Stapled ß-Hairpins Featuring 4-Mercaptoproline.

Peptides constrained by intramolecular cross-links, especially stapled α-helices, have emerged as versatile scaffolds for drug development. However, there are fewer examples of similarly constrained scaffolds for other secondary structures. Here, we used a novel computational strategy to identify an optimal staple for antiparallel ß-strands, and then we incorporated that staple within a ß-hairpin peptide. The hairpin uses 4-mercaptoproline as a novel staple component, which contributes to a unique, kinked structure. The stapled hairpins show a high degree of structure in aqueous solution, excellent resistance to degradation in cell lysates, and cytosolic penetration at micromolar concentrations. They also overlay with a unique subset of kinked hairpin motifs at protein-protein interaction interfaces. Thus, these scaffolds represent promising starting points for developing inhibitors of cellular protein-protein interactions.

Formulation and evaluation of itraconazole liposomes for Hedgehog pathway inhibition.

Itraconazole (ITZ) is an FDA-approved antifungal agent that has recently been explored for novel biological properties. In particular, ITZ was identified as a potent inhibitor of the hedgehog (Hh) pathway, a cell signalling pathway that has been linked to a variety of cancers and accounts for ∼25% of paediatric medulloblastoma (MB) cases. To date, there is not a targeted therapeutic option for paediatric MB, resulting in long-term side effects such as hormone deficiency, organ damage and secondary cancers. A primary obstacle for developing targeted therapy for brain ailments is the presence of the blood-brain barrier (BBB), which protects the brain from potentially harmful substances. Due to its size and hydrophobicity, ITZ does not penetrate the BBB. Alternatively, liposomes are being increasingly used within the clinic to increase drug bioavailability, target specificity and BBB permeability. With this in mind, we have successfully developed ITZ-containing liposomes with an optimal size for BBB penetration (<100 nm) and encapsulation efficiency (∼95%) by utilizing a continuous manufacturing approach-turbulent coaxial jet in co-flow. Our preliminary in vitro data demonstrate that these liposomes inhibit the Hh pathway, albeit at a reduced level in comparison to free ITZ. (196/250 words).

Structure-Activity Relationships for Itraconazole-Based Triazolone Analogues as Hedgehog Pathway Inhibitors.

The Food and Drug Administration-approved antifungal agent, itraconazole (ITZ), has been increasingly studied for its novel biological properties. In particular, ITZ inhibits the hedgehog (Hh) signaling pathway and has the potential to serve as an anticancer chemotherapeutic against several Hh-dependent malignancies. We have extended our studies on ITZ analogues as Hh pathway inhibitors through the design, synthesis, and evaluation of novel des-triazole ITZ analogues that incorporate modifications to the triazolone/side chain region of the scaffold. Our overall results suggest that the triazolone/side chain region can be replaced with various functionalities (hydrazine carboxamides and meta-substituted amides) resulting in improved potency when compared to ITZ. Our studies also indicate that the stereochemical orientation of the dioxolane ring is important for both potent Hh pathway inhibition and compound stability. Finally, our studies suggest that the ITZ scaffold can be successfully modified in terms of functionality and stereochemistry to further improve its anti-Hh potency and physicochemical properties.

Repurposing the Clinically Efficacious Antifungal Agent Itraconazole as an Anticancer Chemotherapeutic.

Itraconazole (ITZ) is an FDA-approved member of the triazole class of antifungal agents. Two recent drug repurposing screens identified ITZ as a promising anticancer chemotherapeutic that inhibits both the angiogenesis and hedgehog (Hh) signaling pathways. We have synthesized and evaluated first- and second-generation ITZ analogues for their anti-Hh and antiangiogenic activities to probe more fully the structural requirements for these anticancer properties. Our overall results suggest that the triazole functionality is required for ITZ-mediated inhibition of angiogenesis but that it is not essential for inhibition of Hh signaling. The synthesis and evaluation of stereochemically defined des-triazole ITZ analogues also provides key information as to the optimal configuration around the dioxolane ring of the ITZ scaffold. Finally, the results from our studies suggest that two distinct cellular mechanisms of action govern the anticancer properties of the ITZ scaffold.