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Since RNAi (RNA interference) was first discovered 20 years ago, researchers from both academic institutions and pharmaceutical companies have been hoping to develop this technology into new therapies that can treat diseases, but so far there is no one. Related drugs are listed.


Fortunately, however, Alnylam Pharmaceuticals, a pioneer in the RNAi field in December 2017, filed a patisiran listing application with the FDA and EMA for the treatment of a rare, potentially fatal disease: hereditary transthyretin amyloidosis ( Hereditary transthyretin-mediated (ATTR) amyloidosis).


The launch of the drug will be a landmark victory in RNAi’s decade-long exploration journey. In fact, RNAi drugs are very different from traditional drugs. If we want to understand the mechanism of RNAi, we must also talk about nematodes.


The secret of nematodes

Genetic information in DNA can be copied in the form of mRNA and used to guide the synthesis of proteins. In other words, the genetic information in DNA determines the structure of the protein. The flow of genetic information from DNA to RNA to proteins is called the central law (here the traditional definition). Proteins are found in almost all life processes, such as enzymes that digest food, receptors that receive signals in the brain, and antibodies that fight bacterial infections.

This basic rule is very conservative in the evolutionary process and follows the same laws from bacteria to humans. However, in the 1990s, molecular biologists observed a series of unexplained phenomena during the experiment. One of the most famous is an experiment conducted by a plant biologist.

Botanists hope to deepen petal color of Petunia flowers by artificially introducing genes that produce red pigments, but they found that this operation did not make the color of the petals deeper, but instead it completely disappeared and turned white. No one at the time knew the reason for this phenomenon. And Andrew Fire and Craig Mello also discovered similar weird phenomena when studying nematodes.

Fire and Mello have been studying how the genes of Caenorhabditis elegans are regulated. They found that the nematode did not produce obvious behavioral changes after injection of mRNA encoding muscle proteins. But after the injection of antisense mRNA that can pair with the sense sequence (we generally refer to the sequence in the mRNA as the sense sequence), the nematode also does not produce obvious responses.

But Fire and Mello discovered that if the injection contained both the sense and antisense strands of mRNA, they discovered that the nematode showed a strange, distorted state of motion. And, interestingly, nematodes that are completely deficient in this gene also exhibit the same state. In the end what happened?

We know that when the sense and antisense RNA molecules meet, they will combine to form a double-stranded RNA. Is this double-stranded RNA molecule silencing the corresponding DNA? To test this hypothesis, Fire and Mello injected double-stranded RNA into the nematode. They found that the injection of multiple double-stranded RNAs can silence genes that contain corresponding genetic information, which means that these gene codes cannot guide protein production.

On February 19, 1998, they published their research on Nature, and the publication of this paper also marked the birth of a new research area of ​​RNA interference.

In the following years, as the research progressed, the detailed mechanism of RNAi gradually became clear: double-stranded RNA could bind Dicer and cut it into fragments and bind to another protein complex RISC. One strand of the double-stranded RNA is then removed, leaving only one strand to bind to the RISC and used to detect mRNA that can pair with it. When the mRNA is successfully paired with the RNA fragment on the RISC, the mRNA binds to the RISC complex and is cleaved and degraded by the complex. In this way, the gene corresponding to the mRNA is also silenced.

RNAi is an important defense method for organisms against viruses and foreign genes. It can also participate in the regulation of the expression of organisms’ own genes. Since its birth, RNAi has been developed as a powerful molecular tool that can control gene expression. However, the power of RNAi is far more than this, and it also has a very broad application prospect in the field of medicine.

Many diseases in humans are caused by abnormal protein accumulation in the structure or function. Many traditional medicines exert their effects by affecting the function of proteins, such as enzyme inhibitors that affect catalytic activity. RNAi drugs may directly inhibit the production of abnormal proteins by silencing related genes.

Rise and fall

Given the powerful power of RNAi, many people hope that these drugs will become the next breakthrough in the pharmaceutical industry. In 2002, after completing the analysis of the basic mechanism of RNAi, some biotechnology companies focused on the field of RNAi began to appear. These startups quickly captured the enthusiasm and imagination of investors and gained a lot of capital.

When Alnylam was founded, there were only a few researchers but there were numerous patent lawyers. This was actually because the RNAi technology was so hot at that time that they wanted to apply for as many patents as possible. It was an era full of bubbles. It seems that most people have an irrational passion for RNAi.

After that, large pharmaceutical companies entered the market and began to collect the area where the madness and hope coexist. In 2005, Novartis purchased the rights to use the Alnylam technology platform for hundreds of millions of dollars. Alnylam’s main competitor, Sirna Therapeutics, has also reached various agreements with GSK and other companies.

In 2006, Fire and Mello won the Nobel Prize for their contributions in the field of RNAi. What followed was another brutal growth in the field. Venture capital has long since flooded into this crowded area, and the funds of large pharmaceutical companies have flooded. That’s when Merck acquired Sirna for $1.1 billion.

These news made many people excited. They think that since there is so much money flowing into this area, it means that the prospect of RNAi is bright and there will be a drug market in the near future. At that time, many people did not know what they were excited about and where the future of this technology was.

Maybe reality is always crueler than you think. It soon became clear that RNAi drugs still had a long way to go before they were launched. There are still many problems that have not yet been properly solved, such as how accurately these drugs can be delivered to target cells.

Many of the earlier clinical trials were actually unreasonable. Some pharmaceutical companies pushed drugs into clinical studies without perfect preclinical research in order to compete for the title of “the first clinical entry.” Moreover, some drugs cause more dangerous side effects in clinical trials, and these side effects have not been found in preclinical animal experiments. It is clear that this type of drug cannot be easily delivered to target cells, which causes great problems in the efficacy and side effects of the drug.

Perhaps because of the systemic delivery of these drugs, the enthusiasm of the pharmaceutical industry for RNAi is fading. The subsequent announcement of the results of clinical studies has brought a heavy blow to the field. Clinical trials of naked RNAi drugs to treat ocular diseases such as age-related macular degeneration have all failed, and lipid nanoparticles (LNPs) for systemic administration can entrap RNA to prevent the drug from being degraded before reaching the target, but this approach There is still a need for extremely high drug doses to produce weak effects, and these drugs are only effective for liver-related diseases. RNAi drugs have no effect on the respiratory system and cardiovascular diseases.

With the release of clinical trial results, large pharmaceutical companies have gradually begun to leave. At the end of 2010, Novartis and Roche stopped cooperation with Alnylam and Alnylam was forced to live on the verge of death. For the field of RNAi, this is probably the critical moment of life and death.

Even if Merck has been publicly claiming that the company still retains enthusiasm for RNAi in the following years, the public R&D pipeline has never made any progress in RNAi therapy projects. Eventually in 2014 Merck sold Sirna’s IP to Alnylam at a one-sixth of the initial purchase price.

At the time, Alnylam, struggling on the death line, had a dedication that was incompatible with the corporate culture of large pharmaceutical companies. But in fact, Alnylam has the ability to insist that the support and cooperation with Sanofi can not be separated. In 2012 Alnylam and Sanofi first reached an agreement. In 2014, Sanofi acquired a 12% stake in Alnylam for US$700 million, and acquired some of the company’s proprietary rights outside North America and Western Europe. In January this year, the two sides also conducted a reorganization of the transaction. Alnylam obtained worldwide ownership of patisiran and ALN-TTRsc02, and Sanofi acquired the right to global commercialization of the drug fitusiran, a phase III clinical drug.


With the soon-to-be approved first RNAi drug, the biopharmaceutical industry is also recognizing this technology. For example, in 2016, Amgen reached an agreement with Arrowhead Pharmaceuticals (the company’s acquisition of Roche’s RNAi project in 2011) to conduct research on cardiovascular diseases with a transaction volume of up to 675 million US dollars. In 2017, Boehringer also cooperated with Dicerna to jointly conduct drug research in the field of nonalcoholic fatty liver disease and other chronic liver diseases.

In addition to the re-entry of capital, the technology in this area is also rapidly iterating. Patisiran uses LNP drug delivery via intravenous administration, but now Alnylam and other RNA-related companies have completely abandoned this type of drug delivery. The state-of-the-art technology is probably linked by RNAi and N-acetylgalactosamine (GalNAc) or similar ligands, giving the drug a better therapeutic index, allowing subcutaneous administration, increasing doses and reducing side effects.

Partial use of GalNAc-coupled RNAi technology has entered Phase III clinical studies. These drugs are more likely to enter the liver and bind to receptors in the liver. Even with statins, there are more than $50 billion in liver-related disease. In other words, even if RNAi drugs can only be applied to liver-related diseases, the market for this drug will not be too small. However, it is still very challenging to apply RNAi to other therapeutic areas.

In fact, there are many potential drug targets in the liver. For these targets, they show strong drug-like properties compared to the antibody class RNAi. Clinical trials at this stage have also confirmed that the technology is relatively safe, the therapeutic index is relatively high, and the duration of efficacy is long. In fact, the patisrian achieved all primary and secondary endpoints in its pivotal phase III clinical trial. After 18 months of drug injection, some patients experienced some common mild or moderate side effects, including injection site reactions. Peripheral edema. Furthermore, the clinical trial results of the GalNAc-conjugated drug ALN-TTRsc02 should theoretically be superior to the patisrian.

But for practitioners in the pharmaceutical industry, the application of RNAi in liver disease is likely to be just the beginning. Future RNAi may also be successful in the treatment of other types of diseases, such as cancer, peripheral nervous system diseases, etc. Some pharmaceutical companies are also trying to use naked RNAi to treat kidney and eye diseases.

And new drug delivery technology may be able to make RNAi drugs even higher. For example, Codiak BioSciences is trying to use exosomes to deliver oligonucleotide drugs. LNP is a synthetic lipid carrier, and exosomes are naturally occurring substances in organisms. Codiak’s exosome-based drugs are expected to enter clinical research later this year. It is worth noting that this drug is a RNAi targeting KRAS. In addition, Arrowhead is using another new technology. They are developing RNAi inhalers to make them work in the lungs.

Exosomes seem promising, and some large pharmaceutical companies and biotech companies are using exosomes to solve the delivery problems of other types of drugs. Because RNAi is also only a branch of generalized oligonucleotide technology, this technology also includes antisense and exon skip drugs, virus-mediated gene therapy (see: Thirty years of gene therapy listed Lu), mRNA-based drugs, and CRISPR technology that can be genetically edited (see: Why CRISPR Must Have Nominees?). The development of these technologies and the drug delivery system used by them can also promote the further development of RNAi drugs.

In fact, pharmaceutical companies have also achieved some success in the field of oligonucleotide drugs. For example, Ionis Pharmaceuticals/Sanofi’s high-cholesterol treatment drug mipomersen has been approved for listing. But even so, there are still numerous unsolved problems in this area.

The development process of science and technology is always full of hardships and hardships. Now there are many pharmaceutical companies that make monoclonal antibodies in China, but how many people really understand how many drug-resistant drugs have gone from lab to pharmacy shelves for 20 years? Tribulation. Our understanding of the life sciences continues to deepen, and new developments in the pharmaceutical field will follow, such as CAR-T, PROTAC, and Waston, which uses artificial intelligence. When faced with such new technologies, we always unconsciously generate some unrealistic illusions that the tumors will be cured soon and that artificial intelligence can replace doctors.

But the reality is always cruel. Many times we cannot predict what kind of difficulties we will encounter and what setbacks we will encounter. It’s very difficult to make new drugs. It’s really hard because we’ve been fighting against such exquisite organisms. I don’t know if such a complex human body structure has evolved billions of years ago, or it was created by God. I only know that as a practitioner in the pharmaceutical industry, in the face of such a complex system, there should be more awe. After all, it’s not easy to succeed.

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