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. 2020 Aug 15;26(16):4360-4368.
doi: 10.1158/1078-0432.CCR-20-0175. Epub 2020 Apr 6.

Biomarkers Associated with Beneficial PD-1 Checkpoint Blockade in Non-Small Cell Lung Cancer (NSCLC) Identified Using High-Plex Digital Spatial Profiling

Jon Zugazagoitia  1 , Swati Gupta  2 , Yuting Liu  2 , Kit Fuhrman  3 , Scott Gettinger  4 , Roy S Herbst  4 , Kurt A Schalper  2   4 , David L Rimm  2   4
Affiliations

Biomarkers Associated with Beneficial PD-1 Checkpoint Blockade in Non-Small Cell Lung Cancer (NSCLC) Identified Using High-Plex Digital Spatial Profiling

Jon Zugazagoitia et al. Clin Cancer Res. .

Abstract

Purpose: Only a minority of patients with advanced non-small cell lung cancer (NSCLC) truly benefits from single-agent PD-1 checkpoint blockade, and more robust predictive biomarkers are needed.

Experimental design: We assessed tumor samples from 67 immunotherapy-treated NSCLC cases represented in a tissue microarray, 53 of whom had pretreatment samples and received monotherapy. Using GeoMx Digital Spatial Profiling System (NanoString Technologies), we quantified 39 immune parameters simultaneously in four tissue compartments defined by fluorescence colocalization [tumor (panCK+), leucocytes (CD45+), macrophages (CD68+), and nonimmune stroma].

Results: A total of 156 protein variables were generated per case. In the univariate unadjusted analysis, we found 18 markers associated with outcome in spatial context, five of which remained significant after multiplicity adjustment. In the multivariate analysis, high levels of CD56 and CD4 measured in the CD45 compartment were the only markers that were predictive for all clinical outcomes, including progression-free survival (PFS, HR: 0.24, P = 0.006; and HR: 0.31, P = 0.011, respectively), and overall survival (OS, HR: 0.26, P = 0.014; and HR: 0.23, P = 0.007, respectively). Then, using an orthogonal method based on multiplex immunofluorescence and cell counting (inForm), we validated that high CD56+ immune cell counts in the stroma were associated with PFS and OS in the same cohort.

Conclusions: This pilot scale discovery study shows the potential of the digital spatial profiling technology in the identification of spatially informed biomarkers of response to PD-1 checkpoint blockade in NSCLC. We identified a number of relevant candidate immune predictors in spatial context that deserve validation in larger independent cohorts.

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Conflict of interest statement

Potential conflict of interest statement

J. Zugazagoitia has received consulting honoraria from Guardant Health, speaker fees from Roche, Pfizer, Guardant Health, and NanoString, and travel fees from Roche.

Kit Fuhrman is NanoString employee.

Roy Herbst has served as a consultant for Abbvie Pharmaceuticals, AstraZeneca, Biodesix, Bristol-Myers Squibb, Eli Lilly and Company, EMD Serrano, Genentech/Roche, Heat Biologics, Loxo Oncology, Merck and Company, Nektar, NextCure, Novartis, Pfizer, Sanofi, Seattle Genetics, Shire PLC, Spectrum Pharmaceuticals, Symphogen, and Tesaro. He has received research support from AstraZeneca, Eli Lilly and Company, and Merck and Company

Kurt Schalper has served as a consultant, advisor or served on a Scientific Advisory Board for Clinical Alemana de Santiago, Celgene, Moderna Therapeutics and Shattuck Labs, Agenus, Torque Therapeutics, Pierre-Fabre, Dynamo Therapeutics, EMD Serono, Astra Zeneca. He has received research funding from Genoptix/Navigate (Novartis), Vasculox/Tioma, Tesaro, Onkaido Therapeutics, Takeda Pharmaceuticals, Surface Oncology, Pierre-Fabre Research Institute, Merck, Bristol-Myers Squibb, Astra Zeneca, Eli Lilly.

David Rimm has served as a consultant, advisor or served on a Scientific Advisory Board for Amgen, Astra Zeneca, Agendia, Biocept, BMS, Cell Signaling Technology, Cepheid, Daiichi Sankyo, GSK, Lilly, Merck, NanoString, Perkin Elmer, PAIGE, Ventana and Ultivue. He has received research funding or instrument support from Astra Zeneca, Cepheid, NanoString, Navigate/Novartis, NextCure, Lilly, Ultivue, and Perkin Elmer.

The remaining authors declare no conflicts of interest.

Figures

Figure 1.
Figure 1.
Representative TMA spots showing the fluorescence image (A) and the compartmentalized image created by fluorescence co-localization (B) using GeoMx DSP (scale bar = 100 μm)
Figure 2.
Figure 2.
CD56 and CD4 expression in the CD45 compartment measured by digital counts and their association with outcome. A-B. Histogram showing the distribution of CD56 digital counts (A) and CD4 digital counts (B) in YTMA404 (n = 42); C-D. PFS according to CD56 digital counts in the CD45 compartment (top tertile) (C) and CD4 digital counts in the CD45 compartment (median) (D) (n = 42); E-F. OS according to CD56 digital counts in the CD45 compartment (top tertile) (E) and CD4 digital counts in the CD45 compartment (median) (F) (n = 42)
Figure 3.
Figure 3.
Orthogonal validation of CD56+/CK− cell counts assessed by Inform as predictors of outcome in YTMA404 cohort. A. Representative images acquired with Vectra Polaris microscope showing CD56 staining pattern in four NSCLC cases (scale bar = 100 μm). CK+ tumor cells are shown in green, CD3+ T cells in red, and CD56+ cells in white. Orange arrows indicate CD56+ NK cells, yellow arrows indicate CD3+/CD56+ NKT cells. Panel 3 illustrates a NSCLC case with strong CD56 positivity in the tumor compartment, and the red asterisk highlights CD56+/CK+ tumor cells; B. Distribution of CD56+/CK− cells in the tumor compartment (B), the stromal compartment, (C) and entire TMA spot (D) (n = 42); E-F. PFS (E) and OS (F) according to CD56+/CK− cell counts (top tertile) in YTMA404 cohort (n = 42)
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References

    1. Doroshow DB, Sanmamed MF, Hastings K, Politi K, Rimm DL, Chen L, et al. Immunotherapy in Non-Small Cell Lung Cancer: Facts and Hopes. Clin Cancer Res. 2019;25(15):4592–602. - PMC - PubMed
    1. Stack EC, Wang C, Roman KA, Hoyt CC. Multiplexed immunohistochemistry, imaging, and quantitation: a review, with an assessment of Tyramide signal amplification, multispectral imaging and multiplex analysis. Methods. 2014;70(1):46–58. - PubMed
    1. Lu S, Stein JE, Rimm DL, Wang DW, Bell JM, Johnson DB, et al. Comparison of Biomarker Modalities for Predicting Response to PD-1/PD-L1 Checkpoint Blockade: A Systematic Review and Meta-analysis. JAMA Oncol. 2019. [Epub Ahead of Print]. - PMC - PubMed
    1. Merritt CR, Ong GT, Church S, Barker K, Geiss G, Hoang M, et al. High multiplex, digital spatial profiling of proteins and RNA in fixed tissue using genomic detection methods. BioRxiv 2019. [Epub Ahead of Print]. - PubMed
    1. Huang W, Hennrick K, Drew S. A colorful future of quantitative pathology: validation of Vectra technology using chromogenic multiplexed immunohistochemistry and prostate tissue microarrays. Hum Pathol. 2013;44(1):29–38. - PubMed

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