The Leibniz Institute for Analytical Sciences (ISAS) carried out a three‑year project (01 August 2020 – 28 February 2023, with a cost‑neutral extension) funded under the German BMBF grant number 16GW0262K. The aim was to validate the dimerisation interface of the mitogen‑activated protein kinase ERK1/2 as a therapeutic target for heart failure, cancer and RASopathies, to crystallise and model the ERK‑peptide complex, to develop low‑molecular‑weight inhibitors of the ERK‑dimer complex, and to establish assays for future optimisation of lead compounds. The project was led by ISAS and involved close collaboration with the University Hospital Würzburg (Armin Wiegering), the University Hospital Göttingen, and the research partner LDC, which supplied small‑molecule candidates.

Technically, the team first established a 96‑well cardiotoxicity screening platform that measures cell viability, arrhythmia incidence and heart rhythm in cultured cardiomyocytes. Using this platform, the ERK‑inhibiting peptide EDI (derived from the ERK‑dimer interface) and several small molecules identified by LDC were characterised for their effects on cardiac cells. In parallel, a transgenic mouse line was generated that ubiquitously expresses the EDI peptide, allowing in‑vivo assessment of potential side effects.

In vitro studies demonstrated that EDI selectively inhibited proliferation of H1703 lung‑cancer cells, which depend on the MAPK cascade, while having no effect on H196 cells that are less MAPK‑dependent. Western blot analysis confirmed that EDI reduced phosphorylation of ERK at threonine 188 (pERK T188) in H1703 cells but not in H196 cells, supporting the hypothesis that the peptide targets the pathological ERK‑dimer interface without disrupting essential cardiac ERK signalling.

For cancer relevance, human colon‑carcinoma organoids were established at Würzburg, lentiviral vectors were produced, and the organoids were exposed to EDI and the LDC‑derived compounds. Preliminary data indicate that the peptide and selected small molecules suppress organoid growth, suggesting translational potential.

The RASopathy component involved Raf1‑L613V mutant mice, a model of Noonan syndrome. These mice were treated with an AAV9‑EDI vector or crossed with the EDI‑expressing line. Longitudinal monitoring and survival analysis revealed that Raf1 mutants exhibit a significantly increased heart weight after transverse aortic constriction (TAC) compared with controls, whereas EDI‑expressing mice show a reduced heart weight under the same hypertrophic stimulus. Histological and functional cardiac parameters are still being analysed but appear promising. Collaboration with Göttingen enabled the screening of EDI and lead compounds in induced pluripotent‑stem‑cell‑derived cardiomyocytes for further in‑vivo validation.

In 2022, cardiotoxicity assays compared adenoviral‑delivered EDI with low‑molecular‑weight inhibitors, including DEL‑22379, a compound reported by another group. DEL‑22379 displayed an antihypertrophic effect and inhibited pERK T188 similarly to EDI, indicating that small‑molecule inhibition of the ERK‑dimer interface is feasible.

The project faced delays due to the COVID‑19 pandemic, mouse breeding setbacks, and personnel changes, which postponed some in‑vitro work. A cost‑neutral extension allowed the team to recover lost time and complete the planned assays. Overall, the study provides strong evidence that targeting the ERK‑dimer interface can selectively inhibit pathological ERK signalling in heart failure, cancer, and RASopathies while sparing essential cardiac functions, and it establishes a robust platform for further drug development.