Abstract 2155: High-plex spatial profiling analysis of multidrug CIVO microdose studies in cancer patients
Authors: Deutsch Gary B. , Pollack Seth M. , Thompson Matthew J. , Gundle Kenneth R. , Bertout Jessica A. , Frazier Jason P. , Beirne Emily , Grenley Marc O. , Gong JingJing , Liang Yan , Beechem Joseph M. , Klinghoffer Richard A. , Maki Robert G. 
Published: August 2019

Background: CIVO microdosing studies performed in patient tumors in situ allow drug developers to assess localized tumor and microenvironment responses to multiple agents without having to expose patients to high systemic drug levels. By concentrating microdoses of multiple different drugs within a living tumor in situ, it is possible to compare tumor and immune responses in spatially resolved regions of the same tumor. This type of early phase (Phase 0) clinical study represents a new path for drug developers to gain insight into drug efficacy, tumor associated immune cell modulation, biomarker discovery and validation, and microenvironment interactions for new drugs earlier in the drug development process. Traditionally, analysis of FFPE samples from these microdosing studies involve routine immunohistochemistry, immunofluorescence, and in situ assays that reveal changes in protein and RNA expression. These assays offer tissue wide protein and gene expression information but have limited multiplexing capabilities and dynamic range, as well as rapidly consume precious trial samples. Novel technologies such as NanoString’s GeoMxTM Digital Spatial Profiler (DSP) enable high-plex spatially resolved analysis of proteins and RNA transcripts in single FFPE tissue sections. In this proof of principle study, we utilized GeoMxTM Digital Spatial Profiler for protein and RNA expression on single FFPE sections from patient sarcoma tumors that were microdosed with multiple FDA approved drugs.

Methods: Single FFPE sections from microdosed patient tumor samples were IF stained for DNA and CD3 and whole slide imaged. 600-micron and 100-micron diameter regions of interests (ROIs) were selected for DSP analysis within drug and control microinjection sites. Imaging and barcode counts were performed using GeoMxTM DSP and nCounter systems.

Results: DSP protein analysis highlighted phospho-S6 and phospho-ERK upregulation in response to doxorubicin compared to vehicle site. Additionally, ROIs sampled along the doxorubicin exposure gradient showed a dose-dependent reduction of phosphorylation of both S6 and ERK proteins. DSP RNA analysis revealed drug specific transcript regulation of multiple genes in microdosed tumors, including upregulation of chemokines CXCL9 and CXCL10 at sites of doxorubicin and aldesleukin injection but not to other chemotherapy agents.

Conclusions: Early phase CIVO microdosing studies combined with high-plex DSP opens the door to generating multi-omics data for multiple microdosed drugs within small patient studies. Analysis of protein and RNA expression using DSP enabled collection of targeted region of interest high density data from single FFPE sections, conserving precious patient biopsy samples. Through continued expansion of the GeoMxTM DSP analyte panels, collecting an ever-increasing depth of protein and gene expression data is possible in Phase 0 CIVO microdosing studies.

Citation Format: Gary B. Deutsch, Seth M. Pollack, Matthew J. Thompson, Kenneth R. Gundle, Jessica A. Bertout, Jason P. Frazier, Emily Beirne, Marc O. Grenley, JingJing Gong, Yan Liang, Joseph M. Beechem, Richard A. Klinghoffer, Robert G. Maki. High-plex spatial profiling analysis of multidrug CIVO microdose studies in cancer patients [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 2155.