American Association for Cancer Research
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Figures S1-12, Table 1 from BRAF Inhibition Stimulates Melanoma-Associated Macrophages to Drive Tumor Growth

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journal contribution
posted on 2023-03-31, 19:09 authored by Tao Wang, Min Xiao, Yingbin Ge, Clemens Krepler, Eric Belser, Alfonso Lopez-Coral, Xiaowei Xu, Gao Zhang, Rikka Azuma, Qin Liu, Rui Liu, Ling Li, Ravi K. Amaravadi, Wei Xu, Giorgos Karakousis, Tara C. Gangadhar, Lynn M. Schuchter, Melissa Lieu, Sanika Khare, Molly B. Halloran, Meenhard Herlyn, Russel E. Kaufman

Figures S1-12, Table 1. Figure S1: Macrophages confer melanoma cell resistance to PLX4720 and Dabrafenib. Figure S2: Macrophages activate MAPK signaling, but do not activate CRAF, ARAF, NF- κB, PDGFRβ and STAT3 signaling in melanoma cells. Figure S3: Effects of growth factors on melanoma cell growth in the presence of PLX4720. Figure S4: Effects of VEGF on BRAFi-induced growth inhibition and cell death. Figure S5: The effect of macrophages on expression of VEGF receptors in melanoma cells. Figure S6. Blockade of VEGF signaling with a VEGFR inhibitor, brivanib alaninate reverses macrophagemediated resistance to PLX4720. Figure S7: Blockade of VEGF signaling with an anti-VEGF antibody reverses macrophagemediated resistance to PLX4720. Figure S8: BRAF inhibition elicits potent effects in macrophages. Figure S9: Effects of combination therapy of PLX4720 and GW2580 on melanoma growth in a xenograft model. Figure S10: Targeting macrophages with a M-CSFR inhibitor, GW2580, has a modest effect on melanoma cell growth and cell death. Figure S11: Effects of GW2580 and/or PLX4720 on tumor angiogenesis and macrophage infiltration. Figure S12: Positive control for Ki67 and CD163 staining. Table 1. Summary of melanoma patient clinical information



Purpose: To investigate the roles of melanoma-associated macrophages in melanoma resistance to BRAF inhibitors (BRAFi).Experimental Design: An in vitro macrophage and melanoma cell coculture system was used to investigate whether macrophages play a role in melanoma resistance to BRAFi. The effects of macrophages in tumor resistance were examined by proliferation assay, cell death assay, and Western blot analyses. Furthermore, two mouse preclinical models were used to validate whether targeting macrophages can increase the antitumor activity of BRAFi. Finally, the number of macrophages in melanoma tissues was examined by immunohistochemistry.Results: We demonstrate that in BRAF-mutant melanomas, BRAFi paradoxically activate the mitogen-activated protein kinase (MAPK) pathway in macrophages to produce VEGF, which reactivates the MAPK pathway and stimulates cell growth in melanoma cells. Blocking the MAPK pathway or VEGF signaling then reverses macrophage-mediated resistance. Targeting macrophages increases the antitumor activity of BRAFi in mouse and human tumor models. The presence of macrophages in melanomas predicts early relapse after therapy.Conclusions: Our findings demonstrate that macrophages play a critical role in melanoma resistance to BRAFi, suggesting that targeting macrophages will benefit patients with BRAF-mutant melanoma. Clin Cancer Res; 21(7); 1652–64. ©2015 AACR.

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