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Despite improved knowledge regarding disease causality, new drug targets, and enabling technologies, the attrition rate for compounds entering clinical trials has remained consistently high for several decades, with an average 90% failure rate. These failures are manifested in an inability to reproduce efficacy findings from animal models in humans and/or the occurrence of unexpected safety issues, and reflect failures in T1 translation. Similarly, an inability to sequentially demonstrate compound efficacy and safety in Phase IIa, IIb, and III clinical trials represents failures in T2 translation. Accordingly, T1 and T2 translation are colloquially termed 'valleys of death'. Since T2 translation dealt almost exclusively with clinical trials, T3 and T4 translational steps were added, with the former focused on facilitating interactions between laboratory- and population-based research and the latter on 'real world' health outcomes. Factors that potentially lead to T1/T2 compound attrition include: the absence of biomarkers to allow compound effects to be consistently tracked through development; a lack of integration/'de-siloing' of the diverse discipline-based and technical skill sets involved in drug discovery; the industrialization of drug discovery, which via volume-based goals often results in quantity being prioritized over quality; inadequate project governance and strategic oversight; and flawed decision making based on unreliable/irreproducible or incomplete data. A variety of initiatives have addressed this problem, including the NIH National Center for Advancing Translational Sciences (NCATS), which has focused on bringing an unbiased academic perspective to translation, to potentially revitalize the process. This commentary provides an overview of the basic concepts involved in translation, along with suggested changes in the conduct of biomedical research to avoid valleys of death, including the use of Translational Scoring as a tool to avoid translational attrition and the impact of the FDA Accelerated Approval Pathway in lowering the hurdle for drug approval. © 2021 Wiley Periodicals LLC.

People are constantly exposed to phthalates, due to their common use in the production of plastics, pharmaceuticals, cosmetics and skin care products. The ability of phthalates to disrupt endocrine signaling, leading to developmental, reproductive and metabolic defects, has been studied, yet how phthalates interfere with these biological functions is still unclear. To uncover DBP interacting molecular pathways, we raised Drosophila melanogaster on food containing dibutyl phthalate (DBP) at various concentrations. Whole transcriptome analysis of adult Drosophila reveals that DBP exposure throughout development disrupts the expression of genes central to circadian rhythm regulation, including increased expression of vrille (vri, human NFIL3), timeless (tim, human TIMELESS) and period (per, human PER3), with decreased expression of Pigment-dispersing factor (Pdf). DBP exposure also alters the expression of the evolutionarily conserved nuclear receptor Hormone receptor-like in 38 (Hr38, human NR4A2), which is known to regulate Pdf expression. Furthermore, behavioral assays determined that exposing Drosophila to DBP throughout development modifies the circadian rhythm of adults. Although DBP inhibits the expression of signaling systems regulating vision, including Rh5 and Rh6, two light-sensing G-protein coupled receptors involved in the daily resetting of circadian rhythm, it does not influence eye development. Circadian rhythm genes are well conserved from flies to humans; therefore, we tested the effect of DBP exposure on human breast cells (MCF10A) and demonstrate that, similar to the fruit fly model, this exposure disrupts circadian rhythm (BMAL1 expression) at doses that promote the proliferation and migration ability of MCF10A cells. Our results are the first to provide comprehensive evidence that DBP interferes with circadian rhythm in both adult Drosophila and human cells, which may help to explain the broad physiological action of phthalates. De två första författarna delar förstaförfattarskapet

The assessment of behavioral outcomes is a central component of neuroscientific research, which has required continuous technological innovations to produce more detailed and reliable findings. In this article, we provide an in-depth review on the progress and future implications for three model organisms (mouse, rat, and Drosophila) essential to our current understanding of behavior. By compiling a comprehensive catalog of popular assays, we are able to compare the diversity of tasks and usage of these animal models in behavioral research. This compilation also allows for the evaluation of existing state-of-the-art methods and experimental applica- tions, including optogenetics, machine learning, and high-throughput behavioral assays. We go on to discuss novel apparatuses and inter-species analyses for centrophobism, feeding behavior, aggression and mating par- adigms, with the goal of providing a unique view on comparative behavioral research. The challenges and recent advances are evaluated in terms of their translational value, ethical procedures, and trustworthiness for behavioral research.

Implantable drug depots have the capacity to locally meet therapeutic requirements by maximizing local drug efficacy and minimizing potential systemic side effects. Tubular organs including the gastrointestinal tract, respiratory tract and vasculature all manifest with endoluminal disease. The anatomic distribution of localized drug delivery for these organs using existing therapeutic modalities is limited. Application of local depots in a circumferential and extended longitudinal fashion could transform our capacity to offer effective treatment across a range of conditions. Here we report the development and application of a kirigami-based stent platform to achieve this. The stents comprise a stretchable snake-skin-inspired kirigami shell integrated with a fluidically driven linear soft actuator. They have the capacity to deposit drug depots circumferentially and longitudinally in the tubular mucosa of the gastrointestinal tract across millimetre to multi-centimetre length scales, as well as in the vasculature and large airways. We characterize the mechanics of kirigami stents for injection, and their capacity to engage tissue in a controlled manner and deposit degradable microparticles loaded with therapeutics by evaluating these systems ex vivo and in vivo in swine. We anticipate such systems could be applied for a range of endoluminal diseases by simplifying dosing regimens while maximizing drug on-target effects through the sustained release of therapeutics and minimizing systemic side effects.

Cell mediated immunity of the honey bee (Apis mellifera) involves the activity of several hemocyte populations, currently defined by morphological features and lectin binding characteristics. The objective of the present study was to identify molecular markers capable of characterizing subsets of honey bee hemocytes. We developed and employed monoclonal antibodies with restricted reactions to functionally distinct hemocyte subpopulations. Melanizing cells, known as oenocytoids, were defined by an antibody to prophenoloxidase, aggregating cells were identified by the expression of Hemolectin, and phagocytic cells were identified by a marker expressed on granulocytes. We anticipate that this combination of antibodies not only allows for the detection of functionally distinct hemocyte subtypes, but will help to further the exploration of hematopoietic compartments, as well as reveal details of the honey bee cellular immune defense against parasites and microbes.

Alzheimer’s Disease (AD) therapeutics based on the amyloid hypothesis have repeatedly failed in clinical trials. Together with numerous reports that amyloid is present in brains from aged individuals without cognitive dysfunction, this suggests that the association of amyloid with AD is collateral rather than causal. However, the preeminence of the amyloid hypothesis has resulted in the ‘systematic …thwart[ing of] alternative approaches’ to AD/dementia driven by a ‘cabal’ of amyloid acolytes who have effectively controlled the ideas funded and published, which startups received venture investment and which programs were advanced in biopharmaceutical companies where they consulted. As a result, dementia research is estimated to be 15–30 years behind where it could be with conflicting data ignored in favor of the amyloid dogma and clinical trial failures being ascribed to faulty design or inadequacies in the compound selection process including flawed animal models. Major concerns regarding the precise diagnosis of AD/dementia and conflicting views on the validated status of fluid biomarker assays have resulted in trials that included patients with unknown amyloid pathologies. With the failure of the amyloid approach, emerging data on the role(s) of vascular, mitochondrial and synaptic network dysfunction, infection, diabetes, sleep, hearing loss, the gut microbiome and neuroinflammation/ innate immune function as dementia targets are driving research in new directions bolstered by recent findings on the genetic, omics and systems biology associated with AD/dementia. In moving forward, lessons learnt from the amyloid debacle should be used to enhance the objective identification of AD/dementia therapeutics as a multifactorial disease syndrome.

Bisphenol A diglycidyl ether (BADGE), a derivative of bisphenol A (BPA), is widely used in the manufacture of epoxy resins as well as a coating on food containers. Recent studies have demonstrated the adverse effects of BADGE on reproduction and development in rodents and amphibians, but how BADGE affects biological activity is not understood. To gain a better understanding of the biological effects of BADGE exposure during development, we used the model organism Drosophila melanogaster and performed whole transcriptome sequencing. Interestingly, when Drosophila are raised on food containing BADGE, genes having significantly increased transcript numbers are enriched for those involved in regulating cell proliferation, including DNA replication and cell cycle control. Furthermore, raising larvae on BADGE-containing food induces hemocyte (blood cell) over-proliferation. This effect can be stimulated with even lower concentrations of BADGE if the hemocytes are already primed for cell proliferation by the expression of dominant active Ras GTPase. We conclude that chronic exposure to the xenobiotic BADGE throughout development can induce cell proliferation. Electronic supplementary material The online version of this article (10.1007/s11356-020-08899-7) contains supplementary material, which is available to authorized users.

The only drugs currently approved for the treatment of Alzheimer's Disease (AD) are four acetylcholinesterase inhibitors and the NMDA antagonist memantine. Apart from these drugs, which have minimal to no clinical benefit, the 40-year search for effective therapeutics to treat AD has resulted in a clinical failure rate of 100% not only for compounds that prevent brain amyloid deposition or remove existing amyloid plaques but also those acting by a variety of other putative disease-associated mechanisms. This indicates that the preclinical data generated from current AD targets to support the selection, optimization, and translation of new chemical entities (NCEs) and biologics to clinical trials is seriously compromised. While many of these failures reflect flawed hypotheses or a lack of adequate characterization of the preclinical pharmacodynamic and pharmacokinetic (PD/PK) properties of lead NCEs-including their bioavailability and toxicity-the conceptualization, validation, and interrogation of the current animal models of AD represent key limitations. The overwhelming majority of these AD models are transgenic, based on aspects of the amyloid hypothesis and the genetics of the familial form of the disease. As a result, these generally lack construct and predictive validity for the sporadic form of the human disease. The 170 or so transgenic models, perhaps the largest number ever focused on a single disease, use rodents, mainly mice, and in addition to amyloid also address aspects of tau causality with more complex multigene models including other presumed causative factors together with amyloid. This overview discusses the current animal models of AD in the context of both the controversies surrounding the causative role of amyloid in the disease and the need to develop validated models of cognitive function/dysfunction that more appropriately reflect the phenotype(s) of human aged-related dementias. © 2019 by John Wiley & Sons, Inc.

Edwardsiella andrillae is a sea anemone (Cnidaria: Anthozoa: Actiniaria) only known to live embedded in the ice at the seawater interface on the underside of the Ross Ice Shelf, Antarctica. Although the anatomy and morphological characteristics of E. andrillae have been described, the adaptations of this species to the under-ice ecosystem have yet to be examined. One feature that may be important to the physiology and ecology of E. andrillae is its microbiome, which may play a role in health and survival, as has been deduced in other metazoans, including anthozoans. Here we describe the microbiome of five specimens of E. andrillae, compare the diversity we recovered to that known for temperate anemones and another Antarctic cnidarian, and consider the phylogenetic and functional implications of microbial diversity for these animals. The E. andrillae microbiome was relatively low in diversity, with seven phyla detected, yet included substantial phylogenetic novelty. Among the five anemones investigated, the distribution of microbial taxa varied; this trait appears to be shared by many anthozoans. Most importantly, specimens either appeared to be dominated by Proteobacteria-affiliated members or by deeply branching Tenericute sequences. There were few closely related sequence types that were common to temperate and Antarctic sea anemone microbiomes, the exception being an Acinetobacter-related representative. Similar observations were made between microbes associated with E. andrillae and an Antarctic soft coral; however, there were several closely-related, low abundance Gammaproteobacteria in both Antarctic microbiomes, particularly from the soft coral, that are also commonly detected in Southern Ocean seawater. Although this preliminary study leaves open many questions concerning microbiome diversity and its role in host ecology, we identify major lineages of microbes (e.g., diverse deep-branching Alphaproteobacteria, Epsilonproteobacteria, and divergent Tenericutes affiliates) that may play critical roles, and we highlight the current understanding and the need for future studies of sea anemone-microbiome relationships.

CEP-32215 is a new, potent, selective, and orally bioavailable inverse agonist of the histamine H3 receptor (H3R) with drug-like properties. High affinity in human (hH3R Ki = 2.0 ± 0.2 nM) and rat (rH3R Ki = 3.6 ± 0.7 nM) H3R radioligand binding assays was demonstrated. Potent functional antagonism (Kb = 0.3 ± 0.1 nM) and inverse agonism (EC50 = 0.6 ± 0.2 nM) were demonstrated in [(35)S]guanosine 5(')-O-(γ-thio)-triphosphate binding assays. Oral bioavailability and dose-related exposure was consistent among rat, dog, and monkey. After oral dosing, occupancy of H3R by CEP-32215 was estimated by the inhibition of ex vivo binding in rat cortical slices (ED50 = 0.1 mg/kg p.o.). Functional antagonism in brain was demonstrated by the inhibition of R-α-methylhistamine-induced drinking in the rat dipsogenia model (ED50 = 0.92 mg/kg). CEP-32215 significantly increased wake duration in the rat EEG model at 3-30 mg/kg p.o. Increased motor activity, sleep rebound or undesirable events (such as spike wave or seizure activity) was not observed following doses up to 100 mg/kg p.o., indicating an acceptable therapeutic index. CEP-32215 may have potential utility in the treatment of a variety of sleep disorders. This article is part of the Special Issue entitled 'Histamine Receptors'.

Transmembrane protein 18 (TMEM18) is an ill-described, obesity-related gene, but few studies have explored its molecular function. We found single-nucleotide polymorphism data, suggesting that TMEM18 may be involved in the regulation/physiology of metabolic syndrome based on associations with insulin, homeostatic model assessment-β (HOMAβ), triglycerides, and blood sugar. We then found an ortholog in the Drosophila genome, knocked down Drosophila Tmem18 specifically in insulin-producing cells, and tested for its effects on metabolic function. Our results suggest that TMEM18 affects substrate levels through insulin and glucagon signaling, and its downregulation induces a metabolic state resembling type 2 diabetes. This work is the first to experimentally describe the metabolic consequences of TMEM18 knockdown, and further supports its association with obesity.

Phthalate diesters are commonly used as industrial plasticisers, as well as in cosmetics and skin care products, as a result people are constantly exposed to these xenobiotics. Recent epidemiological studies have found a correlation between circulating phthalate levels and type 2 diabetes, whereas animal studies indicate that phthalates are capable of disrupting endocrine signaling. Nonetheless, how phthalates interfere with metabolic function is still unclear. Here, we show that feeding Drosophila males the xenobiotic dibutyl phthalate (DBP) affects conserved insulin- and glucagon-like signaling. We report that raising flies on food containing DBP leads to starvation resistance, increased lipid storage, hyperglycemia, and hyperphagia. We go on to show that the starvation-resistance phenotype can be rescued by overexpression of the glucagon analogue adipokinetic hormone (Akh). Furthermore, although acute DBP exposure in adult flies is able to affect insulin levels, only chronic feeding influences Akh expression. We establish that raising flies on DBP-containing food or feeding adults DBP food affects the expression of homologous genes involved in xenobiotic and lipid metabolism (AHR [Drosophila ss], NR1I2 [Hr96], ABCB1 [MDR50], ABCC3 [MRP], and CYP3A4 [Cyp9f2]). Finally, we determined that the expression of these genes is also influenced by Akh. Our results provide comprehensive evidence that DBP can disrupt metabolism in Drosophila males, by regulating genes involved in glucose, lipid, and xenobiotic metabolism.

Concerns regarding the reliability of biomedical research outcomes were precipitated by two independent reports from the pharmaceutical industry that documented a lack of reproducibility in preclinical research in the areas of oncology, endocrinology, and hematology. Given their potential impact on public health, these concerns have been extensively covered in the media. Assessing the magnitude and scope of irreproducibility is limited by the anecdotal nature of the initial reports and a lack of quantitative data on specific failures to reproduce published research. Nevertheless, remediation activities have focused on needed enhancements in transparency and consistency in the reporting of experimental methodologies and results. While such initiatives can effectively bridge knowledge gaps and facilitate best practices across established and emerging research disciplines and therapeutic areas, concerns remain on how these improve on the historical process of independent replication in validating research findings and their potential to inhibit scientific innovation.

Alzheimer’s disease (AD) was differentiated from senile dementia (SD) in 1910 due to its early onset and pathological severity. In 1976, this distinction was upended when SD was redesignated as AD to focus efforts and funding in dementia-related research. AD then became conflated with amyloid plaques and, to a lesser degree, neurofibrillary tangles complicating efforts in understanding dementia causality and its treatment. The resultant four-decade search for therapies-based almost exclusively on amyloid was an exercise in futility. While dementia is a complex, multifactorial syndrome, AD is viewed as a homogeneous, linear disease. An amyloid-agnostic approach is necessary to discover therapeutics for age-related dementias.

Deposits of amyloid plaques and neurofibrillary tangles of aggregated tau in the brain represent key hallmarks of the neurodegenerative disorder, Alzheimer’s Disease (AD) and form the basis of the major hypotheses of AD causality. To date, therapeutics that reduce brain amyloid in AD patients have demonstrated no effect in reversing the associated decline in cognition or function indicating that the amyloid hypothesis is either incorrect or that there is a point when the disease becomes independent of Aβ production or is refractory to any type of therapeutic intervention. The clinical failures of inhibitors of tau aggregation, neurotransmitter modulators and drugs repurposed from AD-associated disease indications tend to support this latter viewpoint. Current understanding of AD causality is thus incomplete, a situation that has been compounded by a debate on whether AD is a singularly distinct form of dementia and by the dogmatic promotion of hypotheses over actual clinical data. The latter has repeatedly led to compounds lacking efficacy in Phase II trials being advanced into Phase III where their lack of efficacy is routinely recapitulated. This Commentary, the first of two, discusses amyloid and tau as putative drug targets for AD in the context of the prevalence and economic and social impact of this insidious neurodegenerative disease.

Compounds targeted for the treatment of Alzheimer's Disease (AD) have consistently failed in clinical trials despite evidence for target engagement and pharmacodynamic activity. This questions the relevance of compounds acting at current AD drug targets - the majority of which reflect the seminal amyloid and, to a far lesser extent, tau hypotheses - and limitations in understanding AD causality as distinct from general dementia. The preeminence of amyloid and tau led to many alternative approaches to AD therapeutics being ignored or underfunded to the extent that their causal versus contributory role in AD remains unknown. These include: neuronal network dysfunction; cerebrovascular disease; chronic, local or systemic inflammation involving the innate immune system; infectious agents including herpes virus and prion proteins; neurotoxic protein accumulation associated with sleep deprivation, circadian rhythm and glymphatic/meningeal lymphatic system and blood-brain-barrier dysfunction; metabolic related diseases including diabetes, obesity hypertension and hypocholesterolemia; mitochondrial dysfunction and environmental factors. As AD has become increasingly recognized as a multifactorial syndrome, a single treatment paradigm is unlikely to work in all patients. However, the biomarkers required to diagnose patients and parse them into mechanism/disease-based sub-groups remain rudimentary and unvalidated as do non-amyloid, non-tau translational animal models. The social and economic impact of AD is also discussed in the context of new FDA regulatory draft guidance and a proposed biomarker-based Framework (re)-defining AD and its stages as part of the larger landscape of treating dementia via the 2013 G8 initiative to identify a disease-modifying therapy for dementia/AD by 2025.

Many results reported in the biomedical research literature cannot be independently reproduced, undermining the basic foundations of science. This overview is intended for researchers who are committed to improving the quality and integrity of biomedical science by raising awareness of both the sources of irreproducibility, and activities specifically targeted to address the issue. The irreproducibility of biomedical research is due to a variety of factors, known and unknown, that markedly influence experimental outcomes. Among the known factors are inadequate training or mentoring, or experimenter incompetence. These may result in flawed experimental design and execution that reflect a lack of planning, leading to an underpowering of a study, an absence of appropriate controls, selective data analysis, inappropriate statistical analysis, and/or investigator bias or fraud. Another frequently overlooked source of irreproducibility is failure to ensure that the equipment used is performing up to manufacturer specifications. Failure to validate/authenticate the experimental reagents is another source of irreproducibility. These include compounds, cell lines, antibodies, and the animal models used in a study. This overview discusses how a lack of validation of research reagents negatively impact experimental reproducibility, and provides guidelines to avoid these pitfalls. © 2018 by John Wiley & Sons, Inc.

The identification of genetic variants responsible for behavioral variation is an enduring goal in biology, with wide-scale ramifications, ranging from medical research to evolutionary theory on personality syndromes. Here, we use for the first time a large-scale genetical genomics analysis in the brains of chickens to identify genes affecting anxiety as measured by an open field test. We combine quantitative trait locus (QTL) analysis in 572 individuals and expression QTL (eQTL) analysis in 129 individuals from an advanced intercross between domestic chickens and Red Junglefowl. We identify 10 putative quantitative trait genes affecting anxiety behavior. These genes were tested for an association in the mouse Heterogeneous Stock anxiety (open field) data set and human GWAS data sets for bipolar disorder, major depressive disorder, and schizophrenia. Although comparisons between species are complex, associations were observed for four of the candidate genes in mice and three of the candidate genes in humans. Using a multimodel approach we have therefore identified a number of putative quantitative trait genes affecting anxiety behavior, principally in chickens but also with some potentially translational effects as well. This study demonstrates that chickens are an excellent model organism for the genetic dissection of behavior.

Purpose: To provide a comprehensive and current review on the available experimental animal models of retinal vein occlusion (RVO) and to identify their strengths and limitations with the purpose of helping researchers to plan preclinical studies on RVO.Methods: A systematic review of the literature on experimental animal models of RVO was undertaken. Medline, SCOPUS, and Web of Science databases were searched. Studies published between January 1, 1965, and March 31, 2017, and that met the inclusion criteria were reviewed. The data extracted included animal species used, methods of inducing RVO, and the clinical and histopathologic features of the models, especially in relation to strengths, limitations, and faithfulness to clinical sequelae.Results: A total of 128 articles fulfilling the inclusion criteria were included. Several species were used to model human branch and central RVO (BRVO; CRVO) with nonhuman primates being the most common, followed by rodents and pigs. BRVO and CRVO were most commonly induced by laser photocoagulation and all models showed early features of clinical disease, including retinal hemorrhages and retinal edema. These features made many of the models adequate for studying the acute phase of BRVO and CRVO, although macular edema, retinal ischemia, and neovascular complications were observed in only a few experimental animal models (laser-induced model in rodents, pigs, and nonhuman primates, diathermy-induced model in pigs, and following intravitreal injection of PD0325901 in rabbits for BRVO; and in the laser-induced model in rodents, rabbits, and nonhuman primates, diathermy-induced model in nonhuman primates, following permanent ligation of the central retinal vein in nonhuman primates, and with intravitreal injection of thrombin in rabbits for CRVO).Conclusions: Experimental animal models of RVO are available to study the pathogenesis of this disease and to evaluate diagnostic/prognostic biomarkers and to develop new therapeutics. Data available suggest laser-induced RVO in pigs and rodents to be overall the best models of BRVO and the laser-induced RVO rodents the best model for CRVO.

In children with severe haemophilia A, inhibitors to factor VIII (FVIII) usually develop during the first 50 treatment exposure days and are classified as low or high titre depending on the peak inhibitor titre being greater or less than 5 Bethesda units/mL (BU/mL). Classification of the inhibitor may change with time, as some low-titre inhibitors progress to high titre following re-exposure to FVIII concentrate. The aim of this study was to investigate potential risk factors for such a progression in children with severe haemophilia A and newly diagnosed inhibitors. This study was a follow-up study of the PedNet Registry and included 260 children with severe haemophilia A and inhibitors born between 1990 and 2009 and recruited consecutively from 31 haemophilia centres. Clinical and laboratory data were collected from the date of each child's first positive inhibitor test for at least 3 years. At the time of first positive inhibitor test, 49% (n = 127) had low-titre inhibitors, with 50% of them progressing to high titre and only 25% maintaining low titres. The FVIII gene (F8) mutation type was known in 247 patients (95%), and included 202 (82%) null mutations. The progression to high-titre inhibitors was associated with null F8 mutations (odds ratio [OR]: 2.6; 95% confidence interval [CI]: 1.0–6.5), family history of inhibitors (OR: 7.2; 95% CI: 1.8–28.4) and the use of high-dose immune tolerance induction, defined as ≥100 IU FVIII concentrate/kg/d (OR: 3.9; 95% CI: 1.5–10.0). These results suggest that high-dose immune tolerance induction should be avoided as the initial strategy in patients who develop low-titre FVIII inhibitors.

Many neuronal circuits contain, or are influenced by, colocalized neurotransmitters, which are suggested to fine-tune circuit function. Much of our knowledge about cotransmission derives from studies in non-mammalian model systems, although a growing number of studies have provided insights into the physiological role of cotransmission in more complex mammalian circuits, including the spinal cord. Here we discuss the current picture of colocalized neurotransmitters in spinal circuits of non-mammalian and mammalian systems, as well as the progression towards understanding of their functional roles. We delineate both evolutionarily conserved and divergent aspects of spinal cotransmission and highlight open questions in the area.

Efforts to address reproducibility concerns in biomedical research include: initiatives to improve journal publication standards and peer review; increased attention to publishing methodological details that enable experiments to be reconstructed; guidelines on standards for study design, implementation, analysis and execution; meta-analyses of multiple studies within a field to synthesize a common conclusion and; the formation of consortia to adopt uniform protocols and internally reproduce data. Another approach to addressing reproducibility are Reproducibility Initiatives (RIs), well-intended, high-profile, systematically peer-vetted initiatives that are intended to replace the traditional process of scientific self-correction. Outcomes from the RIs reported to date have questioned the usefulness of this approach, particularly when the RI outcome differs from other independent self-correction studies that have reproduced the original finding. As a failed RI attempt is a single outcome distinct from the original study, it cannot provide any definitive conclusions necessitating additional studies that the RI approach has neither the ability nor intent of conducting making it a questionable replacement for self-correction. A failed RI attempt also has the potential to damage the reputation of the author of the original finding. Reproduction is frequently confused with replication, an issue that is more than semantic with the former denoting “similarity” and the latter an “exact copy” – an impossible outcome in research because of known and unknown technical, environmental and motivational differences between the original and reproduction studies. To date, the RI framework has negatively impacted efforts to improve reproducibility, confounding attempts to determine whether a research finding is real.

Maneuvering safely through the environment is central to survival of all animals. The ability to do this depends on learning and remembering locations. This capacity is encoded in the brain by two systems: one using cues outside the organism (distal cues), allocentric navigation, and one using self-movement, internal cues and sometimes proximal cues, egocentric navigation. Allocentric navigation involves the hippocampus, entorhinal cortex, and surrounding structures (e.g., subiculum); in humans this system encodes declarative memory (allocentric, semantic, and episodic, i.e., memory for people, places, things, and events). This form of memory is assessed in laboratory animals by many methods, but predominantly the Morris water maze (MWM). Egocentric navigation involves the dorsal striatum and connected structures; in humans this system encodes routes and integrated paths and when over-learned becomes implicit or procedural memory. Several allocentric methods for rodents are reviewed and compared with the MWM with particular focus on the Cincinnati water maze (CWM). MWM advantages include minimal training, no food deprivation, ease of testing, reliable learning, insensitivity to differences in body weight and appetite, absence of non-performers, control methods for performance effects, repeated testing capability and other factors that make this test well-suited for regulatory studies. MWM limitations are also reviewed. Evidence-based MWM design and testing methods are presented. On balance, the MWM is arguably the preferred test for assessing learning and memory in basic research and regulatory studies and the CWM is recommended if two tests can be accommodated so that both allocentric (MWM) and egocentric (CWM) learning and memory can be effectively and efficiently assessed.

The greatest challenge standing in the way of effective in vivo siRNA delivery is creating a delivery vehicle that mediates a high degree of efficacy with a broad therapeutic window. Key structure-activity relationships of a poly(amide) polymer conjugate siRNA delivery platform were explored to discover the optimized polymer parameters that yield the highest activity of mRNA knockdown in the liver. At the same time, the poly(amide) backbone of the polymers allowed for the metabolism and clearance of the polymer from the body very quickly, which was established using radiolabeled polymers to demonstrate the time course of biodistribution and excretion from the body. The fast degradation and clearance of the polymers provided for very low toxicity at efficacious doses, and the therapeutic window of this poly(amide)-based siRNA delivery platform was shown to be much broader than a comparable polymer platform. The results of this work illustrate that the poly(amide) platform has a promising future in the development of a siRNA-based drug approved for human use.

In Drosophila, serotonin (5-HT) regulates aggression, mating behaviour and sleep/wake behaviour through different receptors. Currently, how these various receptors are themselves regulated is still not completely understood. The KCTD12-family of proteins, which have been shown to modify G-protein-coupled receptor (GPCR) signalling in mammals, are one possibility of auxiliary proteins modulating 5-HT receptor signalling. The KCTD12-family was found to be remarkably conserved and present in species from C. elegans to humans. The Drosophila KCTD12 homologue Kctd12-like (Ktl) was highly expressed in both the larval and adult CNS. By performing behavioural assays in male Drosophila, we now reveal that Ktl is required for proper male aggression and mating behaviour. Previously, it was shown that Ktl is in a complex with the Drosophila 5-HT receptor 5-HT7, and we observed that both Ktl and the 5-HT1A receptor are required in insulin-producing cells (IPCs) for proper adult male behaviour, as well as for hyperaggressive activity induced by the mammalian 5-HT1A receptor agonist 8-hydroxy-2-dipropylaminotetralin-hydrobromide. Finally, we show that Ktl expression in the IPCs is necessary to regulate locomotion and normal sleep/wake patterns in Drosophila, but not the 5-HT1A receptor. Similar to what was observed with mammalian KCTD12-family members that interact physically with a GPCR receptor to regulate desensitization, in Drosophila Ktl may function in GPCR 5-HT receptor pathways to regulate their signalling, which is required for proper adult male behaviour.

The application of small interfering (si)RNAs as potential therapeutic agents requires safe and effective methods for their delivery to the cytoplasm of the target cells and tissues. Recent studies have shown significant progress in the development of targeting reagents that facilitate the recognition of, and siRNA delivery to, specific cell types. Among recently reported delivery approaches, polymers with amphipathic properties have been used to enable endosome escape and cytosolic delivery. Here, we describe a linear amphipathic poly(amido amine) polymer conjugate system for the efficient siRNA delivery in vitro and in vivo. This polymer contains a novel amine bearing bis-acrylamide monomer designed for increasing amine density, which resulted in substantial improvement in liver uptake and RNAi activity compared to our previously reported poly(amido amine disulfide) polymer.1 The activity for this liver targeted delivery system was demonstrated in rodents and nonhuman primates.

A pharmacological experiment is typically conducted to: i) test or expand a hypothesis regarding the potential role of a target in the mechanism(s) underlying a disease state using an existing drug or tool compound in normal and/or diseased tissue or animals; or ii) characterize and optimize a new chemical entity (NCE) targeted to modulate a specific disease-associated target to restore homeostasis as a potential drug candidate. Hypothesis testing necessitates an intellectually rigorous, null hypothesis approach that is distinct from a high throughput fishing expedition in search of a hypothesis. In conducting an experiment, the protocol should be transparently defined along with its powering, design, appropriate statistical analysis and consideration of the anticipated outcome (s) before it is initiated. Compound-target interactions often involve the direct study of phenotype(s) unique to the target at the cell, tissue or animal/human level. However, in vivo studies are often compromised by a lack of sufficient information on the compound pharmacokinetics necessary to ensure target engagement and also by the context-free analysis of ubiquitous cellular signaling pathways downstream from the target. The use of single tool compounds/drugs at one concentration in engineered cell lines frequently results in reductionistic data that have no physiologically relevance. This overview, focused on trends in the peer-reviewed literature, discusses the execution and reporting of experiments and the criteria recommended for the physiologically-relevant assessment of target engagement to identify viable new drug targets and facilitate the advancement of translational studies.

The receptor concept, now more than century old, remains the core concept in understanding the mechanisms of disease causality and drug action. Originally formulated in the early 1900s, receptor theory has evolved in both detail and complexity as the tools of molecular biology and increasingly sophisticated research technologies have facilitated the study of drug/ligand receptor interactions at the molecular level. The result has been a more detailed and nuanced understanding of the parameters of drug action that together with knowledge of the pleotropic nature of cellular targets and associated signaling pathways has reshaped the receptor concept. Added to the basic agonist/antagonist view of drug/ligand function, concepts like allosterism, inverse agonism, biased signaling and compound residence time have provided a broader understanding of compound-target interactions to better inform drug discovery efforts. The iteration of descriptive data from pharmacological experiments to provide generic indices of affinity and efficacy can now be used to predict expected drug/compound effects in other cell/organ systems where the sensitivity of the latter to drug effects (either via expression levels of cell surface receptors or variances in the efficiency with which those receptors are coupled to cytosolic metabolic processes) can be accommodated.

Animal models of disease represent the pinnacle of hierarchical research efforts to validate targets and compounds for therapeutic intervention. Yet models of asthma, particularly in the mouse, which, for practical reasons, has become the sine qua non of asthma research, have been a bone of contention for decades. With barely a nod to their limitations and an extensive history of translational failures, they continue to be used for target identification and to justify the clinical evaluation of new compounds. Recent improvements - including sensitization directly to the airways; use of more relevant allergens; development of a chronic rather than short-term condition; utilization of techniques to measure lung function beyond uninterpretable measures of airway hyperresponsiveness - are laudable but cannot bridge the chasm between the models and the myriad complexities of the human disorder and multiple asthma endophenotypes. While further model developments are necessary, including recognition of key environmental factors beyond allergens, the judicious integration with newer ex vivo and in vitro techniques, including human precision-cut lung slices, reprograming of patient-derived induced pluripotent stem cells and fibroblasts to epithelial and smooth muscle cells, and use of other clinical samples to create a more holistic depiction of activities, might improve their translational success.

In Drosophila, the monoamine octopamine, through mechanisms that are not completely understood, regulates both aggression and mating behavior. Interestingly, our study demonstrates that the Drosophila obesity-linked homologs Transcription factor AP-2 (TfAP-2; TFAP2B in humans) and Tiwaz (Twz; KCTD15 in humans) interact to modify male behavior by controlling the expression of Tyramine β-hydroxylase and Vesicular monanime transporter, genes necessary for octopamine production and secretion. Furthermore, we reveal that octopamine in turn regulates aggression through the Drosophila cholecystokinin satiation hormone homolog Drosulfakinin (Dsk). Finally, we establish that TfAP-2 is expressed in octopaminergic neurons known to control aggressive behavior and that TfAP-2 requires functional Twz for its activity. We conclude that genetically manipulating the obesity-linked homologs TfAP-2 and Twz is sufficient to affect octopamine signaling, which in turn modulates Drosophila male behavior through the regulation of the satiation hormone Dsk.

The translational sciences represent the core element in enabling and utilizing the output from the biomedical sciences and to improving drug discovery metrics by reducing the attrition rate as compounds move from preclinical research to clinical proof of concept. Key to understanding the basis of disease causality and to developing therapeutics is an ability to accurately diagnose the disease and to identify and develop safe and effective therapeutics for its treatment. The former requires validated biomarkers and the latter, qualified targets. Progress has been hampered by semantic issues, specifically those that define the end product, and by scientific issues that include data reliability, an overt reductionistic cultural focus and a lack of hierarchically integrated data gathering and systematic analysis. A necessary framework for these activities is represented by the discipline of pharmacology, efforts and training in which require recognition and revitalization.

Exposure to bisphenol A (BPA) in rodents was shown to induce obesity, yet the mechanism by which BPA might induce obesity is still unclear. We employed the genetically tractable model organism, Drosophila melanogaster, to test the effects of raising them on food containing various concentrations of BPA. Of note, raising males on food containing BPA were susceptible to starvation, possibly by inhibiting their ability to perform lipolysis during starvation, leading to significantly increased lipid content after 24 hr of fasting. Furthermore, feeding males with BPA significantly inhibited the expression of insulin-like peptides. From these results, we conclude that BPA may inhibit lipid recruitment during starvation in Drosophila.

In this study, the distribution of steroid hormones, phytoestrogens, and estrogenic activity was thoroughly characterized within the anaerobic waste lagoon of a typical commercial swine sow operation. Three independent rounds of sampling were conducted in June 2009, April 2010, and February 2011. Thirty-seven analytes in lagoon slurry and sludge were assessed using LC/MS-MS, and yeast estrogen screen was used to determine estrogenic activity. Of the hormone analytes, steroidal estrogens were more abundant than androgens or progesterone, with estrone being the predominant estrogen species. Conjugated hormones were detected only at low levels. The isoflavone metabolite equol was by far the predominant phytoestrogen species, with daidzein, genistein, formononetin, and coumestrol present at lower levels. Phytoestrogens were often more abundant than steroidal estrogens, but contributed minimally toward total estrogenic activity. Analytes were significantly elevated in the solid phases of the lagoon; although low observed log KOC values suggest enhanced solubility in the aqueous phase, perhaps due to dissolved or colloidal organic carbon. The association with the solid phase, as well as recalcitrance of analytes to anaerobic degradation, results in a markedly elevated load of analytes and estrogenic activity within lagoon sludge. Overall, findings emphasize the importance of adsorption and transformation processes in governing the fate of these compounds in lagoon waste, which is ultimately used for broadcast application as a fertilizer.

The biochemical perturbations in diabetes mellitus (DM) create the conditions for the production of free radicals, the consequence of which is increased oxidative stress. Evidence has accrued over the past 2 decades that suggests that oxidative stress is an important pathogenetic factor in the development of diabetic retinopathy (DR). Experimental data show that the use of strategies that ameliorate oxidative stress can prevent and retard the development of DR in the animal model. Clinical observations also suggest that reducing oxidative stress may help to reverse pathological manifestations of DR. The present article constitutes an examination of the role of antioxidants in the management of DR and the current state of clinically relevant knowledge.

The worldwide incidence of Alzheimer's disease (AD) is increasing with estimates that 115 million individuals will have AD by 2050, creating an unsustainable healthcare challenge due to a lack of effective treatment options highlighted by multiple clinical failures of agents designed to reduce the brain amyloid burden considered synonymous with the disease. The amyloid hypothesis that has been the overarching focus of AD research efforts for more than two decades has been questioned in terms of its causality but has not been unequivocally disproven despite multiple clinical failures, This is due to issues related to the quality of compounds advanced to late stage clinical trials and the lack of validated biomarkers that allow the recruitment of AD patients into trials before they are at a sufficiently advanced stage in the disease where therapeutic intervention is deemed futile. Pursuit of a linear, reductionistic amyloidocentric approach to AD research, which some have compared to a religious faith, has resulted in other, equally plausible but as yet unvalidated AD hypotheses being underfunded leading to a disastrous roadblock in the search for urgently needed AD therapeutics. Genetic evidence supporting amyloid causality in AD is reviewed in the context of the clinical failures, and progress in tau-based and alternative approaches to AD, where an evolving modus operandi in biomedical research fosters excessive optimism and a preoccupation with unproven, and often ephemeral, biomarker/genome-based approaches that override transparency, objectivity and data-driven decision making, resulting in low probability environments where data are subordinate to self propagating hypotheses.

The Nimrod gene cluster, located on the second chromosome of Drosophila melanogaster, is the largest synthenic unit of the Drosophila genome. Nimrod genes show blood cell specific expression and code for phagocytosis receptors that play a major role in fruit fly innate immune functions. We previously identified three homologous genes (vajk-1, vajk-2 and vajk-3) located within the Nimrod cluster, which are unrelated to the Nimrod genes, but are homologous to a fourth gene (vajk-4) located outside the cluster. Here we show that, unlike the Nimrod candidates, the Vajk proteins are expressed in cuticular structures of the late embryo and the late pupa, indicating that they contribute to cuticular barrier functions.

Because colorectal cancer (CRC) remains a leading cause of cancer mortality worldwide, more accessible screening tests are urgently needed to identify early stage lesions. We hypothesized that highly sensitive, metabolic profile analysis of stool samples will identify metabolites associated with early stage lesions and could serve as a noninvasive screening test. We therefore applied traveling wave ion mobility mass spectrometry (TWIMMS) coupled with ultraperformance liquid chromatography (UPLC) to investigate metabolic aberrations in stool samples in a transgenic model of premalignant polyposis aberrantly expressing the gene encoding the high mobility group A (Hmga1) chromatin remodeling protein. Here, we report for the first time that the fecal metabolome of Hmga1 mice is distinct from that of control mice and includes metabolites previously identified in human CRC. Significant alterations were observed in fatty acid metabolites and metabolites associated with bile acids (hypoxanthine xanthine, taurine) in Hmga1 mice compared to controls. Surprisingly, a marked increase in the levels of distinctive short, arginine-enriched, tetra-peptide fragments was observed in the transgenic mice. Together these findings suggest that specific metabolites are associated with Hmga1-induced polyposis and abnormal proliferation in intestinal epithelium. Although further studies are needed, these data provide a compelling rationale to develop fecal metabolomic analysis as a noninvasive screening tool to detect early precursor lesions to CRC in humans.

Zizimin-related (Zir), a Rho guanine nucleotide exchange factor (RhoGEF) homologous to the mammalian Dock-C/Zizimin-related family, was identified in a screen to find new genes involved in the Drosophila melanogaster cellular immune response against eggs from the parasitoid wasp Leptopilina boulardi. RhoGEFs activate Rho-family GTPases, which are known to be central regulators of cell migration, spreading and polarity. When a parasitoid wasp is recognized as foreign, multiple layers of circulating immunosurveillance cells (haemocytes) should attach to the egg. In Zir mutants this process is disrupted and lamellocytes, a haemocyte subtype, fail to properly encapsulate the wasp egg. Furthermore, macrophage-like plasmatocytes exhibit a strong reduction in their ability to phagocytise Escherichia coli and Staphylococcus aureus bacteria. During encapsulation and phagocytosis Zir genetically interacts with two Rho-family GTPases, Rac2 and Cdc42. Finally, Zir is dispensable for the humoral immune response against bacteria. We propose that Zir is necessary to activate the Rho-family GTPases Rac2 and Cdc42 during the Drosophila cellular immune response.

With the current explosion of knowledge on the role of mitochondrial dysfunction in the genesis of various human disease states, there is an increased interest in targeting mitochondrial processes, pathways, and proteins for drug discovery efforts in cancer and cardiovascular, metabolic, and central nervous system diseases, the latter including autism and neurodegenerative diseases. We provide an update on understanding the central role of the mitochondrion in ATP and reactive oxygen species production and in controlling cell death pathways.

Genome-wide association studies have identified a number of genes associated with human body weight. While some of these genes are large fields within obesity research, such as MC4R, POMC, FTO and BDNF, the majority do not have a clearly defined functional role explaining why they may affect body weight. Here, we searched biological databases and discovered 33 additional genes associated with human obesity (CADM2, GIPR, GPCR5B, LRP1B, NEGR1, NRXN3, SH2B1, FANCL, GNPDA2, HMGCR, MAP2K5, NUDT3, PRKD1, QPCTL, TNNI3K, MTCH2, DNAJC27, SLC39A8, MTIF3, RPL27A, SEC16B, ETV5, HMGA1, TFAP2B, TUB, ZNF608, FAIM2, KCTD15, LINGO2, POC5, PTBP2, TMEM18, TMEM160). We find that the majority have orthologues in distant species, such as D. melanogaster and C. elegans, suggesting that they are important for the biology of most bilateral species. Intriguingly, signalling cascade genes and transcription factors are enriched among these obesity genes, and several of the genes show properties that could be useful for potential drug discovery. In this review, we demonstrate how information from several distant model species, interactomics and signalling pathway analysis represents an important way to better understand the functional diversity of the surprisingly high number of molecules that seem to be important for human obesity.

The successful transition of drug-like new chemical entities from discovery to clinical trials coupled with real-time feedback from the latter represents a key element for success in drug discovery. Now designated as T1 translational medicine, this process has, similar to other recent solutions to improve productivity, been hyped as a novel discipline, despite the fact that many of its component activities have existed in the pharmacological sciences for many decades. Instead of proselytizing translational medicine, the priority is to improve the quality of the science and decision-making processes involved in advancing compounds to ensure that what is translated has value.

Preclinical characterization of new chemical entities (NCEs) in terms of efficacy, safety and their pharmacokinetic (PK), pharmacodynamic and pharmaceutical properties, is key to advancing appropriate compounds to clinical trials. The use of high throughput synthetic and screening methodologies has frequently led to NCE characterization becoming highly reductionistic, to the extent that compounds are often selected without adequate characterization. Classical, null hypothesis-based approaches involving the use of concentration/dose response curves and antagonists have been replaced by more qualitative approaches that limit NCE characterization. The return to a more integrated, hierarchical and pharmacologically driven approach will aid in ensuring that the NCEs advanced to clinical status are better understood, strengthening the process and predictivity of the translational approach in drug discovery.

Drosophila larvae react against eggs from the endoparasitoid wasp Leptopilina boulardi by surrounding them in a multilayered cellular capsule. Once a wasp egg is recognized as foreign, circulating macrophage-like cells, known as plasmatocytes, adhere to the invader. After spreading around the wasp egg, plasmatocytes form cellular junctions between the cells, effectively separating the egg from the hemocoel. Next, a second sub-type of circulating immunosurveillance cell (hemocyte), known as lamellocytes, adhere to either the wasp egg or more likely the plasmatocytes surrounding the egg. From these events, it is obvious that adhesion and cell shape change are an essential part of Drosophila's cellular immune response against parasitoid wasp eggs. To date, very few genes have been described as being necessary for a proper anti-parasitization response in Drosophila. With this in mind, we performed a directed genetic miniscreen to discover new genes required for this response. Many of the genes with an encapsulation defect have mammalian homologues involved in cellular adhesion, wound healing, and thrombosis, including extracellular matrix proteins, cellular adhesion molecules, and small GTPases.

Optimization of a novel series of pyridazin-3-one histamine H(3) receptor (H(3)R) antagonists/inverse agonists identified 6-{4-[3-(R)-2-methylpyrrolidin-1-yl)propoxy]phenyl}-2H-pyridazin-3-one (8a, CEP-26401; irdabisant) as a lead candidate for potential use in the treatment of attentional and cognitive disorders. 8a had high affinity for both human (K(i) = 2.0 nM) and rat (K(i) = 7.2 nM) H(3)Rs with greater than 1000-fold selectivity over the hH(1)R, hH(2)R, and hH(4)R histamine receptor subtypes and against an in vitro panel of 418 G-protein-coupled receptors, ion channels, transporters, and enzymes. 8a demonstrated ideal pharmaceutical properties for a CNS drug in regard to water solubility, permeability and lipophilicity and had low binding to human plasma proteins. It weakly inhibited recombinant cytochrome P450 isoforms and human ether-a-go-go-related gene. 8a metabolism was minimal in rat, mouse, dog, and human liver microsomes, and it had good interspecies pharmacokinetic properties. 8a dose-dependently inhibited H(3)R agonist-induced dipsogenia in the rat (ED(50) = 0.06 mg/kg po). On the basis of its pharmacological, pharmaceutical, and safety profiles, 8a was selected for preclinical development. The clinical portions of the single and multiple ascending dose studies assessing safety and pharmacokinetics have been completed allowing for the initiation of a phase IIa for proof of concept.

The need to develop effective treatments for Alzheimer's disease has been confounded by repeated clinical failures where promising new chemical entities that have been extensively characterized in preclinical models of Alzheimer's disease have failed to show efficacy in the human disease state. This has been attributed to: the selection of drug targets that have yet to be shown as causal to the disease as distinct from being the result of the disease process, a lack of congruence in the animal models of Alzheimer's disease, wild-type and transgenic, to the human disease, and the enrollment of patients in proof of concept clinical trials who are at too advanced a stage of the disease to respond to any therapeutic. The development of validated biomarkers that can be used for disease diagnosis and progression is anticipated to improve patient enrollment in clinical trials, to develop new animal models and to identify new disease targets for drug discovery. The present review assesses the status of current efforts in developing CSF biomarkers for Alzheimer's disease and briefly discusses the status of CSF biomarker efforts in schizophrenia, depression, Parkinson's disease and multiple sclerosis.

The studies described here are the first to evaluate the in vitro and in vivo properties of (111)In-CHX-A''-panitumumab for radioimmunotherapy (alpha- and beta(-)-emitters) and radioimmunoimaging (single photon emission computed tomography and positron emission tomography).Twenty-seven human carcinoma cell lines were analysed for expression of epidermal growth factor receptors by flow cytometry. Panitumumab was conjugated with CHX-A''-DTPA (diethylenetriamine-pentaacetic acid) and radiolabelled with (111)In. Immunoreactivity of the CHX-A''-DTPA-panitumumab and (111)In-CHX-A''-DTPA-panitumumab was evaluated by radioimmunoassays. Tumour targeting was determined in vivo by direct quantitation of tumour and normal tissues and by gamma-scintigraphy.For 26 of 27 human tumour cell lines, 95% of the cells expressed epidermal growth factor receptors over a range of intensity. Immunoreactivity of panitumumab was retained after modification with CHX-A''-DTPA. Radiolabelling of the immunoconjugate with (111)In was efficient with a specific activity of 19.5 +/- 8.9 mCi.mg(-1) obtained. Immunoreactivity and specificity of binding of the (111)In-panitumumab was shown with A431 cells. Tumour targeting by (111)In-panitumumab was demonstrated in athymic mice bearing A431, HT-29, LS-174T, SHAW or SKOV-3 s.c. xenografts with little uptake observed in normal tissues. The (111)In-panitumumab was also evaluated in non-tumour-bearing mice. Pharmacokinetic studies compared the plasma retention time of the (111)In-panitumumab in both non-tumour-bearing and A431 tumour-bearing mice. Tumour targeting was also visualized by gamma-scintigraphy.Panitumumab can be efficiently radiolabelled with (111)In with high labelling yields. Based on the efficiency in tumour targeting and low normal tissue uptake, panitumumab may be an effective targeting component for radioimmunodiagnostic and radioimmunotherapeutic applications.

The history of drug discovery spans approximately 200,000 years. For much of this time, the identification of therapeutic agents was empirical, with the shift to a more hypothesis-driven approach occurring in the late 19th century. Since then, the objective has changed from identifying an active drug and its mechanism of action to determining therapeutic potential only after identifying drug-like compounds that interact with a target site. Although the emphasis on target identification, or "targephilia," has yielded novel drugs, overall it appears to have slowed the drug discovery process, especially for compounds used in treating central nervous system (CNS) disorders. This is because the "targephilic" approach requires a good understanding of target physiology and its integration with the target organ, with a hierarchical integration from in vitro cellular and functional tissue studies to animal models that reasonably predict human responses. Because the majority of CNS drugs were discovered empirically, drug discovery in this area appears less amenable to target-based approaches than it seems for other types of therapeutics. Improving the success rate in CNS drug discovery requires a more pharmacometric-based approach, with a renewed emphasis on defining basic CNS function in intact animals and a more systematic in vivo screening of novel structures. Efforts must also be directed toward defining the sites of action of existing CNS drugs to aid in the design of second-generation agents with improved efficacy and safety.

Drosophila is an extremely useful model organism for understanding how innate immune mechanisms defend against microbes and parasitoids. Large foreign objects trigger a potent cellular immune response in Drosophila larva. In the case of endoparasitoid wasp eggs, this response includes hemocyte proliferation, lamellocyte differentiation and eventual encapsulation of the egg. The encapsulation reaction involves the attachment and spreading of hemocytes around the egg, which requires cytoskeletal rearrangements, changes in adhesion properties and cell shape, as well as melanization of the capsule. Guanine nucleotide metabolism has an essential role in the regulation of pathways necessary for this encapsulation response. Here, we show that the Drosophila inosine 5'-monophosphate dehydrogenase (IMPDH), encoded by raspberry (ras), is centrally important for a proper cellular immune response against eggs from the parasitoid wasp Leptopilina boulardi. Notably, hemocyte attachment to the egg and subsequent melanization of the capsule are deficient in hypomorphic ras mutant larvae, which results in a compromised cellular immune response and increased survival of the parasitoid.

Neuronal nicotinic acetylcholine receptors (nAChRs) have been a target for drug discovery efforts, primarily for CNS indications, for the past two decades. While nicotine and related natural products have been used for smoking cessation in various formulations (e.g., gum, spray, patches), it was only in 2006 with the launch of varenicline (Chantix) by Pfizer for smoking cessation that a new chemical entity (NCE) originating from a rational medicinal chemistry effort targeting neuronal AChRs was approved. The current overview outlines the chronology of drug discovery efforts in nAChRs from the cloning of the receptor family in the 1980s, to initial research efforts at SIBIA, R.J. Reynolds and Abbott, to the current industry-wide interest in nAChR agonists as novel therapeutics for pain, schizophrenia and Alzheimer's Disease. Key events in the evolution of the nAChR field were the development of high throughput electrophysiological screening tools that provided the means to enable lead optimization efforts in medicinal chemistry and the discovery by John Daly at the NIH of the frog alkaloid, epibatidine, that provided the framework for the discovery of ABT-594, an alpha4beta2 agonist that is 200 times more potent than morphine as an analgesic. Over the next decade, it is anticipated that additional NCEs including antagonists and allosteric modulators (both positive and negative), interacting with various nAChR subtypes, will be advanced to the clinic in areas of high unmet medical need, e.g., pain, neurodegeneration, to provide novel medications with improved efficacy.