John S. Hill

A First-in-Human Study of Conatumumab in Adult Patients with Advanced Solid Tumors

Purpose: To determine the safety, tolerability, pharmacokinetics, and maximum tolerated dose (MTD) of conatumumab, an investigational, fully human monoclonal agonist antibody against human death receptor 5, in patients with advanced solid tumors. Experimental Design: In the dose-escalation phase, patients received escalating intravenous doses of conatumumab (0.3, 1, 3, 10, or 20 mg/kg, 3–9 per cohort) every 2 weeks. In the dose-expansion phase, 10 patients with colorectal cancer (CRC) and 7 with non–small cell lung cancer (NSCLC) received 20 mg/kg of conatumumab every 2 weeks. Results: Thirty-seven patients received 1 or more doses of conatumumab. Conatumumab seemed to be well tolerated; there were no dose-limiting toxicities. Of adverse events possibly related to treatment, only 3 patients (8%) had a grade 3 event (fatigue and/or elevated lipase), and no anticonatumumab antibodies were detected. An MTD was not reached. Conatumumab exhibited dose linear kinetics from 3 to 20 mg/kg, with a mean terminal half-life of 13 to 19 days. One patient with NSCLC (0.3 mg/kg) had a confirmed partial response (PR) at week 32 (38% reduction in tumor size), with further reduction (48%) by week 96; this patient remains on conatumumab after 4.2 years with a sustained PR. Fourteen patients had a best response of stable disease, 2 for 32 weeks or more. One patient with CRC (0.3 mg/kg) and stable disease for 24 weeks had a 24% reduction in tumor size by RECIST (Response Evaluation Criteria in Solid Tumors) and a 35% reduction in the sum of standardized uptake values of all lesions measured by [18F]fluorodeoxyglucose positron emission tomographic scan. Changes in tumor levels of activated caspase-3 did not appear to be associated with tumor response. Conclusions: Conatumumab can be administered safely up to the target dose of 20 mg/kg every 2 weeks. Clin Cancer Res; 16(23); 5883–91. ©2010 AACR.

AMG 595, an Anti-EGFRvIII Antibody–Drug Conjugate, Induces Potent Antitumor Activity against EGFRvIII-Expressing Glioblastoma

Epidermal growth factor receptor variant III (EGFRvIII) is a cancer-specific deletion mutant observed in approximately 25% to 50% of glioblastoma multiforme (GBM) patients. An antibody drug conjugate, AMG 595, composed of the maytansinoid DM1 attached to a highly selective anti-EGFRvIII antibody via a noncleavable linker, was developed to treat EGFRvIII-positive GBM patients. AMG 595 binds to the cell surface and internalizes into the endo-lysosomal pathway of EGFRvIII-expressing cells. Incubation of AMG 595 with U251 cells expressing EGFRvIII led to potent growth inhibition. AMG 595 treatment induced significant tumor mitotic arrest, as measured by phospho-histone H3, in GBM subcutaneous xenografts expressing EGFRvIII. A single intravenous injection of AMG 595 at 17 mg/kg (250 μg DM1/kg) generated complete tumor regression in the U251vIII subcutaneous xenograft model. AMG 595 mediated tumor regression in the D317 subcutaneous xenograft model that endogenously expresses EGFRvIII. Finally, AMG 595 treatment inhibited the growth of D317 xenografts orthotopically implanted into the brain as determined by magnetic resonance imaging. These results demonstrate that AMG 595 is a promising candidate to evaluate in EGFRvIII-expressing GBM patients. Mol Cancer Ther; 14(7); 1614–24. ©2015 AACR.

Measurement of conatumumab-induced apoptotic activity in tumors by fine needle aspirate sampling†

Conatumumab is a monoclonal antibody specific for death receptor 5 (DR5) that activates caspases leading to DNA fragmentation and tumor‐cell death. Like other Tumor Necrosis Factor‐Related Apoptosis‐Inducing Ligand (TRAIL) receptor therapies, conatumumab is currently being evaluated in clinical trials across a variety of tumor types. However, molecular evidence of on‐target drug activity in tumors is often an elusive goal for clinical investigation. Here we evaluated a translational approach using a relevant biopsy method, fine needle aspirates (FNAs), to study the on‐target pharmacodynamics of conatumumab pre‐clinically. As detected by laser scanning cytometry, drug‐induced caspase‐3 activation in FNA biopsies of Colo205 xenografts correlated well with activated caspase‐3 in conventional section‐based samples. Furthermore, in tumor‐bearing mice, surrogate assays of serum caspase‐3/7 activity and serum drug exposure correlated with in situ caspase‐3 activation. We found that one advantage of FNA sampling over other sampling techniques was the ability to measure caspase activity on a per cell basis using DNA content information. To adapt the utility of FNAs for measuring pharmacodynamic markers in humans, detection of activated caspase‐3 was multiplexed with EpCAM to characterize mock and clinical FNAs from colorectal and nonsmall cell lung cancer patients. These data suggest that FNA sampling is a practical method to cytometrically evaluate tumors for pharmacological impact in a clinical setting.

Rapid affinity measurement of protein–protein interactions in a microfluidic platform

A rapid screening method has been developed to determine binding affinities for protein-ligand interactions using the Gyrolab workstation, a commercial microfluidic platform developed to accurately and precisely quantify proteins in solution. This method was particularly suited for assessing the high-affinity interactions that have become typical of therapeutic antibody-antigen systems. Five different commercially available antibodies that bind digoxin and a digoxin-bovine serum albumin (BSA) conjugate with high affinity were rigorously evaluated by this method and by the more conventional kinetic exclusion assay (KinExA) method. Binding parameter values obtained using Gyrolab were similar to those recovered from KinExA. However, the total experimental time for 20 binding affinity titrations, with each titration covering 12 data points in duplicate, took approximately 4h by the Gyrolab method, which reduced the experimental duration by more than 10-fold when compared with the KinExA method. This rapid binding analysis method has significant applications in the screening and affinity ranking selection of antibodies from a very large pool of candidates spanning a wide range of binding affinities from the low pM to μM range.

First-in-Human Study of AMG 820, a Monoclonal Anti-Colony-Stimulating Factor 1 Receptor Antibody, in Patients with Advanced Solid Tumors

Purpose: Binding of colony-stimulating factor 1 (CSF1) ligand to the CSF1 receptor (CSF1R) regulates survival of tumor-associated macrophages, which generally promote an immunosuppressive tumor microenvironment. AMG 820 is an investigational, fully human CSF1R antibody that inhibits binding of the ligands CSF1 and IL34 and subsequent ligand-mediated receptor activation. This first-in-human phase I study evaluated the safety, pharmacokinetics, pharmacodynamics, and antitumor activity of AMG 820. Experimental Design: Adult patients with relapsed or refractory advanced solid tumors received intravenous AMG 820 0.5 mg/kg once weekly or 1.5 to 20 mg/kg every 2 weeks until disease progression, adverse event (AE), or consent withdrawal. Results: Twenty-five patients received ≥1 dose of AMG 820. AMG 820 was tolerated up to 20 mg/kg; the MTD was not reached. One dose-limiting toxicity was observed (20 mg/kg; nonreversible grade 3 deafness). Most patients (76%) had treatment-related AEs; the most common were periorbital edema (44%), increased aspartate aminotransferase (AST; 28%), fatigue (24%), nausea (16%), increased blood alkaline phosphatase (12%), and blurred vision (12%). No patients had serious or fatal treatment-related AEs; 28% had grade ≥3 treatment-related AEs. Grade 3 AST elevations resolved when treatment was withheld. AMG 820 showed linear pharmacokinetics, with minimal accumulation (<2-fold) after repeated dosing. Pharmacodynamic increases in serum CSF1 concentrations and reduced numbers of skin macrophages were observed. Best response was stable disease in 8 patients (32%). Conclusions: AMG 820 was tolerated with manageable toxicities up to 20 mg/kg every 2 weeks. Pharmacodynamic response was demonstrated, and limited antitumor activity was observed. Clin Cancer Res; 23(19); 5703–10. ©2017 AACR.

Abstract CT137: First-in-human study of AMG 820, a monoclonal anti-CSF-1R (c-fms) antibody, in patients (pts) with advanced solid tumors

Background: Tumor-associated macrophages (TAMs) may contribute to tumor growth and invasion and promote an immunosuppressive tumor microenvironment, making them an attractive target for cancer immunotherapy. TAMs can be activated by the binding of colony stimulating factor-1 (CSF-1) to its receptor CSF-1R (c-fms). AMG 820 is an investigational, fully human antagonistic antibody to CSF-1R that inhibits ligand-induced receptor activation. In this open-label, sequential dose-escalation study, we evaluated the safety, tolerability, pharmacokinetics (PK), pharmacodynamics (PD), and efficacy of AMG 820. Methods: Key eligibility criteria: age ? 18 years, advanced solid tumors relapsed or refractory to standard treatment, measurable disease per RECIST 1.1, ECOG performance status ? 2, and no primary central nervous system tumors/metastases. AMG 820 was administered by IV infusion once weekly at a starting dose of 0.5 mg/kg (cohort 1). Dosing frequency was later amended to once every 2 weeks (Q2W) with planned escalation to 1.5, 3, 6, 10, 20, and 30 mg/kg (cohorts 2–7), following a 3 + 3 design until a maximum tolerated dose (MTD, highest dose at which Results: A total of 25 pts received ? 1 dose of AMG 820: 3 pts at 0.5, 1.5, and 6 mg/kg; 4 pts at 3 and 10 mg/kg; 8 pts at 20 mg/kg. Median (range) age: 63 (48–80) years. Women: 56%. ECOG 0/1/2: 20%/76%/4%. MTD was not reached. One DLT was observed (20 mg/kg): nonreversible grade 3 hearing impairment in a female pt with anal cancer with confounding factors (prior treatment [tx] with cisplatin). Most pts (76%) had tx-related adverse events (TRAEs). Most common TRAEs (in > 10% of pts overall) were periorbital edema (11, 44%), increased aspartate aminotransferase (7, 28%), fatigue (6, 24%), nausea (4, 16%), increased blood alkaline phosphatase (3, 12%), and blurred vision (3, 12%). Grade ? 3 TRAEs were observed in 28% of pts. There were no serious or fatal TRAEs. After the first dose of AMG 820 at 20 mg/kg, mean estimates of Cmax and AUClast (AUC from time 0 to time of last sample) were 619 μg/mL and 89,200 μg•hr/mL, respectively. Both AMG 820 AUClast and Cmax increased linearly with dose, with minimal accumulation ( Conclusions: AMG 820 was tolerated with manageable toxicities up to 20 mg/kg Q2W and showed linear PK, with PD response in CSF-1 serum levels and tx-related changes in skin macrophages. These results and an increasing understanding of the role of TAMs in tumor immunosuppression provide the rationale for combining AMG 820 with other immunotherapies. Citation Format: Kyriakos Papadopoulos, Larry Gluck, Lainie P. Martin, Anthony J. Olszanski, Gateree Ngarmchamnanrith, Erik Rasmussen, Benny Amore, Dirk Nagorsen, John S. Hill, Joe Stephenson. First-in-human study of AMG 820, a monoclonal anti-CSF-1R (c-fms) antibody, in patients (pts) with advanced solid tumors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr CT137.

Abstract 2351: CDK4/FLT3 dual inhibitors as potential therapeutics for acute myeloid leukemia.

CDK4 is a cyclin D dependent kinase that promotes cell cycle progression in a broad range of tumor types by phosphorylating the tumor suppressor retinoblastoma protein (Rb) and releasing transcription factor E2F. Critical involvement of the cyclin D-CDK4-Rb pathway in carcinogenesis is strongly supported by a large amount of genetic evidence. In addition, promoter methylation with consequent silencing of expression of the CDK4 inhibitor, p15, has been reported in 44-60% of acute myeloid leukemia (AML) patients. It is also well established that constitutive activation of the tyrosine kinase FLT3 via mutation contributes to the development of AML, with 30% of AML carrying such activating mutations. FLT3 tyrosine kinase inhibitors used as single agents reduce peripheral blood and bone marrow blasts in only a minority of AML patients, and the effect tends to be transient. This may be due to insufficient FLT3 inhibition, the selection of drug-resistant clones, or the independence of the cell on FLT3 signaling for proliferation and survival. In preclinical models, a synergistic effect of CDK4 inhibition and FLT3 inhibition resulting in increased apoptosis of AML cell lines was reported (Wang et al., Blood, 2007). From a HTS hit through SAR optimization led to AM-5992, a potent and orally bioavailable dual inhibitor of CDK4 and FLT3 including all FLT3 mutants reported to date. AM-5992 inhibits the proliferation of a panel of human tumor cell lines including MDA-MB-435(Rb+), colo-205(Rb+), U937(FLT3WT) and induced cell death in MOLM13(FLT3ITD), MV4-11(FLT3ITD), and even in MOLM13(FLT3ITD, D835Y) which exhibits resistance to a number of FLT3 inhibitors currently under clinical development. In mouse models of leukemia using cells with the FLT3ITD mutation, AM-5992 treatment at 150 mpk qd on days 6-16 after leukemia cell injection significantly reduced the leukemia burden and prolonged survival 11 days over that of vehicle controls. Collectively, these data support the hypothesis that simultaneously inhibition of CDK4 and FLT3 may improve the durability of clinical response in AML; and consequently that this hypothesis should be tested in the clinic. Citation Format: Zhihong Li, Kang Dai, Kathleen Keegan, Ji Ma, Mark Ragains, Jacob Kaizerman, Dustin McMinn, Jiasheng Fu, Benjamin Fisher, Michael Gribble, Lawrence R. McGee, John Eksterowicz, Cong Li, Lingming Liang, Margaret Weidner, Justin Huard, Robert Cho, Timothy Carlson, Grace M. Alba, David Hollenback, John Hill, Darrin Beaupre, Alexander Kamb, Dineli Wickramasinghe, Julio C. Medina. CDK4/FLT3 dual inhibitors as potential therapeutics for acute myeloid leukemia. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2351. doi:10.1158/1538-7445.AM2013-2351