Elevated NR2F1 underlies the persistence of invasive disease after treatment of BRAF-mutant melanoma
Manoela Tiago, Timothy J. Purwin, Casey D. Stefanski, Renaira Oliveira da Silva, Mitchell E. Fane, Yash Chhabra, Jelan I. Haj, Jessica L.F. Teh, Rama Kadamb, Weijia Cai, Sheera R. Rosenbaum, Vivian Chua, Nir Hacohen, Michael A. Davies, Jessie Villanueva, Inna Chervoneva, Ashani T. Weeraratna, Dan A. Erkes, Claudia Capparelli, Julio A. Aguirre-Ghiso, Andrew E. Aplin
Manoela Tiago, Timothy J. Purwin, Casey D. Stefanski, Renaira Oliveira da Silva, Mitchell E. Fane, Yash Chhabra, Jelan I. Haj, Jessica L.F. Teh, Rama Kadamb, Weijia Cai, Sheera R. Rosenbaum, Vivian Chua, Nir Hacohen, Michael A. Davies, Jessie Villanueva, Inna Chervoneva, Ashani T. Weeraratna, Dan A. Erkes, Claudia Capparelli, Julio A. Aguirre-Ghiso, Andrew E. Aplin
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Research Article Oncology

Elevated NR2F1 underlies the persistence of invasive disease after treatment of BRAF-mutant melanoma

Abstract

Despite the success of targeted inhibitors in cutaneous melanoma, therapeutic responses are limited by the aged tumor microenvironment and drug-tolerant residual cells. Given the similarities between drug tolerance and cellular dormancy, we studied the dormancy marker, nuclear receptor subfamily 2 group F member 1 (NR2F1), in response to BRAF-V600E inhibitors (BRAFi) plus MEK inhibitors (MEKi) in BRAF-mutant melanoma models. Transcriptomic analysis of melanoma patient samples treated with BRAFi + MEKi showed increased NR2F1. NR2F1 was highly expressed in the drug-tolerant invasive cell state of minimal residual disease in patient-derived and mouse-derived xenografts on BRAFi + MEKi. NR2F1 over-expression was sufficient to reduce BRAFi + MEKi effects on tumor growth in vivo, and cell proliferation, death, and invasion in vitro. Effects were linked to genes involved in mTORC1 signaling. These cells were sensitive to the combination of BRAFi, MEKi plus rapamycin. Melanomas from aged mice, known to exhibit decreased responses to BRAFi + MEKi, displayed higher levels of NR2F1 compared to tumors from young mice. Depleting NR2F1 in an aged mouse melanomas improved the response to targeted therapy. These findings show high NR2F1 expression in ‘invasive-state’ residual cells and that targeting NR2F1-high cells with mTORC1 inhibitors may improve outcomes in patients with melanoma.

Authors

Manoela Tiago, Timothy J. Purwin, Casey D. Stefanski, Renaira Oliveira da Silva, Mitchell E. Fane, Yash Chhabra, Jelan I. Haj, Jessica L.F. Teh, Rama Kadamb, Weijia Cai, Sheera R. Rosenbaum, Vivian Chua, Nir Hacohen, Michael A. Davies, Jessie Villanueva, Inna Chervoneva, Ashani T. Weeraratna, Dan A. Erkes, Claudia Capparelli, Julio A. Aguirre-Ghiso, Andrew E. Aplin

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Figure 1

NR2F1 is highly expressed in melanoma lesions of patients on BRAFi + MEKi therapy.

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NR2F1 is highly expressed in melanoma lesions of patients on BRAFi + MEK...
(A) Normalized expression of NR2F1 for baseline or before treatment (Pre) and early during treatment (On) tumor samples from patients who received BRAFi + MEKi combination therapy in datasets from Song et al. (32) and Kwong et al. (30). (B) Box plot of NR2F1 RNA-Seq gene expression data for melanoma cell lines categorized across cell states according to the dataset from Tsoi et al. (33). (C) Violin plot of NR2F1 expression levels by cell state in a scRNA-Seq dataset of PDX melanoma following BRAFi + MEKi treatment, according to data from Rambow et al. (12). Cell types present during MRD are shown. NCSC, neural crest stem cell; SMC, “starved-like” melanoma cell. (D) Tumor volume in mice bearing 1205Lu-tdTomato–labeled xenografts following continuous BRAFi + MEKi (PLX4720 200 ppm + PD0325901 7 ppm) for 3 weeks. (E) Representative images of detection of tdTomato fluorescence representing tumor size in xenografts after 3 weeks on BRAFi + MEKi therapy and plot showing the mean nuclear intensity of NR2F1 protein expression in 1205Lu-tdTomato cells by immunofluorescence of tumor xenografts compared with the no-drug-treatment control group. Scale bars: 100 µm. Data are presented as the mean ± SD. *P < 0.05 and ***P < 0.001, by unpaired, 2-tailed Student’s t test.