Small molecule inhibition of GDC-0449 refractory smoothened mutants and downstream mechanisms of drug resistance.
Inappropriate Hedgehog (Hh) signaling has been directly linked to medulloblastoma (MB), a common malignant brain tumor in children. GDC-0449 is an Hh pathway inhibitor (HPI) currently under clinical investigation as an anticancer agent. Treatment of a MB patient with GDC-0449 initially regressed tumors, but this individual ultimately relapsed with a D473H resistance mutation in Smoothened (SMO), the molecular target of GDC-0449. To explore the role of the mutated aspartic acid residue in SMO function, we substituted D473 with every amino acid and found that all functional mutants were resistant to GDC-0449, with positively charged residues conferring potential oncogenic properties. Alanine scan mutagenesis of SMO further identified E518 as a novel prospective mutation site for GDC-0449 resistance. To overcome this form of acquired resistance, we screened a panel of chemically diverse HPIs and identified several antagonists with potent in vitro activity against these GDC-0449-resistant SMO mutants. The bis-amide compound 5 was of particular interest, as it was able to inhibit tumor growth mediated by drug resistant SMO in a murine allograft model of MB. However, focal amplifications of the Hh pathway transcription factor Gli2 and the Hh target gene cyclin D1 (Ccnd1) were observed in two additional resistant models, indicating that resistance may also occur downstream of SMO. Importantly, these HPI resistant MB allografts retained their sensitivity to PI3K inhibition, presenting additional opportunities for the treatment of such tumors.
Department of Molecular Biology, Genentech Inc., South San Francisco, California 94080, USA.
Gerrit J P
GJ
Dijkgraaf
Bruno
B
Alicke
Lasse
L
Weinmann
Thomas
T
Januario
Kristina
K
West
Zora
Z
Modrusan
Dan
D
Burdick
Richard
R
Goldsmith
Kirk
K
Robarge
Dan
D
Sutherlin
Suzie J
SJ
Scales
Stephen E
SE
Gould
Robert L
RL
Yauch
Frederic J
FJ
de Sauvage
D000813:Anilides / Q000494:pharmacology*; D000818:Animals; D045744:Cell Line, Tumor; D002528:Cerebellar Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D057809:HEK293 Cells; D006801:Humans; D008527:Medulloblastoma / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008819:Mice, Nude; D008822:Mice, Transgenic; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D015513:Oncogene Proteins / Q000096:biosynthesis / Q000235:genetics; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D043562:Receptors, G-Protein-Coupled / Q000037:antagonists & inhibitors* / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D000072086:Smoothened Receptor; D015534:Trans-Activators / Q000096:biosynthesis / Q000235:genetics; D000071676:Zinc Finger Protein GLI1
D000813:Anilides; D015513:Oncogene Proteins; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt; D011725:Pyridines; D012333:RNA, Messenger; D043562:Receptors, G-Protein-Coupled; D000072086:Smoothened Receptor; D015534:Trans-Activators; D000071676:Zinc Finger Protein GLI1; C538724:HhAntag691; C491023:SMO protein, human; C491022:Smo protein, mouse