Annual Review of Pharmacology and Toxicology - Volume 57, 2017

Development of scientific creativity is often tied closely to mentorship. In my case, two years with Julius Axelrod, the sum total of my research training, was transformative. My mentoring generations of graduate students and postdoctoral fellows has been as nurturing for me as it has been for them. Work in our lab over fifty years has covered the breadth of neurotransmitters and related substances, focusing on the discovery and characterization of novel messenger molecules. I can't conceptualize a more rewarding professional life.

Major advances in scientific discovery and insights can result from the development and use of new techniques, as exemplified by the work of Solomon Snyder, who writes a prefatory article in this volume. The Editors have chosen “New Methods and Novel Therapeutic Approaches in Pharmacology and Toxicology” as the Theme for a number of articles in this volume. These include ones that review the development and use of new experimental tools and approaches (e.g., nanobodies and techniques to explore protein-protein interactions), new types of therapeutics (e.g., aptamers and antisense oligonucleotides), and systems pharmacology, which assembles (big) data derived from omics studies together with information regarding drugs and patients. The application of these new methods and therapeutic approaches has the potential to have a major impact on basic and clinical research in pharmacology and toxicology as well as on patient care.

Ligand-induced activation of G protein–coupled receptors (GPCRs) is a key mechanism permitting communication between cells and organs. Enormous progress has recently elucidated the structural and dynamic features of GPCR transmembrane signaling. Nanobodies, the recombinant antigen–binding fragments of camelid heavy-chain-only antibodies, have emerged as important research tools to lock GPCRs in particular conformational states. Active-state stabilizing nanobodies have elucidated several agonist-bound structures of hormone-activated GPCRs and have provided insight into the dynamic character of receptors. Nanobodies have also been used to stabilize transient GPCR transmembrane signaling complexes, yielding the first structural insights into GPCR signal transduction across the cellular membrane. Beyond their in vitro uses, nanobodies have served as conformational biosensors in living systems and have provided novel ways to modulate GPCR function. Here, we highlight several examples of how nanobodies have enabled the study of GPCR function and give insights into potential future uses of these important tools.

Protein-protein interactions are fundamental for virtually all functions of the cell. A large fraction of these interactions involve short peptide motifs, and there has been increased interest in targeting them using peptide-based therapeutics. Peptides benefit from being specific, relatively safe, and easy to produce. They are also easy to modify using chemical synthesis and molecular biology techniques. However, significant challenges remain regarding the use of peptides as therapeutic agents. Identification of peptide motifs is difficult, and peptides typically display low cell permeability and sensitivity to enzymatic degradation. In this review, we outline the principal high-throughput methodologies for motif discovery and describe current methods for overcoming pharmacokinetic and bioavailability limitations.

Aptamers are single-stranded nucleic acid molecules that bind to and inhibit proteins and are commonly produced by systematic evolution of ligands by exponential enrichment (SELEX). Aptamers undergo extensive pharmacological revision, which alters affinity, specificity, and therapeutic half-life, tailoring each drug for a specific clinical need. The first therapeutic aptamer was described 25 years ago. Thus far, one aptamer has been approved for clinical use, and numerous others are in preclinical or clinical development. This review presents a short history of aptamers and SELEX, describes their pharmacological development and optimization, and reviews potential treatment of diseases including visual disorders, thrombosis, and cancer.

Recent studies have led to a greater appreciation of the diverse roles RNAs play in maintaining normal cellular function and how they contribute to disease pathology, broadening the number of potential therapeutic targets. Antisense oligonucleotides are the most direct means to target RNA in a selective manner and have become an established platform technology for drug discovery. There are multiple molecular mechanisms by which antisense oligonucleotides can be used to modulate RNAs in cells, including promoting the degradation of the targeted RNA or modulating RNA function without degradation. Antisense drugs utilizing various antisense mechanisms are demonstrating therapeutic potential for the treatment of a broad variety of diseases. This review focuses on some of the advances that have taken place in translating antisense technology from the bench to the clinic.

Protein homeostasis networks are highly regulated systems responsible for maintaining the health and productivity of cells. Whereas therapeutics have been developed to disrupt protein homeostasis, more recently identified techniques have been used to repurpose homeostatic networks to effect degradation of disease-relevant proteins. Here, we review recent advances in the use of small molecules to degrade proteins in a selective manner. First, we highlight all-small-molecule techniques with direct clinical application. Second, we describe techniques that may find broader acceptance in the biomedical research community that require little or no synthetic chemistry. In addition to serving as innovative research tools, these new approaches to control intracellular protein levels offer the potential to develop novel therapeutics targeting proteins that are not currently pharmaceutically vulnerable.

Stem cells are critical to maintaining steady-state organ homeostasis and regenerating injured tissues. Recent intriguing reports implicate extracellular vesicles (EVs) as carriers for the distribution of morphogens and growth and differentiation factors from tissue parenchymal cells to stem cells, and conversely, stem cell–derived EVs carrying certain proteins and nucleic acids can support healing of injured tissues. We describe approaches to make use of engineered EVs as technology platforms in therapeutics and diagnostics in the context of stem cells. For some regenerative therapies, natural and engineered EVs from stem cells may be superior to single-molecule drugs, biologics, whole cells, and synthetic liposome or nanoparticle formulations because of the ease of bioengineering with multiple factors while retaining superior biocompatibility and biostability and posing fewer risks for abnormal differentiation or neoplastic transformation. Finally, we provide an overview of current challenges and future directions of EVs as potential therapeutic alternatives to cells for clinical applications.

Drug discovery and development is a lengthy and expensive process. Although no one, simple, single solution can significantly accelerate this process, steps can be taken to avoid unnecessary delays. Using the development of antiviral therapies as a model, we describe options for acceleration that cover target selection, assay development and high-throughput screening, hit confirmation, lead identification and development, animal model evaluations, toxicity studies, regulatory issues, and the general drug discovery and development infrastructure. Together, these steps could result in accelerated timelines for bringing antiviral therapies to market so they can treat emerging infections and reduce human suffering.

There are many challenges along the path to the approval of new drugs to treat CNS disorders, one of the greatest areas of unmet medical need with a large societal burden and health-care impact. Unfortunately, over the past two decades, few CNS drug approvals have succeeded, leading many pharmaceutical companies to deprioritize this therapeutic area. The reasons for the failures in CNS drug discovery are likely to be multifactorial. However, selecting the most biologically plausible molecular targets that are relevant to the disorder is a critical first step to improve the probability of success. In this review, we outline previous methods for identifying and validating novel targets for CNS drug discovery, and, cognizant of previous failures, we discuss potential new strategies that may improve the probability of success of developing novel treatments for CNS disorders.

Safe and efficacious vaccines are arguably the most successful medical interventions of all time. Yet the ongoing discovery of new pathogens, along with emergence of antibiotic-resistant pathogens and a burgeoning population at risk of such infections, imposes unprecedented public health challenges. To meet these challenges, innovative strategies to discover and develop new or improved anti-infective vaccines are necessary. These approaches must intersect the most meaningful insights into protective immunity and advanced technologies with capabilities to deliver immunogens for optimal immune protection. This goal is considered through several recent advances in host-pathogen relationships, conceptual strides in vaccinology, and emerging technologies. Given a clear and growing risk of pandemic disease should the threat of infection go unmet, developing vaccines that optimize protective immunity against high-priority and antibiotic-resistant pathogens represents an urgent and unifying imperative.

Proprotein convertase subtilisin/kexin type-9 (PCSK9) is a secreted zymogen expressed primarily in the liver. PCSK9 circulates in plasma, binds to cell surface low-density lipoprotein (LDL) receptors, is internalized, and then targets the receptors to lysosomal degradation. Studies of naturally occurring PCSK9 gene variants that caused extreme plasma LDL cholesterol (LDL-C) deviations and altered atherosclerosis risk unleashed a torrent of biological and pharmacological research. Rapid progress in understanding the physiological regulation of PCSK9 was soon translated into commercially available biological inhibitors of PCSK9 that reduced LDL-C levels and likely also cardiovascular outcomes. Here we review the swift evolution of PCSK9 from novel gene to drug target, to animal and human testing, and finally to outcome trials and clinical applications. In addition, we explore how the genetics-guided path to PCSK9 inhibitor development exemplifies a new paradigm in pharmacology. Finally, we consider some potential challenges as PCSK9 inhibition becomes established in the clinic.

Systems pharmacology aims to holistically understand mechanisms of drug actions to support drug discovery and clinical practice. Systems pharmacology modeling (SPM) is data driven. It integrates an exponentially growing amount of data at multiple scales (genetic, molecular, cellular, organismal, and environmental). The goal of SPM is to develop mechanistic or predictive multiscale models that are interpretable and actionable. The current explosions in genomics and other omics data, as well as the tremendous advances in big data technologies, have already enabled biologists to generate novel hypotheses and gain new knowledge through computational models of genome-wide, heterogeneous, and dynamic data sets. More work is needed to interpret and predict a drug response phenotype, which is dependent on many known and unknown factors. To gain a comprehensive understanding of drug actions, SPM requires close collaborations between domain experts from diverse fields and integration of heterogeneous models from biophysics, mathematics, statistics, machine learning, and semantic webs. This creates challenges in model management, model integration, model translation, and knowledge integration. In this review, we discuss several emergent issues in SPM and potential solutions using big data technology and analytics. The concurrent development of high-throughput techniques, cloud computing, data science, and the semantic web will likely allow SPM to be findable, accessible, interoperable, reusable, reliable, interpretable, and actionable.

Widespread acceptance of evidence-based medicine has led to the proliferation of clinical practice guidelines as the primary mode of communicating current best practices across a range of chronic diseases. Despite overwhelming evidence supporting the benefits of their use, there is a long history of poor uptake by providers. Nonadherence to clinical practice guidelines is referred to as clinical inertia and represents provider failure to initiate or intensify treatment despite a clear indication to do so. Here we review evidence for the ubiquity of clinical inertia across a variety of chronic health conditions, as well as the organizational and system, patient, and provider factors that serve to maintain it. Limitations are highlighted in the emerging literature examining interventions to reduce clinical inertia. An evidence-based framework to address these limitations is proposed that uses behavior change theory and advocates for shared decision making and enhanced guideline development and dissemination.

Cannabis enables and enhances the subjective sense of well-being by stimulating the endocannabinoid system (ECS), which plays a key role in modulating the response to stress, reward, and their interactions. However, over time, repeated activation of the ECS by cannabis can trigger neuroadaptations that may impair the sensitivity to stress and reward. This effect, in vulnerable individuals, can lead to addiction and other adverse consequences. The recent shift toward legalization of medical or recreational cannabis has renewed interest in investigating the physiological role of the ECS as well as the potential health effects, both adverse and beneficial, of cannabis. Here we review our current understanding of the ECS and its complex physiological roles. We discuss the implications of this understanding vis-á-vis the ECS's modulation of stress and reward and its relevance to mental disorders in which these processes are disrupted (i.e., addiction, depression, posttraumatic stress disorder, schizophrenia), along with the therapeutic potential of strategies to manipulate the ECS for these conditions.

Originally, organophosphorus (OP) toxicology consisted of acetylcholinesterase inhibition by insecticides and chemical threat agents acting as phosphorylating agents for serine in the catalytic triad, but this is no longer the case. Other serine hydrolases can be secondary OP targets, depending on the OP structure, and include neuropathy target esterase, lipases, and endocannabinoid hydrolases. The major OP herbicides are glyphosate and glufosinate, which act in plants but not animals to block aromatic amino acid and glutamine biosynthesis, respectively, with safety for crops conferred by their expression of herbicide-tolerant targets and detoxifying enzymes from bacteria. OP fungicides, pharmaceuticals including calcium retention agents, industrial chemicals, and cytochrome P450 inhibitors act by multiple noncholinergic mechanisms, often with high potency and specificity. One type of OP-containing fire retardant forms a highly toxic bicyclophosphate γ-aminobutyric acid receptor antagonist upon combustion. Some OPs are teratogenic, mutagenic, or carcinogenic by known mechanisms that can be avoided as researchers expand knowledge of OP chemistry and toxicology for future developments in bioregulation.

Despite the unprecedented Ebola virus outbreak response in West Africa, no Ebola medical countermeasures have been approved by the US Food and Drug Administration. However, multiple valuable lessons have been learned about the conduct of clinical research in a resource-poor, high risk–pathogen setting. Numerous therapeutics were explored or developed during the outbreak, including repurposed drugs, nucleoside and nucleotide analogues (BCX4430, brincidofovir, favipiravir, and GS-5734), nucleic acid–based drugs (TKM-Ebola and AVI-7537), and immunotherapeutics (convalescent plasma and ZMapp). We review Ebola therapeutics progress in the aftermath of the West Africa Ebola virus outbreak and attempt to offer a glimpse of a path forward.

Protease-activated receptors (PARs) are a unique class of G protein–coupled receptors (GPCRs) that transduce cellular responses to extracellular proteases. PARs have important functions in the vasculature, inflammation, and cancer and are important drug targets. A unique feature of PARs is their irreversible proteolytic mechanism of activation that results in the generation of a tethered ligand that cannot diffuse away. Despite the fact that GPCRs have proved to be the most successful class of druggable targets, the development of agents that target PARs specifically has been challenging. As a consequence, researchers have taken a remarkable diversity of approaches to develop pharmacological entities that modulate PAR function. Here, we present an overview of the diversity of therapeutic agents that have been developed against PARs. We further discuss PAR biased signaling and the influence of receptor compartmentalization, posttranslational modifications, and dimerization, which are important considerations for drug development.

Macroautophagy (hereafter called autophagy) is a vacuolar, lysosomal pathway for catabolism of intracellular material that is conserved among eukaryotic cells. Autophagy plays a crucial role in tissue homeostasis, adaptation to stress situations, immune responses, and the regulation of the inflammatory response. Blockade or uncontrolled activation of autophagy is associated with cancer, diabetes, obesity, cardiovascular disease, neurodegenerative disease, autoimmune disease, infection, and chronic inflammatory disease. During the past decade, researchers have made major progress in understanding the three levels of regulation of autophagy in mammalian cells: signaling, autophagosome formation, and autophagosome maturation and lysosomal degradation. As we discuss in this review, each of these levels is potentially druggable, and, depending on the indication, may be able to stimulate or inhibit autophagy. We also summarize the different modulators of autophagy and their potential and limitations in the treatment of life-threatening diseases.

Herbal supplements are generally considered safe; however, drug disposition is influenced by the interactions of herbal supplements and food constituents with transport and metabolic processes. Although the interference of herbal supplements with drug metabolism has been studied extensively, knowledge of how they interact with the drug transport processes is less advanced. Therefore, we describe here specific examples of experimental and human interaction studies of herbal supplement components with drug transporters addressing, for example, organic anion transporting polypeptides or P-glycoprotein, as such interactions may lead to severe side effects and altered drug efficacy. Hence, it is clearly necessary to increase the awareness of the clinical relevance of the interference of herbal supplements with the drug transport processes.