Molly M. Hood

Discovery of 1-(3,3-Dimethylbutyl)-3-(2-fluoro-4-methyl-5-(7-methyl-2-(methylamino)pyrido[2,3-d]pyrimidin-6-yl)phenyl)urea (LY3009120) as a Pan-RAF Inhibitor with Minimal Paradoxical Activation and Activity against BRAF or RAS Mutant Tumor Cells

The RAS-RAF-MEK-MAPK cascade is an essential signaling pathway, with activation typically mediated through cell surface receptors. The kinase inhibitors vemurafenib and dabrafenib, which target oncogenic BRAF V600E, have shown significant clinical efficacy in melanoma patients harboring this mutation. Because of paradoxical pathway activation, both agents were demonstrated to promote growth and metastasis of tumor cells with RAS mutations in preclinical models and are contraindicated for treatment of cancer patients with BRAF WT background, including patients with KRAS or NRAS mutations. In order to eliminate the issues associated with paradoxical MAPK pathway activation and to provide therapeutic benefit to patients with RAS mutant cancers, we sought to identify a compound not only active against BRAF V600E but also wild type BRAF and CRAF. On the basis of its superior in vitro and in vivo profile, compound 13 was selected for further development and is currently being evaluated in phase I clinical studies.

Altiratinib Inhibits Tumor Growth, Invasion, Angiogenesis, and Microenvironment-Mediated Drug Resistance via Balanced Inhibition of MET, TIE2, and VEGFR2.

Altiratinib (DCC-2701) was designed based on the rationale of engineering a single therapeutic agent able to address multiple hallmarks of cancer (1). Specifically, altiratinib inhibits not only mechanisms of tumor initiation and progression, but also drug resistance mechanisms in the tumor and microenvironment through balanced inhibition of MET, TIE2 (TEK), and VEGFR2 (KDR) kinases. This profile was achieved by optimizing binding into the switch control pocket of all three kinases, inducing type II inactive conformations. Altiratinib durably inhibits MET, both wild-type and mutated forms, in vitro and in vivo. Through its balanced inhibitory potency versus MET, TIE2, and VEGFR2, altiratinib provides an agent that inhibits three major evasive (re)vascularization and resistance pathways (HGF, ANG, and VEGF) and blocks tumor invasion and metastasis. Altiratinib exhibits properties amenable to oral administration and exhibits substantial blood–brain barrier penetration, an attribute of significance for eventual treatment of brain cancers and brain metastases. Mol Cancer Ther; 14(9); 2023–34. ©2015 AACR.

Switch control pocket inhibitors of p38-MAP kinase. Durable type II inhibitors that do not require binding into the canonical ATP hinge region

Switch control pocket inhibitors of p38-alpha kinase are described. Durable type II inhibitors were designed which bind to arginines (Arg67 or Arg70) that function as key residues for mediating phospho-threonine 180 dependant conformational fluxing of p38-alpha from an inactive type II state to an active type I state. Binding to Arg70 in particular led to potent inhibitors, exemplified by DP-802, which also exhibited high kinase selectivity. Binding to Arg70 obviated the requirement for binding into the ATP Hinge region. X-ray crystallography revealed that DP-802 and analogs induce an enhanced type II conformation upon binding to either the unphosphorylated or the doubly phosphorylated form of p38-alpha kinase.

The Selective Tie2 Inhibitor Rebastinib Blocks Recruitment and Function of Tie2Hi Macrophages in Breast Cancer and Pancreatic Neuroendocrine Tumors

Tumor-infiltrating myeloid cells promote tumor progression by mediating angiogenesis, tumor cell intravasation, and metastasis, which can offset the effects of chemotherapy, radiation, and antiangiogenic therapy. Here, we show that the kinase switch control inhibitor rebastinib inhibits Tie2, a tyrosine kinase receptor expressed on endothelial cells and protumoral Tie2-expressing macrophages in mouse models of metastatic cancer. Rebastinib reduces tumor growth and metastasis in an orthotopic mouse model of metastatic mammary carcinoma through reduction of Tie2+ myeloid cell infiltration, antiangiogenic effects, and blockade of tumor cell intravasation mediated by perivascular Tie2Hi/Vegf-AHi macrophages in the tumor microenvironment of metastasis (TMEM). The antitumor effects of rebastinib enhance the efficacy of microtubule inhibiting chemotherapeutic agents, either eribulin or paclitaxel, by reducing tumor volume, metastasis, and improving overall survival. Rebastinib inhibition of angiopoietin/Tie2 signaling impairs multiple pathways in tumor progression mediated by protumoral Tie2+ macrophages, including TMEM-dependent dissemination and angiopoietin/Tie2-dependent angiogenesis. Rebastinib is a promising therapy for achieving Tie2 inhibition in cancer patients. Mol Cancer Ther; 16(11); 2486–501. ©2017 AACR.

Abstract 2690: DCC-2618 is a potent inhibitor of wild-type and mutant KIT, including refractory Exon 17 D816 KIT mutations, and exhibits efficacy in refractory GIST and AML xenograft models

Introduction: KIT kinase mutations are causative of a number of human cancers, including gastrointestinal stromal tumors (GIST), systemic mastocytosis (SM), mast cell leukemia (MCL), and subtypes of melanoma and acute myeloid leukemia (AML). DCC-2618 is a robust Type II switch pocket control inhibitor which potently inhibits exon 17 KIT mutations that are resistant to conventional TKIs. Experimental procedures: DCC-2618 was tested for inhibition of KIT isoforms using a standard PK/LDH coupled spectrophotometric assay. CHO cells were transiently transfected to express mutant KIT or PDGFRα constructs. Transfected cells were treated with a range of DCC-2618 and levels of phosphorylated KIT or PDGFRα in cell lysates were determined by ELISA or western blot. Cell proliferation of several cell lines was measured using the fluorescent dye resazurin. Experiments were performed in triplicate. In vivo xenograft models were performed at Molecular Imaging, Inc. (Ann Arbor, MI) or Molecular Response, LLC (San Diego, CA). Summary of results: DCC-2618 inhibited various forms of KIT with nanomolar potency: WT (IC50 4 nM), V654A (8 nM), T670I (18 nM), D816H (5 nM), D816V (14 nM). In CHO cells transiently transfected with both single and double (primary/secondary) KIT mutants, DCC-2618 robustly inhibited exon 17, exon 9/13, exon 9/14, and exon 9/17 KIT mutants, as well as exon 11/17 KIT mutants, including exon 17 D816V, D816G, D820A, D820E, D820Y, N822K, N822Y, N822H, and Y823D primary or secondary mutations. DCC-2618 inhibited wild type KIT phosphorylation in the MO7e cell line (IC50 36 nM). DCC-2618 potently inhibited KIT activation in human GIST cell lines, including GIST T1 (exon 11 deletion, IC50 2 nM), GIST 430 (exon 11 deletion/exon 13 V654A, IC50 7 nM), and GIST 48 (exon 11 V560D/exon 17 D820A, IC50 53 nM). In the murine mastocytosis P815 cell line expressing the exon 17 D816Y mutation, DCC-2618 potently inhibited cell proliferation (IC50 2 nM). In vivo, DCC-2618 administration at 50 mg/kg afforded an ED90 for inhibition of KIT phosphorylation in the GIST T1 xenograft model, corresponding to an EC90 concentration of ∼ 470 ng/mL. When give twice daily, this oral dose resulted in almost complete tumor stasis. This dose of DCC-2618 produced tumor regressions in a patient derived xenograft (PDX) GIST expressing KIT exon 11 delW557K558/exon 17 Y823D, and also in a KIT exon 17 N822K AML xenograft model. Conclusion: DCC-2618 is a potent inhibitor of singly and doubly mutated KIT characterized by primary exon 9 or exon 11 mutations paired with secondary mutations in exons 13, 14 or 17. DCC-2618 inhibits exon 17 mutations, including the D816V mutation refractory to currently marketed KIT inhibitors. DCC-2618 has the potential to treat KIT mutant-driven cancers including GIST, systemic mastocytosis, AML, or melanoma. DCC-2618 has been selected for formal IND-enabling clinical development. Citation Format: Bryan D. Smith, Molly M. Hood, Scott C. Wise, Michael D. Kaufman, Wei-Ping Lu, Thomas Rutkoski, Daniel L. Flynn, Michael C. Heinrich. DCC-2618 is a potent inhibitor of wild-type and mutant KIT, including refractory Exon 17 D816 KIT mutations, and exhibits efficacy in refractory GIST and AML xenograft models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2690. doi:10.1158/1538-7445.AM2015-2690

A Novel Combination of Wheat Peptides and Fucoidan Attenuates Ethanol-Induced Gastric Mucosal Damage through Anti-Oxidant, Anti-Inflammatory, and Pro-Survival Mechanisms

Gastritis or peptic ulcer is believed to affect about half of people worldwide. Traditional medications can lead to adverse effects, therefore, alternative nutritional strategies are needed to prevent the development of gastric mucosal damage. A novel combination of two food-grade ingredients, wheat peptides and fucoidan (WPF), was prepared to treat male Sprague Dawley rats for 30 days before gastric mucosal damage was induced by oral administration of ethanol. The serum levels of biomarkers were determined by enzyme-linked immunosorbent assay. Biomarkers in stomach tissue were analyzed using immunohistochemistry. In addition, human gastric epithelial cell line (GES-1) was used to investigate protein expression by Western blot. WPF could attenuate ethanol-induced gastric mucosal damage in an inverse dose-dependent manner, with both ulcer index and pathological index improved. WPF increased superoxide dismutase level and decreased malondialdehyde level. WPF also decreased the levels of interleukin-8, platelet-activating factor, and Caspase 3, while increasing the levels of prostaglandin E-2, epidermal growth factor (EGF), and EGF receptor (EGFR). Furthermore, phosphorylation of EGFR and extracellular signal–regulated kinases was induced by WPF in GES-1 cells. In conclusion, the novel combination of wheat peptides and fucoidan attenuated ethanol-induced gastric mucosal damage in rats through anti-oxidant, anti-inflammatory, and pro-survival mechanisms.

Abstract A53: The specific FMS kinase inhibitor, DCC-3014, durably inhibits FMS kinase in vivo and blocks cancer bone invasiveness

Within the tumor microenvironment, tumor-associated macrophages (TAMs) rely on signaling through FMS kinase to promote tumor growth, angiogenesis, vasculogenesis, and metastasis. In addition, FMS-expressing macrophages communicate with the adaptive immune system, which can lead to cancer immunotolerance. Osteoclasts also depend on FMS kinase for differentiation and growth, and play a direct role in the ability of a tumor to metastasize to the bone and progress. Taken together, FMS kinase is an excellent target for small molecule therapy of tumor growth and bone metastasis. Using Deciphera Pharmaceuticals9 approach to kinase inhibition, potent and specific FMS kinase inhibitors have been identified that achieve single digit nanomolar inhibition in biochemical and cellular assays of FMS activity. Due to their binding mode, these inhibitors retain potency in the presence of high ATP concentrations and also exhibit long off-rates from FMS kinase. Development candidate DCC-3014 is highly selective, inhibiting no other kinases within 100-fold of FMS potency, including related kinases KIT, PDGFRs, FLT3, and VEGFR2. DCC-3014 exhibits potent and durable inhibition of FMS in vivo in a pharmacokinetic/pharmacodynamic model. In a variety of cancer models, DCC-3014 or related analogs slow tumor growth, decrease tumor-promoting macrophages, and protect against osteolytic bone invasion. DCC-3014 is currently undergoing formal preclinical development. Citation Format: Bryan D. Smith, Michael D. Kaufman, Cynthia B. Leary, Molly M. Hood, Wei-Ping Lu, Benjamin A. Turner, Subha Vogeti, Scott C. Wise, Daniel L. Flynn. The specific FMS kinase inhibitor, DCC-3014, durably inhibits FMS kinase in vivo and blocks cancer bone invasiveness. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr A53. doi:10.1158/1538-7445.CHTME14-A53

Abstract B78: Rebastinib, a small molecule TIE2 kinase inhibitor, prevents primary tumor growth and lung metastasis in the PyMT breast cancer model

Rebastinib is an inhibitor of tyrosine kinases TIE2 and ABL1. A clinical trial in chronic myeloid leukemia (CML) to inhibit BCR-ABL was previously performed but efficacy was modest and development in CML is not continuing. Rebastinib is approximately 50 times more potent as an inhibitor of TIE2 than BCR-ABL. Biomarker analysis of patients from the CML trial of rebastinib revealed that a majority displayed significant increases in circulating angiopoietin 2 (ANG2) levels. Increases in ANG2, a TIE2 ligand, have been previously reported to correlate with inhibition of the TIE2 pathway. The hypoxic tumor environment engendered by radiation therapy, cytotoxic chemotherapy, or anti-angiogenic treatments (such as anti-VEGF therapies) leads to rebound tumor vascularization by the recruitment of pro-vasculogenic TIE2 expressing monocytes (TEMs) from the bone marrow to these hypoxic tumor sites. TEMs are believed to play an important role in the revascularization of tumors after these treatments, leading to progression of residual tumor. Thus, inhibition of TEM recruitment and activity could lead to better patient outcomes. The polyoma middle-T antigen (PyMT) syngeneic mouse breast cancer model utilizes the mouse mammary tumor virus (MMTV) promoter, a breast specific promoter, to express PyMT in mouse breast tissue. In this model, primary breast cancers spontaneously occur, proliferate, and metastasize (mainly to the lungs) and lead to the death of the mice. Unlike xenografts, the PyMT model utilizes fully immunocompetent mice and metastasis in this model is known to be modulated by TEMs. Rebastinib has been evaluated in a PyMT mouse where primary tumors were allowed to reach 800 mg in size before starting treatment. Rebastinib therapy resulted in a significant decrease in the growth rate of the primary breast tumor (75%), a 71% reduction in lung metastases, a decrease in the levels of tumoral TIE2 staining by immunohistochemical analysis (IHC) and caused the remaining tumor to become necrotic. Combining rebastinib with paclitaxel resulted in a synergistic response with 90% inhibition of tumor growth and a 93% reduction of lung metastases. This presentation will focus on the biochemical, cellular and in vivo activity of rebastinib as a novel small molecule inhibitor of TIE2 kinase and its potential for prevention of primary tumor rebound and metastasis in combination with first line interventional therapy. A clinical trial exploring this activity is planned for 2013. Citation Format: Bryan D. Smith, Molly M. Hood, Michael D. Kaufman, Mark Berger, Daniel L. Flynn, Scott C. Wise. Rebastinib, a small molecule TIE2 kinase inhibitor, prevents primary tumor growth and lung metastasis in the PyMT breast cancer model. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr B78.

Abstract PR01: Rebastinib, a selective TIE2 kinase inhibitor, decreases TIE2-expressing macrophages, reduces metastasis, and increases survival in murine cancer models

In the tumor microenvironment, TIE2 expression on tissue macrophages, bone marrow derived TIE2-expressing monocytes (TEMs), osteoclasts, and vascular endothelial cells promotes tumor invasiveness, dissemination, and metastasis. Additionally, a subset of TIE2-expressing macrophages, located within specialized vascular structures known as tumor microenvironment for metastases (TMEMs), are linked to intravasation of cancer cells into circulation and dissemination to metastatic sites. Rebastinib is a picomolar inhibitor of TIE2 kinase, and exhibits an extraordinarily long off-rate from TIE2, measured to be over 24 hours in a cell-based assay. Herein, we examine the efficacy of rebastinib in the polyoma middle-T antigen (PyMT) syngeneic mouse breast cancer model. In this model, PyMT breast cancer cells are implanted in the mammary fat pad, and primary tumor growth leads to lung metastasis, which is known to be modulated by TEMs and TMEM vascular structures. We examined multiple dosing schedules of rebastinib in combination with anti-tubulin agents (ATAs). Rebastinib treatment in this model significantly ablated TEMs in the primary tumor stroma and caused a significant decrease in lung metastases. Furthermore, the combination of rebastinib with ATAs, even with once or twice weekly oral dosing of rebastinib, led to a significant further decrease in lung metastases compared to single-agent treatment with ATAs. Rebastinib also enhanced the activity of ATAs in reducing primary tumor growth and regrowth of tumor post-resection. TIE2 inhibition with targeted therapy represents a novel treatment approach for metastatic breast cancer and other cancers that rely on TEMs and TMEMs for growth and metastasis. As such, rebastinib has been selected for further clinical development in solid tumors with a Phase 1b trial being planned for 2014. This abstract is also presented as Poster A5. Citation Format: Daniel L. Flynn, Michael D. Kaufman, Cynthia B. Leary, Molly M. Hood, Wei-Ping Lu, Benjamin A. Turner, Scott C. Wise, Marc S. Rudoltz, Bryan D. Smith. Rebastinib, a selective TIE2 kinase inhibitor, decreases TIE2-expressing macrophages, reduces metastasis, and increases survival in murine cancer models. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr PR01. doi:10.1158/1538-7445.CHTME14-PR01

Abstract 3594: Conformational control of FMS kinase for treatment of human malignancies

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL FMS kinase is involved in the process of osteoclast maturation. Osteoclasts have been shown to play a role not only in arthritic diseases but also in a cancers ability to metastasize to the bone. This is especially true for breast, lung and prostate cancer. Taken together the ability to prevent osteoclast maturation and accumulation through FMS inhibition should be a good target for small molecule therapy of bone metastasis. Using Deciphera Pharmaceuticals approach to kinase inhibition, compounds have been designed that potently inhibit FMS kinase. These inhibitors can be highly selective with lead compounds inhibiting only a single kinase within 20 fold of FMS activity in a 300 kinase profile. A second class broadens the profile to include anti-angiogenic kinase targets in addition to potent FMS inhibition. This presentation will highlight the attributes and development status of these compounds for treatment of human malignancies. Decipheras FMS program has afforded potent inhibitors that achieve single digit nanomolar inhibition in biochemical assays of FMS activity. The inhibitors retain this same level of potency in the presence of high (5mM) ATP concentrations. These same compounds are also selective with some inhibiting as few as three kinases within 50 fold of FMS potency. In addition, proliferation and FMS phosphoprotein assays performed with M-NFS-60 and THP-1 cells have demonstrated excellent inhibitory profiles with achieved potencies in the low nanomolar range. In functional osteoclast differentiation models, key compounds have demonstrated single digit nanomolar inhibition as assessed by TRAP assays. In vivo evaluation of the inhibitors produced excellent tolerance in two week MTD studies. Potent and durable efficacy in pharmacokinetic/pharmacodynamic xenograft models was also observed demonstrating on target effects. In vivo models of bone invasion coupled with non-invasive translational image-based biomarkers potentially provide a powerful method for visualization and quantification of osteoclast activity and FMS inhibition. Early results in imaging of bone invasion will be presented using micro CT and fluorescent activateable probes. Decipheras FMS inhibitors show acceptable ADME properties in cell permeability, cytochrome p450 inhibition, microsomal clearance and are orally bioavailable in rat and dog. Key prototype compounds have been evaluated in a two week rat tolerability study. These data demonstrate that Decipheras technology has been used to identify potent and selective FMS inhibitors to be developed for treatment of cancers where metastasis to bone is an issue. 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 3594. doi:10.1158/1538-7445.AM2011-3594