SEP 26, 2019 06:00 AM PDT
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Profiling Focal Areas of FFPE Tissue: Spatial Transcriptomics and the Challenge of Low Input Samples

Presented At Cell Biology 2019
C.E. CREDITS: P.A.C.E. CE | Florida CE
Speakers
  • Co-Founder and CSO, BioSpyder Technologies, Inc., Professor, University of Arizona College of Pharmacy
    Biography
      Bruce began his research career at the NIH, within the Laboratory of Clinical Investigation, NIAID, developing novel technology to monitor the function of cells and identify the basis for disease. He continued that through his work at Ciba-Geigy (now Novartis) where he worked with management to bring molecular biology out of an institute and into the drug discovery project team for the first time within that company, and implemented novel assays to accelerate the drug discovery programs he led, including novel testing of samples to demonstrate efficacy in the human through Phase I. He joined Selectide, a leader in combinatorial chemistry during the infancy of this new paradigm, as the VP of Research and Dev. and transformed it from a peptide company to a small molecule combinatorial chemistry leader, resulting it its sale to Marion Merrell Dow (now Sanofi). After serving as its center director. He left what had become Hoechst Marion Roussel to found SIDDCO around a novel combinatorial chemistry consortium services business plan and then founded HTG Molecular (now trading under the symbol HTGM), inventing (US 2014/0235460) and leading the development of a novel gene expression platform, marketed as the EdgeSeq platform. He left to co-found BioSpyder and develop its TempO-Seq® sequencing-based platform and applications. All of BioSpyder's research program, and initially all its staff until commercial sales began, was supported by grants and contracts he wrote and/or was key, and grant support continues to fund the research program. This effort resulted in the development of the TempO-Seq™ targeted sequencing platform (US 9,856,521; US 9,938,566; US 9,957,550), launched in 2016 (1,2). Launched assays include a human whole transcriptome assay (measuring 38,000+ RefSeq ID's), human surrogate assay (~2700 genes, the EPA S1500V 2, (Mav, et. al. PLOSone, 2018; doi.org/10.1371/journal.pone.0191105), human pan cancer assay of ~5,000 genes, and rat and mouse whole transcriptome and surrogate assays. Commercial kits include kits for the measurement from cells and frozen tissue (1,2) or FFPE tissue (3,4). TempO-Seq has enabled high throughput generation of QSAR-quality dose response data using a focused gene panel (5 and Ramaiahgri, et al, Tox. Sci. 2019. doi.org/10.1093/toxsci/kfz065) for use in quantitative safety assessment drug discovery, or mechanistic/mode of action studies. Validation of applications for single cells using FACS and Digital Spatial Molecular Profiling (DSMP) permitting profiling from H&E or antibody-stained FFPE tissue down to focal areas of 20 μm are ongoing. Diagnostic applications are being pursued.

    Abstract:

    Spatial transcriptomics methods permit gene expression from focal areas within a tissue to be profiled while maintaining the morphologic context of the tissue microenvironment. This presentation will address considerations based on the needs of a study for selecting the best fit to what are new methods still in their infancy. These  important considerations start with; i) whether focal profiling will provide data that profiling of bulk tissue will not; ii) is it necessary to profile single cells or are larger focal areas such as single glands sufficient; iii) what is the size of such histologic regions and Is it necessary to have histologically discrete profiling without cross-contamination from adjacent histologic regions or cells; iv) What is the throughput required, just a few sections or lots of sections from 100+ tissues, animals, or subjects, and within each section, how many focal areas; v) Is it desirable to measure splice variants, snRNA, long non-coding RNA, histones - if so, then a 3’ biased assay method would be unsuited for the need vi) What tissue type does the method use, and how does that match up with what the investigator has available - Frfsh frozen, FFPE, H&E stained, antibody-stained; vii) What is the sensitivity – can all the genes measured from a bulk sample, including low expressed ones be measured; viii) What is the dynamic range; i.e. can a broad range of expressed genes from low to high be measured quantitatively at the same time. As methods continue in their development, the choice will also be whether the investigator wants to measure a panel of antigens at the same time as gene expression. If so, can this be from different serial sections, or does it have to be from the same section, same focal area. And of course, there is the question of cost. Different methods will be presented within the context of these questions, and spatial profiling data using TempO-Seq® will be provided to demonstrate several of these considerations. This data will also demonstrate the potential of spatial profiling in understanding the specificity and context of gene expression within the tissue microenvironment.


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