Abstract 3310: Extracellular vesicles budding from stromal macrophages in the blood of metastatic non-small cell lung cancer patients correlates with poor survival
Authors: Moran Jillian A. , Lin Steven H. , Edelman Martin J. , Lopez Pablo , He Jianzhong , Qiao Yawei , Xu Ting , Liao Zhongxing , Gardner Kirby P. , Tang Cha-Mei , Adams Daniel L. 
Published: April 2023

Abstract
Purpose: Cancer extracellular vesicles (EVs) are involved in cellular communication, tumor growth, progression, and metastasis in cancer. The origins of EVs, their formation, and potential clinical use as biomarkers are not well understood. Recently, budding of extracellular structures on Cancer Associated Macrophage-Like Cells [CAMLs] a specific subtype of phagocytic circulating stromal cells has been observed in metastatic non-small cell lung carcinoma (mNSCLC) patients. In this prospective analysis of n=40 mNSCLC samples, we enumerated EV budding on CAMLs to determine if their formation had an effect on clinical outcomes. These preliminary results suggest that EV budding from a specific subtype of circulating tumor associated macrophage prognosticates for worse clinical outcome which may serve as the mechanism for cancer EV formation and spread throughout the body.
Patients and Methods: We initiated a single blind prospective pilot study to evaluate extracellular budding on the CAMLs of mNSCLC patients from blood samples obtained prior to therapy to determine their prevalence and clinical utility. Anonymized blood was procured and filtered to isolate CAMLs and stained for cytokeratin, CD45, CD31 and PD-L1. EV budding was observed as small (≤1 µm) bulbous protrusions from the cell periphery. EVs were quantified and compared against patient progression free survival (PFS) and overall survival (OS) with hazard ratios (HRs) at 24 months by censored univariate analysis. The imaged EVs were also characterized by their PD-L1 biomarker expression.
Results: CAMLs were identified in 88% (n=35/40) of all samples, with EV budding identified in 60% (n=21/35) of CAMLs. These EVs appeared with tumor positive proteins (i.e. CD31, CD45, PD-L1 and cytokeratin). With a minimum of 24 months of follow-up, it was determined that the presence of EV budding in patients’ CAMLs was associated with significantly worse PFS (HR=4.00, 95%CI=1.4-12, p=0.0251) and borderline significant OS (HR=3.57, 95%CI=1.1-12, p=0.0747).
Conclusions: EV budding found on phagocytic stromal cells found in the blood appear with tumor positive biomarkers and predict poor survival. These findings suggest that CAMLs are an origin cell for some cancer EVs. Larger validation studies and cross comparison of PD-L1 on EVs as it related to immunotherapy response is ongoing.
Citation Format: Jillian A. Moran, Steven H. Lin, Martin J. Edelman, Pablo Lopez, Jianzhong He, Yawei Qiao, Ting Xu, Zhongxing Liao, Kirby P. Gardner, Cha-Mei Tang, Daniel L. Adams. Extracellular vesicles budding from stromal macrophages in the blood of metastatic non-small cell lung cancer patients correlates with poor survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3310.