Effects of DNA topoisomerase IIα splice variants on acquired drug resistance

Abstract

DNA topoisomerase IIα (170 kDa, TOP2α/170) induces transient DNA double-strand breaks in proliferating cells to resolve DNA topological entanglements during chromosome condensation, replication, and segregation. Therefore, TOP2α/170 is a prominent target for anticancer drugs whose clinical efficacy is often compromised due to chemoresistance. Although many resistance mechanisms have been defined, acquired resistance of human cancer cell lines to TOP2α interfacial inhibitors/poisons is frequently associated with a reduction of Top2α/170 expression levels. Recent studies by our laboratory, in conjunction with earlier findings by other investigators, support the hypothesis that a major mechanism of acquired resistance to TOP2α-targeted drugs is due to alternative RNA processing/splicing. Specifically, several TOP2α mRNA splice variants have been reported which retain introns and are translated into truncated TOP2α isoforms lacking nuclear localization sequences and subsequent dysregulated nuclear-cytoplasmic disposition. In addition, intron retention can lead to truncated isoforms that lack both nuclear localization sequences and the active site tyrosine (Tyr805) necessary for forming enzyme-DNA covalent complexes and inducing DNA damage in the presence of TOP2α-targeted drugs. Ultimately, these truncated TOP2α isoforms result in decreased drug activity against TOP2α in the nucleus and manifest drug resistance. Therefore, the complete characterization of the mechanism(s) regulating the alternative RNA processing of TOP2α pre-mRNA may result in new strategies to circumvent acquired drug resistance. Additionally, novel TOP2α splice variants and truncated TOP2α isoforms may be useful as biomarkers for drug resistance, prognosis, and/or direct future TOP2α-targeted therapies.

CRISPR/Cas9 Genome Editing of the Human Topoisomerase IIαIntron 19 5′ Splice Site Circumvents Etoposide Resistance in Human Leukemia K562 Cells

“…Resistance to TOP2a interfacial poisons is frequently associated with a reduction of TOP2a/170 expression levels or its altered subcellular localization (Ganapathi and Ganapathi, 2013;Capelôa et al, 2020). We previously demonstrated that acquired resistance to etoposide in a human K562 leukemia cell line, K/VP.5, is associated with decreased TOP2a/170 mRNA/protein expression levels and a dramatically increased expression of a novel TOP2a mRNA (University of California Santa Cruz Genome Browser accession number MH936673), which retains a processed intron 19 (I19) and encodes a 90-kDa TOP2a isoform now designated TOP2a/90 (Kanagasabai et al, 2017;Elton et al, 2020). Importantly, the TOP2a/90 isoform heterodimerizes with TOP2a/170, resulting in a dominant-negative effect with respect to etoposide-induced covalent TOP2a-DNA complexes, DNA damage, and cytotoxicity (Kanagasabai et al, 2018), thereby functioning as a resistance determinant.…”

“…Although most intron-retaining mRNA transcripts are susceptible to nuclear intron detention (Boutz et al, 2015) or nonsense-mediated decay (Kurosaki and Maquat 2016), some intron-retaining transcripts leave the nucleus and undergo translation to produce new protein isoforms with novel functions (Li et al, 2016;Uzor et al, 2018;Shoubridge et al, 2019;Wang and Buolamwini, 2019). This process seems to occur in a number of TOP2a intronretaining mRNA variants that are translated into novel truncated TOP2a isoforms and play a role in chemoresistance (Harker et al, 1995;Yu et al, 1997;Mo and Beck, 1997;Kanagasabai et al, 2017Kanagasabai et al, , 2018Elton et al, 2020).…”

Targeting DNA topoisomerases: past & future

“…A characteristic of cancer cells is increased or otherwise altered PTMs modulating enzyme activity, and Top2α is no exception. In this issue, Lotz and Lamour [3] reviewed the interplay between Top2α PTMs and drug resistance. Top2α activity is modulated by multiple phosphorylation sites and cancer cells phosphorylate Top2α at a number of sites not identified in nonmalignant cells including the conserved catalytic tyrosine (Y805), which is phosphorylated in Jurkat cells and K562 acute leukemia cells.…”

“…Cells deficient in Top2α expression or nuclear localization are relatively resistant to Top2 poisons. Elton et al [ 4 ] have identified alternative splice variants of the Top2α protein that do not undergo nuclear localization and cells expressing these variants are resistant to Top2α poisons. In this issue, Elton et al [ 4 ] review Top2α splice variants and the role of alternative splicing as a cause of drug resistance that predominates in some cancer cell lines.…”

“…Elton et al [ 4 ] have identified alternative splice variants of the Top2α protein that do not undergo nuclear localization and cells expressing these variants are resistant to Top2α poisons. In this issue, Elton et al [ 4 ] review Top2α splice variants and the role of alternative splicing as a cause of drug resistance that predominates in some cancer cell lines. Alternative splicing is an important mechanism affecting drug activity that demands consideration in addition to drug efflux mechanisms.…”