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  • Poster presentation
  • Open Access

Utilizing quantitative immunohistochemistry for relationship analysis of tumor microenvironment of head and neck cancer patients

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Journal for ImmunoTherapy of Cancer20142 (Suppl 3) :P258

  • Published:


  • Tumor Microenvironment
  • Arginase
  • Neck Squamous Cell Carcinoma
  • Neck Cancer Patient
  • Relationship Analysis


Analysis of tumor-infiltrating immune cells using quantitative immunohistochemistry (IHC) has proved to be a powerful prognostic biomarker in colon cancer [1, 2]. Similar observations have been made in patients with oral, head and neck squamous cell carcinoma (OHNSCC), where CD8 infiltration is associated with prolonged survival [3]. Recently, advancements are made in multiplex imaging and relationship analysis to better delineate suppressive mechanisms within the tumor microenvironment, which may direct immune interventions that augment tumor-specific immune response.


The purpose of this investigation was to apply multiplex immunohistochemistry and objective assessment techniques to identify biomarkers that correlate with HPV status, T cell infiltrate, and patient survival. Relationships analysis between immune markers and tumor cells will also be performed to examine the dynamic interactions that occur within the tumor microenvironment.


92 subjects with biopsy-proven OHNSCC from different sub-sites underwent surgery with curative intent and were enrolled into this prospective, IRB approved protocol. Formalin-fixed-paraffin-embedded (FFPE) samples of patients' primary tumor or metastatic lymph nodes are obtained and stained for markers including CD4, CD8, CD137, CD163, interferon-gamma, arginase I, PD-L1, and class I, using the PerkinElmer Opal system. Images are scanned and analyzed using PerkinElmer Vectra system. Single stains are being done simultaneously using Ventana Benchmark XT and analyzed using Definiens platform.


Preliminary results analyzed from 24 patients showed positive correlation between CD8 immune infiltrate within the tumor and HPV status (P = 0.05). Level of Arg1 within the tumor microenvironment showed a stronger correlation with HPV status (Figure 1), and inversely correlated with CD8 infiltrate (P = 0.03). Interestingly, the number of IFN-γ positive CD8 cells has no correlation with PD-L1 status in the subset of the patients that we have analyzed (Figure 2) - implying potential constitutive expression of PD-L1 in a subset of these patients.
Figure 1
Figure 1

Arg1: HPV correlation.

Figure 2
Figure 2

% IFN+ CD8 vs PD-L1 expression.


While still early, the technique is reproducible and can provide useful information on the relationships between various cells within the tumor microenvironment. Planned studies will assess the interplay between these markers in larger cohorts of patients with long-term follow-up, which aims to provide insights that may be exploited to develop novel therapeutic strategies that will improve outcomes of patients with OHNSCC.


Steve and Cindy Harder, Nancy and Wes Lematta, Robert W. and Elsie Franz, Lynn and Jack Loacker and The Chiles foundation.

Authors’ Affiliations

Earle A. Chiles Research Institute, Providence Cancer Center; Department of Cancer Biology, Oregon Health & Science University, Portland, OR, USA
Oral, Head and Neck Cancer Program and Clinic, Providence Cancer Center, OR, USA
UbiVac, USA
Molecular Microbiology and Immunology, OHSU, Portland, Portland, OR, OR, USA


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  2. Galon J, Costes A, Sanchez-Cabo F, Kirilovsky A: Type, density, and location of immune cells within human colorectal tumors predict clinical outcome. Science. 2006, 313-Sept 29Google Scholar
  3. Pretscher D, Distel LV, Grabenbauer GG, Wittlinger M, Buettner M, Niedobitek G: Distribution of immune cells in head and neck cancer: CD8+ T-cells and CD20+ B-cells in metastatic lymph nodes are associated with favourable outcome in patients with oro- and hypopharyngeal carcinoma. BMC Cancer. 2009Google Scholar


© Feng et al.; licensee BioMed Central Ltd. 2014

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver ( applies to the data made available in this article, unless otherwise stated.