Current Medicinal Chemistry (v.18, #27)

Aptamers, typically selected through systematic evolution of ligands by exponential enrichment (SELEX), have quickly emerged as aversatile class of agents with tremendous potential for a wide range of biomedical applications. Often regarded as chemical antibodies,aptamers can fold into well-defined 3D structures and bind to their target molecules with high affinity and specificity. To date, aptamers havebeen selected against a wide variety of targets such as proteins, phospholipids, sugars, nucleic acids, metal ions, dyes, whole cells, amongmany others.During the last two decades since aptamers were first selected through SELEX in 1990, an aptamer that binds to human vascular endothelialgrowth factor (VEGF) has been approved by the United States Food and Drug Administration for clinical use in treating age-relatedmacular degeneration (AMD). A number of aptamers against other molecular targets are currently in clinical investigation. To provide a centralizedresource for scientists who are either new to or working in the area of aptamer-based research, I have organized this special issue ofCurrent Medicinal Chemistry. An international ensemble of experts in the field was invited to write a total of twelve review articles, focusingon the selection, modification, and biomedical application of aptamers.In the first review article, Dr. Li and co-workers introduced the identification, modification, and working mechanism of aptamers againstcell-surface receptors. Next, Dr. Tan and co-workers reviewed the various methods of SELEX-based selection of aptamers, such as protein-SELEX and cell-SELEX. The major disadvantage of aptamers is that they are typically not stable enough for biomedical applications. Notonly are DNA/RNA aptamers susceptible to nuclease degradation, peptide aptamers can also be digested by proteases. In this regard, Dr. Caiand co-workers gave a comprehensive summary of different chemical modifications that have been employed to increase the stability ofDNA, RNA, and peptide aptamers.The following two review articles were focused on proteases and protein kinases, respectively. Two world-renowned experts in the field,Dr. Andreasen and Dr. de Franciscis, each gave a superb overview on their specific topic. Next, Dr. Zhang and co-workers provided a summaryof the applications of aptamers in nervous system disorders, while Dr. Fang and co-workers focused on aptamers selected against moleculartargets that are involved in cardiovascular diseases.Since aptamers can bind to target molecules with high affinity and specificity, they are excellent candidates for recognition elements inbiosensors. Dr. Gao and co-workers contributed a comprehensive review on the development of aptamer-based fluorescent biosensors, withemphasis on the design as well as properties such as sensitivity and specificity. The same properties that make aptamers suitable for biosensingalso make them amenable for tumor-targeted drug delivery and molecular imaging applications, which are the focuses of the two followingreview articles.Over the last two decades since its initial discovery, big strides have been made in aptamer-based research. Dr. Lupold and co-workersdiscussed the current status of aptamers in clinical trials, as well as some promising aptamers in pre-clinical development. Another class ofaptamers besides the traditional DNA/RNA aptamers is peptide aptamers, which are combinatorial protein molecules composed of a shortpeptide region inserted within a scaffold protein. The short peptide region is responsible for target binding, while the scaffold protein helps toenhance the binding affinity and specificity through conformational restriction. In the last review article, Dr. Zhang and co-workers gave athorough review on the structure, selection, and application of peptide aptamers in biological studies as well as in therapeutics.The future of aptamer-based research is becoming increasingly bright and we are starting to reap the fruits of two decades of aptamerbasedresearch. However, significant challenges lie ahead before more aptamer-based agents can reach clinical trials and eventually the dayto-day management of patients, as detailed in many of these review articles. I am truly grateful to all authors for their tremendous effort inthis special issue, which I am confident will help moving the field forward.

Aptamers are synthetic oligonucleotides selected from pools of random-sequence oligonucleotides which bind to a wide rangeof biomolecular targets with high affinity and specificity. Compared with antibodies, aptamers exhibit significant advantages includingsmall size, easy synthesis and modification, as well as low immunogenicity. Many of the aptamers also show inhibition of their targets,making them potential therapeutic and targeting reagents in clinical applications. Compared with aptamers against intracellular proteinsand molecules, however, the identification of aptamers against cell-surface receptors and receptor-related antigens is more difficult, dueto the complex cellular environment in which receptors are located, and also the unique conformations and compositions of receptors tokeep their activity. In this review, we will introduce the identification, modification and working mechanism of aptamers against cellsurfacereceptors. Based on the different characteristics of target receptors and selection strategies used, the identified aptamers show distinctbinding affinity with recombinant targets or specific cell lines which express receptors on the surface in vitro. Some of the in vivoexperiments also indicate that aptamers have the capability of inhibiting the overexpressing receptor-related tumor growth, working aspotential anti-tumor therapeutic drugs. Despite of the difficulties during the selection of receptor aptamers and the study of their workingmechanism during the present time, it is possible that in the future aptamers will increasingly exhibit therapeutic and diagnostic utility.

Recent Developments in Protein and Cell-Targeted Aptamer Selection and Applications by Jun Liu, Mingxu You, Ying Pu, Huixia Liu, Mao Ye, Weihong Tan (4117-4125).
Because of their easily modified chemical structures and wide range of targets, aptamers are ideal candidates for various applications,such as biomarker discovery, target diagnosis, molecular imaging, and drug delivery. Aptamers are oligonucleotide sequencesthat can bind to their targets specifically via unique three dimensional (3-D) structures. Usually, aptamers are obtained from repeatedrounds of in vitro or in vivo selection termed SELEX (Systematic Evolution of Ligands by EXponential enrichment), which can generateaptamers with high affinity and specificity for many kinds of targets, such as biomedically important proteins and even cancer cells. Inthis review, some basic principles and recent developments in the design of SELEX process are discussed, hopefully to provide someguidelines towards performing more efficient aptamer isolation procedures. Moreover, the biomedical and bioanalytical applications ofaptamers are further reviewed, based on some smart biochemical modifications of these oligonucleotide structures.

Improving the Stability of Aptamers by Chemical Modification by R.E. Wang, H. Wu, Y. Niu, J. Cai (4126-4138).
Ever since the invention of SELEX (systematic evolution of ligands by exponential enrichment), there has been rapid developmentfor aptamers over the last two decades, making them a promising approach in therapeutic applications as either drug candidatesor diagnostic tools. For therapeutic purposes, a durable performance of aptamers in biofluids is required, which is, however, hampered bythe lack of stability of most aptamers. Not only are the nucleic acid aptamers susceptible to nucleases, the peptide aptamers are also subjectiveto degradation by proteases. With the advancement of chemical biology, numerous attempts have been made to overcome this obstacle,many resulting in significant improvements in stability. In this review, chemical modifications to increase the stability of threemain types of aptamers, DNA, RNA and peptide are comprehensively summarized. For nucleic acid aptamers, development of modifiedSELEX coupled with mutated polymerase is discussed, which is adaptive to a number of modifications in aptamers and in a large extentfacilitates the research of aptamer-modifications. For peptide aptamers, approaches in molecular biology with introduction of stabilizingprotein as well as the switch of scaffold protein are included, which may represent a future direction of chemical conjugations toaptamers.

Nucleic Acid Aptamers Against Proteases by D.M. Dupont, L.M. Andersen, K.A. Botkjaer, P.A. Andreasen (4139-4151).
Proteases are potential or realized therapeutic targets in a wide variety of pathological conditions. Moreover, proteases areclassical subjects for studies of enzymatic and regulatory mechanisms. We here review the literature on nucleic acid aptamers selectedwith proteases as targets. Designing small molecule protease inhibitors of sufficient specificity has proved a daunting task. Aptamersseem to represent a promising alternative. In our review, we concentrate on biochemical mechanisms of aptamer selection, proteinaptamerrecognition, protease inhibition, and advantages of aptamers for pharmacological intervention with pathophysiological functionsof proteases. Aptamers can be selected so that they bind their targets highly specifically and with affinities corresponding to KD values inthe nM range. Aptamers can be selected so that they recognize their targets conformation-specifically, for instance with vastly differentaffinities to zymogen and active enzyme forms. Furthermore, aptamers can be selected to inhibit the enzyme activity of the target proteases,but also to inhibit functionally important exosite interactions, for instance cofactor binding. Several protease-inhibiting aptamers, directedagainst blood coagulation factors, are in clinical trials as anticoagulant drugs. Several of the studies on protease-binding aptamershave been pioneering and trend-setting in the field. The work with protease-binding aptamers also demonstrates many interesting examplesof non-standard selection strategies and of new principles for regulating the activity of the inhibitory action of aptamers of generalinterest to researchers working with nucleic acid aptamers.

Nucleic Acid Aptamers Against Protein Kinases by L. Cerchia, V. de Franciscis (4152-4158).
Deregulation of kinase function has been implicated in several important diseases, including cancer, neurological and metabolicdisorders. Because of their key role in causing disease, kinases have become one of the most intensively pursued classes of drugtargets. To date, several monoclonal antibodies (mAbs) and small-molecule inhibitors have been approved for the treatment of cancer.Aptamers are short structured single stranded RNA or DNA ligands that bind at high affinity to their target molecules and are nowemerging as promising molecules to target specific cancer epitopes in clinical diagnosis and therapy. Further, because of their high specificityand low toxicity aptamers will likely reveal among the most promising molecules for in vivo targeted recognition as therapeutics ordelivery agents for nanoparticles, small interfering RNAs bioconjugates, chemotherapeutic cargos and molecular imaging probes.In this article, we discuss recent advances in the development of aptamers targeting kinase proteins.

Aptamers: Selection, Modification and Application to Nervous System Diseases by Y. Yang, X. Ren, H.J. Schluesener, Z. Zhang (4159-4168).
Aptamers are nonnaturally occurring oligonucleotides generated by the SELEX (Systematic Evolution of Ligands by Exponentialenrichment) process. Due to their unique three-dimensional structures, aptamers can bind to various targets, ranging from smallcompounds to cells and tissues, with high affinity and specificity. While first reported in 1990, aptamers have become useful tools in thebiomedical field because of their unique characteristics, such as easy and quick preparation, cost-effectiveness, small size, versatility, etal. Recently various chemical modifications have been introduced to enhance aptamers stability in the body fluids and their bioavailabilityin animals, which have pushed aptamer closer to therapeutic and diagnostic application. This review provides an overview of the aptamermodifications and their application in the nervous system disorders.

Aptamers as Therapeutics in Cardiovascular Diseases by P. Wang, Y. Yang, H. Hong, Y. Zhang, W. Cai, D. Fang (4169-4174).
With many advantages over other therapeutic agents such as monoclonal antibodies, aptamers have recently emerged as anovel and powerful class of ligands with excellent potential for diagnostic and therapeutic applications. Typically generated through SystematicEvolution of Ligands by EXponential enrichment (SELEX), aptamers have been selected against a wide range of targets such asproteins, phospholipids, sugars, nucleic acids, as well as whole cells. DNA/RNA aptamers are single-stranded DNA/RNA oligonucleotides(with a molecular weight of 5-40 kDa) that can fold into well-defined 3D structures and bind to their target molecules with high affinityand specificity. A number of strategies have been adopted to synthesize aptamers with enhanced in vitro/in vivo stability, aiming atpotential therapeutic/diagnostic applications in the clinic. In cardiovascular diseases, aptamers can be developed into therapeutic agentsas anti-thrombotics, anti-coagulants, among others. This review focuses on aptamers that were selected against various molecular targetsinvolved in cardiovascular diseases: von Willebrand factor (vWF), thrombin, factor IX, phospholamban, P-selectin, platelet-derivedgrowth factor, integrin ..v..3, CXCL10, vasopressin, among others. With continued effort in the development of aptamer-based therapeutics,aptamers will find their niches in cardiovascular diseases and significantly impact clinical patient management.

Aptamer-Based Fluorescent Biosensors by R.E. Wang, Y. Zhang, J. Cai, W. Cai, T. Gao (4175-4184).
Selected from random pools of DNA or RNA molecules through systematic evolution of ligands by exponential enrichment(SELEX), aptamers can bind to target molecules with high affinity and specificity, which makes them ideal recognition elements in thedevelopment of biosensors. To date, aptamer-based biosensors have used a wide variety of detection techniques, which are briefly summarizedin this article. The focus of this review is on the development of aptamer-based fluorescent biosensors, with emphasis on theirdesign as well as properties such as sensitivity and specificity. These biosensors can be broadly divided into two categories: those usingfluorescently-labeled aptamers and others that employ label-free aptamers. Within each category, they can be further divided into signalonand signal-off sensors. A number of these aptamer-based fluorescent biosensors have shown promising results in biological samplessuch as urine and serum, suggesting their potential applications in biomedical research and disease diagnostics.

Tumor-Targeted Drug Delivery with Aptamers by Y. Zhang, H. Hong, W. Cai (4185-4194).
Cancer is one of the leading causes of death around the world. Tumor-targeted drug delivery is one of the major areas in cancerresearch. Aptamers exhibit many desirable properties for tumor-targeted drug delivery, such as ease of selection and synthesis, highbinding affinity and specificity, low immunogenicity, and versatile synthetic accessibility. Over the last several years, aptamers havequickly become a new class of targeting ligands for drug delivery applications. In this review, we will discuss in detail about aptamerbaseddelivery of chemotherapy drugs (e.g. doxorubicin, docetaxel, daunorubicin, and cisplatin), toxins (e.g. gelonin and various photodynamictherapy agents), and a variety of small interfering RNAs. Although the results are promising which warrants enthusiasm foraptamer-based drug delivery, tumor homing of aptamer-based conjugates after systemic injection has only been achieved in one report.Much remains to be done before aptamer-based drug delivery can reach clinical trials and eventually the day-to-day management of cancerpatients. Therefore, future directions and challenges in aptamer-based drug delivery are also discussed.

Molecular Imaging with Nucleic Acid Aptamers by H. Hong, S. Goel, Y. Zhang, W. Cai (4195-4205).
With many desirable properties such as ease of synthesis, small size, lack of immunogenicity, and versatile chemistry, aptamersrepresent a class of targeting ligands that possess tremendous potential in molecular imaging applications. Non-invasive imaging ofvarious disease markers with aptamer-based probes has many potential clinical applications such as lesion detection, patient stratification,treatment monitoring, etc. In this review, we will summarize the current status of molecular imaging with aptamer-based probes. First,fluorescence imaging will be described which include both direct targeting and activatable probes. Next, we discuss molecular magneticresonance imaging and targeted ultrasound investigations using aptamer-based agents. Radionuclide-based imaging techniques (singlephotonemission computed tomography and positron emission tomography) will be summarized as well. In addition, aptamers have alsobeen labeled with various tags for computed tomography, surface plasmon resonance, dark-field light scattering microscopy, transmissionelectron microscopy, and surface-enhanced Raman spectroscopy imaging. Among all molecular imaging modalities, no single modality isperfect and sufficient to obtain all the necessary information for a particular question. Thus, a multimodality probe has also been constructedfor concurrent fluorescence, gamma camera, and magnetic resonance imaging in vivo. Although the future of aptamer-based molecularimaging is becoming increasingly bright and many proof-of-principle studies have already been reported, much future effort needsto be directed towards the development of clinically translatable aptamer-based imaging agents which will eventually benefit patients.

Nucleic Acid Aptamers: Clinical Applications and Promising New Horizons by X. Ni, M. Castanares, A. Mukherjee, S.E. Lupold (4206-4214).
Aptamers are a special class of nucleic acid molecules that are beginning to be investigated for clinical use. These smallRNA/DNA molecules can form secondary and tertiary structures capable of specifically binding proteins or other cellular targets; theyare essentially a chemical equivalent of antibodies. Aptamers have the advantage of being highly specific, relatively small in size, andnon-immunogenic. Since the discovery of aptamers in the early 1990s, great efforts have been made to make them clinically relevant fordiseases like cancer, HIV, and macular degeneration. In the last two decades, many aptamers have been clinically developed as inhibitorsfor targets such as vascular endothelial growth factor (VEGF) and thrombin. The first aptamer based therapeutic was FDA approved in2004 for the treatment of age-related macular degeneration and several other aptamers are currently being evaluated in clinical trials.With advances in targeted-therapy, imaging, and nanotechnology, aptamers are readily considered as potential targeting ligands becauseof their chemical synthesis and ease of modification for conjugation. Preclinical studies using aptamer-siRNA chimeras and aptamer targetednanoparticle therapeutics have been very successful in mouse models of cancer and HIV. In summary aptamers are in several stagesof development, from pre-clinical studies to clinical trials and even as FDA approved therapeutics. In this review, we will discuss the currentstate of aptamers in clinical trials as well as some promising aptamers in pre-clinical development.

Peptide Aptamers with Biological and Therapeutic Applications by J. Li, S. Tan, X. Chen, C.-Y. Zhang, Y. Zhang (4215-4222).
Peptide aptamers are combinatorial protein molecules with specific bind affinity to given target proteins under intracellularconditions. The typical structure of peptide aptamers is a short peptide region inserted within a scaffold protein. The short peptide regionis responsible for binding with its target protein and the scaffold protein helps to enhance the binding affinity and specificity through restrictionon the conformation of the binding peptide. This unique structural feature allows peptide aptamers to bind with their target proteinswith strong affinity and high specificity. Applications of peptide aptamers thus vary from in vitro detection of various proteins in acomplex mixture to in vivo modulation on proteins and cell functions. Peptide aptamers have also been considered as therapeutic moleculesbecause of their anticancer and antivirus activity. Due to the importance of peptide aptamers, a general review on the structure, selectionand applications of peptide aptamers in biological study as well as in therapeutics will be presented in this paper.

Current Trends in β-Lactam Based β-Lactamases Inhibitors by S. Biondi, S. Long, M. Panunzio, W.L. Qin (4223-4236).
The introduction of antibiotics to treat bacterial infections either by killing or blocking their growth has been accompanied bythe development of resistance mechanism that allows the bacteria to survive and proliferate. In particular the successive series of β-lactams have selected several generations of β-lactamases including ESBLs, AmpC β-lactamases, KPC carbapenamases in Enterobacteriaceae,the metallo β-lactamases VIMs and IMPs, and very recently the threatening NDM-1 that confers resistance to virtually any clinicallyused antibiotic. The increasing use of carbapenems due to the spread of resistance to other existing antibacterial agents has facilitatedthe spread of resistance, especially in Acinetobacter spp. due to OXA- and metallo-carbapenemases. The pharmaceutical industry,that abandoned this field at the end of the nineties, is now trying to recover by developing some novel β-lactam antibiotics and novel β-lactamase-inhibitors, the latter to be used in combination with new as well as seasoned β-lactam antibiotics. This article provides a surveyof patent and scientific literature for β-lactamase inhibitors discovered in the period 2006-2010.

Novel Insights into Targeting ATP-Binding Cassette Transporters for Antitumor Therapy by L. Gatti, G. Cossa, G.L. Beretta, N. Zaffaroni, P. Perego (4237-4249).
ATP-binding cassette (ABC) transporters are a large family of proteins implicated in physiological cellular functions. Selectedcomponents of the family play a well-recognized role in extruding conventional cytotoxic antitumor agents and molecularly targeteddrugs from cells. Some lines of evidence also suggest links between transporters and tumor cell survival, in part unrelated to efflux.However, the study of the precise mechanisms regulating the function of drug transporters (e.g., posttranslational modifications such asglycosylation) is still in its infancy. A better definition of the molecular events clarifying the regulation of transporter levels includingregulation by microRNAs may contribute to provide new molecular tools to target such a family of transporters. The present review focuseson the biological aspects that implicate ABC transporters in resistance of tumor cells, including cancer stem cells. Molecular analysisof well-known preclinical systems as well as of cancer stem cell models supports the notion that ABC transporters represent amenabletargets for modulation of the efficacy of antitumor agents endowed with different molecular features. Recent achievements regarding tumorcell biology are expected to provide a rationale for developing novel inhibitors that target ABC transporters implicated in drug resistance.

Recent progresses in cancer therapy suggest the importance of targeting more than one protein targets or signaling pathways.In events of stresses including the therapeutic treatments, damaged proteins are either repaired by heat shock proteins or ubiquitin-taggedfor proteasome-dependent protein degradation. Heat shock proteins mediated protein protection and cell signaling, as well as the ubiquitin-proteasomal degradation are thus central to cellular homeostasis, and are reported to play substantial roles in tumor cells’ rapidmetabolismand stimuli-resistance. The up-regulated heat shock protein 90 (HSP90), heat shock protein 70 (HSP70) and 26S proteasomein cancer cells have been thereby recognized as important drug targets and are under intensive studies in recent years. While most researchfocuses on each target in a separate manner, simultaneous inhibition of more than one target results in an enhanced efficacy, especiallyin single-drug-resistant cancer cell line. In this review, current development of chemical inhibitors for these three core targets issummarized respectively and the progress on related simultaneous inhibitions has been discussed. In a perspective view, combined inhibitionsof HSP 90/70 and the 26S proteasome could be a promising approach in cancer therapy and may suggest a future direction fordrug-screening.