{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,8]],"date-time":"2026-07-08T04:17:02Z","timestamp":1783484222607,"version":"3.55.0"},"reference-count":12,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2026,7,8]],"date-time":"2026-07-08T00:00:00Z","timestamp":1783468800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2026,7,8]],"date-time":"2026-07-08T00:00:00Z","timestamp":1783468800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/501100006692","name":"Universit\u00e0 degli Studi di Torino","doi-asserted-by":"crossref","id":[{"id":"10.13039\/501100006692","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Mol Biol Rep"],"published-print":{"date-parts":[[2026,12]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Polymorphisms in the TATA-box of the UGT1A1 promoter are responsible for Gilbert syndrome and play a key role in irinotecan-related toxicity. Reliable, rapid, and cost-effective genotyping methods are therefore required in routine molecular diagnostics.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods and Results<\/jats:title>\n                    <jats:p>We performed an analytical validation of a High-Resolution Melting (HRM) assay designed to discriminate UGT1A1 TATA-box alleles. A total of 106 neonatal clinical samples were analyzed to assess genotype distribution under routine diagnostic conditions. Analytical validation was performed using plasmid controls, reference genomic DNA samples, and sequencing-confirmed in-house samples. Plasmid-derived TA5, TA6, TA7, and TA8 genotypes and their heterozygous combinations were tested in quintuplicate. Reference genomic DNA samples carrying rare or non-observed genotypes were tested in triplicate, and 30 in-house samples representative of TA6\/TA6, TA6\/TA7, and TA7\/TA7 genotypes were tested in triplicate and confirmed by Sanger sequencing. The HRM assay discriminated the tested genotypes based on melting temperature (Tm) and curve morphology. HRM genotype calls showed complete concordance with expected or reference genotypes. Intra-test analysis of the TA6\/TA7 standard showed a mean Tm of 43.36\u00a0\u00b0C, SD of 0.20\u00a0\u00b0C, and CV of 0.46%, while inter-test analysis showed identical Tm values across replicates.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>The UGT1A1 TATA-box HRM assay showed reproducible genotype discrimination in the tested validation panel and may represent a practical approach for routine UGT1A1 promoter genotyping.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1007\/s11033-026-12240-9","type":"journal-article","created":{"date-parts":[[2026,7,8]],"date-time":"2026-07-08T04:09:54Z","timestamp":1783483794000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Analytical validation of a high-resolution melting assay for UGT1A1 TATA-box polymorphisms"],"prefix":"10.1007","volume":"53","author":[{"given":"Stefano","family":"Gambarino","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anna","family":"Clemente","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anna","family":"Pau","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cristina","family":"Calvi","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paola","family":"Montanari","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1273-3178","authenticated-orcid":false,"given":"Ilaria","family":"Galliano","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Massimiliano","family":"Bergallo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2026,7,8]]},"reference":[{"key":"12240_CR1","doi-asserted-by":"publisher","first-page":"492","DOI":"10.1006\/bbrc.2002.6683","volume":"292","author":"J Sugatani","year":"2002","unstructured":"Sugatani J, Yamakawa K, Yoshinari K, Machida T, Takagi H, Mori M et al (2002) Identification of a defect in the UGT1A1 gene promoter and its association with hyperbilirubinemia. Biochem Biophys Res Commun 292:492\u2013497. https:\/\/doi.org\/10.1006\/bbrc.2002.6683","journal-title":"Biochem Biophys Res Commun"},{"key":"12240_CR2","doi-asserted-by":"publisher","first-page":"1171","DOI":"10.1056\/NEJM199511023331802","volume":"333","author":"PJ Bosma","year":"1995","unstructured":"Bosma PJ, Chowdhury JR, Bakker C, Gantla S, de Boer A, Oostra BA et al (1995) The genetic basis of the reduced expression of bilirubin UDP-glucuronosyltransferase 1 in Gilbert\u2019s syndrome. N Engl J Med 333:1171\u20131175. https:\/\/doi.org\/10.1056\/NEJM199511023331802","journal-title":"N Engl J Med"},{"key":"12240_CR3","doi-asserted-by":"publisher","first-page":"324","DOI":"10.1038\/clpt.2014.126","volume":"96","author":"C Guillemette","year":"2014","unstructured":"Guillemette C, L\u00e9vesque \u00c9, Rouleau M (2014) Pharmacogenomics of human uridine diphospho-glucuronosyltransferases and clinical implications. Clin Pharmacol Ther 96:324\u2013339. https:\/\/doi.org\/10.1038\/clpt.2014.126","journal-title":"Clin Pharmacol Ther"},{"key":"12240_CR4","doi-asserted-by":"publisher","first-page":"1382","DOI":"10.1200\/JCO.2004.07.173","volume":"22","author":"F Innocenti","year":"2004","unstructured":"Innocenti F, Undevia SD, Iyer L, Chen PX, Das S, Kocherginsky M et al (2004) Genetic variants in the UDP-glucuronosyltransferase 1A1 gene predict the risk of severe neutropenia of irinotecan. J Clin Oncol 22:1382\u20131388. https:\/\/doi.org\/10.1200\/JCO.2004.07.173","journal-title":"J Clin Oncol"},{"key":"12240_CR5","doi-asserted-by":"publisher","first-page":"1389","DOI":"10.1111\/bcp.13574","volume":"84","author":"P Riera","year":"2018","unstructured":"Riera P, Salazar J, Virgili AC, Tobe\u00f1a M, Sebio A, Gallano P et al (2018) Relevance of CYP3A4*20, UGT1A1*37 and UGT1A1*28 variants in irinotecan-induced severe toxicity. Br J Clin Pharmacol 84:1389\u20131392. https:\/\/doi.org\/10.1111\/bcp.13574","journal-title":"Br J Clin Pharmacol"},{"key":"12240_CR6","doi-asserted-by":"publisher","first-page":"9","DOI":"10.1016\/j.ejca.2020.09.007","volume":"141","author":"EC Hulshof","year":"2020","unstructured":"Hulshof EC, Deenen MJ, Guchelaar HJ, Gelderblom H (2020) Pre-therapeutic UGT1A1 genotyping to reduce the risk of irinotecan-induced severe toxicity: Ready for prime time. Eur J Cancer 141:9\u201320. https:\/\/doi.org\/10.1016\/j.ejca.2020.09.007","journal-title":"Eur J Cancer"},{"key":"12240_CR7","doi-asserted-by":"publisher","first-page":"549","DOI":"10.2353\/jmoldx.2008.080036","volume":"10","author":"U Ehmer","year":"2008","unstructured":"Ehmer U, Lankisch TO, Erichsen TJ, Kalthoff S, Freiberg N, Wehmeier M et al (2008) Rapid allelic discrimination by TaqMan PCR for the detection of the Gilbert\u2019s syndrome marker UGT1A1*28. J Mol Diagn 10:549\u2013552. https:\/\/doi.org\/10.2353\/jmoldx.2008.080036","journal-title":"J Mol Diagn"},{"key":"12240_CR8","doi-asserted-by":"publisher","first-page":"250","DOI":"10.1016\/j.ymeth.2010.01.013","volume":"50","author":"M Erali","year":"2010","unstructured":"Erali M, Wittwer CT (2010) High resolution melting analysis for gene scanning. Methods 50:250\u2013261. https:\/\/doi.org\/10.1016\/j.ymeth.2010.01.013","journal-title":"Methods"},{"key":"12240_CR9","doi-asserted-by":"publisher","first-page":"860","DOI":"10.1002\/humu.21019","volume":"30","author":"RH Vossen","year":"2009","unstructured":"Vossen RH, Aten E, Roos A, den Dunnen JT (2009) High-resolution melting analysis (HRMA): More than just sequence variant screening. Hum Mutat 30:860\u2013866. https:\/\/doi.org\/10.1002\/humu.21019","journal-title":"Hum Mutat"},{"key":"12240_CR10","doi-asserted-by":"publisher","first-page":"853","DOI":"10.1373\/49.6.853","volume":"49","author":"CT Wittwer","year":"2003","unstructured":"Wittwer CT, Reed GH, Gundry CN, Vandersteen JG, Pryor RJ (2003) High-resolution genotyping by amplicon melting analysis using LCGreen. Clin Chem 49:853\u2013860. https:\/\/doi.org\/10.1373\/49.6.853","journal-title":"Clin Chem"},{"key":"12240_CR11","doi-asserted-by":"publisher","first-page":"396","DOI":"10.1373\/49.3.396","volume":"49","author":"CN Gundry","year":"2003","unstructured":"Gundry CN, Vandersteen JG, Reed GH, Pryor RJ, Chen J, Wittwer CT (2003) Amplicon melting analysis with labeled primers: A closed-tube method for differentiating homozygotes and heterozygotes. Clin Chem 49:396\u2013406. https:\/\/doi.org\/10.1373\/49.3.396","journal-title":"Clin Chem"},{"key":"12240_CR12","doi-asserted-by":"publisher","first-page":"219","DOI":"10.1586\/erm.09.84","volume":"10","author":"JL Montgomery","year":"2010","unstructured":"Montgomery JL, Sanford LN, Wittwer CT (2010) High-resolution DNA melting in clinical diagnostics. Expert Rev Mol Diagn 10:219\u2013240. https:\/\/doi.org\/10.1586\/erm.09.84","journal-title":"Expert Rev Mol Diagn"}],"container-title":["Molecular Biology Reports"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1007\/s11033-026-12240-9.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/article\/10.1007\/s11033-026-12240-9","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1007\/s11033-026-12240-9.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,8]],"date-time":"2026-07-08T04:09:55Z","timestamp":1783483795000},"score":1,"resource":{"primary":{"URL":"https:\/\/link.springer.com\/10.1007\/s11033-026-12240-9"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,8]]},"references-count":12,"journal-issue":{"issue":"1","published-print":{"date-parts":[[2026,12]]}},"alternative-id":["12240"],"URL":"https:\/\/doi.org\/10.1007\/s11033-026-12240-9","relation":{},"ISSN":["0301-4851","1573-4978"],"issn-type":[{"value":"0301-4851","type":"print"},{"value":"1573-4978","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,8]]},"assertion":[{"value":"27 February 2026","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"18 June 2026","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"8 July 2026","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"The clinical data analyzed in this study were generated as part of routine diagnostic testing. All data were anonymized prior to their use for analytical validation. The study did not involve any additional sampling, intervention or interaction with patients. According to institutional procedures, studies based exclusively on anonymized routine diagnostic data do not require approval by an ethics committee.","order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Institutional Review Board Statement"}},{"value":"Not applicable.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Informed Consent"}},{"value":"M.B., S.G., A.P. and A.C. are affiliated with or collaborate with BioMole S.r.l, the company providing the UGT1A1 TATA-Box HRM kit used in this study. Data analysis and interpretation were performed by the listed authors using anonymized primary HRM and sequencing results. Authors not affiliated with BioMole S.r.l. had access to the anonymized original results and participated in data interpretation. The company did not influence the interpretation of the data, the decision to submit the manuscript, or the preparation of the final conclusions. I.G., C.C., P.M. declare no conflict of interest.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"<!--Emphasis Type='Bold' removed-->Conflicts of Interest"}}],"article-number":"1103"}}