RESUMO
Nearly 60% of acute myeloid leukemia (AML) patients with the t(8;21)(q22;q22) translocation fail to achieve long-term disease-free survival. Our previous studies demonstrated that oridonin selectively induces apoptosis of t(8;21) leukemia cells and causes cleavage of AML1-ETO oncoprotein resulting from t(8;21), but the underlying mechanisms remain unclear. We show that oridonin interacted with glutathione and thioredoxin/thioredoxin reductase to increase intracellular reactive oxygen species, which in turn activated caspase-3 in t(8;21) cells. Moreover, oridonin bound AML1-ETO, directing the enzymatic cleavage at aspartic acid 188 via caspase-3 to generate a truncated AML1-ETO (ΔAML1-ETO) and preventing the protein from further proteolysis. ΔAML1-ETO interacted with AML1-ETO and interfered with the trans-regulatory functions of remaining AML1-ETO oncoprotein, thus acting as a tumor suppressor that mediates the anti-leukemia effect of oridonin. Furthermore, oridonin inhibited the activity of c-Kit(+) leukemia-initiating cells. Therefore, oridonin is a potential lead compound for molecular target-based therapy of leukemia.
Assuntos
Cromossomos Humanos Par 21/genética , Cromossomos Humanos Par 8/genética , Subunidade alfa 2 de Fator de Ligação ao Core/antagonistas & inibidores , Diterpenos do Tipo Caurano/farmacologia , Leucemia Mieloide Aguda/genética , Proteínas de Fusão Oncogênica/antagonistas & inibidores , Translocação Genética/efeitos dos fármacos , Proteínas Supressoras de Tumor/metabolismo , Animais , Caspase 3/metabolismo , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Ativação Enzimática/efeitos dos fármacos , Humanos , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Leucemia Mieloide Aguda/enzimologia , Camundongos , Camundongos Endogâmicos C57BL , Modelos Biológicos , Terapia de Alvo Molecular , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Proteínas de Fusão Oncogênica/genética , Estabilidade Proteica/efeitos dos fármacos , Proteína 1 Parceira de Translocação de RUNX1 , Espécies Reativas de Oxigênio/metabolismoRESUMO
The full-length AML1-ETO (AE) fusion gene resulting from t(8;21)(q22;q22) in human acute myeloid leukemia (AML) is not sufficient to induce leukemia in animals, suggesting that additional mutations are required for leukemogenesis. We and others have identified activating mutations of C-KIT in nearly half of patients with t(8;21) AML. To test the hypothesis that activating C-KIT mutations cooperate with AE to cause overt AML, we generated a murine transduction and transplantation model with both mutated C-KIT and AE. To overcome the intracellular transport block of human C-KIT in murine cells, we engineered hybrid C-KIT (HyC-KIT) by fusing the extracellular and transmembrane domains of the murine c-Kit in-frame to the intracellular signaling domain of human C-KIT. We showed that tyrosine kinase domain mutants HyC-KIT N822K and D816V, as well as juxtamembrane mutants HyC-KIT 571+14 and 557-558Del, could transform murine 32D cells to cytokine-independent growth. The protein tyrosine kinase inhibitor dasatinib inhibited the proliferation of 32D cells expressing these C-KIT mutants, with potency in the low nanomolar range. In mice, HyC-KIT N822K induced a myeloproliferative disease, whereas HyC-KIT 571+14 induces both myeloproliferative disease and lymphocytic leukemia. Interestingly, coexpression of AE and HyC-KIT N822K led to fatal AML. Our data have further enriched the two-hit model that abnormalities of both transcription factor and membrane/cytosolic signaling molecule are required in AML pathogenesis. Furthermore, dasatinib prolonged lifespan of mice bearing AE and HyC-KIT N822K-coexpressing leukemic cells and exerted synergic effects while combined with cytarabine, thus providing a potential therapeutic for t(8;21) leukemia.
Assuntos
Transformação Celular Neoplásica , Subunidade alfa 2 de Fator de Ligação ao Core , Leucemia Mieloide Aguda , Mutação , Proteínas de Fusão Oncogênica , Proteínas Proto-Oncogênicas c-kit , Animais , Antimetabólitos Antineoplásicos/farmacologia , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Transformação Celular Neoplásica/patologia , Cromossomos Humanos Par 21/genética , Cromossomos Humanos Par 21/metabolismo , Cromossomos Humanos Par 8/genética , Cromossomos Humanos Par 8/metabolismo , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Citarabina/farmacologia , Humanos , Leucemia Linfoide/genética , Leucemia Linfoide/metabolismo , Leucemia Linfoide/patologia , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos Transgênicos , Células NIH 3T3 , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas c-kit/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Proteína 1 Parceira de Translocação de RUNX1 , Translocação Genética/genéticaRESUMO
All-trans retinoic acid (ATRA)/arsenic trioxide (ATO) combination-based therapy has benefitted newly diagnosed acute promyelocytic leukemia (APL) in short-term studies, but the long-term efficacy and safety remained unclear. From April 2001, we have followed 85 patients administrated ATRA/ATO with a median follow-up of 70 months. Eighty patients (94.1%) entered complete remission (CR). Kaplan-Meier estimates of the 5-year event-free survival (EFS) and overall survival (OS) for all patients were 89.2% +/- 3.4% and 91.7% +/- 3.0%, respectively, and the 5-year relapse-free survival (RFS) and OS for patients who achieved CR (n = 80) were 94.8% +/- 2.5% and 97.4% +/- 1.8%, respectively. Upon ATRA/ATO, prognosis was not influenced by initial white blood cell count, distinct PML-RARalpha types, or FLT3 mutations. The toxicity profile was mild and reversible. No secondary carcinoma was observed, and 24 months after the last dose of ATRA/ATO, patients had urine arsenic concentrations well below the safety limit. These results demonstrate the high efficacy and minimal toxicity of ATRA/ATO treatment for newly diagnosed APL in long-term follow-up, suggesting a potential frontline therapy for de novo APL.