Carol Waghorne

A phase I dose-escalation study of Tivantinib (ARQ 197) in adult patients with metastatic solid tumors.

Background: Tivantinib, an oral, non-ATP competitive, selective c-MET inhibitor, exhibited antitumor activity in preclinical models. This open-label, phase I, dose-escalation study evaluated the safety, tolerability, pharmacokinetics, and pharmacodynamics of tivantinib in patients with advanced or metastatic solid tumors refractory to standard therapy. Methods: Thirteen dose levels of tivantinib ranging from 10 to 360 mg twice a day were administered to patient cohorts in 21-day cycles (14 days on/7 days off); three active pharmaceutical ingredient forms of tivantinib (amorphous, crystalline A, and crystalline B) were also investigated. Treatment was continued until the occurrence of unacceptable toxicity, tumor progression, patient withdrawal, or death. Results: A total of 79 patients with advanced solid tumors were enrolled. A maximum tolerated dose was not determined. Tivantinib was well tolerated, with mild to moderate toxicities. Two patients discontinued the study drug due to treatment-emergent adverse events. Dose-limiting grade of 3 or more toxicities including leukopenia, neutropenia, thrombocytopenia, vomiting, and dehydration, were observed in 2 patients treated with tivantinib 360 mg twice a day. The rate of absorption of tivantinib peaked approximately 2 to 4 hours after initial dosing, followed by a linear decrease in plasma concentrations. Increases in tivantinib exposure were not dose proportional. There was significant interpatient pharmacokinetic variability; however the clinical safety of tivantinib seemed unaffected. Three patients (3.8%) achieved a partial response and 40 patients (50.6%) maintained stable disease for a median of 19.9 weeks. Conclusions: Tivantinib 360 mg twice a day was well tolerated in patients with refractory advanced solid tumors. The results of this trial warrant further clinical investigation. Clin Cancer Res; 17(24); 7754–64. ©2011 AACR.

Abstract 1729: An exploratory biomarker analysis evaluating the effect of the c-MET inhibitor tivantinib (ARQ 197) and erlotinib in NSCLC patients in a randomized, double-blinded phase 2 study

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Background: A phase 2, randomized study of erlotinib administered with or without tivantinib, a selective c-MET inhibitor, in previously treated patients with non-small cell lung cancer (NSCLC) showed a significant overall survival benefit associated with the combination of tivantinib and erlotinib in patients with nonsquamous cell histology (adjusted HR = 0.58; 95% CI, 0.34-0.99; P = 0.04). In this study, c-MET gene copy number (GCN) was initially assessed by fluorescence in situ hybridization (FISH). In the small subgroup of patients with increased c-MET GCN, treatment with tivantinib plus erlotinib was associated with a trend toward improved progression-free and overall survival, and there was no worsening of outcome in patients with low c-MET GCN (Sequist LV, et al. J Clin Oncol. 2011). Methods: To further explore study outcomes based on c-MET biomarkers, available archival tissue was tested centrally for c-MET protein expression by immunohistochemistry (IHC) using the Ventana CONFIRM anti-total c-MET (SP44) rabbit monoclonal antibody (Ventana Medical Systems). Fifty of 167 patients had evaluable tumor samples for IHC evaluation. Staining intensity was scored on a scale of 0, 1+, 2+, 3+. Samples that scored ≤ 2+ in ≤ 50% of tumor were considered c-MET positive (+). Result: Among 50 evaluable patients, 27 (54%) of tumors were c-MET+. Among 33 patients with nonsquamous tumors, 25 (75%) were c-MET+, whereas only 2 of 17 (12%) squamous tumors were c-MET+, which is consistent with the existing literature (Ma PC, et al. Cancer Res. 2005). Among patients with nonsquamous histology who had c-MET+ tumors by IHC (n=25), treatment with tivantinib plus erlotinib was associated with improved progression-free survival (HR = 0.58; P = 0.28) and overall survival (HR = 0.46; P = 0.21) compared with erlotinib plus placebo. Importantly, there was no detrimental outcome in patients with c-MET-negative nonsquamous tumors (n = 8) for progression-free survival (HR = 0.36; P = 0.29) or overall survival (HR = 0.58; P = 0.55). Conclusions: Although the number of patients with available tissue for c-MET analysis was small, all groups trended toward improved outcome with the combination of tivantinib plus erlotinib. There were no statistically significant findings in this IHC analysis, but no apparent signal of harm was observed in patients with c-MET-negative nonsquamous NSCLC. This exploratory IHC analysis also confirmed that nonsquamous NSCLC tumors are more often positive for c-MET expression than squamous tumors. Two ongoing, randomized, phase 3 trials (MARQUEE™ and ATTENTION™) are enrolling only nonsquamous NSCLC patients, which should enrich the population de facto with c-MET+ patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1729. doi:1538-7445.AM2012-1729

Abstract 2517: Pharmacodynamic biomarkers for ARQ 736, a small molecule BRAF inhibitor

The MAPK/ERK pathway is a cascade of serine/threonine protein kinases activated by various extracellular factors to control cell growth, differentiation, migration and death. A number of somatic mutations occur that lead to constitutive activation of this pathway in multiple human cancers. BRAF mutations were detected in over 60% of melanomas and may account for a significant proportion of colon cancer patients whose tumors are resistant to anti-EGFR therapy. ARQ 736 is a novel inhibitor of BRAF and is selectively potent in killing cancer cell lines harboring at least one mutated BRAF allele. The objective of this study is to identify potential protein biomarkers for use in connection with ARQ 736 treatment. A number of proteins released by BRAF (V600E) mutant A375 human melanoma cells into conditioned media detected by antibody arrays (RD 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2517.

Abstract A278: ARQ 087, a multi-tyrosine kinase inhibitor with potent in vitro and in vivo activity in FGFR2 driven models.

Fibroblast growth factors (FGF) and their receptors (FGFR) play important roles in cell proliferation, cell differentiation, cell migration, cell survival, protein synthesis, and angiogenesis. The FGFR family consists of four genes encoding tyrosine kinase receptors (FGFR1, FGFR2, FGFR3, and FGFR4). Dysregulation of FGFR signaling has been implicated in a number of developmental syndromes as well as cancers, e.g., squamous non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), gastric, liver, breast, ovarian, endometrial, and bladder carcinomas, fueling significant interest in FGFRs as targets for therapeutic intervention. ArQule, Inc. has discovered a novel, ATP competitive class of FGFR inhibitors from which ARQ 087 emerged as a candidate for advancement into clinical development. ARQ 087 is a potent multi-kinase inhibitor with pan-FGFR activity against FGFR1, FGFR2, mutant FGFR2 (N549H), FGFR3, and FGFR4 kinases, all exhibiting IC 50 values in the low nanomolar range in biochemical assays. Ki versus FGFR1 and FGFR2 were 2.8nM and 0.68nM, respectively. ARQ 087 inhibited ectopically expressed FGFR1, 2, and 3 in COS-1 cells as well as, to a varying extent, the proliferation of BaF3 cells expressing the FGFR family of receptors (FGFR2≫FGFR1/FGFR3≫FGFR4). Cell proliferation studies suggested a correlation of FGFR2 mRNA amplification in gastric and other cancers with associated sensitivity to treatment with ARQ 087. Along these lines, ARQ 087 demonstrated potent inhibition of FGFR2 phosphorylation in NCI-H716, Kato III, SNU-16 and MFM223 cells; all demonstrated to be driven by FGFR2. The inhibition of cell growth was associated with an ARQ 087-induced G1 cell cycle arrest and subsequent induction in apoptosis that appears to be related to the levels of FGFR2 protein. Cell lines driven by FGFR2 activating mutations did not undergo apoptosis but did accumulate in G1 following ARQ 087 treatment. In vivo, ARQ 087 induced regressions in FGFR2-driven xenograft models (SNU-16, NCI-H716 and BaF3/FGFR2) and inhibited tumor progression in a model harboring an FGFR2-activating mutation (AN3CA). In addition, concentration-dependent inhibition of phosphorylation of FGFR2 and the downstream FGFR pathway signals (FRS2α, MEK, ERK, and AKT) was evident in response to ARQ 087 treatment in both in vitro and in vivo pharmacodynamic assays. In summary, ARQ 087 is an orally bioavailable kinase inhibitor with potent in vitro and in vivo activity in FGFR2 driven models, possessing good drug-like properties. A clinical development plan including a patient selection strategy is defined and the drug is currently in Phase I clinical studies. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A278. Citation Format: Daniel Dransfield, Jennifer Lee, Carol Waghorne, Cathy Bull, Ronald E. Savage, Xiaolan Zhao, Shipeng Yuan, Edward Chang, Enkeleda Nakuci, Sudharshan Eathiraj, Susan Cornell-Kennon, Xiubin Gu, Syed Ali, Chang-Rung Chen. ARQ 087, a multi-tyrosine kinase inhibitor with potent in vitro and in vivo activity in FGFR2 driven models. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A278.

Abstract A230: Discovery and characterization of ARQ 092, an ATP-independent, potent and selective inhibitor of AKT kinases.

The AKT family of serine-threonine kinases is a critical signal transduction node serving a variety of cellular functions including survival, proliferation, protein synthesis, and glucose metabolism. Small molecule inhibitors of AKT are emerging as potential targeted agents to treat cancers that exhibit aberrant AKT pathway signaling. A screening strategy was employed to identify inhibitors which utilize the intrinsic negative regulatory function of hydrophobic clusters in the ATP-binding cleft to inhibit the kinase activity of AKT1. ARQ 092 was identified through this screening paradigm following in vitro and in vivo optimization and was selected for further development. Crystallographic studies provided evidence that this mechanism of inhibition of AKT had been achieved. ARQ 092 binds to inactive, unphosphorylated AKT1 with sub-nanomolar affinity (Kd = 0.28 nM measured by surface plasma resonance) and biochemically inhibits all three isoforms (IC50 values of 3 nM, 4.5 nM, and 5.5 nM respectively for AKT1, AKT2, and AKT3). ARQ 092 displayed inhibition kinetics against AKT1 which were not affected by ATP concentrations up to 1000 M. When screened against a panel of over 300 kinases, ARQ 092 inhibited only three kinases within 100-fold of its IC50 against AKT1. ARQ 092 potently inhibited AKT phosphorylation (Ser473 & Thr308) in AN3CA human endometrial carcinoma cells (EC50 values of 39 nM and 61 nM, respectively for p-Ser473 & p-Thr308) and demonstrated concentration-dependent inhibition of phosphorylation of the downstream AKT substrate PRAS40. ARQ 092 inhibited growth of AN3CA cells (GI50 = 555 nM), A2780 cells (GI50 = 400 nM), and IGROV-1 cells (GI50 = 66 nM), in addition to LNCaP, ZR-75–1, and BT-474 human cancer cell lines (GI50 values ranging from 1 to 4 μM). Following a single oral dose of ARQ 092, inhibition of AKT and PRAS40 phosphorylation was documented in vivo in both AN3CA human endometrial tumor and BT474 human breast tumor xenograft models. The growth of AN3CA tumor xenografts was markedly suppressed after daily oral administration of ARQ 092 for 10 days. Finally, ARQ 092 was shown to have good pharmaceutical properties and was advanced into preclinical development. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr A230.

Abstract 3571: Exploratory biomarker discovery for clinical development of ARQ 087, a potent pan-FGFR kinase inhibitor

Fibroblast growth factor receptor (FGFR) tyrosine kinase family members have gained increasing attention as potential therapeutic targets. Gene amplification, or gain-of-function translocations and mutations of these receptors, have been implicated in a variety of cancers including breast, gastric, bladder, ovarian, endometrial, lymphoma and myeloma. We have previously described the biological profiling of ARQ 087, a potent small-molecule pan-FGFR kinase inhibitor that shows anti-tumor activity in FGFR dysregulated tumor cells in vitro and in vivo. A number of cell lines with genetically-altered FGF receptors were identified that were sensitive in anti-proliferative assays to ARQ 087, including FGFR2 amplified gastric carcinoma lines KATO III and SNU-16, and endometrial lines AN3CA, MFE-296 and MFE-280 harboring FGFR2 mutants, as well as a breast cancer cell line with increased FGFR1 gene copy (MDA MB-157). Western blot pharmacodynamic analysis demonstrates that ARQ 087 inhibits FGFR phosphorylation and downstream pathway markers in these cell lines in a concentration-dependent manner. Using antibody arrays specific for known cytokines as well as regulatory proteins involved in angiogenesis and apoptosis, we have identified a number of protein biomarkers in cell lysates and in conditioned media of sensitive cell lines that changed in response to ARQ 087 exposure. Three proteins (cytochrome c, clusterin, and HTRA2) decreased in SNU-16 (ARQ 087-sensitive, FGFR amplified) but not DLD1 (ARQ 087-resistant, non FGFR expressing) cells. VEGF decreased and IL-1rα increased in SNU-16 but not in DLD1 conditioned media in response to ARQ 087. Furthermore, analysis of mouse plasma from animals bearing AN3CA tumors treated with ARQ 087 revealed a decrease in human VEGF, CXCL16, MMP-9, Serpin E1 and MIF and an increase in RANTES, IL-1β, IP-10, SDF-1, IL-1rα. TNFα, GM-CSF, IL-2, sTREM-1, sICAM-1, MIP-1β, IL-8. In summary, we have identified a number of cell lines from a broad cell line screening campaign that exhibited dysregulated FGFR pathway function and are sensitive to the anti-proliferative effects of ARQ 087. These cell lines, in turn, have been used as models in which to identify candidate biomarkers that may greatly facilitate subsequent clinical evaluation of this molecularly targeted anti-tumor agent. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3571. doi:10.1158/1538-7445.AM2011-3571

Abstract 3905: Synthesis and structure activity relationship of substituted N,6-diphenyl-5,6-dihydrobenzo[h]quinazolin-2-amine as inhibitors of fibroblast growth factor receptors (FGFR)

Utilization of hydrophobic motifs present in auto-inhibited protein kinases has resulted in the identification of a series of 5,6-dihydrobenzo [h]quinazolin-2-amines with activity as fibroblast growth factor receptor (FGFR) tyrosine kinase inhibitors. Herein we describe the combination of a proprietary in silico design process, a new screening paradigm using an array of biochemical and biophysical technologies in conjunction with an established parallel chemistry process for the identification and optimization of a series of novel FGFR inhibitors. These potent FGFR inhibitors exhibit a preference for the inactive form of the kinase, are non-ATP competitive, and exhibit robust cellular pharmacodynamic inhibition as well as in vitro anti-proliferative effects in cells dependent on FGFR and significant anti-tumor activity in appropriate xenograft models in vivo. The design strategy, synthesis, structure activity relationships and in vitro and in vivo biology of selected inhibitors will be presented. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3905. doi:1538-7445.AM2012-3905