Antitumor activity of a phenoxazine compound, 2-amino-4,4α-dihydro-4α,7-dimethyl-3H-phenoxazine-3-one against human B cell and T cell lymphoblastoid cell lines: induction of mixed types of cell death, apoptosis, and necrosis

摘要

Purpose: We studied the antitumor activity ofn2-amino-4,4a-dihydro-4a,7-dimethyl-3H-phenoxazine-3-none (Phx), which was synthesized by the reactions ofn2-amino-5-methylphenol with bovine hemoglobin, onnhuman B cell lymphoblastoid cell lines, P3HR-1 and Rajinderived from African Burkitt’s lymphoma, and thenhuman T cell lymphoblastoid cell line Molt-4. We alsonstudied whether Phx might cause apoptosis and necrosisnin these cells. Methods: We evaluated cell viability andnapoptosis and necrosis of the cells in the presence of Phx,nby using agarose gel electrophoresis, flow cytometry, andnfluorescence microscopy. Results: Phx suppressed thenviability of P3HR-1, Raji, and Molt-4 cells, though thensuppression patterns were different, i.e., Phx suppressednthe viability of P3HR-1, Raji, and Molt-4 cells at highernconcentrations, while the drug enhanced the viability ofnRaji cells, but not those of P3HR-1 and Molt-4 cellsnat lower concentrations. To investigate which type ofncell death– apoptosis or necrosis – is induced bynPhx, induction of DNA ladder, phosphatidylserinenexternalization, and propidium iodide-permeable cellsnwere examined in Phx-treated cells. Although Phx did notninduce DNA ladder formation, it induced the phosphatidylserinenexternalization and propidium iodidepermeablencells, suggesting that Phx caused a mixed typenof cell death, both apoptosis and necrosis. The populationnof early stage apoptotic cells was dominant in Rajincells, and that of the late stage apoptoticecrotic cellsnwas dominant in Molt-4 cells after 72-htreatment withnPhx. The population of the early stage apoptotic cells andnthe late stage apoptoticecrotic cells was almost equal innP3HR-1 cells in the presence of Phx, though the populationnof bothtypes of cells increased with time. Thennuclear morphological analysis of Phx-treated Raji,nP3HR-1, and Molt-4 cells also showed that Phx inducesnapoptosis. Conclusions: The present results suggest thatnPhx shows antitumor activity against human B cell-derivednand T cell-derived lymphoblastoid cell lines, innvitro, causing apoptosis and necrosis.