Half-life extension via ABD-fusion leads to higher tumor uptake of an affibody-drug conjugate compared to PAS- and XTENylation.

A critical parameter during the development of protein therapeutics is to endow them with suitable pharmacokinetic and pharmacodynamic properties. Small protein drugs are quickly eliminated by kidney filtration, and in vivo half-life extension is therefore often desired. Here, different half-life extension technologies were studied where PAS polypeptides (PAS300, PAS600), XTEN polypeptides (XTEN288, XTEN576), and an albumin binding domain (ABD) were compared for half-life extension of an anti-human epidermal growth factor receptor 2 (HER2) affibody-drug conjugate. The results showed that extension with the PAS or XTEN polypeptides or the addition of the ABD lowered the affinity for HER2 to some extent but did not negatively affect the cytotoxic potential. The half-lives in mice ranged from 7.3 h for the construct including PAS300 to 11.6 h for the construct including PAS600. The highest absolute tumor uptake was found for the construct including the ABD, which was 60 to 160% higher than the PASylated or XTENylated constructs, even though it did not have the longest half-life (9.0 h). A comparison of the tumor-to-normal-organ ratios showed the best overall performance of the ABD-fused construct. In conclusion, PASylation, XTENylation, and the addition of an ABD are viable strategies for half-life extension of affibody-drug conjugates, with the best performance observed for the construct including the ABD.

Synthesis and Preclinical Evaluation of Urea-Based Prostate-Specific Membrane Antigen-Targeted Conjugates Labeled with 177Lu.

177Lu-labeled small-molecule prostate-specific membrane antigen (PSMA) targeted tracers are therapeutic agents for metastatic castration-resistant prostate cancer. Optimizing molecular design holds the potential to further enhance the pharmacokinetic properties of PSMA-targeted agents while preserving their potent therapeutic effects. In this study, six novel N-[N-[(S)-1,3-dicarboxypropyl]carbamoyl]-(S)-l-lysine (DCL) urea-based PSMA ligand 2,2',2″,2‴-(1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrayl)tetraacetic acid conjugates were synthesized. These conjugates feature polypeptide linkers containing the Phe-Phe peptide sequence and an aromatic fragment at the ε-NH-Lys group of the DCL fragment. The synthesis yielded products with satisfactory yields ranging from 60% to 72%, paving the way for their preclinical evaluation. The labeling of the new variants of urea-based PSMA inhibitors provided a radiochemical yield of over 95%. The 177Lu-labeled conjugates demonstrated specific and moderate affinity binding to PSMA-expressing human cancer cells PC3-pip in vitro and specific accumulation in PSMA-expressing xenografts in vivo. Based on the results, both the lipophilicity and the type of substituent in the linker significantly influence the binding properties of the PSMA inhibitor and its biodistribution profile. Specifically, the studied variants containing a bromine substituent or a hydroxyl group introduced into the aromatic fragment of the phenylalanyl residue in DCL exhibit higher affinities to PSMA compared to variants with only a chlorine-substituted aromatic fragment or variants without any substituents. The [177Lu]Lu-13C with the bromine substituent was characterized by the highest activity accumulation in blood, salivary glands, muscle, bone, and gastrointestinal tract and had inasmuch as an unfavorable pharmacokinetic profile. The negative charge of the carboxyl group in the phenyl moiety of the [177Lu]Lu-13A variant has demonstrated a positive effect on reducing the retention of activity in the liver and the kidneys (the ratio of tumor to kidneys was 1.3-fold). Low accumulation in normal tissues in vivo indicates that this novel PSMA-targeting inhibitor is a promising radioligand.

Smart Graphene Textiles for Biopotential Monitoring: Laser-Tailored Electrochemical Property Enhancement.

While most of the research in graphene-based materials seeks high electroactive surface area and ion intercalation, here, we show an alternative electrochemical behavior that leverages graphene's potential in biosensing. We report a novel approach to fabricate graphene/polymer nanocomposites with near-record conductivity levels of 45 Ω sq-1 and enhanced biocompatibility. This is realized by laser processing of graphene oxide in a sandwich structure with a thin (100 µm) polyethylene terephthalate film on a textile substrate. Such hybrid materials exhibit high conductivity, low polarization, and stability. In addition, the nanocomposites are highly biocompatible, as evidenced by their low cytotoxicity and good skin adhesion. These results demonstrate the potential of graphene/polymer nanocomposites for smart clothing applications.

Comparative Preclinical Evaluation of HYNIC-Modified Designed Ankyrin Repeat Proteins G3 for the 99mTc-Based Imaging of HER2-Expressing Malignant Tumors.

HER2 status determination is a necessary step for the proper choice of therapy and selection of patients for the targeted treatment of cancer. Targeted radiotracers such as radiolabeled DARPins provide a noninvasive and effective way for the molecular imaging of HER2 expression. This study aimed to evaluate tumor-targeting properties of three 99mTc-labeled DARPin G3 variants containing Gly-Gly-Gly-Cys (G3C), (Gly-Gly-Gly-Ser)3-Cys ((G3S)3C), or Glu-Glu-Glu-Cys (E3C) amino acid linkers at the C-terminus and conjugated to the HYNIC chelating agent, as well as to compare them with the clinically evaluated DARPin G3 labeled with 99mTc(CO)3 using the (HE)3-tag at the N-terminus. The labeling of DARPin G3-HYNIC variants provided radiochemical yields in the range of 50-80%. Labeled variants bound specifically to human HER2-expressing cancer cell lines with affinities in the range of 0.5-3 nM. There was no substantial influence of the linker and HYNIC chelator on the binding of 99mTc-labeled DARPin G3 variants to HER2 in vitro; however, [99mTc]Tc-G3-(G3S)3C-HYNIC had the highest affinity. Comparative biodistribution of [99mTc]Tc-G3-G3C-HYNIC, [99mTc]Tc-G3-(G3S)3C-HYNIC, [99mTc]Tc-G3-E3C-HYNIC, and [99mTc]Tc-(HE)3-G3 in healthy CD1 mice showed that there was a strong influence of the linkers on uptake in normal tissues. [99mTc]Tc-G3-E3C-HYNIC had an increased retention of activity in the liver and the majority of other organs compared to the other conjugates. The tumor uptake of [99mTc]Tc-G3-(G3S)3C-HYNIC and [99mTc]Tc-(HE)3-G3 in Nu/j mice bearing SKOV-3 xenografts was similar. The specificity of tumor targeting in vivo was demonstrated for both tracers. [99mTc]Tc-G3-(G3S)3C-HYNIC provided comparable, although slightly lower tumor-to-lung, tumor-to spleen and tumor-to-liver ratios than [99mTc]Tc-(HE)3-G3. Radiolabeling of DARPin G3-HYNIC conjugates with 99mTc provided the advantage of a single-step radiolabeling procedure; however, the studied HYNIC conjugates did not improve imaging contrast compared to the 99mTc-tricarbonyl-labeled DARPin G3. At this stage, [99mTc]Tc-(HE)3-G3 remains the most promising candidate for the clinical imaging of HER2-overexpressing cancers.

Nanoparticles Partially Restore Bacterial Susceptibility to Antibiotics.

The growing resistance of bacteria to antibiotics is one of the main public health problems nowadays. The influence of silver nanoparticle (AgNP) pretreatment of 220 cows with mastitis on the susceptibility of Staphylococcus epidermidis bacteria to 31 antibiotics was studied. The obtained results were compared with the previous results for Escherichia coli, Streptococcus dysgalactiae, and Staphylococcus aureus. For all four bacteria, an increase in susceptibility (9.5-21.2%) to 31 antibiotics after cow treatment with AgNPs was revealed, while after first-line antibiotic drug treatment as expected, the susceptibility decreased (11.3-27.3%). These effects were explained by (1) the increase in the contribution of isolates with efflux effect after antibiotic treatments and its decrease after AgNP treatment and (2) the changes in bacteria adhesion and anti-lysozyme activity after these treatments. The effect of the increasing antibacterial activity of antibiotics after AgNP treatment was the most pronounced in the case of E. coli and was minimal in the case of S. epidermidis. With AgNP treatment, the time of recovery decreased by 26.8-48.4% compared to the time of recovery after treatment with the first-line antibiotic drugs. The AgNP treatment allows for achieving the partial restoration of the activity of antibiotics.

Piezoelectric and Dielectric Electrospun Fluoropolymer Membranes for Oral Mucosa Regeneration: A Comparative Study.

Wound healing of the oral mucosa is an urgent problem in modern dental surgical practice. This research article presents and compares the findings of the investigations of the structural, physicochemical, and biological characteristics of two types of polymeric membranes used for the regeneration of oral mucosa. The membranes were prepared from poly(tetrafluoroethylene) (PTFE) and a copolymer of vinylidene fluoride and tetrafluoroethylene (VDF-TeFE) and analyzed via scanning electron microscopy, atomic force microscopy, X-ray diffraction analysis, and Fourier transform infrared spectroscopy. Investigation results obtained indicate that both types of membranes are composed of thin fibers: (0.57 ± 0.25) µm for PTFE membranes and (0.43 ± 0.14) µm for VDF-TeFE membranes. Moreover, the fibers of VDF-TeFE membranes exhibit distinct piezoelectric properties, which are confirmed by piezoresponse force microscopy and X-ray diffraction. Both types of membranes are hydrophobic: (139.7 ± 2.5)° for PTFE membranes and (133.5 ± 2.0)° for VDF-TeFE membranes. In vitro assays verify that both membrane types did not affect the growth and division of mice fibroblasts of the 3T3-L1 cell line, with a cell viability in the range of 88-101%. Finally, in vivo comparative experiments carried out using Wistar rats demonstrate that the piezoelectric VDF-TeFE membranes have a high ability to regenerate oral mucosa.

Evaluation of affinity matured Affibody molecules for imaging of the immune checkpoint protein B7-H3.

B7-H3 (CD276), an immune checkpoint protein, is a promising molecular target for immune therapy of malignant tumours. Sufficient B7-H3 expression level is a precondition for successful therapy. Radionuclide molecular imaging is a powerful technique for visualization of expression levels of molecular targets in vivo. Use of small radiolabelled targeting proteins would enable high-contrast radionuclide imaging of molecular targets if adequate binding affinity and specificity of an imaging probe could be provided. Affibody molecules, small engineered affinity proteins based on a non-immunoglobulin scaffold, have demonstrated an appreciable potential in radionuclide imaging. Proof-of principle of radionuclide visualization of expression levels of B7-H3 in vivo was demonstrated using the [99mTc]Tc-AC12-GGGC Affibody molecule. We performed an affinity maturation of AC12, enabling selection of clones with higher affinity. Three most promising clones were expressed with a -GGGC (triglycine-cysteine) chelating sequence at the C-terminus and labelled with technetium-99m (99mTc). 99mTc-labelled conjugates bound to B7-H3-expressing cells specifically in vitro and in vivo. Biodistribution in mice bearing B7-H3-expressing SKOV-3 xenografts demonstrated improved imaging properties of the new conjugates compared with the parental variant [99mTc]Tc-AC12-GGGC. [99mTc]Tc-SYNT-179 provided the strongest improvement of tumour-to-organ ratios. Thus, affinity maturation of B7-H3 Affibody molecules could improve biodistribution and targeting properties for imaging of B7-H3-expressing tumours.

Neuroprotective Effects of Tryptanthrin-6-Oxime in a Rat Model of Transient Focal Cerebral Ischemia.

The activation of c-Jun N-terminal kinase (JNK) plays an important role in stroke outcomes. Tryptanthrin-6-oxime (TRYP-Ox) is reported to have high affinity for JNK and anti-inflammatory activity and may be of interest as a promising neuroprotective agent. The aim of this study was to investigate the neuroprotective effects of TRYP-Ox in a rat model of transient focal cerebral ischemia (FCI), which involved intraluminal occlusion of the left middle cerebral artery (MCA) for 1 h. Animals in the experimental group were administered intraperitoneal injections of TRYP-Ox 30 min before reperfusion and 23 and 47 h after FCI. Neurological status was assessed 4, 24, and 48 h following FCI onset. Treatment with 5 and 10 mg/kg of TRYP-Ox decreased mean scores of neurological deficits by 35-49 and 46-67% at 24 and 48 h, respectively. At these doses, TRYP-Ox decreased the infarction size by 28-31% at 48 h after FCI. TRYP-Ox (10 mg/kg) reduced the content of interleukin (IL) 1ß and tumor necrosis factor (TNF) in the ischemic core area of the MCA region by 33% and 38%, respectively, and attenuated cerebral edema by 11% in the left hemisphere, which was affected by infarction, and by 6% in the right, contralateral hemisphere 24 h after FCI. TRYP-Ox reduced c-Jun phosphorylation in the MCA pool at 1 h after reperfusion. TRYP-Ox was predicted to have high blood-brain barrier permeability using various calculated descriptors and binary classification trees. Indeed, reactive oxidant production was significantly lower in the brain homogenates from rats treated with TRYP-Ox versus that in control animals. Our data suggest that the neuroprotective activity of TRYP-Ox may be due to the ability of this compound to inhibit JNK and exhibit anti-inflammatory and antioxidant activity. Thus, TRYP-Ox may be considered a promising neuroprotective agent that potentially could be used for the development of new treatment strategies in cerebral ischemia.

Synthesis, 123I-Radiolabeling Optimization, and Initial Preclinical Evaluation of Novel Urea-Based PSMA Inhibitors with a Tributylstannyl Prosthetic Group in Their Structures.

Prostate-specific membrane antigen (PSMA) has been identified as a target for the development of theranostic agents. In our current work, we describe the design and synthesis of novel N-[N-[(S)-1,3-dicarboxypropyl]carbamoyl]-(S)-L-lysine (DCL) urea-based PSMA inhibitors with a chlorine-substituted aromatic fragment at the lysine ε-nitrogen atom, a dipeptide including two phenylalanine residues in the L-configuration as the peptide fragment of the linker, and 3- or 4-(tributylstannyl)benzoic acid as a prosthetic group in their structures for radiolabeling. The standard compounds [127I]PSMA-m-IB and [127I]PSMA-p-IB for comparative and characterization studies were first synthesized using two alternative synthetic approaches. An important advantage of the alternative synthetic approach, in which the prosthetic group (NHS-activated esters of compounds) is first conjugated with the polypeptide sequence followed by replacement of the Sn(Bu)3 group with radioiodine, is that the radionuclide is introduced in the final step of synthesis, thereby minimizing operating time with iodine-123 during the radiolabeling process. The obtained DCL urea-based PSMA inhibitors were radiolabeled with iodine-123. The radiolabeling optimization results showed that the radiochemical yield of [123I]PSMA-p-IB was higher than that of [123I]PSMA-m-IB, which were 74.9 ± 1.0% and 49.4 ± 1.2%, respectively. The radiochemical purity of [123I]PSMA-p-IB after purification was greater than 99.50%. The initial preclinical evaluation of [123I]PSMA-p-IB demonstrated a considerable affinity and specific binding to PC-3 PIP (PSMA-expressing cells) in vitro. The in vivo biodistribution of this new radioligand [123I]PSMA-p-IB showed less accumulation than [177Lu]Lu-PSMA-617 in several normal organs (liver, kidney, and bone). These results warrant further preclinical development, including toxicology evaluation and experiments in tumor-bearing mice.

Textile Electronics with Laser-Induced Graphene/Polymer Hybrid Fibers.

The concept of wearables is rapidly evolving from flexible polymer-based devices to textile electronics. The reason for this shift is the ability of textiles to ensure close contact with the skin, resulting in comfortable, lightweight, and compact "always with you" sensors. We are contributing to this polymer-textile transition by introducing a novel and simple way of laser intermixing of graphene with synthetic fabrics to create wearable sensing platforms. Our hybrid materials exhibit high electrical conductivity (87.6 ± 36.2 Ω/sq) due to the laser reduction of graphene oxide and simultaneous laser-induced graphene formation on the surface of textiles. Furthermore, the composite created between graphene and nylon ensures the durability of our materials against sonication and washing with detergents. Both of these factors are essential for real-life applications, but what is especially useful is that our free-form composites could be used as-fabricated without encapsulation, which is typically required for conventional laser-scribed materials. We demonstrate the exceptional versatility of our new hybrid textiles by successfully recording muscle activity, heartbeat, and voice. We also show a gesture sensor and an electrothermal heater embedded within a single commercial glove. Additionally, the use of these textiles could be extended to personal protection equipment and smart clothes. We achieve this by implementing self-sterilization with light and laser-induced functionalization with silver nanoparticles, which results in multifunctional antibacterial textiles. Moreover, incorporating silver into such fabrics enables their use as surface-enhanced Raman spectroscopy sensors, allowing for the direct analysis of drugs and sweat components on the clothing itself. Our research offers valuable insights into simple and scalable processes of textile-based electronics, opening up new possibilities for paradigms like the Internet of Medical Things.

Revealing the Second and the Third Causes of AgNPs Property to Restore the Bacterial Susceptibility to Antibiotics.

The increase in bacterial resistance to antibiotics is a global problem for public health. In our previous works, it was shown that the application of AgNPs in cow mastitis treatment increased S. aureus and S. dysgalactiae susceptibility to 31 antibiotics due to a decrease in the bacterial efflux effect. The aim of the present work was to shed light on whether the change in adhesive and anti-lysozyme activities caused by AgNPs also contribute to the restoration of bacterial susceptibility to antibiotics. In vivo sampling was performed before and after cow mastitis treatments with antibiotics or AgNPs. The isolates were identified, and the adhesive and anti-lysozyme activities were assessed. These data were compared with the results obtained for in vitro pre-treatment of reference bacteria with AgNPs or antibiotics. The present study revealed that bacterial treatments in vitro and in vivo with AgNPs: (1) decrease the bacterial ability to adhere to cells to start an infection and (2) decrease bacterial anti-lysozyme activity, thereby enhancing the activity of lysozyme, a natural "antibiotic" present in living organisms. The obtained data contribute to the perspective of the future application of AgNPs for recovering the activity of antibiotics rapidly disappearing from the market.

Lithium and Microorganisms: Biological Effects and Mechanisms.

This review covers the lithium effects on microorganisms, including gut and soil bacteria. Available studies of the biological effects of lithium salts have revealed a wide range of different effects of lithium cations on various microorganisms, but so far, the study of this direction has not been summarized enough. Here we consider the confirmed and various plausible mechanisms of lithium action on microorganisms. Special emphasis is placed on assessing the effect of lithium ions under oxidative stress and adverse environmental conditions. The impact of lithium on the human microbiome is also being reviewed and discussed. Controversial effects of lithium have been shown, including the inhibitory and stimulating effects of lithium on bacterial growth.

A Comparative Study of Cancer Cells Susceptibility to Silver Nanoparticles Produced by Electron Beam.

INTRODUCTION: Silver nanoparticles (AgNPs) have a wide range of bioactivity, which is highly dependent on particle size, shape, stabilizer, and production method. Here, we present the results of studies of AgNPs cytotoxic properties obtained by irradiation treatment of silver nitrate solution and various stabilizers by accelerating electron beam in a liquid medium. METHODS: The results of studies of morphological characteristics of silver nanoparticles were obtained by transmission electron microscopy, UV-vis spectroscopy, and dynamic light scattering measurements. MTT test, alamar blue test, flow cytometry, and fluorescence microscopy were used to study the anti-cancer properties. As biological objects for standard tests, adhesive and suspension cell cultures of normal and tumor origin, including prostate cancer, ovarian cancer, breast cancer, colon cancer, neuroblastoma, and leukemia, were studied. RESULTS: The results showed that the silver nanoparticles obtained by irradiation with polyvinylpyrrolidone and collagen hydrolysate are stable in solutions. Samples with different stabilizers were characterized by a wide average size distribution from 2 to 50 nm and low zeta potential from -7.3 to +12.4 mV. All AgNPs formulations showed a dose-dependent cytotoxic effect on tumor cells. It has been established that the particles obtained with the combination of polyvinylpyrrolidone/collagen hydrolysate have a relatively more pronounced cytotoxic effect in comparison to samples stabilized with only collagen or only polyvinylpyrrolidone. The minimum inhibitory concentrations for nanoparticles were less than 1 µg/mL for various types of tumor cells. It was found that neuroblastoma (SH-SY5Y) is the most susceptible, and ovarian cancer (SKOV-3) is the most resistant to the action of silver nanoparticles. The activity of the AgNPs formulation prepared with a mixture of PVP and PH studied in this work was higher that activity of other AgNPs formulations reported in the literature by about 50 times. CONCLUSIONS: The results indicate that the AgNPs formulations synthesized with an electron beam and stabilized with polyvinylpyrrolidone and protein hydrolysate deserve deep study for their further use in selective cancer treatment without harming healthy cells in the patient organism.

Molecular Plasmonic Silver Forests for the Photocatalytic-Driven Sensing Platforms.

Structural electronics, as well as flexible and wearable devices are applications that are possible by merging polymers with metal nanoparticles. However, using conventional technologies, it is challenging to fabricate plasmonic structures that remain flexible. We developed three-dimensional (3D) plasmonic nanostructures/polymer sensors via single-step laser processing and further functionalization with 4-nitrobenzenethiol (4-NBT) as a molecular probe. These sensors allow ultrasensitive detection with surface-enhanced Raman spectroscopy (SERS). We tracked the 4-NBT plasmonic enhancement and changes in its vibrational spectrum under the chemical environment perturbations. As a model system, we investigated the sensor's performance when exposed to prostate cancer cells' media over 7 days showing the possibility of identifying the cell death reflected in the environment through the effects on the 4-NBT probe. Thus, the fabricated sensor could have an impact on the monitoring of the cancer treatment process. Moreover, the laser-driven nanoparticles/polymer intermixing resulted in a free-form electrically conductive composite that withstands over 1000 bending cycles without losing electrical properties. Our results bridge the gap between plasmonic sensing with SERS and flexible electronics in a scalable, energy-efficient, inexpensive, and environmentally friendly way.

Sweetened Alkylated Verdazyls Effectively Kill Cancer Cells under Light Irradiation.

The development of novel photo-dynamic therapy (PDT) agents enabling to treat the oxygen-deficient tumors is the emerging tasks for the modern medicinal chemistry. Herein, we describe the design and preparation of water-soluble agents for PDT which generate active radical species upon light irradiation. Two conjugates of carbohydrates with 1,2,4,6-substituted-1,4-dihydro-1,2,4,5-tetrazin-3(2H)-ones (AlkVZs) demonstrated high oxygen-independent cytotoxicity on PC-3 and Jurkat cancer cells under light irradiation combining with low toxicity in the dark. The efficacy of prepared compounds was estimated using MTT and Alamar Blue tests as well as microscopic dead/live colored images and flow cytometry. The analysis of obtained results reveals the influence of sugar moiety on the activity of AlkVZs. We believe that obtained compounds have high potency as platform for design of new agents for photo-dynamic therapy.

Mathematical Model of Antibiotic Resistance Determinants' Stability Under Space Flight Conditions.

Increasing antibiotic resistance (AR) poses dangers of treatment complications and even treatment failure to astronauts. An AR determinant is a gene of resistance carried by bacteria. This article considers the issue of the stability of AR determinants and the influence of manned spaceflight conditions on this characteristic. A phenomenological model has been developed that makes it possible to evaluate the integral value of the stability of determinants of AR in bacteria as a function of time. Based on experimental results obtained during implementation of the SALYUT 7 space program, the stability of determinants of AR in Escherichia coli strains isolated before and after a spaceflight in 16 astronauts was evaluated. In addition, an assessment was made of the integral value of the stability of determinants of AR in bacteria during in vitro experiments, both in spaceflight and terrestrial conditions, after preincubation in space. The calculation using the developed phenomenological model showed that the stability of AR determinants in E. coli bacteria isolated from astronauts before the spaceflight is 33% higher than after the flight. The in vitro experiment carried out on board the International Space Station showed the opposite situation-an increase in the stability of AR determinants by 33% in cultures that have been in space compared with terrestrial control. This indicates an additional influence on the stability of determinants and of the astronaut's immune system, as well as space conditions. The common result in these two types of studies is the experimental fact that the largest number of bacteria, in space conditions, had two determinants of AR. The importance of fighting bacteria with two determinants is that at least three different antibiotics are required to have an effect. This circumstance makes it possible to predict a possible strategy for the use of antibiotics in autonomous spaceflights.

Comparative Preclinical Evaluation of Peptide-Based Chelators for the Labeling of DARPin G3 with 99mTc for Radionuclide Imaging of HER2 Expression in Cancer.

Non-invasive radionuclide imaging of human epidermal growth factor receptor type 2 (HER2) expression in breast, gastroesophageal, and ovarian cancers may stratify patients for treatment using HER2-targeted therapeutics. Designed ankyrin repeat proteins (DARPins) are a promising type of targeting probe for radionuclide imaging. In clinical studies, the DARPin [99mTc]Tc-(HE)3-G3 labeled using a peptide-based chelator His-Glu-His-Glu-His-Glu ((HE)3), provided clear imaging of HER2 expressing breast cancer 2-4 h after injection. The goal of this study was to evaluate if the use of cysteine-containing peptide-based chelators Glu-Glu-Glu-Cys (E3C), Gly-Gly-Gly-Cys (G3C), and Gly-Gly-Gly-Ser-Cys connected via a (Gly-Gly-Gly-Ser)3-linker (designated as G3-(G3S)3C) would further improve the contrast of imaging using 99mTc-labeled derivatives of G3. The labeling of the new variants of G3 provided a radiochemical yield of over 95%. Labeled G3 variants bound specifically to human HER2-expressing cancer cell lines with affinities in the range of 1.9-5 nM. Biodistribution of [99mTc]Tc-G3-G3C, [99mTc]Tc-G3-(G3S)3C, and [99mTc]Tc-G3-E3C in mice was compared with the biodistribution of [99mTc]Tc-(HE)3-G3. It was found that the novel variants provide specific accumulation in HER2-expressing human xenografts and enable discrimination between tumors with high and low HER2 expression. However, [99mTc]Tc-(HE)3-G3 provided better contrast between tumors and the most frequent metastatic sites of HER2-expressing cancers and is therefore more suitable for clinical applications.

Pulsed Vacuum Arc Deposition of Nitrogen-Doped Diamond-like Coatings for Long-Term Hydrophilicity of Electrospun Poly(ε-caprolactone) Scaffolds.

The surface hydrophobicity of poly(ε-caprolactone) electrospun scaffolds prevents their interactions with cells and tissue integration. Although plasma treatment of scaffolds enhances their hydrophilicity, this effect is temporary, and the hydrophobicity of the scaffolds is restored in about 30 days. In this communication, we report a method for hydrophilization of poly(ε-caprolactone) electrospun scaffolds for more than 6 months. To that end, diamond-like coating was deposited on the surface of the scaffolds in a nitrogen atmosphere using pulsed vacuum arc deposition with sputtering of graphite target. This approach allows for a single-side hydrophilization of the scaffold (water contact angle of 22 ± 3° vs. 126 ± 2° for pristine PCL scaffold) and preserves its structure. With increased nitrogen pressure in the chamber, sp3-hybridized carbon content decreased twice (sp2/sp3 ratio decreased from 1.06 to 0.52), which demonstrates the possibility of tailoring the content of carbon in sp2 and sp3 hybridization state. Nitrogen content in the deposited coatings was found at 16.1 ± 0.9 at.%. In vitro tests with fibroblast cell culture did not reveal any cytotoxic compounds in sample extracts.

Neuroprotective Effects of the Lithium Salt of a Novel JNK Inhibitor in an Animal Model of Cerebral Ischemia-Reperfusion.

The c-Jun N-terminal kinases (JNKs) regulate many physiological processes, including inflammatory responses, morphogenesis, cell proliferation, differentiation, survival, and cell death. Therefore, JNKs represent attractive targets for therapeutic intervention. In an effort to develop improved JNK inhibitors, we synthesized the lithium salt of 11H-indeno[1,2-b]quinoxaline-11-one oxime (IQ-1L) and evaluated its affinity for JNK and biological activity in vitro and in vivo. According to density functional theory (DFT) modeling, the Li+ ion stabilizes the six-membered ring with the 11H-indeno[1,2-b]quinoxaline-11-one (IQ-1) oximate better than Na+. Molecular docking showed that the Z isomer of the IQ-1 oximate should bind JNK1 and JNK3 better than (E)-IQ-1. Indeed, experimental analysis showed that IQ-1L exhibited higher JNK1-3 binding affinity in comparison with IQ-1S. IQ-1L also was a more effective inhibitor of lipopolysaccharide (LPS)-induced nuclear factor-κB/activating protein 1 (NF-κB/AP-1) transcriptional activity in THP-1Blue monocytes and was a potent inhibitor of proinflammatory cytokine production by MonoMac-6 monocytic cells. In addition, IQ-1L inhibited LPS-induced c-Jun phosphorylation in MonoMac-6 cells, directly confirming JNK inhibition. In a rat model of focal cerebral ischemia (FCI), intraperitoneal injections of 12 mg/kg IQ-1L led to significant neuroprotective effects, decreasing total neurological deficit scores by 28, 29, and 32% at 4, 24, and 48 h after FCI, respectively, and reducing infarct size by 52% at 48 h after FCI. The therapeutic efficacy of 12 mg/kg IQ-1L was comparable to that observed with 25 mg/kg of IQ-1S, indicating that complexation with Li+ improved efficacy of this compound. We conclude that IQ-1L is more effective than IQ-1S in treating cerebral ischemia injury and thus represents a promising anti-inflammatory compound.

Evaluation of an Affibody-Based Binder for Imaging of Immune Check-Point Molecule B7-H3.

Radionuclide molecular imaging could provide an accurate assessment of the expression of molecular targets in disseminated cancers enabling stratification of patients for specific therapies. B7-H3 (CD276) is a transmembrane protein belonging to the B7 superfamily. This protein is overexpressed in different types of human malignancies and such upregulation is generally associated with a poor clinical prognosis. In this study, targeting properties of an Affibody-based probe, AC12, containing a -GGGC amino acid sequence as a chelator (designated as AC12-GGGC) labelled with technetium-99m (99mTc) were evaluated for imaging of B7-H3-expressing tumours. AC12-GGGC was efficiently labelled with 99mTc. [99mTc]Tc-AC12-GGGC bound specifically to B7-H3 expressing cells in vitro with affinities in nanomolar range. In mice bearing B7-H3-expressing xenografts, [99mTc]Tc-AC12-GGGC showed tumour uptake of 2.1 ± 0.5 %ID/g at 2 h after injection. Its clearance from blood, normal organs and tissues was very rapid. This new targeting agent, [99mTc]Tc-AC12-GGGC, provided high tumour-to-blood ratio already at 2 h (8.2 ± 1.9), which increased to 11.0 ± 0.5 at 4 h after injection. Significantly (p < 0.05) higher tumour-to-liver and higher tumour-to-bone ratios at 2 h in comparison with 4 h after injection were observed. Thus, [99mTc]Tc-AC12-GGGC could be a promising candidate for further development.