DNA synthesis occur without mitosis. Recent data suggest that the APC/C is also involved in differentiation and metabolism, and plays important roles in postmitotic cells such as neurons. Thus, the APC/C is not only critical for anaphase onset but also regulates many other cellular processes during G1/S or in quiescent cells.


Assuntos
Anáfase , Caderinas/metabolismo , Proteínas de Ciclo Celular/metabolismo , Neurônios/fisiologia , Fuso Acromático/metabolismo , Complexos Ubiquitina-Proteína Ligase/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ciclossomo-Complexo Promotor de Anáfase , Animais , Antígenos CD , Caderinas/genética , Proteínas Cdc20 , Proteínas de Ciclo Celular/genética , Diferenciação Celular , Replicação do DNA , Regulação da Expressão Gênica no Desenvolvimento , Instabilidade Genômica , Humanos , Camundongos , Ligação Proteica , Fuso Acromático/genética , Complexos Ubiquitina-Proteína Ligase/genética , Ubiquitina-Proteína Ligases/genética
13.
bioRxiv ; 2023 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-37808711

RESUMO

Lung adenocarcinoma (LUAD), commonly driven by KRAS mutations, is responsible for 7% of all cancer mortality. The first allele-specific KRAS inhibitors were recently approved in LUAD, but clinical benefit is limited by intrinsic and acquired resistance. LUAD predominantly arises from alveolar type 2 (AT2) cells, which function as facultative alveolar stem cells by self-renewing and replacing alveolar type 1 (AT1) cells. Using genetically engineered mouse models, patient-derived xenografts, and patient samples we found inhibition of KRAS promotes transition to a quiescent AT1-like cancer cell state in LUAD tumors. Similarly, suppressing Kras induced AT1 differentiation of wild-type AT2 cells upon lung injury. The AT1-like LUAD cells exhibited high growth and differentiation potential upon treatment cessation, whereas ablation of the AT1-like cells robustly improved treatment response to KRAS inhibitors. Our results uncover an unexpected role for KRAS in promoting intra-tumoral heterogeneity and suggest targeting alveolar differentiation may augment KRAS-targeted therapies in LUAD. Significance: Treatment resistance limits response to KRAS inhibitors in LUAD patients. We find LUAD residual disease following KRAS targeting is composed of AT1-like cancer cells with the capacity to reignite tumorigenesis. Targeting the AT1-like cells augments responses to KRAS inhibition, elucidating a therapeutic strategy to overcome resistance to KRAS-targeted therapy.

14.
JTO Clin Res Rep ; 3(1): 100256, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34984405

RESUMO

INTRODUCTION: Somatic KRAS mutations occur in 25% of patients with NSCLC. Treatment with MEK inhibitor monotherapy has not been successful in clinical trials to date. Compensatory activation of FGFR1 was identified as a mechanism of trametinib resistance in KRAS-mutant NSCLC, and combination therapy with trametinib and ponatinib was synergistic in in vitro and in vivo models. This study sought to evaluate this drug combination in patients with KRAS-mutant NSCLC. METHODS: A phase 1 dose escalation study of trametinib and ponatinib was conducted in patients with advanced NSCLC with KRAS mutations. A standard 3-plus-3 dose escalation was done. Patients were treated with the study therapy until intolerable toxicity or disease progression. RESULTS: A total of 12 patients with KRAS-mutant NSCLC were treated (seven at trametinib 2 mg and ponatinib 15 mg, five at trametinib 2 mg and ponatinib 30 mg). Common toxicities observed were rash, diarrhea, and fever. Serious adverse events potentially related to therapy were reported in five patients, including one death in the study and four cardiovascular events. Serious events were observed at both dose levels. Of note, 75% (9 of 12) were assessable for radiographic response and no confirmed partial responses were observed. The median time on study was 43 days. CONCLUSIONS: In this phase 1 study, in patients with KRAS-mutant advanced NSCLC, combined treatment with trametinib and ponatinib was associated with cardiovascular and bleeding toxicities. Exploring the combination of MEK and FGFR1 inhibition in future studies is potentially warranted but alternative agents should be considered to improve safety and tolerability.

15.
Biochem Soc Trans ; 38(Pt 1): 65-71, 2010 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20074037

RESUMO

The APC/C (anaphase-promoting complex/cyclosome) is an E3 ubiquitin ligase that targets specific substrates for degradation by the 26S proteasome. APC/C activity depends on two cofactors, namely Cdc20 (cell division cycle 20) and Cdh1, which select the appropriate targets for ubiquitination. It is well established that APC/C is a target of the SAC (spindle assembly checkpoint) during mitosis and has critical roles in controlling the protein levels of major regulators of mitosis and DNA replication. In addition, recent studies have suggested new cell-cycle-independent functions of APC/C in non-mitotic cells and specifically in neuronal structure and function. Given the relevant functions of APC/C in cell proliferation and neuronal physiology, modulating APC/C activity may have beneficial effects in the clinic.


Assuntos
Ciclo Celular/fisiologia , Complexos Ubiquitina-Proteína Ligase/fisiologia , Ciclossomo-Complexo Promotor de Anáfase , Animais , Antígenos CD , Caderinas/metabolismo , Proteínas Cdc20 , Proteínas de Ciclo Celular/metabolismo , Quinases Ciclina-Dependentes/metabolismo , Humanos , Neurônios/citologia , Neurônios/fisiologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Fuso Acromático/metabolismo
16.
Mol Cancer Ther ; 18(7): 1323-1334, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31068384

RESUMO

FGFR1 was recently shown to be activated as part of a compensatory response to prolonged treatment with the MEK inhibitor trametinib in several KRAS-mutant lung and pancreatic cancer cell lines. We hypothesize that other receptor tyrosine kinases (RTK) are also feedback-activated in this context. Herein, we profile a large panel of KRAS-mutant cancer cell lines for the contribution of RTKs to the feedback activation of phospho-MEK following MEK inhibition, using an SHP2 inhibitor (SHP099) that blocks RAS activation mediated by multiple RTKs. We find that RTK-driven feedback activation widely exists in KRAS-mutant cancer cells, to a less extent in those harboring the G13D variant, and involves several RTKs, including EGFR, FGFR, and MET. We further demonstrate that this pathway feedback activation is mediated through mutant KRAS, at least for the G12C, G12D, and G12V variants, and wild-type KRAS can also contribute significantly to the feedback activation. Finally, SHP099 and MEK inhibitors exhibit combination benefits inhibiting KRAS-mutant cancer cell proliferation in vitro and in vivo These findings provide a rationale for exploration of combining SHP2 and MAPK pathway inhibitors for treating KRAS-mutant cancers in the clinic.


Assuntos
Acrilonitrila/análogos & derivados , Compostos de Anilina/uso terapêutico , Neoplasias/genética , Proteína Tirosina Fosfatase não Receptora Tipo 11/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Acrilonitrila/farmacologia , Acrilonitrila/uso terapêutico , Compostos de Anilina/farmacologia , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Camundongos , Camundongos Nus , Neoplasias/metabolismo , Transfecção , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Cell Death Differ ; 25(5): 828-840, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29229993

RESUMO

PP2A is a major tumor suppressor whose inactivation is frequently found in a wide spectrum of human tumors. In particular, deletion or epigenetic silencing of genes encoding the B55 family of PP2A regulatory subunits is a common feature of breast cancer cells. A key player in the regulation of PP2A/B55 phosphatase complexes is the cell cycle kinase MASTL (also known as Greatwall). During cell division, inhibition of PP2A-B55 by MASTL is required to maintain the mitotic state, whereas inactivation of MASTL and PP2A reactivation is required for mitotic exit. Despite its critical role in cell cycle progression in multiple organisms, its relevance as a therapeutic target in human cancer and its dependence of PP2A activity is mostly unknown. Here we show that MASTL overexpression predicts poor survival and shows prognostic value in breast cancer patients. MASTL knockdown or knockout using RNA interference or CRISPR/Cas9 systems impairs proliferation of a subset of breast cancer cells. The proliferative function of MASTL in these tumor cells requires its kinase activity and the presence of PP2A-B55 complexes. By using a new inducible CRISPR/Cas9 system in breast cancer cells, we show that genetic ablation of MASTL displays a significant therapeutic effect in vivo. All together, these data suggest that the PP2A inhibitory kinase MASTL may have both prognostic and therapeutic value in human breast cancer.


Assuntos
Neoplasias da Mama/enzimologia , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Proteínas Associadas aos Microtúbulos/biossíntese , Proteínas de Neoplasias/metabolismo , Proteína Fosfatase 2/metabolismo , Proteínas Serina-Treonina Quinases/biossíntese , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/mortalidade , Neoplasias da Mama/terapia , Linhagem Celular Tumoral , Feminino , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos Nus , Proteínas Associadas aos Microtúbulos/antagonistas & inibidores , Proteínas Associadas aos Microtúbulos/genética , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Proteína Fosfatase 2/genética , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética
18.
Science ; 362(6421): 1416-1422, 2018 12 21.
Artigo em Inglês | MEDLINE | ID: mdl-30573629

RESUMO

Molecularly targeted therapies aim to obstruct cell autonomous programs required for tumor growth. We show that mitogen-activated protein kinase (MAPK) and cyclin-dependent kinase 4/6 inhibitors act in combination to suppress the proliferation of KRAS-mutant lung cancer cells while simultaneously provoking a natural killer (NK) cell surveillance program leading to tumor cell death. The drug combination, but neither agent alone, promotes retinoblastoma (RB) protein-mediated cellular senescence and activation of the immunomodulatory senescence-associated secretory phenotype (SASP). SASP components tumor necrosis factor-α and intercellular adhesion molecule-1 are required for NK cell surveillance of drug-treated tumor cells, which contributes to tumor regressions and prolonged survival in a KRAS-mutant lung cancer mouse model. Therefore, molecularly targeted agents capable of inducing senescence can produce tumor control through non-cell autonomous mechanisms involving NK cell surveillance.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Citostáticos/uso terapêutico , Citotoxicidade Imunológica , Vigilância Imunológica , Células Matadoras Naturais/imunologia , Neoplasias Pulmonares/tratamento farmacológico , Aminopiridinas/farmacologia , Aminopiridinas/uso terapêutico , Animais , Apoptose , Benzimidazóis/farmacologia , Benzimidazóis/uso terapêutico , Senescência Celular , Citostáticos/farmacologia , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Quinases Ativadas por Mitógeno , Terapia de Alvo Molecular , Mutação , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Proteínas Proto-Oncogênicas p21(ras)/genética , Purinas/farmacologia , Purinas/uso terapêutico , Piridinas/farmacologia , Piridinas/uso terapêutico , Piridonas/farmacologia , Piridonas/uso terapêutico , Pirimidinonas/farmacologia , Pirimidinonas/uso terapêutico , Proteína do Retinoblastoma/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
19.
Nat Biotechnol ; 35(6): 577-582, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28459450

RESUMO

Colorectal cancer (CRC) is a leading cause of death in the developed world, yet facile preclinical models that mimic the natural stages of CRC progression are lacking. Through the orthotopic engraftment of colon organoids we describe a broadly usable immunocompetent CRC model that recapitulates the entire adenoma-adenocarcinoma-metastasis axis in vivo. The engraftment procedure takes less than 5 minutes, shows efficient tumor engraftment in two-thirds of mice, and can be achieved using organoids derived from genetically engineered mouse models (GEMMs), wild-type organoids engineered ex vivo, or from patient-derived human CRC organoids. In this model, we describe the genotype and time-dependent progression of CRCs from adenocarcinoma (6 weeks), to local disseminated disease (11-12 weeks), and spontaneous metastasis (>20 weeks). Further, we use the system to show that loss of dysregulated Wnt signaling is critical for the progression of disseminated CRCs. Thus, our approach provides a fast and flexible means to produce tailored CRC mouse models for genetic studies and pre-clinical investigation.


Assuntos
Neoplasias Colorretais/genética , Modelos Animais de Doenças , Edição de Genes/métodos , Genes Neoplásicos/genética , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/secundário , Transplante de Órgãos/métodos , Animais , Carcinogênese/genética , Linhagem Celular Tumoral , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Feminino , Masculino , Camundongos , Camundongos Transgênicos , Metástase Neoplásica
20.
Artigo em Inglês | MEDLINE | ID: mdl-28057848

RESUMO

Rapid and affordable tumor profiling has led to an explosion of genomic data that is facilitating the development of new cancer therapies. The potential of therapeutic strategies aimed at inactivating the oncogenic lesions that contribute to the aberrant survival and proliferation of tumor cells has yielded remarkable success in some malignancies such as BRAF-mutant melanoma and BCR-ABL expressing chronic myeloid leukemia. However, the direct inhibition of several well-established oncoproteins in some of these cancers is not possible or produces only transient benefits. Functional genomics represents a powerful approach for the identification of vulnerabilities linked to specific genetic alterations and has provided substantial insights into cancer signaling networks. Still, as inhibition of gene function can have diverse effects on both tumor and normal tissues, information on the potency of target inhibition on tumor growth as well as the toxic side effects of target inhibition are also needed. Here, we discuss our RNA interference (RNAi) pipeline for cancer target discovery based on our optimized short-hairpin RNA (shRNA) tools for negative selection screens and inducible RNAi platform that, in combination with embryonic stem cell (ESC)-based genetically engineered mouse models (GEMMs), enable deep in vivo target validation.


Assuntos
Modelos Animais de Doenças , Células-Tronco Embrionárias , Interferência de RNA/fisiologia , RNA Interferente Pequeno/genética , Animais , Proliferação de Células/genética , Células-Tronco Embrionárias/citologia , Humanos , Transdução de Sinais/genética
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