Acquisition of Genetic Aberrations During the Progression of High-Grade Intraepithelial Lesions/Micro-Invasive Squamous Cancers to Widely Invasive Cervical Sauamous Cell Cancer

PIK3CA is the most commonly mutated gene in cervical cancer, and frequency of detection increased with tumor stage. 1~10 Recent data showed that driver mutations are present already in early invasive disease,11 while all HSILs lacked somatic mutations.12 Early invasive tumor cells acquire a survival advantage in a nutrient-deficient microenvironment including a migratory phenotype.3 PIK3CA E545K mutations enhance tumor cell proliferation by promoting glucose metabolism,13 reduce dependence on growth factors under cell culture conditions,14 and provide tumor cells with migratory (metastatic) potential via cytoskeletal reorganization.15 Not only mutations, but also methylation or amplifications of PIK3CA E545K and/ or gains of chromosome arm 3q, promote cervical carcinogenesis3,16,17 via phosphorylation of numerous proteins18 that enhance cell motility and invasion by regulating signal transduction from the cell membrane to cytoskeletal proteins.19 To document acquisition of genetic aberrations during carcinogenesis in individual patients, we searched our archives dating back 30 years for patients with HSILs or pTla SCCs without lymphovascular space involvement (LVSI) who presented with recurrences or invasive SCC after more than 1 year. pTla SCCs without LVSI were considered equivalent to HSILs for the outset of analysis, as both lesions are treated identically, and both HSILs and pTla SCCs without LVSI are believed to have no metastatic potential. Other HSILs, however, may arise from a second type of CK7+ reserve cells of embryological Miillerian epithelial origin with the highest concentration near the isthmus that can differentiate toward both glandular and squamous epithelium.21,22 MATERIALS AND METHODS We searched the archive of the Diagnostic and Research Institute of Pathology, Medical University Graz, Graz, Austria, for patients with a diagnosis of HPV-induced HSIL/pTla SCC followed by invasive cervical SCC after at least a 1-year interval. Sequencing was done on an Ion Genestudio S5 XL benchtop sequencer (Thermo Fisher) to a length of 200 base pairs, and initial data were analyzed using the Ion Torrent Suite Software Plug-ins (Thermo Fisher, open source, general public license, https://github.com/ iontorrent/). Called variants were annotated using open source software ANNOVAR23 and SnpEff.24 All coding, nonsynonymous mutations were further evaluated and visually inspected using the Integrated Genomics Viewer,25 and variant calls resulting from tech