May 30, 2024

Tumor-specific proteins and antigens have long been considered ideal targets for cancer treatment, and the medical community has developed a large number of targeted drugs for them. Unfortunately, the expression of some targets in non-tumor tissues limits their effectiveness, while highly tumor-specific targets are often difficult to develop into drugs due to their intracellular distribution. How to ensure specificity while ensuring drug efficacy is the most concerned issue in the field of cancer treatment. Antibody-drug conjugates and nuclear drugs have rapidly gained favor in the pharmaceutical field due to their extremely high specificity and strong killing power. However, in vivo, a phenomenon called "synthetic lethality" has gradually begun to emerge in the field.

"Synthetic lethality" refers to the phenomenon that when two genes are mutated or inactivated at the same time, it will lead to cell death. Mutation of either gene alone will not affect cell survival. If we can find similar synthetic lethal genes in tumor cells, we can kill tumor cells with extremely high specificity.

On April 24, 2024, Vividion Therapeutics and Novartis published papers in Nature titled "Chemoproteomic discovery of a covalent allosteric inhibitor of WRN helicase" and "Discovery of WRN inhibitor HRO761 with synthetic lethality in MSI cancers," respectively, reporting their latest progress on WRN helicase synthetic lethal drugs.

Challenge the Impossible

There are a large number of specific structures and proteins in tumor cells, but due to various limitations, we cannot develop targets targeting them. They are also called "undruggable targets." Vividion Therapeutics is a company that challenges the impossible. Through its unique chemical proteomics platform, Vividion's scientists can study the interactions between small molecules and proteins with extremely high resolution. By finding specific cysteine binding sites, Vividion has successfully found a series of small molecule drug candidates that can target undruggable targets.

As a DNA helicase, WRN has been previously found to be synthetic lethal with microsatellite instability (MSI) tumor cells. Through its chemical proteomics technology, Vividion found 11 potential druggable cysteine sites and developed VVD-109063, a small molecule that can covalently bind to the C727 site, with an IC50 of 8μM. However, in subsequent cell experiments, VVD-109063 performed poorly, and researchers found that this was due to the high ATP concentration in living cells. Through molecular structure optimization, Vividion's scientists finally obtained VVD-133214, which has strong WRN inhibitory activity at the cellular level.

Action Mechanism of VVD-133214

Subsequently, Vividion's scientists studied the mechanism of action of VVD-133214. Researchers analyzed the crystal structures of the WRN ATPase domain after binding to ADP, ATP, and VVD-133214+ADP, and found that VVD-133214 can weakly bind to the hydrophobic pocket between RecA_1 and RecA_2 in the ATPase domain. Although this weak binding is reversible, it is sufficient to cause a large conformational change, making WRN form an extremely stable conformation. Researchers speculate that this stable structure makes WRN lose its original helicase activity, which is the origin of the inhibitory effect of VVD-133214.

In Vitro Activity of VVD-133214

After finding the mechanism of action of the drug, the next step is naturally to study its efficacy.

In MSI-carrying HHCT-116 cells, Vividion's scientists studied the effect of VVD-133214 on cell growth. Compared with microsatellite stable (MSS) cells, VVD-133214 can greatly reduce cell growth in MSI, while in HHCT-116 cells carrying drug-resistant mutations, VVD-133214 completely loses the effect of inhibiting cell growth. The vast majority of MSI tumor cell lines are sensitive to VVD-133214, while all MSS cells are insensitive to VVD-133214. Only 24 hours of incubation can achieve a long-lasting inhibitory effect, and removing the culture medium containing the drug after incubation does not restore cell growth.

MSI tumor cells rely on WRN to replicate after replication of unstable damaged DNA. Morphological observations revealed that after VVD-133214 treatment, the nucleus tends to enlarge, which is similar to the morphological changes observed in previously reported WRN synthetic lethal phenomena. In MSI cells treated with VVD-133214, researchers also found an upregulation of DNA damage markers, causing cells to arrest in G2 phase and eventually undergo apoptosis. These data demonstrate that VVD-133214 leads to tumor cell death by suppressing WRN activity, resulting in DNA breaks and chromosomal integrity defects. In addition, VVD-133214 not only inhibits WRN activity but also significantly reduces WRN levels, which researchers believe is one of the results of DNA damage.

The above results demonstrate the in vitro specificity and efficacy of VVD-133214 as a synthetic lethal drug.

In Vivo Performance of VVD-133214

Finally, Vividion's scientists used mouse models to confirm the in vivo antitumor efficacy of VVD-133214. An oral dose of only 5 mg/kg daily achieved a tumor penetration rate of 95% in animal models and maintained a good blood level for 8 hours. In mouse tumors, researchers observed a significant decrease in WRN levels, but not in WRN-rich organs such as the pancreas. In terms of specificity and efficacy, the in vivo performance of VVD-133214 was consistent with in vitro experiments.

Subsequently, researchers investigated the antitumor activity of VVD-133214 in mouse models. Compared with the control group, VVD-133214 exhibited strong tumor suppressive effect in mouse models of MSI tumor cells. VVD-133214 failed to inhibit tumor growth in MSS and WRN action site mutant tumor models. This was also reproduced in PDX models based on human tumor tissues. More importantly, VVD-133214 was able to exert the same strong tumor suppressive effect in tumor tissues of patients who had undergone chemotherapy or immunotherapy, suggesting that VVD-133214 has broad application prospects.

The synthetic lethal approach targeting WRN was further validated in another Nature paper published in the same issue. Novartis's WRN inhibitor HRO761 showed similar results.

Genscript is honored to contribute to the conquest of "undruggable" targets. The study used Genscript's BacuVance baculovirus-insect protein expression service.

Reference

[1] Baltgalvis, K.A., Lamb, K.N., Symons, K.T. et al. Chemoproteomic discovery of a covalent allosteric inhibitor of WRN helicase. Nature (2024). https://doi.org/10.1038/s41586-024-07318-y

[2] Ferretti, S., Hamon, J., de Kanter, R. et al. Discovery of WRN inhibitor HRO761 with synthetic lethality in MSI cancers. Nature (2024). https://doi.org/10.1038/s41586-024-07350-y