What are the different types of drugs available for circular RNA?

Introduction to Circular RNA
Circular RNAs (circRNAs) are a unique class of RNA molecules characterized by a covalently closed loop structure that distinguishes them from their linear counterparts. Unlike traditional mRNAs, circRNAs lack 5′ caps and 3′ tails, which renders them exceptionally resistant to exonuclease degradation. This structural stability, along with notable cell- and tissue-specific expression patterns, has made circRNAs a subject of increasing interest in both basic biological research and clinical applications.

Definition and Characteristics
circRNAs are produced predominantly through a process known as back-splicing, where a downstream splice donor site ligates with an upstream splice acceptor site, forming a continuous loop. Their biosynthesis may arise from exonic, intronic, or exon–intron sequences, and in some cases, circRNAs can encode proteins in a cap-independent manner. These molecules can vary widely in size, ranging from a few hundred to several thousand nucleotides, and yield diverse functions, including acting as microRNA (miRNA) sponges, interacting with RNA-binding proteins (RBPs), or regulating transcription. Their enhanced stability compared to linear RNAs positions them as promising candidates for therapeutic interventions.

Role in Cellular Functions
Beyond serving as potential biomarkers for diseases, circRNAs participate actively in cellular regulatory networks. They are known to regulate gene expression by sequestering miRNAs, influencing protein translation, and possibly serving as templates for protein synthesis when conditions permit. In cancer biology, for instance, certain circRNAs act either as oncogenes or tumor suppressors by modulating signaling pathways and the activity of crucial cellular regulators. In addition, circRNAs have been implicated in controlling immune responses, drug resistance, and overall cellular homeostasis, giving them a multifaceted role in both normal physiology and disease states.

Drug Categories Targeting Circular RNA
Over the years, several drug categories have been developed or are under investigation to modulate the function and expression of circRNAs. These therapeutic approaches often target either the circRNAs themselves or rely on using circRNAs as a platform for therapeutic intervention. The major drug categories include small molecule drugs, RNA-based therapeutics, and antisense oligonucleotides—each leveraging distinct mechanisms to interact with or alter circRNA behavior.

Small Molecule Drugs
Small molecule drugs targeting RNA have traditionally been developed to interact with specific RNA structures, modulating RNA processing, splicing, or translation. For circRNAs, the unique looped structure and secondary conformations open up novel avenues for small molecule intervention. These compounds are designed to bind specific regions of the circRNA, thereby altering its interaction with miRNAs, RBPs, or other components of cellular machinery. For instance, by binding to regions responsible for back-splicing or by influencing interactions within the circRNA, these molecules may modulate its stability or function in disease processes.

Moreover, recent efforts aimed at targeting structured RNA domains have led to the development of RNA-targeted small molecules. These agents are designed using high-throughput screening strategies that identify compounds capable of selectively binding structured regions within circRNAs. The binding of small molecules to circRNAs can hinder or promote the recruitment of proteins or other RNAs, thereby modulating the circRNA’s function in post-transcriptional regulation. In the context of drug discovery for circRNAs, chemical modifications and scaffold optimizations have been crucial to attain binding affinity, specificity, and favorable drug-like properties. This strategy, although still in an early phase of development for circRNAs, holds promise as part of a broader approach in RNA-targeted pharmacology.

RNA-based Therapeutics
RNA-based therapeutics encompass a wide range of modalities that exploit the inherent properties of RNA molecules for therapeutic benefit. In the realm of circRNAs, this broad category includes approaches where circRNAs are used as drugs themselves or as carriers of information (such as coding for proteins or peptides). RNA-based therapeutics targeting circRNAs or employing synthetic circRNAs include:

1. Circular RNA Vaccines
Synthetic circRNA molecules have recently been explored as vaccines due to their improved stability and enhanced protein expression profiles. For example, circular RNA vaccines are engineered to encode antigenic polypeptides that, once delivered to cells, can be efficiently translated without the need for a 5′ cap. This self-circularization leads to prolonged translation and a more robust immunogenicity profile compared to linear mRNA vaccines. Such vaccines benefit from a simplified production process and an inherent lower risk of triggering unwanted immune responses.

2. Therapeutic Protein Replacement Platforms
By leveraging the translation potential of circRNAs, companies are developing synthetic circRNAs for applications beyond vaccination. These platforms can be designed to express therapeutic proteins for conditions such as enzyme deficiencies, targeted protein replacement therapy, or even for generating bio-PROTACs in cancer, where the circRNA encodes a protein that induces targeted degradation of oncogenic proteins. The enhanced durability and expression efficiency of circRNAs make them a compelling candidate for addressing diseases where traditional protein therapy might fail.

3. Exogenous circRNA Delivery Systems
The field also includes therapeutic strategies where exogenous circRNAs are delivered to cells to modulate gene expression networks. In these cases, circRNAs can function as sponges to sequester microRNAs or as regulators of gene transcription directly. Their stability in the extracellular environment and reduced immunogenicity compared to linear RNA supports their use as therapeutic agents, providing an alternative avenue to conventional antisense or siRNA-based interventions.

Antisense Oligonucleotides
Antisense oligonucleotides (ASOs) represent a mature class of RNA-targeted therapeutics that involves short, synthetic nucleic acid sequences designed to bind specific RNA targets through Watson-Crick base pairing. ASOs can be tailored to interact with circRNAs by selectively targeting the unique back-splice junctions—a feature that distinguishes circRNAs from their linear counterparts.

1. Mechanism in the Context of circRNAs
ASOs designed to target circRNAs typically aim to modulate circular RNA function by either blocking miRNA binding sites or promoting circRNA degradation by engaging RNase H when bound to the circRNA. Given the high stability of circRNAs, ASOs often need to be chemically modified to improve binding affinity, cellular uptake, and nuclease resistance. Additionally, they can be designed as gapmers, where a central DNA segment flanked by modified ribonucleotides enhances the ability to recruit RNase H for targeted RNA cleavage.

2. Therapeutic Advantages and Challenges
The primary advantage of ASOs directed at circRNAs is their specificity. By focusing on the back-splice junction, ASOs avoid off-target effects on the parental linear mRNA, thus improving precision. However, challenges remain in terms of delivery, as the circRNA’s circular structure might sometimes demand innovative delivery systems to ensure adequate intracellular uptake. Moreover, while antisense methods have been extensively applied in other areas, such as targeting mRNA in cancer or neurological disorders, their application toward circRNA-specific interventions is gaining momentum and represents a promising frontier for targeted gene modulation.

Mechanisms of Action
Drugs designed to target circRNAs employ several sophisticated mechanisms that either disrupt the function of circRNAs or harness their properties for therapeutic benefit. These mechanisms are based on either direct interactions with the circRNAs or modulation of the downstream pathways regulated by circRNAs, ensuring precise control over cellular functions and disease progression.

Interaction with Circular RNA
Small molecules and ASOs interact with circRNAs primarily through sequence-specific and structure-specific mechanisms. Small molecules are designed to recognize unique secondary or tertiary structures within the circRNA. By binding to these regions, a drug can sterically hinder the interactions between the circRNA and its cellular partners, such as miRNAs or RNA-binding proteins. This disruption can either prevent the circRNA from acting as a sponge or alter its normal localization within the cell. Conversely, ASOs directly bind to the unique back-splice junctions or specific functional domains within the circRNA, thereby blocking binding sites for other cellular factors or inducing targeted degradation.

Furthermore, RNA-based therapeutics that employ exogenous circRNAs function by mimicking natural circRNA interactions. When administered, these synthetic circRNAs can compete with endogenous circRNAs for binding to proteins or miRNAs, altering gene regulatory networks in a manner that is therapeutically beneficial.

Modulation of Circular RNA Functions
The modulation of circRNA functions can be approached from two angles. On one hand, drugs can downregulate or degrade aberrant circRNAs that contribute to disease pathology. For example, in cancer, overexpressed oncogenic circRNAs may be targeted for degradation using ASOs, alleviating their sponging effects on tumor-suppressor miRNAs.

On the other hand, therapeutic strategies might aim to upregulate beneficial circRNAs that act as tumor suppressors or modulators of critical cellular processes. Synthetic circRNAs can be administered to re-establish regulatory balance, promote protein expression through cap-independent translation, or act as adjuvants in immune responses. This dual approach—either inhibiting pathogenic circRNAs or supplementing protective ones—exemplifies the versatility of the drug design strategies available for circRNA modulation.

The underlying mechanisms involve not only the primary binding and molecular recognition events but also downstream modulation of signaling pathways, protein interactions, and translational control. By targeting specific molecular interactions, these drugs can influence the dynamic equilibrium between pathological and protective cellular phenotypes.

Therapeutic Applications
The diverse functions of circRNAs in cellular regulation make them attractive therapeutic targets for several diseases, especially in areas with complex molecular underpinnings such as cancer and neurological disorders. Here, we discuss how different drug categories targeting circRNAs are being applied to tackle these conditions.

Cancer Treatment
Cancer remains one of the primary fields where circRNA-targeted therapeutics are receiving significant attention. Many circRNAs have been identified as either oncogenic drivers or tumor suppressors in various cancers. For instance, specific circRNAs modulate pathways that govern cell proliferation, apoptosis, and metastasis. By deploying ASOs that target overexpressed oncogenic circRNAs, researchers aim to restore the normal regulatory balance, thereby inhibiting tumor growth.

In addition, RNA-based therapeutics using exogenous circRNAs have shown promise in cancer immunotherapy models. Synthetic circRNAs that encode tumor antigens can act as vaccines, stimulating robust immune responses against cancer cells. These circular RNA vaccines offer advantages such as prolonged antigen expression and an improved safety profile compared to their linear counterparts. Moreover, small molecule drugs designed to interact with and modulate circRNA function could potentially disrupt the oncogenic regulatory circuits, adding a layer of precision in cancer treatment.

The combination of these strategies—using ASOs for knockdown of deleterious circRNAs, RNA-based vaccines for antigen presentation, and small molecules for modulating circRNA structure—allows for a multi-pronged attack on cancer, addressing the disease from several molecular angles.

Neurological Disorders
Neurological disorders, including Parkinson’s disease and Alzheimer’s disease, have also emerged as key targets for circRNA-based therapeutics. The stability and specific expression patterns of circRNAs in neural tissues render them particularly attractive for addressing neurodegenerative conditions. In experimental models of Parkinson’s disease, for example, circRNAs implicated in neurodegeneration are being targeted by ASOs and RNA interference strategies to restore normal neuronal function.

Furthermore, synthetic circRNAs can be engineered to express neuroprotective proteins or factors critical for neuronal survival, offering a novel strategy for protein replacement therapy in neurodegenerative disorders. The long-term stability of circRNAs ensures sustained therapeutic effects, which is especially important in chronic conditions where consistent protein expression can mitigate progression.

Additionally, small molecule drugs that modulate circRNA-protein interactions in neural tissues could potentially alter the neurodegenerative cascade. Such drugs can fine-tune the expression and activity of circRNAs involved in inflammatory responses, synaptic regulation, and neuronal apoptosis, thereby providing a new tool in the therapeutic arsenal against neurological diseases.

Current Research and Future Directions
The field of circRNA-targeted drugs is rapidly evolving, with several exciting developments in preclinical and clinical settings. Current research focuses not only on refining the existing drug categories but also on addressing challenges in delivery, specificity, and off-target effects. Future research directions aim to build upon these strategies to produce even safer and more effective therapies.

Ongoing Clinical Trials
Recent years have seen a surge in clinical trials targeting various RNA molecules, and circRNA-based interventions are beginning to follow suit. For instance, early-phase clinical studies have investigated the efficacy and safety of circular RNA vaccines for cancer treatment, capitalizing on their favorable immunogenicity and protein expression profiles. Similarly, ongoing trials are evaluating the knockdown of oncogenic circRNAs in cancer patients using ASOs designed for high specificity by targeting back-splice junctions.

Moreover, RNA-based therapeutic platforms using synthetic circRNAs are currently being studied for their ability to produce sustained therapeutic protein expression in vivo. These clinical trials are essential for validating preclinical findings, optimizing dosing regimens, improving delivery platforms (such as lipid nanoparticles), and assessing long-term effects. The outcomes of these studies are expected to provide pivotal insights into the translational potential of circRNA drugs and help streamline regulatory pathways for future approvals.

Future Research Directions
Looking forward, research in the area of circRNA therapeutics is poised to explore several avenues for improvement and innovation:

1. Enhanced Compound Screening and Optimization
There is an increasing need for advanced screening methods capable of identifying small molecule compounds with high affinity and specificity for circRNA targets. This involves leveraging high-throughput sequencing, structural prediction algorithms, and computational modeling to design molecules that precisely interact with circRNA secondary and tertiary structures. Improving these processes will drive the rational design of next-generation small molecule drugs that can selectively modulate circRNA functions.

2. Improved Delivery Technologies
One of the core challenges remains the safe and efficient delivery of circRNA-based therapeutics. Future research will need to concentrate on the development of novel delivery systems, including nanoparticle formulations, viral vectors, and conjugation technologies that can target specific tissues and cell types with minimal off-target effects. This is especially critical for neurological disorders, where crossing the blood–brain barrier represents a major hurdle.

3. Combinatorial Therapeutic Strategies
The integration of circRNA-targeted drugs with other therapeutic modalities, including immunotherapies, chemotherapies, and conventional targeted therapies, could provide synergistic benefits. For example, combining a circular RNA vaccine with checkpoint inhibitors might enhance the anti-tumor immune response in cancer patients. There is also potential for using circRNA modulation in combination with antisense or miRNA-based treatments to achieve a more comprehensive suppression of oncogenic pathways.

4. Understanding circRNA Biology at a Deeper Level
Future studies must continue to elucidate the biological functions and regulatory networks of circRNAs in various tissues and disease contexts. This includes mapping circRNA–protein and circRNA–miRNA interactions comprehensively, investigating their roles in cellular metabolism, and understanding how changes in circRNA levels affect disease outcomes. Such fundamental insights will inform the design of more effective therapeutic agents.

5. Personalized Medicine Approaches
Given the cell-type and tissue-specific expression patterns of circRNAs, personalized medicine offers an exciting direction for future research. By profiling circRNAs in individual patients, clinicians could tailor therapies to target specific aberrations in the circRNA landscape. This personalized approach holds the promise of increasing treatment efficacy and reducing side effects.

6. Safety and Toxicity Studies
As with any new therapeutic modality, thorough evaluation of long-term safety and toxicity is crucial. Future research is likely to focus on comprehensive in vivo studies that assess the immunogenicity and potential off-target effects of circRNA therapeutics, whether they are based on small molecules, ASOs, or synthetic circRNAs. These investigations will be vital for securing regulatory approvals and for the broader clinical adoption of these novel drugs.

Conclusion
In summary, the landscape of drugs available for targeting circular RNAs is multifaceted and rapidly evolving. Researchers are exploring three major categories of circRNA-targeted drugs:

1. Small molecule drugs are being designed to interact with and modulate the unique secondary and tertiary structures of circRNAs. These compounds aim to disrupt pathogenic interactions between circRNAs and other cellular molecules, thereby rebalancing dysregulated gene expression.

2. RNA-based therapeutics exploit the intrinsic properties of circRNAs by employing synthetic circRNAs as vaccines, protein replacement platforms, or modulators of gene regulatory networks. The durability and improved immunogenic profiles of circRNAs render them particularly attractive for applications such as cancer treatment and immune modulation.

3. Antisense oligonucleotides (ASOs) offer a highly specific means of targeting circRNAs by binding to the unique back-splice junctions. By doing so, these ASOs can block or degrade pathogenic circRNAs, thereby restoring normal cellular functions—a strategy that is especially promising for diseases like cancer and neurological disorders.

The mechanisms of action for these drugs involve direct binding to circRNAs, interference with their interactions with miRNAs and RNA-binding proteins, and modulation of downstream signaling pathways. Therapeutically, these approaches have the potential to revolutionize treatment paradigms in cancer by suppressing oncogenic circuits and in neurological diseases by ensuring sustained expression of neuroprotective proteins. Current research is focused on refining delivery methods, optimizing molecular designs, and integrating combinatorial strategies, while ongoing clinical trials promise to shed light on the safety and efficacy of these novel therapeutics.

In conclusion, the development of drugs targeting circRNAs represents a promising frontier in personalized and precision medicine. The diverse strategies—ranging from small molecule inhibitors to RNA-based vaccines and specifically designed antisense oligonucleotides—offer innovative approaches to modulate circRNA function. As research deepens our understanding of circRNA biology, and as clinical trials begin to validate these therapies, we are likely to witness a significant transformation in the management of complex diseases. The future direction of circRNA-targeted therapeutics will undoubtedly benefit from interdisciplinary collaborations, advanced delivery technologies, and a commitment to rigorous preclinical and clinical investigation, ultimately paving the way for transformative treatments across a broad spectrum of human diseases.