Abstract
BackgroundA molecular linkage between the MAPK and the LKB1-AMPK energy sensor pathways suggests that combined MAPK oncogene inhibition and metabolic modulation of AMPK would be more effective than either manipulation alone in melanoma cell lines.Materials and methodsThe combination of the BRAF inhibitor vemurafenib (formerly PLX4032) and metformin were tested against a panel of human melanoma cell lines with defined BRAF and NRAS mutations for effects on viability, cell cycle and apoptosis. Signaling molecules in the MAPK, PI3K-AKT and LKB1-AMPK pathways were studied by Western blot.ResultsSingle agent metformin inhibited proliferation in 12 out of 19 cell lines irrespective of the BRAF mutation status, but in one NRASQ61K mutant cell line it powerfully stimulated cell growth. Synergistic anti-proliferative effects of the combination of metformin with vemurafenib were observed in 6 out of 11 BRAFV600E mutants, including highly synergistic effects in two BRAFV600E mutant melanoma cell lines. Antagonistic effects were noted in some cell lines, in particular in BRAFV600E mutant cell lines resistant to single agent vemurafenib. Seven out of 8 BRAF wild type cell lines showed marginally synergistic anti-proliferative effects with the combination, and one cell line had highly antagonistic effects with the combination. The differential effects were not dependent on the sensitivity to each drug alone, effects on cell cycle or signaling pathways.ConclusionsThe combination of vemurafenib and metformin tended to have stronger anti-proliferative effects on BRAFV600E mutant cell lines. However, determinants of vemurafenib and metformin synergism or antagonism need to be understood with greater detail before any potential clinical utility of this combination.
Highlights
Exclusive mutations in NRAS and BRAF provide oncogenic driver signals in melanoma of skin origin through the constitutive activation of the mitogen-activated protein kinase (MAPK) pathway [1,2,3]
In agreement with prior studies [6,16], single agent vemurafenib induced the expected growth inhibition restricted to BRAFV600E mutant cell lines, while NRASQ61 or double wild type cell lines were resistant (Table 1 Figure 1a)
The differential effects of this combination could not be attributed to known oncogenic mutations nor differential modulation of MAPK, PI3K/AKT or LKB1AMPK signaling pathways
Summary
Exclusive mutations in NRAS and BRAF provide oncogenic driver signals in melanoma of skin origin through the constitutive activation of the mitogen-activated protein kinase (MAPK) pathway [1,2,3]. LKB1, together with low energy conditions, activates the AMP-activated protein kinase (AMPK), which results in inhibition of cell growth and proliferation. In BRAFV600E mutant melanoma cells there is an uncoupling of the LKB1-AMPK complex [9,10] These data suggest that AMPK can no longer be phosphorylated and activated by LKB1, resulting in AMPK being unable to inhibit cell growth, proliferation and survival. This uncoupling of the LKB1-AMPK complex allows BRAFV600E oncogene-driven cancer cells to become resistant to energy stress and avoid apoptosis [9]. A molecular linkage between the MAPK and the LKB1-AMPK energy sensor pathways suggests that combined MAPK oncogene inhibition and metabolic modulation of AMPK would be more effective than either manipulation alone in melanoma cell lines
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