Safety and efficacy of a biomimetic monolayer of permanently bound multiphosphonic acid molecules on dental implants: 3 years post-loading results from a pilot quadruple-blinded randomised controlled trial.

PURPOSE: To evaluate the safety and clinical efficacy of a novel surface treatment (SurfLink, Nano Bridging Molecules, Gland, Switzerland) on titanium dental implants. SurfLink consists of a monolayer of permanently bound multiphosphonic acid molecules, which mimics the surface of naturally occurring hydroxyapatite. MATERIALS AND METHODS: Twenty-three patients requiring at least two single dental implants had their sites randomised according to a split-mouth design to receive one titanium grade 4 implant treated with SurfLink and one untreated control implant. Additional SurfLink-treated implants were placed if needed. Implants were submerged for 3 months in mandibles and 6 months in maxillae, then loaded with definitive metal-ceramic crowns and followed for 3 years after loading. Outcome measures were crown/implant failures, any complication, radiographic peri-implant marginal bone level changes and marginal bleeding. RESULTS: Three patients dropped out but all remaining patients were followed up to 3-years post-loading. No implant failed. Complications were reported for three patients, affecting both types of implant in two patients and only SurfLink implant in one patient. No differences for complications between the two implant types was observed (McNemar test, P = 1, difference in proportions = -0.04, 95% CI: -0.22 to 0.14). No bleeding was observed when a periodontal probe was run in the peri-implant soft tissues around any of the implants, with the exception of three implants affected by peri-implantitis. There were no statistically significant differences in marginal bone level changes between the two groups (at 3 years post-loading P = 0.86, mean difference = -0.05; SD = 1.15; 95% CI: -0.56 to 0.47). CONCLUSIONS: Medium-term data (3-years post-loading) of implants with a biomimetic monolayer of permanently bound multiphosphonic acid molecules (SurfLink surface treatment) presented no safety issues. Clinical healing in both control and SurfLink-treated implant groups was uneventful and did not differ significantly. More challenging clinical situations need to be investigated to evaluate the real effectiveness of this surface treatment. Conflict-of-interest statement: Marco Esposito and Ivan Dojcinovic declare that they have no conflict of interest, and they acted as consultants for Nano Bridging Molecules (NBM). Sabrina Buchini, Péter Péchy and Björn-Owe Aronsson are employed at NBM. NBM, the manufacturer of the SurfLink surface treatment, supported and carefully monitored this trial without interfering with the presentation of its results.

A novel multi-phosphonate surface treatment of titanium dental implants: a study in sheep.

The aim of the present study was to evaluate a new multi-phosphonate surface treatment (SurfLink®) in an unloaded sheep model. Treated implants were compared to control implants in terms of bone to implant contact (BIC), bone formation, and biomechanical stability. The study used two types of implants (rough or machined surface finish) each with either the multi-phosphonate Wet or Dry treatment or no treatment (control) for a total of six groups. Animals were sacrificed after 2, 8, and 52 weeks. No adverse events were observed at any time point. At two weeks, removal torque showed significantly higher values for the multi-phosphonate treated rough surface (+32% and +29%, Dry and Wet, respectively) compared to rough control. At 52 weeks, a significantly higher removal torque was observed for the multi-phosphonate treated machined surfaces (+37% and 23%, Dry and Wet, respectively). The multi-phosphonate treated groups showed a positive tendency for higher BIC with time and increased new-old bone ratio at eight weeks. SEM images revealed greater amounts of organic materials on the multi-phosphonate treated compared to control implants, with the bone fracture (from the torque test) appearing within the bone rather than at the bone to implant interface as it occurred for control implants.

Tuning mechanism-based inactivators of neuraminidases: mechanistic and structural insights.

3-Fluorosialosyl fluorides are inhibitors of sialidases that function by the formation of a long-lived covalent active-site adduct and have potential as therapeutics if made specific for the pathogen sialidase. Surprisingly, human Neu2 and the Trypanosoma cruzi trans-sialidase are inactivated more rapidly by the reagent with an equatorial fluorine at C3 than by its axial epimer, with reactivation following the same pattern. To explore a possible stereoelectronic basis for this, rate constants for spontaneous hydrolysis of the full series of four 3-fluorosialosyl fluorides were measured, and ground-state energies for each computed. The alpha (equatorial) anomeric fluorides hydrolyze more rapidly than their beta anomers, consistent with their higher ground-state energies. However ground-state energies do not explain the relative spontaneous reactivities of the 3-fluoro-epimers. The three-dimensional structures of the two 3-fluoro-sialosyl enzyme intermediates of human Neu2 were solved, revealing key stabilizing interactions between Arg21 and the equatorial, but not the axial, fluorine. Because of changes in geometry these interactions will increase at the transition state, likely explaining the difference in reaction rates.

Safety and efficacy of a biomimetic monolayer of permanently bound multi-phosphonic acid molecules on dental implants: 1 year post-loading results from a pilot quadruple-blinded randomised controlled trial.

PURPOSE: To evaluate the safety and clinical efficacy of a novel surface treatment (SurfLink®, Nano Bridging Molecules, Gland, Switzerland) on titanium dental implants. SurfLink consists of a monolayer of permanently bound multi-phosphonic acid molecules, which mimics the surface of naturally occurring hydroxyapatite. MATERIALS AND METHODS: Twenty-three patients requiring at least two single dental implants had their sites randomised according to a split-mouth design to receive one titanium grade 4 implant treated with SurfLink and one untreated control implant. Additional SurfLink-treated implants were placed if needed. Implants were submerged for 3 months in mandibles and 6 months in maxillae, were loaded with definitive metal-ceramic crowns, and followed up for 1 year after loading. Outcome measures were crown/implant failures, any complication, radiographic peri-implant marginal bone level changes and marginal bleeding. RESULTS: One patient dropped out after abutment connection. All remaining patients were followed up to 1 year post-loading. No implant failed and only 1 postoperative complication (pain) occurred, but it may not have been related to the implant treatment. No bleeding was observed when a periodontal probe was used to examine the peri-implant soft tissues around the implants. There were no statistically significant differences in marginal bone level changes between the two groups (P = 0.057, mean difference = -0.27, SE = 0.13; 95% CI -0.55 to 0.01). CONCLUSIONS: Preliminary short-term data (1 year post-loading) of implants with a biomimetic monolayer of permanently bound multi-phosphonic acid molecules (SurfLink surface treatment) presented no safety issues. Clinical healing in both the control and SurfLink-treated implant group was uneventful and did not differ significantly between groups. More challenging clinical situations need to be investigated to evaluate the real effectiveness of this surface treatment.

Mechanism-based covalent neuraminidase inhibitors with broad-spectrum influenza antiviral activity.

Influenza antiviral agents play important roles in modulating disease severity and in controlling pandemics while vaccines are prepared, but the development of resistance to agents like the commonly used neuraminidase inhibitor oseltamivir may limit their future utility. We report here on a new class of specific, mechanism-based anti-influenza drugs that function through the formation of a stabilized covalent intermediate in the influenza neuraminidase enzyme, and we confirm this mode of action with structural and mechanistic studies. These compounds function in cell-based assays and in animal models, with efficacies comparable to that of the neuraminidase inhibitor zanamivir and with broad-spectrum activity against drug-resistant strains in vitro. The similarity of their structure to that of the natural substrate and their mechanism-based design make these attractive antiviral candidates.

Kinetic and mechanistic analysis of Trypanosoma cruzi trans-sialidase reveals a classical ping-pong mechanism with acid/base catalysis.

The trans-sialidase from Trypanosoma cruzi catalyzes the transfer of a sialic acid moiety from sialylated donor substrates to the terminal galactose moiety of lactose and lactoside acceptors to yield alpha-(2,3)-sialyllactose or its derivatives with net retention of anomeric configuration. Through kinetic analyses in which the concentrations of two different donor aryl alpha-sialoside substrates and the acceptor substrate lactose were independently varied, we have demonstrated that this enzyme follows a ping-pong bi-bi kinetic mechanism. This is supported for both the native enzyme and a mutant (D59A) in which the putative acid/base catalyst has been replaced by the demonstration of the half-reaction in which a sialyl-enzyme intermediate is formed. Mass spectrometric analysis of the protein directly demonstrates the formation of a covalent intermediate, while the observation of release of a full equivalent of p-nitrophenol by the mutant in a pre-steady state burst provides further support. The active site nucleophile is confirmed to be Tyr342 by trapping of the sialyl-enzyme intermediate using the D59A mutant and sequencing of the purified peptic peptide. The role of D59 as the acid/base catalyst is confirmed by chemical rescue studies in which activity is restored to the D59A mutant by azide and a sialyl azide product is formed.

High-throughput screening methodology for the directed evolution of glycosyltransferases.

Engineering of glycosyltransferases (GTs) with desired substrate specificity for the synthesis of new oligosaccharides holds great potential for the development of the field of glycobiology. However, engineering of GTs by directed evolution methodologies is hampered by the lack of efficient screening systems for sugar-transfer activity. We report here the development of a new fluorescence-based high-throughput screening (HTS) methodology for the directed evolution of sialyltransferases (STs). Using this methodology, we detected the formation of sialosides in intact Escherichia coli cells by selectively trapping the fluorescently labeled transfer products in the cell and analyzing and sorting the resulting cell population using a fluorescence-activated cell sorter (FACS). We screened a library of >10(6) ST mutants using this methodology and found a variant with up to 400-fold higher catalytic efficiency for transfer to a variety of fluorescently labeled acceptor sugars, including a thiosugar, yielding a metabolically stable product.

Recent improvements in antigene technology.

DNA triple-helix-based approaches to control and modulate cellular functions on the level of genomic DNA (antigene technology) suffered in the past from a stepmother-like treatment in comparison to the flourishing field of oligonucleotide-based control of translation (antisense technology). This was mostly due to lack of affinity of triplex-forming oligonucleotides to their DNA target, to sequence restriction constraints imposed by the triple helical recognition motifs and by open questions to the accessibility of the target DNA. Recent developments in the area have brought about new bases that specifically recognize pyrimidine-purine inversion sites as well as sugar modifications, for example, the 2'-aminoethoxy-oligonucleotides or oligonucleotides based on the locked nucleic acid sugar unit, which greatly enhance triplex stability and alleviate in part the sequence restriction constraints. With this, sequence-specific genomic DNA manipulation is starting to become a useful tool in biotechnology.

New nucleoside analogues for the recognition of pyrimidine-purine inversion sites.

A new nucleoside designed to enhance triplex stability has been synthesised in 15 steps starting from sugar 2. This pathway contains the sugar derivative 9 which is a useful intermediate for the introduction of other natural and unnatural bases into the 2'-aminoethoxy nucleoside containing scaffold.

A catalytic antibody programmed for torsional activation of amide bond hydrolysis.

Amidase antibody 312d6, obtained against the sulfonamide hapten 4 a that mimics the transition state for hydrolysis of a distorted amide, accelerates the hydrolysis of the corresponding amides 1 a-3 a by a factor of 10(3) at pH 8. The mechanisms of both the uncatalyzed and antibody-catalyzed reactions were studied. Between pH 8 and 12 the uncatalyzed hydrolysis of N-toluoylindoles 1 a and 3 a shows a simple first-order dependence on [OH(-)], while hydrolysis of 3 a is zeroth-order in [OH(-)] below pH 8. The pH profile for hydrolysis of the corresponding tryptophan amide 2 a is more complex due to the dissociation of the zwitterion into an anion with pK(a) 9.74; hydrolysis of the zwitterionic and the anionic form of 2 a both show simple first-order dependence on [OH(-)]. Absence of (18)O exchange between H(2) (18)O/(18)OH(-) and the substrate, a normal SKIE for both 1 a (k(H)/k(D)=1.12) and 3 a (k(H)/k(D)=1.24) and the value of the Hammett constant rho for hydrolysis of p-substituted amides 3 a-e are consistent with an ester-like mechanism in which formation of the tetrahedral intermediate is rate-determining and the amine departs as anion. The 312d6-catalyzed hydrolysis of 3 a was studied between pH 7.5 and 9, and its independence of pH in this range indicates that water is the reacting nucleophile. Hydrolysis of 3 a is only partially inhibited by the sulfonamide hapten, and this indicates that non-specific catalysis by the protein accompanies the specific process. Only the nonspecific process is observed in the hydrolysis of amides 3 with para substituents other than methyl. Binding studies on the corresponding series of p-substituted sulfonamides 5 a-e confirm the high specificity of antibody 312d6 for p-methyl substituted substrates.