The antitumor mechanism of di-2-pyridylketone 2-pyridine carboxylic acid hydrazone and its copper complex in ROS generation and topoisomerase inhibition, and hydrazone involvement in oxygen-catalytic iron mobilization

Abstract

Iron depletion and stimulation of iron-dependent free radical damage is a rapidly developing field for chelation therapy, but the iron mobilization from ferritin by chelators has received less attention. In this study, the di-2-pyridylketone 2-pyridine carboxylic acid hydrazone (DPPCAH) and its copper complex was prepared and characterized by NMR and MS spectra. The proliferation inhibition assay showed that both DPPCAH and its copper complex exhibited selectively proliferation inhibition for HepG2 (IC50, 4.6 ± 0.2 µM for DPPACH and 1.3 ± 0.2 µM for its copper complex), but less inhibition for HCT-116 cell line (IC50, >100 µM for DPPACH and 7.8 ± 0.4 µM for its copper complex). The mechanistic studies revealed that DPPACH could remove iron from ferritin in a oxygen-catalytic manner, and contributed to redox activity of labile iron pool (LIP), that is less reported for the chelators that possess significant biological activity. The reactive oxygen species (ROS) generation and DNA cleavage assay in vitro and in vivo showed that both DPPACH-Fe(II) and DPPACH-Cu were redox-active species, indicating that ROS may mediate their antitumor activity. Further study revealed that both DPPACH and its copper complex displayed certain degree of inhibition of type II topoisomerase (Top) which contributed to their antitumor activity. Thus, the mechanism that iron mobilization by DPPACH from ferritin contributed to LIP was proposed, and both DPPACH and its copper complex were involved in ROS generation and Top II inhibition for their antitumor activities.

Calcium release induced by 2-pyridinecarboxaldehyde thiosemicarbazone and its copper complex contributes to tumor cell death

“…ROS detection. H 2 DCF-AM was converted to dichlorofluorescein (DCF) according to a previously described method (21). Briefly, 0.25 ml of 2 mM H 2 DCF-AM in absolute ethanol was added to 2.0 ml of 10 mM NaOH and allowed to stand at room temperature for 30 min.…”

Palmitic Acid Curcumin Ester Facilitates Protection of Neuroblastoma against Oligomeric Aβ40 Insult

“…S5b-c), indicating that only P-curcumin possessed [37]. We questioned whether the "cultivated" curcumin had characteristics similar to those of curcumin; thus, we evaluated the effect of oligomer Aβ (in the study, Aβ represent Aβ 40 , unless otherwise indicated) on cellular fluorescence. Aβ oligomer has been identified as one of the main components in the brain of patients with AD; therefore, we prepared the oligomer according to a method described in the literature [38].…”

“…Next, the stained cells were washed with serum-free culture medium. Then, 100 μL of the cell culture was transferred to individual polymerase chain reaction (PCR) tubes and the test compound (or positive control) was added, followed by incubation for 5 h. Next, 10 μM oligomer Aβ 40 (with or without copper chloride) was added and incubated for 24 h. The cell suspension was used directly for ROS detection (excitation at 488 nm and emission at 525 nm) using an FC-960 spectrofluorometer.…”

“…Before use, the HFIP-pre-treated Aβ was resuspended in dimethyl sulfoxide (DMSO) and sonicated for 15 min. In order to prepare oligomer Aβ 40 , the above Aβ 40 solution was rocked on a shaker (500 rpm) for 24 h, as described previously [39]. The resulting oligomer Aβ 40 solution was stored at 4 °C and used within 7 days for cell viability and other assays.…”

“…The assessment of ROS production was conducted as previously reported [40]. In brief, H 2 DCF-DA was first converted to dichlorofluorescein (DCF) by NaOH; the neutralized hydrolysate was used for the in vitro assay.…”