RESUMO
DNA damage and cytoplasmic DNA induce type-1 interferon (IFN-1) and potentiate responses to immune checkpoint inhibitors. Our prior work found that inhibitors of the DNA damage response kinase ATR (ATRi) induce IFN-1 and deoxyuridine (dU) incorporation by DNA polymerases, akin to antimetabolites. Whether and how dU incorporation is required for ATRi-induced IFN-1 signaling is not known. Here, we show that ATRi-dependent IFN-1 responses require uracil DNA glycosylase (UNG)-initiated base excision repair and STING. Quantitative analyses of nine distinct nucleosides reveals that ATRi induce dU incorporation more rapidly in UNG wild-type than knockout cells, and that induction of IFN-1 is associated with futile cycles of repair. While ATRi induce similar numbers of micronuclei in UNG wild-type and knockout cells, dU containing micronuclei and cytoplasmic DNA are increased in knockout cells. Surprisingly, DNA fragments containing dU block STING-dependent induction of IFN-1, MHC-1, and PD-L1. Furthermore, UNG knockout sensitizes cells to IFN-γ in vitro , and potentiates responses to anti-PD-L1 in resistant tumors in vivo . These data demonstrate an unexpected and specific role for dU-rich DNA in suppressing STING-dependent IFN-1 responses, and show that UNG-deficient tumors have a heightened response to immune checkpoint inhibitors. STATEMENT OF SIGNIFICANCE: Antimetabolites disrupt nucleotide pools and increase dU incorporation by DNA polymerases. We show that unrepaired dU potentiates responses to checkpoint inhibitors in mouse models of cancer. Patients with low tumor UNG may respond to antimetabolites combined with checkpoint inhibitors, and patients with high tumor UNG may respond to UNG inhibitors combined with checkpoint inhibitors.
RESUMO
Inhibitors of the DNA damage signaling kinase ATR increase tumor cell killing by chemotherapies that target DNA replication forks but also kill rapidly proliferating immune cells including activated T cells. Nevertheless, ATR inhibitor (ATRi) and radiotherapy (RT) can be combined to generate CD8+ T cell-dependent antitumor responses in mouse models. To determine the optimal schedule of ATRi and RT, we determined the impact of short-course versus prolonged daily treatment with AZD6738 (ATRi) on responses to RT (days 1-2). Short-course ATRi (days 1-3) plus RT caused expansion of tumor antigen-specific, effector CD8+ T cells in the tumor-draining lymph node (DLN) at 1 week after RT. This was preceded by acute decreases in proliferating tumor-infiltrating and peripheral T cells and a rapid proliferative rebound after ATRi cessation, increased inflammatory signaling (IFN-ß, chemokines, particularly CXCL10) in tumors, and an accumulation of inflammatory cells in the DLN. In contrast, prolonged ATRi (days 1-9) prevented the expansion of tumor antigen-specific, effector CD8+ T cells in the DLN, and entirely abolished the therapeutic benefit of short-course ATRi with RT and anti-PD-L1. Our data argue that ATRi cessation is essential to allow CD8+ T cell responses to both RT and immune checkpoint inhibitors.
Assuntos
Neoplasias , Animais , Camundongos , Neoplasias/patologia , Sulfonamidas , Imunidade , Antígenos de NeoplasiasRESUMO
ATR kinase is a central regulator of the DNA damage response (DDR) and cell cycle checkpoints. ATR kinase inhibitors (ATRi's) combine with radiation to generate CD8+ T cell-dependent responses in mouse models of cancer. We show that ATRi's induce cyclin-dependent kinase 1 (CDK1)-dependent origin firing across active replicons in CD8+ T cells activated ex vivo while simultaneously decreasing the activity of rate-limiting enzymes for nucleotide biosynthesis. These pleiotropic effects of ATRi induce deoxyuridine (dU) contamination in genomic DNA, R loops, RNA-DNA polymerase collisions, and interferon-α/ß (IFN-α/ß). Remarkably, thymidine rescues ATRi-induced dU contamination and partially rescues death and IFN-α/ß expression in proliferating CD8+ T cells. Thymidine also partially rescues ATRi-induced cancer cell death. We propose that ATRi-induced dU contamination contributes to dose-limiting leukocytopenia and inflammation in the clinic and CD8+ T cell-dependent anti-tumor responses in mouse models. We conclude that ATR is essential to limit dU contamination in genomic DNA and IFN-α/ß expression.
Assuntos
Linfócitos T CD8-Positivos , Proteína Quinase CDC2 , Animais , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Proteína Quinase CDC2/metabolismo , Morte Celular , Linhagem Celular Tumoral , DNA , Dano ao DNA , DNA Polimerase Dirigida por DNA/metabolismo , Desoxiuridina , Genômica , Interferon-alfa/metabolismo , Interferon beta , Camundongos , Nucleotídeos/metabolismo , Inibidores de Proteínas Quinases/farmacologia , RNA , Timidina/farmacologiaRESUMO
Human lung adenocarcinoma (LUAD) in current or former smokers exhibits a high tumor mutational burden (TMB) and distinct mutational signatures. Syngeneic mouse models of clinically relevant smoking-related LUAD are lacking. We established and characterized a tobacco-associated, transplantable murine LUAD cell line, designated FVBW-17, from a LUAD induced by the tobacco carcinogen 4-(methylnitrosoamino)-1-(3-pyridyl)-1-butanone in the FVB/N mouse strain. Whole-exome sequencing of FVBW-17 cells identified tobacco-associated KrasG12D and Trp53 mutations and a similar mutation profile to that of classic alkylating agents with a TMB greater than 500. FVBW-17 cells transplanted subcutaneously, via tail vein, and orthotopically generated tumors that were histologically similar to human LUAD in FVB/N mice. FVBW-17 tumors expressed programmed death ligand 1 (PD-L1), were infiltrated with CD8+ T cells, and were responsive to anti-PD-L1 therapy. FVBW-17 cells were also engineered to express green fluorescent protein and luciferase to facilitate detection and quantification of tumor growth. Distant metastases to lung, spleen, liver, and kidney were observed from subcutaneously transplanted tumors. This potentially novel cell line is a robust representation of human smoking-related LUAD biology and provides a much needed preclinical model in which to test promising new agents and combinations, including immune-based therapies.
Assuntos
Adenocarcinoma de Pulmão/induzido quimicamente , Antígeno B7-H1/metabolismo , Carcinógenos/toxicidade , Neoplasias Pulmonares/induzido quimicamente , Nitrosaminas/toxicidade , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/imunologia , Animais , Antígeno B7-H1/antagonistas & inibidores , Linhagem Celular Tumoral , Feminino , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/imunologia , Masculino , Camundongos , Mutação , Neoplasias Experimentais/induzido quimicamente , Neoplasias Experimentais/genética , Neoplasias Experimentais/imunologia , Fumaça/efeitos adversos , Nicotiana/toxicidade , Microambiente Tumoral/genética , Microambiente Tumoral/imunologiaRESUMO
Multiple studies have associated the transcription factor IRF1 with tumor-suppressive activities. Here, we report an opposite tumor cell-intrinsic function of IRF1 in promoting tumor growth. IRF1-deficient tumor cells showed reduced tumor growth in MC38 and CT26 colon carcinoma and B16 melanoma mouse models. This reduction in tumor growth was dependent on host CD8+ T cells. Detailed profiling of tumor-infiltrating leukocytes did not show changes in the various T-cell and myeloid cell populations. However, CD8+ T cells that had infiltrated IRF1-deficieint tumors in vivo exhibited enhanced cytotoxicity. IRF1-deficient tumor cells lost the ability to upregulate PD-L1 expression in vitro and in vivo and were more susceptible to T-cell-mediated killing. Induced expression of PD-L1 in IRF1-deficient tumor cells restored tumor growth. These results indicate differential activity of IRF1 in tumor escape.
Assuntos
Antígeno B7-H1/genética , Regulação Neoplásica da Expressão Gênica , Imunomodulação , Fator Regulador 1 de Interferon/metabolismo , Animais , Antígeno B7-H1/metabolismo , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Humanos , Memória Imunológica , Imunomodulação/genética , Fator Regulador 1 de Interferon/genética , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Melanoma Experimental , Camundongos , Camundongos Knockout , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
DNA-damaging chemotherapy and radiation therapy are integrated into the treatment paradigm of the majority of cancer patients. Recently, immunotherapy that targets the immunosuppressive interaction between programmed death 1 (PD-1) and its ligand PD-L1 has been approved for malignancies including non-small cell lung cancer, melanoma, and head and neck squamous cell carcinoma. ATR is a DNA damage-signaling kinase activated at damaged replication forks, and ATR kinase inhibitors potentiate the cytotoxicity of DNA-damaging chemotherapies. We show here that the ATR kinase inhibitor AZD6738 combines with conformal radiation therapy to attenuate radiation-induced CD8+ T cell exhaustion and potentiate CD8+ T cell activity in mouse models of Kras-mutant cancer. Mechanistically, AZD6738 blocks radiation-induced PD-L1 upregulation on tumor cells and dramatically decreases the number of tumor-infiltrating Tregs. Remarkably, AZD6738 combines with conformal radiation therapy to generate immunologic memory in complete responder mice. Our work raises the possibility that a single pharmacologic agent may enhance the cytotoxic effects of radiation while concurrently potentiating radiation-induced antitumor immune responses.
Assuntos
Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/efeitos da radiação , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/radioterapia , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/farmacologia , Sulfóxidos/farmacologia , Adenocarcinoma de Pulmão/tratamento farmacológico , Adenocarcinoma de Pulmão/imunologia , Adenocarcinoma de Pulmão/radioterapia , Animais , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Antígeno B7-H1/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Quimiorradioterapia , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/radioterapia , Humanos , Indóis , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/efeitos da radiação , Camundongos , Camundongos Endogâmicos BALB C , Morfolinas , Neoplasias Experimentais/imunologia , Inibidores de Proteínas Quinases/farmacocinética , Proteínas Proto-Oncogênicas p21(ras)/genética , Pirimidinas/farmacocinética , Radioterapia Conformacional , Sulfonamidas , Sulfóxidos/farmacocinéticaRESUMO
We show that ATM kinase inhibition using AZ31 prior to 9 or 9.25 Gy total body irradiation (TBI) reduced median time to moribund in mice to 8 days. ATR kinase inhibition using AZD6738 prior to TBI did not reduce median time to moribund. The striking finding associated with ATM inhibition prior to TBI was increased crypt loss within the intestine epithelium. ATM inhibition reduced upregulation of p21, an inhibitor of cyclin-dependent kinases, and blocked G1 arrest after TBI thereby increasing the number of S phase cells in crypts in wild-type but not Cdkn1a(p21CIP/WAF1)-/- mice. In contrast, ATR inhibition increased upregulation of p21 after TBI. Thus, ATM activity is essential for p21-dependent arrest while ATR inhibition may potentiate arrest in crypt cells after TBI. Nevertheless, ATM inhibition reduced median time to moribund in Cdkn1a(p21CIP/WAF1)-/- mice after TBI. ATM inhibition also increased cell death in crypts at 4 h in Cdkn1a(p21CIP/WAF1)-/-, earlier than at 24 h in wild-type mice after TBI. In contrast, ATR inhibition decreased cell death in crypts in Cdkn1a(p21CIP/WAF1)-/- mice at 4 h after TBI. We conclude that ATM activity is essential for p21-dependent and p21-independent mechanisms that radioprotect intestinal crypts and that ATM inhibition promotes GI syndrome after TBI.
Assuntos
Síndrome Aguda da Radiação/tratamento farmacológico , Fase G1/efeitos dos fármacos , Gastroenteropatias/tratamento farmacológico , Mucosa Intestinal/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Lesões Experimentais por Radiação/tratamento farmacológico , Protetores contra Radiação/farmacologia , Animais , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Feminino , Raios gama/efeitos adversos , Indóis , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efeitos da radiação , Camundongos , Camundongos Endogâmicos C57BL , Morfolinas , Inibidores de Proteínas Quinases/farmacocinética , Inibidores de Proteínas Quinases/uso terapêutico , Pirimidinas/farmacocinética , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Quinolinas/farmacocinética , Quinolinas/farmacologia , Quinolinas/uso terapêutico , Protetores contra Radiação/farmacocinética , Protetores contra Radiação/uso terapêutico , Sulfonamidas , Sulfóxidos/farmacocinética , Sulfóxidos/farmacologia , Sulfóxidos/uso terapêuticoRESUMO
The ATM kinase inhibitor AZ31 and ATR kinase inhibitor AZD6738 are in various phases of preclinical and clinical evaluation for their ability to potentiate chemoradiation. To support the preclinical evaluation of their pharmacokinetics, we developed and validated an LC-MS/MS assay for the simultaneous quantification of AZ31 and AZD6738 in mouse plasma. A "dilute and shoot" method was used to precipitate proteins from a sample volume of 50µL. Chromatographic separation was achieved using a Phenomenex Polar-RP column and a gradient mobile phase consisting of methanol-water with 0.1% formic acid. Detection was accomplished using a Waters Quattro Micro mass spectrometer in positive ionization mode. The assay utilizing 50µL sample was linear from 10 to 5000ng/mL and determined to be both accurate (-8.2 to 8.6%) and precise (<5.4% CV) and achieved the criteria for U.S. FDA guidance for bioanalytical method validation. Quantification was achieved in mouse tissue homogenate using a separate 200µL sample preparation. This LC-MS/MS assay will be essential for determining the tissue distribution and pharmacokinetics in future mouse studies.
Assuntos
Ligas/química , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Cromatografia Líquida/métodos , Plasma/química , Pirimidinas/química , Sulfóxidos/química , Espectrometria de Massas em Tandem/métodos , Animais , Bioensaio/métodos , Estabilidade de Medicamentos , Formiatos/química , Indóis , Metanol/química , Camundongos , Morfolinas , Pirimidinas/sangue , Reprodutibilidade dos Testes , Sulfonamidas , Sulfóxidos/sangueRESUMO
ATR and ATM are DNA damage signaling kinases that phosphorylate several thousand substrates. ATR kinase activity is increased at damaged replication forks and resected DNA double-strand breaks (DSBs). ATM kinase activity is increased at DSBs. ATM has been widely studied since ataxia telangiectasia individuals who express no ATM protein are the most radiosensitive patients identified. Since ATM is not an essential protein, it is widely believed that ATM kinase inhibitors will be well-tolerated in the clinic. ATR has been widely studied, but advances have been complicated by the finding that ATR is an essential protein and it is widely believed that ATR kinase inhibitors will be toxic in the clinic. We describe AZD6738, an orally active and bioavailable ATR kinase inhibitor. AZD6738 induces cell death and senescence in non-small cell lung cancer (NSCLC) cell lines. AZD6738 potentiates the cytotoxicity of cisplatin and gemcitabine in NSCLC cell lines with intact ATM kinase signaling, and potently synergizes with cisplatin in ATM-deficient NSCLC cells. In contrast to expectations, daily administration of AZD6738 and ATR kinase inhibition for 14 consecutive days is tolerated in mice and enhances the therapeutic efficacy of cisplatin in xenograft models. Remarkably, the combination of cisplatin and AZD6738 resolves ATM-deficient lung cancer xenografts.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Proteínas Mutadas de Ataxia Telangiectasia/deficiência , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Cisplatino/administração & dosagem , Neoplasias Pulmonares/tratamento farmacológico , Inibidores de Proteínas Quinases/administração & dosagem , Pirimidinas/administração & dosagem , Sulfóxidos/administração & dosagem , Administração Oral , Animais , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Disponibilidade Biológica , Carcinoma Pulmonar de Células não Pequenas/enzimologia , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Senescência Celular/efeitos dos fármacos , Desoxicitidina/administração & dosagem , Desoxicitidina/análogos & derivados , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Feminino , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Humanos , Indóis , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Camundongos Nus , Morfolinas , Inibidores de Proteínas Quinases/farmacocinética , Pirimidinas/farmacocinética , Interferência de RNA , Sulfonamidas , Sulfóxidos/farmacocinética , Transfecção , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , GencitabinaRESUMO
Recent clinical data in lung cancer suggests that epigenetically targeted therapy may selectively enhance chemotherapeutic sensitivity. There have been few if any studies rigorously evaluating this hypothesized priming effect. Here we describe a series of investigations testing whether epigenetic priming with azacitidine and entinostat increases sensitivity of NSCLC to cytotoxic agents. We noted no differences in chemosensitivity following treatment with epigenetic therapy in in vitro assays of viability and colony growth. Using cell line and patient derived xenograft (PDX) models, we also observed no change in responsiveness to cisplatin in vivo. In select models, we noted differential responses to irinotecan treatment in vivo. In vitro epigenetic therapy prior to tumor implantation abrogated response of H460 xenografts to irinotecan. Conversely, in vitro epigenetic therapy appeared to sensitize A549 xenografts (tumor growth inhibition 51%, vs. 22% in mock-pretreated control). In vivo epigenetic therapy enhanced the response of adenocarcinoma PDX to irinotecan. Taken together, these data do not support broadly applicable epigenetic priming in NSCLC. Priming effects may be context-specific, dependent on both tumor and host factors. Further preclinical study is necessary to determine whether, and in which contexts, priming with epigenetic therapy has potential to enhance chemotherapeutic efficacy in NSCLC patients.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Azacitidina/farmacologia , Benzamidas/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Piridinas/farmacologia , Animais , Camptotecina/administração & dosagem , Camptotecina/análogos & derivados , Linhagem Celular Tumoral , Sobrevivência Celular , Metilação de DNA , Avaliação Pré-Clínica de Medicamentos , Epigênese Genética , Feminino , Histonas/química , Humanos , Irinotecano , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Transplante de NeoplasiasRESUMO
Innovative therapies are needed for advanced Non-Small Cell Lung Cancer (NSCLC). We have undertaken a genomics based, hypothesis driving, approach to query an emerging potential that epigenetic therapy may sensitize to immune checkpoint therapy targeting PD-L1/PD-1 interaction. NSCLC cell lines were treated with the DNA hypomethylating agent azacytidine (AZA - Vidaza) and genes and pathways altered were mapped by genome-wide expression and DNA methylation analyses. AZA-induced pathways were analyzed in The Cancer Genome Atlas (TCGA) project by mapping the derived gene signatures in hundreds of lung adeno (LUAD) and squamous cell carcinoma (LUSC) samples. AZA up-regulates genes and pathways related to both innate and adaptive immunity and genes related to immune evasion in a several NSCLC lines. DNA hypermethylation and low expression of IRF7, an interferon transcription factor, tracks with this signature particularly in LUSC. In concert with these events, AZA up-regulates PD-L1 transcripts and protein, a key ligand-mediator of immune tolerance. Analysis of TCGA samples demonstrates that a significant proportion of primary NSCLC have low expression of AZA-induced immune genes, including PD-L1. We hypothesize that epigenetic therapy combined with blockade of immune checkpoints - in particular the PD-1/PD-L1 pathway - may augment response of NSCLC by shifting the balance between immune activation and immune inhibition, particularly in a subset of NSCLC with low expression of these pathways. Our studies define a biomarker strategy for response in a recently initiated trial to examine the potential of epigenetic therapy to sensitize patients with NSCLC to PD-1 immune checkpoint blockade.
Assuntos
Antimetabólitos Antineoplásicos/farmacologia , Azacitidina/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/imunologia , Imunoterapia/métodos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Antígeno B7-H1/biossíntese , Antígeno B7-H1/genética , Antígeno B7-H1/imunologia , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Metilação de DNA/efeitos dos fármacos , Epigenômica , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Receptor de Morte Celular Programada 1/imunologia , Transdução de Sinais , Regulação para Cima/efeitos dos fármacosRESUMO
INTRODUCTION: Epigenetics refers to heritable modifications of DNA and associated chromatin components that influence gene expression without altering DNA coding sequence. Epigenetic dysregulation is a central contributor to oncogenesis and is increasingly a focus of interest in cancer therapeutic research. Two key levels of aberrant epigenetic control are DNA methylation and histone acetylation. Primary regulators of these epigenetic changes include DNA methyltransferases (DNMTs) and histone deacetylases (HDACs). AREAS COVERED: This review focuses on epigenetic changes in non-small-cell lung cancer and recent preclinical and clinical studies targeting these changes. DNMT inhibitors were previously explored at or near maximally tolerated doses, levels at which these agents are cytotoxic but have suboptimal effects on DNA methylation. Use of these inhibitors at substantially lower doses, in combination with HDAC inhibitors, can promote re-expression of silenced tumor suppressor genes, can result in major clinical responses and may alter tumor responsiveness to subsequent cytotoxic therapies. EXPERT OPINION: Combinatorial epigenetic therapy has demonstrated encouraging clinical activity, but many relevant questions remain. Global strategies influencing the epigenome may have both positive and potential negative long-term effects on cancer progression. Further clinical investigation of this approach, including exploratory studies to define predictive biomarkers, is warranted.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Metilases de Modificação do DNA/antagonistas & inibidores , Epigênese Genética , Inibidores de Histona Desacetilases/uso terapêutico , Neoplasias Pulmonares/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/enzimologia , Resistencia a Medicamentos Antineoplásicos , Humanos , Neoplasias Pulmonares/enzimologiaRESUMO
UNLABELLED: Epigenetic alterations are strongly associated with the development of cancer. We conducted a phase I/II trial of combined epigenetic therapy with azacitidine and entinostat, inhibitors of DNA methylation and histone deacetylation, respectively, in extensively pretreated patients with recurrent metastatic non-small cell lung cancer. This therapy is well tolerated, and objective responses were observed, including a complete response and a partial response in a patient who remains alive and without disease progression approximately 2 years after completing protocol therapy. Median survival in the entire cohort was 6.4 months (95% CI 3.8-9.2), comparing favorably with existing therapeutic options. Demethylation of a set of 4 epigenetically silenced genes known to be associated with lung cancer was detectable in serial blood samples in these patients and was associated with improved progression-free (P = 0.034) and overall survival (P = 0.035). Four of 19 patients had major objective responses to subsequent anticancer therapies given immediately after epigenetic therapy. SIGNIFICANCE: This study demonstrates that combined epigenetic therapy with low-dose azacitidine and entinostat results in objective, durable responses in patients with solid tumors and defines a blood-based biomarker that correlates with clinical benefit.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/terapia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/terapia , Idoso , Idoso de 80 Anos ou mais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacocinética , Azacitidina/administração & dosagem , Azacitidina/efeitos adversos , Benzamidas/administração & dosagem , Benzamidas/efeitos adversos , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Estudos de Coortes , Terapia Combinada , Metilação de DNA/efeitos dos fármacos , Progressão da Doença , Intervalo Livre de Doença , Epigênese Genética , Feminino , Terapia Genética , Histonas/metabolismo , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Masculino , Pessoa de Meia-Idade , Piridinas/administração & dosagem , Piridinas/efeitos adversosRESUMO
Shroom is an actin-associated determinant of cell morphology that is required for neural tube closure in both mice and frogs. Shroom regulates this process by causing apical constriction of epithelial cells via a pathway involving myosin II. Here we report on characterization of the Shroom-related proteins Apxl and KIAA1202 and their role in cell architecture. Shroom, Apxl, and KIAA1202 exhibit differing abilities to interact with the actin cytoskeleton. In fibroblasts, Shroom readily associates with actin stress fibers and induces bundling, Apxl is found on cortical actin, and KIAA1202 is localized to a cytoplasmic population of F-actin. In epithelial cells, Apxl and KIAA1202 do not induce apical constriction as Shroom does, but have the capacity to do so if targeted to the apical junctional complex. To determine whether the activity of Shroom-like proteins is conserved in invertebrates, we have tested the ability of the lone Shroomrelated protein in Drosophila, CG8603, to activate the constriction pathway. A chimeric protein consisting of the Shroom targeting domain and the Drosophila protein elicits constriction. Finally, we show that Apxl is involved in regulating the cytoskeletal organization and architecture of endothelial cells. We predict that the ability of Shroom-like proteins to regulate cellular morphology is conserved in evolution and is regulated in part by subcellular localization.