Advancements in Antisense Oligonucleotide Therapy: A Potential Breakthrough in Hepatocellular Carcinoma Treatment


Hepatocellular carcinoma (HCC) is a malignancy that continues to challenge the medical community with its rising incidence and formidable prognosis. Current treatment options are limited, and the development of effective therapies has been hampered by the complex biology of this disease. antisense oligonucleotides (ASOs), a class of synthetic molecules designed to modulate gene expression at the mRNA level, have recently garnered significant attention due to their precision in targeting specific genes or pathways.

In a new study published in the peer-reviewed  Molecular Therapy – Nucleic Acids Journal  led by Yufei Pan, Yuejie Zhu, Huantong Li, Hua Guo, Qianyi He, Zhu Guan, and Zhenjun Yang from the School of Pharmaceutical Sciences at Peking University in collaboration with Jing Guan from Guizhou University and Yujing Gao from Chengdu University of Traditional Chinese Medicine  conducted a comprehensive investigation into the potential therapeutic efficacy of ASOs, particularly focusing on one specific ASO named CT102, in the context of HCC.

Central to the understanding of ASO therapy’s impact on HCC is the comprehensive analysis of transcriptomic and proteomic alterations induced by these molecules. The authors conducted a rigorous evaluation, revealing thousands of mRNA and protein changes within HCC cells upon exposure to CT102. These changes were not haphazard but orchestrated, signifying a meticulous orchestration of gene expression. Furthermore, the modulation of multiple signaling pathways became apparent, illustrating the pleiotropic nature of ASO intervention. Of particular significance was the identification of 13 upregulated and 5 downregulated genes and proteins by CT102, signaling its dual role in gene regulation. These differentially expressed genes were enriched in pathways intricately linked to tumorigenesis. It is noteworthy that aside from the well-documented influence of ASOs on tyrosine kinase receptor-mediated signal transduction via IGF1R, other pathways, including PPAR, P53, DNA replication, and mismatch repair, exhibited notable alterations. This multifaceted impact on signaling pathways underscores the potential of ASOs to exert comprehensive control over cancer cell biology. They explored the concept of chemical modification to enhance the specificity and efficacy of ASO therapy. By modifying CT102 with 2ʹ-OMOE, they sought to improve its activity, ultimately achieving a higher level of target gene silencing.

The subcellular localization of ASOs has been a topic of considerable interest. In this study, approximately 25% of differentially expressed proteins were found to localize in the nucleus following ASO treatment. These nuclear-resident proteins were engaged in pivotal cellular processes, including transcription, phosphorylation, phosphatase activation, and DNA/RNA binding. The nucleus, long considered the “control center” of the cell, is a nexus for regulating gene expression. The ability of modified CT102s to influence nuclear proteins suggests their profound impact on intracellular processes.

Remarkably, the authors also uncovered the capacity of modified CT102s to decrease the expression of IGF1R mRNA in both the nucleus and cytoplasm. Additionally, several genes with functional roles in tumorigenesis, such as GAS2, POLA2, LGALS2, and IGFBP1, were significantly affected. The mechanistic basis for these observations is intriguing, potentially involving imperfect base pairing or other complex interactions. These findings highlight the versatility of ASOs, not confined to mere mRNA binding but extending to multifaceted roles within the cellular nucleus. The authors also investigated the potential of glycoconjugation as a strategy to further enhance the therapeutic effects of ASOs. Glycoconjugates were evaluated for their ability to promote cell proliferation inhibition and target gene silencing, potentially through increased cellular uptake via glycosyl receptors on the cell surface. Recognizing the challenges of in vivo delivery of ASOs, the researchers developed a cytidinyl lipid-based delivery system known as DCP (Delivery Carrier Platform). This platform was designed to optimize the transfection of ASO CT102 into HCC cell lines. They systematically examined various formulations to achieve efficient and targeted delivery to the liver. The researchers focused on the PI3K-AKT signaling pathway, which is frequently dysregulated in cancer, including HCC. They evaluated how CT102 and its derivative conjugates affected the synthesis of the target protein IGF1R and how this inhibition impacted downstream signaling within the PI3K-AKT pathway.

The study investigated the complex mechanisms underlying ASO action within the nucleus. It discovered that modified CT102s not only influenced the PI3K-AKT pathway but also affected DNA replication, anti-inflammation, the immune microenvironment, and various other cellular processes crucial for tumor growth. Additionally, they identified interactions between ASOs and other mRNAs via imperfect matches, leading to the downregulation of related proteins and the induction of apoptotic processes in tumor cells.

The study’s findings underscore the potency of ASOs when subjected to chemical modification. By introducing 2ʹ-OMOE modification, the researchers achieved a substantial improvement in the activity of gapmer CT102. This modification translated into a remarkable silencing level of target genes, reaching nearly 80% at a concentration of 100 nM. Such a high level of target affinity is a promising development, as it suggests the potential for robust gene regulation. Further enhancing the therapeutic potential of ASOs, the authors explored glycoconjugates as a means to promote cell proliferation inhibition and target gene silencing. Glycoconjugates are known to interact with glycosyl receptors on the cell surface, facilitating cellular uptake. This strategy exhibited significant promise in vitro and, more importantly, in vivo, as it allowed for reduced dosing intervals and lower doses, ultimately enhancing the anti-tumor efficacy.

The PI3K-AKT signaling pathway, frequently dysregulated in various cancers, emerged as a focal point in this study. CT102 and its derivative conjugates, notably Glu-CT102MOE5, effectively curtailed the synthesis of the target protein IGF1R. This inhibition reverberated downstream, suppressing the PI3K-AKT pathway and, consequently, impeding tumor growth. The role of this pathway in tumorigenesis is well-established, making it an attractive target for therapeutic intervention.

Interestingly, the study compared ASOs and siRNAs targeting IGF1R mRNA. While both exhibited similar target gene silencing abilities, ASOs displayed a distinct advantage in terms of their antiproliferative capabilities. This observation hints at the multifaceted roles of ASOs within cells, which extend beyond mere gene silencing. ASOs, it appears, are endowed with the capacity to influence multiple signaling pathways and cellular processes, thereby offering a more comprehensive approach to inhibiting tumor development.

The precise mechanisms underlying the actions of ASOs have long intrigued researchers. This study offers valuable insights into the multifaceted roles that ASOs play within the nucleus, a domain where gene regulation is orchestrated with utmost precision. Transcriptomic and proteomic analyses revealed that modified CT102s not only impacted the PI3K-AKT pathway but also exerted influence over DNA replication, anti-inflammation, the immune microenvironment, and various other processes central to tumor growth. Intriguingly, ASOs were found to interact with other mRNAs via imperfect matches, resulting in the downregulation of related proteins and the induction of apoptotic processes within tumor cells. This nuanced mechanism suggests that ASOs extend beyond the scope of traditional mRNA targeting, providing a more intricate and versatile approach to cancer therapy.

In conclusion, the findings of Professor Zhenjun Yang and colleagues presented herein illuminate a path forward in the treatment of HCC, offering not only hope for patients but also avenues for further research and clinical application. The study’s revelations regarding the complex mechanisms of ASO action within the nucleus emphasize the need for continued exploration into the intricate biology of cancer cells and the potential of ASOs to unlock novel therapeutic strategies.


Pan Y, Guan J, Gao Y, Zhu Y, Li H, Guo H, He Q, Guan Z, Yang Z. Modified ASO conjugates encapsulated with cytidinyl/cationic lipids exhibit more potent and longer-lasting anti-HCC effects. Mol Ther Nucleic Acids, 2023, 32: 807-821. doi: 10.1016/j.omtn.2023.04.028.

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