Combined VEGFR and MAPK pathway inhibition in angiosarcoma

Michael J Wagner,Eric C Holland,Jean H Siedel,Anil K Sood,Robert Dood,Vinod Ravi,Monika Haemmerle,Alexander J Lazar,Lingegowda S Mangala,Archana S Nagaraja,Pritha Chanana,Yasmin A Lyons,Wei-Lien Wang

doi:10.1038/s41598-021-88703-9

Abstract

Angiosarcoma is an aggressive malignancy of endothelial cells that carries a high mortality rate. Cytotoxic chemotherapy can elicit clinical responses, but the duration of response is limited. Sequencing reveals multiple mutations in angiogenesis pathways in angiosarcomas, particularly in vascular endothelial growth factor (VEGFR) and mitogen-activated protein kinase (MAPK) signaling. We aimed to determine the biological relevance of these pathways in angiosarcoma. Tissue microarray consisting of clinical formalin-fixed paraffin embedded tissue archival samples were stained for phospho- extracellular signal-regulated kinase (p-ERK) with immunohistochemistry. Angiosarcoma cell lines were treated with the mitogen-activated protein kinase kinase (MEK) inhibitor trametinib, pan-VEGFR inhibitor cediranib, or combined trametinib and cediranib and viability was assessed. Reverse phase protein array (RPPA) was performed to assess multiple oncogenic protein pathways. SVR angiosarcoma cells were grown in vivo and gene expression effects of treatment were assessed with whole exome RNA sequencing. MAPK signaling was found active in over half of clinical angiosarcoma samples. Inhibition of MAPK signaling with the MEK inhibitor trametinib decreased the viability of angiosarcoma cells. Combined inhibition of the VEGF and MAPK pathways with cediranib and trametinib had an additive effect in in vitro models, and a combinatorial effect in an in vivo model. Combined treatment led to smaller tumors than treatment with either agent alone. RNA-seq demonstrated distinct expression signatures between the trametinib treated tumors and those treated with both trametinib and cediranib. These results indicate a clinical study of combined VEGFR and MEK inhibition in angiosarcoma is warranted.

Highlights

Abbreviations B-Raf Proto-Oncogene (BRAF) B-Raf proto-oncogene dual specificity protein phosphatase (DUSP) Dual specificity protein phosphatase Dulbecco’s Modified Eagle Medium (DMEM) Dulbecco’s modified eagle medium early growth response 1 (EGR1) Early growth response 1 ETV5 ETS variant transcription factor 5 FCGBP Fc fragment of IgG binding protein fibroblast growth factor receptor 2 (FGFR2) Fibroblast growth factor receptor 2 FLT4 FMS related receptor tyrosine kinase 4 GSEA Gene set enrichment analysis HRAS H-Ras Proto-Oncogene integrin subunit alpha 6 (ITGA6) Integrin subunit alpha 6 IL2 Interleukin 2
MAPK pathway alterations have been identified in angiosarcoma clinical samples with mutations in K-Ras Proto-Oncogene (KRAS), H-Ras Proto-Oncogene (HRAS), N-Ras Proto-Oncogene (NRAS), B-Raf Proto-Oncogene (BRAF), Raf-1 Proto-Oncogene (CRAF), mitogen-activated protein kinase 1 (MAPK1) and Neurofibromin 1 (NF1)[7]
We hypothesized that MAPK signaling would be active in clinical angiosarcoma samples, and that targeting the MAPK pathway would be effective against angiosarcoma cells

Summary

Introduction

Abbreviations BRAF B-Raf proto-oncogene DUSP Dual specificity protein phosphatase DMEM Dulbecco’s modified eagle medium EGR1 Early growth response 1 ETV5 ETS variant transcription factor 5 FCGBP Fc fragment of IgG binding protein FGFR2 Fibroblast growth factor receptor 2 FLT4 FMS related receptor tyrosine kinase 4 GSEA Gene set enrichment analysis HRAS H-Ras Proto-Oncogene ITGA6 Integrin subunit alpha 6 IL2 Interleukin 2. Initial efforts to identify drivers of angiosarcoma have revealed several recurrent aberrations in angiogenesis pathways[2,3] These include activating mutations in kinase insert domain receptor (KDR) and phospholipase C gamma 1 (PLCG1), and loss of function mutation in receptor-type tyrosine-protein phosphatase beta (PTPRB)[4,5]. The recurrent R707Q mutation in PLCG1 leads to canonical activation of the MAPK pathway[8] and leads to acquired resistance to VEGFR inhibition[9] In spite of these potentially targetable mutations, targeted agents modulating angiogenesis pathways in angiosarcoma clinical studies have overall been disappointing with response rates less than 20%10–12. We sought to further investigate the potential for targeting the MAPK pathway in angiosarcoma with dual inhibition of the VEGFR-MAPK axis

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Discussion

Conclusion