ALBERT

Description: Abstract 601 Background. DCC-2036 is a novel and potent tyrosine kinase inhibitor (TKI) which binds to a novel region called the switch pocket, thereby preventing BCR-ABL from adopting a conformationally active state. Efficacy against multiple imatinib-resistant BCR-ABL mutants has been demonstrated both in vitro and in vivo (Chan et al., Cancer Cell 2011;19:556). Importantly, DCC-2036 retains full potency against the T315I mutant in preclinical efficacy studies. Methods. This study was designed to find the maximal tolerated dose (MTD) of DCC-2036 when administered daily as a single-agent on a 28-day cycle. Eligible patients included adults with Ph+ CML/ALL who were refractory/intolerant to ≥2 TKI's or were T315I positive. Initially DCC-2036 capsules were administered orally once daily (QD) at increasing dose levels. Only 1 patient was enrolled in each of the lowest dose cohorts of 57mg QD and 114 mg QD. For higher doses, 3– 6 patients were enrolled into each ascending dose cohort with standard dose limiting toxicity (DLT) rules evaluating safety in cycle 1 to determine dose escalation. A transition from unformulated capsules (C) to formulated tablets (T) occurred after the 1200 mg QD dose level. Paired blood samples were obtained for PK and PD assessments. Results. 30 patients (16 males, 14 females; median age 59, range 31 – 80) with CML including 19 in Chronic (CP); 8 in Accelerated (AP) and 3 in Blast (BP) Phase were enrolled. Enrolled patients had received 1–6 prior CML treatments, and 11 patients had the T315I mutation. To date, a total of 212.5 (median 5.6; range 0.2 – 23.4) 28-day cycles were administered over 10 dose levels either as C (7 dose levels) or T (3 dose levels). The 7 C dose levels were studied first and included 57 mg QD through 1200 mg QD. Following transition to T, evaluation continued with 100 mg QD, 100 mg twice daily (BID), and 200 mg BID. Two reversible DLTs (Grade 3 peripheral neuropathy and Grade 4 lower extremity weakness) occurred during the initial treatment cycle at the 200 mg T BID dose level. Evaluation of 6 patients at the 150 mg T BID dose level determined that dose to be the MTD. Preliminary safety data show that other Grade (Gr) 3/4 adverse events (AEs) were Gr 3 slurred speech and Gr 3 eruptive nevi. Gr 1/2 AEs included dry mouth, constipation, diarrhea, paresthesias, and retinal vein occlusion. There was 1 case of Gr 2 pancreatitis that recurred on rechallenge in a patient with previous pancreatitis with nilotinib. Preliminary responses include one major molecular response in a CP patient with T315I mutation who started on capsules and transitioned to 100 mg T QD. There was one complete cytogenetic response in a CP patient at 100 mg T BID, and one partial cytogenetic response in a CP patient who started on capsules and transitioned to 100 mg T BID. One patient with AP CML and T315I mutation had a complete hematologic response at 450 mg C QD. Another patient with AP CML had a partial hematologic response after receiving 200 mg BID for 1 cycle and then downdosing to 100 mg T BID. Four out of 8 patients receiving 100 mg tablets and evaluable for efficacy (completed 3 cycles of treatment) had responses. PK results indicate dose-related, nonlinear increases in both peak plasma concentration (Cmax) and exposure (AUC). PD results reveal both acute and steady state post-treatment reductions in phospho-protein levels on Days 1 and 8. Marked reductions in pSTAT5 and pCRKL have been observed in subjects with both CP and AP and appear to be required for clinical response. Conclusion. The MTD of DCC-2036 tablets is 150 mg BID. Preliminary results suggest that DCC-2036 is well tolerated and has anti-leukemia activity in subjects with refractory CML and T315I positive disease. PD results are consistent with inhibition of BCR-ABL signaling in this first-in-man study of a switch pocket tyrosine kinase inhibitor. Disclosures: Cortes: Novartis: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Ariad: Consultancy, Research Funding; Chemgenex: Consultancy, Research Funding; Deciphera Pharmaceuticals: Research Funding. Bixby:Novartis: Speakers Bureau; Bristol Myers Squibb: Speakers Bureau; GlaxoSmithKline: Speakers Bureau. Rafferty:Deciphera Pharmaceuticals: Employment. Berger:Deciphera Pharmaceuticals: Employment. Wise:Deciphera Pharmaceuticals LLC: Employment. Rutkoski:Deciphera Pharmaceuticals: Employment. Smith:Deciphera Pharmaceuticals: Employment. Van Etten:Deciphera Pharmaceuticals: Consultancy, Research Funding.

Description: Therapy with ATP-competitive ABL kinase inhibitors, including imatinib (Gleevec), dasatinib (Sprycel), and nilotinib (Tasigna), has revolutionized the treatment of chronic myeloid leukemia (CML), but a substantial proportion of patients develop resistance to these agents. A major mechanism of acquired resistance is mutations in the ABL kinase domain that render BCR-ABL insensitive to the drug; in particular, mutation of the ABL kinase “gatekeeper” Thr315 residue to Ile (T315I) confers pan-resistance to all ATP-competitive ABL inhibitors. To address this need, we developed and characterized a novel chemical class of compounds that bind to five structural pockets involved in the endogenous “switch” mechanism used by the ABL kinase to conformationally control its activity state. Using a structure-based drug design approach, diversity in these “switch pockets” between kinases can be exploited to develop inhibitors with high potency and specificity. The resulting ABL inhibitors have several attractive features, as they: act through a non-ATP-competitive mechanism; avoid steric clash with Ile315; inhibit purified ABL that is either unphosphorylated (switch-off) or phosphorylated (switch-on) at Tyr393 with IC50 of 0.8–4.0 nM; co-crystallize with phospho-Y393-ABL WT and T315I; and have very prolonged residency time on the kinase (off-rate ∼400 min vs. 3 min for imatinib). A lead compound, DCC-2036, is highly selective for ABL, FLT3, and SRC family kinases, and has favorable pharmacokinetic and toxicity profiles in animals. In vitro, DCC-2036 inhibited proliferation and induced apoptosis of Ba/F3 cells expressing BCR-ABL WT or several common imatinib-resistant mutants (Y253F, T315I, M351T) with IC50 from 5 to 25 nM, without appreciable inhibition of parental Ba/F3 cells in IL-3 (IC50 〉1000 nM). In Balb/c mice injected with Ba/F3-BCR-ABL T315I cells, pharmacodynamic studies indicated 〉8 hour inhibition of phospho-Stat5 and phospho-ABL in the leukemic cells following a single oral 100 mg/kg dose of DCC-2036, whereas daily dosing of DCC-2036 significantly prolonged survival. In the mouse retroviral bone marrow transduction/transplantation model of CML, DCC-2036 at 100 mg/kg/d also reduced peripheral blood leukocyte counts and significantly prolonged survival vs. vehicle-treated control mice. These results demonstrate that ABL switch pocket inhibitors are a promising new pharmacologic weapon for the treatment of de novo and drug-resistant CML, including BCR-ABL T315I.

Description: Abstract 2611 Approximately 30% of acute myeloid leukemia (AML) patients have activating mutations in FLT3, commonly internal tandem duplication (ITD) mutations, which are associated with poor survival. Although FLT3 tyrosine kinase inhibitors (TKI) such as AC220 can induce remissions, resistance-causing mutations in FLT3-ITD are known to impair the in vitro activity of first and second generation FLT3 TKIs. DCC2036 is a unique switch-pocket, non-ATP competitive (allosteric) inhibitor with low nanomolar inhibitory concentration 50 (IC50) activity against a number of tyrosine kinases including FLT3 (1.7 nM), TRKA (7.0 nM), TIE-2 (2.7 nM) and BCR-ABL (2.0 nM). DCC2036 has shown promising activity in a phase I/II clinical trial in chronic myeloid leukemia (CML), where plasma concentrations of 350 nM of DCC2036 have been safely achieved. DCC-2036 has induced clinical and molecular remissions in patients with TKI-resistant CML expressing the ‘gate-keeper’ T315I BCR-ABL mutation, as well as demonstrated activity against mutations that cause BCR-ABL conformational escape resistant (Cancer Cell. 2011;19:556). Here, we evaluated the in vitro activity of DCC2036 against FLT3-ITD in cell line model systems. In the FLT3-ITD expressing human leukemia MV4-11 and MOLM-13 cells, treatment with DCC2036 (20 to 500 nM for 24 hours) dose-dependently induced cell cycle G1-phase accumulation with decline in the S and G2/M phases. Exposure to 50 to 500 nM DCC2036 for 48 to 72 hours also dose-dependently induced apoptosis of 30 to 80 % of MV4-11 and MOLM-13, as well as induced 30 to 50% apoptosis of patient-derived primary AML cells with FLT3-ITD (n =4). This was associated with dose-dependent decline in the levels of p-FLT3, p-STAT5, p-AKT, p-ERK1/2 and Bcl-xL levels but increase in the levels of BIM and p27. In contrast, treatment with DCC2036 induced significantly lower level of apoptosis (1000nM). DCC2036 retained some activity against the clinically relevant FLT3-ITD gatekeeper mutation F691L and F691I (IC50 49 nM and 34 nM), and was similarly active against the activation loop mutations Y842C and Y842H (IC50 26–28 nM). The activation loop mutations D835V and D835Y, which are commonly detected in patients with loss of response to AC220 and are hypothesized to destabilize the kinase inactive “DFG-out” conformation, were substantially less sensitive to DCC2036 (IC50 233 nM and 196 nM, respectively). Based on our previous findings (Blood. 2005;105:1768) that FLT3-ITD is a heat shock protein (hsp) 90 client oncoprotein, we also determined the effect of co-treatment with the non-geldanamycin hsp90 inhibitor AUY922 (5 to 10 nM) (Novartis Pharmaceuticals) against the cultured and primary FLT3-ITD expressing AML cells. Co-treatment with AUY922 significantly improved the activity of DCC2036 against primary AML cells (p 〈 .05). These findings demonstrate that DCC2036 exhibits potent activity against cultured and primary AML cells with FLT-3-ITD, as well as against cellular models of FLT3-ITD with AC220-resistant gatekeeper and select activation loop mutations. The molecular basis of resistance to DCC2036 conferred by activation loop mutations at D835 is under investigation. Co-treatment with DCC-2036 and the hsp90 inhibitor AUY922 exerted higher lethal activity against cultured and primary AML cells with FLT3-ITD. Disclosures: Wise: Deciphera Pharmaceuticals LLC: Employment. Reyes:Millennium, Sanofi Aventis: Consultancy. Berger:Deciphera Pharmaceuticals: Employment. Rutkoski:Deciphera Pharmaceuticals: Employment.

Description: Abstract 1740 KIT is a receptor protein tyrosine kinase which undergoes dimerization, autophysophyralation, and activation upon binding of its ligand, stem cell factor (SCF). Such activation is critical for the growth, differentiation and survival of mast cells. By enhancing Fcƒ'RI-mediated degranulation and inducing chemotaxis, KIT also contributes to mast cell functional responses. Targeting KIT is thus an attractive approach for the management of proliferative mast cell disorders. Activating mutations in the KIT tyrosine kinase domain, most notably KIT D816V, are commonly observed in the myeloproliferative disorder, systemic mastocytosis. The D816 mutation is located within the activating-switch component of KIT. These D816 mutations render the activating-switch of KIT constitutively active and capable of binding into the cognate switch pocket of KIT. Such aberrant switch function aggressively fluxes KIT to a catalytically active conformation. Switch pocket inhibitors block access to this switch pocket, thus inhibiting the ability of the mutationally activated D816 KIT mutants to adopt active conformations. Most tyrosine kinase inhibitors in clinical development target the KIT ATP binding pocket whose structure is often conserved among kinases resulting in off-target inhibition. In contrast, switch pockets are more unique among kinases. Utilizing rational drug design, the diversity in switch pockets that KIT uses to switch between the inactive and active conformations may be targeted to fine-tune selectivity of an inhibitor against the rest of the kinome. In this study, two such KIT “switch pocket” (SP) inhibitors, DP-2976 and DCC-2618, were examined for their effects on KIT activation, mast cell activation and mast cell proliferation; in comparison to the known ATP binding pocket inhibitors, imatinib and PKC412. To explore the effects of the SP inhibitors on KIT activity, wild type KIT and KIT D816V were expressed in 293T cells and their KIT autophosphorylation status assessed. DP-2976 and DCC-2618 blocked the inherent autophosphorylation of both wild type and KIT D816V in the transfected cells in the nanomolar range. Similar results were observed on constiutively active KIT in HMC 1.1 and 1.2 cells; and in CD34+ derived human mast cells in which KIT phosphorylation was induced by exposure to SCF (10 ng/ml). SP inhibitors had minimal effects on FcεRI mediated mast cell degranulation. They did completely block the SCF potentiation of FceRI-mediated degranulation. In contrast, PKC 412 inhibited degrangulation in an SCF independent manner suggesting a more generalized kinase targeting. To assess the anti-neoplastic activity of the SP inhibitors, they were next examined in HMC1.1 and HMC1.2 cell proliferation assays. Both DP-2976 and DCC-2618 significantly reduced cell numbers through induction of apoptosis with low nanomolar IC50 values. To examine whether such observations translate into clinical efficacy, ex vivo studies were performed. The survival of primary bone marrow mast cells from mastocytosis patients was significantly reduced when exposed to the SP inhibitors; a reduction comparable to PKC412. Overall, DP-2976 and DCC-2618 “switch pocket” inhibitors offer a novel and potent KIT inhibition profile whose selectivity and dual suppression of SCF enhanced mast cell activation and KIT D816V neoplastic proliferation may provide significant therapeutic benefits. Disclosures: Wise: Deciphera Pharmaceuticals LLC: Employment. Flynn:Deciphera Pharmaceuticals LLC: Employment, Equity Ownership.