OligoY - a new class of oligonucleotide-based therapeutic means
Lead Research Organisation:
Sixfold Bioscience Ltd
Department Name: Head office
Abstract
A number of RNA drugs have been developed to combat polygenic diseases such as cancer. The early enthusiasm for the promise of RNAi therapeutics was quickly weakened by the failure or limited efficacy in early clinical trials.1 Decades of studies on modification of nucleic acids which can affect pairing and chemical stability, conformation and interactions with a proteins and enzymes involved in uptake, transport or processing of targets have been the main focus of the oligonucleotide therapeutic field. While many RNA-based drugs are in development, there are still significant barriers to efficient RNA-based treatment strategies including delivery of the drug to a particular site or tissue, off-target effects and longevity of the treatment.
RNAi, antisense and splice-switching oligonucleotides are the main class of therapeutic still under screening. RNAi drugs, although being showing promising result in clinical trial and having some representative drugs already FDA approved, are intrinsically presenting some limitations due to their modality of action. The RNAi and miRNA pathway share common proteins, like Ago2, which are involved in miRNA maturation. Hence, siRNA drugs compete with endogenous miRNAs for Ago2 binding, limiting the efficiency of all siRNA drugs. To compensate this, the concentration of the RNA drug must be carefully optimized avoiding an excess that could saturate the RISC machinery and inhibiting the normal miRNA pathway.
The proposed project investigates the design of a new drug RNA drug, 'OligoY', which blocks the most essential mechanism in cell proliferation, DNA replication. DNA replication is the cell process of producing two identical copies of DNA from one DNA template. It is the most essential process in cell division but its initiation is still poorly understood. DNA replication in eukaryotes employs several extracellular signals to coordinate the specialized compartment of the multicellular organisms. OligoYs act indirectly on replication through the binding with a family of non-coding RNA, Y RNAs, specifically blocking the interaction of Y RNA with ORC (Origin Replication complex) during the replication initiation step. OligoYs are complexes formed by two strands of chemically modified RNAs, complementary to the Y RNA target, and a third element (peptide, aptamer, antibody, POD) conferring cell-selectivity. To test their application as drug, they will be initially evaluated in several cancer cell lines. OligoYs are potentially the first example of an oligonucleotide-based drug that acts on the DNA replication mechanism, bypassing all the drawbacks associated to variations of the disease during treatment and between patients.
RNAi, antisense and splice-switching oligonucleotides are the main class of therapeutic still under screening. RNAi drugs, although being showing promising result in clinical trial and having some representative drugs already FDA approved, are intrinsically presenting some limitations due to their modality of action. The RNAi and miRNA pathway share common proteins, like Ago2, which are involved in miRNA maturation. Hence, siRNA drugs compete with endogenous miRNAs for Ago2 binding, limiting the efficiency of all siRNA drugs. To compensate this, the concentration of the RNA drug must be carefully optimized avoiding an excess that could saturate the RISC machinery and inhibiting the normal miRNA pathway.
The proposed project investigates the design of a new drug RNA drug, 'OligoY', which blocks the most essential mechanism in cell proliferation, DNA replication. DNA replication is the cell process of producing two identical copies of DNA from one DNA template. It is the most essential process in cell division but its initiation is still poorly understood. DNA replication in eukaryotes employs several extracellular signals to coordinate the specialized compartment of the multicellular organisms. OligoYs act indirectly on replication through the binding with a family of non-coding RNA, Y RNAs, specifically blocking the interaction of Y RNA with ORC (Origin Replication complex) during the replication initiation step. OligoYs are complexes formed by two strands of chemically modified RNAs, complementary to the Y RNA target, and a third element (peptide, aptamer, antibody, POD) conferring cell-selectivity. To test their application as drug, they will be initially evaluated in several cancer cell lines. OligoYs are potentially the first example of an oligonucleotide-based drug that acts on the DNA replication mechanism, bypassing all the drawbacks associated to variations of the disease during treatment and between patients.
Organisations
People |
ORCID iD |
Publications
Bonetti A
(2023)
RNA Structure and Function
Ghidini A
(2023)
The RNA-Recognition Pathway: An Overlooked Transportation Mechanism for Extracellular and Therapeutic RNAs
in GEN Biotechnology
Sixfold Bioscience
(2023)
A revolution in targeted RNA delivery
in Nature Research, Advertisement Feature
| Description | We have achieved most of the milestones of the objectives indicated in the original work plan. Objective 2 was recently started and the library of oligos is currently being tested on a immortalized cell line. The collaborations with Petzold and NMR has been changed to Prof Ramos at UCL, but is currently on hold until we have a hit from the first screening in cells. Two of the major findings are: 1. Bacterial RNA to be used as a model for oligo targeting 2. Disruption of human Y RNA1 by TFO and ssOligos (both in vitro and in cells) Both projects are currently being brought forward from the team and we are currently scoping the landscape for potential patents. Objective 1: the design, synthesis and in vitro assay of the first library of OligoY sequences has been carried out. The bioconjugation of the two oligonucleotides strands through click chemistry was tested but this resulted in poor yield. Although we were able to collect enough material for an initial characterisation of the RNA target binding, we are currently looking for an alternative reaction. The first library of OligoY has been tested for binding of the target RNA (Y RNA1) through DSF melting denaturation assay. The constructs showed high affinity for the folder hairpin target. We are currently working on differentiating the two binding events, the duplex and the triplex formation. The SPR binding assay could not be run since we are still currently applying for access to the Imperial facility, who owns the instrument. The first library has been tested in hypotriploid alveolar basal epithelial cells and one of the compounds has shown a delay in cell proliferation after four days of incubation. Objective 4: we have analysed and benchmarked the field of bacteria targeting. We have identified the main limitations we may face during the design of the bacteria-targeting oligos. We have chosen some classes of bacterial RNAs as targets which are highly structured as OligoY. We used Escherichia_coli_str_K-12_substr_MG1655 as a model and after running an alignment with human RNA, we found a unique sequence stretch which we can target with an oligonucleotide. We have designed a library of modified Oligos which will be tested for in vitro target binding. |
| Exploitation Route | Obj 1: Aside from the scientific benefits, the knowledge gained in this project could contribute more broadly to societal and economic benefits for UK by the engineering of new therapeutic tools, which could be tailor-designed for specific diseases. The chosen target for the described OligoY molecule is common to all cells, and thus has the benefit of decreasing the potential variability of drug responses among patients and diseases stages. Obj 4: Discovery and development of new antibacterial drugs against multidrug resistant bacterial strains have become increasingly urgent. Many studies have focused on combatting viral infections with RNA therapies. Currently, most viral infections do not have specific drugs, and rely on preventive vaccination to combat the pathogen. Therefore, RNA-based solutions are attractive, as there is the potential to directly and specifically halt expression of the viral genes required for the infection to spread. Oligonucleotides show immense potential to control the spread of resistant microbes due to its high specificity of action, little risk to human gene expression, and easy design and synthesis to target any possible gene. Obj 5: Although the field of biophysics and mechanistic biology are well-established in terms of protein characterization, structural prediction, and molecular interactions, the field of RNA-therapeutics has only recently transitioned to an industrial scale. Consequently, many of the prevalent techniques used in small molecule (SM)/protein interactions and protein/protein interactions are not directly applicable to RNA. This creates challenges and necessitates which require the development of specialised approaches specifically tailored for RNA-therapeutics. We designed a comprehensive pipeline to be executed over the year, initially deploying assays like SEC and FIDA to qualitatively confirm the presence of Mergo:protein complexes in a smaller subset of Mergos. Following this, we explored more quantitative and specific assays to identify the proteins involved. Subsequently, we expanded our analysis to a larger set of Mergos, enabling us to compare various scaffolds and assess their differences and similarities. We are currently working on understanding how the different biological matrices influence the assays and which component may need further confirmation. |
| Sectors | Chemicals Healthcare Pharmaceuticals and Medical Biotechnology |
| Description | Project outputs have led to securing further funding (A4i project with MHRA in 2023). Further, funding has generated key data, contributing to Sixfold's ongoing due-diligence investor data room to secure Series A financing. This key data has formed part of larger data packages shared with potential partners interested in future collaborations. Engagement with both academic and industrial scientists facilitated knowledge transfer (discussing challenges and advancements in oligonucleotide therapeutics development). |
| First Year Of Impact | 2023 |
| Sector | Pharmaceuticals and Medical Biotechnology |
| Impact Types | Societal Economic |
| Description | Analysis for Innovators |
| Amount | £79,989 (GBP) |
| Funding ID | 10074412 |
| Organisation | Innovate UK |
| Sector | Public |
| Country | United Kingdom |
| Start | 07/2023 |
| End | 01/2024 |
| Title | Code developed for alignment bacterial RNA target and human RNA genome |
| Description | The team has developed a code for aligning the bacterial RNA of E. coli and human and found a unique RNA sequence which contains a stretch of nucleotide that can be targeted with our technology We have designed a library of oligos which is now in the synthesis phase. |
| Type Of Material | Technology assay or reagent |
| Year Produced | 2023 |
| Provided To Others? | No |
| Impact | The alignment has allowed us to find a unique bacterial RNA target for which we have designed the oligo library |
| Title | Differential scanning fluorimetry (DSF) for RNA target disruption in vitro |
| Description | The team has worked on the design and synthesis of a library of OligoY. We have tested the strategy for the bioconjugation of the oligo strands and obtained acceptable yield for the first in vitro tests. A Differential scanning fluorimetry (DSF) which takes advantage of Ribogreen dye binding properties to RNA, has been used to measure the interaction of the oligos with the target RNA (Y RNA1) |
| Type Of Material | Technology assay or reagent |
| Year Produced | 2022 |
| Provided To Others? | No |
| Impact | We are testing in vitro the target engagement allowing for a high throughput assessment of the oligo libraries before proceding to experiment in cells reducing the expenses and accelerating the process. |
| Description | BIO-Europe Spring 2023 |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Industry/Business |
| Results and Impact | BIO-Europe Spring 2023 is Europe's largest partnering conference serving the global biotech industry. The annual international event hosted its edition in the city of Basel from 20 to 23 March 2023, expecting over 1,000 companies and more than 2,000 visitors and key decision-makers from across the world. |
| Year(s) Of Engagement Activity | 2023 |
| URL | https://baselarea.swiss/events/bio-europe-spring/ |
| Description | Embedding natural RNA-protein binding mechanism in RNA-based delivery scaffolds. |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Industry/Business |
| Results and Impact | The inaugural ELRIG meeting on Therapeutic OLIGOs combined with the 8th European Chemical Biology Symposium (ECBS) was held in May 2023 at the AstraZeneca R&D site in Gothenburg. This interactive 3-day event hosted leading scientists from academia, industry and the vendor community to discuss recent advancements in the discovery of therapeutic OLIGOs and in the Chemical Biology research field in Europe and internationally. The ECBS part of the meeting is co-organized by EU-OPENSCREEN and the EuChemS Division of Chemistry in Life Sciences, and includes contributions from key academic chemical biology research groups in Europe. |
| Year(s) Of Engagement Activity | 2023 |
| URL | https://elrig.org/portfolio/elrigs-therapeutic-oligos-european-chemical-biology-symposium-2023/ |
| Description | MRes in Drug Discovery and Development, seminar on current advances and challenges in RNA (or nucleic acid) therapeutics |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | Local |
| Primary Audience | Undergraduate students |
| Results and Impact | MRes in Drug Discovery and Development, delivered a seminar on current advances and challenges in RNA (or nucleic acid) therapeutics. The content included target discovery/validation, case studies of current/novel therapies, delivery technologies, horizon scanning for future solutions, etc. It was a new seminar for the course. It was a 1-2 hours contact time in one session. The course intake is diverse and includes chemists, biochemists, pharmacologists, biologists, etc. so the seminar did not assume in-depth prior knowledge. |
| Year(s) Of Engagement Activity | 2023 |
| Description | MRes student project supervised by the team |
| Form Of Engagement Activity | Participation in an open day or visit at my research institution |
| Part Of Official Scheme? | No |
| Geographic Reach | Local |
| Primary Audience | Undergraduate students |
| Results and Impact | The MRes student was supervised and involved in the listed activities: 1. Assay development: it was be given the opportunity to learn a wide variety of state-of-the-art techniques as well as analytical tools. 2. Multidisciplinary teamwork: exposed to working environment with scientist with wide variety of backgrounds and expertise from core organic chemistry, preclinical sciences, DNA/ RNA nanotechnology to machine learning. 3. Communication: invited to participate, and present, in weekly scientific presentations and bi-weekly journal clubs. 4. Project management: take responsibility for planning and executing the project, and you will receive hands-on project management training and mentorship from day one. 5. Mentorship: given expert guidance and constant mentorship from the lead investigators. This will allow you to troubleshoot quickly so your project is delivered to a high standard. 6. Industry experience: get to experience life at a rapidly growing startup and given the opportunity to see your ideas and research make a real impact on how Sixfold develops its technologies. |
| Year(s) Of Engagement Activity | 2023 |
| Description | Oligonucleotide Therapeutics Society Mentorship Program |
| Form Of Engagement Activity | A formal working group, expert panel or dialogue |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Postgraduate students |
| Results and Impact | The purpose of the Mentor Program is to connect early career scientist in our community with established scientific professionals to support & encourage the mentees' in their career. Once matched, the mentor-mentee pair can take the experience in any direction they desire, including discussion of career goals, personal development, skill enhancement, networking, introductions, and more. |
| Year(s) Of Engagement Activity | 2023,2024 |
| URL | https://www.oligotherapeutics.org/the-mentorship-program/ |
| Description | RNA THERAPEUTICS & DELIVERY EUROPE 2023 |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Industry/Business |
| Results and Impact | RNA leaders, experts and distinguished scientists in Europe, delivering breakthrough research, technologies & connecting global pharma, biotech and academia for high-level discussions on the latest innovations within the formulation and delivery of RNA-based therapeutics. |
| Year(s) Of Engagement Activity | 2023 |
| URL | https://ebrc.org/rna-therapeutics-delivery-europe-2023/ |
| Description | SLAS2024 International Conference and Exhibition |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Industry/Business |
| Results and Impact | SLAS is a global, non-profit professional community made up of scientists from academia, government and industry who are collectively focused on leveraging the power of technology to achieve scientific objectives. The primary part of SLAS mission to unite great minds in life sciences and technology to transform research. |
| Year(s) Of Engagement Activity | 2024 |
| URL | https://www.slas.org/events-calendar/slas2024-international-conference-and-exhibition/ |
| Description | Scaling the Edge |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Industry/Business |
| Results and Impact | This programme helps businesses accelerate product and process innovations using the market leading Strategyzer Toolkit to rapidly validate their ideas with the market. This programme works with multi-disciplinary teams from within existing businesses using Strategyzer's Lean methodology to define clear Value Propositions, winning Business Models and uses modern Digital Testing methodologies to gather rapid and accurate market feedback on the likely success of new business ideas. This process de-risks the chances of market failure, increases speed to market and potential profitability. We have worked on a pitch simulation which will be simulated externally the first week of April, where we will have a video depicting the oligo platform and show results from a Google keyword search volume data. Through the program me and one of my colleagues got access to an in person participation to a prestigious techshow in Basel, the BIO Europe Spring where we will interact with potential collaborators and partners. |
| Year(s) Of Engagement Activity | 2023 |
| URL | https://www.innovateukedge.ukri.org/Scale-with-us |