Abstract
Mutant serine/threonine protein kinase B-Raf (BRAF) protein is expressed in over half of all melanoma tumors. Although BRAF inhibitors (BRAFi) elicit rapid anti-tumor responses in the majority of patients with mutant BRAF melanoma, the tumors inevitably relapse after a short time. We hypothesized that polyamines are essential for tumor survival in mutant BRAF melanomas. These tumors rely on both polyamine biosynthesis and an upregulated polyamine transport system (PTS) to maintain their high intracellular polyamine levels. We evaluated the effect of a novel arylpolyamine (AP) compound that is cytotoxic upon cellular entry via the increased PTS activity of melanoma cells with different BRAF mutational status. Mutant BRAF melanoma cells demonstrated greater PTS activity and increased sensitivity to AP compared to wild type BRAF (BRAFWT) melanoma cells. Treatment with an inhibitor of polyamine biosynthesis, α-difluoromethylornithine (DFMO), further upregulated PTS activity in mutant BRAF cells and increased their sensitivity to AP. Furthermore, viability assays of 3D spheroid cultures of mutant BRAF melanoma cells demonstrated greater resistance to the BRAFi, PLX4720, compared to 2D monolayer cultures. However, co-treatment with AP restored the sensitivity of melanoma spheroids to PLX4720. These data indicate that mutant BRAF melanoma cells are more dependent on the PTS compared to BRAFWT melanoma cells, resulting in greater sensitivity to the PTS-targeted cytotoxic AP compound.
Highlights
Melanoma is a highly aggressive tumor with poor prognosis in the metastatic stage
We tested whether mutant BRAFV600E cells were more sensitive to increasing concentrations of the cytotoxic polyamine transport ligand, AP (Figure 1), compared to BRAFWT
This observation was reflected by the greater polyamine transport activity in BRAFV600E melanoma cells compared to BRAFWT
Summary
Melanoma is a highly aggressive tumor with poor prognosis in the metastatic stage. Multiple oncogenic mutations (including genes encoding serine/threonine protein kinase B-Raf (BRAF), the neuroblastoma RAS homolog (NRAS), and the proto-oncogene receptor tyrosine protein kinase KIT). Multiple oncogene-encoded proteins, including c-MYC and RAS, are known to upregulate processes, including cellular proliferation, differentiation, chromatin remodeling, hypusinationkey of polyamine biosynthetic enzymes [7,9,10]. Polyamine uptake is upregulated in many tumor types, especially in melanoma tumor cells when These intracellular polyamine levels are maintained via tightly-regulated biosynthetic, catabolic, and compared normal cells [11,20]. Polyamine uptake is tumor upregulated in many tumor types,with especially in oncogenic mutations have a greatly increased need for polyamines compared to normal cells to meet melanoma tumor cells when compared to normal cells [11,20]. The mutant BRAF-driven cytotoxic polyamine compounds, which selectively target melanoma cells withselectively high polyamine polyamine addiction can be targeted by cytotoxic polyamine compounds, which target import activity.
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