Molecular Determinants of EphA2 and EphB2 Antagonism Enable the Design of Ligands with Improved Selectivity.

With the aim of identifying novel antagonists selective for the EphA receptor family, a combined experimental and computational approach was taken to investigate the molecular basis of the recognition between a prototypical Eph-ephrin antagonist (UniPR1447) and two representative receptors of the EphA and EphB subfamilies, namely, EphA2 and EphB2 receptors. The conformational free-energy surface (FES) of the binding state of UniPR1447 within the ligand binding domain of EphA2 and EphB2, reconstructed from molecular dynamics (MD) simulations performed on the microsecond time scale, was exploited to drive the design and synthesis of a novel antagonist selective for EphA2 over the EphB2 receptor. The availability of compounds with this pharmacological profile will help discriminate the importance of these two receptors in the insurgence and progression of cancer.

Numerical Simulation of the Heat Transfer in the Cryoprobe of an Innovative Apparatus for Cryosurgery.

Cryosurgery is a rapidly developing discipline, alternative to conventional surgical techniques, used to destroy cancer cells by the action of low temperatures. Currently, the refrigeration is obtained via the adiabatic expansion of gases in probes used for surgeries, with the need of inherently dangerous pressurized vessels. The proposed innovative prototypal apparatus aims to reach the cryosurgical temperatures exploiting a closed-loop refrigeration system, avoiding the hazardous presence of pressurized vessels in the operating room. This study preliminarily examines the technical feasibility of the cryoablation with this machine focusing the attention on the cryoprobe design. Cryoprobe geometry and materials are assessed and the related heat transfer taking place during the cryoablation process is simulated with the aid of the computational fluid dynamics software ANSYS®Fluent. Parametric analyses are carried out varying the length of the collecting tubes and the inlet velocity of the cold carrier fluid in the cryoprobe. The values obtained for physical quantities such as the temperature reached in the treated tissue, the width of the obtained cold front, and the maximum pressure required for the cold carrier fluid are calculated and discussed in order to prove the effectiveness of the experimental apparatus and develop the machine further.

Discovery of a new 1-(phenylsulfonyl)-1H-indole derivative targeting the EphA2 receptor with antiproliferative activity on U251 glioblastoma cell line.

In our continuing effort devoted at developing agents targeting the EphA2 receptor by means of protein-protein interaction (PPI) inhibitors, we report here the design and synthesis of a new class of l-ß-homotryptophan conjugates of 3-ß-hydroxy-Δ5-cholenic acid bearing a set of arylsulfonyl substituents at the indole nitrogen atom. An extensive structure-activity relationship (SAR) analysis indicates that the presence of a bulky lipophilic moiety at the indole nitrogen is fundamental for improving potency on the EphA2 receptor, while abrogating activity on the EphB1-EphB3 receptor subtypes. A rational exploration, guided by the combined application of an experimental design on σp and π physicochemical descriptors and docking simulations, led to the discovery of UniPR1454, a 1-(4-(trifluoromethyl)phenyl)sulfonyl derivative acting as potent and competitive EphA2 antagonist able to inhibit ephrin-A1 dependent signals and to reduce proliferation of glioblastoma (U251) cell line at micromolar concentration.

Protein-Protein Interaction Inhibitors Targeting the Eph-Ephrin System with a Focus on Amino Acid Conjugates of Bile Acids.

The role of the Eph-ephrin system in the etiology of pathological conditions has been consolidated throughout the years. In this context, approaches directed against this signaling system, intended to modulate its activity, can be strategic therapeutic opportunities. Currently, the most promising class of compounds able to interfere with the Eph receptor-ephrin protein interaction is composed of synthetic derivatives of bile acids. In the present review, we summarize the progresses achieved, in terms of chemical expansions and structure-activity relationships, both in the steroidal core and the terminal carboxylic acid group, along with the pharmacological characterization for the most promising Eph-ephrin antagonists in in vivo settings.

Metabolic Soft Spot and Pharmacokinetics: Functionalization of C-3 Position of an Eph-Ephrin Antagonist Featuring a Bile Acid Core as an Effective Strategy to Obtain Oral Bioavailability in Mice.

UniPR129, an L-ß-homotryptophan conjugate of the secondary bile acid lithocholic acid (LCA), acts as an effective protein-protein interaction (PPI) inhibitor of the Eph-ephrin system but suffers from a poor oral bioavailability in mice. To improve UniPR129 bioavailability, a metabolic soft spot, i.e., the 3α-hydroxyl group on the LCA steroidal ring, was functionalized to 3-hydroxyimine. In vitro metabolism of UniPR129 and 3-hydroxyimine derivative UniPR500 was compared in mouse liver subcellular fractions, and main metabolites were profiled by high resolution (HR-MS) and tandem (MS/MS) mass spectrometry. In mouse liver microsomes (MLM), UniPR129 was converted into several metabolites: M1 derived from the oxidation of the 3-hydroxy group to 3-oxo, M2-M7, mono-hydroxylated metabolites, M8-M10, di-hydroxylated metabolites, and M11, a mono-hydroxylated metabolite of M1. Phase II reactions were only minor routes of in vitro biotransformation. UniPR500 shared several metabolic pathways with parent UniPR129, but it showed higher stability in MLM, with a half-life (t1/2) of 60.4 min, if compared to a t1/2 = 16.8 min for UniPR129. When orally administered to mice at the same dose, UniPR500 showed an increased systemic exposure, maintaining an in vitro valuable pharmacological profile as an EphA2 receptor antagonist and an overall improvement in its physico-chemical profile (solubility, lipophilicity), if compared to UniPR129. The present work highlights an effective strategy for the pharmacokinetic optimization of aminoacid conjugates of bile acids as small molecule Eph-ephrin antagonists.