{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T04:26:25Z","timestamp":1760243185096,"version":"build-2065373602"},"reference-count":39,"publisher":"MDPI AG","issue":"1","license":[{"start":{"date-parts":[[2014,1,10]],"date-time":"2014-01-10T00:00:00Z","timestamp":1389312000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/3.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Sensors"],"abstract":"<jats:p>Human topoisomerase I (hTopI) is an essential cellular enzyme. The enzyme is often upregulated in cancer cells, and it is a target for chemotherapeutic drugs of the camptothecin (CPT) family. Response to CPT-based treatment is dependent on hTopI activity, and reduction in activity, and mutations in hTopI have been reported to result in CPT resistance. Therefore, hTOPI gene copy number, mRNA level, protein amount, and enzyme activity have been studied to explain differences in cellular response to CPT. We show that Rolling Circle Enhanced Enzyme Activity Detection (REEAD), allowing measurement of hTopI cleavage-religation activity at the single molecule level, may be used to detect posttranslational enzymatic differences influencing CPT response. These differences cannot be detected by analysis of hTopI gene copy number, mRNA amount, or protein amount, and only become apparent upon measuring the activity of hTopI in the presence of CPT. Furthermore, we detected differences in the activity of the repair enzyme tyrosyl-DNA phosphodiesterase 1, which is involved in repair of hTopI-induced DNA damage. Since increased TDP1 activity can reduce cellular CPT sensitivity we suggest that a combined measurement of TDP1 activity and hTopI activity in presence of CPT will be the best determinant for CPT response.<\/jats:p>","DOI":"10.3390\/s140101195","type":"journal-article","created":{"date-parts":[[2014,1,13]],"date-time":"2014-01-13T03:27:49Z","timestamp":1389583669000},"page":"1195-1207","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":20,"title":["Topoisomerase I as a Biomarker: Detection of Activity at the Single Molecule Level"],"prefix":"10.3390","volume":"14","author":[{"given":"Joanna","family":"Proszek","sequence":"first","affiliation":[{"name":"Department of Pathology, Aarhus University Hospital, Aarhus C 8000, Denmark"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Amit","family":"Roy","sequence":"additional","affiliation":[{"name":"Department of Molecular Biology and Genetics, Aarhus University, Aarhus C 8000, Denmark"},{"name":"Department of Biotechnology, National Institute of Pharmaceutical Education & Research (NIPER), Hajipur-844 101, Bihar, India"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ann-Katrine","family":"Jakobsen","sequence":"additional","affiliation":[{"name":"Department of Pathology, Aarhus University Hospital, Aarhus C 8000, Denmark"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Rikke","family":"Fr\u00f8hlich","sequence":"additional","affiliation":[{"name":"Department of Molecular Biology and Genetics, Aarhus University, Aarhus C 8000, Denmark"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Birgitta","family":"Knudsen","sequence":"additional","affiliation":[{"name":"Department of Molecular Biology and Genetics, Aarhus University, Aarhus C 8000, Denmark"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Magnus","family":"Stougaard","sequence":"additional","affiliation":[{"name":"Department of Pathology, Aarhus University Hospital, Aarhus C 8000, Denmark"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2014,1,10]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"369","DOI":"10.1146\/annurev.biochem.70.1.369","article-title":"DNA topoisomerases: Structure, function, and mechanism","volume":"70","author":"Champoux","year":"2001","journal-title":"Annu. Rev. Biochem."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1196","DOI":"10.1007\/s00595-011-4546-7","article-title":"Topoisomerase I expression in tumors as a biological marker for CPT-11 chemosensitivity in patients with colorectal cancer","volume":"41","author":"Ikeguchi","year":"2011","journal-title":"Surg. Today"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1397","DOI":"10.2165\/11591720-000000000-00000","article-title":"Management strategies for recurrent platinum-resistant ovarian cancer","volume":"71","author":"Naumann","year":"2011","journal-title":"Drugs"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"242","DOI":"10.1016\/j.ctrv.2010.07.005","article-title":"Topotecan for relapsed small cell lung cancer: A systematic review and economic evaluation","volume":"37","author":"Hartwell","year":"2011","journal-title":"Cancer Treat. Rev."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"125","DOI":"10.1517\/14656566.2012.643870","article-title":"Oxaliplatin: A review of approved uses","volume":"13","author":"Stein","year":"2012","journal-title":"Expert Opin. Pharmacother."},{"key":"ref_6","first-page":"309","article-title":"Chemotherapy for locally advanced and metastatic gastric cancer: State of the art and future perspectives","volume":"14","author":"Bittoni","year":"2010","journal-title":"Eur. Rev. Med. Pharmacol. Sci."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"461","DOI":"10.1097\/COC.0b013e3181b4e3b0","article-title":"Irinotecan plus bolus\/infusional 5-Fluorouracil and leucovorin in patients with pretreated advanced pancreatic carcinoma: A multicenter experience of the Gruppo Oncologico Italia Meridionale","volume":"33","author":"Gebbia","year":"2010","journal-title":"Am. J. Clin. Oncol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"163","DOI":"10.1016\/j.critrevonc.2009.08.001","article-title":"Review role of topotecan in gynaecological cancers: current indications and perspectives","volume":"74","author":"Lorusso","year":"2010","journal-title":"Crit. Rev. Oncol.\/Hematol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"629","DOI":"10.1007\/s00280-013-2235-5","article-title":"CPT-11 as a second-line treatment for patients with advanced\/metastatic gastric cancer who failed S-1 (CCOG0702)","volume":"72","author":"Mochizuki","year":"2013","journal-title":"Cancer Chemother. Pharmacol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"15387","DOI":"10.1073\/pnas.242259599","article-title":"The mechanism of topoisomerase I poisoning by a camptothecin analog","volume":"99","author":"Staker","year":"2002","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_11","first-page":"6919","article-title":"Involvement of nucleic acid synthesis in cell killing mechanisms of topoisomerase poisons","volume":"50","author":"Beardmore","year":"1990","journal-title":"Cancer Res."},{"key":"ref_12","first-page":"2116","article-title":"Topoisomerase I-related parameters and camptothecin activity in the colon carcinoma cell lines from the National Cancer Institute anticancer screen","volume":"55","author":"Goldwasser","year":"1995","journal-title":"Cancer Res."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"26441","DOI":"10.1074\/jbc.272.42.26441","article-title":"Trapping of mammalian topoisomerase I and recombinations induced by damaged DNA containing nicks or gaps. Importance of DNA end phosphorylation and camptothecin effects","volume":"272","author":"Pourquier","year":"1997","journal-title":"J. Biol. Chem."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"7792","DOI":"10.1074\/jbc.272.12.7792","article-title":"Effects of uracil incorporation, DNA mismatches, and abasic sites on cleavage and religation activities of mammalian topoisomerase I","volume":"272","author":"Pourquier","year":"1997","journal-title":"J. Biol. Chem."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"620","DOI":"10.1016\/j.dnarep.2013.04.015","article-title":"DNA repair mechanisms in dividing and non-dividing cells","volume":"12","author":"Iyama","year":"2013","journal-title":"DNA Repair"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"161","DOI":"10.1016\/j.cllc.2011.03.013","article-title":"A phase II study of carboplatin and irinotecan in extensive stage small-cell lung cancer","volume":"12","author":"Horn","year":"2011","journal-title":"Clin. Lung Cancer"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"347","DOI":"10.1007\/s10549-013-2476-3","article-title":"A systematic review on topoisomerase 1 inhibition in the treatment of metastatic breast cancer","volume":"138","author":"Kumler","year":"2013","journal-title":"Breast Cancer Res. Treat."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"905","DOI":"10.1056\/NEJM200009283431302","article-title":"Irinotecan plus fluorouracil and leucovorin for metastatic colorectal cancer. Irinotecan Study Group","volume":"343","author":"Saltz","year":"2000","journal-title":"New Engl. J. Med."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"508","DOI":"10.1038\/bjc.1996.394","article-title":"Variation in topoisomerase I gene copy number as a mechanism for intrinsic drug sensitivity","volume":"74","author":"McLeod","year":"1996","journal-title":"Br. J. Cancer"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"84","DOI":"10.3109\/00365521.2013.856464","article-title":"Assessment of the topoisomerase I gene copy number as a predictive biomarker of objective response to irinotecan in metastatic colorectal cancer","volume":"49","author":"Nygard","year":"2013","journal-title":"Scand. J. Gastroenterol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"335","DOI":"10.1002\/(SICI)1097-0215(19970127)70:3<335::AID-IJC15>3.0.CO;2-E","article-title":"CPT-11 in human colon-cancer cell lines and xenografts: Characterization of cellular sensitivity determinants","volume":"70","author":"Jansen","year":"1997","journal-title":"Int. J. Cancer"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"593","DOI":"10.1093\/nar\/21.3.593","article-title":"Characterization of an altered DNA catalysis of a camptothecin-resistant eukaryotic topoisomerase I","volume":"21","author":"Gromova","year":"1993","journal-title":"Nucleic Acids Res."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"6782","DOI":"10.1073\/pnas.0801038105","article-title":"Mutations in topoisomerase I as a self-resistance mechanism coevolved with the production of the anticancer alkaloid camptothecin in plants","volume":"105","author":"Sirikantaramas","year":"2008","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"223","DOI":"10.1021\/nn800509b","article-title":"Single-molecule detection of human topoisomerase I cleavage-ligation activity","volume":"3","author":"Stougaard","year":"2009","journal-title":"ACS Nano"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1492","DOI":"10.1054\/bjoc.2001.2095","article-title":"Colorectal cancer genomics: Evidence for multiple genotypes which influence survival","volume":"85","author":"Rooney","year":"2001","journal-title":"Br. J. Cancer"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e60613","DOI":"10.1371\/journal.pone.0060613","article-title":"Mechanisms of topoisomerase I (TOP1) gene copy number increase in a stage III colorectal cancer patient cohort","volume":"8","author":"Smith","year":"2013","journal-title":"PloS One"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"56","DOI":"10.1016\/S0165-4608(00)00242-9","article-title":"Characterization of the topoisomerase I locus in human colorectal cancer","volume":"121","author":"Boonsong","year":"2000","journal-title":"Cancer Genet. Cytogenet."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"737","DOI":"10.1016\/0020-711X(90)90009-R","article-title":"Phosphorylation sites for type N II protein kinase in DNA-topoisomerase I from calf thymus","volume":"22","author":"Coderoni","year":"1990","journal-title":"Int. J. Biochem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"9418","DOI":"10.1016\/S0021-9258(19)38865-9","article-title":"Phosphorylation of mammalian DNA topoisomerase I and activation by protein kinase C","volume":"265","author":"Pommier","year":"1990","journal-title":"J. Biol. Chem."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"133","DOI":"10.1016\/j.bbapap.2005.03.007","article-title":"Proteomic analysis of complexes formed by human topoisomerase I","volume":"1749","author":"Czubaty","year":"2005","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"871","DOI":"10.1016\/S1074-5521(01)00059-X","article-title":"Directed evolution to increase camptothecin sensitivity of human DNA topoisomerase I","volume":"8","author":"Scaldaferro","year":"2001","journal-title":"Chem. Biol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"3305","DOI":"10.1074\/jbc.M705781200","article-title":"Mutation of Gly721 alters DNA topoisomerase I active site architecture and sensitivity to camptothecin","volume":"283","author":"Bjornsti","year":"2008","journal-title":"J. Biol. Chem."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"55618","DOI":"10.1074\/jbc.M405042200","article-title":"TDP1 overexpression in human cells counteracts DNA damage mediated by topoisomerases I and II","volume":"279","author":"Barthelmes","year":"2004","journal-title":"J. Biol. Chem."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"27","DOI":"10.1016\/j.bcp.2011.09.021","article-title":"Role of tyrosyl-DNA phosphodiesterase 1 and inter-players in regulation of tumor cell sensitivity to topoisomerase I inhibition","volume":"83","author":"Perego","year":"2012","journal-title":"Biochem. Pharmacol."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"230","DOI":"10.1016\/j.bios.2013.04.019","article-title":"Real-time detection of TDP1 activity using a fluorophore-quencher coupled DNA-biosensor","volume":"48","author":"Jensen","year":"2013","journal-title":"Biosens. Bioelectron."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"303","DOI":"10.1016\/j.lungcan.2006.10.019","article-title":"Increased expression and activity of repair genes TDP1 and XPF in non-small cell lung cancer","volume":"55","author":"Liu","year":"2007","journal-title":"Lung Cancer"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"704","DOI":"10.1021\/bi101110e","article-title":"Protein kinase CK2 is a central regulator of topoisomerase I hyperphosphorylation and camptothecin sensitivity in cancer cell lines","volume":"50","author":"Bandyopadhyay","year":"2011","journal-title":"Biochemistry"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"e50427","DOI":"10.1371\/journal.pone.0050427","article-title":"CK2-mediated hyperphosphorylation of topoisomerase I targets serine 506, enhances topoisomerase I-DNA binding, and increases cellular camptothecin sensitivity","volume":"7","author":"Bandyopadhyay","year":"2012","journal-title":"PLoS One"},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Stougaard, M., and Ho, Y.P. (2013). DNA-based nanosensors for next-generation clinical diagnostics via detection of enzyme activity. Expert Rev. Mol. Diagn., 1\u20133.","DOI":"10.1586\/14737159.2014.863151"}],"container-title":["Sensors"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1424-8220\/14\/1\/1195\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T21:07:16Z","timestamp":1760216836000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1424-8220\/14\/1\/1195"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2014,1,10]]},"references-count":39,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2014,1]]}},"alternative-id":["s140101195"],"URL":"https:\/\/doi.org\/10.3390\/s140101195","relation":{},"ISSN":["1424-8220"],"issn-type":[{"type":"electronic","value":"1424-8220"}],"subject":[],"published":{"date-parts":[[2014,1,10]]}}}