Multicenter, phase 1 study of etavopivat (FT-4202) treatment for up to 12 weeks in patients with sickle cell disease.

Etavopivat is an investigational, once-daily, oral, selective erythrocyte pyruvate kinase (PKR) activator. A multicenter, randomized, placebo-controlled, double-blind, 3-part, phase 1 study (https://clinicaltrials.gov/study/NCT03815695) was conducted to characterize the safety and clinical activity of etavopivat. Thirty-six patients with sickle cell disease (SCD) were enrolled into 4 cohorts: one single-dose; two multiple ascending doses; one open-label [OL]. In the OL cohort, 15 patients (median age 33.0 [range, 17‒55] years received 400-mg etavopivat once daily for 12 weeks; 14 completed treatment. Consistent with the mechanism of PKR activation, increases in ATP and decreases in 2,3 diphosphoglycerate were observed and sustained over 12 weeks' treatment. This translated clinically to an increase in hemoglobin (mean maximal increase 1.6 [range, 0.8‒2.8] g/dL), with >1 g/dL increase in 11 (73%) patients during treatment. Additionally, oxygen tension at which hemoglobin is 50% saturated was reduced (P=.0007) with concomitant shift in point-of-sickling (P=.0034) to lower oxygen tension in oxygen-gradient ektacytometry. Hemolysis markers (absolute reticulocyte count, indirect bilirubin, lactate dehydrogenase) decreased from baseline, along with matrix metalloproteinase-9 and erythropoietin. In the OL cohort, adverse events (AEs) were mostly grade 1/2, consistent with underlying SCD; 5 patients had serious AEs. Vaso-occlusive pain episode was the most common treatment-emergent AE (n=7) in the OL cohort. In this first study of etavopivat in SCD, 400 mg once daily for 12 weeks was well-tolerated, resulting in rapid and sustained increases in hemoglobin, improved RBC physiology, and decreased hemolysis.

Safety, Pharmacokinetics, and Pharmacodynamics of Etavopivat (FT-4202), an Allosteric Activator of Pyruvate Kinase-R, in Healthy Adults: A Randomized, Placebo-Controlled, Double-Blind, First-in-Human Phase 1 Trial.

Etavopivat (FT-4202) is an orally administered, small-molecule allosteric activator of erythrocyte pyruvate kinase-R (PKR) in clinical development for the treatment of sickle cell disease and other hemoglobin disorders. This randomized, placebo-controlled, double-blind, first-in-human combination single-ascending dose and multiple-ascending dose phase 1 trial (NCT03815695) evaluated the safety and pharmacokinetics/pharmacodynamics of etavopivat in 90 healthy adult subjects. In 4 single-ascending dose cohorts, 8 participants were randomized 3:1 to a single oral dose of either etavopivat (n = 6) or placebo (n = 2). In four 14-day multiple-ascending dose cohorts, 12 participants were randomized 3:1 to 14 days of etavopivat (n = 9) or placebo (n = 3). In these studies, most treatment-emergent adverse events were of mild severity (grade 1) and none led to study discontinuation. Etavopivat exhibited a linear and time-independent pharmacokinetic profile (at doses ≤400 mg) and elicited the expected pharmacodynamic effects of PKR activation (decreased 2,3-diphosphoglycerate and increased adenosine triphosphate) and evidence of improved hemoglobin-oxygen affinity. In addition, pharmacodynamic responses were durable with effects continuing for 48 to 72 hours after the last dose, thereby supporting once-daily dosing. Food appeared to have no clinically meaningful effects on etavopivat exposure, thus facilitating administration with or without food. In conclusion, the evaluation of etavopivat in healthy subjects demonstrated proof of mechanism (PKR activation) without significant adverse events. This study also allowed for the selection of dose levels, projected to have an acceptable safety profile and provide therapeutic benefit, for evaluation in future trials in patients with sickle cell disease.

Etavopivat, a Pyruvate Kinase Activator in Red Blood Cells, for the Treatment of Sickle Cell Disease.

Etavopivat is an investigational, oral, small molecule activator of erythrocyte pyruvate kinase (PKR) in development for the treatment of sickle cell disease (SCD) and other hemoglobinopathies. PKR activation is proposed to ameliorate the sickling of SCD red blood cells (RBCs) through multiple mechanisms, including reduction of 2,3-diphosphoglycerate (2,3-DPG), which consequently increases hemoglobin (Hb)-oxygen affinity; increased binding of oxygen reduces sickle hemoglobin polymerization and sickling. In addition, PKR activation increases adenosine triphosphate (ATP) produced via glycolytic flux, which helps preserve membrane integrity and RBC deformability. We evaluated the pharmacodynamic response to etavopivat in nonhuman primates (NHPs) and in healthy human subjects and evaluated the effects in RBCs from patients with SCD after ex vivo treatment with etavopivat. A single dose of etavopivat decreased 2,3-DPG in NHPs and healthy subjects. Hb-oxygen affinity was significantly increased in healthy subjects after 24 hours. After daily dosing of etavopivat over 5 consecutive days in NHPs, ATP was increased by 38% from baseline. Etavopivat increased Hb-oxygen affinity and reduced sickling in RBCs collected from patients with SCD with either homozygous hemoglobin S or hemoglobin S and C disease. Collectively, these results demonstrate the ability of etavopivat to decrease 2,3-DPG and increase ATP, resulting in increased Hb-oxygen affinity and improved sickle RBC function. Etavopivat is currently being evaluated in clinical trials for the treatment of SCD. SIGNIFICANCE STATEMENT: Etavopivat, a small molecule activator of the glycolytic enzyme erythrocyte pyruvate kinase, decreased 2,3-diphosphoglycerate in red blood cells (RBCs) from nonhuman primates and healthy subjects and significantly increased hemoglobin (Hb)-oxygen affinity in healthy subjects. Using ex vivo RBCs from donors with sickle cell disease (SCD) (homozygous hemoglobin S or hemoglobin S and C genotype), etavopivat increased Hb-oxygen affinity and reduced sickling under deoxygenation. Etavopivat shows promise as a treatment for SCD that could potentially reduce vaso-occlusion and improve anemia.

Discovery and Preclinical Characterization of BIIB091, a Reversible, Selective BTK Inhibitor for the Treatment of Multiple Sclerosis.

Multiple Sclerosis is a chronic autoimmune neurodegenerative disorder of the central nervous system (CNS) that is characterized by inflammation, demyelination, and axonal injury leading to permeant disability. In the early stage of MS, inflammation is the primary driver of the disease progression. There remains an unmet need to develop high efficacy therapies with superior safety profiles to prevent the inflammation processes leading to disability. Herein, we describe the discovery of BIIB091, a structurally distinct orthosteric ATP competitive, reversible inhibitor that binds the BTK protein in a DFG-in confirmation designed to sequester Tyr-551, an important phosphorylation site on BTK, into an inactive conformation with excellent affinity. Preclinical studies demonstrated BIB091 to be a high potency molecule with good drug-like properties and a safety/tolerability profile suitable for clinical development as a highly selective, reversible BTKi for treating autoimmune diseases such as MS.

Inhibition of fatty acid synthase with FT-4101 safely reduces hepatic de novo lipogenesis and steatosis in obese subjects with non-alcoholic fatty liver disease: Results from two early-phase randomized trials.

AIMS: To assess the therapeutic potential of fatty acid synthase (FASN) inhibition with FT-4101, a potent, selective, orally bioavailable, small-molecule by (a) evaluating the dose-response of single FT-4101 doses (3, 6 and 9 mg) on hepatic de novo lipogenesis (DNL) in healthy participants (Study 1) and (b) demonstrating the safety, tolerability and efficacy on hepatic steatosis after 12 weeks of FT-4101 dosing in patients with non-alcoholic fatty liver disease (NAFLD; Study 2). MATERIALS AND METHODS: In Study 1, three sequential cohorts of healthy men (n = 10/cohort) were randomized to receive a single dose of FT-4101 (n = 5/cohort) or placebo (n = 5/cohort) followed by crossover dosing after 7 days. Hepatic DNL was assessed during fructose stimulation from 13 C-acetate incorporation. In Study 2, men and women with NAFLD (n = 14) randomly received 12 weeks of intermittent once-daily dosing (four cycles of 2 weeks on-treatment, followed by 1 week off-treatment) of 3 mg FT-4101 (n = 9) or placebo (n = 5). Steady-state DNL based on deuterated water labelling, hepatic steatosis using magnetic resonance imaging-proton density fat fraction and sebum lipids and circulating biomarkers were assessed. RESULTS: Single and repeat dosing of FT-4101 were safe and well tolerated. Single FT-4101 doses inhibited hepatic DNL dose-dependently. Twelve weeks of 3 mg FT-4101 treatment improved hepatic steatosis and inhibited hepatic DNL. Decreases in sebum sapienate content with FT-4101 at week 11 were not significant compared to placebo and rebounded at week 12. Biomarkers of liver function, glucose and lipid metabolism were unchanged. CONCLUSIONS: Inhibition of FASN with 3 mg FT-4101 safely reduces hepatic DNL and steatosis in NAFLD patients.

Abnormal brain structure and behavior in MyD88-deficient mice.

While the original protein Toll in Drosophila melanogaster regulates both host defense and morphogenesis, the role of its ortholog Toll-like receptors (TLRs), the interleukin 1 receptor (IL-1R) family, and the associated signaling pathways in mammalian brain development and structure is poorly understood. Because the adaptor protein myeloid differentiation primary response protein 88 (MyD88) is essential for downstream signaling of most TLRs and IL-1R, we systematically investigated the effect of MyD88 deficiency on murine brain structure during development and on behavior. In neonatal Myd88-/- mice, neocortical thickness was reduced, while density of cortical neurons was increased. In contrast, microglia, astrocyte, oligodendrocyte, and proliferating cell numbers were unchanged in these mice compared to wild-type mice. In adult Myd88-/- mice, neocortical thickness was unaltered, but neuronal density in neocortex and hippocampus was increased. Neuron arborization was less pronounced in adult Myd88-/- mice compared to wild-type animals. In addition, numbers of microglia and proliferating cells were increased in the neocortex and subventricular zone, respectively, with unaltered astrocyte and oligodendrocyte numbers, and myelinization was enhanced in the adult Myd88-/- neocortex. These morphologic changes in the brain of adult Myd88-/- mice were accompanied by specific behavioral traits, such as decreased locomotor activity, increased anxiety-like behavior, but normal day/light activity, satisfactory learning, short- and long-term spatial memory, potential cognitive inflexibility, and increased hanging and locomotor behavior within their home cage. Taken together, MyD88 deficiency results in morphologic and cellular changes in the mouse brain, as well as in altered natural and specific behaviors. Our data indicate a pathophysiological significance of MyD88 for mammalian CNS development, structure, and function.

Discovery of BIIB068: A Selective, Potent, Reversible Bruton's Tyrosine Kinase Inhibitor as an Orally Efficacious Agent for Autoimmune Diseases.

Autoreactive B cell-derived antibodies form immune complexes that likely play a pathogenic role in autoimmune diseases. In systemic lupus erythematosus (SLE), these antibodies bind Fc receptors on myeloid cells and induce proinflammatory cytokine production by monocytes and NETosis by neutrophils. Bruton's tyrosine kinase (BTK) is a non-receptor tyrosine kinase that signals downstream of Fc receptors and plays a transduction role in antibody expression following B cell activation. Given the roles of BTK in both the production and sensing of autoreactive antibodies, inhibitors of BTK kinase activity may provide therapeutic value to patients suffering from autoantibody-driven immune disorders. Starting from an in-house proprietary screening hit followed by structure-based rational design, we have identified a potent, reversible BTK inhibitor, BIIB068 (1), which demonstrated good kinome selectivity with good overall drug-like properties for oral dosing, was well tolerated across preclinical species at pharmacologically relevant doses with good ADME properties, and achieved >90% inhibition of BTK phosphorylation (pBTK) in humans.

Efficacy and Pharmacodynamic Modeling of the BTK Inhibitor Evobrutinib in Autoimmune Disease Models.

Because of its role in mediating both B cell and Fc receptor signaling, Bruton's tyrosine kinase (BTK) is a promising target for the treatment of autoimmune diseases such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). Evobrutinib is a novel, highly selective, irreversible BTK inhibitor that potently inhibits BCR- and Fc receptor-mediated signaling and, thus, subsequent activation and function of human B cells and innate immune cells such as monocytes and basophils. We evaluated evobrutinib in preclinical models of RA and SLE and characterized the relationship between BTK occupancy and inhibition of disease activity. In mouse models of RA and SLE, orally administered evobrutinib displayed robust efficacy, as demonstrated by reduction of disease severity and histological damage. In the SLE model, evobrutinib inhibited B cell activation, reduced autoantibody production and plasma cell numbers, and normalized B and T cell subsets. In the RA model, efficacy was achieved despite failure to reduce autoantibodies. Pharmacokinetic/pharmacodynamic modeling showed that mean BTK occupancy in blood cells of 80% was linked to near-complete disease inhibition in both RA and SLE mouse models. In addition, evobrutinib inhibited mast cell activation in a passive cutaneous anaphylaxis model. Thus, evobrutinib achieves efficacy by acting both on B cells and innate immune cells. Taken together, our data show that evobrutinib is a promising molecule for the chronic treatment of B cell-driven autoimmune disorders.

Discovery and Optimization of a Novel Series of Highly Selective JAK1 Kinase Inhibitors.

Janus kinases (JAKs) have been demonstrated to be critical in cytokine signaling and have thus been implicated in both cancer and inflammatory diseases. The JAK family consists of four highly homologous members: JAK1-3 and TYK2. The development of small-molecule inhibitors that are selective for a specific family member would represent highly desirable tools for deconvoluting the intricacies of JAK family biology. Herein, we report the discovery of a potent JAK1 inhibitor, 24, which displays ∼1000-fold selectivity over the other highly homologous JAK family members (determined by biochemical assays), while also possessing good selectivity over other kinases (determined by panel screening). Moreover, this compound was demonstrated to be orally bioavailable and possesses acceptable pharmacokinetic parameters. In an in vivo study, the compound was observed to dose dependently modulate the phosphorylation of STAT3 (a downstream marker of JAK1 inhibition).

Preclinical QSP Modeling in the Pharmaceutical Industry: An IQ Consortium Survey Examining the Current Landscape.

A cross-industry survey was conducted to assess the landscape of preclinical quantitative systems pharmacology (QSP) modeling within pharmaceutical companies. This article presents the survey results, which provide insights on the current state of preclinical QSP modeling in addition to future opportunities. Our results call attention to the need for an aligned definition and consistent terminology around QSP, yet highlight the broad applicability and benefits preclinical QSP modeling is currently delivering.

Broad analgesic activity of a novel, selective M1 agonist.

Although the muscarinic receptor family has long been a source of potentially compelling targets for small molecule drug discovery, it was difficult to achieve agonist selectivity within the family. A new class of M1 muscarinic agonists has emerged, and these compounds have been characterized as agonists that activate the receptor at an allosteric site. Members of this class of M1 agonists have been shown to be selective across the muscarinic receptors. However, upon introduction of a novel pharmacologic mechanism, it is prudent to ensure that no new off-target activities have arisen, particularly within the context of in vivo experiments. Reported here, is the in vitro and in vivo characterization of a novel M1 agonist tool compound, PPBI, and demonstrations that the primary biological effects of PPBI are mediated through M1. PPBI reverses d-amphetamine locomotor activity, but fails to do so in transgenic mice that do not express M1. PPBI also reverses a natural deficit in a rat cognition model at a level of exposure which also activates cortical circuitry. Most notably, PPBI is analgesic in a variety of rat and mouse models and the analgesic effect of PPBI is reversed by an M1-preferring antagonist and an M1-selective toxin. Finally, the pharmacokinetic/pharmacodynamic measures of PPBI are compared across multiple endpoints which highlights that activity in models of psychosis and pain require higher exposures than that required in the cognition model.

Translational pharmacokinetic-pharmacodynamic analysis in the pharmaceutical industry: an IQ Consortium PK-PD Discussion Group perspective.

With inadequate efficacy being the primary cause for the attrition of drug candidates in clinical development, the need to better predict clinical efficacy earlier in the drug development process has increased in importance in the pharmaceutical industry. Here, we review current applications of translational pharmacokinetic-pharmacodynamic (PK-PD) modeling of preclinical data in the pharmaceutical industry, including best practices. Preclinical translational PK-PD modeling has been used in many therapeutic areas and has been impactful to drug development. The role of preclinical translational PK-PD modeling in drug discovery and development will continue to evolve and broaden, given that its broad implementation in the pharmaceutical industry is relatively recent and many opportunities still exist for its further application.

M2698 is a potent dual-inhibitor of p70S6K and Akt that affects tumor growth in mouse models of cancer and crosses the blood-brain barrier.

Dysregulated PI3K/Akt/mTOR (PAM) pathway signaling occurs in ~30% of human cancers, making it a rational target for new therapies; however, the effectiveness of some PAM pathway inhibitors, such as mTORC rapalogs, may be compromised by a compensatory feedback loop leading to Akt activation. In this study, the p70S6K/Akt dual inhibitor, M2698 (previously MSC2363318A), was characterized as a potential anti-cancer agent through examination of its pharmacokinetic, pharmacodynamic and metabolic properties, and anti-tumor activity. M2698 was highly potent in vitro (IC50 1 nM for p70S6K, Akt1 and Akt3 inhibition; IC50 17 nM for pGSK3ß indirect inhibition) and in vivo (IC50 15 nM for pS6 indirect inhibition), and relatively selective (only 6/264 kinases had an IC50 within 10-fold of p70S6K). Orally administered M2698 crossed the blood-brain barrier in rats and mice, with brain tumor exposure 4-fold higher than non-disease brain. Dose-dependent inhibition of target substrate phosphorylation was observed in vitro and in vivo, indicating that M2698 blocked p70S6K to provide potent PAM pathway inhibition while simultaneously targeting Akt to overcome the compensatory feedback loop. M2698 demonstrated dose-dependent tumor growth inhibition in mouse xenograft models derived from PAM pathway-dysregulated human triple-negative (MDA-MB-468) and Her2-expressing breast cancer cell lines (MDA-MB-453 and JIMT-1), and reduced brain tumor burden and prolonged survival in mice with orthotopically implanted U251 glioblastoma. These findings highlight M2698 as a promising PAM pathway inhibitor whose unique mechanism of action and capacity to pass the blood-brain barrier warrant clinical investigation in cancers with PAM pathway dysregulation, and those with central nervous system involvement.

Preclinical pharmacokinetic/pharmacodynamic modeling and simulation in the pharmaceutical industry: an IQ consortium survey examining the current landscape.

The application of modeling and simulation techniques is increasingly common in preclinical stages of the drug discovery and development process. A survey focusing on preclinical pharmacokinetic/pharmacodynamics (PK/PD) analysis was conducted across pharmaceutical companies that are members of the International Consortium for Quality and Innovation in Pharmaceutical Development. Based on survey responses, ~68% of companies use preclinical PK/PD analysis in all therapeutic areas indicating its broad application. An important goal of preclinical PK/PD analysis in all pharmaceutical companies is for the selection/optimization of doses and/or dose regimens, including prediction of human efficacious doses. Oncology was the therapeutic area with the most PK/PD analysis support and where it showed the most impact. Consistent use of more complex systems pharmacology models and hybrid physiologically based pharmacokinetic models with PK/PD components was less common compared to traditional PK/PD models. Preclinical PK/PD analysis is increasingly being included in regulatory submissions with ~73% of companies including these data to some degree. Most companies (~86%) have seen impact of preclinical PK/PD analyses in drug development. Finally, ~59% of pharmaceutical companies have plans to expand their PK/PD modeling groups over the next 2 years indicating continued growth. The growth of preclinical PK/PD modeling groups in pharmaceutical industry is necessary to establish required resources and skills to further expand use of preclinical PK/PD modeling in a meaningful and impactful manner.

Lifespan based pharmacokinetic-pharmacodynamic model of tumor growth inhibition by anticancer therapeutics.

Accurate prediction of tumor growth is critical in modeling the effects of anti-tumor agents. Popular models of tumor growth inhibition (TGI) generally offer empirical description of tumor growth. We propose a lifespan-based tumor growth inhibition (LS TGI) model that describes tumor growth in a xenograft mouse model, on the basis of cellular lifespan T. At the end of the lifespan, cells divide, and to account for tumor burden on growth, we introduce a cell division efficiency function that is negatively affected by tumor size. The LS TGI model capability to describe dynamic growth characteristics is similar to many empirical TGI models. Our model describes anti-cancer drug effect as a dose-dependent shift of proliferating tumor cells into a non-proliferating population that die after an altered lifespan TA. Sensitivity analysis indicated that all model parameters are identifiable. The model was validated through case studies of xenograft mouse tumor growth. Data from paclitaxel mediated tumor inhibition was well described by the LS TGI model, and model parameters were estimated with high precision. A study involving a protein casein kinase 2 inhibitor, AZ968, contained tumor growth data that only exhibited linear growth kinetics. The LS TGI model accurately described the linear growth data and estimated the potency of AZ968 that was very similar to the estimate from an established TGI model. In the case study of AZD1208, a pan-Pim inhibitor, the doubling time was not estimable from the control data. By fixing the parameter to the reported in vitro value of the tumor cell doubling time, the model was still able to fit the data well and estimated the remaining parameters with high precision. We have developed a mechanistic model that describes tumor growth based on cell division and has the flexibility to describe tumor data with diverse growth kinetics.

Discovery of BI 224436, a Noncatalytic Site Integrase Inhibitor (NCINI) of HIV-1.

An assay recapitulating the 3' processing activity of HIV-1 integrase (IN) was used to screen the Boehringer Ingelheim compound collection. Hit-to-lead and lead optimization beginning with compound 1 established the importance of the C3 and C4 substituent to antiviral potency against viruses with different aa124/aa125 variants of IN. The importance of the C7 position on the serum shifted potency was established. Introduction of a quinoline substituent at the C4 position provided a balance of potency and metabolic stability. Combination of these findings ultimately led to the discovery of compound 26 (BI 224436), the first NCINI to advance into a phase Ia clinical trial.

Pathology and Neurotoxicity in Dogs after Repeat Dose Exposure to a Serotonin 5-HT1B Inhibitor.

AZD3783, a cationic amphiphilic drug and a potent inhibitor of the 5-hydroxytryptamine (5-HT1B) receptor, was explored as a potential treatment for depression. To support clinical trials, repeat dose toxicity studies in rats and dogs were conducted. Here we report toxicity findings in dogs after dosing from 1 to 3 months. In the 1-month study, there were minimal neuronal vacuolation in the brain, a marked increase in liver enzymes accompanied by hepatocellular degeneration/necrosis and phospholipidosis (PLD), and PLD/cholecystitis in the gallbladder of animals dosed at 47 mg/kg/day. In the 3-month study, neurotoxicity resulted in euthanasia of one animal dosed at 30 mg/kg/day after 86 days. Extensive pathologic changes were seen in all animals in retina epithelium (inclusion bodies), brain (neuronal vacuolation, degeneration, or necrosis and nerve fiber degeneration), spinal ganglia (vacuolation, degeneration, or necrosis), as well as sciatic and optic nerves (degeneration). Pigment-laden macrophages were observed in the lung, kidney, liver, gallbladder, bone marrow, gastrointestinal tract, and lymphoid tissues. Also seen were vitrel and retinal hemorrhage in the eyes. A brain concentration and pathology study showed that the concentration of AZD3783 in the brain was approximately 4 times higher than in the plasma after 4 weeks of dosing, however, they were similar in all regions examined, and did not correlate with areas with pathologic findings. Our findings with AZD3783 in dogs have not been reported previously with other CNS compounds that effect through serotonergic pharmacology.

Diagnostic yield of endoscopy in patients with abdominal complaints: incremental value of faecal calprotectin on guidelines of appropriateness.

BACKGROUND: European Panel on the Appropriateness of Gastrointestinal Endoscopy (EPAGE) criteria have been developed to increase diagnostic yield, but their predictive value is limited. We investigated the incremental diagnostic value of faecal calprotectin to EPAGE criteria. METHODS: In a post-hoc analysis of a prospective study, EPAGE criteria were applied to 298 of 575 (51.8%) patients who had undergone esophagogastroduodenoscopy (EGD), colonoscopy or both for abdominal complaints at the Division of Gastroenterology & Hepatology at the University Hospital Basel in Switzerland. Faecal calprotectin was measured in stool samples collected within 24 hours before the investigation using an enzyme-linked immunosorbent assay. Final endoscopic diagnoses were blinded to calprotectin values. RESULTS: Of 149 EGDs and 224 colonoscopies, 17.6% and 14.7% respectively were judged inappropriate by EPAGE criteria. Appropriate or uncertain indications revealed more endoscopic findings in both EGD (46.3% vs. 23.1%, P = 0.049) and colonoscopy (23.6% vs. 6.1%, P = 0.041) than inappropriate indications. Median calprotectin levels were higher (81.5 µg/g, interquartile range 26-175, vs. 10 µg/g, IQR 10-22, P < 0.001) and testing was more often positive (>50 µg/g) in patients with endoscopic findings, both in EGD (58.2% vs. 33.0%, P = 0.005) and in colonoscopy (57.3% vs. 7.4%, P < 0.001). The use of faecal calprotectin in addition to EPAGE criteria improved the risk reclassification of patients by endoscopic findings. The calculated net reclassification index was 37.8% (P = 0.002) for EGD and 110.9% (P <0.001) for colonoscopy, thus improving diagnostic yield to 56.8% and 70.2%, respectively. CONCLUSIONS: The use of faecal calprotectin in addition to EPAGE criteria improved diagnostic yield in patients with abdominal complaints.

Preclinical profile of BI 224436, a novel HIV-1 non-catalytic-site integrase inhibitor.

BI 224436 is an HIV-1 integrase inhibitor with effective antiviral activity that acts through a mechanism that is distinct from that of integrase strand transfer inhibitors (INSTIs). This 3-quinolineacetic acid derivative series was identified using an enzymatic integrase long terminal repeat (LTR) DNA 3'-processing assay. A combination of medicinal chemistry, parallel synthesis, and structure-guided drug design led to the identification of BI 224436 as a candidate for preclinical profiling. It has antiviral 50% effective concentrations (EC50s) of <15 nM against different HIV-1 laboratory strains and cellular cytotoxicity of >90 µM. BI 224436 also has a low, ∼2.1-fold decrease in antiviral potency in the presence of 50% human serum and, by virtue of a steep dose-response curve slope, exhibits serum-shifted EC95 values ranging between 22 and 75 nM. Passage of virus in the presence of inhibitor selected for either A128T, A128N, or L102F primary resistance substitutions, all mapping to a conserved allosteric pocket on the catalytic core of integrase. BI 224436 also retains full antiviral activity against recombinant viruses encoding INSTI resistance substitutions N155S, Q148H, and E92Q. In drug combination studies performed in cellular antiviral assays, BI 224436 displays an additive effect in combination with most approved antiretrovirals, including INSTIs. BI 224436 has drug-like in vitro absorption, distribution, metabolism, and excretion (ADME) properties, including Caco-2 cell permeability, solubility, and low cytochrome P450 inhibition. It exhibited excellent pharmacokinetic profiles in rat (clearance as a percentage of hepatic flow [CL], 0.7%; bioavailability [F], 54%), monkey (CL, 23%; F, 82%), and dog (CL, 8%; F, 81%). Based on the excellent biological and pharmacokinetic profile, BI 224436 was advanced into phase 1 clinical trials.