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Innovative assay for microRNAs analysis

Periodic Reporting for period 1 - LIVELMIA (Innovative assay for microRNAs analysis)

Reporting period: 2019-09-01 to 2020-02-29

miRNA (microRNA) is a recently discovered type of biomolecules implicated in numerous biological systems and in common human diseases (e.g. cancer, aging and degenerative diseases, etc.). In the past decade, the research on miRNAs has soared with over 60,000 papers deposited on PubMed related to the use of miRNA. To give an example of the value of miRNA analysis, it has recently been shown that classification of tumour subtypes by miRNA is more accurate than the gold standard classification by protein . The following is a small list outlining the value and importance of miRNA analysis in medicine:
• Used or believed to act as a biomarker to detect and quantify the progression of many diseases including but not limited to Cancer, Heart Diseases, Kidney Diseases, Nervous system, etc.
• Serve as early biomarkers for the evaluation of drug efficacy and drug safety
• Aid in the establishment of anti-viral drugs and/or vaccines for a number of viruses such as the herpesvirus.
While the value of miRNA analysis is clear, miRNA is extremely difficult to analyse. To date, the three major profiling methods are qPCR (RT-PCR), Microarray and NextGen Sequencing (NGS), however each method suffers from limitations thereby limiting its use to research only. The following list outlines the customer pain and therefore the key requirements in miRNA analysis:
• Analysis directly on the sample – The current solutions are unable to work directly on the sample, which, in medicine, are mostly plasma, serum and cells. So before performing assays current solutions require an extra process to extract/purify and prepare miRNA. This procedure is complicated by the ubiquitous presence of ribonuclease enzymes in cells and tissues, which can rapidly degrade microRNA. Extraction/Purification is also needed because current solutions are easily influenced by sample contaminants (i.e. proteins, salts).
• Throughput – Current solution require too much time for a single analysis, thereby limiting their throughput: qPCR – 6 hours for a single analysis including 3 hours of hands-on work by an expert; Microarray - 2 days for a single analysis including 6 hours of hands-on work by an expert; NGS - 2 days to 2 weeks for a single analysis including 6 hours of hands-on work by an expert.
• Specificity – relates to the ability for an analysis to provide accurate results over a range of differing samples. All of the three major analytic methods available today require an extraction/purification stage which is a major source for introducing errors thereby reducing specificity. In fact some studies have found that this step can introduce unexpected variation, more so than the analytical method itself (eg. qPCR) , this increases the risk of having to repeat the analysis for statistically valid data and decreases the reproducibility.
The culmination of these pitfalls above result in higher costs, lower availability and to an extent lower confidence on the ability for miRNA to act as a stable bio-marker in its various uses.
The objective of Phase 1 will be to determine the feasibility, viability and profitability of the product on our company, and to identify the risks and the best strategies for commercialisation. We also aim to point out functional and organisational gaps that need to be addressed and to set up mitigation actions. Our objectives and the activities we plan to undertake in this feasibility study are aligned and consistent with the expected impact of the project in several ways.
Bioridis was set-up to solve the customer pain identified above. Our CEO and Scientific Director Andrea Tortori Ph.D has studied the use of miRNA as a biomarker in Alziehmer’s Disease and immediately saw the value of miRNA but was aware of the current pitfalls in current analytical techniques and set on a mission to develop a solution that is easy to use, cost effective and provides the necessary properties for its use in medical applications. LIVELMIA is the result of 3 years of high-risk research that has resulted in an innovative product with a very high-potential which we have patented in 2018 and we are now looking to commercialize. We have studied the market very well and we are aware that no other company has anything that can compete on the level of LIVELMIA in terms of throughput and cost effectiveness. In our development, we made sure that our product is able to deliver a result in less than 50% of the time the competition requires, works directly on the sample and does not require the use of expensive equipment for long periods and has a very high specificity, a key requirement for the use of miRNA as a biomarker in diagnostic medicine today. LIVELMIA is an affordable and easy to use kit that cuts down the total time to 3 hours. This allows multiple analyses per work day (of 8 hours) with the same personnel and instrumentation, thus allowing to reduce waiting lists for laboratory analysis and get results quickly for health research and diagnostics without expensive investments. For example, with LIVELMIA it is possible to run 2 cycles of analysis for instrument/day, instead with RT-PCR it is possible to run 1 analysis cycle for the instrument/day.
LIVELMIA is supplied to our customers in a kit. The kit is made up of a number of sample holders (called wells) in which we anchor our probe molecules that capture the target miRNA in the sample. Once a sample (blood, plasma, cells) are placed in the well, the users adds our labelling buffer. The labelling buffer translates the level of miRNA in the sample into a change in light absorption of certain colour. This is then measured, instantly, using a spectrophotometer which is standard equipment which any lab would have.
Our Novelty - The novelty of the LIVELMIA assay is based on the design of our probes which improves the recognition of miRNAs. These are based on a new analogue of nucleic acids accompanies by a specific patented procedure. The components that make up LIVELMIA are:
a) Probes designed for high specificity and sensitivity for targeting of specific miRNAs with a signalling system
b) Optimized reagents (buffer solutions) and analytical procedure to reduce overall time and cost;
c) A platform for rapid design of kits targeting new miRNAs of clinical interests.
Overall, the unique features of LIVELMIA assay are ideal for miRNAs analysis. Our innovation, when compared to the competitors:
• Saves time/ease of use: No sample extraction/purification and preparation, analysis directly on the sample;
• Provides high biological and chemical stability: LIVELMIA’s components do not degrade in a biological sample
• LIVELMIA is the only method that directly protects miRNAs from degradation by sample components;
• High affinity and specificity: increased reliability, consistency, accuracy of miRNA analysis thanks to approximately 25% greater affinity and specificity for the target;
• Easy standardization: Works efficiently in different environments and difficult sample types (living cells, blood, plasma, etc).