Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2019 Nov 18;58(22):15659-15670.
doi: 10.1021/acs.inorgchem.9b02780. Epub 2019 Oct 28.

ATP7B Binds Ruthenium(II) p-Cymene Half-Sandwich Complexes: Role of Steric Hindrance and Ru-I Coordination in Rescuing the Sequestration

Kallol Purkait, Ruturaj, Arindam Mukherjee, Arnab Gupta

ATP7B Binds Ruthenium(II) p-Cymene Half-Sandwich Complexes: Role of Steric Hindrance and Ru-I Coordination in Rescuing the Sequestration

Kallol Purkait et al. Inorg Chem. .

Abstract

Ruthenium(II/III) complexes are predicted to be efficient alternatives to platinum drug-resistant cancers but have never been investigated for sequestration and efflux by Cu-ATPases (ATP7A or ATP7B) overexpressed in resistant cancer cells, although a major cause of platinum drug resistance is found to be sequestration of platinum chemotherapeutic agents by thiol donors glutathione (GSH) or the Cys-X-X-Cys (CXXC) motifs in the Cu-ATPases in cytosol. Here, we show for the first time that ATP7B efficiently sequesters ruthenium(II) η6-p-cymene complexes. We present seven complexes, [RuII(η6-p-cym)(L)X](PF6) (1-7; L = L1-L3, X = Cl, Br, and I), out of which two resists deactivation by the cellular thiol, glutathione (GSH). The results show that Ru-I coordination and a moderate steric factor increase resistance to GSH and the CXXC motif. RuII-I-coordinated 3 and 7 showed resistance to sequestration by ATP7B. 3 displays highest resistance against GSH and does not trigger ATP7B trafficking in the liver cancer cell line. It escapes ATP7B-mediated sequestration and triggers apoptosis. Thus, with a suitable bidentate ligand and iodido leaving group, RuII(η6-p-cym) complexes may display strong kinetic inertness to inhibit the ATP7B detoxification pathway. Inductively coupled plasma mass spectrometry data show higher retention of 3 and 7 inside the cell with time compared to 4, supporting ATP7B-mediated sequestration.

PubMed Disclaimer

Conflict of interest statement

Notes

The authors declare no competing financial interest.

Figures

Figure 1
Figure 1
Chemical structures of complexes 1−7.
Figure 2
Figure 2
Molecular structures of complexes 2, 5, and 6 with 50% probability level thermal ellipsoids obtained from SCXRD. In 6, only one molecule from the asymmetric unit is displayed for clarity. Solvent molecules, hydrogen atoms, and counteranions are omitted for clarity.
Figure 3
Figure 3
Speciation and stability of the complexes in the presence of 25 equiv of GSH at pH 7.4 in the presence of 4 mM NaCl.
Figure 4
Figure 4
Apoptosis induction of MDA-MB-231 cells by complex 3. The four quadrants are as follows: lower left, viable cells; lower right, early apoptosis; upper right, late apoptosis; upper left, necrosis.
Figure 5
Figure 5
Levels of ATP7B mRNA in HepG2 in response to CDDP and ruthenium complex 4. Cells were treated with complexes for 24 h (i). Subsequently, cells were incubated in a fresh complex-free medium for 12 h (ii). Finally, cells were again incubated in a ruthnium-containing medium for 24 h (iii).
Figure 6
Figure 6
Viability (%) of HepG2 cells determined by NR assay after 48 h of treatment with 4 in the presence of 50 μM CuCl2 in normoxic condition in comparison to treatment with only 4. The plots provided are for one independent experiment out of the three independent experiments performed with each concentration.
Figure 7
Figure 7
Levels of HIF-1α in a HepG2 immunoblot showing protein levels in response to ruthenium compound 4 in the presence or absence of copper. GAPDH is taken as the control.
Figure 8
Figure 8
Effect of ruthenium compounds on ATP7B transport. Colocalization of ATP7B (green) and the TGN (red) in response to ruthenium complexes (3 and 4), CDDP, copper, and disodium bathophenanthrolinedisulfonate (BCS). See Figure S24 for large-field images and for complexes 6 and 7. The bar represents 5 μm.
Figure 9
Figure 9
Bacterial length as a function of the stress induced by ruthenium complexes. Bacteria-expressing GST-N-ATP7B or GST was subjected to treatment by ruthenium complexes (3 and 4), and CDDP and bacterial length alterations were recorded. The bar represents 10 μm. (Large images are provided in Figure S25.)
Figure 10
Figure 10
Total cellular ruthenium content measured at two intervals of time: after immediate removal and after 24 h for complexes 3, 4, and 7. The total drug treatment time was 4 h at 37 °C, and the cells used were HepG2. (A) Plot of the ruthenium content in cells after 0 and 24 h of drug removal measured by ICP-MS. (B) Ratio of the cellular ruthenium concentration showing that the retention order for 3 and 7 is quite a bit higher than 4.
See this image and copyright information in PMC

References

    1. Noffke AL, Habtemariam A, Pizarro AM, Sadler PJ. Designing organometallic compounds for catalysis and therapy. Chem Commun. 2012;48(43):5219–5246. - PubMed
    1. Hanif M, Hartinger CG. Anticancer metallodrugs: where is the next cisplatin? Future Med Chem. 2018;10(6):615–617. - PubMed
    1. Kenny RG, Marmion CJ. Toward Multi-Targeted Platinum and Ruthenium Drugs-A New Paradigm in Cancer Drug Treatment Regimens? Chem Rev. 2019;119(2):1058–1137. - PubMed
    1. Murray BS, Menin L, Scopelliti R, Dyson PJ. Conformational control of anticancer activity: the application of arene-linked dinuclear ruthenium(ii) organometallics. Chem Sci. 2014;5(6):2536–2545.
    1. Clavel CM, Paunescu E, Nowak-Sliwinska P, Griffioen AW, Scopelliti R, Dyson PJ. Modulating the Anticancer Activity of Ruthenium(II)-Arene Complexes. J Med Chem. 2015;58(8):3356–3365. - PubMed