Abstract 1992: Changes in gene expression of stromal and epithelial cells during prostate cancer progression
Authors: Babic Milos , Imler Elliot , Shepard Peter , Yeakley Joanne , Nagle Raymond , Seligmann Bruce 
Published: July 2017

Abstract
Prostatic intraepithelial neoplasia (PIN) is a histologic abnormality which arises within the secretory epithelium of prostate glands, without disrupting the layer of basal cells which separates the epithelium from the surrounding stroma. PINs are generally considered to be a step towards development of prostatic adenocarcinoma (PCA). Molecular steps in the transition from normal glands to PIN to PCA are poorly understood, in part due to the difficulty of accessing and characterizing individual PINs without contamination from surrounding tissue. Furthermore, stromal cells surrounding pre-cancerous glands may play an active role in cancer development and progression, but are difficult to profile.
We utilized the ultra-sensitive and quantitative TempO-Seq gene expression assay of the whole transcriptome to profile microdisections of stromal vs. epithelial cells in archival prostatectomy FFPE tissue. TempO-Seq is highly insensitive to RNA degradation (measurements of intact RNA RIN=9.1 to degraded RNA RIN=3.0 correlate with R2=0.97) and does not require RNA extraction, allowing measurement of both soluble and insoluble cross-linked RNA. This allowed deparaffinization and H&E staining of prostate sections prior to the assay, so that morphology was clearly visible and subsections could be readily extracted and profiled. Microdissections were performed on individual 5 µm FFPE sections of prostatectomies from patients with Gleason sum scores of 6, containing PIN alongside PCA. We removed an ~300 µm diameter region of stroma surrounding individual normal, PIN, and PCA glands, followed by targeted dissection of the gland epithelium itself which was lysed and assayed separately. Stroma and epithelial samples from 3 adjacent sections of the same glands were pooled together (adding up to ~0.005 mm3 tissue/replicate), and then profiled.
We compared the whole transcriptome expression profile of stroma vs. epithelium at every stage of progression. Our results indicate that stroma and epithelium undergo separate and distinct changes in gene expression patterns throughout the development of PCA, most notably in genes involved in cell-cell signaling. Specifically, we found that changes in cAMP signaling in stromal cells accompanied the shift from normal glands to PIN. Furthermore, changes in androgen signaling pathways were present in stromal cells as well as in epithelium during the transition from PIN to invasive carcinoma.
These results shed light on the molecular progression to PCA and demonstrate the feasibility of profiling gene expression at the level of tissue microstructure to elucidate changes within the tissue microenvironment. Since archival FFPE can be used, studies of progression can be carried out and the findings translated to diagnosis and prognosis as well as to the investigation of new therapeutic approaches.
Citation Format: Milos Babic, Elliot Imler, Peter Shepard, Joanne Yeakley, Raymond Nagle, Bruce Seligmann. Changes in gene expression of stromal and epithelial cells during prostate cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1992. doi:10.1158/1538-7445.AM2017-1992