Recent advances on the inhibition of human solute carriers: Therapeutic implications and mechanistic insights

Solute carriers (SLCs) are membrane transport proteins tasked with mediating passage of hydrophilic molecules across lipid bilayers. Despite the extensive roles played in all aspects of human biology, SLCs remain vastly under-explored as therapeutic targets. In this brief review, we first discuss a few successful cases of drugs that exert their mechanisms of action through inhibition of human SLCs, and introduce select examples of human SLCs that have untapped therapeutic potential. We then highlight two recent structural studies which uncovered detailed structural mechanisms of inhibition exhibited against two different human major facilitator superfamily (MFS) transporters of clinical relevance.     

Mechanisms of intrinsic and acquired resistance to kinase‐targeted therapies

Cancer drugs that target pivotal signaling molecules required for malignant cell survival and growth have demonstrated striking antitumor activities in appropriately selected patient populations. Unfortunately, however, therapeutic responses are often of limited duration, typically 6–12 months, because of emergence of drug‐resistant subclones of tumor cells. In this review, we highlight several of the mechanisms of emergent resistance to several kinase‐targeted small molecule therapies used in melanoma, non‐small cell lung cancer (NSCLC) and other solid tumors as illustrative examples. We discuss the implications of these findings for the development of new treatment strategies to delay or prevent the onset of drug resistance.     

Deciphering the molecular mechanisms of anti-tubulin plant derived drugs

Even though commercialized anticancer drugs are now produced by pharmaceutical companies, most of them were originally obtained from natural sources, and more particularly from plants. Indeed, many structurally diverse compounds isolated from plants or marine flora have been purified and synthesized for their anticancer bioactivity. Among these, several molecules belong to the class of anticancer drugs which target the microtubule cytoskeleton, either by stabilizing it or destabilizing it. To characterize the activity of these drugs and to understand in which physiological context they are more likely to be used as therapeutic agents, it is necessary to fully determine their interaction with tubulin. Understanding the molecular basis of their effects on microtubule cytoskeleton is an important step in designing analogs with greater pharmacological activity and with fewer side effects. In addition, knowing the molecular mechanism of action of each drug that is already used in chemotherapy protocols will also help to find strategies to circumvent resistance. By taking examples of known anti-tubulin plant derived drugs, we show how identification of microtubule targeting agents and further characterization of their activity can be achieved combining biophysical and biochemical approaches. We also illustrate how continuing in depth study of molecules with already known primary mechanisms of action can lead to the discovery of new targets or biomarkers which can open new perspectives in anticancer strategies.     

Site-selective reactions of imperfectly matched DNA with small chemical molecules: applications in mutation detection

The last decade has witnessed many exciting scientific publications associated with site-selective reactions of small chemical molecules with imperfectly matched DNA. Typical examples are carbodiimide, hydroxylamine, potassium permanganate, osmium tetroxide, chemical tagging probes, biotinylated, chemiluminescent and fluorescent probes, and all of them selectively react with imperfectly matched DNA. More recently, some therapeutic agents including DNA intercalating drugs and groove binders have been found to promote the in vivo repair system to recognize and repair the mismatch more effectively. The results have established a novel method for detection of mismatches. Development of new chemical reactions for detection of imperfectly matched DNA and mutations is a rapidly growing field and has attracted significant interest of scientists from both chemical and biological fields and it is the main focus of this review.     

The enzyme as drug: application of enzymes as pharmaceuticals

Enzymes as drugs have two important features that distinguish them from all other types of drugs. First, enzymes often bind and act on their targets with great affinity and specificity. Second, enzymes are catalytic and convert multiple target molecules to the desired products. These two features make enzymes specific and potent drugs that can accomplish therapeutic biochemistry in the body that small molecules cannot. These characteristics have resulted in the development of many enzyme drugs for a wide range of disorders.     

Druggable Protein Interaction Sites Are More Predisposed to Surface Pocket Formation than the Rest of the Protein Surface

Despite intense interest and considerable effort via high-throughput screening, there are few examples of small molecules that directly inhibit protein-protein interactions. This suggests that many protein interaction surfaces may not be intrinsically “druggable” by small molecules, and elevates in importance the few successful examples as model systems for improving our fundamental understanding of druggability. Here we describe an approach for exploring protein fluctuations enriched in conformations containing surface pockets suitable for small molecule binding. Starting from a set of seven unbound protein structures, we find that the presence of low-energy pocket-containing conformations is indeed a signature of druggable protein interaction sites and that analogous surface pockets are not formed elsewhere on the protein. We further find that ensembles of conformations generated with this biased approach structurally resemble known inhibitor-bound structures more closely than equivalent ensembles of unbiased conformations. Collectively these results suggest that “druggability” is a property encoded on a protein surface through its propensity to form pockets, and inspire a model in which the crude features of the predisposed pocket(s) restrict the range of complementary ligands; additional smaller conformational changes then respond to details of a particular ligand. We anticipate that the insights described here will prove useful in selecting protein targets for therapeutic intervention.     

What Can We Learn from the Evolution of Protein-Ligand Interactions to Aid the Design of New Therapeutics?

Efforts to increase affinity in the design of new therapeutic molecules have tended to lead to greater lipophilicity, a factor that is generally agreed to be contributing to the low success rate of new drug candidates. Our aim is to provide a structural perspective to the study of lipophilic efficiency and to compare molecular interactions created over evolutionary time with those designed by humans. We show that natural complexes typically engage in more polar contacts than synthetic molecules bound to proteins. The synthetic molecules also have a higher proportion of unmatched heteroatoms at the interface than the natural sets. These observations suggest that there are lessons to be learnt from Nature, which could help us to improve the characteristics of man-made molecules. In particular, it is possible to increase the density of polar contacts without increasing lipophilicity and this is best achieved early in discovery while molecules remain relatively small.     

Yeast "N"-hybrid systems for protein-protein and drug-protein interaction discovery

The majority of small molecule drugs act on protein targets to exert a therapeutic function. It has become apparent in recent years that many small molecule drugs act on more than one particular target and consequently, approaches which profile drugs to uncover their target binding spectrum have become increasingly important. Classical yeast two-hybrid systems have mainly been used to discover and characterize protein-protein interactions, but recent modifications and improvements have opened up new routes towards screening for small molecule-protein interactions. Such yeast "n"-hybrid systems hold great promise for the development of drugs which interfere with protein-protein interactions and for the discovery of drug-target interactions. In this review, we discuss several yeast two-hybrid based approaches with applications in drug discovery and describe a protocol for yeast three-hybrid screening of small molecules to identify their direct targets.     

Recent progress on the development of antibiotics from the genus <Emphasis Type="Italic">Micromonospora</Emphasis>

The emergence of a large number of antimicrobialresistant organisms is a cause for concern. Nature is historically the source of drugs; indeed a considerable number of drugs have been developed from microorganisms, and are now used daily in the treatment of infectious diseases. However, the introduction to the pharmaceutical market of new therapeutic molecules has decreased during the last two decades. In this review, the genus Micromonospora is proposed as a biofactory for production of new antibiotics. The genus Micromonospora has been investigated extensively and more than 100 antibiotics have been isolated from diverse Micromonospora strains. In addition, recent developments in analytical, biological and bioinformatics screening tools used in the discovery of new therapeutic compounds are described. It may be possible to revive formerly used antibiotics produced by Micromonospora and study of this genus may facilitate discovery of novel bioactive molecules.     

Characterizing the role of the dark kinome in neurodegenerative disease – A mini review

BackgroundDrugs that modulate previously unexplored targets could potentially slow or halt the progression of neurodegenerative diseases. Several candidate proteins lie within the dark kinome, those human kinases that have not been well characterized. Much of the kinome (~80%) remains poorly studied, and these targets likely harbor untapped biological potential.Scope of reviewThis review highlights the significance of kinases as mediators of aberrant pathways in neurodegeneration and provides examples of published high-quality small molecules that modulate some of these kinases.Major conclusionsThere is a need for continued efforts to develop high-quality chemical tools to illuminate the function of understudied kinases in the brain. Potent and selective small molecules enable accurate pairing of an observed phenotype with a protein target.General significanceThe examples discussed herein support the premise that validation of therapeutic hypotheses surrounding kinase targets can be accomplished via small molecules and they can serve as the basis for disease-focused drug development campaigns.     

Review physical and chemical aspects of stabilization of compounds in silk

The challenge of stabilization of small molecules and proteins has received considerable interest. The biological activity of small molecules can be lost as a consequence of chemical modifications, while protein activity may be lost due to chemical or structural degradation, such as a change in macromolecular conformation or aggregation. In these cases, stabilization is required to preserve therapeutic and bioactivity efficacy and safety. In addition to use in therapeutic applications, strategies to stabilize small molecules and proteins also have applications in industrial processes, diagnostics, and consumer products like food and cosmetics. Traditionally, therapeutic drug formulation efforts have focused on maintaining stability during product preparation and storage. However, with growing interest in the fields of encapsulation, tissue engineering, and controlled release drug delivery systems, new stabilization challenges are being addressed; the compounds or protein of interest must be stabilized during: (1) fabrication of the protein or small molecule-loaded carrier, (2) device storage, and (3) for the duration of intended release needs in vitro or in vivo. We review common mechanisms of compound degradation for small molecules and proteins during biomaterial preparation (including tissue engineering scaffolds and drug delivery systems), storage, and in vivo implantation. We also review the physical and chemical aspects of polymer-based stabilization approaches, with a particular focus on the stabilizing properties of silk fibroin biomaterials.     

Biomolecular engineering and drug development

Biomolecular engineering is a technology to create novel structures of high-value biomolecules for use in medicine and industry, through the directed alteration of proteins and/or biologically active molecules in living cells to produce a novel biometabolites as well as engineered protein itself. For the development of new drugs by biomolecular engineering, desired biomolecules have to be rationally designed based on their structure-stability/structure-activity relationship, and then screened through well-established mutation and selection program. Over the past decade, there has been significant progress in mutation and selection methodology; DNA shuffling technology mimicking natural evolution for artificial DNA recombination and phage-displayed combinatorial peptide library for rapid selection of proteins expressed from mutated genes. Bioinformatic tools including functional genomics and proteomics have been also developed for the ready access to the information related to the protein-function and genome-protein, leading to the design and identification of new drug targets. Throughout the use of an enormous amount of bioinformatic databases, many protein/peptide drugs and biometabolite molecules have been designed. The candidates of new drugs are monoclonal antibodies, vaccines, enzymes, antibiotics, therapeutic peptides, and so on. Two humanized monoclonal antibodies approved by FDA became the first line of drugs designed by biomolecular engineering approach. They are Herceptin and Synagis, for the treatment of breast cancer and pediatric respiratory syncytial viral infection, respectively. Many more newly engineered biomolecules are under developing for medicinal application. Some clinical trials for therapeutic applications are now in progress, and very positive results are already anticipated.     

Diseases of protein aggregation and the hunt for potential pharmacological agents

Protein aggregation is a ubiquitous phenomenon significant to all aspects of science. Notably, the formation of protein aggregates is frequently encountered in biochemical research and biopharmaceutical industry. Formation of protein aggregates is generally regarded to be associated with partially folded intermediate species that are susceptible to self-association due to the exposure of hydrophobic core. Evidence supports the concept that the formation of aggregates in vitro is a generic property of proteins. In human etiology, more than 20 different devastating human diseases have been reported to be associated with protein aggregation. Although protein aggregation diseases have been the center of intense research, much remains to be learned regarding the underlying molecular mechanisms. In this review, the general background information on protein aggregation is first provided. Next, we summarize the properties, characteristics and causes of protein aggregates. Finally, from the perspectives of epidemiology, pathogenesis, existing mechanisms, relevant hypotheses, and current as well as potential therapeutic approaches, two examples of protein aggregation diseases, Alzheimer's disease and cataract, are briefly discussed. Importantly, while a variety of molecules have been suggested, the effective therapeutic drugs for curing the diseases involving protein aggregation have yet to be identified. We believe that a better understanding of the mechanisms of protein aggregation process and an extensive investigation into the drug penetration, efficacy, and side effects will certainly aid in developing the successful pharmacological agents for these diseases.     

Three's company: two or more unrelated receptors pair with the same ligand

Intercellular communication relies on signal transduction mediated by extracellular ligands and their receptors. Although the ligand-receptor interaction is usually a two-player event, there are selective examples of one polypeptide ligand interacting with more than one phylogenetically unrelated receptor. Likewise, a few receptors interact with more than one polypeptide ligand, and sometimes with more than one coreceptor, likely through an interlocking of unique protein domains. Phylogenetic analyses suggest that for certain triumvirates, the matching events could have taken place at different evolutionary times. In contrast to a few polypeptide ligands interacting with more than one receptor, we found that many small nonpeptide ligands have been paired with two or more plasma membrane receptors, nuclear receptors, or channels. The observation that many small ligands are paired with more than one receptor type highlights the utilitarian use of a limited number of cellular components during metazoan evolution. These conserved ligands are ubiquitous cell metabolites likely favored by natural selection to establish novel regulatory networks. They likely possess structural features useful for designing agonistic and antagonistic drugs to target diverse receptors.     

Structural insights and biomedical potential of IgNAR scaffolds from sharks

In addition to antibodies with the classical composition of heavy and light chains, the adaptive immune repertoire of sharks also includes a heavy-chain only isotype, where antigen binding is mediated exclusively by a small and highly stable domain, referred to as vNAR. In recent years, due to their high affinity and specificity combined with their small size, high physicochemical stability and low-cost of production, vNAR fragments have evolved as promising target-binding scaffolds that can be tailor-made for applications in medicine and biotechnology. This review highlights the structural features of vNAR molecules, addresses aspects of their generation using immunization or in vitro high throughput screening methods and provides examples of therapeutic, diagnostic and other biotechnological applications.     

Structural insights and biomedical potential of IgNAR scaffolds from sharks

In addition to antibodies with the classical composition of heavy and light chains, the adaptive immune repertoire of sharks also includes a heavy-chain only isotype, where antigen binding is mediated exclusively by a small and highly stable domain, referred to as vNAR. In recent years, due to their high affinity and specificity combined with their small size, high physicochemical stability and low-cost of production, vNAR fragments have evolved as promising target-binding scaffolds that can be tailor-made for applications in medicine and biotechnology. This review highlights the structural features of vNAR molecules, addresses aspects of their generation using immunization or in vitro high throughput screening methods and provides examples of therapeutic, diagnostic and other biotechnological applications.     

Green Alternatives in Treatment of Liver Diseases: the Challenges of Traditional Medicine and Green Nanomedicine

Over the last decade, liver diseases have become a global problem, with approximately two million deaths per year. The high increase in the mortality rate of these diseases is mostly related to the limitations in the understanding of the evolutionary clinical cases of liver diseases, the low delivery of drugs in the liver, the non-specific administration of drugs, and the side effects generated at the systemic level by conventional therapeutic agents. Today it is common knowledge that phytochemicals have a high curative potential, even in the prevention and/or reversibility of liver disorders; however, even using these green molecules, researchers continue to deal with the same challenges implemented with conventional therapeutic agents, which limits the pharmacological potential of these friendly molecules. On the other hand, the latest advances in nanotechnology have proven that the use of nanocarriers as a delivery system for green active ingredients, as well as conventional ones, increases the pharmacological potential of these active ingredients due to their physicochemical characteristics (size, Zeta potential, etc.,) moldable depending on the therapeutic objective; in addition to the above, it should be noted that in recent years, nanoparticles have been developed for the specific delivery of drugs towards a specific target (stellar cells, hepatocytes, Kupffer cells), depending on the clinical state of the disease in the patient. The present review addresses the challenges of traditional medicine and green nanomedicine as alternatives in the treatment of liver diseases.     

Electropermeabilization, a physical method for the delivery of therapeutic molecules into cells

Electropermeabilization designates the use of short high-voltage pulses to overcome the barrier of the cell membrane. A position-dependent reversible local membrane permeabilization is induced leading to an exchange of hydrophilic molecules across the membrane. This permeabilized state can be used to load cells with therapeutic molecules. In the case of small molecules, such as anticancer drugs, transfer occurs through simple diffusion. In the case of DNA, transfer occurs through a multi-step mechanism, a process that involves the electrophoretically driven association of the DNA molecule with the destabilised membrane and then its passage.     

Recent Efforts in Identification of Privileged Scaffolds as Antiviral Agents

Viral infections are the most important health concern nowadays to mankind, which is unexpectedly increasing the health complications and fatality rate worldwide. The recent viral infection outbreak developed a pressing need for small molecules that can be quickly deployed for the control/treatment of re-emerging or new emerging viral infections. Numerous viruses, including the human immunodeficiency virus (HIV), hepatitis, influenza, SARS-CoV-1, SARS-CoV-2, and others, are still challenging due to emerging resistance to known drugs. Therefore, there is always a need to search for new antiviral small molecules that can combat viral infection with new modes of action. This review highlighted recent progress in developing new antiviral molecules based on natural product-inspired scaffolds. Herein, the structure-activity relationship of the FDA-approved drugs along with the molecular docking studies of selected compounds have been discussed against several target proteins. The findings of new small molecules as neuraminidase inhibitors, other than known drug scaffolds, Anti-HIV and SARS-CoV are incorporated in this review paper.