Internal Ribosome Entry Site (IRES)-Mediated Translation and Its Potential for Novel mRNA-Based Therapy Development

Abstract

Many conditions can benefit from RNA-based therapies, namely, those targeting internal ribosome entry sites (IRESs) and their regulatory proteins, the IRES trans-acting factors (ITAFs). IRES-mediated translation is an alternative mechanism of translation initiation, known for maintaining protein synthesis when canonical translation is impaired. During a stress response, it contributes to cell reprogramming and adaptation to the new environment. The relationship between IRESs and ITAFs with tumorigenesis and resistance to therapy has been studied in recent years, proposing new therapeutic targets and treatments. In addition, IRES-dependent translation initiation dysregulation is also related to neurological and cardiovascular diseases, muscular atrophies, or other syndromes. The participation of these structures in the development of such pathologies has been studied, yet to a far lesser extent than in cancer. Strategies involving the disruption of IRES-ITAF interactions or the modification of ITAF expression levels may be used with great impact in the development of new therapeutics. In this review, we aim to comprehend the current data on groups of human pathologies associated with IRES and/or ITAF dysregulation and their application in the designing of new therapeutic approaches using them as targets or tools. Thus, we wish to summarise the evidence in the field hoping to open new promising lines of investigation toward personalised treatments.

Keywords: IRES trans-acting factors; IRES-based multicistronic vectors; RNA-based therapies; antisense oligonucleotides; internal ribosome entry sites.

Figure 1

Cap-dependent versus internal ribosome entry site (IRES)-dependent translation initiation. (A): Canonical 5′ cap-dependent translation initiation. The canonical eukaryotic translation initiation depends on the recognition of the cap structure at the 5′ end of transcripts by the small ribosomal subunit (the 40S). The binding of several eukaryotic initiation factors (eIF1, eIF1A, eIF3, and eIF5) to the 40S subunit and the simultaneous formation of the ternary complex, composed of eukaryotic initiation factor (eIF) 2 bound to guanosine triphosphate (GTP) and initiator methionyl-tRNA, allows the assembly of the 43S pre-initiation complex (PIC). Simultaneously, a group of eIF4 factors is responsible for some of the interactions that eventually lead to mRNA activation. eIF4E binds to the 5′ cap and consequently to eIF4G, and eIF4A interacts with eIF4G:eIF4E, thus forming the trimeric eIF4F complex. Once these connections have been established, the 43S PIC binds to the cap structure at the mRNA 5′ end, becoming the 48S initiation complex, which in turn scans the 5′ untranslated region (UTR) until reaching the first initiation codon in a favourable context. After the codon recognition, 60S subunit joining and consequent 80S formation are induced, hence promoting further elongation and peptide synthesis. At this stage, eIF2 recycling is required to enable another round of translation initiation. (B): IRES-dependent translation initiation. This alternative mode of translation initiation does not need cap recognition nor the scanning of the 5′ UTR. Instead, there are some elements, the internal ribosome entry sites, which are intricate mRNA secondary structures usually located within the 5′ UTR of the transcript, which can directly recruit the 40S subunit to the vicinity of the initiation codon. This binding does not require complete assistance from eIFs, happening with the help of just a few eIFs or some IRES trans-acting factors (ITAFs), RNA-binding proteins that regulate IRES activity, either activating or repressing it. Once again, after the recognition of the initiation codon, both ribosomal subunits are assembled and ready for elongation, thus leading to peptide synthesis.

Figure 2

A model proposing the use of antisense oligonucleotides (ASOs) to modulate internal ribosome entry site (IRES) or IRES trans-acting factor (ITAF) activity and further protein expression. (A) An ASO targeting an IRES responsible for translating a pathogenic protein would lead to the disruption of IRES activity and hence hinder protein synthesis. (B) ASO targeting activator ITAF mRNA. If the ITAF enhances IRES activity, disrupting its expression would lead to impaired IRES activity and subsequent protein synthesis inhibition. (C) ASO targeting inhibitory ITAF mRNA. If the ITAF represses IRES activity, disrupting its expression would allow IRES activation and consequent regular protein synthesis.

Figure 3

Mechanism of action of RNA-based therapies using internal ribosome entry sites (IRESs) as targets or tools. (A) IRESs as targets. RNA-based drugs, which can be composed of different RNA molecules, such as antisense oligonucleotides (ASOs), antagonists, or small-molecule inhibitors, are delivered into the bloodstream and carried to specific cells. Such RNA compounds bind to the mRNA and promote alterations in the IRES-mediated translation of pathogenic proteins, by blocking or cleaving the IRES structure, disrupting the interactions between the IRES element and both the ribosome and IRES trans-acting factors (ITAFs) and blocking the activity of ITAFs. These modifications lead to protein synthesis impairment, which may be crucial for the treatment or prevention of several pathologies, besides cancer. (B) IRESs as tools. Circular RNAs with IRESs, and several IRES-based viral vectors, have been used to produce non-pathogenic proteins with a therapeutic role. These vectors are injected into the bloodstream and then bound to specific cells. Once expressed, they can promote the simultaneous expression of more than one protein. The expression of these vectors allows for maintaining or enhancing the expression levels of proteins with important biomedical properties that present a positive effect on different conditions, constituting an important method of gene therapy and strategy of treatment for several diseases.