Abstract 1364: Spatially-resolved in situ expression profiling using the GeoMx™ Cancer Transcriptome Atlas panel in FFPE tissue
Authors: Hoang Margaret L. , Kriner Michelle , Zhou Zoey , Norgaard Zach , Sorg Kristina , Merritt Chris , Piazza Erin , Ross Marty , Fropf Robin , Saraf Nileshi , Danaher Patrick , Rhodes Michael , Beechem Joseph 
Published: September 2020

Abstract
The emerging field of spatial genomics represents a significant advance for biology. To drive new discoveries in spatial genomics and immuno-oncology, we introduce the GeoMx Cancer Transcriptome Atlas (CTA) Panel for comprehensive spatial analysis of cancer pathways using the Nanostring GeoMx Digital Spatial Profiler (DSP). We demonstrate profiling of 1600+ immuno-oncology targets in the tumor, microenvironment, and immune compartments of archival FFPE tissue sections, coupled to downstream Next Generation Sequencing (NGS) readout to enable high-throughput workflows. High-plex spatial RNA molecular profiling with GeoMx CTA was performed as follows:
1. Photocleavable DNA oligonucleotides tags were coupled to 8000+ in situ hybridization probes targeting 1600+ genes. These reagents were allowed to bind targets directly on slide-mounted FFPE tissue sections.
2. ROIs were identified and selected using GeoMx DSP, and ROI-specific oligonucleotide tags were released using ultraviolet exposure.
3. Released oligonucleotide tags from each ROI were collected and deposited into designated wells on a microtiter plate, allowing well indexing of each ROI during NGS library preparation.
4. After indexing, the entire plate was pooled into a single tube for purification and then sequenced on an Illumina instrument.
5. NGS reads were processed into digital counts and mapped back to each ROI, generating a map of transcript activity within the tissue architecture.
We compared data from experiments in which bulk RNA-seq and GeoMx DSP using the CTA Panel were performed on the same samples. Overall, we found good correlation between pseudo-bulk GeoMx CTA (sum of ROIs) and RNA-seq from the same tissue specimen. Individually, however, each ROI showed a distinct expression pattern from bulk, and ROI expression patterns clustered based on similar tissue morphology. Importantly, GeoMx CTA was able to detect a higher number of genes with low expression within the microenvironment and immune spatial compartment compared to bulk RNA-seq, providing a detailed look at the anti-tumor immune response. Lastly, we profiled similar tissues using a novel 18000+ gene whole transcriptome panel and found further enrichment of low-expressers relative to RNA-seq, revealing novel spatial biology previously masked by bulk assays. Together, these data demonstrate that GeoMx offers high sensitivity for genome-scale expression profiling while preserving critical information about tissue architecture. GeoMx DSP technology is for Research Use Only and not for use in diagnostic procedures.
Citation Format: Margaret L. Hoang, Michelle Kriner, Zoey Zhou, Zach Norgaard, Kristina Sorg, Chris Merritt, Erin Piazza, Marty Ross, Robin Fropf, Nileshi Saraf, Patrick Danaher, Michael Rhodes, Joseph Beechem. Spatially-resolved in situ expression profiling using the GeoMx™ Cancer Transcriptome Atlas panel in FFPE tissue [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1364.