After a half-century pursuit, scientists at the Massachusetts Institute of Technology (MIT) have successfully synthesized verticillin A in a laboratory setting for the first time, unlocking a potent molecule initially identified from fungi over fifty years ago, which has long tantalized researchers with its potential as an anticancer agent. The achievement marks a significant milestone in medicinal chemistry, overcoming formidable synthetic hurdles that had previously rendered this complex natural product inaccessible for extensive study and therapeutic development.
Verticillin A’s notoriety stems from its exceptionally intricate molecular framework, a labyrinth of fused rings and precisely oriented chemical centers that presented a monumental challenge to chemists. Even minor structural variations compared to related compounds dramatically escalated the difficulty of its creation. Professor Mohammad Movassaghi of MIT highlighted this complexity, stating, "We now possess a much deeper understanding of how these subtle structural alterations can profoundly amplify the synthetic challenge." He further elaborated on the implications of this breakthrough: "We now have the capability not only to access these compounds for the first time, more than fifty years after their initial isolation, but also to engineer a multitude of designed variants. This opens the door for more in-depth and targeted investigations."
Preliminary laboratory evaluations utilizing human cancer cell lines revealed a particularly promising activity of one verticillin A derivative against diffuse midline glioma, a challenging pediatric brain cancer. The research team, however, prudently emphasizes that extensive further investigation is imperative to ascertain its clinical viability.
The pivotal study, published in the esteemed Journal of the American Chemical Society, was spearheaded by senior authors Professor Mohammad Movassaghi from MIT and Jun Qi, an associate professor of medicine affiliated with the Dana-Farber Cancer Institute, Boston Children’s Cancer and Blood Disorders Center, and Harvard Medical School. The leading author of the publication is Walker Knauss, a doctoral candidate at MIT. Key contributions also came from Xiuqi Wang, a medicinal chemist and chemical biologist at Dana-Farber, and Mariella Filbin, the research director within the Pediatric Neurology-Oncology Program at Dana-Farber/Boston Children’s Cancer and Blood Disorders Center, underscoring a collaborative, multidisciplinary approach.
The inherent difficulty in producing verticillin A stemmed from its natural origins. First documented in 1970, the molecule was originally discovered in fungi, where it serves as a defense mechanism against microbial invaders. While verticillin A and its fungal relatives have been recognized for their potential antimicrobial and anticancer properties, their complex structures had historically impeded efficient laboratory synthesis, limiting their exploration for therapeutic purposes.
Prior to this breakthrough, a significant step toward understanding verticillin A’s structural intricacies was achieved in 2009 when Movassaghi’s laboratory reported the synthesis of (+)-11,11′-dideoxyverticillin A. This related compound, featuring ten rings and eight stereogenic centers—carbon atoms meticulously bonded to four distinct chemical groups that must be arranged in a precise three-dimensional orientation—represented a substantial synthetic achievement in itself. Nevertheless, the parent compound, verticillin A, remained elusive. The crucial difference between the two molecules lies in the presence of two oxygen atoms in verticillin A. While seemingly minor, these additions profoundly altered the molecule’s reactivity and stability during synthetic processes.
"Those two oxygen atoms significantly constrain the available window for performing chemical transformations," explained Movassaghi. "They render the molecule considerably more fragile and sensitive. Consequently, despite years of methodological advancements in synthesis, this compound continued to present a formidable obstacle."
The synthetic strategy for both verticillin A and its dideoxy analog involves the meticulous joining of two identical molecular subunits to form a dimer. In the earlier synthesis of (+)-11,11′-dideoxyverticillin A, this dimerization step was strategically performed towards the conclusion of the synthetic sequence, followed by the formation of four critical carbon-sulfur bonds. However, attempts to replicate this exact sequence for verticillin A proved unsuccessful. The late-stage introduction of the carbon-sulfur bonds failed to yield the desired stereochemical outcome, necessitating a fundamental reevaluation and redesign of the entire reaction pathway.
"Our learning from this endeavor was that the precise timing of each chemical event is absolutely paramount," Movassaghi emphasized. "We were compelled to substantially alter the order in which the bond-forming reactions were executed."
The revised and ultimately successful synthetic route commences with a derivative of an amino acid, specifically beta-hydroxytryptophan. From this starting point, the researchers progressively constructed the molecule in carefully orchestrated stages. This involved the sequential addition of various chemical functional groups, including alcohols, ketones, and amides, while maintaining rigorous control over the stereochemistry at every juncture. To facilitate this precise stereochemical control, the team ingeniously incorporated a precursor containing two carbon-sulfur bonds and a disulfide linkage early in the synthesis. Given the inherent sensitivity of disulfide bonds, they were temporarily protected by conversion into a disulfide pair. This "masking" strategy ensured the structural integrity of the molecule throughout subsequent chemical transformations. Once the dimerization was complete, the disulfide-containing groups were then regenerated.
"The dimerization process in this synthesis is particularly noteworthy due to the remarkable complexity of the substrates being joined," Movassaghi commented. "These substrates possess an exceptionally dense array of functional groups and stereochemical features."
In total, the refined synthetic pathway comprises sixteen distinct chemical steps, beginning with beta-hydroxytryptophan and culminating in the successful production of verticillin A.
With verticillin A finally within reach, the researchers were also empowered to adapt their synthetic methodology to generate a range of derivatives. A specialized team at the Dana-Farber Cancer Institute undertook the crucial task of evaluating these novel compounds against various forms of diffuse midline glioma (DMG). DMG represents a rare and aggressive form of brain cancer, often characterized by limited therapeutic options and a poor prognosis.
The most striking biological effects were observed in DMG cell lines that exhibit elevated levels of a protein known as EZHIP. This protein plays a significant role in regulating DNA methylation, a fundamental epigenetic mechanism, and has previously been identified as a potential therapeutic target for DMG. Professor Jun Qi elaborated on the significance of this observation: "Identifying the specific targets of these compounds will be instrumental in advancing our comprehension of their mechanisms of action. More importantly, it will enable us to optimize these compounds from the Movassaghi laboratory to achieve greater target specificity, thereby facilitating the development of novel therapeutic strategies."
The verticillin derivatives appear to exert their influence by interacting with EZHIP in a manner that enhances DNA methylation. This epigenetic modification can drive cancer cells toward programmed cell death, a critical process for tumor suppression. The most potent compounds identified in these experiments were N-sulfonylated derivatives of both (+)-11,11′-dideoxyverticillin A and verticillin A. The addition of an N-sulfonyl group, a functional moiety containing sulfur and oxygen, was found to significantly improve the molecular stability of these compounds.
"While the natural product itself may not be the most potent agent, the ability to synthesize it has propelled us to a point where we can now create and study these derivatives," Movassaghi stated.
Looking ahead, the research team at Dana-Farber intends to conduct further rigorous studies to definitively elucidate the precise mechanisms by which these verticillin derivatives operate. Their ultimate goal is to evaluate these promising compounds in preclinical animal models of pediatric brain cancers.
"Natural compounds have historically served as invaluable sources for drug discovery, and we are committed to thoroughly assessing the therapeutic potential of these molecules," Qi affirmed. "This endeavor will involve the synergistic integration of our expertise across chemistry, chemical biology, cancer biology, and clinical patient care. We have also characterized our lead molecules across a broad spectrum of over 800 cancer cell lines, which will provide valuable insights into their functional roles in other cancer types."
The foundational research enabling this significant advancement was generously supported by funding from the National Institute of General Medical Sciences, the Ependymoma Research Foundation, and the Curing Kids Cancer Foundation, highlighting the collaborative spirit and diverse support network essential for pioneering scientific discovery.
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