A canthin-6-one derivative induces cell death by apoptosis/necroptosis-like with DNA damage in acute myeloid cells
Heron F V Torquato1, Manoel Trindade Rodrigues Junior2, Cauê Santos Lima3
1Departamento de Bioquímica, Universidade Federal de São Paulo, R. Três de Maio 100, 04044-020 São Paulo, SP, Brazil; Faculdade de Farmácia, Centro Universitário Braz Cubas, 08773-380 Mogi das Cruzes, SP, Brazil; Faculdade de Ciências Farmacêuticas, Alimentos e Nutrição, Universidade Federal de Mato Grosso do Sul, 79070-900 Campo Grande, MS, Brazil.
Abstract:
Natural products have long been considered a relevant source of new antitumor agents. Despite advances in the treatment of younger patients with acute myeloid leukemia (AML), the prognosis of elderly patients remains poor, with a high frequency of relapse. The cytotoxicity of canthin-6-one alkaloids has been extensively studied in different cell types, including leukemic strains. Among the canthin-6-one analogs tested, 10-methoxycanthin-6-one (Mtx-C) showed the highest cytotoxicity in the malignant AML cells Kasumi-1 and KG-1. Thus, we evaluated the cytotoxicity and cell death mechanisms related to Mtx-C using the EC50 (80 µM for Kasumi-1 and 36 µM for KG-1) treatment for 24 h. Our results identify reactive oxygen species production, mitochondrial depolarization, annexin V-FITC/7-AAD double staining, caspase cleave and upregulation of mitochondria-dependent apoptosis proteins (Bax, Bim, Bik, Puma and phosphorylation of p53) for both cell lineages. However, downregulation of Bcl-2 and the simultaneous execution of the apoptotic and necroptotic programs associated with the phosphorylation of the proteins receptor-interacting serine/threonine-protein kinase 3 and mixed lineage kinase domain-like pseudokinase occurred only in Kasumi-1 cells. About the lasted events, Kasumi-1 cell death was inhibited by pharmacological agents such as Zvad-FMK and necrostatin-1. The underlying molecular mechanisms of Mtx-C still include participation in the DNA damage and stress-signaling pathways involving p38 and c-Jun N-terminal mitogen-activated protein kinases and interaction with DNA. Thus, Mtx-C represents a promising tool for the development of new antileukemic molecules.
Insights
10-methoxycanthin-6-one (Mtx-C) demonstrates potent anti-leukemic activity against acute myeloid leukemia (AML) cells by inducing apoptosis and necroptosis. This natural product shows promise for developing novel antileukemic therapies.
Area of Science:
- Natural product chemistry
- Cancer biology
- Pharmacology
Background:
- Natural products are vital sources for novel antitumor agents.
- Elderly acute myeloid leukemia (AML) patients face poor prognoses and high relapse rates.
- Canthin-6-one alkaloids exhibit cytotoxicity against various cell types, including leukemia.
Purpose of the Study:
- To evaluate the cytotoxicity and cell death mechanisms of 10-methoxycanthin-6-one (Mtx-C) in AML cell lines.
- To identify specific pathways and molecular targets affected by Mtx-C treatment.
- To assess the potential of Mtx-C as a novel antileukemic therapeutic agent.
Main Methods:
- Cytotoxicity assays using Kasumi-1 and KG-1 AML cell lines treated with Mtx-C at EC50 concentrations.
- Analysis of reactive oxygen species (ROS) production and mitochondrial membrane potential.
- Flow cytometry with Annexin V-FITC/7-AAD staining to detect apoptosis.
- Western blot analysis for caspase cleavage and apoptosis/necroptosis-related protein expression.
- Inhibition studies using Zvad-FMK and necrostatin-1 to elucidate cell death pathways.
Main Results:
- Mtx-C exhibited significant cytotoxicity against Kasumi-1 and KG-1 cells.
- Apoptosis induction was observed in both cell lines, characterized by ROS production, mitochondrial depolarization, caspase activation, and upregulation of pro-apoptotic proteins (Bax, Bim, Bik, Puma) and p53 phosphorylation.
- Kasumi-1 cells uniquely displayed Bcl-2 downregulation and simultaneous apoptosis and necroptosis execution, involving RIPK3 and MLKL phosphorylation.
- Mtx-C treatment involved DNA damage, stress-signaling pathways (p38, JNK), and direct DNA interaction.
Conclusions:
- Mtx-C effectively induces cell death in AML cells through multiple mechanisms, including apoptosis and necroptosis.
- The compound's ability to trigger distinct cell death programs in different AML cell lines highlights its complex mode of action.
- Mtx-C represents a promising candidate for the development of new antileukemic drugs, particularly for patients with poor prognoses.
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