Release date: 2017-12-01
When Novartis advanced clinical trials of the new drug LMI070 for spinal muscular atrophy (SMA), this small molecule of preclinical data gave them a surprise. LMI070 was discovered by phenotypic screen, but researchers have been unable to find a way to increase the expression of survival motor neuron protein (SMN) for a long time. The absence of SMN is the cause of SMA. The data suggest that LMI070 acts through an unexpected mechanism that binds to RNA targets.
“This surprised me,†said Dr. Rajeev Sivasankaran, head of the rare disease at the Novartis Neuroscience Group. His team found that LMI070 binds to the splicing machinery of the cell and the mRNA precursor (pre-mRNA) produced by the SMN2 gene, thereby regulating exon splicing and enhancing functional protein production. “Everyone is very excited because we have discovered an innovative mechanism that selectively targets the splicing process with small molecules,†he said.
The drug is now in a clinical phase 2 trial of SMA, which represents the frontier of targeting small RNA targets using small molecules. Small molecules can be used to target multiple RNA targets, including RNA complexes, mRNA, and non-coding RNA. Merck (MSD) and Pfizer also independently identified small molecules that target RNA through phenotypic screening, and they are beginning to increase their exploration in this area. The recently established biotechnology companies, including Arrakis Therapeutics and Ribometrix, focus on finding out how to use the fruits of this cutting-edge science to target targets that cannot be used in other ways.
â–² Dr. Matt Disney (Source: Scripps Research Institute)
For Dr. Matt Disney of the Scripps Research Institute in Florida, this represents affirmation of his many years of research. Dr. Disney is a pioneer in the field of small-molecule targeting of RNA targets. He used to publish his own research at scientific conferences and often followed the debates of famous scientists. Because these scientists believe that drugs that selectively target RNA are unlikely to succeed. “Frankly, I have been defeated several times in the process of discussing with them. Because they are scientists I respect very much, they are still the same,†he said. But in September, the first conference on small-molecule-targeted RNA, held at the New York Academy of Sciences (NYAS), attracted more than 200 participants from academia and industry. “One of the things that I am most proud of is that we have finally provided enough data to show that it is possible to target drugs by targeting RNA!â€
Pioneer case of small molecule targeting RNA
A few decades ago, scientists studying infectious diseases knew that small molecules could regulate RNA targets. Aminoglycosides such as gentamicin and streptomycin, which save people's lives, inhibit protein synthesis by inhibiting the function of bacterial ribosomes. Ribosomes are molecular machines constructed from ribosomal RNA and proteins.
The ribosome is a large complex with many cracks and pockets in the structure that allow small molecules to bind to it. The amount of ribosomal RNA expressed is very high and is expressed in all cells, making it easy to be a target for small molecules. These relatively easy RNA targets make it possible to see targeted RNAs that may present unexpected opportunities. For example, Pfizer has inadvertently discovered a small molecule that binds to human ribosomes. It selectively binds to the initial polypeptide chain generated by PCSK9 transcription, thereby preventing the synthesis of PCSK9 protein.
Subsequent studies have found that RNA with less complex structures such as mRNA or non-coding RNA can also be a target. Merck has designed a signaling pathway-based phenotypic screening experiment a few years ago to try to find small molecules that shut down riboflavin synthesis and inhibit bacterial growth. This experiment screened a small molecule called ribocil that binds to a non-coding RNA structural unit called riboswitch, which reduces the translation of bacterial mRNA, reduces riboflavin synthesis, and inhibits cell growth.
Preclinical studies in this project found that bacteria can quickly develop resistance to ribocil, so the study did not continue. But Merck's drug discovery staff is able to optimize small molecule lead compounds for RNA targets almost as much as traditional small molecule screening programs. This insight has allowed Merck to expand its investment in this area.
"We don't just focus on one type of RNA or a disease field, we are exploring this field very widely," said Dr. Noreen Rizvi, a postdoctoral researcher at Merck. She is responsible for assessing the technical feasibility and potential opportunities for targeting different RNA targets. “So far we have identified more than 40 different RNA targets from different biological fields,†she said. “We are developing different techniques to explore the RNA domain and transcriptional subgroups.â€
â–² Dr. Noreen Rizvi (Source: Merck East official website)
When Novartis, Pfizer, and Merck encountered their first small RNA compound that regulates RNA through phenotypic screening, other researchers have begun to adopt a directed discovery strategy. For example, Dr. Disney has developed a screening strategy that screens non-coding RNA libraries and small molecule libraries for small and non-coding RNAs that produce strong interactions. The main goal of this study is to help understand the properties of interactions between small molecules and RNA, and if his findings can be used for medical purposes, it is a icing on the cake.
The study led him to discover a small molecule called targaprimir-96 that binds to the microRNA-96 precursor (pri-miR-96). pri-miR-96 undergoes RNA mutation to generate miR-96, a carcinogenic miRNA that reduces the activity of FOXO1 and plays an important role in causing breast cancer. Targaprimir-96 blocks the production of miR-96 by binding to pri-miR-96, increases FOXO1 activity and can cause apoptosis in tumor cells.
The same strategy led Dr. Disney's team to discover targapremir-210, a small molecule that regulates the production of miR-210, which can be increased in triple-negative breast cancer cells via the hypoxia inducible factor (HIF) signaling pathway. The occurrence of apoptosis.
"This result has finally convinced people to target RNA from a medical point of view," said Dr. Kevin Weeks, an RNA scientist at the University of North Carolina (UNC) and a co-founder of Ribometrix. Say.
â–² Dr. Kevin Weeks (Source: UNC)
Professor Disney is now conducting lead optimization experiments on these compounds, and he is working with biotech companies to try to discover innovative drugs by targeting other RNAs.
RNA target evaluation
Although the existing research results in a proof of principle for small molecule-targeted RNA, there are still many unresolved problems in this field.
One of the important issues is how to classify and evaluate RNA targets. Some RNAs have better drug-making properties, and researchers need to find out which RNAs they should put more energy into. The current preliminary guidelines for RNA selection are beginning to take shape. "If you specifically look for areas of RNA that produce complex structures, they are also more likely to produce special structures that can be targeted," Dr. Weeks said. He has developed a chemical and bioinformatics approach to assess RNA structure.
But small molecules also need to produce biological effects, said Dr. Michael Gilman, CEO of Arrakis. Arrakis completed a $38 million Series A round of financing earlier this year, including support from Pfizer and Celgene. Usually when drug developers discover that small molecules can bind to the active site of a protein, they are more certain that these small molecules will play a role. However, RNA targets usually do not have "active sites". This research area needs to find other reliable methods to predict when small molecules bind to RNA can affect biological function, which may include regulation of RNA folding, mRNA splicing and ribose. Body processing.
The first step in Arrakis' drug development process is the use of bioinformatics to identify RNA targets based on predicted structural features. Because the accuracy of these techniques has not been validated, research teams often perform physical experiments quickly to verify that the predictions are accurate. These experiments are typically adapted from a protein-based drug development project process. “To be honest, we don’t know if these algorithms are correct,†Dr. Gilman said. “This means that the speed at which we screen RNA needs to be much faster than the rate at which proteins are normally screened.â€
Next year the company plans to conduct 1,000 high-throughput screening experiments to find candidate compounds that bind to RNA targets. He added: "We expect that there will be more compounds that bind to the target molecularly, but we don't know how much of it affects biological function."
At the recent NYAS meeting, researchers repeatedly returned to the question of whether it was accumulated and screened for decades and whether the library of peptides used to target proteins was suitable for targeting RNA. Dr. Amanda Hargrove, a chemist at Duke University, tends to use a proven library of existing compounds. Her research team conducted a chemical informatics analysis of 100 RNA-targeting ligands, and concluded that most of these compounds met existing medical chemistry guidelines. Researchers at Novartis, Merck and Arrakis agree. "The compounds we found were not very different from the compounds found in protein screening experiments," said Dr. Rizvi of Merck. This is reassuring because it means that these projects will have fewer risks in terms of solubility, cell permeability and toxicity in future developments.
Dr. Disney does not fully agree. The targaprimir-96 he discovered did not follow the Lipinski's rule of five. This is a set of guidelines that describe the physicochemical properties of a drug and is often used to assess whether a compound can be an oral drug. But there are a lot of drugs approved by the FDA that save the lives of patients and do not meet these five rules. Moreover, since the library of compounds used in screening experiments may be more prone to enrich for compounds that conform to the five rules, the first batch of RNA-targeting compounds discovered by screening also have similar propensities. "We should be very careful not to be biased about the properties of the drug," said Dr. Disney.
Endless possibilities
From the perspective of the medical field, researchers have begun to focus their research on several clear and rational diseases.
Oncology is the first. The inability to treat RAS and MYC proteins in this field has long hampered the development of small molecule drugs that target them, and researchers may have a better chance of success by targeting RNA. Many non-coding RNAs are abnormal in tumor cells, resulting in large changes in protein expression profiles. "We don't need to compete with targeted protein projects," Dr. Weeks concluded. "The hope in the RNA field lies in protein and non-coding RNA that cannot be made into medicine."
â–²Structure of RAS protein (Source: Wikipedia)
The same principle applies to neurological diseases. One research focus here is on mRNAs that are amplified by sequences carrying trinucleotide repeats. Amplification of these sequences results in protein dysfunction. For example, in Huntington disease, CAG amplification occurring on HTT genes and mRNA leads to accumulation of toxic proteins. At the same time, these repetitive sequence amplifications may add more secondary structures to the mRNA that can be targeted, so both Arrakis and Ribometrix use HTT mRNA as their primary research project. Earlier this year, Dr. Disney's team found that small molecules could selectively bind to repeats that cause myotonic dystrophy type 1 (myotonic dystrophy type 1).
Novartis's progress on LMI070 suggests that small molecules can be used to modulate proteins using splicing mechanisms to treat rare genetic diseases.
Infectious diseases are also an important area, especially for bacterial ribosomes and riboswitch. The team led by Dr. Weeks has found potential targets for dengue virus.
But for Dr. Disney, these areas are just the beginning. "The role of RNA in biology is limitless," he said. "I can almost imagine that all diseases are controlled by a certain RNA. "Most of the small molecules that target proteins are antagonists, and their role is to reduce Protein activity. Small molecules that bind to RNA may become versatile drugs. Some may attenuate protein activity by blocking mRNA translation, while others may increase protein activity by shutting down miRNA inhibition.
In some areas, small molecules that target RNA will be complementary to oligonucleotide-based drugs. For example, Ionis Pharmaceuticals and Biogen have received FDA-approved nusinersen to treat the same disease with Novartis's LMI070; Alnylam Pharmaceuticals uses oligonucleotides to reduce protein activity using RNA interference mechanisms; Moderna Therapeutics utilizes Therapeutic therapy for increasing mRNA expression in medical mRNA is undergoing clinical phase 1 trials. However, the advantage of small molecules is that they can be administered orally and can be transported to more organs than oligonucleotides. "I have been cheering on the field of oligonucleotides," Dr. Weeks said. "They provide excellent proof of concept for our research strategy."
Although there are still many challenges in this area, its bright future cannot be ignored, Dr. Weeks said. "In theory, if you can reliably target RNA, you will make a major change in the way you care," he said. "A person who can find a reproducible, scalable mechanism to target RNA will light up." this world!"
Reference materials:
[1] Small pyramid against RNA targets attract big backers
[2] "Nature" official website
Source: WuXi PharmaTech
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