Abstract 5375: Large, polymorphonuclear circulating cancer-associated cell with dual epithelial and macrophage/myeloid phenotype is prognostic in non-small cell lung cancer and has an impact on anti-tumor immune responses
Authors: Manjunath Yariswamy , Suvilesh Kanve N. , Avella Diego M. , Kimchi Eric T. , Staveley-O'Carroll Kevin F. , Deroche Chelsea B. , Li Guangfu , Kaifi Jussuf T. 
Published: September 2020

Purpose: In addition to the FDA-approved definition of a circulating tumor cell (CTC), various phenotypes of circulating cancer-associated cells have been described. A distinct large (&gt;30µm), polymorphonuclear, circulating, cancer-associated cell (CCAC) with a dual epithelial and macrophage/myeloid phenotype has been identified in solid cancer patients. Aims of the study were to identify and characterize these hybrid CCACs in non-small cell lung cancer (NSCLC) patients and to determine their impact on cancer-associated immune responses in vitro and in vivo in a murine model of lung cancer.

Experimental Design: In a prospective trial, 7.5ml of peripheral blood was collected from 167 subjects. CTCs and CCACs were enriched by size-based microfilter and immunostaining was performed (CK 8/18/19, EpCAM, CD14/45, DAPI). To mimic circulating hybrid cells, polyethylene glycol was used to induce fusion of murine RAW 264.7 macrophage cells and Lewis Lung Carcinoma (LLC) cells; fused cells were validated by flow cytometry, electron microscopy and fluorescence microscopy. Immunomodulatory effects of fused cells were determined in LLC-tumor bearing mice.

Results: 78 NSCLC patients and 89 control subjects (69 chronic smokers with and without lung nodules on screening LDCT, 20 healthy never-smokers) were enrolled. Distinct, large, polymorphonuclear CK+/EpCAM+/CD14/45+ CCACs were detected at lower rates than traditional CK+/EpCAM+/CD45- CTCs, but significantly higher in subjects with NSCLC in comparison to controls (p&lt;0.05). CCACs and traditional CTCs were absent in 20 healthy controls. ≥2 CCACs correlated with significantly poorer survival and higher recurrence rates by uni- and multivariate analysis (p&lt;0.05). To support these findings, PEG-induced fusion cells suppressed immune responses decreased T-cell activation and increased Tregs in vitro and in LLC tumor-bearing mice.

Conclusions and future perspectives: These investigations show that CCAC with a dual epithelial and macrophage/myeloid phenotype is associated with aggressive clinical behavior in NSCLC. Pro-tumorigenic mechanisms are suggested by macrophage-tumor cell fusion leading to suppressed T cell-mediated anti-tumor immune responses. In future, the impact of CCACs on tumor progression and immune responses will need to be further studied in NSCLC patients.

CCACs presentCCACs mean (±SEM); median (range)CTCs presentCTCs mean (±SEM); median (range)p valueSubjects analyzed (total)167NSCLC7861 (78.2%)3.12 (±0.37); 2 (0-17)78 (100%)21.95 (±1.24); 19 (8-80)&lt;0.05† (CCACs vs. CTCs)Median Age (range)69 (41-81)Gender• Females (%)41 (52.6%)• Males (%)37 (47.4%)Stage (AJCC 8th ed.)&lt;0.05* (CCACs vs. CTCs)• I39 (50%)26 (66.7%)1.26 (±0.21); 1 (0-5)39 (100%)14.59 (±0.62); 14 (8-28)• II15 (19.2%)11 (73.3%)2.67 (±0.64); 2 (0-7)15 (100%)22.67 (±1.12); 23 (18-33)• III12 (15.4%)12 (100%)6.17 (±0.81); 6 (2-11)12 (100%)29.42 (±1.58); 29 (23-42)• IV12 (15.4%)12 (100%)6.67 (±1.21); 5 (3-17)12 (100%)37.5 (±4.23); 36 (25-80)(p&lt;0.05*)(p&lt;0.05*)Control long-term smoker subjects69Median Age (range)63 (54-74)Gender• Females (%)32 (46.4%)• Males (%)37 (53.6%)No lung nodules14007 (50%)1.36 (±0.45); 0 (0-4)Benign lung nodules5511 (20%)0.33 (±0.09); 0 (0-2)41 (74.5%)3.26 (±0.39); 3 (0-11)Healthy, never-smoker controls200 (0%)00 (0%)0

Citation Format: Yariswamy Manjunath, Kanve N. Suvilesh, Diego M. Avella, Eric T. Kimchi, Kevin F. Staveley-O'Carroll, Chelsea B. Deroche, Guangfu Li, Jussuf T. Kaifi. Large, polymorphonuclear circulating cancer-associated cell with dual epithelial and macrophage/myeloid phenotype is prognostic in non-small cell lung cancer and has an impact on anti-tumor immune responses [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5375.