Paclitaxel (PTX) is among the most commonly used first-line drugs for cancer chemotherapy. However, its poor water solubility and indiscriminate distribution in normal tissues remain clinical ...challenges. Here we design and synthesize a highly water-soluble nucleolin aptamer-paclitaxel conjugate (NucA-PTX) that selectively delivers PTX to the tumor site. By connecting a tumor-targeting nucleolin aptamer (NucA) to the active hydroxyl group at 2' position of PTX via a cathepsin B sensitive dipeptide bond, NucA-PTX remains stable and inactive in the circulation. NucA facilitates the uptake of the conjugated PTX specifically in tumor cells. Once inside cells, the dipeptide bond linker of NucA-PTX is cleaved by cathepsin B and then the conjugated PTX is released for action. The NucA modification assists the selective accumulation of the conjugated PTX in ovarian tumor tissue rather than normal tissues, and subsequently resulting in notably improved antitumor activity and reduced toxicity.
Aptamers are oligonucleotide sequences with a length of about 25–80 bases which have abilities to bind to specific target molecules that rival those of monoclonal antibodies. They are attracting ...great attention in diverse clinical translations on account of their various advantages, including prolonged storage life, little batch-to-batch differences, very low immunogenicity, and feasibility of chemical modifications for enhancing stability, prolonging the half-life in serum, and targeted delivery. In this Review, we demonstrate the emerging aptamer discovery technologies in developing advanced techniques for producing aptamers with high performance consistently and efficiently as well as requiring less cost and resources but offering a great chance of success. Further, the diverse modifications of aptamers for therapeutic applications including therapeutic agents, aptamer–drug conjugates, and targeted delivery materials are comprehensively summarized.
Aptamers are short single-stranded DNA, RNA, or synthetic Xeno nucleic acids (XNA) molecules that can interact with corresponding targets with high affinity. Owing to their unique features, including ...low cost of production, easy chemical modification, high thermal stability, reproducibility, as well as low levels of immunogenicity and toxicity, aptamers can be used as an alternative to antibodies in diagnostics and therapeutics. Systematic evolution of ligands by exponential enrichment (SELEX), an experimental approach for aptamer screening, allows the selection and identification of in vitro aptamers with high affinity and specificity. However, the SELEX process is time consuming and characterization of the representative aptamer candidates from SELEX is rather laborious. Artificial intelligence (AI) could help to rapidly identify the potential aptamer candidates from a vast number of sequences. This review discusses the advancements of AI pipelines/methods, including structure-based and machine/deep learning-based methods, for predicting the binding ability of aptamers to targets. Structure-based methods are the most used in computer-aided drug design. For this part, we review the secondary and tertiary structure prediction methods for aptamers, molecular docking, as well as molecular dynamic simulation methods for aptamer-target binding. We also performed analysis to compare the accuracy of different secondary and tertiary structure prediction methods for aptamers. On the other hand, advanced machine-/deep-learning models have witnessed successes in predicting the binding abilities between targets and ligands in drug discovery and thus potentially offer a robust and accurate approach to predict the binding between aptamers and targets. The research utilizing machine-/deep-learning techniques for prediction of aptamer-target binding is limited currently. Therefore, perspectives for models, algorithms, and implementation strategies of machine/deep learning-based methods are discussed. This review could facilitate the development and application of high-throughput and less laborious in silico methods in aptamer selection and characterization.
Background
Skeletal muscle atrophy induced by either aging (sarcopenia) or mechanical unloading is associated with serious health consequences. Long non‐coding RNAs (lncRNAs) are implicated as ...important regulators in numerous physiological and pathological processes.
Methods
Microarray analysis was performed to identify the differentially expressed lncRNAs in skeletal muscle between adult and aged mice. The most decreased lncRNA in aged skeletal muscle was identified. The C2C12 mouse myoblast cells were used to assess the biological function of the lncRNA in vitro. The target microRNA of lncRNA and the target protein of microRNA were predicted by bioinformatics analysis and validated in vitro. Furthermore, the biology function of the lncRNA in vivo was investigated by local overexpression or knockdown the lncRNA in skeletal muscle. The therapeutic effect of the lncRNA overexpression in age‐related or mechanical unloading‐induced muscle atrophy was also evaluated.
Results
We identified a novel lncRNA (muscle anabolic regulator 1, MAR1) which was highly expressed in mice skeletal muscle and positively correlated with muscle differentiation and growth in vitro and in vivo. We predicted and validated that microRNA‐487b (miR‐487b) was a direct target of MAR1. We also predicted and validated that Wnt5a, an important regulator during myogenesis, was a target of miR‐487b in C2C12 cells. Our findings further demonstrated that enforced MAR1 expression in myoblasts led to derepression of Wnt5a. Moreover, MAR1 promoted skeletal muscle mass/strength and Wnt5a protein level in mice. Enforced MAR1 expression in mice attenuated muscle atrophy induced by either aging or unloading.
Conclusions
The newly identified lncRNA MAR1 acts as a miR‐487b sponge to regulate Wnt5a protein, resulting in promoting muscle differentiation and regeneration. MAR1 could be a novel therapeutic target for treating muscle atrophy induced by either aging or mechanical unloading.
Abstract
Sclerostin negatively regulates bone formation by antagonizing Wnt signalling. An antibody targeting sclerostin for the treatment of postmenopausal osteoporosis was approved by the U.S. Food ...and Drug Administration, with a boxed warning for cardiovascular risk. Here we demonstrate that sclerostin participates in protecting cardiovascular system and inhibiting bone formation via different loops. Loop3 deficiency by genetic truncation could maintain sclerostin’s protective effect on the cardiovascular system while attenuating its inhibitory effect on bone formation. We identify an aptamer, named aptscl56, which specifically targets sclerostin loop3 and use a modified aptscl56 version, called Apc001PE, as specific in vivo pharmacologic tool to validate the above effect of loop3. Apc001PE has no effect on aortic aneurysm and atherosclerotic development in
ApoE
−/−
mice and
hSOST
ki
.ApoE
−/−
mice with angiotensin II infusion. Apc001PE can promote bone formation in
hSOST
ki
mice and ovariectomy-induced osteoporotic rats. In summary, sclerostin loop3 cannot participate in protecting the cardiovascular system, but participates in inhibiting bone formation.
Aptamers are promising therapeutic and diagnostic agents for various diseases due to their high affinity and specificity against target proteins. Structural determination in combination with multiple ...biochemical and biophysical methods could help to explore the interacting mechanism between aptamers and their targets. Regrettably, structural studies for aptamer-target interactions are still the bottleneck in this field, which are facing various difficulties. In this review, we first reviewed the methods for resolving structures of aptamer-protein complexes and for analyzing the interactions between aptamers and target proteins. We summarized the general features of the interacting nucleotides and residues involved in the interactions between aptamers and proteins. Challenges and perspectives in current methodologies were discussed. Approaches for determining the binding affinity between aptamers and target proteins as well as modification strategies for stabilizing the binding affinity of aptamers to target proteins were also reviewed. The review could help to understand how aptamers interact with their targets and how alterations such as chemical modifications in the structures affect the affinity and function of aptamers, which could facilitate the optimization and translation of aptamers-based theranostics.
Several virtual screening models are proposed to screen small molecules only targeting primary miRNAs without selectivity. Few attempts have been made to develop virtual screening strategies for ...discovering small molecules targeting mature miRNAs. Mature miRNAs and their specific target mRNA can form unique functional loops during argonaute (AGO)‐mediated miRNA–mRNA interactions, which may serve as potential targets for small‐molecule drug discovery. Thus, a loop‐based and AGO‐incorporated virtual screening model is constructed for targeting the loops. The previously published studies have found that miR‐214 can target ATF4 to inhibit osteoblastic bone formation, whereas miR‐214 can target TRAF3 to promote osteoclast activity. By using the virtual model, the top ten candidate small molecules targeting miR‐214‐ATF4 mRNA interactions and top ten candidate small molecules targeting miR‐214‐TRAF3 mRNA interactions are selected, respectively. Based on both in vitro and in vivo data, one small molecule can target miR‐214‐ATF4 mRNA to promote ATF4 protein expression and enhance osteogenic potential, whereas one small molecule can target miR‐214‐TRAF3 mRNA to promote TRAF3 protein expression and inhibit osteoclast activity. These data indicate that the loop‐based and AGO‐incorporated virtual screening model can help to obtain small molecules specifically targeting miRNA–mRNA interactions to rescue bone phenotype in genetically modified mice.
It is found that mature miRNA and their specific target mRNA can form unique functional loops during argonaute (AGO)‐mediated miRNA–mRNA interactions. Thus, a loop‐based and AGO‐incorporated virtual screening model is constructed for screening small molecules targeting the loops. By using the virtual model, two small molecules are obtained that can specifically target miR‐214‐ATF4 mRNA interactions and miR‐214‐TRAF3 mRNA interactions, respectively.
Emerging evidence indicates that osteoclasts direct osteoblastic bone formation. MicroRNAs (miRNAs) have a crucial role in regulating osteoclast and osteoblast function. However, whether miRNAs ...mediate osteoclast-directed osteoblastic bone formation is mostly unknown. Here, we show that increased osteoclastic miR-214-3p associates with both elevated serum exosomal miR-214-3p and reduced bone formation in elderly women with fractures and in ovariectomized (OVX) mice. Osteoclast-specific miR-214-3p knock-in mice have elevated serum exosomal miR-214-3p and reduced bone formation that is rescued by osteoclast-targeted antagomir-214-3p treatment. We further demonstrate that osteoclast-derived exosomal miR-214-3p is transferred to osteoblasts to inhibit osteoblast activity in vitro and reduce bone formation in vivo. Moreover, osteoclast-targeted miR-214-3p inhibition promotes bone formation in ageing OVX mice. Collectively, our results suggest that osteoclast-derived exosomal miR-214-3p transfers to osteoblasts to inhibit bone formation. Inhibition of miR-214-3p in osteoclasts may be a strategy for treating skeletal disorders involving a reduction in bone formation.
Rheumatoid arthritis (RA) is a chronic inflammatory disorder characterized by progressive bone erosion. Leflunomide is originally developed to suppress inflammation via its metabolite A77 1726 to ...attenuate bone erosion. However, distinctive responsiveness to Leflunomide is observed among RA individuals. Here we show that Leflunomide exerts immunosuppression but limited efficacy in RA individuals distinguished by higher serum C-reactive protein (CRP
, CRP
), whereas the others with satisfactory responsiveness to Leflunomide show lower CRP (CRP
, CRP
). CRP inhibition decreases bone erosion in arthritic rats. Besides the immunomodulation via A77 1726, Leflunomide itself induces AHR-ARNT interaction to inhibit hepatic CRP production and attenuate bone erosion in CRP
arthritic rats. Nevertheless, high CRP in CRP
rats upregulates HIF1α, which competes with AHR for ARNT association and interferes Leflunomide-AHR-CRP signaling. Hepatocyte-specific HIF1α deletion or a HIF1α inhibitor Acriflavine re-activates Leflunomide-AHR-CRP signaling to inhibit bone erosion. This study presents a precision medicine-based therapeutic strategy for RA.
Currently, major concerns about the safety and efficacy of RNA interference (RNAi)-based bone anabolic strategies still exist because of the lack of direct osteoblast-specific delivery systems for ...osteogenic siRNAs. Here we screened the aptamer CH6 by cell-SELEX, specifically targeting both rat and human osteoblasts, and then we developed CH6 aptamer-functionalized lipid nanoparticles (LNPs) encapsulating osteogenic pleckstrin homology domain-containing family O member 1 (Plekho1) siRNA (CH6-LNPs-siRNA). Our results showed that CH6 facilitated in vitro osteoblast-selective uptake of Plekho1 siRNA, mainly via macropinocytosis, and boosted in vivo osteoblast-specific Plekho1 gene silencing, which promoted bone formation, improved bone microarchitecture, increased bone mass and enhanced mechanical properties in both osteopenic and healthy rodents. These results indicate that osteoblast-specific aptamer-functionalized LNPs could act as a new RNAi-based bone anabolic strategy, advancing the targeted delivery selectivity of osteogenic siRNAs from the tissue level to the cellular level.
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