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Flow Cytometric Analysis of Mitochondrial Reactive Oxygen Species in Murine Hematopoietic Stem and Progenitor Cells and MLL-AF9 Driven Leukemia
Published on: September 5, 2019
Involvement of reactive oxygen species in adaphostin-induced cytotoxicity in human leukemia cells
Joya Chandra1, Jennifer Hackbarth, Son Le
1Division of Oncology Research, Guggenheim 1301, Mayo Clinic, 200 First St, SW, Rochester, MN 55901, USA.
Abstract:
Adaphostin (NSC 680410), an analog of the tyrphostin AG957, was previously shown to induce Bcr/abl down-regulation followed by loss of clonogenic survival in chronic myelogenous leukemia (CML) cell lines and clinical samples. Adaphostin demonstrated selectivity for CML myeloid progenitors in vitro and remained active in K562 cells selected for imatinib mesylate resistance. In the present study, the mechanism of action of adaphostin was investigated in greater detail in vitro. Initial studies demonstrated that adaphostin induced apoptosis in a variety of Bcr/abl- cells, including acute myelogenous leukemia (AML) blasts and cell lines as well as chronic lymphocytic leukemia (CLL) samples. Further study demonstrated that adaphostin caused intracellular peroxide production followed by DNA strand breaks and, in cells containing wild-type p53, a typical DNA damage response consisting of p53 phosphorylation and up-regulation. Importantly, the antioxidant N-acetylcysteine (NAC) blunted these events, whereas glutathione depletion with buthionine sulfoximine (BSO) augmented them. Collectively, these results not only outline a mechanism by which adaphostin can damage both myeloid and lymphoid leukemia cells, but also indicate that this novel agent might have a broader spectrum of activity than originally envisioned.
Insights
Adaphostin induces leukemia cell death by causing DNA damage and apoptosis. This novel agent shows potential for treating various leukemia types, including imatinib-resistant chronic myelogenous leukemia.
Area of Science:
- Oncology
- Molecular Biology
- Pharmacology
Background:
- Adaphostin, a tyrphostin analog, previously showed efficacy against chronic myelogenous leukemia (CML) by down-regulating Bcr/abl.
- It maintained activity in imatinib-resistant K562 cells, suggesting broader therapeutic potential.
Purpose of the Study:
- To investigate the detailed mechanism of action of adaphostin in vitro.
- To determine if adaphostin's effects extend beyond CML to other leukemia types.
Main Methods:
- In vitro studies using various leukemia cell lines and clinical samples.
- Assessment of apoptosis, intracellular peroxide production, and DNA damage response.
- Evaluation of the role of oxidative stress using N-acetylcysteine (NAC) and buthionine sulfoximine (BSO).
Main Results:
- Adaphostin induced apoptosis in Bcr/abl-negative cells, including acute myelogenous leukemia (AML) and chronic lymphocytic leukemia (CLL) samples.
- The drug triggered intracellular peroxide production and DNA strand breaks.
- A DNA damage response involving p53 phosphorylation and up-regulation was observed in wild-type p53 cells.
- Antioxidant NAC reduced adaphostin's effects, while glutathione depletion with BSO enhanced them.
Conclusions:
- Adaphostin induces leukemia cell death through oxidative stress and DNA damage pathways.
- This mechanism is effective in both myeloid and lymphoid leukemia cells.
- Adaphostin may possess a broader spectrum of anti-leukemia activity than initially anticipated.
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