DECONSTRUCTING ONC201 TO DESIGN NEW MITOCHONDRIAL ClpP ACTIVATORS FOR PEDIATRIC H3K27-ALTERED DIFFUSE MIDLINE GLIOMA

Diffuse Midline Glioma (DMG) with H3K27 alteration is one of the most aggressive and therapeutically challenging pediatric brain tumors. Characterized by a lysine-to-methionine substitution at position 27 in histone H3 (H3K27M), this epigenetic mutation leads to global chromatin remodeling and aberrant gene expression, contributing to a dismal prognosis and limited treatment options. As conventional therapies have failed to significantly improve survival, there is an urgent need for novel therapeutic strategies that target the unique vulnerabilities of this disease.1 Recent studies have identified the mitochondrial protease ClpP as a promising therapeutic target in cancer, including DMG. ClpP is part of the ClpXP/ClpP proteolytic complex, which maintains mitochondrial proteostasis by degrading misfolded or damaged proteins. When pathologically activated by small molecules, ClpP triggers uncontrolled degradation of essential mitochondrial matrix proteins, leading to mitochondrial dysfunction, disruption of metabolic homeostasis, and ultimately, cell death. This mechanism is particularly effective in tumor cells that are highly dependent on mitochondrial integrity, making ClpP hyperactivation a compelling anti cancer strategy.2 ONC201, a first-in-class imipridone compound, was identified as a ClpP-binding molecule capable of inducing mitochondrial stress and anti-tumor effects in multiple cancer models, including DMG. ONC201 induced regression of the primary thalamic lesion in a limited number of DMG patients; however, despite this initial clinical promise, it is unfortunately not effective in all cases, underscoring the need to identify alternative molecules for non-responding patients.3 By fully deconstructing the imipridone core and discarding its rigid tricyclic scaffold, we have rationally designed and synthesized a new generation of compounds with novel chemical frameworks which were subjected to pharmacochemical profiling, encompassing stability, membrane permeability, and ClpP affinity. The novel derivatives retain high selectivity for ClpP and exhibit enhanced mitochondrial protease activation, resulting in potent degradation of mitochondrial matrix proteins and profound disruption of mitochondrial function.