Development and Characterization of Phospha Sugar Derivatives, DMPP and TMPP, with Anti-Leukemic Effects
Authors: Nakamura Satoki , Yamashita Mitsuji , Yokota Daisuke , Hirano Isao , Ono Takaaki , Fujie Michio , Shibata Kiyoshi , Niimi Taishi , Suyama Takuya , Maddali Kasthuraiah , Asai Kazuhide , Yamashita Junko , Iguchi Yukiko , Ohnishi Kazunori 
Published: April 2020

Abstract
[Background] Sugar derivatives, whose oxygen atom in the hemiacetal ring is replaced by a carbon, nitrogen, sulfur atom, and etc., are celled as pseudo sugars. The synthesis and bioassay of these pseudo sugars are well investigated and are known as potential bioactive materials. Phospha sugar, which is one of pseudo sugars, having a phosphorus atom instead of the oxygen atom in the hemiacetal ring, is little known. The synthesis and bioassay of phospha sugars are not well studied in detail. We synthesized the 2,3,4-tribromo-3-methyl-1-phenyl phospholane 1-oxide (TMPP) and 2,3-dibromo-3-methyl-1-phenyl phospholane 1-oxide (DMPP) by the nucleophilic substitution reactions, and found their anti-leukemic activities.
[Purpose] The aims of the present study were to evaluate the inhibition of proliferation and induction of apoptosis in leukemia cell treated with TMPP or DMPP, and defines the target molecules for TMPP in leukemia cells.
[Methods] The cells used in this study were human leukemia cell lines, K562, U937, and YRK2 cells. For proliferation analysis, MTT assays were performed in leukemia cells treated with TMPP. For cell cycle analysis, flow cytometory analysis was performed in leukemia cells treated with TMPP or DMPP by PI staining. For analysis of mitotic regulatory proteins (p27, p21, Skp2, Cdc25B, Cyclin D1, Survivin, and Aurora kinase B), Western blotting was performed in leukemia cells treated with TMPP. For colony analysis, the colonies of CFU-GEMM, CFU-GM, and BFU-E were counted in AML stem/progenitor cells treated with TMPP.
[Results] In leukemia cell lines, DMPP and TMPP significantly inhibited the cell proliferation, and TMPP more strongly inhibited the cell proliferation than DMPP. 10 μM TMPP significantly induced G2/M phase arrest, and 25 μM TMPP induced apoptosis in leukemia cells. In leukemia cells, 10μM TMPP reduced protein of Aurora kinase B, Survivin, Cyclin D1, Skp2 KIS, and FoxM1, while increased protein expression of p21and p27 by western blot analysis. Moreover, 25μM TMPP activated caspase-3 and caspase-9, cleaved PARP, and reduced Bcl-2. Moreover, the treatment with TMPP decreased the counts of CFU-GEMM, CFU-GM and BFU-E by depletion of FoxM1 expression.
[Conclusions] In this study, we report in the first time the possibility of phospha sugar derivatives as anti-leukemic agents in therapy for leukemias, and analysis of their characterizations. DMPP and TMPP significantly inhibited the proliferation, and induced apoptosis of leukemia cells. DMPP and TMPP induced cell cycle arrest by suppression of FoxM1 expression and apoptosis by Bcl-2 down-regulation in leukemia cells. Moreover, TMPP inhibited colony formation of leukemia progenitors. Therefore, TMPP has new agents with anti-leukemic effects by regulation of cell cycle and apoptosis in leukemia therapy.