Rajendran Rahul, Beck Rachel C, Waskasi Morteza M, Kelly Brian D, Bauer Daniel R
Roche Diagnostics Solutions, (Ventana Medical Systems, Inc.), Tucson, AZ, USA.
J Pathol Inform. 2023 Nov 21;15:100352. doi: 10.1016/j.jpi.2023.100352. eCollection 2024 Dec.
As our understanding of the tumor microenvironment grows, the pathology field is increasingly utilizing multianalyte diagnostic assays to understand important characteristics of tumor growth. In clinical settings, brightfield chromogenic assays represent the gold-standard and have developed significant trust as the first-line diagnostic method. However, conventional brightfield tests have been limited to low-order assays that are visually interrogated. We have developed a hybrid method of brightfield chromogenic multiplexing that overcomes these limitations and enables high-order multiplex assays. However, how compatible high-order brightfield multiplexed images are with advanced analytical algorithms has not been extensively evaluated. In the present study, we address this gap by developing a novel 6-marker prostate cancer assay that targets diverse aspects of the tumor microenvironment such as prostate-specific biomarkers (PSMA and p504s), immune biomarkers (CD8 and PD-L1), a prognostic biomarker (Ki-67), as well as an adjunctive diagnostic biomarker (basal cell cocktail) and apply the assay to 143 differentially graded adenocarcinoma prostate tissues. The tissues were then imaged on our spectroscopic multiplexing imaging platform and mined for proteomic and spatial features that were correlated with cancer presence and disease grade. Extracted features were used to train a UMAP model that differentiated healthy from cancerous tissue with an accuracy of 89% and identified clusters of cells based on cancer grade. For spatial analysis, cell-to-cell distances were calculated for all biomarkers and differences between healthy and adenocarcinoma tissues were studied. We report that p504s positive cells were at least 2× closer to cells expressing PD-L1, CD8, Ki-67, and basal cell in adenocarcinoma tissues relative to the healthy control tissues. These findings offer a powerful insight to understand the fingerprint of the prostate tumor microenvironment and indicate that high-order chromogenic multiplexing is compatible with digital analysis. Thus, the presented chromogenic multiplexing system combines the clinical applicability of brightfield assays with the emerging diagnostic power of high-order multiplexing in a digital pathology friendly format that is well-suited for translational studies to better understand mechanisms of tumor development and growth.
随着我们对肿瘤微环境的认识不断加深，病理学领域越来越多地利用多分析物诊断检测来了解肿瘤生长的重要特征。在临床环境中，明场显色检测是金标准，作为一线诊断方法已获得了高度信任。然而，传统的明场检测仅限于通过视觉观察的低阶检测。我们开发了一种明场显色多重检测的混合方法，克服了这些局限性，实现了高阶多重检测。然而，高阶明场多重图像与先进分析算法的兼容性尚未得到广泛评估。在本研究中，我们通过开发一种新型的6标记前列腺癌检测方法来填补这一空白，该方法针对肿瘤微环境的不同方面，如前列腺特异性生物标志物（PSMA和p504s）、免疫生物标志物（CD8和PD-L1）、预后生物标志物（Ki-67）以及辅助诊断生物标志物（基底细胞混合物），并将该检测方法应用于143例不同分级的前列腺腺癌组织。然后在我们的光谱多重成像平台上对这些组织进行成像，并挖掘与癌症存在和疾病分级相关的蛋白质组学和空间特征。提取的特征用于训练一个UMAP模型，该模型区分健康组织和癌组织的准确率为89%，并根据癌症分级识别细胞簇。对于空间分析，计算了所有生物标志物的细胞间距离，并研究了健康组织和腺癌组织之间的差异。我们报告称，相对于健康对照组织，腺癌组织中p504s阳性细胞与表达PD-L1、CD8、Ki-67和基底细胞的细胞的距离至少近2倍。这些发现为理解前列腺肿瘤微环境的特征提供了有力的见解，并表明高阶显色多重检测与数字分析兼容。因此，所展示的显色多重检测系统以一种对数字病理学友好的形式，将明场检测的临床适用性与高阶多重检测的新兴诊断能力结合起来，非常适合转化研究，以更好地理解肿瘤发生和生长的机制。
