Update on GIST Research Michael Heinrich, M.D.

Overview PDGFRA D842V mutant GIST Clinical background Lack of effective conventional therapy Development of an active compound Pre-clinical studies Newly opened phase 2 study Update on other OHSU GIST Clinical Studies

KIT and PDGFRA Mutations in >2000 GISTs (Heinrich-Corless Lab) Overall Mutation Frequency: 86% KIT (78.5%) Exon 14 (rare) PDGFRA (7.5% total) Exon 12 (2%) Exon 18 (5.5%) (35% of KIT-WT) Exon 9 (9%) Over 80% of GIST have KIT gene mutations. Exon 11: Mutations in the intracellular juxtamembrane region occur in mast cell tumors and GIST. Comprise ~70% of all mutations in GIST Are generally point mutations (missense), in-frame deletions, or duplications Are associated with constitutive ligand-independent receptor dimerization and activation of the kinase domain (gain-of-function mutations) Occur more frequently in high-risk GIST than in intermediate-risk GIST (and are rarely or never found in leiomyomas or leiomyosarcomas) Predict a poor prognosis: 49% 5-year patient survival rate vs 86% 5-year survival rate of patients with non–exon 11 mutations Exon 9: Mutations in the extracellular juxtamembrane region involve a similar activation mechanism. Exon 13: Mutations in the split tyrosine kinase domain also occur in GIST. Other KIT mutations have been identified in other diseases. Exon 17: Mutations in the phosphotransferase domain occur in mast cell and germ cell tumors and result in tyrosine kinase activation that requires no dimerization. Exon 17 mutations render these tumors resistant to imatinib mesylate. Exon 2: A mutation in the proximal extracellular domain has been identified in 2 hematologic disorders: myelofibrosis and chronic myeloid leukemia (CML). Seven percent of GIST have PDGFRA gain-of-function intragenic mutations. Exon 11 (67%) Exon 13 (1%) Exon 17 (1%) Taniguchi M, Nishida T, Hirota S, et al. Effect of c-kit mutation on prognosis of gastrointestinal stromal tumors. Cancer Res. 1999;59:4297-4300. Hirota S, Isozaki K, Moriyama Y, et al. Gain-of-function mutations of c-kit in human gastrointestinal stromal tumors. Science. 1998;279:577-580. Lasota J, Jasinski M, Sarlomo-Rikala M, Miettinen M. Mutations in exon 11 of c-Kit occur preferentially in malignant versus benign gastrointestinal stromal tumors and do not occur in leiomyomas or leiomyosarcomas. Am J Pathol. 1999;154:53-60. Heinrich MC, Rubin BP, Longley BJ, Fletcher JA. Biology and genetic aspects of gastrointestinal stromal tumors: KIT activation and cytogenetic alterations. Hum Pathol. 2002;33:484-495. Corless CL, Heinrich MC, Duensing A, et al. PDGFRA and KIT gain-of-function mutations are alternative oncogenic mechanisms in gastrointestinal stromal tumors (GIST). Proc Am Assoc Cancer Res. 2003;44. Abstract R4447.

PDGFRA exon 12 and 14 Mutations (2-3% of Adult GIST) PDGFRA exon 12 and 14 mutations are extremely sensitive to imatinib in vitro Sensitivity is similar to KIT exon 11 mutations Most commonly found in gastric tumors and may be associated with a reduced risk of recurrence Limited clinical data, but appears sensitive to imatinib Exon 12 Exon 14 Heinrich et al. JCO 21:4342, 2003 Debiec-Rychter et al. EJC 40:689. 2004 Debiec-Rychter et al. EJC 42:1093, 2006 Heinrich et al. JCO 26:5360, 2008 Corless et al. JCO 23:5357 2005

PDGFRA exon 18 D842V (~5% of Adult GISTs) PDGFRA D842V is the single most common PDGFRA mutation Vast majority are found in gastric GISTs May be associated with a lower overall risk of recurrence Resistant to imatinib in vitro Heinrich et al. JCO 21:4342, 2003

PDGFRA exon 18 D842V Imatinib sensitive Imatinib resistant In vitro, PDGFRA D842V mutation is resistant to imatinib, sunitinib, nilotinib, and sorafenib ? Sensitive to high dose dasatinib in vitro Biron et al. Abstract 10051, ASCO 2010 Heinrich et al. JCO 26:5360, 2008 Debiec-Rychter et al. Clin Cancer Res. 14:5749, 2008

Imatinib Treatment of D842V-mutant GIST PDGFRA D842V-mutant GIST are under represented in the major imatinib clinical studies These tumors may be less aggressive and have a lower incidence of recurrence These tumors are often “KIT-negative” and were therefore not eligible for clinical studies Imatinib clinical studies Phase 3 studies 0/8 patients responded B2222 Phase 2 study 0/3 patients responded Sunitinib clinical studies 0/3 responses for primary PDGFRA D842V mutant GIST 0/1 responses for secondary PDGFRA D842V mutant GIST

PDGFRA D842V Mutant GIST Survey of European GIST centers 0/19 patients with PDGFRA D842V mutant GIST had a response to imatinib Progression free survival of 2.8 months Median survival of 12.7 months To date, 0/33 patients in the literature with PDGFRA D842V mutation have responded to imatinib This particular subtype of GIST has not benefited from the “Gleevec revolution”

Dosing Recommendations for KIT-mutant GIST Imatinib 400 mg Imatinib 800 mg Previous studies have identified recurrent sites of gain-of-function mutations of KIT in approximately 80-85% of GISTs. The most common site of KIT mutation is mutation of the juxtamembrane domain encoded by KIT exon 11 in 67% of GISRs, the next most common site is a portion of the extracellular domain encoded by KIT exon 9 mutated in 10% of GISTs. Mutations of KIT exons 13 and 17 are rare. More recently, studies have identified homologous mutations in the closely related platelet derived growth factor alpha (PDGFRA) protein in about 8% of GISTs. These mutations are most commonly located in exons 12 and 18. Exon 9 (9.1%) Exon 11(59.6%) Exon 13 (1.9%) Exon 17 (0.8%)

Dosing Recommendations for PDGFRA-mutant GIST Imatinib 400 mg Imatinib 800 mg D842V Clinical study Exon 12 (2.5%) Not D842V IM-sensitive Exon 14 (0.5%) Not D842V IM-resistant Clinical study Exon 18 (6.9%)

Pre-clinical development of a PDGFRA D842V mutant inhibitor Studies performed in partnership with AROG Pharmaceuticals (Dallas, Texas) First experiment Sepember 2010! Testing a novel PDGFRA inhibitor (crenolanib) against GIST-relevant mutations Crenolanib has already undergone phase I testing Good bioavailability and pharmacokinetics Well tolerated with a similar side effect profile to other similar TKIs Pfizer discontinued clinical development due to the large number of TKIs that were being developed

PDGFRA WT IC 50 Crenolanib: 11 nM IC50 Imatinib: 9 nM

D842V IC 50 Crenolanib: 7nM IC50 Imatinib: 720 nM

V561D + D842V IC 50 Crenolanib: 18 nM IC50 Imatinib: >1000 nM

Phase 2 Study of Crenolanib Primary or secondary PDGFRA D842V Open at Fox Chase Cancer Center (2 patients enrolled) Scheduled to open at OHSU by end of this month (3 patients awaiting study opening)

Other OHSU GIST trials (opening soon) Phase 3 study of regorafenib as 3rd or 4th line therapy Phase 2 study of HSP90 inhibitor + imatinib Monoclonal antibody to PDGFRA Ongoing (but closed to enrollment) Regorafenib phase 2 study Persist-5 study (5 years of adjuvant imatinib for high-risk resected GIST)

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