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In the late 1980s, Brian Bingham was still a Ph.D. He carries his bags to the Florida Keys every month. Among the mangroves that surround the island, there is the sea squirts that the marine biologist has been studying, and it is also the best place to study the sea squirt.

Sea squirts can multiply and grow on the roots of mangroves swaying in sea water. Bingham carefully marked the roots of the trees with fluorescent pink bands so that he could still find them the next time he came. Every month, he photographs the roots of the trees and records changes in the sea squirt population in the area.

One day, however, when Bingham docked the boat on the shore, he found that a group of divers were busy salvaging the sea squirt and putting them in the bag.

Bingham was puzzled. At the time, there were not many people who knew the creature of the Ecteinascidia turbinate. Why are there so many people now collecting this sea squirt in the mangroves?


Birth of Trabexidine

In the 1950s and 1960s, the National Cancer Institute conducted a series of screening projects to screen the biological activities of plants and marine organisms, and the biological activity screening of Caribbean sea squirt extracts was also part of the project.

In 1969, scientists first discovered that crude ethanol extracts from the Caribbean sea squirt have antitumor activity. However, due to the limitations of the conditions at the time, the separation and structural identification of ecteinascidins had to wait until many years later.

In 1990, KL Rinehart Laboratories at the University of Illinois isolated six ascidins from the Caribbean sea squirt, and one of the ascidin Trabexidine had the highest extraction yield (0.0001%).


Subsequent studies have found that the mechanism of action of Trabexidine is unique. Unlike paclitaxel and eribulin inhibiting the action of microtubules, trastidine can block cell division by binding to DNA: two fused tetrahydroisoquinoline rings can interact with specific sequences of DNA minor groove Covalently binds to it, causing abnormalities in the double helix of DNA and interfering with the normal function of XPG protein in the TC-NER system, thereby inducing DNA double-strand breaks and leading to cell death.

Trebexidine binds to DNA-XPG complex

Spanish pharmaceutical company PharmaMar purchased the patent for trebutidine from the University of Illinois in about 1994. They then began to try to extract the Trabexidine from the cultured sea squirt. PharmaMar has a 250-ton Caribbean sea squirt, but due to the complexity of the separation and purification process, the yield of this extraction method is extremely low, and only one gram of Trabexidine is extracted per ton of Caribbean sea squirt.

Although the yield is extremely low, the amount of Trabexidine required for preclinical studies is not large, so the extraction of Trabexidine from cultured sea squirts can also meet the needs of the time. However, in 1997, as PharmaMar decided to advance trebutidine to clinical research, they had to carry out larger sea squirt cultures to ensure adequate supply of compounds.

At the same time, Rinehart is also considering how to fully synthesize the compound, so he asked Harvard chemist E. J. Corey for help.

In addition to the unique mechanism of action, the complex meridional molecular structure has been fascinated by chemists. In 1994, Corey asked his laboratory’s postdoctoral fellow David Gin to try a full synthesis study. Two years later, Gin completed the work.

Corey once said that the success of the synthesis of trometidine is a feat, and this research has also given Gin unlimited glory in the academic field.

This fully synthetic strategy was published in JACS in 1996. This synthetic route divides the chemical structure of Trabexidine into four fragments and uses the Mannich reaction to construct the skeleton structure of the compound. All synthetic routes included 36 steps with a total yield of 0.72%.

Analysis of the inverse synthetic route of Trabexidine

Harvard University then sold the fully synthetic patent to PharmaMar. For PharmaMar, successful clinical development without full synthesis will be extremely difficult.

Preclinical animal model experiments showed that Trabexidine had extensive antitumor activity. However, the researchers did not find the sensitivity of soft tissue sarcoma to Trabexidine in preclinical studies. In the clinical phase I study, the researchers found that patients with soft tissue sarcoma were able to respond or stabilize the disease after using Trabexidine, and the clinical phase II study once again confirmed the effectiveness of Trabexidine in the treatment of soft tissue sarcoma.

But by the time of the Phase III clinical study, PharmaMar once again faced pressure from the supply of compounds. Although the full synthetic route provided by Coley Laboratories provides sufficient medicine to complete early clinical development, the synthetic route is difficult to scale up, even if the supply of Phase III clinical requirements is difficult, not to mention industrial production after the product goes on the market. . So PharmaMar had to look for other drug synthesis options.

In 1999, PharmaMar’s team of scientists, Carmen Cuevas, found an efficient compound synthesis solution. They used the biofermentation of Pseudomonas fluorescens to produce the antibiotic safracin B, which possesses a fused ring structure similar to that of ectoin.

The use of safracin B as a starting material greatly simplifies the Corey synthesis route. Starting from safracin B, the final product, trimetine, was obtained after 21 steps of reaction with a total yield of 0.96%.


Difficult industrial production

The semi-synthesis strategy based on safracin B temporarily solved the obstacles encountered in the amplification synthesis of Trabexidine, paving the way for product launch.

In 2007, Yondelis was approved by the European Union for the treatment of soft tissue sarcomas. The incidence of soft tissue sarcoma is about 1.28/100,000-1.72/100,000, accounting for 0.73%-0.81% of all adult malignant tumors, and 6.5% of all malignant tumors in children younger than 15 years old.

Surgery is the only curetable and most important treatment for soft tissue sarcoma, but there are still many sarcoma patients who need chemotherapy. For example, patients with high-grade soft tissue sarcoma have a metastasis of 10% at the time of initial diagnosis, and even if the local control of the tumor is good, 40% – 50% of patients will have local recurrence after surgery, and more than 50% of patients will have distant Transfer.

The market for Trabexidine is a new treatment option for patients with anthracycline and IFO drug failure, as well as patients with advanced soft tissue sarcoma who are not suitable for these drugs.

In 2015, trometidine was approved by the FDA for the treatment of patients with unresectable or metastatic liposarcoma or leiomyosarcoma who had received an anthracycline treatment. In addition to soft tissue sarcoma, trebutidine has been approved by regulatory agencies in Europe, Canada and other regions for the treatment of platinum-sensitive recurrent ovarian cancer.

Although the Cuevas team temporarily solved the problem of product amplification and synthesis through semi-synthesis strategy before the listing of trobe, the industrial production of trebutidine still faces many obstacles after the product launch, and there are still many other products including quality control. Improve the technical difficulties of solving. This is also an important reason for the annual production of the trebutidine to remain at a very low level.

In addition, safracin B has a low fermentation unit and poor stability of the product itself. Only a few hundred grams of safracin B can be extracted per ton of fermentation broth, and the subsequent semi-synthetic reaction route still exceeds 20 steps. The overall synthesis yield is less than 1 %, so the semi-synthetic strategy also has a lot of optimization space. How to improve the total yield, shorten the reaction route, reduce the protection base and the use of toxic reagents such as n-Bu3SnH is still a problem to be solved in the process of industrial production exploration.

The patent for trometidine has long expired (WO1987007610). In fact, whether it is soft tissue sarcoma or ovarian cancer, Trabexidine has a large market in the world. However, for such a drug with extremely complicated chemical structure, even if there are still many obstacles in the industrial production of the original research company, there are not many pharmaceutical companies that have the ability to overcome many difficulties and complete the industrial production of drugs.


Borui Biomedical is one of the few pharmaceutical companies in the world to explore and overcome industrial production barriers in the world, and will soon submit the DMF (drug master file).


Many natural products derived from the ocean have very novel chemical structures and unique pharmacological mechanisms, but the extraction of natural products to obtain the compounds required for the drug development process is often very inefficient. For these complex natural products, the total synthesis or semi-synthesis of the compound is likely to be a key factor restricting the development of related drugs.


However, for a complex drug, even after the full synthetic route is opened, industrial production after the drug is marketed often needs to overcome a lot of technical obstacles. It is not easy for pharmaceutical companies to be able to complete the industrial production of drugs with such complex structures.

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