Abstract 4709: Extensive intratumor proteogenomic heterogeneity revealed by multiregion sampling in a high-grade serous ovarian tumor specimen
Authors: Hunt Allison L. , Bateman Nicholas W. , Wang Guisong , Parikh Niyati , Oliver Julie , Mitchell Dave , Gist Glenn , Hood Brian L. , Zhou Ming , Blanton Brian , Conrads Kelly A. , Hamilton Chad A. , Darcy Kathleen M. , Shriver Craig D. , Casablanca Yovanni , Maxwell George L. , Conrads Thomas P. 
Published: August 2019

Abstract
We generated 200 consecutive thin sections from a high-grade serous ovarian carcinoma (HGSOC) tissue specimen and laser microdissected (LMD) four spatially separated tumor “core” regions throughout the depth of the tissue block to examine proteogenomic intratumor heterogeneity (ITH). Tumor epithelium and stroma were each LMD harvested at 150µm intervals throughout the block and mixed epithelial and stromal (e.g. whole tumor) samples were harvested from additional adjacent thin sections; the remaining tissue was cryopulverized. Mass spectrometry-based proteomics quantified 6,053 proteins and 4,225 phosphosites and RNA sequencing mapped to 20,785 transcripts. Unsupervised hierarchical cluster analysis of the 1,018 and 584 differentially abundant transcripts and proteins (median absolute deviation &gt; 1) demonstrated clustering by LMD sample types collected, with tumor cores and cross-sectional tumor epithelium samples versus mixed epithelium/stroma and stroma samples co-clustering. The cryopulverized proteome classified independently and represented a completely unique sample. Surprisingly, many proteins and transcripts previously classified as ovarian tumor biomarkers, many of which correlate with poor clinical prognosis, were absent in the LMD harvested tumor epithelium, yet were strongly expressed in the LMD enriched stroma. Comparison of our data with existing immunoreactive, differentiated, proliferative, and mesenchymal molecular subtypes, the overall protein and transcript abundance of discrete tumor collections inversely correlated with the mesenchymal HGSOC subtype, which is associated with the worst overall patient survival, whereas the LMD enriched stroma collections were, unexpectedly, most positively correlated. While there was overall concordance between the proteomic and transcriptomic data for each LMD collection type, transcript abundance from LMD enriched stroma was most strongly correlated with the cognate protein products from the mixed epithelial/stromal LMD harvest than to stroma, potentially explainable by the secretory nature of ovarian stromal cells and inclusion of extracellular matrix proteins in mixed LMD collections. Proteins measured from the cryopulverized tissue were overall negatively correlated with LMD harvested tumor epithelium, and notably had limited detection of the ovarian cancer biomarker CA-125. This proteogenomic analysis reveals stark molecular heterogeneity in the cellular admixture of the HGSOC tumor microenvironment and underscores the need to account for compartmental ITH in molecular profiling analyses.
Citation Format: Allison L. Hunt, Nicholas W. Bateman, Guisong Wang, Niyati Parikh, Julie Oliver, Dave Mitchell, Glenn Gist, Brian L. Hood, Ming Zhou, Brian Blanton, Kelly A. Conrads, Chad A. Hamilton, Kathleen M. Darcy, Craig D. Shriver, Yovanni Casablanca, George L. Maxwell, Thomas P. Conrads. Extensive intratumor proteogenomic heterogeneity revealed by multiregion sampling in a high-grade serous ovarian tumor specimen [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4709.