The study, published in Nature Communications, provides important new insight into how tumour metabolism influences drug action and could accelerate the development of more precise cancer treatments.
Schematic showing how the CBH-002 probe detects different types of PRMT5 inhibitors and senses changes in cellular metabolic state of SAM. Credit: Rothweiler et al., Nat Comm 2025
The work centres on PRMT5, a key enzyme that regulates gene expression and is an important target in oncology. Around 10-15% of human cancers lack a metabolic gene called MTAP, causing a molecule known as MTA to accumulate within tumour cells. This altered metabolic landscape changes the way PRMT5 responds to potential drugs and creates a therapeutic vulnerability that researchers hope to exploit.
Until now, tools to directly quantify how PRMT5 inhibitors behave in the distinct metabolic environments of tumour versus normal cells have been lacking. Existing approaches provided only indirect readouts or relied on studying purified proteins outside their natural cellular context.
The University of Oxford team designed and developed CBH-002, a cell-permeable small molecule BRET probe that binds to a genetically encoded PRMT5-NanoLuc biosensor to report drug target engagement in live cells.
Dr Elisabeth Mira Rothweiler with NanoBRET assay plates used to measure PRMT5 inhibitor binding in live cancer cells at Oxford's Centre for Medicines Discovery
Dr Elisabeth Mira Rothweiler, Postdoctoral Researcher, Centre for Medicines Discovery, University of Oxford, and co-first author, says: "CBH-002 could measure various PRMT5 inhibitor types in live cells, prompting us to test its sensitivity to the cofactor SAM. When we discovered the probe's ability to sense metabolite levels, it established its utility as a metabolic biosensor. Working with Promega, we showed how MTA influences drug selectivity, revealing why certain inhibitors are so effective in MTAP-deleted cancers.”
In a controlled pair of colorectal cancer cell lines that differed only in MTAP status, the team found that next-generation MTA-uncompetitive inhibitors bound PRMT5 orders of magnitude more tightly in MTAP-deleted cells than in normal cells. This dramatic shift in affinity mirrors previously reported selective anti-proliferative activity in MTAP-deleted cancer models.
Professor Kilian Huber, co-senior author of the study, said: “Our biosensor lets us examine, in living cells, how different PRMT5 inhibitors behave under the specific metabolic conditions that make some tumours uniquely vulnerable. This provides unprecedented insight into why certain inhibitors are much more effective in cancers lacking MTAP, and paves the way for highly targeted cancer treatment in the future. It’s like turning on the lights inside the cell so we can finally see which key actually fits the lock.”
Using CBH-002, the researchers characterised several classes of PRMT5 inhibitors. The biosensor distinguished between different modes of drug binding - whether inhibitors compete with natural metabolites, require their presence, or behave differently when MTA accumulates.
Understanding how a cell’s metabolic state affects drug engagement is essential for the development of precision medicines. The approach established in this study may guide the design and optimisation of future PRMT5 inhibitors specifically tailored to MTAP-deleted tumours.
Ani Michaud, Sr Research Scientist at Promega, and co-first author of the Nature Communications paper said: “The methods in this study enable us to characterise inhibitors that bind much more tightly in tumour cells with MTAP deletions. To our knowledge, this is the first time anyone has characterised this type of uncompetitive inhibitor mechanism directly in live cells.”
“This work underscores the value of research collaborations between academia and industry,” concludes Matt Robers, Associate Director of R&D at Promega and co-senior author of the study. “By combining our complementary expertise in chemical biology and assay design, we were able to dissect how cooperativity can drive cancer cell selectivity. These findings have real potential to guide the development of future precision medicines.”
This study was the result of collaboration between The Nuffield Department of Medicine’s Centre for Medicines Discovery at the University of Oxford, Promega, and the Center for Advanced Study of Drug Action at Stony Brook University, with additional contributions from researchers at Boston University and the Structural Genomics Consortium at the University of Toronto.