Date: 8th July 2021
One of the biggest challenges in cancer research is finding suitable targets to direct therapies. Many known cancer-driving molecules are ‘undruggable’ as their type, shape or location prevent drugs from binding to them. Changes in cellular growth and identity during cancer progression are driven by specific gene expression signature, and these are programmed by transcription factors (TFs) and RNA-binding proteins (RBPs). Although cancer-associated TF mutations have been studied for decades, RBPs have been in the main overlooked as drivers of disease and as therapeutically relevant targets. Now, researchers have identify 57 RBP candidates with distinct roles in supporting MYC-driven breast cancer, one of which, YTHDF2 when inhibited triggered apoptosis in triple-negative breast cancer cells and tumours.
RNA-binding proteins (RBPs) are critical regulators of post-transcriptional gene expression, and aberrant RBP-RNA interactions can promote cancer progression. In general, high oncogenic growths rates in cancer cells require increased levels of transcription and global pre-RNA synthesis controlled by TFs, which increases the cell’s dependence on post-transcriptional processing by RBPs. This regulation of processing by RBPs includes splicing, polyadenylation, translation, subcellular localization, and turnover.
One of the primary oncogenic drivers of cancer gene expression in a wide range of cancers is MYC, it has long been viewed as a promising target for anti-cancer drugs however, several features have unfortunately rendered it undruggable. However, orthogonal methods to control other candidates that support MYC-driven oncogenic pathways may prove to be an alternative and viable strategy.
Now, researchers at the University of California, San Diego, US, led by Gene Yeo, have identified 57 RBP candidates that maintained MYC-driven cell survival using a pooled CRISPR-Cas9 screen. Of which one, called YTHDF2, maintained mRNA homeostasis by limiting the number of translating mRNAs. Upon depletion, YTHDF2 triggered apoptosis in triple-negative breast cancer cells and shrank tumours nearly 10-fold.
The team started by screening for RBP dependencies in cancer using a CRISPR-Cas9 lentiviral library containing 10 single-guide RNAs (sgRNAs) for each of 1,078 RBPs, along with other control sgRNAs. The library was then transduced into human mammary epithelial cells (HMECs) expressing a MYC estrogen receptor (ER) fusion (MYC-ER HMECs), when the cells were induced they found 57 RBPs, that when inhibited, led to the death of the cancerous cells.
As MYC-amplified cells typically contain increased quantities of mRNA, candidates that regulate RNA stability and turnover were of particular interest. One stood out to the team, YTHDF2, which they then validated, showing loss of YTHDF2 expression using short hairpin RNAs also caused cell death in MYC-induced HMECs.
To test how safe it might be to treat cancer by inhibiting YTHDF, the team generated inducible multi-tissue knockout mice for YTHDF2. Systematical genetic depletion resulted in no gross physiological abnormalities or changes in body weight for at least 4 weeks after gene knockdown, suggesting that in non-cancerous tissues, depletion of YTHDF2 was safe.
But would YTHDF2 inhibition negatively affects tumour growth in vivo? To answer this, the researchers genetically removed YTHDF2 from an aggressive form of cancer – human triple-negative breast tumour cells, which were then implanted into mice. After initial tumour engraftment they found reduced growth rates and tumours that were ~10 fold in volume smaller than in the control cohort. Upon further analysis, this shrinkage in tumour size was concomitant with fewer proliferating cells, increased apoptosis and reduced host angiogenic vascular endothelial cell markers.
Lastly the team used a new technique that they had recently developed called Surveying Targets by APOBEC-Mediated Profiling (STAMP), a method to assess translation which allowed them to measure previously invisible interactions between RBPs and RNA molecules. This technique revealed that YTHDF2-deficient cancer cells died by stress-induced apoptosis. They found that the deficient cancer cells contained unique translatomes displaying increased translation of tumorigenic and apoptotic transcripts whilst also lacking translation of cell cycle regulators which are normally important for maintaining oncogenic proliferation.
Conclusions and future applications
The team here have discovered a new class of molecules that could act as potential new drug targets for difficult-to-treat breast cancer. By inhibiting RBPs, a previously overlooked family of molecules, it might provide a new approach to cancer treatments, which may extended to other types of cancer. The study revealed that RBP-RNA interactions are selectively essential for the growth and survival of tumour cells, but not somatic tissues, and therefore targeting RBPs, in particular YTHDF2, holds great promise for minimal toxicity and high specificity.
The team will now be investigating RBP vulnerabilities in other MYC-driven cancers derived from different tissue origins to determine whether this is a feature in all of these types of cancer.
Yeo is co-founder of several start-up biotech companies, and first author Jaclyn Einstein will join one company spun out from the Yeo lab to explore YTHDF2’s druggability.
The discovery of a new set of drug targets for aggressive, difficult-to-treat cancer is an exciting one. With an unprecedented plethora of next generation tools now becoming available to target and deliver cancer fighting drugs and therapies aggressive cancers are starting to become far more treatable. We have seen tools developed for the treatment of the aggressive brain cancer, glioblastoma, such as synthetic nanoparticles and immunocytokines. Researchers have discovered and ‘off’ switch for aggressive breast cancer targeted by short hairpin RNA delivered by lentivirus which suppressed tumour growth as well as macro- and micro-metastasis. Novel ionic liquids functioning as drug carriers have induced local tumour ablation in the liver, and living biotherapeutics are also being explored for fighting cancer. With such a rich wealth of tools available, it should be possible to target RBPs and advance treatments for MYC-driven cancers in the near future.
For more information please see the press release at UC San Diego
Einstein, J.M., Perelis, M., Chaim, I.A., Meena, J.K., Nussbacher, J.K., Tankka, A.T., Yee, B.A., Li, H., Madrigal, A.A., Neill, N.J., et al. (2021). Inhibition of YTHDF2 triggers proteotoxic cell death in MYC-driven breast cancer. Molecular Cell.
https://doi.org/10.1016/j.molcel.2021.06.014
