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Pancreatic ductal adenocarcinoma remains a highly aggressive and untreatable cancer. There is a need to develop a new PDAC-associated antigen-targeting drug delivery system to tackle this disease. We validated choosing ZIP4 as a putative target in PDAC theranostics. We developed a nanosystem composed of a fluorescent polystyrene core coated with gold nanoparticles onto which a ZIP4-specific polyclonal antibody is attached. The polystyrene core's fluorescence properties allow the nanosystem tracking by intravital imaging. We also developed two ZIP4-expressing cell lines by stably transfecting HEK293 and RWP1 cells with a ZIP4-coding plasmid that simultaneously provides cells with puromycin resistance. We studied the cell internalisation of the as-synthesised nanoparticles and demonstrated that ZIP4-expressing HEK293 and ZIP4-expressing RWP1 cells tended to take up more ZIP4-targeting nanoparticles. Moreover, we observed that ZIP4-targeting nanoparticles accumulated more in ZIP4-expressing HEK293 and RWP1 tumours when injected intravenously in a subcutaneous xenograft and an orthotopic in vivo model, respectively. Furthermore, the administration of these nanoparticles did not induce any significant systemic toxicity as determined by histological analysis of all organs. Altogether, these results provide the first evidence of the feasibility of using a ZIP4-targeting nanosystem further to design efficient therapeutic and diagnostic tools for PDAC. [ABSTRACT FROM AUTHOR]

Background and Purpose: The transient receptor potential vanilloid 4 (TRPV4) cation channel participates in multiple physiological processes and is also at the core of different diseases, making this channel an interesting pharmacological target with therapeutic potential. However, little is known about the structural elements governing its inhibition. Experimental Approach We have now combined in silico drug discovery and molecular dynamics simulation based on Xenopus tropicalis xTRPV4 structure with functional studies measuring cell Ca2+ influx mediated by human TRPV4 channel to characterize the binding site of known TRPV4 inhibitors and to identify novel small molecule channel modulators. Key Results: We have found that the inhibitor HC067047 binds to a pocket conformed by residues from S2–S3 linker (xTRPV4‐D542), S4 (xTRPV4‐M583 and Y587 and S5 (xTRPV4‐D609 and F613). This pocket was also used for structure‐based virtual screening in the search of novel channel modulators. Forty potential hits were selected based on the lower docking scores (from ~250,000 compounds) and their effect upon TRPV4 functionally tested. Three were further analysed for stability using molecular dynamics simulation and functionally tested on TRPV4 channels carrying mutations in the binding pocket. Compound NSC151066, shown to require residue xTRPV4‐M583 for its inhibitory effect, presented an IC50 of 145 nM and demonstrated to be an effective antiviral against Zika virus with a potency similar to HC067047. Conclusion and Implications: Together, we propose structural insights into the inhibition of TRPV4 and how this information can be used for the design of novel channel modulators. LINKED ARTICLES: This article is part of a themed issue on Structure Guided Pharmacology of Membrane Proteins (BJP 75th Anniversary). To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.14/issuetoc [ABSTRACT FROM AUTHOR]

Calcium signaling in the brain is fundamental to the learning and memory process and there is evidence to suggest that its dysfunction is involved in the pathological pathways underlying Alzheimer’s disease (AD). Recently, the calcium hypothesis of AD has received support with the identification of the non-selective Ca2+-permeable channel CALHM1. A genetic polymorphism (p. P86L) in CALHM1 reduces plasma membrane Ca2+ permeability and is associated with an earlier age-at-onset of AD. To investigate the role of CALHM1 variants in early-onset AD (EOAD), we sequenced all CALHM1 coding regions in three independent series comprising 284 EOAD patients and 326 controls. Two missense mutations in patients (p.G330D and p.R154H) and one (p.A213T) in a control individual were identified. Calcium imaging analyses revealed that while the mutation found in a control (p.A213T) behaved as wild-type CALHM1 (CALHM1-WT), a complete abolishment of the Ca2+ influx was associated with the mutations found in EOAD patients (p.G330D and p.R154H). Notably, the previously reported p. P86L mutation was associated with an intermediate Ca2+ influx between the CALHM1-WT and the p.G330D and p.R154H mutations. Since neither expression of wild-type nor mutant CALHM1 affected amyloid ß-peptide (Aß) production or Aß-mediated cellular toxicity, we conclude that rare genetic variants in CALHM1 lead to Ca2+ dysregulation and may contribute to the risk of EOAD through a mechanism independent from the classical Aß cascade. [ABSTRACT FROM AUTHOR]