A homozygous mutation of human tyrosyl-DNA phosphodiesterase 1 (TDP1) causes the neurodegenerative syndrome, spinocerebellar ataxia with axonal neuropathy (SCAN1). TDP1 hydrolyzes the phosphodiester bond between DNA 3′-end and a tyrosyl moiety within trapped topoisomerase I (Top1)-DNA covalent complexes (Top1cc). TDP1 is critical for mitochondrial DNA (mtDNA) repair; however, the role of mitochondria remains largely unknown for the etiology of SCAN1. We demonstrate that mitochondria in cells expressing SCAN1-TDP1 (TDP1H493R) are selectively trapped on mtDNA in the regulatory non-coding region and promoter sequences. Trapped TDP1H493R-mtDNA complexes were markedly increased in the presence of the Top1 poison (mito-SN38) when targeted selectively into mitochondria in nanoparticles. TDP1H493R-trapping accumulates mtDNA damage and triggers Drp1-mediated mitochondrial fission, which blocks mitobiogenesis. TDP1H493R prompts PTEN-induced kinase 1–dependent mitophagy to eliminate dysfunctional mitochondria. SCAN1-TDP1 in mitochondria creates a pathological state that allows neurons to turn on mitophagy to rescue fit mitochondria as a mechanism of survival.

Camptothecin (CPT) selectively traps topoisomerase 1-DNA cleavable complexes (Top1cc) to promote anticancer activity. Here, we report the design and synthesis of a new class of neutral porphyrin derivative 5,10-bis(4-carboxyphenyl)-15, 20-bis(4-dimethylaminophenyl)porphyrin (compound 8) as a potent catalytic inhibitor of human Top1. In contrast to CPT, compound 8 reversibly binds with the free enzyme and inhibits the formation of Top1cc and promotes reversal of the preformed Top1cc with CPT. Compound 8 induced inhibition of Top1cc formation in live cells was substantiated by fluorescence recovery after photobleaching (FRAP) assays. We established that MCF7 cells treated with compound 8 trigger proteasome-mediated Top1 degradation, accumulate higher levels of reactive oxygen species (ROS), PARP1 cleavage, oxidative DNA fragmentation, and stimulate apoptotic cell death without stabilizing apoptotic Top1-DNA cleavage complexes. Finally, compound 8 shows anticancer activity by targeting cellular Top1 and preventing the enzyme from directly participating in the apoptotic process.

To overcome chemical limitations of camptothecin (CPT), we report design, synthesis, and validation of a quinoline-based novel class of topoisomerase 1 (Top1) inhibitors and establish that compound 28 (N-(3-(1H-imidazol-1-yl)­propyl)-6-(4-methoxyphenyl)-3-(1,3,4-oxadiazol-2-yl)­quinolin-4-amine) exhibits the highest potency in inhibiting human Top1 activity with an IC50 value of 29 ± 0.04 nM. Compound 28 traps Top1–DNA cleavage complexes (Top1ccs) both in the in vitro cleavage assays and in live cells. Point mutation of Top1-N722S fails to trap compound 28-induced Top1cc because of its inability to form a hydrogen bond with compound 28. Unlike CPT, compound 28 shows excellent plasma serum stability and is not a substrate of P-glycoprotein 1 (permeability glycoprotein) advancing its potential anticancer activity. Finally, we provide evidence that compound 28 overcomes the chemical instability of CPT in human breast adenocarcinoma cells through generation of persistent and less reversible Top1cc-induced DNA double-strand breaks as detected by γH2AX foci immunostaining after 5 h of drug removal.

Selective trapping of human topoisomerase 1 (Top1) on the DNA (Top1 cleavage complexes; Top1cc) by specific Top1-poisons triggers DNA breaks and cell death. Poly(ADP-ribose) polymerase 1 (PARP1) is an early nick sensor for trapped Top1cc. New mechanistic insights have been developed in recent years to rationalize the importance of PARP1 beyond the repair of Top1-induced DNA breaks. This review summarizes the progress in the molecular mechanisms of trapped Top1cc-induced DNA damage, PARP1 activation at DNA damage sites, PAR-dependent regulation of Top1 nuclear dynamics, and PARP1-associated molecular network for Top1cc repair. Finally, we have discussed the rationale behind the synergy between the combination of Top1 poison and PARP inhibitors in cancer chemotherapies, which is independent of the ‘PARP trapping’ phenomenon.

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