DISCUSSION
The RAS-RAF-MEK-ERK cascade (RAS-RAF mitogen-activated protein kinase;
MAPK) is the key regulatory pathway for cell growth, proliferation,
differentiation, and apoptosis. Signaling through this pathway typically
occurs through different plasma membrane growth factor receptors that
trigger RAS family GTPases, including HRAS, NRAS and KRAS. Activated RAS
proteins can recruit members of the RAF kinase family (ARAF, BRAF, and
CRAF) to the plasma membrane, complex with and activate them. Activated
RAF kinases signal downstream to other MEK/ERK cascade elements to over
150 downstream targets, nuclear and cytosolic. The MAPK pathway is
frequently dysregulated in cancer, often via mutations of its
intracellular components or activation of growth factor receptor
tyrosine kinases. BRAF is the most potent form of RAF
kinases13. T1799A transversion mutation in BRAF
(BRAFv600) accounts for more than 80% of all known BRAF mutations and
leads to hyperactivation of the MAPK pathway12,14 that
results in deregulated downstream signaling and consequently,
unregulated cell proliferation and survival, which contributes to
oncogenesis. Patients harboring BRAF v600 mutations are treated by BRAF
inhibitors, such as vemurafenib, dabrafenib and encorafenib. However,
other BRAF mutations like Class III BRAF mutations were recently
identified as being resistant to these agents.
In our patient, we identified two somatic mutations in the RAS pathway
genes (BRAF N581I and HRAS G13V), which are mutually
exclusive15. BRAF N581I is a Class III hotspot
mutation that lies within the protein kinase domain of the Braf protein
(UniProt.org). Class III BRAF mutants display low kinase activity or are
kinase-dead. They require the co-existence of other upstream mutations
to give the oncogenic signaling. They trigger the MAPK pathway through
enhanced RAS binding and subsequent RAS-dependent CRAF activation
resulting in elevated ERK signaling and are susceptible to ERK-mediated
feedback16–18. (Supplementary Figure 4).
One study reported the cooperation of the BRAF and HRAS in histiocytic
sarcoma. In this study, the BRAF mutation Phe595Leu (class III)
co-occurred with HRAS (Q61R). The co-occurrence of these two mutations
causes a higher oncogenic signal than in the case of BRAF
alone19. This study also highlighted the efficacy of
Pan-RAF inhibitors, such as sorafenib and AZ628, and the MEK inhibitor
trametinib in interfering with the MEK/ERK phosphorylation driven by the
cooperative activity of BRAF (F595L) and oncogenic RAS. Another study
introduced the KIN-2787 as a potent and selective small-molecule pan-RAF
inhibitor specifically designed to inhibit Class II and III BRAF
dimers18.
Class III mutations are sensitive to the inhibition of RAS activation by
inhibitors of receptor tyrosine kinases. A multicenter analysis
suggested that anti-EGFR therapy has promising effects for some patients
with metastatic colorectal cancer that is not v600 BRAF mutated. The
study demonstrated that half of the patients with kinase-impaired and
RAS-dependent BRAF mutations responded to the therapy, whereas almost
none of those with kinase-activating and RAS-independent mutations
did20. In contrast, current RAF inhibitors are not
expected to effectively inhibit the ERK pathway in these tumors. It was
reported that RAF inhibitor vemurafenib failed to inhibit ERK signaling
in tumor cells that express class III BRAF mutants. However, cell lines
harboring Class III mutations are sensitive to classic RAS/MAPK singling
inhibitors 21.