Selective cytotoxicity of 3-amino-l-tyrosine correlates with peroxidase activity

Abstract

In the presence of 3-amino-L-tyrosine (3-AT), abundant brown pigment forms in human HL-60 cells, but not in a variety of other cell lines, which are reported to be lower in mean myeloperoxidase (MPO) content than HL-60. Cells were assessed for peroxidase activity with an ABTS-based colorimetric assay and compared to values obtained with known amounts of human myeloperoxidase. HL-60 cells were estimated to contain the equivalent of 37.1 ng myeloperoxidase/10(6) cells versus 26.1 and 5.0 ng/10(6) cells for human K562 and murine RAW 264.7 cell lines, respectively. HL-60 cells exhibited a nearly 60% inhibition of proliferation and > 70% reduction in cell viability after 4 d of culture in the presence of 100 microg 3-AT per ml. Higher concentrations of 3-AT (up to 400 microg/ml) for 4 d reduced HL-60 proliferation by 80% and decreased viability to 1-3%. Comparable levels of cytotoxicity were achieved in KG-1 cells after 7 d with 200 or 400 microg 3-AT per ml. K562 cells exhibited a 40% reduction in cell number after 7 d with 400 microg 3-AT per ml, but concentrations less than 400 microg/ml did not significantly affect K562 proliferation. K562 viability remained unchanged with doses of 3-AT up to 400 microg/ml. RAW 264.7 cells exhibited unchanged viability and proliferation in the presence of 3-AT at concentrations up to 400 microg 3-AT per ml. K562, KG-1, and RAW 264.7 cells exhibited no evidence of brown pigment formation in the presence of 3-AT and medium containing 10% fetal bovine serum. However, RAW 264.7 cells that were converted to protein-free medium and exposed to 3-AT exhibited intense brown pigment in some cell nuclei. A high percentage of HL-60 cells treated with 3-AT exhibited membrane blebbing, pyknosis, and nuclear fragmentation, which was not observed among other 3-AT-treated cell lines. A mechanism involving toxic intermediates of peroxidase-mediated "aminomelanin" formation is hypothesized.

Peroxidase-Mediated Bioactivation of Hydroxylated Metabolites of Carbamazepine and Phenytoin

“…It is also likely that the murine monocyte/ macrophage RAW 264.7 cells used in this study contain peroxidase (Bruno et al, 1999). The hypothesis of bioactivation by peroxidases in different organs to generate reactive metabolites is consistent with the clinical observation that many IDRs associated with CBZ and PHN involve several organs (Askmark and Wiholm, 1990).…”

The percentage of myeloperoxidase-positive blast cells is a strong independent prognostic factor in acute myeloid leukemia, even in the patients with normal karyotype

“…It was recently reported that a drug-induced cytotoxicity correlated with amount of MPO in HL-60, an AML cell line. 28 More recently, it was shown that H 2 O 2 -induced apoptosis is mediated by MPO in HL-60. 29 H 2 O 2 has been assumed to be an intracellular mediator in the toxicity of chemotherapy.…”

Protein Tyrosine Nitration in Cytokine-activated Murine Macrophages

“…6, suggesting that a peroxidase/nitrite pathway rather than peroxynitrite was responsible for tyrosine nitration in our experiments. Among several heme peroxidases, myeloperoxidase appeared to be a likely candidate catalyzing nitrite/H 2 O 2 -dependent tyrosine nitration, but it is unclear whether this enzyme occurs in RAW 264.7 macrophages (35,36). We attempted to clarify the possible involvement of myeloperoxidase in protein nitration using the fairly selective myeloperoxidase inhibitor 4-aminobenzoic acid hydrazide (32).…”