Ditercalinium chloride, a pro-anticancer drug, intimately associates with mammalian mitochondrial DNA and inhibits its replication

Abstract

Ditercalinium chloride was originally synthesized for use as an anticancer drug and was then found to deplete mitochondrial DNA. Ethidium bromide is widely used to deplete mitochondrial DNA and produce mitochondrial DNA-less cell lines. Although ethidium bromide is used in the case of human cell lines, it frequently fails to deplete mitochondrial DNA in mouse cells. In contrast, ditercalinium chloride can deplete mitochondrial DNA in both mouse and human cells. However, little is known of the mechanisms by which ditercalinium chloride depletes mitochondrial DNA. Here, we show that ditercalinium chloride inhibits human DNA polymerase gamma activity as efficiently as does ethidium bromide. Ethidium bromide accumulates much less in mouse B82 cells, as compared with findings in human HeLa cells, whereas ditercalinium chloride accumulates in both to a similar extent. This poor accumulation of ethidium bromide may, in part, account for the resistance. Ethidium bromide distributes diffusely in the mitochondria of HeLa cells, while ditercalinium chloride distributes granularly and hence may be strongly associated with mitochondrial DNA. Each granular spot presumably represents one mitochondrial DNA nucleoid. In support of this idea, ditercalinium chloride co-localizes with Twinkle, a mitochondrial helicase and is assumed to associate with mitochondrial DNA. This close association of ditercalinium chloride with mitochondrial DNA may contribute to the mitochondrial DNA-depleting activity.

Ethidium bromide as a marker of mtDNA replication in living cells

“…19 Moreover, it has been demonstrated that in living cells EB stains mtDNA in nucleoids with fluorescence intensity that varies from one nucleoid to the other. 15,17 A similar result was also obtained for DC, 18 which is also an intercalating nucleic acid fluorescent probe. It has been also reported that EB blocks mtDNA replication, 20,21 inhibits DNA polymerase γ, 22 and that a long-term exposition to EB is used to deplete human cells of their mtDNA.…”

“…In living cells, nucleoids have been studied by noninvasive fluorescence microscopy, targeting fluorescent proteins to nucleoids, 10,14 or using supravital nucleic acid fluorescent probes, like ethidium bromide (EB) 10,[15][16][17] and ditercalinium chloride (DC). 18 We should recall that EB is an intercalating nucleic acid probe widely used to quantify the DNA content in fixed cells. On the contrary, it has been shown that in living cells nuclear DNA accessibility to EB is restricted only to a few DNA regions in active replication and/or transcription.…”

Tuning the cellular uptake and cytotoxicity properties of oligonucleotide intercalator-functionalized mesoporous silica nanoparticles with human cervical cancer cells HeLa

“…However, it has been reported in the literature that these molecules can penetrate damaged plasmic and nuclear membranes of animal cells [20,21]. They are genotoxic and mutagenic because of their strong binding affinity to nuclear DNAs and RNAs via intercalation, and thereby blocking the gene replication and transcription processes [22][23][24][25][26][27]. Ideally, if one could find ways to carry these strong oligonucleotide intercalators through the intact plasmic membrane of a healthy live cell, while preventing them from entering the nucleus, it would be possible to design a phenanthridinium-based system for the selective sequestration of cytoplasmic oligonucleotides, i.e., messenger RNAs.…”

Other Anticancer Drugs Targeting DNA and DNA-Associated Enzymes

“…40 In addition, it is assumed to associate with mitochondrial DNA, inhibiting its replication. 41 Elinafide (LU 79553) is a bis intercalator derived from the naphthalimide pharmacophore 42 that exhibited excellent antitumor activity and reached phase I clinical trials, 43 showing anti-neoplastic activity in ovarian cancer, breast cancer, and mesothelioma. Many mechanistic studies on elinafide and its analogs have been undertaken, 44 but this drug has neuromuscular dose-limiting toxicity that has halted its clinical development.…”