Preclinical Properties Research Articles

Merestinib blocks Mnk kinase activity in acute myeloid leukemia progenitors and exhibits antileukemic effects in vitro and in vivo

AbstractMitogen-activated protein kinase interacting protein kinases (Mnks) play important roles in the development and progression of acute myeloid leukemia (AML) by regulating eukaryotic translation initiation factor 4E (eIF4E) activation. Inhibiting Mnk1/2-induced phosphorylation of eIF4E may represent a unique approach for the treatment of AML. We provide evidence for antileukemic effects of merestinib, an orally bioavailable multikinase inhibitor with suppressive effects on Mnk activity. Our studies show that merestinib effectively blocks eIF4E phosphorylation in AML cells and suppresses primitive leukemic progenitors from AML patients in vitro and in an AML xenograft model in vivo. Our findings provide evidence for potent preclinical antileukemic properties of merestinib and support its clinical development for the treatment of patients with AML.

Abstract 4859: Identifying high quality, potent and selective inhibitors of ATM kinase: Discovery of AZD0156

Abstract Ataxia telangiectasia mutant (ATM) is a serine/threonine protein kinase from the phosphatidylinositol 3-kinase-related kinase (PIKK) family of protein kinases (also comprising ATR, DNA-PKcs, mTOR etc.) and plays a crucial role in the cellular DNA damage response signalling activated by DNA double strand breaks (DSB). Activated ATM promotes DNA repair and S/G1-cell cycle checkpoints to prevent premature mitosis, maintain genomic integrity and promote appropriate cell survival or death pathways. DSBs arise intrinsically through the collapse of stalled replication forks, which are induced by a wide range of chemotherapies, or extrinsically through exposure to ionizing radiation. Therefore, ATM inhibition represents an exciting clinical opportunity as a target to hyper-sensitize tumors to chemo/radiotherapy. Herein, we describe our efforts to identify multiple, novel series of ATM inhibitors. We will describe our optimization efforts with particular attention given to improving the potency of the compounds in cellular systems and the selectivity of the compounds over other closely related proteins (e.g. ATR, mTOR etc.). We will also describe our efforts to optimize both the physicochemical properties of the molecules as well as the pharmacokinetic profile to enable low dose, oral administration in the clinic. These efforts culminated in the discovery of the clinical candidate AZD0156, a first in class orally available ATM inhibitor. AZD0156 shows sub-nanomolar potency in cell based assays of ATM inhibition with selectivities of greater than 1000 fold over other members of the PIKK family of enzymes. AZD0156 is a permeable, highly soluble compound with excellent preclinical pharmacokinetic properties including oral bioavailability. AZD0156 shows robust efficacy in mouse xenograft models after oral administration when combined with DSB inducing agents. AZD0156 is currently undergoing early clinical assessment. Citation Format: Kurt G. Pike. Identifying high quality, potent and selective inhibitors of ATM kinase: Discovery of AZD0156. [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 4859.

Integrated Strategy for Use of Positron Emission Tomography in Nonhuman Primates to Confirm Multitarget Occupancy of Novel Psychotropic Drugs: An Example with AZD3676.

Positron emission tomography (PET) is widely applied in central nervous system (CNS) drug development for assessment of target engagement in vivo. As the majority of PET investigations have addressed drug interaction at a single binding site, findings of multitarget engagement have been less frequently reported and have often been inconsistent with results obtained in vitro. AZD3676 [N,N-dimethyl-7-(4-(2-(pyridin-2-yl)ethyl)piperazin-1-yl) benzofuran-2-carboxamide] is a novel combined serotonin (5-hydroxytryptamine) 5-HT1A and 5-HT1B receptor antagonist that was developed for the treatment of cognitive impairment in Alzheimer's disease. Here, we evaluated the properties of AZD3676 as a CNS drug by combining in vitro and ex vivo radioligand binding techniques, behavioral pharmacology in rodents, and PET imaging in nonhuman primates. Target engagement in the nonhuman primate brain was assessed in PET studies by determination of drug-induced occupancy using receptor-selective radioligands. AZD3676 showed preclinical properties consistent with CNS drug potential, including nanomolar receptor affinity and efficacy in rodent models of learning and memory. In PET studies of the monkey brain, AZD3676 inhibited radioligand binding in a dose-dependent manner with similar affinity at both receptors. The equally high affinity at 5-HT1A and 5-HT1B receptors as determined in vivo was not predicted from corresponding estimates obtained in vitro, suggesting more than 10-fold selectivity for 5-HT1A versus 5-HT1B receptors. These findings support the further integrated use of PET for confirmation of multitarget occupancy of CNS drugs. Importantly, earlier introduction of PET studies in nonhuman primates may reduce future development costs and the requirement for animal experiments in preclinical CNS drug development programs.

Preclinical Characterization of the Novel HCV NS3 Protease Inhibitor GS-9256

GS-9256 is an inhibitor of HCV NS3 protease with a macrocyclic structure and novel phosphinic acid pharmacophore. Key preclinical properties of GS-9256 including in vitro antiviral activity, cross-resistance and pharmacokinetic properties were investigated in non-human species. In genotype (GT) 1b Huh-luc cells with a replicon encoding luciferase, GS-9256 had a mean 50% effective concentration (EC50) value of 20.0 nM, with minimal cytotoxicity. Antiviral activity was similar in a number of additional GT1b and GT1a replicon cell lines. Similar potency was observed in chimeric replicons encoding the NS3 protease of GT1 clinical isolates. GS-9256 was less active in GT2a replicon cells (14.2-fold increase in EC50). Additive to synergistic in vitro antiviral activity was observed when GS-9256 was combined with other agents including interferon-α, ribavirin, NS5B polymerase inhibitors GS-6620 and tegobuvir, as well as the NS5A inhibitor ledipasvir. GS-9256 retained wild-type activity against all tested NS5B and NS5A inhibitor resistance mutations. GS-9256 was metabolically stable in microsomes and hepatocytes of tested species, including rodents, dogs and humans. GS-9256 had high bioavailability in mice (near 100%) and moderate bioavailability in rats (14%), dogs (21%) and monkeys (14%). Elimination half-lives were approximately 2 h in mice, 0.6 h in rats, 5 h in dogs and 4 h in monkey. A study in bile duct-cannulated rats indicated that the major route of elimination is through biliary excretion of unmetabolized GS-9256. GS-9256 showed a favourable preclinical profile supportive of clinical development for the treatment of chronic HCV infection in GT1 patients.

Discovery of MK-7145, an Oral Small Molecule ROMK Inhibitor for the Treatment of Hypertension and Heart Failure.

ROMK, the renal outer medullary potassium channel, is involved in potassium recycling at the thick ascending loop of Henle and potassium secretion at the cortical collecting duct in the kidney nephron. Because of this dual site of action, selective inhibitors of ROMK are expected to represent a new class of diuretics/natriuretics with superior efficacy and reduced urinary loss of potassium compared to standard-of-care loop and thiazide diuretics. Following our earlier work, this communication will detail subsequent medicinal chemistry endeavors to further improve lead selectivity against the hERG channel and preclinical pharmacokinetic properties. Pharmacological assessment of highlighted inhibitors will be described, including pharmacodynamic studies in both an acute rat diuresis/natriuresis model and a subchronic blood pressure model in spontaneous hypertensive rats. These proof-of-biology studies established for the first time that the human and rodent genetics accurately predict the in vivo pharmacology of ROMK inhibitors and supported identification of the first small molecule ROMK inhibitor clinical candidate, MK-7145.

Targeting the hypoxic fraction of tumours using hypoxia-activated prodrugs

The presence of a microenvironment within most tumours containing regions of low oxygen tension or hypoxia has profound biological and therapeutic implications. Tumour hypoxia is known to promote the development of an aggressive phenotype, resistance to both chemotherapy and radiotherapy and is strongly associated with poor clinical outcome. Paradoxically, it is recognised as a high-priority target and one of the therapeutic strategies designed to eradicate hypoxic cells in tumours is a group of compounds known collectively as hypoxia-activated prodrugs (HAPs) or bioreductive drugs. These drugs are inactive prodrugs that require enzymatic activation (typically by 1 or 2 electron oxidoreductases) to generate cytotoxic species with selectivity for hypoxic cells being determined by (1) the ability of oxygen to either reverse or inhibit the activation process and (2) the presence of elevated expression of oxidoreductases in tumours. The concepts underpinning HAP development were established over 40 years ago and have been refined over the years to produce a new generation of HAPs that are under preclinical and clinical development. The purpose of this article is to describe current progress in the development of HAPs focusing on the mechanisms of action, preclinical properties and clinical progress of leading examples.

In Vitro Antiviral Activity and Resistance Profile Characterization of the Hepatitis C Virus NS5A Inhibitor Ledipasvir.

Ledipasvir (LDV; GS-5885), a component of Harvoni (a fixed-dose combination of LDV with sofosbuvir [SOF]), is approved to treat chronic hepatitis C virus (HCV) infection. Here, we report key preclinical antiviral properties of LDV, including in vitro potency, in vitro resistance profile, and activity in combination with other anti-HCV agents. LDV has picomolar antiviral activity against genotype 1a and genotype 1b replicons with 50% effective concentration (EC50) values of 0.031 nM and 0.004 nM, respectively. LDV is also active against HCV genotypes 4a, 4d, 5a, and 6a with EC50 values of 0.11 to 1.1 nM. LDV has relatively less in vitro antiviral activity against genotypes 2a, 2b, 3a, and 6e, with EC50 values of 16 to 530 nM. In vitro resistance selection with LDV identified the single Y93H and Q30E resistance-associated variants (RAVs) in the NS5A gene; these RAVs were also observed in patients after a 3-day monotherapy treatment. In vitro antiviral combination studies indicate that LDV has additive to moderately synergistic antiviral activity when combined with other classes of HCV direct-acting antiviral (DAA) agents, including NS3/4A protease inhibitors and the nucleotide NS5B polymerase inhibitor SOF. Furthermore, LDV is active against known NS3 protease and NS5B polymerase inhibitor RAVs with EC50 values equivalent to those for the wild type.

Preclinical pharmacology and pharmacokinetics of CERC-301, a GluN2B-selective N-methyl-D-aspartate receptor antagonist.

The preclinical pharmacodynamic and pharmacokinetic properties of 4‐methylbenzyl (3S, 4R)‐3‐fluoro‐4‐[(Pyrimidin‐2‐ylamino) methyl] piperidine‐1‐carboxylate (CERC‐301), an orally bioavailable selective N‐methyl‐D‐aspartate (NMDA) receptor subunit 2B (GluN2B) antagonist, were characterized to develop a translational approach based on receptor occupancy (RO) to guide CERC‐301 dose selection in clinical trials of major depressive disorder. CERC‐301 demonstrated high‐binding affinity (K i, 8.1 nmol L−1) specific to GluN2B with an IC 50 of 3.6 nmol L−1 and no off‐target activity. CERC‐301 efficacy was demonstrated in the forced swim test with an efficacy dose (ED 50) of 0.3–0.7 mg kg−1 (RO, 30–50%); increase in locomotor activity was observed at ED 50 of 2 mg kg−1, corresponding to an RO of 75%. The predicted 50% RO concentration (Occ50) in humans was 400 nmol L−1, similar to that predicted for rat, dog, and monkey (300, 200, and 400 nmol L−1, respectively). Safety pharmacology and neurotoxicity studies raised no specific safety concerns. A first‐in‐human study in healthy males demonstrated a dose‐proportional pharmacokinetic profile, with T max of ~1 h and t 1/2 of 12–17 h. Based on the preclinical and pharmacodynamic data, doses of ≥8 mg in humans are hypothesized to have an acceptable safety profile and result in clinically relevant peak plasma exposure.

Discovery of highly potent and selective Bruton's tyrosine kinase inhibitors: Pyridazinone analogs with improved metabolic stability.

BTK inhibitor GDC-0834 (1) was found to be rapidly metabolized in human studies, resulting in a suspension of clinical trials. The primary route of metabolism was through cleavage of the acyclic amide bond connecting the terminal tetrahydrobenzothiophene with the central linker aryl ring. SAR studies were focused on reducing metabolic cleavage of this amide, and resulted in the identification of several central aryl linker substituents that conferred improved stability. The most promising substituted aryl linkers were then incorporated into an optimized pyridazinone scaffold, resulting in the identification of lead analog 23, possessing improved potency, metabolic stability and preclinical properties.

Identification and Characterization of GAL-021 as a Novel Breathing Control Modulator.

The authors describe the preclinical pharmacological properties of GAL-021, a novel peripheral chemoreceptor modulator. The ventilatory effects of GAL-021 were characterized using tracheal pneumotachometry (n = 4 to 6), plethysmography (n = 5 to 6), arterial blood gas analyses (n = 6 to 11), and nasal capnography (n = 3 to 4) in naive animals and those subjected to morphine-induced respiratory depression. Morphine analgesia in rats was evaluated by tail-flick test (n = 6). Carotid body involvement in GAL-021 ventilatory effects was assessed by comparing responses in intact and carotid sinus nerve-transected rats. Hemodynamic effects of GAL-021 were evaluated in urethane-anesthetized rats (n = 7). The pharmacological profile of GAL-021 in vitro was investigated using radioligand binding, enzyme inhibition, and cellular electrophysiology assays. GAL-021 given intravenously stimulated ventilation and/or attenuated opiate-induced respiratory depression in rats, mice, and nonhuman primates, without decreasing morphine analgesia in rats. GAL-021 did not alter mean arterial pressure but produced a modest increase in heart rate. Ventilatory stimulation in rats was attenuated by carotid sinus nerve transection. GAL-021 inhibited KCa1.1 in GH3 cells, and the evoked ventilatory stimulation was attenuated in Slo1 mice lacking the pore-forming α-subunit of the KCa1.1 channel. GAL-021 behaved as a breathing control modulator in rodents and nonhuman primates and diminished opioid-induced respiratory depression without compromising opioid analgesia. It acted predominantly at the carotid body, in part by inhibiting KCa1.1 channels. Its preclinical profile qualified the compound to enter clinical trials to assess effects on breathing control disorders such as drug (opioid)-induced respiratory depression and sleep apnea.

Preclinical properties and human in vivo assessment of 123I-ABC577 as a novel SPECT agent for imaging amyloid-β.

Non-invasive imaging of amyloid-β in the brain, a hallmark of Alzheimer's disease, may support earlier and more accurate diagnosis of the disease. In this study, we assessed the novel single photon emission computed tomography tracer (123)I-ABC577 as a potential imaging biomarker for amyloid-β in the brain. The radio-iodinated imidazopyridine derivative (123)I-ABC577 was designed as a candidate for a novel amyloid-β imaging agent. The binding affinity of (123)I-ABC577 for amyloid-β was evaluated by saturation binding assay and in vitro autoradiography using post-mortem Alzheimer's disease brain tissue. Biodistribution experiments using normal rats were performed to evaluate the biokinetics of (123)I-ABC577. Furthermore, to validate (123)I-ABC577 as a biomarker for Alzheimer's disease, we performed a clinical study to compare the brain uptake of (123)I-ABC577 in three patients with Alzheimer's disease and three healthy control subjects. (123)I-ABC577 binding was quantified by use of the standardized uptake value ratio, which was calculated for the cortex using the cerebellum as a reference region. Standardized uptake value ratio images were visually scored as positive or negative. As a result, (123)I-ABC577 showed high binding affinity for amyloid-β and desirable pharmacokinetics in the preclinical studies. In the clinical study, (123)I-ABC577 was an effective marker for discriminating patients with Alzheimer's disease from healthy control subjects based on visual images or the ratio of cortical-to-cerebellar binding. In patients with Alzheimer's disease, (123)I-ABC577 demonstrated clear retention in cortical regions known to accumulate amyloid, such as the frontal cortex, temporal cortex, and posterior cingulate. In contrast, less, more diffuse, and non-specific uptake without localization to these key regions was observed in healthy controls. At 150 min after injection, the cortical standardized uptake value ratio increased by ∼ 60% in patients with Alzheimer's disease relative to healthy control subjects. Both healthy control subjects and patients with Alzheimer's disease showed minimal (123)I-ABC577 retention in the white matter. These observations indicate that (123)I-ABC577 may be a useful single photon emission computed tomography imaging maker to identify amyloid-β in the human brain. The availability of an amyloid-β tracer for single photon emission computed tomography might increase the accessibility of diagnostic imaging for Alzheimer's disease.

ChemInform Abstract: Cyclin Dependent Kinase (CDK) Inhibitors as Anticancer Drugs

AbstractReview: 78 refs.

Evaluation of the Safety and Benefit of Phase I Oncology Trials for Patients With Primary CNS Tumors.

Patients with high-grade gliomas (HGG) are frequently excluded from first-in-human solid tumor trials because of perceived poor prognosis, excessive toxicities, concomitant drug interactions, and poor efficacy. We conducted an analysis of outcomes from select, single-agent phase I studies in patients with HGG. We compared outcomes to pooled analysis of published studies in solid tumors with various molecular and cytotoxic drugs evaluated as single agents or as combinations. Individual records of patients with recurrent HGG enrolled onto Adult Brain Tumor Consortium trials of single-agent, cytotoxic or molecular agents from 2000 to 2008 were analyzed for baseline characteristics, toxicities, responses, and survival. Our analysis included 327 patients with advanced, refractory HGG who were enrolled onto eight trials involving targeted molecular (n=5) and cytotoxic (n=3) therapies. At enrollment, patients had a median Karnofsky performance score of 90 and median age of 52 years; 62% were men, 63% had glioblastoma, and the median number of prior systemic chemotherapies was one. Baseline laboratory values were in an acceptable range to meet eligibility criteria. Patients were on the study for a median of two cycles (range, <one to 56 cycles), and 96% were evaluable for primary end points. During cycle 1, grade≥3 nonhematologic and grade≥4 hematologic toxicities were 5% (28 of 565 adverse events) and 0.9% (five of 565 adverse events), respectively, and 66% of these occurred at the highest dose level. There was one death attributed to drug. Overall response rate (complete and partial response) was 5.5%. Median progression-free and overall survival times were 1.8 and 6 months, respectively. Patients with HGG who meet standard eligibility criteria may be good candidates for solid tumor phase I studies with single-agent molecular or cytotoxic drugs with favorable preclinical rationale and pharmacokinetic properties in this population.

Abstract 4531: Effects of drug load on therapeutic index for antibody-maytansinoid conjugates

Abstract Antibody-drug conjugates (ADCs) are being actively pursued as a new modality to treat cancer following the regulatory approval of Adcetris@ and Kadcyla@. ADCs consist of a cytotoxic agent, or drug, conjugated to a targeting antibody (Ab) through a linker. The two approved ADCs (and most ADCs now in the clinic) are heterogeneous conjugates with an average molar drug to Ab ratio (DAR) of 3-4 (potentially ranging from 0-8 for individual molecules). To understand the effects of different DAR ranges on the preclinical properties of ADCs using a maytansinoid cytotoxic agent, we prepared a series of conjugates with a cleavable linker (M9346A-sulfo-SPDB-DM4 targeting folate receptor alpha) or an uncleavable linker (J2898A-SMCC-DM1 targeting epidermal growth factor receptor) with varying DAR, and evaluated their biochemical characteristics, in vivo stability, efficacy, and tolerability. Both M9346A-sulfo-SPDB-DM4 and J2898A-SMCC-DM1 conjugates with low (average ∼2, range 0-4) to very high DAR (average 10, range 7-14) were prepared in good yield, high monomer content and low free drug levels. At constant Ab concentration, the in vitro potency consistently increased with increasing DAR. We then characterized the in vivo disposition of these ADCs. First, pharmacokinetic analysis showed that conjugates with an average DAR below ∼6 had comparable clearance rates, but at an average DAR around 9-10 rapid clearance was observed. Biodistribution studies in mice showed that these 9-10 DAR conjugates rapidly distribute to the liver, with the maximum%ID/g for this organ at 24-28% compared with 7-10% for lower DAR conjugates (all at 2-6 h post-injection). We further studied the efficacy of these ADCs in mouse xenograft models at both constant antibody and constant maytansinoid doses. At constant Ab dose, conjugates with average DAR ∼6 are more efficacious than conjugates with DAR ∼3. However, DAR 9-10 conjugates are either less or similarly active compared to DAR 6 conjugates, consistent with the observed rapid clearance. At constant maytansinoid dose, conjugates with 2 - 6 DAR show similar efficacy, while those with 9-10 DAR were again less active. Tolerability studies monitoring body weight loss show that conjugates with different DAR, even as high as 9-10, have comparable tolerability based on total administered maytansinoid dose. In summary, our preclinical findings on tolerability and efficacy suggest that maytansinoid conjugates with DAR in the range between 2 - 6 have better therapeutic index than conjugates with very high DAR (∼9-10). These very high DAR ADCs suffer from decreased efficacy likely due to faster clearance. These results support the use of DAR 3-4 for maytansinoid ADCs, but suggest the exploration of lower or higher DAR depending on the biology of the target antigen. Citation Format: Xiuxia Sun, Jose F. Ponte, Nicholas C. Yoder, Jennifer Coccia, Leanne Lanieri, Rassol Laleau, Qifeng Qiu, Rui Wu, Erica Hong, Megan Bogalhas, Lintao Wang, Erin K. Maloney, Olga Ab, Hans K. Erickson, Thomas A. Keating, Ravi Chari, John M. Lambert. Effects of drug load on therapeutic index for antibody-maytansinoid conjugates. [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 4531. doi:10.1158/1538-7445.AM2015-4531

Novel Ranking System for Identifying Efficacious Anti-Influenza Virus PB2 Inhibitors.

Through antigenic drift and shifts, influenza virus infections continue to be an annual cause of morbidity in healthy populations and of death among elderly and at-risk patients. The emergence of highly pathogenic avian influenza viruses such as H5N1 and H7N9 and the rapid spread of the swine-origin H1N1 influenza virus in 2009 demonstrate the continued need for effective therapeutic agents for influenza. While several neuraminidase inhibitors have been developed for the treatment of influenza virus infections, these have shown a limited window for treatment initiation, and resistant variants have been noted in the population. In addition, an older class of antiviral drugs for influenza, the adamantanes, are no longer recommended for treatment due to widespread resistance. There remains a need for new influenza therapeutic agents with improved efficacy as well as an expanded window for the initiation of treatment. Azaindole compounds targeting the influenza A virus PB2 protein and demonstrating excellent in vitro and in vivo properties have been identified. To evaluate the in vivo efficacy of these PB2 inhibitors, we utilized a mouse influenza A virus infection model. In addition to traditional endpoints, i.e., death, morbidity, and body weight loss, we measured lung function using whole-body plethysmography, and we used these data to develop a composite efficacy score that takes compound exposure into account. This model allowed the rapid identification and ranking of molecules relative to each other and to oseltamivir. The ability to identify compounds with enhanced preclinical properties provides an opportunity to develop more-effective treatments for influenza in patients.

Cyclin dependent kinase (CDK) inhibitors as anticancer drugs.

Sustained proliferative capacity is a hallmark of cancer. In mammalian cells proliferation is controlled by the cell cycle, where cyclin-dependent kinases (CDKs) regulate critical checkpoints. CDK4 and CDK6 are considered highly validated anticancer drug targets due to their essential role regulating cell cycle progression at the G1 restriction point. This review provides an overview of recent advances on cyclin dependent kinase inhibitors in general with special emphasis on CDK4 and CDK6 inhibitors and compounds under clinical evaluation. Chemical structures, structure activity relationships, and relevant preclinical properties will be described.

The preclinical pharmacology of the high affinity anti-IL-6R Nanobody® ALX-0061 supports its clinical development in rheumatoid arthritis.

IntroductionThe pleiotropic cytokine interleukin-6 (IL-6) plays an important role in the pathogenesis of different diseases, including rheumatoid arthritis (RA). ALX-0061 is a bispecific Nanobody® with a high affinity and potency for IL-6 receptor (IL-6R), combined with an extended half-life by targeting human serum albumin. We describe here the relevant aspects of its in vitro and in vivo pharmacology.MethodsALX-0061 is composed of an affinity-matured IL-6R-targeting domain fused to an albumin-binding domain representing a minimized two-domain structure. A panel of different in vitro assays was used to characterize the biological activities of ALX-0061. The pharmacological properties of ALX-0061 were examined in cynomolgus monkeys, using plasma levels of total soluble (s)IL-6R as pharmacodynamic marker. Therapeutic effect was evaluated in a human IL-6-induced acute phase response model in the same species, and in a collagen-induced arthritis (CIA) model in rhesus monkeys, using tocilizumab as positive control.ResultsALX-0061 was designed to confer the desired pharmacological properties. A 200-fold increase of target affinity was obtained through affinity maturation of the parental domain. The high affinity for sIL-6R (0.19 pM) translated to a concentration-dependent and complete neutralization of sIL-6R in vitro. In cynomolgus monkeys, ALX-0061 showed a dose-dependent and complete inhibition of hIL-6-induced inflammatory parameters, including plasma levels of C-reactive protein (CRP), fibrinogen and platelets. An apparent plasma half-life of 6.6 days was observed after a single intravenous administration of 10 mg/kg ALX-0061 in cynomolgus monkeys, similar to the estimated expected half-life of serum albumin. ALX-0061 and tocilizumab demonstrated a marked decrease in serum CRP levels in a non-human primate CIA model. Clinical effect was confirmed in animals with active drug exposure throughout the study duration.ConclusionsALX-0061 represents a minimized bispecific biotherapeutic of 26 kDa, nearly six times smaller than monoclonal antibodies. High in vitro affinity and potency was demonstrated. Albumin binding as a half-life extension technology resulted in describable and expected pharmacokinetics. Strong IL-6R engagement was shown to translate to in vivo effect in non-human primates, demonstrated via biomarker deregulation as well as clinical effect. Presented results on preclinical pharmacological properties of ALX-0061 are supportive of clinical development in RA.Electronic supplementary materialThe online version of this article (doi:10.1186/s13075-015-0651-0) contains supplementary material, which is available to authorized users.

119P - Reversible Versus Irreversible EGFR Tyrosine-Kinase Inhibitors for Non-Small-Cell Lung Cancer (NSCLC): a Preclinical and Pharmacokinetic Comparison

Aim: The small-molecule EGFR tyrosine-kinase inhibitors afatinib, gefitinib, erlotinib, dacomitinib and AZD-9291 were developed to treat advanced NSCLC, and are classed as reversible (erlotinib, gefitinib) or irreversible (afatinib, dacomitinib, AZD-9291). Physicochemical, preclinical and clinical ADME properties were compared in an effort to differentiate these EGFR inhibitors.

Potential application of curcumin and its analogues in the treatment strategy of patients with primary epithelial ovarian cancer.

Recent findings on the molecular basis of ovarian cancer development and progression create new opportunities to develop anticancer medications that would affect specific metabolic pathways and decrease side systemic toxicity of conventional treatment. Among new possibilities for cancer chemoprevention, much attention is paid to curcumin—A broad-spectrum anticancer polyphenolic derivative extracted from the rhizome of Curcuma longa L. According to ClinicalTrials.gov at present there are no running pilot studies, which could assess possible therapeutic benefits from curcumin supplementation to patients with primary epithelial ovarian cancer. Therefore, the goal of this review was to evaluate potential preclinical properties of curcumin and its new analogues on the basis of in vivo and in vitro ovarian cancer studies. Curcumin and its different formulations have been shown to display multifunctional mechanisms of anticancer activity, not only in platinum-resistant primary epithelial ovarian cancer, but also in multidrug resistant cancer cells/xenografts models. Curcumin administered together with platinum-taxane chemotherapeutics have been reported to demonstrate synergistic effects, sensitize resistant cells to drugs, and decrease their biologically effective doses. An accumulating body of evidence suggests that curcumin, due to its long-term safety and an excellent profile of side effects should be considered as a beneficial support in ovarian cancer treatment strategies, especially in patients with platinum-resistant primary epithelial recurrent ovarian cancer or multidrug resistant disease. Although the prospect of curcumin and its formulations as anticancer agents in ovarian cancer treatment strategy appears to be challenging, and at the same time promising, there is a further need to evaluate its effectiveness in clinical studies.

Vortioxetine, a novel antidepressant with multimodal activity: review of preclinical and clinical data.

Vortioxetine, a novel antidepressant for the treatment of major depressive disorder (MDD), is a 5-HT3, 5-HT7 and 5-HT1D receptor antagonist, 5-HT1B receptor partial agonist, 5-HT1A receptor agonist and serotonin (5-HT) transporter (SERT) inhibitor. Here we review its preclinical and clinical properties and discuss translational aspects. Vortioxetine increases serotonergic, noradrenergic, dopaminergic, cholinergic, histaminergic and glutamatergic neurotransmission in brain structures associated with MDD. These multiple effects likely derive from its interaction with 5-HT-receptor-mediated negative feedback mechanisms controlling neuronal activity. In particular, 5-HT3 receptors may play a prominent role, since their blockade i) increases pyramidal neuron activity by removing 5-HT3 receptor-mediated excitation of GABA interneurons, and ii) augments SSRI effects on extracellular 5-HT. However, modulation of the other 5-HT receptor subtypes also likely contributes to vortioxetine's pharmacological effects. Preclinical animal models reveal differences from SSRIs and SNRIs, including antidepressant-like activity, increased synaptic plasticity and improved cognitive function. Vortioxetine had clinical efficacy in patients with MDD: 11 placebo-controlled studies (including one in elderly) with efficacy in 8 (7 positive, 1 supportive), 1 positive active comparator study plus a positive relapse prevention study. In two positive studies, vortioxetine was superior to placebo in pre-defined cognitive outcome measures. The clinically effective dose range (5-20mg/day) spans ~50 to >80% SERT occupancy. SERT and 5-HT3 receptors are primarily occupied at 5mg, while at 20mg, all targets are likely occupied at functionally relevant levels. The side-effect profile is similar to that of SSRIs, with gastrointestinal symptoms being most common, and a low incidence of sexual dysfunction and sleep disruption possibly ascribed to vortioxetine's receptor modulation.