Abstract
Most currently available small molecule inhibitors of DNA replication lack enzymatic specificity, resulting in deleterious side effects during use in cancer chemotherapy and limited experimental usefulness as mechanistic tools to study DNA replication. Towards development of targeted replication inhibitors, we have focused on Mcm2-7 (minichromosome maintenance protein 2–7), a highly conserved helicase and key regulatory component of eukaryotic DNA replication. Unexpectedly we found that the fluoroquinolone antibiotic ciprofloxacin preferentially inhibits Mcm2-7. Ciprofloxacin blocks the DNA helicase activity of Mcm2-7 at concentrations that have little effect on other tested helicases and prevents the proliferation of both yeast and human cells at concentrations similar to those that inhibit DNA unwinding. Moreover, a previously characterized mcm mutant (mcm4chaos3) exhibits increased ciprofloxacin resistance. To identify more potent Mcm2-7 inhibitors, we screened molecules that are structurally related to ciprofloxacin and identified several that compromise the Mcm2-7 helicase activity at lower concentrations. Our results indicate that ciprofloxacin targets Mcm2-7 in vitro, and support the feasibility of developing specific quinolone-based inhibitors of Mcm2-7 for therapeutic and experimental applications.
Highlights
As cancer cells demonstrate uncontrolled proliferation relative to most non-cancer cells, DNA replication has traditionally been an important target for cancer chemotherapy
We provide evidence that ciprofloxacin inhibits the activity of the budding yeast Mcm2-7 helicase both biochemically and in cell culture
Our experiments largely focus on yeast, we demonstrated that ciprofloxacin inhibits the viability of human cells at roughly similar concentrations
Summary
As cancer cells demonstrate uncontrolled proliferation relative to most non-cancer cells, DNA replication has traditionally been an important target for cancer chemotherapy. Such therapeutics are frequently nonspecific and mutagenic, as they either chemically modify the DNA to block replication fork progression or trap deleterious Topo II (topoisomerase II)/DNA double-strand break intermediates [1]. Not surprisingly, these therapies have multiple toxic side effects (reviewed in [2]). As eukaryotic DNA replication is a complex process that is poorly understood at a mechanistic level, the development of targeted small molecule inhibitors of specific replication factors would be of significant research utility
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