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A narrative review synthesizes evidence on HKG selection in endometrial cancer studies, highlighting limitations of commonly used GAPDH, particularly in normal endometrium and endometrial cancer contexts. Findings emphasize that using only one HKG can distort RT-qPCR conclusions, and consensus on validated multi-gene combinations is critical across endometrial cancer histotypes and grades.",{"@graph":69,"@context":123},[70,84,106],{"@type":71,"itemListElement":72},"BreadcrumbList",[73,77,79,82],{"item":74,"name":75,"@type":76,"position":8},"https://docshare.wps.com","Home","ListItem",{"item":78,"name":9,"@type":76,"position":14},"https://docshare.wps.com/document/",{"item":80,"name":35,"@type":76,"position":81},"https://docshare.wps.com/document/healthcare/",3,{"item":83,"name":65,"@type":76,"position":19},"https://docshare.wps.com/document/selecting-optimal-housekeeping-genes-for-rt-qpcr-in-endometrial-cancer-studies-a-narrative-review/372674/",{"url":83,"name":65,"@type":85,"image":86,"author":91,"headline":65,"publisher":94,"fileFormat":97,"inLanguage":63,"description":67,"dateModified":98,"datePublished":99,"encodingFormat":97,"isAccessibleForFree":100,"interactionStatistic":101},"DigitalDocument",{"url":87,"@type":88,"width":89,"height":90},"https://docshare.wps.com/thumbnails/selecting-optimal-housekeeping-genes-for-rt-qpcr-in-endometrial-cancer-studies-a-narrative-review/372674.png","ImageObject",300,407,{"name":92,"@type":93},"Elsa","Person",{"url":74,"name":95,"@type":96},"DocShare","Organization","application/pdf","2026-10-24","2026-09-23",true,{"@type":102,"interactionType":103,"userInteractionCount":105},"InteractionCounter",{"@type":104},"ViewAction",14,{"@type":107,"mainEntity":108},"FAQPage",[109,115,119],{"name":110,"@type":111,"acceptedAnswer":112},"Why are housekeeping genes essential in RT-qPCR for endometrial cancer studies?","Question",{"text":113,"@type":114},"They serve as internal controls to normalize target gene expression across different samples. 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Relying on only one HKG, such as GAPDH, can contribute to broad discrepancies in published RT-qPCR results.","https://schema.org",{"og:url":83,"og:type":125,"og:title":65,"og:site_name":95,"og:description":67},"article",{"robots":127,"canonical":83},"index,follow",{"doc_id":129,"site_id":62},372674,1790244877,{"code":4,"msg":5,"data":132},{"doc_id":129,"user_id":133,"nickname":92,"user_avatar":134,"doc_module":4,"category_id":34,"category_name":35,"doc_title":65,"doc_description":67,"doc_content":135,"file_id":136,"file_url":137,"file_type":138,"file_size":139,"view_count":105,"is_deleted":4,"is_public":8,"is_downloadable":8,"audit_status":8,"page_count":140,"language":141,"language_code":63,"site_id":62,"html_lang":63,"table_of_contents":142,"faqs":143,"seo_title":144,"seo_description":67,"update_tm":145,"read_time":146},137455077381,"https://ap-avatar.wpscdn.com/davatar_994ba38a5ba835b3df7d355c54d3ed8d","Review  \nSelecting Optimal Housekeeping Genes for RT-qPCR in Endometrial Cancer Studies: A Narrative Review  \nMaciej Jó´zwik 1,*, Iwona Sidorkiewicz 2, Joanna Wojtkiewicz 3, Stanisław Sulkowski 4, Andrzej Semczuk 5 and Marcin Jó´zwik 6  \nAcademic Editors: Apostolos Zaravinos and Maria Montserrat Rivera Del Álamo  \nReceived: 17 July 2025  \nRevised: 20 August 2025  \nAccepted: 28 August 2025  \nPublished: 4 September 2025  \nCitation: Jó´zwik, M.; Sidorkiewicz, I.; Wojtkiewicz, J.; Sulkowski, S.; Semczuk, A.; Jó´zwik, M. Selecting Optimal Housekeeping Genes for RT-qPCR in Endometrial Cancer Studies: A Narrative Review. Int. J.  \nMol. Sci. 2025, 26, 8610. [https://](https://)[ ](https://)[doi.org/10.3390/ijms26178610](doi.org/10.3390/ijms26178610)  \n[Copyright:](Copyright:) © 2025 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ([https://creativecommons.org/](https://creativecommons.org/)[ ](https://creativecommons.org/)[licenses/by/4.0/](licenses/by/4.0/)) .  \n1 Department of Gynecology and Gynecologic Oncology, Medical University of Białystok, 15-276 Białystok, Poland  \n2 Clinical Research Support Centre, Medical University of Białystok, 15-276 Białystok, Poland; [iwona.sidorkiewicz@umb.edu.pl](iwona.sidorkiewicz@umb.edu.pl)  \n3 Department of Human Physiology and Pathophysiology, Faculty of Medicine, Collegium Medicum, University of Warmia and Mazury in Olsztyn, 10-082 Olsztyn, Poland; [joanna.wojtkiewicz@uwm.edu.pl](joanna.wojtkiewicz@uwm.edu.pl)  \n[4](4 Department of General Pathomorphology)[ Department of General Pathomorphology](4 Department of General Pathomorphology), [Medical University of Bia](Medical University of Bia)ł[ystok](ystok), [15-269 Bia](15-269 Bia)ł[ystok](ystok), [Poland](Poland); [stanislaw.sulkowski@umb.edu.pl](stanislaw.sulkowski@umb.edu.pl)  \n5 IInd Department of Gynecological Surgery and Gynecological Oncology, Lublin Medical University, 20-090 Lublin, Poland; [andrzej.semczuk@umlub.pl](andrzej.semczuk@umlub.pl)  \n6 Department of Gynecology and Obstetrics, Collegium Medicum, University of Warmia and Mazury in Olsztyn, 10-045 Olsztyn, Poland; [marcin.jozwik@uwm.edu.pl](marcin.jozwik@uwm.edu.pl)  \n* [Correspondence: maciej.jozwik@umb.edu.pl](Correspondence: maciej.jozwik@umb.edu.pl)  \nAbstract  \nDetailed analysis of gene expression by real time-quantitative polymerase chain reaction (RT-qPCR) has become a widespread method. To normalize the expression of target genes, this approach relies on constitutively expressed internal controls known as housekeeping genes (HKGs) . Their proper selection is a critically important methodological step, since all the studied gene expression will be recalculated based on HKG expression. This concise review aims to discuss the selection of HKGs for endometrial cancer (EC) studies. We draw attention to the fact that the commonly used gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is unsuitable as a HKG for research on the normal endometrium, EC, as well as many other tissues. In contrast, accumulating evidence suggests that GAPDHis a pan-cancer marker and an EC marker. Work on GAPDH overexpression inECin relation to overall and relapse-free survival is lacking. Both original research and overviews indicate that at least two HKGs should be used for target gene expression recalculations, a rarely applied technical aspect of final data processing. The insufficiently careful selection in many studies of only one HKG, e.g., GAPDH, can be held responsible for broad discrepancies in published results obtained by this RT-qPCR technique. We provide an account of the discrepancies reported for sex hormone receptors expression in EC. Achieving consensus on the selection and validation of HKGs for research on this cancer is of crucial importance. Ideally, this trusted gene combination should be universal for any EC histotype and grade, irrespect","cbCaicPTMIgZP9Vq","https://ap.wps.com/l/cbCaicPTMIgZP9Vq","pdf",640924,26,"English","# Introduction\n# Housekeeping genes and their selection principles\n## Stability assumptions and definitions\n# GAPDH limitations in endometrial cancer studies\n# Recommended use and validation of multiple HKGs\n# Consequences of insufficient HKG selection\n## Discrepancies in sex hormone receptor expression reports\n# Consensus needs for universal HKG panels","[{\"question\":\"Why are housekeeping genes essential in RT-qPCR for endometrial cancer studies?\",\"answer\":\"They serve as internal controls to normalize target gene expression across different samples. All recalculations of target gene expression depend on the selected HKG expression stability.\"},{\"question\":\"What issue is highlighted with GAPDH as a housekeeping gene?\",\"answer\":\"GAPDH is described as unsuitable for studies involving normal endometrium and endometrial cancer, and accumulating evidence suggests it functions as a pan-cancer and endometrial cancer marker.\"},{\"question\":\"How many housekeeping genes should be used for RT-qPCR data recalculation?\",\"answer\":\"The review indicates that at least two HKGs should be used. Relying on only one HKG, such as GAPDH, can contribute to broad discrepancies in published RT-qPCR results.\"}]","Selecting Optimal Housekeeping Genes for RT-qPCR in Endometrial Cancer Studies - A Narrative Review | PDF",1790195093,66]