RESUMO
Checkpoint kinase 1 inhibitors (CHK1i) have shown impressive single-agent efficacy in treatment of certain tumors, as monotherapy or potentiators of chemotherapy in clinical trials, but the sensitive tumor types and downstream effectors to dictate the therapeutic responses to CHK1i remains unclear. In this study we first analyzed GDSC (Genomics of Drug Sensitivity in Cancer) and DepMap database and disclosed that hematologic malignancies (HMs) were relatively sensitive to CHK1i or CHK1 knockdown. This notion was confirmed by examining PY34, a new and potent in-house selective CHK1i, which exhibited potent anti-HM effect in vitro and in vivo, as single agent. We demonstrated that the downregulation of c-Myc and its signaling pathway was the common transcriptomic profiling response of sensitive HM cell lines to PY34, whereas overexpressing c-Myc could partially rescue the anticancer effect of PY34. Strikingly, we revealed the significant correlations between downregulation of c-Myc and cell sensitivity to PY34 in 17 HM cell lines and 39 patient-derived cell (PDC) samples. Thus, our results demonstrate that HMs are more sensitive to CHK1i than solid tumors, and c-Myc downregulation could represent the CHK1i efficacy in HMs.
Assuntos
Proteínas de Ligação a DNA/antagonistas & inibidores , Regulação para Baixo/efeitos dos fármacos , Neoplasias Hematológicas/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Fatores de Transcrição/antagonistas & inibidores , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Quinase 1 do Ponto de Checagem/antagonistas & inibidores , Quinase 1 do Ponto de Checagem/deficiência , Quinase 1 do Ponto de Checagem/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Neoplasias Hematológicas/metabolismo , Neoplasias Hematológicas/patologia , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos Nus , Camundongos SCID , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Inibidores de Proteínas Quinases/química , Relação Estrutura-Atividade , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Checkpoint kinase 1 (CHK1) is critical for S-phase fidelity and preventing premature mitotic entry in the presence of DNA damage. Tumor cells have developed a strong dependence on CHK1 for survival, and hence, this kinase has developed into a promising drug target. Chk1 deficiency in mice results in blastocyst death due to G2/M checkpoint failure showing that it is an essential gene and may be difficult to target therapeutically. Here, we show that chemical inhibition of CHK1 kills murine and human hematopoietic stem and progenitor cells (HSPCs) by the induction of BCL2-regulated apoptosis. Cell death in HSPCs is independent of p53 but requires the BH3-only proteins BIM, PUMA, and NOXA. Moreover, Chk1 is essential for definitive hematopoiesis in the embryo. Noteworthy, cell death inhibition in HSPCs cannot restore blood cell formation as HSPCs lacking CHK1 accumulate DNA damage and stop dividing. Moreover, conditional deletion of Chk1 in hematopoietic cells of adult mice selects for blood cells retaining CHK1, suggesting an essential role in maintaining functional hematopoiesis. Our findings establish a previously unrecognized role for CHK1 in establishing and maintaining hematopoiesis.
Assuntos
Apoptose/genética , Células da Medula Óssea/metabolismo , Quinase 1 do Ponto de Checagem/genética , Hematopoese/genética , Células-Tronco Hematopoéticas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Animais , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Proteína 11 Semelhante a Bcl-2/genética , Proteína 11 Semelhante a Bcl-2/metabolismo , Benzodiazepinonas/farmacologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/patologia , Linhagem Celular Tumoral , Quinase 1 do Ponto de Checagem/antagonistas & inibidores , Quinase 1 do Ponto de Checagem/deficiência , Embrião de Mamíferos , Feto , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Hematopoese/efeitos dos fármacos , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células-Tronco Hematopoéticas/patologia , Humanos , Linfoma/genética , Linfoma/metabolismo , Linfoma/patologia , Melanoma Experimental/genética , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Camundongos Knockout , Cultura Primária de Células , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Pirazóis/farmacologia , Quinolinas/farmacologia , Quinuclidinas/farmacologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismoRESUMO
Checkpoint kinase 1 (CHK1) is critical for intrinsic cell cycle control and coordination of cell cycle progression in response to DNA damage. Despite its essential function, CHK1 has been identified as a target to kill cancer cells and studies using Chk1 haploinsufficient mice initially suggested a role as tumor suppressor. Here, we report on the key role of CHK1 in normal B-cell development, lymphomagenesis and cell survival. Chemical CHK1 inhibition induces BCL2-regulated apoptosis in primary as well as malignant B-cells and CHK1 expression levels control the timing of lymphomagenesis in mice. Moreover, total ablation of Chk1 in B-cells arrests their development at the pro-B cell stage, a block that, surprisingly, cannot be overcome by inhibition of mitochondrial apoptosis, as cell cycle arrest is initiated as an alternative fate to limit the spread of damaged DNA. Our findings define CHK1 as essential in B-cell development and potent target to treat blood cancer.
Assuntos
Linfócitos B/enzimologia , Quinase 1 do Ponto de Checagem/fisiologia , Linfoma/enzimologia , Animais , Apoptose , Linfócitos B/citologia , Linfoma de Burkitt/tratamento farmacológico , Linfoma de Burkitt/enzimologia , Linfoma de Burkitt/patologia , Carcinogênese/genética , Carcinogênese/metabolismo , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Quinase 1 do Ponto de Checagem/antagonistas & inibidores , Quinase 1 do Ponto de Checagem/deficiência , Quinase 1 do Ponto de Checagem/genética , Dano ao DNA , Genes myc , Haploinsuficiência , Humanos , Linfoma/etiologia , Linfoma/genética , Linfopoese/genética , Linfopoese/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Modelos Biológicos , Leucemia-Linfoma Linfoblástico de Células Precursoras B/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras B/enzimologia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologiaRESUMO
Genomic instability is the hallmark of cancer. Checkpoint kinase-1 (Chk1) is required for cell cycle delay after DNA damage or blocked DNA replication. Chk1-depleted tumor cells undergo premature mitosis and apoptosis. Here we analyzed the depletion of Chk1 in normal somatic cells in the absence of DNA damage in order to investigate alternative cell cycle checkpoint mechanism(s). By means of adenoviruses, flow cytometry, immunofluorescence and Western blotting, Chk1-depleted mouse embryonic fibroblasts (MEFs) were investigated. Chk1-/- MEFs arrested at the S/G2 boundary of the cell cycle with decreased protein levels of many cell cycle key players. Cyclin B1 was predominantly cytoplasmic. Interestingly, overexpression of nuclear dominant Cyclin B1 leads to nuclear translocation and premature mitosis. Chk1-/- MEFs exhibited the absence of double-strand breaks, yet cells showed delayed DNA damage recovery with pan-nuclear immunostaining pattern of Histone H2AX. Activation of this checkpoint would elicit a senescent-like phenotype. Taken together, our elaborated data revealed the existence of an additional S/M checkpoint functioning via γH2AX signaling and cytoplasmic retention of Cyclin B1 in somatic cells.
