Abstract 36: Changes in cellular and molecular immune markers in the peripheral blood of patients undergoing chemotherapy and radiation for squamous cell carcinoma of head and neck: A prospective pilot study
Authors: Sampath Sagus , Won Hae Jung , Maghami Ellie , Massarelli Erminia , Vora Nayana , Vora Lalit , Kortylewski Marcin 
Published: December 2017

Abstract
Objective: In order to help further development of novel immunotherapeutic targets related to signal-transducer and activator of transcription-3 (STAT3) in squamous cell carcinoma of head and neck (SCCHN), data are needed to help understand changes in immune-mediated signaling during treatment. The purpose of this study was to analyze changes in the mRNA or protein expression of inflammatory or immunosuppressive mediators in peripheral blood mononuclear cells (PBMC) or plasma, in SCCHN patients receiving concurrent chemotherapy and radiation therapy (CRT).
Methods: This was a prospective biospecimen collection study of 15 patients with stage III-IV SCCHN who received CRT with curative intent. Blood specimens were obtained before, during weeks 1, 2, 4, 6, and 7 of CRT, and 6-12 weeks after treatment completion. Plasma and PBMC were separated from patients' blood using Vacutainer CPT tubes (BD Bioscience) within 2 hours after collection by centrifugation at 1800×g for 20 min at room temperature and stored in -80°C. Plasma samples were analyzed for 30 human cytokines and chemokines using Luminex FLEXMAP 3D system at the Clinical Immunobiology Correlative Studies (CICSL) core at City of Hope. mRNA was isolated from PBMCs acquired at the baseline and at the second week draw during RT from 6 patients and expression of 730 genes selected for immune profiling was evaluated using nCounter Immune Cell Profiling Assay, and data were analyzed using nSolver software (Nanostring).
Results: Among the plasma level of 30 cytokines tested by Luminex Assay, IL-6 showed cumulative increase during RT (one-way ANOVA, p=0.046). At 6 week post RT, IL-6 level decreased comparable to baseline level. Plasma level of IFN-gamma was significantly increased at 2 week of RT and the level remained unchanged until 12 week post CRT (p=0.0004). In contrast, we observed pro-inflammatory cytokine IL-12 was significantly decreased during CRT (p=0.0004). Plasma level of a tumor promoting cytokine HGF was significantly increased during and 12 week post CRT (p=0.0035). Comparison of gene expression profiles in PBMC from 6 patients between baseline and the second week of CRT indicates that mRNA expression of 335 immune-regulators are differentially expressed (p&lt;0.05) among 740 tested. Consistent with the plasma cytokine analysis, mRNA level of IL-6 in PBMC was 13.9 fold increased (p=0.003). Subsequently, increase in STAT3 expression was observed (1.7 fold, p=0.002) since IL-6 stimulates Janus kinase (JAK)-STAT pathway. In addition, mRNA expression of Th2 cytokines including IL-10, IL-5, and TGFb1 were significantly increased by greater than 2 folds (p&lt;0.05).
Conclusion: These results suggest CRT affects immune activation in SCCHN patients systemically, which may contribute to immune suppression and disease relapse after treatment. Our data provide an impetus for investigating STAT3-targeted therapies in SCCHN.
Citation Format: Sagus Sampath, Hae Jung Won, Ellie Maghami, Erminia Massarelli, Nayana Vora, Lalit Vora, Marcin Kortylewski. Changes in cellular and molecular immune markers in the peripheral blood of patients undergoing chemotherapy and radiation for squamous cell carcinoma of head and neck: A prospective pilot study [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; April 23-25, 2017; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(23_Suppl):Abstract nr 36.