Inhibition Of T Cell Protein Tyrosine Phosphatase And Proteasome By A Binuclear Copper (Ⅱ) Complex

The T cell protein tyrosine phosphatase is involved in numerous intracellular signal transduction pathways, and is associated with the development of tumor. The ubiquitin-proteasome system is responsible for the degradation of most intracellular proteins, therefore plays an essential regulatory role in numerous cellular processes,and it is also associated with the development of tumor. Therefore, TCPTP and proteasome have emerged as therapeutic targets in cancer. In recent years, some studies have shown that copper complexes can effectively inhibit the activity of TCPTP and proteasome, copper complexes thus attract great interests in developing noval anti-cancer drug.Previously, we investigated that a binuclear copper complex, [Cu2(μ-IDA)(phen)3(NO3)]NO3·4H2O (phen=1,10-phenanthroline, H2IDA =iminodiacetic acid) (1) could potently and selectively inhibit recombinant TCPTP in vitro, and inhibit proliferation and induce apoptosis in human tumor cells. In order to research the selective inhibition of complex 1 on TCPTP in cells and the relationship between the selective inhibition of TCPTP and the inhibition of proliferation as well as induction of apoptosis, we investigated the phosphorylation of TCPTP substrates in three human tumor cells and the inhibition of proteasome in MCF7 cells by complex 1. Main results were obtained as follow:1-, Western blot detecting the phosphorylation levels of PTP substrates indicated that complex 1 potently and selectively inhibited the activity of TCPTP in three tumor cells, resulting in the improvement of TCPTP specific substrate pSrc418 and other substrates EGFR, IR/IGF1R and IRS-1 but PTP IB specific substrate pSrc529. However, the concentration was not the same when the phosphorylation of TCPTP substrates reach to the maximum in different cells. Phosphorylation levels of TCPTP substrates in Hela cells reached to the maximum at 5μM of complex 1 while 10μM in MCF7 and HepG2 cells when they were treated for 7h. In addition complex 1 increased the expression of EGFR, IGF-1R and Src.2、The specific fluorogenic peptide substrate Suc-LLVY-AMC was used to monitor the chymotrypsin-like activity of MCF7 treated with complex 1. The result indicated that complex 1 was failed to inhibit the chymotrypsin-like activity, instead the activity of chymotrypsin-like increased. Moreover, Western blot showed that the ubiquitin-protein decreased, suggesting the increase in the activity of chymotrypsin-like. However, the extracellular chymotrypsin-like activity was not obviously changed when cellular extract was treated directly with complex 1. These results suggest that the promotion of intracellular proteasome activity by complex 1 is not due to the direct activation of complex 1 to chymotrypsin-like activity,maybe result from inhibition of TCPTP, which influence cellular phosphorylation and the expression of proteasome.3、A binuclear zinc complex was synthesized and crystallized. The complex could efficiently inhibit protein tyrosine phosphatase 1B.