Radiation and Cancer Biology

MO 02 - Biology 1 - Immunotherapy

1023 - CX3CR1 Expressing Macrophages Infiltrate the Tumor Microenvironment and Promote Radiation Resistance in a Mouse Model of Lung Cancer

Sunday, September 15
1:50 PM - 1:55 PM
Location: Room W179

CX3CR1 Expressing Macrophages Infiltrate the Tumor Microenvironment and Promote Radiation Resistance in a Mouse Model of Lung Cancer
T. Ben-Mordechai1, Y. Lawrence1,2, Z. Symon1,2, A. Shimoni-Sebag1, S. Appel1, and U. Amit1; 1Radiation Oncology, Sheba Medical Center, Ramat Gan, Israel, 2Sackler School of Medicine, Tel Aviv University, Tel Aviv, Israel

Purpose/Objective(s): Combining radiation and immunotherapy targeting T lymphocytes is extensively studied in lung cancer treatment. Current evidence suggests that only a subset of patients will benefit, while most tumors will eventually develop resistance and progress. Tumor associated macrophages (TAM) are a significant component of the TME capable of both suppressing (M1) and promoting (M2) tumor growth based on their functional state. Chemokine receptor, CX3CR1, plays an important role in macrophage homeostasis and effector functions, however its role in the TME following radiation treatment remains unknown. We hypothesized that macrophages expressing CX3CR1, play a central role in TME after radiation therapy.

Materials/Methods: Mouse lung cancer model was performed by subcutaneously inoculating Lewis Lung Carcinoma (LLC) expressing luciferase (Luc-2) and mCherry cells in CX3CR1GFP/GFP reporter mice and CX3CR1DTR/+ mice. Tumor growth was monitored by bioluminescence Imaging and caliper. TME inflammatory composition was assessed by flow cytometry. Clonogenic and HUVEC assays were used to assess tumor survival and angiogenesis after radiation.

Results: Ten days after tumor irradiation with 8 Gy, irradiated tumors were smaller than non-treated tumors. In-vivo bioluminescent imaging and flow-cytometry demonstrated a significant influx of CX3CR1 expressing cells into the irradiated TME, notably, macrophages (F4/80+ CX3CR1+). To establish the direct effect of CX3CR1 expressing macrophages on tumor growth in-vitro, we performed a clonogenic assay, by co-culturing peritoneal macrophages with irradiated LLC cells. Eliminating CX3CR1 expressing macrophages from the culture (by negative selection), reduced LLC survival fraction by 25% (P=0.005). Tumor growth is highly dependent on angiogenesis and blood supply. Interestingly, HUVEC assay showed reduced tube formation capability after eliminating CX3CR1 expressing macrophages from the co-culture. Finally, to evaluate CX3CR1 depletion effect in-vivo, we injected LLC-mCherry-Luc2 cells subcutaneously into CX3CR1DTR/+ mice, sensitive to the diphtheria toxin, and C57BL/6J control mice. Two weeks after inoculation, tumors were irradiated with 8Gy, and mice were treated every 3 days with diphtheria toxin, leading to reduction in CX3CR1 expressing cells. Three weeks after radiation, CX3CR1 depleted mice showed reduced tumor growth. Furthermore, flow cytometry analysis showed reduction in pro-tumoral M2 macrophage population (F4/80+CD206+), with no difference in T-lymphocyte or programmed cell death-1 expressing cells.

Conclusion: CX3CR1 expressing macrophages invade the TME after radiation therapy and contribute to radiation resistance and lung cancer progression, by promoting tumor survival and angiogenesis. Thus, we purpose a novel strategy to improve radiation sensitivity by targeting the CX3CR1 expressing macrophages in the TME.

Author Disclosure: T. Ben-Mordechai: None. Y. Lawrence: Research Grant; Gateway for Cancer research. Advisory Board; celgene. committee member; RTOG. Z. Symon: None. U. Amit: None.

Uri Amit, MD, PhD, MPH

Sheba Medical Center

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Sheba Medical Center

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