Organic small molecules serve as successful agents in numerous contexts in humans, plants and animals. Through the inhibition of reaction pathways that otherwise drive pathogenesis, they help to cure multiple diseases. The success of small molecules in this regard stems from their ability to effectively interact with the molecule-ligand binding sites that govern important steps of the respective reaction pathway. A range of reaction pathways involve protein-protein interactions. These are hard to tackle with small molecules.

One such pathway is the HIF-1α pathway. HIF, the hypoxia inducible factor, is a transcription factor that is expressed in most oxygen-breathing species. It responds to the decrease of available oxygen (hypoxia) in the cellular environment, thus regulating the response of the cell to oxygen deprivation. Under normal oxygen concentrations in the cell, HIF-1α is degraded after transcription. Under hypoxic conditions, HIF-1α accumulates in the cell and dimerizes with HIF-1β,​ forming a protein heterodimer that complexes with the p300 coactivator in the nucleus. The resulting HIF-1α-p300 complex then binds to the hypoxia response element (HRE) to activate the transcription of genes involved in angiogenesis, physological metabolism, cell growth and survival. Bad enough, some of the resulting HIF-1α gene products turn out to play a role in the genesis and metastasis of tumor cells.

Hence, inhibition of the HIF-1α pathway has become a major target in the fight against cancer. One possible strategy to inhibit this pathway is the inhibition of the HIF-1α-p300 protein-protein interaction. And here we are back at small molecules and their limits when it comes to the inhibition of protein-protein interactions.

Researchers from the University of Macau, Hong Kong Baptist University and the City University of Hong Kong have now reported a transition-metal complex as inhibitor of the described protein-protein interaction.

Transition-metal complexes are promising candidates for inhibitors of protein-protein interactions as they provide a specific three-dimensional shape. They can thus act as globular scaffolds. Moreover, due to the complex chemistry of transition metal ions, transition-metal complexes exhibit rich and diverse molecular properties and binding motifs – a richness and diversity inaccessible to traditional organic small molecules.

From a set of seven promising osmium(II) complexes, the researchers identified one as a particular promising lead for an inhibitor of the HIF-1α-p300 interaction. The identified complex 1, formula [Os(phen-tMe)3](PF6)2, reduces the HRE-driven luciferase activity under hypoxic conditions by 73.6% as inferred from a dual luciferase reporter assay. This makes the newly discovered complex 1 more potent than the known HIF-1α-p300-interaction inhibitor chetomin. Using dose-response assays, a dose-dependent inhibition of HRE-driven luciferase activity under hypoxia with an IC50 of 1.22μM for complex 1 was found. This is particularly impressive when compared with the IC50 of chetomin which is 9.38μM.

Through co-immunoprecipitation experiment, it was shown in cellulo that complex 1 indeed interferes with the HIF-1α-p300 interaction. On the other hand, complex 1 had no significant impact on the protein-protein interaction of the HIF-1α​–HIF-1β​ heterodimer. This supports a specificity of the inhibition process and excludes inhibition as a mere artifact of non-specific hydrophobic interactions. Complex 1 additionally suppresses HIF-1α activity through inhibition of upstream pathways, specifically through inhibition of SRC, SKT and STAT3 phosphorylation.

From a comparison of the proposed seven complexes as well as a preliminary structure-activity relationship analysis, the researchers conclude that the osmium centre coordinates the bioactive structure of the complex. The researchers suspect that the excellent inhibition properties of complex 1 result from the high degree of shape complementarity between complex 1 and the HIF-1α binding site. These experimental findings are corroborated by molecular docking analysis. The structure of complex 1 was obtained from geometry optimization with density functional theory using ORCA. Resolution-of-the-identity was used to accelerate the calculation.

Normally, the environment that develops around tumor tissue is hypoxic. Therefore, the higher degree of effectiveness and cytotoxicity of complex 1 under hypoxic conditions as compared to normal oxygen concentrations adds to the attractiveness of complex 1 as a particularly promising anti-cancer lead.

C. Yang, W. Wang, G. Li, H. Zong, Z. Dong, C. Wong, Daniel Kwong, D. Ma, C. Leung: “Anticancer osmium complex inhibitors of the HIF-1α and p300 protein-protein interaction”, Scientific Reports 7, 42860 (2017)