C2
DARPin-based stabilization of p53 for cancer therapy
Prof. Volker Dötsch, Goethe University Frankfurt; Dr Andreas Jörger, Goethe University Frankfurt
Innovectis GmbH
Challenge
As ‘guardian of the genome’, p53 orchestrates DNA repair, cell-cycle arrest, and apoptosis in response to stress. When p53 is lost or dysfunctional, this causes genomic instability and uncontrolled proliferation, and alterations in p53 are found in more than half of all human cancers. Even in cancer types where p53 mutations are rare, for example acute myeloid leukemia (AML), p53 alterations correlate with poor prognosis. As a transcription factor, p53 has long been considered ‘undruggable’, leaving a major therapeutic gap in oncology. With our approach we address two types of p53-related cancers, namely those driven by certain p53 mutations (e.g. AML) and carcinomas caused by infection with high-risk human papilloma viruses (HPV). TP53-mutated AML has the worst prognosis, with a median overall survival of only 6 months, while HPV-related cervical cancer is the fourth leading cause of cancer-related deaths in women worldwide.
Technology
We developed a Designed Ankyrin Repeat Protein (DARPin) that binds to the DNA binding domain (DBD) of human p53 with very high affinity (3 nM). DARPins are artificial molecules derived from ankyrin repeat proteins that recognize three-dimensional protein epitopes, but unlike conventional antibodies they are very small (~15 kDa). Mutations in p53 often cause destabilization of the DBD, impairing its DNA-binding ability. Our DARPin can bind and stabilize one-third of all known DBD mutants (so-called temperature-sensitive mutants) and restore p53 function. In contrast to small molecule therapeutics such as Rezatapopt, which targets a single point mutation (Y220C), our DARPin reactivates a wide range of temperature-sensitive p53 mutants, covering a large patient population. The same DARPin can also displace the HPV E6 oncoprotein from p53, thereby preventing E6AP-mediated ubiquitination and p53 degradation in HPV-induced cancers. In HPV-positive cervical carcinoma cells, our DARPin restored p53 function and induced apoptosis, while no p53 activation was seen in HPV-negative cells, suggesting low off-target effects.
In in vitro cell culture experiments, we were able to show effective delivery of DARPin mRNA via lipid nanoparticles (LNPs). For several cell types and tissues, a targeted delivery of mRNA has been demonstrated using LNPs with distinct compositions, possibly enabling delivery of the DARPin mRNA specifically to the cancerogenic cells.
Commercial Opportunity
As indicated, our DARPin binds and reactivates a number of p53 mutants that, while individually rare, collectively affect up to 3 million new cancer patients annually. Existing therapeutics are either highly toxic or only target specific p53 mutants, while our DARPin binds to a broad range of clinically relevant pathogenic mutants. Additionally, it might serve as a pan-HPV treatment independed of HPV type. So far, no therapies directed against HPV in HPV-positive cancers are available, and therapy success often is poor. Several DARPins have been shown to cause limited side effects in clinical trials, and our approach of DARPin delivery via mRNA LNPs promises high tissue specificity.
Development Status
In cell culture experiments using patient-derived cell lines, our DARPin successfully activated a p53-based transcriptional program. In vivo studies using xenotransplanted human HPV-positive cancer cell lines in mice have just started.
Patent Situation
Application has been submitted to the European Patent Office (PCT/EP2025/069957 ‘Designed Ankyrin Repeat Proteins binding p53, and uses thereof’).
Further Reading
Münick et al. (2025) Nat Struct Mol Biol 32(5):790-801 doi: 10.1038/s41594-024-01456-7
