Kiriazis, A. | Search bioRxiv

bioRxiv2023

An improved PDE6D inhibitor combines with Sildenafil to synergistically inhibit KRAS mutant cancer cell growth

The trafficking chaperone PDE6D (or PDE) was proposed as a surrogate target for K-Ras, leading to the development of a series of inhibitors that block its prenyl-binding pocket. These inhibitors suffered from low solubility and intracellular potency, preventing their clinical development. Here we developed a highly soluble PDE6D inhibitor (PDE6Di), Deltaflexin3, which has the currently lowest off-target activity, as we demonstrate in dedicated assays. We further increased the K-Ras focus, by exploiting that PKG2-mediated phosphorylation of Ser181 lowers K-Ras binding to PDE6D. Thus, the combination of Deltaflexin3 with the approved PKG2-activator Sildenafil synergistically inhibits cell- and microtumor growth. However, the overall cancer survival of the high PDE6D/ low PKG2 target population is higher than of the group with the opposite signature. Our results therefore suggest re-examining the interplay between PDE6D and K-Ras in cancer, while recommending the development of PDE6Di that plug, rather than stuff the hydrophobic pocket of PDE6D. SignificanceCombinations of a novel PDE6D inhibitor with Sildenafil synergistically focus the inhibition on K-Ras, however, survival data of the target population suggest an interplay of K-Ras and PDE6D that needs further exploration.

biochemistry↗

bioRxiv2018

Asymmetry in catalysis by Thermotoga maritima membrane-bound pyrophosphatase demonstrated by a non-phosphorous allosteric inhibitor

Membrane-bound pyrophosphatases are homodimeric integral membrane proteins that hydrolyse pyrophosphate into orthophosphates, coupled to the active transport of protons or sodium ions across membranes. They are important in the life cycle of bacteria, archaea, plants, and protist parasites, but no homologous proteins exist in vertebrates, making them a promising drug target. Here, we report the first non-phosphorous allosteric inhibitor (Ki of 1.8 {+/-} 0.3 M) of the thermophilic bacterium Thermotoga maritima membrane-bound pyrophosphatase and its bound structure at 3.7 [A] resolution together with the substrate analogue imidodiphosphate. The unit cell contains two protein homodimers, each binding a single inhibitor dimer near the exit channel, creating a hydrophobic clamp that inhibits the movement of {beta}-strand 1-2 during pumping, and thus preventing the hydrophobic gate from opening. This asymmetry of inhibitor binding with respect to each homodimer provide the first clear demonstration of asymmetry in the catalytic cycle of membrane-bound pyrophosphatases.

molecular biology↗