Medizinisches Proteom-Center, Ruhr University Bochum, Bochum, Germany.
Department of Biophysics, Ruhr University Bochum, Bochum, Germany.
Am J Pathol. 2019 Mar;189(3):619-631. doi: 10.1016/j.ajpath.2018.11.018. Epub 2019 Feb 12.
Histopathological differentiation between severe urocystitis with reactive urothelial atypia and carcinoma in situ (CIS) can be difficult, particularly after a treatment that deliberately induces an inflammatory reaction, such as intravesical instillation of Bacillus Calmette-Guèrin. However, precise grading in bladder cancer is critical for therapeutic decision making and thus requires reliable immunohistochemical biomarkers. Herein, an exemplary potential biomarker in bladder cancer was identified by the novel approach of Fourier transform infrared imaging for label-free tissue annotation of tissue thin sections. Identified regions of interest are collected by laser microdissection to provide homogeneous samples for liquid chromatography-tandem mass spectrometry-based proteomic analysis. This approach afforded label-free spatial classification with a high accuracy and without interobserver variability, along with the molecular resolution of the proteomic analysis. Cystitis and invasive high-grade urothelial carcinoma samples were analyzed. Three candidate biomarkers were identified and verified by immunohistochemistry in a small cohort, including low-grade urothelial carcinoma samples. The best-performing candidate AHNAK2 was further evaluated in a much larger independent verification cohort that also included CIS samples. Reactive urothelial atypia and CIS were distinguishable on the basis of the expression of this newly identified and verified immunohistochemical biomarker, with a sensitivity of 97% and a specificity of 69%. AHNAK2 can differentiate between reactive urothelial atypia in the setting of an acute or chronic cystitis and nonmuscle invasive-type CIS.
在严重的膀胱炎伴反应性尿路上皮异型增生和原位癌（CIS）之间进行组织病理学鉴别可能具有挑战性，特别是在经过刻意诱导炎症反应的治疗后，如膀胱内卡介苗灌注。然而，膀胱癌的准确分级对于治疗决策至关重要，因此需要可靠的免疫组织化学生物标志物。本研究通过傅里叶变换红外成像对组织薄片进行无标记组织注释的新方法，确定了膀胱癌的一个潜在的有代表性的生物标志物。通过激光微切割收集感兴趣的区域，为基于液相色谱-串联质谱的蛋白质组学分析提供均质样本。这种方法提供了无标记的空间分类，具有高精度和无观察者变异性，以及蛋白质组学分析的分子分辨率。对膀胱炎和浸润性高级别尿路上皮癌样本进行了分析。通过免疫组织化学在一个小队列中鉴定并验证了三个候选生物标志物，包括低级别尿路上皮癌样本。表现最好的候选标志物 AHNAK2 在一个更大的独立验证队列中进行了进一步评估，其中还包括 CIS 样本。基于这个新鉴定和验证的免疫组织化学生物标志物的表达，可以区分反应性尿路上皮异型增生和 CIS，其敏感性为 97%，特异性为 69%。AHNAK2 可以区分急性或慢性膀胱炎背景下的反应性尿路上皮异型增生和非肌肉浸润型 CIS。
