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The Cytotoxic Action of Cytochrome C/Cardiolipin Nanocomplex (Cyt-CL) on Cancer Cells in Culture

Abstract

Purpose

The effect of existing anti-cancer therapies is based mainly on the stimulation of apoptosis in cancer cells. Here, we have demonstrated the ability of a catalytically-reactive nanoparticle-based complex of cytochrome c with cardiolipin (Cyt-CL) to induce the apoptosis and killing of cancer cells in a monolayer cell culture.

Methods

Cyt-CL nanoparticles were prepared by complexing CytC with different molar excesses of CL. Following characterization, cytotoxicity and apoptosis inducing effects of nanoparticles were investigated. In an attempt to identify the anticancer activity mechanism of Cyt-CL, pseudo-lipoxygenase and lipoperoxidase reaction kinetics were measured by chemiluminescence.

Results

Using chemiluminescence, we have demonstrated that the Cyt-CL complex produces lipoperoxide radicals in two reactions: by decomposition of lipid hydroperoxides, and by lipid peroxidation under the action of H2O2. Antioxidants inhibited the formation of lipid radicals. Cyt-CL nanoparticles, but not the CytC alone, dramatically enhanced the level of apoptosis and cell death in two cell lines: drug-sensitive (A2780) and doxorubicin-resistant (A2780-Adr). The proposed mechanism of the cytotoxic action of Cyt-CL involves either penetration through the cytoplasm and outer mitochondrial membrane and catalysis of lipid peroxidation reactions at the inner mitochondrial membrane, or/and activation of lipid peroxidation within the cytoplasmic membrane.

Conclusions

Here we propose a new type of anticancer nano-formulation, with an action based on the catalytic action of Cyt-CL nanoparticles on the cell membrane and and/or mitochondrial membranes that results in lipid peroxidation reactions, which give rise to activation of apoptosis in cancer cells, including multidrug resistant cells.

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Abbreviations

Bcl-2:

B-cell/lymphoma 2

BCL:

Bovine heart cardiolipin

CytC:

Cytochrome c

Cyt-CL:

Complex of cytochrome с with cardiolipin

IMM and OMM:

Outer and inner mitochondrial membranes

MDR:

Multidrug resistance

PDI:

Polydispersity index

TOCL:

1,1′,2,2′-Tetraoleoyl cardiolipin

PBS:

10 mM NaH2PO4-Na2HPO4 (pH = 7.4)

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Correspondence to Yury A. Vladimirov.

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Vladimirov, Y.A., Sarisozen, C., Vladimirov, G.K. et al. The Cytotoxic Action of Cytochrome C/Cardiolipin Nanocomplex (Cyt-CL) on Cancer Cells in Culture. Pharm Res 34, 1264–1275 (2017). https://doi.org/10.1007/s11095-017-2143-1

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Key Words

  • apoptosis
  • cytotoxicity
  • cytochrome c-cardiolipin complex
  • lipid peroxidation
  • lipid peroxyl radicals