Topically administered purified clinoptilolite-tuff for the treatment of cutaneous wounds: A prospective, randomised phase I clinical trial.

In an ageing society, chronic ulcers pose an increasingly relevant healthcare issue associated with significant morbidity and an increasing financial burden. Hence, there is an unmet medical need for novel, cost-effective therapies that improve healing of chronic cutaneous wounds. This prospective, randomised, open-label, phase I trial investigated the safety and tolerability of topically administered purified clinoptilolite-tuff (PCT), mainly consisting of the naturally occurring zeolite-mineral clinoptilolite, in artificial wounds in healthy male volunteers compared to the standard of care (SoC). We found that topically administered PCT was safe for therapeutic application in acute wounds in healthy male volunteers. No significant differences in wound healing or wound conditions were observed compared to SoC-treated wounds. However, we found a significantly higher proportion of CD68-positive cells and a significantly lower proportion of α-smooth muscle actin-positive cells in PCT-treated wounds. Scanning electron microscopy revealed PCT particles in the restored dermis in some cases. However, these did not impede wound healing or clinical symptoms. Hence, purified PCT could represent an attractive, cost-effective wound treatment promoting the process of healing.

Binding and neutralization of C. difficile toxins A and B by purified clinoptilolite-tuff.

Clostridioides difficile (C. difficile) infection is a major public health problem worldwide. The current treatment of C. difficile-associated diarrhea relies on the use of antibacterial agents. However, recurrences are frequent. The main virulence factors of C. difficile are two secreted cytotoxic proteins toxin A and toxin B. Alternative research exploring toxin binding by resins found a reduced rate of recurrence by administration of tolevamer. Hence, binding of exotoxins may be useful in preventing a relapse provided that the adsorbent is innocuous. Here, we examined the toxin binding capacity of G-PUR®, a purified version of natural clinoptilolite-tuff. Our observations showed that the purified clinoptilolite-tuff adsorbed clinically relevant amounts of C. difficile toxins A and B in vitro and neutralized their action in a Caco-2 intestinal model. This conclusion is based on four independent sets of findings: G-PUR® abrogated toxin-induced (i) RAC1 glucosylation, (ii) redistribution of occludin, (iii) rarefaction of the brush border as visualized by scanning electron microscopy and (iv) breakdown of the epithelial barrier recorded by transepithelial electrical resistance monitoring. Finally, we confirmed that the epithelial monolayer tolerated G-PUR® over a wide range of particle densities. Our findings justify the further exploration of purified clinoptilolite-tuff as a safe agent in the treatment and/or prevention of C. difficile-associated diarrhea.

Clinoptilolite in Dextran Sulphate Sodium-Induced Murine Colitis: Efficacy and Safety of a Microparticulate Preparation.

Background: Clinoptilolite is an aluminium silicate of natural origin; the microporous structure and the net negative charge of its crystal lattice allows for adsorption of ions, toxins, inflammatory mediators, and some microorganisms. We generated 2 preparations of purified clinoptilolite, which differed by about 10-fold in particle size, ie, a standard powder (GHC1) and a microparticulate fraction (GHC2) with a size of 3.6 µm and 0.39 µm (d50) respectively. These were examined for their ability to accelerate the recovery of mice from DSS (dextran sulphate sodium)-induced intestinal inflammation. Methods: Efficacy of clinoptilolite preparations was investigated by administering DSS-treated mice twice daily with 30 mg GHC2 or GHC1 for 5 consecutive days, followed by 5 days of recovery without DSS. To explore the safety of the microparticulate preparation (GHC2), mice were subjected to 4 cycles of DSS-exposure. We specifically verified that clinoptilolite microparticles were not systemically bioavailable by examining the gut tissue and the liver for the accumulation of microparticles by transmission electron microscopy. Results: Treatment of mice with GHC2 was superior to GHC1 and as effective as the reference compound 5-aminosalicylic acid in ameliorating the damage induced by the exposure to DSS. In addition, no clinoptilolite particle was observed in the intestinal epithelial layer, gut-associated lymph follicles, or in the liver. Conclusion: Our observations confirm that a microparticulate preparation of clinoptilolite is safe and effective in a murine model of inflammatory bowel disease and supports the hypothesis that the adsorptive capacity of clinoptilolite is of potential therapeutic relevance.

A mutation in the first ligand-binding repeat of the human very-low-density lipoprotein receptor results in high-affinity binding of the single V1 module to human rhinovirus 2.

Minor group human rhinoviruses (HRVs) bind members of the low-density lipoprotein receptor family for cell entry. The ligand-binding domains of these membrane proteins are composed of various numbers of direct repeats of about 40 amino acids in length. Residues involved in binding of module 3 (V3) of the very-low-density lipoprotein receptor (VLDLR) to HRV2 have been identified by X-ray crystallography (N. Verdaguer, I. Fita, M. Reithmayer, R. Moser, and D. Blaas, Nat. Struct. Mol. Biol. 11:429-434, 2004). Sequence comparisons of the eight repeats of VLDLR with respect to the residues implicated in the interaction between V3 and HRV2 suggested that (in addition to V3) V1, V2, V5, and V6 also fulfill the requirements for interacting with the virus. Using a highly sensitive binding assay employing phage display, we demonstrate that single modules V2, V3, and V5 indeed bind HRV2. However, V1 does not. A single mutation from threonine 17 to proline converted the nonbinding wild-type form of V1 into a very strong binder. We interpret the dramatic increase in affinity by the generation of a hydrophobic patch between virus and receptor; in the presence of threonine, the contact area might be disturbed. This demonstrates that the interaction between virus and its natural receptors can be strongly enhanced by mutation.

Kinetics of thermal denaturation of human rhinoviruses in the presence of anti-viral capsid binders analyzed by capillary electrophoresis.

In vivo, the icosahedral capsid of human rhinoviruses undergoes well-defined transitions during the infection pathway. Native virus, sedimenting at 150S, is converted to subviral particles with a sedimentation coefficient of 135S, which have lost the innermost capsid protein VP4. Upon release of the genomic RNA empty 80S capsids remain. Similar structural modifications are observed in vitro upon exposure to low pH and/or elevated temperature. Virions are stabilized against these transitions by various antiviral compounds, which bind to a hydrophobic pocket in the capsid protein VP1. Using capillary electrophoresis the kinetics of viral decay in the presence of such hydrophobic drugs was investigated. Assuming first-order kinetics, the increase of the time constant reflects the extent of stabilization. Exposure of the virions to 55 degrees C after presaturation with the antivirals increased the time constants (as compared to native virus) by a factor of 8-30, from a few minutes to several ten minutes. Denaturation of the stabilized capsid gave rise to heterogeneous material rather than to defined subviral particles. This was confirmed by electron microscopy and indicates that the structural modification of the virus follows a kinetically well-defined pathway which is disturbed by the drugs resulting in disorganized disruption of the virion.