Protein–protein interaction modulator drug discovery: past efforts and future opportunities using a rich source of low- and high-throughput screening assays

Introduction: Historically, small-molecule drug discovery projects have largely focused on the G-protein-coupled receptor, ion-channel and enzyme target classes. More recently, there have been successes demonstrating that protein–protein interactions (PPIs) can be targeted by small-molecules and that this strategy has the potential to provide appropriate specificity and selectivity. However, a disadvantage is that compounds that modulate PPIs are often associated with relatively weak affinities as the targeted interaction surfaces are often relatively large. Moreover, from a small-molecule screening perspective, a large proportion of the initial screening Hits are often false positives and these need to be identified and excluded in order to focus on genuine modulators of the PPI being investigated.

Areas covered: The authors review previous efforts on PPI modulator drug discovery. Furthermore, they review assays that can be employed in small-molecule screening and/or Hit validation. The PPI assays are categorized as: i) low-throughput target-based biochemical assays, which are primarily employed for Hit validation at the post-screening stage; ii) high-throughput target-based biochemical assays that are suitable for screening campaigns; and iii) cell-based assays, which are suitable for high-throughput screening campaigns and/or Hit validation.

Expert opinion: Modulating the interaction of PPIs offers the potential to develop novel drugs to treat a wide range of diseases. New assay technologies are continually being developed and it is anticipated that these will be able to be directly used for small-molecule screening campaigns in the future.     

Focusing on shared subpockets – new developments in fragment-based drug discovery

Introduction: Protein–protein interactions (PPIs) are important targets for understanding fundamental biology and for the development of therapeutic agents. Based on different physicochemical properties, numerous pieces of software (e.g., POCKETQUERY, ANCHORQUERY and FTMap) have been reported to find pockets on protein surfaces and have applications in facilitating the design and discovery of small-molecular-weight compounds that bind to these pockets.

Areas covered: The authors discuss a pocket-centric method of analyzing PPI interfaces, which prioritize their pockets for small-molecule drug discovery and the importance of multicomponent reaction chemistry as starting points for undruggable targets. The authors also provide their perspectives on the field.

Expert opinion: Only the tight interplay of efficient computational methods capable of screening a large chemical space and fast synthetic chemistry will lead to progress in the rational design of PPI antagonists in the future. Early drug discovery platforms will also benefit from efficient rapid feedback loops from early clinical research back to molecular design and the medicinal chemistry bench.     

Protein-protein interaction inhibitors: advances in anticancer drug design

ABSTRACT

Introduction: Pocket-based drug design has contributed to major scientific breakthroughs in pharmaceutical research and development (R&D). The integrated use of experimental and computational methods, primarily during the early phases of drug discovery, has enabled the development of highly potent and selective small-molecule ligands. In this scenario, the targeting of protein-protein interactions (PPIs) has emerged as an attractive strategy for designing innovative drugs for highly complex diseases, such as cancer.

Areas covered: This article focuses on the use of experimental and computational approaches with a diversity of PPI classes and discusses the relevant advances in the field, primarily for oncological applications. Analyses of the target binding pockets and medicinal chemistry approaches used to develop promising PPI inhibitors are provided, with an emphasis on data reported over the past 2 years.

Expert opinion: PPI drug discovery is a challenging field that depends completely on accurate structural data. The integration of molecular docking, nuclear magnetic resonance and X-ray crystallography is a cornerstone for the current development of effective PPI inhibitors. Although this field has not reached its peak, several compounds have entered clinical trials over the past few years, providing promising perspectives for novel therapies for highly prevalent and life-threatening conditions.     

Recent advances in protein–protein interaction prediction: experimental and computational methods

Introduction: Proteins within the cell act as part of complex networks, which allow pathways and processes to function. Therefore, understanding how proteins interact is a significant area of current research.

Areas covered: This review aims to present an overview of key experimental techniques (yeast two-hybrid, tandem affinity purification and protein microarrays) used to discover protein–protein interactions (PPIs), as well as to briefly discuss certain computational methods for predicting protein interactions based on gene localization, phylogenetic information, 3D structural modeling or primary protein sequence data. Due to the large-scale applicability of primary sequence-based methods, the authors have chosen to focus on this strategy for our review. There is an emphasis on a recent algorithm called Protein Interaction Prediction Engine (PIPE) that can predict global PPIs. The readers will discover recent advances both in the practical determination of protein interaction and the strategies that are available to attempt to anticipate interactions without the time and costs of experimental work.

Expert opinion: Global PPI maps can help understand the biology of complex diseases and facilitate the identification of novel drug target sites. This study describes different techniques used for PPI prediction that we believe will significantly impact the development of the field in a new future. We expect to see a growing number of similar techniques capable of large-scale PPI predictions.     

Protein–protein interactions: developing small-molecule inhibitors/stabilizers through covalent strategies

The development of small-molecule inhibitors or stabilizers of selected protein–protein interactions (PPIs) of interest holds considerable promise for the development of research tools as well as candidate therapeutics. In this context, the covalent modification of selected residues within the target protein has emerged as a promising mechanism of action to obtain small-molecule modulators of PPIs with appropriate selectivity and duration of action. Different covalent labeling strategies are now available that can potentially allow for a rational, ground-up discovery and optimization of ligands as PPI inhibitors or stabilizers. This review article provides a synopsis of recent developments and applications of such tactics, with a particular focus on site-directed fragment tethering and proximity-enabled approaches.     

Exploration of a 14-3-3 PPI Pocket by Covalent Fragments as Stabilizers

The systematic discovery of functional fragments binding to the composite interface of protein complexes is a first critical step for the development of orthosteric stabilizers of protein–protein interactions (PPIs). We have previously shown that disulfide trapping successfully yielded covalent stabilizers for the PPI of 14-3-3 with the estrogen receptor ERα. Here we provide an assessment of the composite PPI target pocket and the molecular characteristics of various fragments binding to a specific subpocket. Evaluating structure–activity relationships highlights the basic principles for PPI stabilization by these covalent fragments that engage a relatively large and exposed binding pocket at the protein/peptide interface with a “molecular glue” mode of action.     

The relationship among proton pump inhibitors,bone disease and fracture

Introduction: There is growing concern about a possible association between the use of proton pump inhibitors (PPIs) and the development of fragility fractures, most notably hip and vertebral fractures. As PPIs are widely used in clinical practice, this association is of paramount clinical importance.

Areas covered: The authors review the published papers analyzing the relationship between PPI use and the occurrence of fragility fractures. The authors also explore the data supporting possible mechanisms through which PPIs may increase the risk of fracture, including the effects of PPIs on calcium homeostasis, bone mineral density and direct effects of PPIs on bone metabolism.

Expert opinion: Overall, though multiple observational studies have demonstrated an association between PPIs and fragility fractures, the lack of a proven mechanism through which PPIs increase the risk of fracture suggests that this association may not be causal. At this time, the authors do not recommend discontinuing PPIs in patients with a history of fracture or those at increased risk of fracture. However, clinicians should still endeavor to avoid using PPIs in situations where benefits are minimal or clinical indications are lacking.     

Evolving drugs in gastroesophageal reflux disease: pharmacologic treatment beyond proton pump inhibitors

Importance of the field: Despite the clinical success of proton pump inhibitors to treat gastroesophageal reflux disease (GERD), for the majority of patients in both gastroenterology and primary care clinics there is still a substantial group of patients (up to 40% in some studies) who do not completely respond symptomatically to a standard dose of proton pump inhibitors (PPIs). Specific explanations for these PPI noncomplete responders included transient lower esophageal sphincter relaxations (TLESRs), sensitivity to weakly acidic and/or alkaline reflux, large volume of reflux and esophageal hypersensitivity. There is a clear need for GERD therapies beyond the PPIs.

Areas covered in this review: These drug classes include the GABA_B receptor agonists (including lesogaberan and arbaclofen placarbil), mGluR5 receptor antagonists, P-CABs, cholecystokinin₂ antagonists and add-on therapies to PPIs including mosapride and rikkunshito.

What the reader will gain: Both physicians and patients are eagerly awaiting the development and FDA approval of a new class of anti-GERD medications targeting distinct mechanisms. This article provides information on pharmacologic strategies, clinical trials and side-effect profiles on several of the most promising and heavily researched compounds being developed today for the treatment of GERD symptoms and inflammation. Hopefully, this important research will help a large group of PPI noncomplete responding patients receive symptomatic relief and reduce esophageal inflammation through a unique pharmacologic mechanism in the near future.

Take home message: The treatment of GERD has greatly improved with the PPI class of therapy. Despite excellent success, there is a sizeable population of patients who do not have adequate response to therapies directed only at acid suppression. Emerging new pharmacologic treatment options show promise in further advancing the treatment success of GERD.     

Fluorination methods for drug discovery and development

ABSTRACT

Introduction: The pivotal role that the element fluorine plays in modulating the properties of bioactive molecules is reflected by the growth of its presence in approved drugs. In 1970, approximately 2% of drugs contained fluorine with this number rising to 25% by 2011. The synthetic chemistry regarding incorporation of fluorine into organic molecules has also evolved over this time with a paradigm shift from harsh, toxic, hazardous reagents utilized primarily by specialist vendors to new deoxyfluorination reagents and metal-mediated techniques capable of the precise introduction of fluorine into complex organic substrates under relatively mild conditions.

Areas covered: This review highlights the importance of fluorinated compounds in drug discovery, and provides an overview on the synthetic strategies and methodologies developed to access them both in discovery and development.

Expert opinion: The development of new reagents for the safe and precise regioselective fluorination of biologically relevant compounds particularly in drug discovery remains a contemporary challenge in organic chemistry. However, significant strides have been made with the development of new deoxyfluorination reagents and the emergence of practical metal-mediated fluorination techniques have enabled the goal of efficient late-stage fluorination of drug-like compounds to be realized, and the extension of these methods for PET-labelling is being investigated.     

Screening-based approaches to identify small molecules that inhibit protein–protein interactions

Introduction: Protein–protein interactions (PPIs) are very attractive targets for drug development as they play important roles in regulating many aspects of pathophysiologies. It has recently been revealed that the functionally important region of most PPIs is small enough to be modulated by small molecules. Thus, many studies in this field have achieved amazing progress, together with diverse and advanced screening technologies.

Areas covered: This article presents screening technologies to identify small molecule inhibitors of PPIs in addition to discussing the suitability of PPIs as molecular targets. The phases in the processes of selecting compounds are discussed and appropriate steps are proposed, including methodologies to test binding affinity, kinetics, structural analysis, and cellular function.

Expert opinion: Targeting PPIs is still a challenging approach in drug development and relatively few small molecules have reached clinical development. Potential candidates should be assessed and optimized by properly using the multiple assay systems to develop ideal small molecule drugs. Although there remain some barriers to be overcome, small molecule inhibitors of PPIs are fascinating and first-in-class as therapeutic agents to treat various diseases.     

Applied metabolomics in drug discovery

ABSTRACT

Introduction: The metabolic profile is a direct signature of phenotype and biochemical activity following any perturbation. Metabolites are small molecules present in a biological system including natural products as well as drugs and their metabolism by-products depending on the biological system studied. Metabolomics can provide activity information about possible novel drugs and drug scaffolds, indicate interesting targets for drug development and suggest binding partners of compounds. Furthermore, metabolomics can be used for the discovery of novel natural products and in drug development. Metabolomics can enhance the discovery and testing of new drugs and provide insight into the on- and off-target effects of drugs.

Areas covered: This review focuses primarily on the application of metabolomics in the discovery of active drugs from natural products and the analysis of chemical libraries and the computational analysis of metabolic networks.

Expert opinion: Metabolomics methodology, both experimental and analytical is fast developing. At the same time, databases of compounds are ever growing with the inclusion of more molecular and spectral information. An increasing number of systems are being represented by very detailed metabolic network models. Combining these experimental and computational tools with high throughput drug testing and drug discovery techniques can provide new promising compounds and leads.     

Evolving pharmacological approaches in gastroesophageal reflux disease

Introduction: Proton pump inhibitors (PPIs) have considerably improved quality of life in patients with gastroesophageal reflux disease (GERD). However, many patients remain symptomatic despite standard PPI therapy.

Areas covered: This review focuses on evolving therapeutic strategies related to the pathophysiological processes of GERD and insufficient response to PPIs. Several clinical trials evaluated new PPI formulations and newer types of acid-suppressive drugs. These studies have evaluated traditional end points in GERD, but have not shown clinical superiority to current PPIs. Novel therapeutic strategies targeting underlying mechanisms of GERD, such as transient lower esophageal sphincter relaxations (TLESRs) and esophageal hypersensitivity, are being developed for add-on therapy to PPIs. Prokinetic drugs may also have some potential in the add-on treatment of GERD with insufficient response to PPIs. Add-on studies are hampered by insufficient information on optimal patient selection and lack of established end points.

Expert opinion: Newer drugs for symptomatic control in GERD have largely focused on improved acid suppression, without evidence of clinical superiority. Drugs targeting esophageal motility and sensitivity to be used as add-onc therapy in PPI insufficient responders have not reached Phase III trials to date, due to difficulties with patient selection, tolerability and end points.     

Proton pump inhibitors for preventing non-steroidal anti-inflammatory drug induced gastrointestinal toxicity: a systematic review

Objective: Proton pump inhibitors (PPIs) are recommended for preventing gastrointestinal lesions induced by non-steroidal anti-inflammatory drugs (NSAIDs). We performed this study: (1) to evaluate the effectiveness and safety of PPIs, (2) to explore the association between effectiveness and potential influential factors, and (3) to investigate the comparative effect of different PPIs.

Methods: MEDLINE, EMBASE, and the Cochrane Library were searched to identify randomized controlled trials comparing different classes of PPIs, or comparing PPIs with placebo, H₂ receptor antagonists or misoprostol in NSAIDs users. Both pairwise meta-analysis and Bayesian network meta-analysis were performed.

Results: Analyses were based on 12,532 participants from 31 trials. PPIs were significantly more effective than placebo in reducing ulcer complications (relative risk [RR]?=?0.29; 95% confidence interval [CI], 0.20 to 0.42) and endoscopic peptic ulcers (RR?=?0.27; 95% CI, 0.22 to 0.33), with no subgroup differences according to class of NSAIDs, ulcer risk, history of previous ulcer disease, Helicobacter pylori infection, or age. To prevent one ulcer complication, 10 high risk patients and 268 moderate risk patients need PPI therapy. Network meta-analysis indicated that the effectiveness of different PPIs in reducing ulcer complications and endoscopic peptic ulcers is generally similar. PPIs significantly reduced gastrointestinal adverse events and the related withdrawals compared to placebo; there is no difference in safety between different PPIs.

Conclusions: PPIs are effective and safe in preventing peptic ulcers and complications in a wide spectrum of patients requiring NSAID therapy. There is no major difference in the comparative effectiveness and safety between different PPIs.     

Proton pump inhibitors: from CYP2C19 pharmacogenetics to precision medicine

Introduction: Proton Pump inhibitors (PPIs) are commonly used for a variety of acid related disorders. Despite the overall effectiveness and safety profile of PPIs, some patients do not respond adequately or develop treatment related adverse events. This variable response among patients is in part due to genotype variability of CYP2C19, the gene encoding the CYP450 (CYP2C19) isoenzyme responsible for PPIs metabolism.

Areas covered: This article provides an overview of the pharmacokinetics and mechanism of action of the currently available PPIs, including the magnitude of CYPC19 contribution to their metabolism. Additionally, the role of CYP2C19 genetic variability in the therapeutic effectiveness or outcomes of PPI therapy is highlighted in details, to provide supporting evidence for the potential value of CYP2C19 genotype-guided approaches to PPI drug therapy.

Expert opinion: There is a large body of evidence describing the impact of CYP2C19 variability on PPIs and its potential role in individualizing PPI therapy, yet, CYP2C19 pharmacogenetics has not been widely implemented into clinical practice. More data are needed but CYP2C19 genotype-guided dosing of PPIs is likely to become increasingly common and is expected to improve clinical outcomes, and minimize side effects related to PPIs.     

Advances in small-molecule therapy for managing angina pectoris in the elderly

ABSTRACT

Introduction: As our population ages, the prevalence of angina is growing, leading to increased morbidity and decreased quality of life. The management of angina in the elderly is challenging due to drug intolerance and/or drug resistance as well as frailty. Over the past decades, many new therapeutic small molecules have been investigated for the management of angina. Although none of these studies have specifically focused on the therapies for the elderly, they offer promising new avenues for the treatment of angina in the elderly.

Areas covered: Herein, the authors provide a review of the recently published literature on the use of small-molecule therapies for angina management in the elderly and provide a brief overview of these therapies.

Expert opinion: A variety of therapeutic classes of existing and newer small molecules are emerging for the management of angina in the elderly. An individualized approach to the management of angina in this growing population is critical for good outcomes. Many small molecules are in their initial stages of clinical use, and further research should be conducted on their utility, especially in the elderly.     

Tuberous sclerosis preclinical studies: timing of treatment, combination of a rapamycin analog (CCI-779) and interferon-gamma, and comparison of rapamycin to CCI-779

Background

Protein-protein interactions (PPIs) are challenging but attractive targets for small chemical drugs. Whole PPIs, called the 'interactome', have been emerged in several organisms, including human, based on the recent development of high-throughput screening (HTS) technologies. Individual PPIs have been targeted by small drug-like chemicals (SDCs), however, interactome data have not been fully utilized for exploring drug targets due to the lack of comprehensive methodology for utilizing these data. Here we propose an integrative in silico approach for discovering candidates for drug-targetable PPIs in interactome data.

Results

Our novel in silico screening system comprises three independent assessment procedures: i) detection of protein domains responsible for PPIs, ii) finding SDC-binding pockets on protein surfaces, and iii) evaluating similarities in the assignment of Gene Ontology (GO) terms between specific partner proteins. We discovered six candidates for drug-targetable PPIs by applying our in silico approach to original human PPI data composed of 770 binary interactions produced by our HTS yeast two-hybrid (HTS-Y2H) assays. Among them, we further examined two candidates, RXRA/NRIP1 and CDK2/CDKN1A, with respect to their biological roles, PPI network around each candidate, and tertiary structures of the interacting domains.

Conclusion

An integrative in silico approach for discovering candidates for drug-targetable PPIs was applied to original human PPIs data. The system excludes false positive interactions and selects reliable PPIs as drug targets. Its effectiveness was demonstrated by the discovery of the six promising candidate target PPIs. Inhibition or stabilization of the two interactions may have potential therapeutic effects against human diseases.     

Indications and safety of proton pump inhibitor drug use in patients with cancer

Introduction: Although the exact prevalence of proton pump inhibitor (PPI) use in cancer patients is not known, it is generally perceived to be widespread. PPIs are generally well tolerated and carry an excellent safety profile. However, increasing and longer term PPI use has raised concerns about the risk of pneumonia, bone fractures and enteric infections, and a possible interaction with clopidogrel that could increase the risk of cardiovascular events.

Areas covered: We conducted a PubMed search of English language articles addressing the safety and adverse events associated with PPI use with particular emphasis in cancer patients.

Expert opinion: PPIs, frequently used in cancer patients, are generally well tolerated and carry an excellent safety profile. PPI-induced acid suppression may increase the risk of Clostridium difficile or other enteric infections, nutritional deficiencies and community acquired pneumonia, all particularly important in cancer patients. The indications for PPI use in cancer patients should be carefully reviewed prior to use.     

An overview of aldehyde oxidase: an enzyme of emerging importance in novel drug discovery

Introduction: Given the rising trend in medicinal chemistry strategy to reduce cytochrome P450-dependent metabolism, aldehyde oxidase (AOX) has recently gained increased attention in drug discovery programs and the number of drug candidates that are metabolized by AOX is steadily growing.

Areas covered: Despite the emerging importance of AOX in drug discovery, there are certain major recognized problems associated with AOX-mediated metabolism of drugs. Intra- and inter-species variations in AOX activity, the lack of reliable and predictive animal models using the common experimental animals, and failure in the predictions of in vivo metabolic activity of AOX using traditional in vitro methods are among these issues that are covered in this article. A comprehensive review of computational human AOX (hAOX) related studies are also provided.

Expert opinion: Following the recent progress in the stem cell field, the authors recommend the application of organoids technology as an effective tool to solve the fundamental problems associated with the evaluation of AOX in drug discovery. The recent success in resolving the hAOX crystal structure can too be another valuable data source for the study of AOX-catalyzed metabolism of new drug candidates, using computer-aided drug discovery methods.