Analysis of inhibition of topoisomerase IIα and cancer cell proliferation by ingenolEZ

Abstract

We previously reported that many ingenol compounds derived from Euphorbia kansui exhibit topoisomerase inhibitory activity and ⁄ or inhibitory activity of cell proliferation. The inhibitory effects of 20-O-(2¢E,4¢Z-decadienoyl) ingenol and 3-O-(2¢E,4¢Z-decadienoyl)-ingenol among these compounds on topoisomerase II activity and on the cell proliferative activity and arrest phase of the cell cycle were studied using a mouse breast cancer (MMT) cell line. Although 20-O-ingenolEZ exerted inhibitory effects on both topoisomerase II activity and cell proliferative activity, 3-O-ingenolEZ exerted inhibitory activity on neither. The 20-O-ingenolEZinduced cell arrest of MMT-cell proliferation led to a cell cycle arrest in the G2 ⁄ M phase. Topoisomerase II inhibition can be divided into the poison and catalytic inhibitor types. A checkpoint mechanism is activated when cells are treated with these topoisomerase II inhibitors. Poison-type inhibition occurs via induction of the DNA damage checkpoint and the catalytic-type inhibition occurs via induction of the DNA-decatenation checkpoint, suggestive of distinct checkpoint reactions. 20-O-ingenolEZ inhibited topoisomerase IIa activity through inhibition of ATPase, and induced DNA-decatenation checkpoint without signaling for phosphorylation of H2AX. (Cancer Sci 2010; 101: 374-378) E uphorbia kansui has been used in herbal remedies employed for treating ascites, leukemia, and some tumors, (1,2,3) and extracts from kansui have been demonstrated to show antitumor activity.(4-6) Many ingenol derivatives from Euphoria have been reported to exhibit antiproliferative and antitumor properties (7)(8)(9) and biologically active ingenol derivatives have also been synthesized. (10) We showed in a previous study that ingenol compounds inhibited topoisomerase II activity and ⁄ or the proliferative activity of cancer cells.(1,11) In cancer chemotherapy, topoisomerase II is a major target for a variety of anticancer drugs. According to their mode of action, these drugs have been divided into two classes: anticancer topoisomerase II poisons, such as adriamycin and etoposide, which stabilize an intermediate in the topoisomerase II reaction in which two topoisomerase II subunits are covalently linked to DNA via a phosphotyrosine linkage. (12,13) This covalent intermediate, termed the cleavable complexes, induces the DNA damage checkpoint with signaling for phosphorylation of H2AX (14) and plays a critical role in the cell killing by anti-topoisomerase II agents. The agents of the second class of topoisomerase II inhibitor do not stabilize the covalent intermediate of the topoisomerase II reaction described above, but inhibit the enzyme at other points of the reaction cycle. (12,15) Since blocking of the enzyme at other points of the reaction cycle would not result in DNA damage, it is thought that this second class of agents arrests cell growth by depriving them of the essential enzymatic activity of topoisomerase II. This second class of inhibitors has been termed catalytic inhibitors to d...

Copper Complexes as Anticancer Agents Targeting Topoisomerases I and II

“…They function by enzymatic activity deprivation and cell cycle arrest in G2 through a decatenation checkpoint distinct from the DNA damage checkpoint. To delay the mitotic entry, an insufficient decatenation engages molecular components from the DDR and the spindle assembly checkpoint (SAC) (Rad9a, ATR, and BRCA1), SUMOylation and phosphorylation of Top2, the p38 and the MAPK pathways, and several decatenation checkpoint effectors but not p53 [ 66 , 132 , 133 , 134 , 135 , 136 ] ( Figure 5 A).…”

3EZ,20Ac-ingenol, a catalytic inhibitor of topoisomerases, downregulates p-Akt and induces DSBs and apoptosis of DT40 cells

“…However, we consider that DNA damage derived from inhibiting DNA decatenation through the inhibition of topo I mediated DNA cleavage by 3EZ,20Ac-ingenol during the G2 phase is the key lesion. We previously reported the effects of the catalytic topo II inhibitor 20- O -ingenolEZ on the cell cycle distributions of MMT cells by flow cytometry (Yoshida et al 2010). At 12 h, 16.5 % of the cell population was in the G2/M phase.…”

“…After 24 and 48 h of treatment, these values were 30 and 27 %, respectively, as compared with 12 % in untreated cells. MMT cells treated with 20- O -ingenolEZ were arrested in G2/M phase, but were not induced in apoptosis, whereas DT40 cells treated with the inhibiter were induced to undergo apoptosis (Yoshida et al 2010; Watanabe et al 2011). In this study, DT40 cells treated with the dual catalytic inhibitor of topo I and II 3EZ,20Ac-ingenol were also selectively arrested in G2/M phase and were induced to undergo apoptosis through signaling by cell cycle checkpoints in G2/M phase.…”

Anticancer Drugs from Traditional Toxic Chinese Medicines

“…These toxic natural compounds can induce cancer cell apoptosis through selective actions on signaling pathways. Camptothecin (Basili and Moro, ), rutaecarpine (Chan et al ., ) and yuanhuacine (Zhang et al ., ) are able to inhibit topoisomerase (Top) I, while ingenol (Yoshida et al ., ) and gambogic acid (Qin et al ., ) can suppress Top II. Vincristine (Groth‐Pedersen et al ., ), gambogic acid (Chen et al ., ) and pseudolaric acid B (Ma et al ., ), as antimitotic drugs, induce microtubule‐destabilizing.…”