Abstract
BackgroundRecent advances in the treatment of melanoma that involve immunotherapy and B-Raf inhibition have revolutionised cancer care for this disease. However, an un-met clinical need remains in B-Raf inhibitor resistant patients where first-generation B-Raf inhibitors provide only short-term disease control. In these cases, B-Raf inhibition leads to paradoxical activation of the C-Raf – MEK – ERK signalling pathway, followed by metastasis. PDE8A has been shown to directly interact with and modulate the cAMP microdomain in the vicinity of C-Raf. This interaction promotes C-Raf activation by attenuating the PKA-mediated inhibitory phosphorylation of the kinase.MethodsWe have used a novel cell-penetrating peptide agent (PPL-008) that inhibits the PDE8A – C-Raf complex in a human malignant MM415 melanoma cell line and MM415 melanoma xenograft mouse model to investigate ERK MAP kinase signalling.ResultsWe have demonstrated that the PDE8A – C-Raf complex disruptor PPL-008 increased inhibitory C-Raf-S259 phosphorylation and significantly reduced phospho-ERK signalling. We have also discovered that the ability of PPL-008 to dampen ERK signalling can be used to counter B-Raf inhibitor-driven paradoxical activation of phospho-ERK in MM415 cells treated with PLX4032 (Vemurafenib). PPL-008 treatment also significantly retarded the growth of these cells. When applied to a MM415 melanoma xenograft mouse model, PPL-008C penetrated tumour tissue and significantly reduced phospho-ERK signalling in that domain.ConclusionOur data suggests that the PDE8A-C-Raf complex is a promising therapeutic treatment for B-Raf inhibitor resistant melanoma.
Highlights
Recent advances in the treatment of melanoma that involve immunotherapy and B-Raf inhibition have revolutionised cancer care for this disease
To determine if PPL-008 conjugates (10 μM) could suppress phospho-Mitogen-activated protein kinase (ERK) signalling in MM415 cells, pERK levels were determined via western blot
PERK was significantly reduced in the human A375 malignant melanoma cell line (BRAF V600E) following PLX treatment (1 μM) (Fig. 1b, lanes 2,3,4) with PPL-008 analogues providing no ERK inhibition as a mono-treatment (Fig. 1b, lanes 11–14 inclusive) or further ERK inhibition as a co-treatment with PLX (Fig. 1b, lanes 7–10 inclusive)
Summary
Recent advances in the treatment of melanoma that involve immunotherapy and B-Raf inhibition have revolutionised cancer care for this disease. An un-met clinical need remains in B-Raf inhibitor resistant patients where first-generation B-Raf inhibitors provide only short-term disease control In these cases, B-Raf inhibition leads to paradoxical activation of the C-Raf – MEK – ERK signalling pathway, followed by metastasis. Peptide mapping of the PDE8A-C-Raf interface allowed for the rational development of a cell penetrating peptide disrupter based on the C-Raf binding site on PDE8A [22, 23] This disrupter was found to inhibit the PDE8A – C-Raf protein-protein interaction (PPI) and significantly increase C-Raf-S259 phosphorylation while concomitantly supressing phospho-ERK signalling. This concept was verified at an organismal level in both PDE8A knock out mice and a drosophila model, where basal ERK activation was attenuated compared to wild type [20]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
