Melanomas acquire resistance to B-RAF(V600E) inhibition by RTK or N-RAS upregulation.
Activating B-RAF(V600E) (also known as BRAF) kinase mutations occur in ∼7% of human malignancies and ∼60% of melanomas. Early clinical experience with a novel class I RAF-selective inhibitor, PLX4032, demonstrated an unprecedented 80% anti-tumour response rate among patients with B-RAF(V600E)-positive melanomas, but acquired drug resistance frequently develops after initial responses. Hypotheses for mechanisms of acquired resistance to B-RAF inhibition include secondary mutations in B-RAF(V600E), MAPK reactivation, and activation of alternative survival pathways. Here we show that acquired resistance to PLX4032 develops by mutually exclusive PDGFRβ (also known as PDGFRB) upregulation or N-RAS (also known as NRAS) mutations but not through secondary mutations in B-RAF(V600E). We used PLX4032-resistant sub-lines artificially derived from B-RAF(V600E)-positive melanoma cell lines and validated key findings in PLX4032-resistant tumours and tumour-matched, short-term cultures from clinical trial patients. Induction of PDGFRβ RNA, protein and tyrosine phosphorylation emerged as a dominant feature of acquired PLX4032 resistance in a subset of melanoma sub-lines, patient-derived biopsies and short-term cultures. PDGFRβ-upregulated tumour cells have low activated RAS levels and, when treated with PLX4032, do not reactivate the MAPK pathway significantly. In another subset, high levels of activated N-RAS resulting from mutations lead to significant MAPK pathway reactivation upon PLX4032 treatment. Knockdown of PDGFRβ or N-RAS reduced growth of the respective PLX4032-resistant subsets. Overexpression of PDGFRβ or N-RAS(Q61K) conferred PLX4032 resistance to PLX4032-sensitive parental cell lines. Importantly, MAPK reactivation predicts MEK inhibitor sensitivity. Thus, melanomas escape B-RAF(V600E) targeting not through secondary B-RAF(V600E) mutations but via receptor tyrosine kinase (RTK)-mediated activation of alternative survival pathway(s) or activated RAS-mediated reactivation of the MAPK pathway, suggesting additional therapeutic strategies.
Division of Dermatology/Department of Medicine, UCLA's Jonsson Comprehensive Cancer Center, 52-121 CHS, Los Angeles, California 90095-1750, USA.
Ramin
R
Nazarian
Hubing
H
Shi
Qi
Q
Wang
Xiangju
X
Kong
Richard C
RC
Koya
Hane
H
Lee
Zugen
Z
Chen
Mi-Kyung
MK
Lee
Narsis
N
Attar
Hooman
H
Sazegar
Thinle
T
Chodon
Stanley F
SF
Nelson
Grant
G
McArthur
Jeffrey A
JA
Sosman
Antoni
A
Ribas
Roger S
RS
Lo
D001483:Base Sequence; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D011905:Genes, ras / Q000235:genetics*; D006801:Humans; D007211:Indoles / Q000494:pharmacology / Q000627:therapeutic use; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008545:Melanoma / Q000188:drug therapy* / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors; D009154:Mutation / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D047428:Protein Kinase Inhibitors / Q000494:pharmacology / Q000627:therapeutic use; D048493:Proto-Oncogene Proteins B-raf / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D020797:Receptor, Platelet-Derived Growth Factor beta / Q000096:biosynthesis / Q000235:genetics; D013449:Sulfonamides / Q000494:pharmacology / Q000627:therapeutic use; D015854:Up-Regulation / Q000187:drug effects; D000077484:Vemurafenib
D007211:Indoles; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; D000077484:Vemurafenib; D020794:Receptor Protein-Tyrosine Kinases; D020797:Receptor, Platelet-Derived Growth Factor beta; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D020929:Mitogen-Activated Protein Kinase Kinases