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In this study, we selected 12 guanidine derivatives from the previously described ligand library and determined their affinity at histamine H 3 and H 4 receptors (H 3 R and H 4 R, respectively). Moreover, we also checked their intrinsic activity toward H 3 R and muscarinic M 1 , M 2 , and M 4 receptors (M 1 R, M 2 R, and M 4 R, respectively). Since ADS1017 has been proved to be the most selective and highly potent H 3 antagonist in our series, we chose it as the lead structure for further biological evaluation. To extend the study of its in vivo efficacy, we proposed an alternative synthetic route that resulted in an increased yield. Interestingly, ADS1017 showed a broad spectrum of analgesic activity in both nociceptive and neuropathic pain models. Finally, as a result of comprehensive analysis of its off-target activity and ADMETox parameters, we confirmed the moderate selectivity of ADS1017 and its promising drug-like properties.

The rise of multidrug-resistant (MDR) bacteria poses a substantial challenge in clinical settings, particularly with the increasing prevalence of ESKAPE pathogens ( E. faecium , S. aureus , K. pneumoniae , A. baumannii , P. aeruginosa , and E. coli ) as critical MDR bacteria. These ESKAPE pathogens have the capability to undermine antibiotic treatments, leading to a high incidence of drug resistance. However, the development of efficient antibacterial agents against ESKAPE pathogens is still in the bottleneck. Herein, the first example of antibacterial carbon dots against ESKAPE pathogens was reported. Onion powder-based carbon dots were melted with poly(hexamethylene biguanide) hydrochloride (PHMB) to obtain guanidinium-functionalized carbon dots (GCDs), which exhibited satisfactory antibacterial activity against all the tested bacteria, including both Gram-positive and Gram-negative bacteria, and even ESKAPE pathogens. The efficient antibacterial ability of GCDs derives from the rupture of the bacterial cell membrane and elevated ROS levels. Safety assessments revealed that GCDs neither trigger detectable drug resistance nor exhibit any cytotoxic effects. Furthermore, GCDs effectively promoted wound healing without observable adverse reactions of mixed MDR bacteria-infected wounds in rats. The GCDs also showed excellent long-term stability. These findings indicate that GCDs hold promise as an efficient antibacterial agent for the treatment of MDR strain-caused clinical infected-wound healing.

Amyloid-β (Aβ) is considered a primary therapeutic target for Alzheimer's disease (AD). However, just eliminating Aβ in patients with AD has exhibited restricted clinical efficacy, possibly failing to address the metabolic abnormalities caused by AD, such as insulin resistance. To address this concern, our research has employed two types of macrocyclic amphiphiles, guanidinium-modified calixarene and cyclodextrin coassembly (GCD), as delivery systems for insulin. This approach aimed to tackle the metabolic dysregulation characteristic of AD in an innovative manner by exploring beyond the conventional strategy of Aβ removal. As a result, GCD and insulin coassembly could effectively improve plaque deposition and insulin resistance. The coassembly could also reduce abnormal neuronal apoptosis and synaptic damage and improve cognitive impairment in 5xFAD mice. Therefore, the GCD and insulin coassembly shows promise as a viable therapeutic option for AD.

Riboswitches represent a class of non-coding RNA that possess the unique ability to specifically bind ligands and, in response, regulate gene expression. A recent report unveiled a type of riboswitch, known as the guanidine-IV riboswitch, which responds to guanidine levels to regulate downstream genetic transcription. However, the precise molecular mechanism through which the riboswitch senses its target ligand and undergoes conformational changes remain elusive. This gap in understanding has impeded the potential applications of this riboswitch. To bridge this knowledge gap, our study investigated the conformational dynamics of the guanidine-IV riboswitch RNA upon ligand binding. We employed single-molecule fluorescence resonance energy transfer (smFRET) to dissect the behaviors of the aptamer, terminator, and full-length riboswitch. Our findings indicated that the aptamer portion exhibited higher sensitivity to guanidine compared to the terminator and full-length constructs. Additionally, we utilized Position-specific Labelling of RNA (PLOR) combined with smFRET to observe, at the single-nucleotide and single-molecule level, the structural transitions experienced by the guanidine-IV riboswitch during transcription. Notably, we discovered that the influence of guanidine on the riboswitch RNA's conformations was significantly reduced after the transcription of 88 nucleotides. Furthermore, we proposed a folding model for the guanidine-IV riboswitch in the absence and presence of guanidine, thereby providing insights into its ligand-response mechanism., Competing Interests: LG, DC, YL No competing interests declared, (© 2024, Gao et al.)

There are some natural products from plants that can prevent and treat disease. Metformin, a derivative of galegine, is the basic drug to treat diabetes. Moreover, this molecule has anticancer properties that inhibit cancer cell growth and proliferation. In this study, the main interactions of galegine and metformin with various cancer-involved proteins, including mitochondrial alpha-glycerophosphate dehydrogenase, yeast NADH dehydrogenase, and transforming growth factor-β1, were surveyed by molecular docking and molecular dynamics simulations. The results showed that each of the proteins makes complexes with the ligands via favorable non-bonded interactions, especially hydrogen bond interactions. There is greater stability for complexes containing galegine based on the root mean square deviation results. The higher structure compactness is also found in galegine receptors than in metformin receptors. Calculation of ΔG binding , using the MM/PBSA methodology, shows that the binding energy values for metformin and galegine in interaction with each of the receptors are almost the same, and galegine has similar binding properties with metformin in interaction with the studied protein receptors. Therefore, galegine, a natural ingredient with better binding properties to cancer-involved proteins than metformin (with various side effects), can be applied as a new drug for cancer treatment., (© 2024 International Union of Biochemistry and Molecular Biology, Inc.)

Dicyandiamide (DCD) reacted with amino acids 1a-f to produce biguanides 2 and 4 and guanidine pyrazolones 3, 5, 6, 7, and 8, according to the reaction. DCD exhibited the following reactions: imidodicarbonimidicdiamide 9, diazocan-2-ylguanidine 10, methyl biguanidylthion 11, N-carbamothioylimidodicarbonimidicdiamide 12, 2-guanidinebenzoimidazole 13a, 2-guanidinylbenzoxazole 13b, and 2-guanidinylbenzothiazol 13c. These reactions were triggered by 6-amino caproic acid, thioacetamide, thiourea, o-aminophenol, o-aminothiophenol, and anthranilic acid, respectively. Compound 2 had the least antimicrobial activity, while compound 13c demonstrated the most antibacterial impact against all bacterial strains. Furthermore, in terms of antiglycation efficacy (AGEs), 12, 11, and 7 were the most effective AGE cross-linking inhibitors. Eight and ten, which showed a considerable inhibition on cross-linking AGEs, come next. Compounds 4 and 6 on the other hand have shown the least suppression of AGE production. The most promising antiglycation scaffolds 8, 11, and 12 in the Human serum albumin (HAS) active site were shown to be able to adopt crucial binding interactions with important amino acids based on the results of in silico molecular docking. The most promising antiglycation compounds 8, 11, and 12 were also shown to have better hydrophilicity, acceptable lipophilicity, gastrointestinal tract absorption (GIT), and blood-brain barrier penetration qualities when their physicochemical properties were examined using the egg-boiled method., Competing Interests: Declarations. Conflict of interest: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Ethical approval: This study has been carried on rabbit and the authors take personal responsibility for knowing their statutory responsibility under the Animal (Scientific Procedures) Act 1986, under the acceptance of ManchesterMetropolitanUniversity and Sohag University., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Vancomycin and norvancomycin have diminished antibacterial efficacy due to acquired or intrinsic resistance from mutations in the terminal dipeptide of lipid II in Gram-positive bacteria or failure to penetrate into the periplasm in Gram-negative bacteria. Herein, we rationally designed and synthesized a series of vancomycin analogues bearing single amine or guanidine functionality, altering various linkers and modification sites, to combat the resistance. Extensive antibacterial screening was performed to delineate a comprehensive SAR. Many derivatives revitalized the activity in vitro, exhibiting a 4-128-fold or 2-16-fold enhancement against the acquired or intrinsic resistance with lower toxicity. Significantly, the optimal compound 4g demonstrated greater pharmacokinetic and pharmacodynamic profiles. Further studies uncovered additional independent and synergistic mechanisms for 4g , including the enhanced membrane activity and augmented inhibition of peptidoglycan biosynthesis via increased lipid II binding, highlighting its potential as a future lead candidate to replenish the glycopeptide antibiotic arsenal.

Diabetes mellitus (DM) is a chronic metabolic disease characterized by hyperglycemia. Type 2 diabetes mellitus (T2DM) represents approximately 90 % of all DM cases and is primarily caused by an imbalance in blood glucose homeostasis due to inadequate insulin secretion or insulin resistance. This study explores the potential therapeutic effects of chitosan guanidine (CSG) on a T2DM mouse model. The findings reveal that CSG significantly enhances oral glucose tolerance (OGTT) and insulin sensitivity (ITT), reduces fasting blood glucose (FBG) levels, and suppresses the expression of proinflammatory cytokines in T2DM mice. These changes improve insulin resistance and diminish inflammation. Additionally, CSG markedly ameliorates lipid metabolism disorders, lowers total cholesterol (TC) and triglyceride (TG) levels, and inhibits hepatic fat accumulation. 16S rRNA and Spearman correlation analyses indicate that CSG promotes the relative abundance of probiotic genera such as Bacteroidota, Patescibacteria, Actinobacteria, and Cyanobacteria. These bacteria are positively correlated with short-chain fatty acids (SCFAs) and high-density lipoprotein cholesterol (HDLC) levels. Conversely, CSG reduces the relative abundance of pathogenic bacteria, including Proteobacteria and Ralstonia, leading to an improved intestinal microbial community composition in T2DM mice and alleviating T2DM symptoms. These results suggest that CSG holds significant potential as a non-insulin therapeutic agent for diabetes management., Competing Interests: Declaration of competing interest We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled. We confirm that the mentioned received grants in the “Acknowledgement” section, did not lead to any conflicts of interest regarding the publication of this manuscript., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The supramolecular palladium complex G-Pd, formed via self-assembly of the Pd-complex of guanidinium pillar[5]arene with Pd 2+ , was used to encapsulate doxorubicin to form G-Pd@DOX. The nanoparticles exhibit responsiveness to glutathione, controlled drug release, the ability to damage mitochondria, and potent anticancer activity while maintaining low toxicity towards normal cells. This work provides a good example for the application of pillararene-based palladium complexes in cancer therapy and is significant for the discovery of new medicines from supramolecular coordination complexes.

Over last years, hydrogels based on natural polymers have attracted considerable interest as materials for wound healing. Herein, hydrogel films based on kappa-carrageenan and guanidinium polyampholytes were prepared by the in situ physical cross-linking with potassium chloride and borax, respectively. The polyampholytes were obtained by a free radical copolymerization of 2,2-diallyl-1,1,3,3-tetraethylguanidinium chloride and unsaturated acids. To characterize the composite films, NMR, FTIR, SEM, TGA, XRD, element analysis and tensile test were used. Ampicillin was incorporated into the hydrogels to enhance wound healing potential. The healing-related characteristics, including swelling ratio, drug release and antimicrobial activity, were assessed. The equilibrium swelling ratios were in the range of 3.9-6.5 depending on the polyampholyte composition. According to the in vitro ampicillin release studies, 30-43 % of ampicillin was released from the hydrogels after 5 h at 37 °C and pH 7.4, with drug release being temperature and pH dependent. The ampicillin-loaded films showed a remarkable antimicrobial effect. The inhibition sizes for Escherichia coli and Staphylococcus aureus were 1.10-1.85 and 1.95-2.60 cm, respectively. Although the bi-polymeric hydrogels were thoroughly characterized, with the in vitro study of their biocidal effects carried out in this work, the in vivo drug release assessment needs to be further explored., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Marina Gorbunova reports financial support was provided by Russian Science Foundation. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Biofouling has been persisting as a global problem due to the difficulties in finding efficient and environmentally friendly antifouling coatings for long-term applications. Initial attachment of bacteria on material surface and subsequent formation of biofilm are the predominate phenomena accounting for subsequent occurrence of biofouling. Among the various factors influencing the bacterial attachment, conditioning layer formed by organic macromolecules usually plays the key role in mediating bacterial attachment through altering physicochemical properties of substrate surface. In this study, a guanidine-modified polysaccharide conditioning layer with the capability of tuning the bacterial attachment is constructed and characterized. Dextran, a polysaccharide widespread in bacteria extracellular polymeric substances (EPS), is oxidized by sodium periodate, and cationic polymer polyhexamethylene guanidine hydrochloride (PHMG) is anchored to oxidized dextran (ODEX) by Schiff base reaction. AFM characterization reveals morphological changes of the polysaccharide conditioning layer from tangled chain to island conformation after the PHMG modification. The guanidine-based dextran conditioning layer promotes attachment of both P. aeruginosa and S. aureus and disrupted bacterial cytomembranes are seen for the attached bacteria due to electrostatic interaction of the electropositive guanidine group with the electronegative bacteria. The guanidine-based dextran conditioning layer shows a low survival ratio of 22 %-34 % and 1 %-4 % for P. aeruginosa and S. aureus respectively after incubation in the bacterial suspension for 72 hours. The results would give insight into further exploring the potential applications of the newly designed polysaccharides conditioning layer for combating occurrence of biofouling., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper, (Copyright © 2024 Elsevier B.V. All rights reserved.)

Colistin (CST) is considered as "agent of last resort" against gram-negative bacteria as feed additive. Its clinical effectiveness has reduced since the emergence of mcr-1 gene in ducks. Isopropoxy benzene guanidine (IBG), a new guanidine derivative, showed positive effects on improving animal weights and alleviating intestinal pathogens, therefore, the objective of this study was to evaluate the effect of this compound supplement with CST in ducks and explore the possibilities in feed additive. A total of fifteen duck-origin Escherichia coli carrying the mcr-1 gene were included in this study. A checkerboard microdilution assay was used to evaluate the in vitro antibacterial activity of IBG combined with CST against mcr-1-positive E. coli. A 3-by-2 time-kill array of IBG (16, 32, and 64 μg/mL) and CST (1/2 MIC and 1/4 MIC) over 24 hours was utilized to characterize the activity of the agents alone and in combination against E. coli strain 1 in vitro. The intestinal colonization model was used to evaluate the in vivo effect of IBG combined with CST. These results indicated that the combination of IBG plus CST showed a synergistic effect against all clinical isolates (FICI < 0.5). The bacterial burden was reduced by more than 2 log 10 CFU/mL when E. coli strain 1 was tested with 1/2 MIC CST plus 64 μg/mL IBG for 24 h. Further experiments in vivo demonstrated that the CST combined with IBG was able to increase duck weights, reduced intestinal pathogenic E. coli and showed a synergistic antibacterial effect. Combination of CST (4 mg/kg b.w.) plus IBG (32 or 64 mg/kg b.w.) achieved 1.84 to 3.29 log 10 CFU/g killing after 7 d of therapy, which was significantly different from that in the challenge control group (p<0.05). In summary, our study demonstrated the potential use of IBG as feed additive for veterinary purposes in ducks and provided new insights into overcoming resistance in the future., Competing Interests: DISCLOSURES The authors declare no conflicts of interest., (Copyright © 2024. Published by Elsevier Inc.)

By performing differential scanning calorimetry(DSC) measurements on RNase A, we studied the stabilization provided by the addition of potassium aspartate(KAsp) or potassium glutamate (KGlu) and found that it leads to a significant increase in the denaturation temperature of the protein. The stabilization proves to be mainly entropic in origin. A counteraction of the stabilization provided by KAsp or KGlu is obtained by adding common denaturants such as urea, guanidinium chloride, or guanidinium thiocyanate. A rationalization of the experimental data is devised on the basis of a theoretical approach developed by one of the authors. The main contribution to the conformational stability of globular proteins comes from the gain in translational entropy of water and co-solute ions and/or molecules for the decrease in solvent-excluded volume associated with polypeptide folding (i.e., there is a large decrease in solvent-accessible surface area). The magnitude of this entropic contribution increases with the number density and volume packing density of the solution. The two destabilizing contributions come from the conformational entropy of the chain, which should not depend significantly on the presence of co-solutes, and from the direct energetic interactions between co-solutes and the protein surface in both the native and denatured states. It is the magnitude of the latter that discriminates between stabilizing and destabilizing agents.

Dynamic covalent polymers (DCPs) that strike a balance between high performance and rapid reconfiguration have been a challenging task. For this purpose, a solution is proposed in the form of a new dynamic covalent supramolecular motif-guanidine urea structure (GUAs). GUAs contain complex and diverse chemical structures as well as unique bonding characteristics, allowing guanidine urea supramolecular polymers to demonstrate advanced physical properties. Noncovalent interaction aggregates (NIAs) have been confirmed to form in GUA-DCPs through multistage H-bonding and π-π stacking, resulting in an extremely high Young's modulus of 14 GPa, suggesting remarkable mechanical strength. Additionally, guanamine urea linkages in GUAs, a new type of dynamic covalent bond, provide resins with excellent malleability and reprocessability. Guanamine urea metathesis is validated using small molecule model compounds, and the temperature dependent infrared and rheological behavior of GUA-DCPs following the dissociative exchange mechanism. Moreover, the inherent photodynamic antibacterial properties are extensively verified by antibacterial experiments. Even after undergoing three reprocessing cycles, the antibacterial rate of GUA-DCPs remains above 99% after 24 h, highlighting their long-lasting antibacterial effectiveness. GUA-DCPs with dynamic nature, tuneable composition, and unique combination of properties make them promising candidates for various technological advancements., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

Biofilm-associated infections remain a tremendous obstacle to the treatment of microbial infections globally. However, the poor penetrability to a dense extracellular polymeric substance matrix of traditional antibacterial agents limits their antibiofilm activity. Here, we show that nanoaggregates formed by self-assembly of amphiphilic borneol-guanidine-based cationic polymers (BGN x - n ) possess strong antibacterial activity and can eliminate mature Staphylococcus aureus ( S. aureus ) biofilms. The introduction of the guanidine moiety improves the hydrophilicity and membrane penetrability of BGN x - n . The self-assembled nanoaggregates with highly localized positive charges are expected to enhance their interaction with negatively charged bacteria and biofilms. Furthermore, nanoaggregates dissociate on the surface of biofilms into smaller BGN x - n polymers, which enhances their ability to penetrate biofilms. BGN x - n nanoaggregates that exhibit superior antibacterial activity have the minimum inhibitory concentration (MIC) of 62.5 μg·mL -1 against S. aureus and eradicate mature biofilms at 4 × MIC with negligible hemolysis. Taken together, this size-variable self-assembly system offers a promising strategy for the development of effective antibiofilm agents.

Parasite-derived new permeation pathways (NPPs) expressed at the red blood cell (RBC) membrane enable Plasmodium parasites to take up nutrients from the plasma to facilitate their survival. Thus, NPPs represent a potential novel therapeutic target for malaria. The putative channel component of the NPP in the human malaria parasite P. falciparum is encoded by mutually exclusively expressed clag3.1/3.2 genes. Complicating the study of the essentiality of these genes to the NPP is the addition of three clag paralogs whose contribution to the P. falciparum channel is uncertain. Rodent malaria P. berghei contains only two clag genes, and thus studies of P. berghei clag genes could significantly aid in dissecting their overall contribution to NPP activity. Previous methods for determining NPP activity in a rodent model have utilised flux-based assays of radioisotope-labelled substrates or patch clamping. This study aimed to ratify a streamlined haemolysis assay capable of assessing the functionality of P. berghei NPPs. Several isotonic lysis solutions were tested for their ability to preferentially lyse infected RBCs (iRBCs), leaving uninfected RBCs (uRBCs) intact. The osmotic lysis assay was optimised and validated in the presence of NPP inhibitors to demonstrate the uptake of the lysis solution via the NPPs. Guanidinium chloride proved to be the most efficient reagent to use in an osmotic lysis assay to establish NPP functionality. Furthermore, following treatment with guanidinium chloride, ring-stage parasites could develop into trophozoites and schizonts, potentially enabling use of guanidinium chloride for parasite synchronisation. This haemolysis assay will be useful for further investigation of NPPs in P. berghei and could assist in validating its protein constituents.

Many virus lysis/transport buffers used in molecular diagnostics, including the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA, contain guanidine-based chaotropic salts, primarily guanidine hydrochloride (GuHCl) or guanidine isothiocyanate (GITC). Although the virucidal effects of GuHCl and GITC alone against some enveloped viruses have been established, standardized data on their optimum virucidal concentrations against SARS-CoV-2 and effects on viral RNA stability are scarce. Thus, we aimed to determine the optimum virucidal concentrations of GuHCl and GITC against SARS-CoV-2 compared to influenza A virus (IAV), another enveloped respiratory virus. We also evaluated the effectiveness of viral RNA stabilization at the determined optimum virucidal concentrations under high-temperature conditions (35°C) using virus-specific real-time reverse transcription polymerase chain reaction. Both viruses were potently inactivated by 1.0 M GITC and 2.5 M GuHCl, but the GuHCl concentration for efficient SARS-CoV-2 inactivation was slightly higher than that for IAV inactivation. GITC showed better viral RNA stability than GuHCl at the optimum virucidal concentrations. An increased concentration of GuHCl or GITC increased viral RNA degradation at 35°C. Our findings highlight the need to standardize GuHCl and GITC concentrations in virus lysis/transport buffers and the potential application of these guanidine-based salts alone as virus inactivation solutions in SARS-CoV-2 and IAV molecular diagnostics., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Hepatitis B virus (HBV) displays remarkable self-assembly capabilities that interest the scientific community and biotechnological industries as HBV is leading to an annual mortality of up to 1 million people worldwide (especially in Africa and Southeast Asia). When the ionic strength is increased, hepatitis B virus-like particles (VLPs) can assemble from dimers of the first 149 residues of the HBV capsid protein core assembly domain (Cp149). Using solution small-angle X-ray scattering, we investigated the disassembly of the VLPs by titrating guanidine hydrochloride (GuHCl). Measurements were performed with and without 1 M NaCl, added either before or after titrating GuHCl. Fitting the scattering curves to a linear combination of atomic models of Cp149 dimer (the subunit) and T = 3 and T = 4 icosahedral capsids revealed the mass fraction of the dimer in each structure in all the titration points. Based on the mass fractions, the variation in the dimer-dimer association standard free energy was calculated as a function of added GuHCl, showing a linear relation between the interaction strength and GuHCl concentration. Using the data, we estimated the energy barriers for assembly and disassembly and the critical nucleus size for all of the assembly reactions. Extrapolating the standard free energy to [GuHCl] = 0 showed an evident hysteresis in the assembly process, manifested by differences in the dimer-dimer association standard free energy obtained for the disassembly reactions compared with the equivalent assembly reactions. Similar hysteresis was observed in the energy barriers for assembly and disassembly and the critical nucleus size. The results suggest that above 1.5 M, GuHCl disassembled the capsids by attaching to the protein and adding steric repulsion, thereby weakening the hydrophobic attraction.

A new series of benzene-sulfonamide derivatives 3a-i was designed and synthesized via the reaction of N-(pyrimidin-2-yl)cyanamides 1a-i with sulfamethazine sodium salt 2 as dual Src/Abl inhibitors. Spectral data IR, 1 H-, 13 C- NMR and elemental analyses were used to confirm the structures of all the newly synthesized compounds 3a-i and 4a-i. Crucially, we screened all the synthesized compounds 3a-i against NCI 60 cancer cell lines. Among all, compound 3b was the most potent, with IC 50 of 0.018 μM for normoxia, and 0.001 μM for hypoxia, compared to staurosporine against HL-60 leukemia cell line. To verify the selectivity of this derivative, it was assessed against a panel of tyrosine kinase EGFR, VEGFR-2, B-raf, ERK, CK1, p38-MAPK, Src and Abl enzymes. Results revealed that compound 3b can effectively and selectively inhibit Src/Abl with IC 50 0.25 μM and Abl inhibitory activity with IC 50 0.08 μM, respectively, and was found to be more potent on these enzymes than other kinases that showed the following results: EGFR IC 50 0.31 μM, VEGFR-2 IC 50 0.68 μM, B-raf IC 50 0.33 μM, ERK IC 50 1.41 μM, CK1 IC 50 0.29 μM and p38-MAPK IC 50 0.38 μM. Moreover, cell cycle analysis and apoptosis performed to compound 3b against HL-60 suggesting its antiproliferative activity through Src/Abl inhibition. Finally, molecular docking studies and physicochemical properties prediction for compounds 3b, 3c, and 3 h were carried out to investigate their biological activities and clarify their bioavailability., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Mellpaladines A-C ( 1-3 ) and dopargimine ( 4 ) are dopamine-derived guanidine alkaloids isolated from a specimen of Palauan Didemnidae tunicate as possible modulators of neuronal receptors. In this study, we isolated the dopargimine derivative 1-carboxydopargimine ( 5 ), three additional mellpaladines D-F ( 6-8 ), and serotodopalgimine ( 9 ), along with a dimer of serotonin, 5,5'-dihydroxy-4,4'-bistryptamine ( 10 ). The structures of these compounds were determined based on spectrometric and spectroscopic analyses. Compound 4 and its congeners dopargine ( 11 ), nordopargimine ( 15 ), and 2-(6,7-dimethoxy-3,4-dihydroisoquinolin-1-yl)ethan-1-amine ( 16 ) were synthetically prepared for biological evaluations. The biological activities of all isolated compounds were evaluated in comparison with those of 1-4 using a mouse behavioral assay upon intracerebroventricular injection, revealing key functional groups in the dopargimines and mellpaladines for in vivo behavioral toxicity. Interestingly, these alkaloids also emerged during a screen of our marine natural product library aimed at identifying antiviral activities against dengue virus, SARS-CoV-2, and vesicular stomatitis Indiana virus (VSV) pseudotyped with Ebola virus glycoprotein (VSV-ZGP).

Glycogen, a complex branched glucose polymer, is responsible for sugar storage in blood glucose homeostasis. It comprises small β particles bound together into composite α particles. In diabetic livers, α particles are fragile, breaking apart into smaller particles in dimethyl sulfoxide, DMSO; they are however stable in glycogen from healthy animals. We postulate that the bond between β particles in α particles involves hydrogen bonding. Liver-glycogen fragility in normal and db/db mice (an animal model for diabetes) is compared using various hydrogen-bond breakers (DMSO, guanidine and urea) at different temperatures. The results showed different degrees of α-particle disruption. Disrupted glycogen showed changes in the mid-infra-red spectrum that are related to hydrogen bonds. While glycogen α-particles are only fragile under harsh, non-physiological conditions, these results nevertheless imply that the bonding between β particles in α particles is different in diabetic livers compared to healthy, and is probably associated with hydrogen bonding., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The chemical diversity of annelids, particularly those belonging to the class Sipuncula, remains largely unexplored. However, as part of a Marine Biodiscovery program in Ireland, the peanut worm Phascolosoma granulatum emerged as a promising source of unique metabolites. The purification of the MeOH/CH 2 Cl 2 extract of this species led to the isolation of six new linear guanidine amides, named phascolosomines A-F ( 1 - 6 ). NMR analysis allowed for the elucidation of their structures, all of which feature a terminal guanidine, central amide linkage, and a terminal isobutyl group. Notably, these guanidine amides were present in unusually high concentrations, comprising ∼3% of the dry mass of the organism. The primary concentration of the phascolosomines in the viscera is similar to that previously identified in linear amides from sipunculid worms and marine fireworms. The compounds from sipunculid worms have been hypothesized to be toxins, while those from fireworms are reported to be defensive irritants. However, screening of the newly isolated compounds for inhibitory bioactivity showed no significant inhibition in any of the assays conducted.

Metabolism and biological functions of the nitrogen-rich compound guanidine have long been neglected. The discovery of four classes of guanidine-sensing riboswitches and two pathways for guanidine degradation in bacteria hint at widespread sources of unconjugated guanidine in nature. So far, only three enzymes from a narrow range of bacteria and fungi have been shown to produce guanidine, with the ethylene-forming enzyme (EFE) as the most prominent example. Here, we show that a related class of Fe 2+ - and 2-oxoglutarate-dependent dioxygenases (2-ODD-C23) highly conserved among plants and algae catalyze the hydroxylation of homoarginine at the C6-position. Spontaneous decay of 6-hydroxyhomoarginine yields guanidine and 2-aminoadipate-6-semialdehyde. The latter can be reduced to pipecolate by pyrroline-5-carboxylate reductase but more likely is oxidized to aminoadipate by aldehyde dehydrogenase ALDH7B in vivo . Arabidopsis has three 2-ODD-C23 isoforms, among which Din11 is unusual because it also accepted arginine as substrate, which was not the case for the other 2-ODD-C23 isoforms from Arabidopsis or other plants. In contrast to EFE, none of the three Arabidopsis enzymes produced ethylene. Guanidine contents were typically between 10 and 20 nmol*(g fresh weight) -1 in Arabidopsis but increased to 100 or 300 nmol*(g fresh weight) -1 after homoarginine feeding or treatment with Din11-inducing methyljasmonate, respectively. In 2-ODD-C23 triple mutants, the guanidine content was strongly reduced, whereas it increased in overexpression plants. We discuss the implications of the finding of widespread guanidine-producing enzymes in photosynthetic eukaryotes as a so far underestimated branch of the bio-geochemical nitrogen cycle and propose possible functions of natural guanidine production., Competing Interests: DF, MS, GF, JH No competing interests declared, (© 2023, Funck et al.)

The progress of the pillar[5]arene chemistry allowed us to set out a new concept on application of the supramolecular assemblies to create antimicrobial films with variable surface morphologies and biological activities. Antibacterial films were derived from the substituted pillar[5]arenes containing nine pharmacophoric guanidine fragments and one thioalkyl substituent. Changing the only thioalkyl fragment in the macrocycle structure made it possible to control the biological activity of the resulting antibacterial coating. Pretreatment of the surface with aqueous solution of the amphiphilic pillar[5]arenes reduced the biofilm thickness by 56 ± 10% of Gram-positive Staphylococcus aureus in the case of the pillar[5]arene containing a thiooctyl fragment and by 52 ± 7% for the biofilm of Gram-negative Klebsiella pneumoniae in the case of pillar[5]arene containing a thiooctadecyl fragment. Meanwhile, the cytotoxicity of the synthesized macrocycles was examined at a concentration of 50 μg/mL, which was significantly lower than that of bis-guanidine-based antimicrobial preparations.

The Neuropeptide FF (NPFF) receptor system is known to modulate opioid actions and has been shown to mediate opioid-induced hyperalgesia and tolerance. The lack of subtype selective small molecule compounds has hampered further exploration of the pharmacology of this receptor system. The vast majority of available NPFF ligands possess a highly basic guanidine group, including our lead small molecule, MES304. Despite providing strong receptor binding, the guanidine group presents a potential pharmacokinetic liability for in vivo pharmacological tool development. Through structure-activity relationship exploration, we were able to modify our lead molecule MES304 to arrive at guanidine-free NPFF ligands. The novel piperidine analogues 8b and 16a are among the few non-guanidine based NPFF ligands known in literature. Both compounds displayed nanomolar NPFF-R binding affinity approaching that of the parent molecule. Moreover, while MES304 was non-subtype selective, these two analogues presented new starting points for subtype selective scaffolds, whereby 8b displayed a 15-fold preference for NPFF1-R, and 16a demonstrated an 8-fold preference for NPFF2-R. Both analogues showed no agonist activity on either receptor subtype in the in vitro functional activity assay, while 8b displayed antagonistic properties at NPFF1-R. The calculated physicochemical properties of 8b and 16a were also shown to be more favorable for in vivo tool design. These results indicate the possibility of developing potent, subtype selective NPFF ligands devoid of a guanidine functionality., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests. Christopher McCurdy reports financial support was provided by National Institutes of Health. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Human African trypanosomiasis (HAT), or sleeping sickness, is a neglected tropical disease with current treatments marred by severe side effects or delivery issues. To identify novel classes of compounds for the treatment of HAT, high throughput screening (HTS) had previously been conducted on bloodstream forms of T. b. brucei, a model organism closely related to the human pathogens T. b. gambiense and T. b. rhodesiense. This HTS had identified a number of structural classes with potent bioactivity against T. b. brucei (IC 50  ≤ 10 μM) with selectivity over mammalian cell-lines (selectivity index of ≥10). One of the confirmed hits was an aroyl guanidine derivative. Deemed to be chemically tractable with attractive physicochemical properties, here we explore this class further to develop the SAR landscape. We also report the influence of the elucidated SAR on parasite metabolism, to gain insight into possible modes of action of this class. Of note, two sub-classes of analogues were identified that generated opposing metabolic responses involving disrupted energy metabolism. This knowledge may guide the future design of more potent inhibitors, while retaining the desirable physicochemical properties and an excellent selectivity profile of the current compound class., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Invasive fungal infections impose a substantial global health burden. They cause more than 1.5 million deaths annually and are insufficiently met by the currently approved antifungal drugs. Antifungal peptides are a promising alternative to existing antifungal drugs; however, they can be challenging to synthesize, and are often susceptible to proteases in vivo . Synthetic polymers which mimic the properties of natural antifungal peptides can circumvent these limitations. In this study, we developed a library of 29 amphiphilic polyacrylamides with different charged units, namely, amines, guanidinium, imidazole, and carboxylic acid groups, representative of the natural amino acids lysine, arginine, histidine, and glutamic acid. Ternary polymers incorporating primary ammonium (lysine-like) or imidazole (histidine-like) groups demonstrated superior activity against Candida albicans and biocompatibility with mammalian cells compared to the polymers containing the other charged groups. Furthermore, a combination of primary ammonium, imidazole, and guanidinium (arginine-like) within the same polymer outperformed the antifungal drug amphotericin B in terms of therapeutic index and exhibited fast C. albicans -killing activity. The most promising polymer compositions showed synergistic effects in combination with caspofungin and fluconazole against C. albicans and additionally demonstrated activity against other clinically relevant fungi. Collectively, these results indicate the strong potential of these easily producible polymers to be used as antifungals.

The global challenge of antibiotic resistance necessitates the introduction of more effective antibiotics. Here we report a potentially general design strategy, exemplified with vancomycin, that improves and expands antibiotic performance. Vancomycin is one of the most important antibiotics in use today for the treatment of Gram-positive infections. However, it fails to eradicate difficult-to-treat biofilm populations. Vancomycin is also ineffective in killing Gram-negative bacteria due to its inability to breach the outer membrane. Inspired by our seminal studies on cell penetrating guanidinium-rich transporters (e.g., octaarginine), we recently introduced vancomycin conjugates that effectively eradicate Gram-positive biofilm bacteria, persister cells and vancomycin-resistant enterococci (with V-r8, vancomycin-octaarginine), and Gram-negative pathogens (with V-R, vancomycin-arginine). Having shown previously that the spatial array (linear versus dendrimeric) of multiple guanidinium groups affects cell permeation, we report here for the first time vancomycin conjugates with dendrimerically displayed guanidinium groups that exhibit superior efficacy and breadth, presenting the best activity of V-r8 and V-R in single broad-spectrum compounds active against ESKAPE pathogens. Mode-of-action studies reveal cell-surface activity and enhanced vancomycin-like killing. The vancomycin-polyguanidino dendrimer conjugates exhibit no acute mammalian cell toxicity or hemolytic activity. Our study introduces a new class of broad-spectrum vancomycin derivatives and a general strategy to improve or expand antibiotic performance through combined mode-of-action and function-oriented design studies.

The occurrence of increased antibiotic resistance has reduced the availability of drugs effective in the control of infectious diseases, especially those caused by various combinations of bacteria and/or fungi that are often associated with poorer patient outcomes. In the hunt for novel antibiotics of interest to treat polymicrobial diseases, molecules bearing guanidine moieties have recently come to the fore in designing and optimizing antimicrobial agents. Due to their remarkable antibacterial and antifungal activities, labdane diterpenes are also attracting increasing interest in antimicrobial drug discovery. In this study, six different guanidines prenylated with labdanic fragments were synthesized and evaluated for their antimicrobial properties. Assays were carried out against both non-resistant and antibiotic-resistant bacteria strains, while their possible antifungal activities have been tested on the yeast Candida albicans. Two of the synthesized compounds, namely labdan-8,13(R)-epoxy-15-oyl guanidine and labdan-8,13(S)-epoxy-15-oyl guanidine, were finally selected as the best candidates for further developments in drug discovery, due to their antimicrobial effects on both Gram-negative and Gram-positive bacterial strains, their fungicide action, and their moderate toxicity in vivo on zebrafish embryos. The study also provides insights into the structure-activity relationships of the guanidine-functionalized labdane-type diterpenoids., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Antimicrobial peptide mimics have been used to kill bacteria and construct antibacterial materials. Precise design and construction of chemical structure are essential for easy access to highly effective antimicrobial peptide mimics. Herein, cationic guanidinium-based polymers (PGXs) with varying hydrophobic structures were synthesized to explore the structure and antibacterial activity relationship of antimicrobial peptide mimics and to construct antibacterial implants. The effect of the hydrophobic chemical structure, including carbon chain length and configuration, on the antimicrobial activities against both Escherichia coli and Staphylococcus aureus was investigated. The antibacterial activities of PGXs improved with increasing alkyl chain length, and PGXs with a straight-chain hydrophobic structure exhibited better bactericidal activities than those with cyclic alkane and aromatic hydrocarbon. Furthermore, PGXs grafted with poly(dimethylsiloxane) (PDMS-PGXs) showed a similar bactericidal change tendency of PGXs in solution. Additionally, the PDMS-PGXs showed potent antibiofilm performance in vitro , which can inhibit bacterial infection in vivo as subcutaneous implants. This study may propose a basis for the precise design and construction of antibacterial materials and provide a promising way of designing biomedical devices and implants with bacterial infection-combating activities.

Chemodynamic therapy (CDT) has emerged as a promising approach to cancer treatment, which can break the intracellular redox state balance and result in severe oxidative damage to biomolecules and organelles with the advantages of being less dependent on external stimulation, having deep tissue-healing abilities, and being resistant to drug resistance. There is considerable interest in developing CDT drugs with high efficiency and low toxicity. In this study, a new guanidinium-based biological metal covalent organic framework (Bio-MCOF), GZHMU-1@Mo, is rationally designed and synthesized as a multifunctional nanocatalyst in tumor cells for enhanced CDT. The DFT calculation and experimental results showed that due to the ability of MoO 4 2- ion to promote electron transfer and increase the redox active site, Cu 3 clusters and MoO 4 2- ions in GZHMU-1@Mo can synergistically catalyze the production of reactive oxygen species (ROS) from oxygen and H 2 O 2 in tumor cells, as well as degrade intracellular reducing substances, GSH and NADH, so as to disrupt the redox balance in tumor cells. Moreover, GZHMU-1@Mo exhibits a potent killing effect on tumor cells under both normal oxygen and anaerobic conditions. Further in vitro and in vivo antiproliferation studies revealed that the GZHMU-1@Mo nanoagent displays a remarkable antiproliferation effect and effectively inhibits tumor growth. Taken together, our study provides an insightful reference benchmark for the rational design of Bio-MCOF-based nanoagents with efficient CDT.

Helicobacter pylori , the world's most common chronic infection-causing pathogen, is responsible for causing gastric ulcers, the fourth-leading cause of cancer-related death globally in 2020. In recent years, the effectiveness of the current treatment regimen (two antibiotics and one proton pump inhibitor) has often been plagued with problems such as resistance and the undesired elimination of commensal bacteria. Herein, we report the synthesis of block and random copolycarbonates, functionalized with cationic guanidinium and anionic acetate functional groups, aimed at selectively killing H. pylori in the acidic environment of the stomach, while remaining nontoxic to the commensal bacteria in the gut. The compositions of the polymers were fine-tuned so that the polymers were readily dispersed in water without any difficulty at both pH 3.0 and 7.4. The self-assembly behavior of the polymers at different pH values by dynamic light scattering showed that the random and block copolymers formed stable micelles in a simulated gastric environment (pH 3.0) while aggregated at pH 7.4. Both polymers demonstrated stronger antibacterial activity against H. pylori than the guanidinium-functionalized homopolymer without any acetate functional group at pH 3.0. The block copolymer was significantly more bactericidal at pH 3.0 across the concentrations tested, as compared to the random copolymer, while it did not show significant toxicity toward rat red blood cells (rRBCs) and HK-2 cells or bactericidal effect toward E. coli (a common gut bacterium) and nor caused aggregation of rRBCs at its effective concentration and at physiological pH of 7.4. Additionally, both the block and random copolymers were much more stable against hydrolysis at pH 3.0 than at pH 7.4. This study provides insight into the influence of both polymer architecture and dynamic assembly on the bioactivities of antimicrobial polymers, where the disassembly of coacervates into narrowly dispersed micelles at pH 3 make them potent antimicrobials aided by the protonated carboxylic acid block.

Antarctic krill suffers from severe water loss after heating, and its quality deteriorates, so it is in urgent need of a green and healthy improver. In this paper, the effects of L-arginine (L-Arg) soaking on the modification of the quality of heat-treated Antarctic krill and the structure of myofibrillar proteins (MPs) in Antarctic krill were investigated. The results showed that L-Arg had an ameliorating effect on heat-treated krill in a concentration-dependent relationship. The water-holding capacity of L-Arg-soaked krill was 1.41 times higher than that of sodium tripolyphosphate (STPP) at an equivalent concentration (80 mM). At 120 mM L-Arg soaked, L* and hardness of krill decreased to 58.31 and 334.81 g, while resilience and moisture content increased to 0.47 and 85.29 % after heating, respectively. The scanning electron microscopy (SEM) results revealed that the tissue state of the pH-corrected groups was better than the control, but not as well as that of the pH-uncorrected groups. pH and the guanidinium group in L-Arg both played roles in promoting the transition of MPs from disordered to ordered secondary structures. This transition reduced the exposure of hydrophobic and sulfhydryl groups in MPs, inhibited the protein aggregation and increased the solubility of MPs to 71.61 %, which ultimately improved the quality of heat-treated krill. It is worth noting that the pH effect had a primary influence on the observed effects, while the guanidinium group made a secondary contribution. These results could broaden the potential application of L-Arg as an improver of the quality of heat-treated krill., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

Antimicrobial peptides (AMPs) often display guanidinium functionalities, and hence robust synthetic procedures are needed to facilitate access to analogues with unnatural homologues of arginine (Arg = R). Initially, a resin-bound Arg/Pro-rich fluoren-9-yl-methyloxycarbonyl-protected fragment (Fmoc-RPRPPR) of the AMP oncocin (i.e., VDKPPYLPRPRPPRRIYNR-NH 2 ) was employed in a comparative on-resin assessment of commercial guanidinylation reagents head-to-head with the recently studied bis-Boc-protected triazole-based reagent, 1H-triazole-1-[N,N'-bis(tert-butoxycarbonyl)]-carboxamidine, which was synthesized by a chromatography-free procedure. This reagent was found to enable quantitative conversion in solid-phase peptide synthesis (SPPS) of peptides displaying homoarginine (Har) residues and/or an N-terminal guanidinium group. SPPS was used to obtain analogues of the 18-mer oncocin with single as well as multiple Arg → Har modifications. In addition, the effect of replacement of proline (Pro) residues in oncocin was explored by incorporating single or multiple trans-4-hydroxy-l-proline (Hyp) or 4,4-difluoro-l-proline (Dfp) residues, which both affected hydrophobicity. The resulting peptide library was tested against both Gram-negative and Gram-positive bacteria. Analysis of the minimal inhibitory concentrations (MICs) showed that analogues, displaying modifications at positions 4, 5 and 12 (originally Pro residues), had retained or slightly improved antimicrobial activity. Next, an oncocin analogue with two stabilizing l-Arg → d-Arg replacements in the C-terminal part was further modified by triple-replacement of Pro by either Dfp or Hyp in positions 4, 5, and 12. The resulting analogue displaying three Pro → Dfp modifications proved to possess the best activity profile: MICs of 1-2 µg/mL against E. coli and Klebsiella pneumoniae, less than 1% hemolysis at 800 µg/mL, and an IC 50 above 1280 µg/mL in HepG2 cells. Thus, incorporation of bis-fluorinated Pro residues appears to constitute a novel tool in structure-activity studies aimed at optimization of Pro-rich AMPs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that have influenced the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Src homology-2 domain containing protein tyrosine phosphatase-2 (SHP2) is a highly attractive therapeutic target for treating Kirsten rat sarcoma viral oncogene (KRAS) mutant cancers. In this work, a series of guanidine-based SHP2 allosteric inhibitors were discovered via virtual screening and rational structural optimization. Notably, lead compound 23 with potent SHP2 inhibitory activity (IC 50 = 17.7 nM) effectively inhibited the proliferation, migration, and invasion of MIA PaCa-2 pancreatic cancer cells. Furthermore, compound 23 featured great in vivo pharmacokinetic properties (AUC po = 4320 nM·h; F = 66.3%) and exhibited significant antitumor efficacy in the MIA PaCa-2 xenograft mouse model. This demonstrates that compound 23 is a potential lead compound for the development of SHP2 allosteric inhibitors to treat KRAS mutant cancers. Moreover, these guanidine-based scaffolds may provide an opportunity to mitigate the potential safety risks of the alkyl amine motif predominately incorporated in current SHP2 allosteric inhibitors.

Medulloblastoma (MB) is the most common pediatric brain tumor. The therapy frequently causes serious side effects, and new selective therapies are needed. MB expresses hyper sialylation, a possible target for selective therapy. The cytotoxic efficacy of a poly guanidine conjugate (GuaDex) incubated with medulloblastoma cell cultures (DAOY and MB-LU-181) was investigated. The cells were incubated with 0.05-8 µM GuaDex from 15 min to 72 h. A fluorometric cytotoxicity assay (FMCA) measured the cytotoxicity. Labeled GuaDex was used to study tumor cell interaction. FITC-label Sambucus nigra confirmed high expression of sialic acid (Sia). Immunofluorescence microscopy was used to visualize the cell F-actin and microtubules. The cell interactions were studied by confocal and fluorescence microscopy. Annexin-V assay was used to detect apoptosis. Cell cycle analysis was done by DNA content determination. A wound-healing migration assay determined the effects on the migratory ability of DAOY cells after GuaDex treatment. IC 50 for GuaDex was 223.4 -281.1 nM. FMCA showed potent growth inhibition on DAOY and MB-LU-181 cells at 5 uM GuaDex after 4 h of incubation. GuaDex treatment induced G2/M phase cell cycle arrest. S. nigra FITC-label lectin confirmed high expression of Sia on DAOY medulloblastoma cells. The GuaDex treatment polymerized the cytoskeleton (actin filaments and microtubules) and bound to DNA, inducing condensation. The Annexin V assay results were negative. Cell migration was inhibited at 0.5 µM GuaDex concentration after 24 h of incubation. GuaDex showed potent cytotoxicity and invasion-inhibitory effects on medulloblastoma cells at low micromolar concentrations. GuaDex efficacy was significant and warrants further studies., (© 2023. The Author(s).)

Guanidines are fascinating small nitrogen-rich organic compounds, which have been frequently associated with a wide range of biological activities. This is mainly due to their interesting chemical features. For these reasons, for the past decades, researchers have been synthesizing and evaluating guanidine derivatives. In fact, there are currently on the market several guanidine-bearing drugs. Given the broad panoply of pharmacological activities displayed by guanidine compounds, in this review, we chose to focus on antitumor, antibacterial, antiviral, antifungal, and antiprotozoal activities presented by several natural and synthetic guanidine derivatives, which are undergoing preclinical and clinical studies from January 2010 to January 2023. Moreover, we also present guanidine-containing drugs currently in the market for the treatment of cancer and several infectious diseases. In the preclinical and clinical setting, most of the synthesized and natural guanidine derivatives are being evaluated as antitumor and antibacterial agents. Even though DNA is the most known target of this type of compounds, their cytotoxicity also involves several other different mechanisms, such as interference with bacterial cell membranes, reactive oxygen species (ROS) formation, mitochondrial-mediated apoptosis, mediated-Rac1 inhibition, among others. As for the compounds already used as pharmacological drugs, their main application is in the treatment of different types of cancer, such as breast, lung, prostate, and leukemia. Guanidine-containing drugs are also being used for the treatment of bacterial, antiprotozoal, antiviral infections and, recently, have been proposed for the treatment of COVID-19. To conclude, the guanidine group is a privileged scaffold in drug design. Its remarkable cytotoxic activities, especially in the field of oncology, still make it suitable for a deeper investigation to afford more efficient and target-specific drugs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Triple-negative breast cancer (TNBC) is one of the subtypes of breast cancer (BC) that is associated with poor survival rates and failure to respond to hormonal and targeted therapies., Objective: The aim of this study was to identify a specific gene at the expression level for TNBC and targeting of this type of breast cancer based on it. Using TCGA database, genes that are particularly high expression in TNBC subtypes compared to other BC subtypes (in terms of receptor status) and normal samples were identified and their sensitivity and specificity were evaluated. Using PharmacoGX and Drug Bank data, drug sensitivity and drug-appropriate genes were identified, respectively. The effects of the identified drug on triple-negative cell lines (MDA-MB-468) were evaluated in comparison with the cell line of other subtypes (MCF7) by apoptosis and MTS tests., Results: Data analyzes showed that the expression level of KCNG1 gene in the TNBC subgroup was significantly higher compared to other BC subtypes from the KCN gene family and ROC results showed that this gene had highest sensitivity and specificity in TNBC subtype. The results of drug resistance and sensitivity showed that an increase in the expression level of KCNG1 was associated with sensitivity to Cisplatin and Oxaliplatin. Moreover, Drug Bank results showed that Guanidine hydrochloride (GuHCl) was a suitable inhibitor for KCNG1. In vitro results showed that the expression level of KCNG1 was higher in MDA-MB-468 compared to MCF7. In addition, the rate of apoptosis in response to GuHCl treatment in MDA-MB-468 cell line as TNBC cell model was higher than MCF7 in the same concentration., Conclusion: This study revealed that GuHCl could be a suitable treatment for TNBC subtype by targeting of KCNG1., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest., (Copyright © 2023. Published by Elsevier Masson SAS.)

Riboswitches function as important translational regulators in bacteria. Comprehensive mutational analysis of transcriptional riboswitches has been used to probe the energetic intricacies of interplay between the aptamer and expression platform, but translational riboswitches have been inaccessible to massively parallel techniques. The guanidine-II (gdm-II) riboswitch is an exclusively translational class. We have integrated RelE cleavage with next-generation sequencing to quantify ligand-dependent changes in translation initiation for all single and double mutations of the Pseudomonas aeruginosa gdm-II riboswitch, a total of more than 23,000 variants. This extensive mutational analysis is consistent with the prominent features of the bioinformatic consensus. These data indicate, unexpectedly, that direct sequestration of the Shine-Dalgarno sequence is dispensable for riboswitch function. Additionally, this comprehensive data set reveals important positions not identified in previous computational and crystallographic studies. Mutations in the variable linker region stabilize alternate conformations. The double mutant data reveal the functional importance of the previously modeled P0b helix formed by the 5' and 3' tails that serves as the basis for translational control. Additional mutations to GU wobble base pairs in both P1 and P2 reveal how the apparent cooperativity of the system involves an intricate network of communication between the two binding sites. This comprehensive examination of a translational riboswitch's expression platform illuminates how the riboswitch is precisely tuned and tunable with regard to ligand sensitivity, the amplitude of expression between ON and OFF states, and the cooperativity of ligand binding., (© 2023 Focht et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.)

The bacterial infection of surgical wounds results in prolonged hospitalization and even death of patients, calling for antibacterial function in modern suture products. To tackle this challenge, cationic guanidine-containing copolymer was synthesized, exhibiting antibacterial potency over 5 log reduction against both Gram-positive S. aureus and Gram-negative E. coli. Furthermore, we developed a double-network silk suture by integrating a guanidine-containing copolymer network into the silk fibroin network. This suture exhibited biocidal activity against S. aureus and E. coli, and superior strength compared to the commercial product in both dry and wet conditions. These results may bring general benefits to public health and medical equipment sustainability., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Nanozymes are effective novel antibacterial agents. However, they still have some shortcomings such as low catalytic efficiency, poor specificity, and non-negligible toxic side effects. Here, we synthesized iridium oxide nanozymes (IrO x NPs) by a one-pot hydrothermal method and used guanidinium peptide-betaine (SNLP/BS-12) to modify the surface of IrO x NPs (SBI NPs) to obtain a high-efficiency and low-toxicity antibacterial agent. In vitro experiments showed that SBI NPs with SNLP/BS12 could enhance IrO x NPs to target bacteria, mediate bacterial surface catalysis and reduce the cytotoxicity of IrO x NPs to mammalian cells. Importantly, SBI NPs were able to effectively alleviate MRSA acute lung infection and effectively promote diabetic wound healing. Accordingly, iridium oxide nanozymes functionalized with guanidinium peptides are expected to be an effective antibiotic candidate in the postantibiotic era.

Although new nematicides have appeared, the demand for new products less toxic and more efficient for the control of plant-parasitic nematodes are still high. Consequently, studies on natural secondary metabolites from plants, to develop new nematicides, have increased. In this work, nineteen extracts from eleven Brazilian plant species were screened for activity against Meloidogyne incognita. Among them, the extracts of Piterogyne nitens showed a potent nematostatic activity. The alkaloid fraction obtained from the ethanol extract of leaves of P. nitens was more active than the coming extract. Due to the promising activity from the alkaloid fraction, three isoprenylated guanidine alkaloids isolated from this fraction, galegine (1), pterogynidine (2), and pterogynine (3) were tested, showing similar activity to the alkaloid fraction, which was comparable to that of the positive control Temik at 250 μg/mL. At lower concentrations (125-50 μg/mL), compound 2 showed to be the most active one. As several nematicides act through inhibition of acetylcholinesterase (AChE), the guanidine alkaloids were also employed in two in vitro AChE assays. In both cases, compound 2 was more active than compounds 1 and 3. Its activity was considered moderated compared to the control (physostigmine). Compound 2 was selected for an in silico study with the electric eel (Electrophorus electricus) AChE, showing to bind mostly to the same site of physostigmine in the AChEs, pointing out that this could be the mechanism of action for this compound. These results suggested that the guanidine alkaloids 1,2 and 3 from P. nitens are promising for the development of new products to control M. incognita, especially guanidine 2, and encourage new investigations to confirm the mechanism of action, as well as to determine the structure-activity relationship of the guanidine alkaloids., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Various environmental compounds are inducers of lung injury. Mitochondria are crucial organelles that can be affected by many lung diseases. NecroX is an indole-derived antioxidant that specifically targets mitochondria. We aimed to evaluate the therapeutic potential and related molecular mechanisms of NecroX in preclinical models of fatal lung injury. We investigated the therapeutic effects of NecroX on two different experimental models of lung injury induced by polyhexamethylene guanidine (PHMG) and bleomycin, respectively. We also performed transcriptome analysis of lung tissues from PHMG-exposed mice and compared the expression profiles with those from dozens of bleomycin-induced fibrosis public data sets. Respiratory exposure to PHMG and bleomycin led to fatal lung injury manifesting extensive inflammation followed by fibrosis. These specifically affected mitochondria regarding biogenesis, mitochondrial DNA integrity, and the generation of mitochondrial reactive oxygen species in various cell types. NecroX significantly improved the pathobiologic features of the PHMG- and bleomycin-induced lung injuries through regulation of mitochondrial oxidative stress. Endoplasmic reticulum stress was also implicated in PHMG-associated lung injuries of mice and humans, and NecroX alleviated PHMG-induced lung injury and the subsequent fibrosis, in part, via regulation of endoplasmic reticulum stress in mice. Gene expression profiles of PHMG-exposed mice were highly consistent with public data sets of bleomycin-induced lung injury models. Pathways related to mitochondrial activities, including oxidative stress, oxidative phosphorylation, and mitochondrial translation, were upregulated, and these patterns were significantly reversed by NecroX. These findings demonstrate that NecroX possesses therapeutic potential for fatal lung injury in humans.

The melanocortin receptors are involved in numerous physiological pathways, including appetite, skin and hair pigmentation, and steroidogenesis. In particular, the melanocortin-3 receptor (MC3R) is involved in fat storage, food intake, and energy homeostasis. Small-molecule ligands developed for the MC3R may serve as therapeutic lead compounds for treating disease states of energy disequilibrium. Herein, three previously reported pyrrolidine bis-cyclic guanidine compounds with five sites for molecular diversity (R1-R5) were subjected to parallel structure-activity relationship studies to identify the common pharmacophore of this scaffold series required for full agonism at the MC3R. The R2, R3, and R5 positions were required for full MC3R efficacy, while truncation of either the R1 or R4 positions in all three compounds resulted in full MC3R agonists. Two additional fragments, featuring molecular weights below 300 Da, were also identified that possessed full agonist efficacy and micromolar potencies at the mMC5R. These SAR experiments may be useful in generating new small-molecule ligands and chemical probes for the melanocortin receptors to help elucidate their roles in vivo and as therapeutic lead compounds.

Cystatin superfamily members, by virtue of their thiol protease regulatory properties, show involvement in myriad physiological processes important for survival and well-being. The current study involves urea-induced denaturation of a novel variant of the cystatin superfamily, rai seed cystatin (RSC), employing a variety of biophysical assays in order to characterize different folding intermediates generated on unfolding. Urea as a denaturant presented the passage of RSC through a series of events resulting in the loss of RSC functional capability, accompanied by changes in the archetype at secondary and tertiary structural levels, as evident from protease inhibitory, UV absorption, and intrinsic fluorescence assays, respectively. ANS fluorescence also revealed routing of RSC through discrete multiple sub-states thus presenting the generation of intermediate states somewhat close to the pre-molten globule and/or molten globule forms of RSC. Furthermore, far-UV circular dichroism analysis revealed a concentration-dependent gradual loss in typical -helical RSC peaks, indicating a nearly 50 % loss in secondary structural elements around 5 M urea treatment. The study also reports the possible role of glycerol in the refolding and/or reactivation of the urea unfolded RSC form. Glycerol presented itself as a potent structural stabilizer as it assisted in the refolding and reactivation of the unfolded RSC in a dosage-dependent manner, concomitantly paving the way for unravelling the mechanistic approach involved in the phenomenon, which can facilitate future studies., Competing Interests: Declaration of Competing Interest The authors declare that there is no conflict of interest in this work., (Copyright © 2023. Published by Elsevier B.V.)

Isopropoxy benzene guanidine (IBG) is a guanidine derivative with antibacterial activity against multidrug-resistant bacteria. A few studies have revealed the metabolism of IBG in animals. The aim of the current study was to identify potential metabolic pathways and metabolites of IBG. The detection and characterization of metabolites were performed with high-performance liquid chromatography tandem mass spectrometry (UHPLC-Q-TOF-MS/MS). Seven metabolites were identified from the microsomal incubated samples by using the UHPLC-Q-TOF-MS/MS system. The metabolic pathways of IBG in the rat liver microsomes involved O-dealkylation, oxygenation, cyclization, and hydrolysis. Hydroxylation was the main metabolic pathway of IBG in the liver microsomes. This research investigated the in vitro metabolism of IBG to provide a basis for the further pharmacology and toxicology of this compound.

The increasing emergence of antibiotic resistance is an urgent threat to global health care; thus, there is a need for new therapeutics. Guanidine is the preferred functional group for antimicrobial design and development. Herein, the potential antibacterial activity of the guanidine derivative isopropoxy benzene guanidine (IBG) against multidrug-resistant (MDR) bacteria was discovered. The synergistic antibacterial activity of IBG and colistin was determined by checkerboard assay, time-killing curve, and mouse experiments. The antibacterial mechanism of IBG was verified in fluorescent probe experiments, intracellular oxidative phosphorylation assays, and transcriptome analysis. The results showed that IBG displays efficient antibacterial activity against Gram-positive pathogens and Gram-negative pathogens with permeabilized outer membranes. Further mechanistic studies showed that IBG triggers cytoplasmic membrane damage by binding to phosphatidylglycerol and cardiolipin, leading to the dissipation of proton motive force and accumulation of intracellular ATP. IBG combined with low levels of colistin enhances bacterial outer membrane permeability and increases the accumulation of reactive oxygen species, as further evidenced by transcriptome analysis. Furthermore, the efficacy of IBG with colistin against MDR Escherichia coli in three infection models was demonstrated. Together, these results suggest that IBG is a promising adjuvant of colistin, providing an alternative approach to address the prevalent infections caused by MDR Gram-negative pathogens. IMPORTANCE As antibiotic discovery stagnates, the world is facing a growing menace from the emergence of bacteria that are resistant to almost all available antibiotics. The key to winning this race is to explore distinctive mechanisms of antibiotics. Thus, novel efficient antibacterial agents and alternative strategies are urgently required to fill the void in antibiotic development. Compared with the large amount of money and time required to develop new agents, the antibiotic adjuvant strategy is a promising approach to inhibit bacterial resistance and increase killing of bacteria. In this study, we found that the guanidine derivatives IBG not only displayed efficient antibacterial activities against Gram-positive bacteria but also restored colistin susceptibility of Gram-negative pathogens as an antibiotic adjuvant. More in-depth study showed that IBG is a potential lead to overcome antibiotic resistance, providing new insight into future antibiotic discovery and development.

To enhance biosafety and reliability in SARS-CoV-2 molecular diagnosis, virus lysis/transport buffers should inactivate the virus and preserve viral RNA under various conditions. Herein, we evaluated the SARS-CoV-2-inactivating activity of guanidine hydrochloride (GuHCl)- and surfactant (hexadecyltrimethylammonium chloride (Hexa-DTMC))-based buffer, Prep Buffer A, (Precision System Science Co., Ltd., Matsudo, Japan) and its efficacy in maintaining the stability of viral RNA at different temperatures using the traditional real-time one-step RT-PCR and geneLEAD VIII sample-to-result platform. Although Prep Buffer A successfully inactivated SARS-CoV-2 in solutions with high and low organic substance loading, there was considerable viral genome degradation at 35 °C compared with that at 4 °C. The individual roles of GuHCl and Hexa-DTMC in virus inactivation and virus genome stability at 35 °C were clarified. Hexa-DTMC alone (0.384%), but not 1.5 M GuHCl alone, exhibited considerable virucidal activity, suggesting that it was essential for potently inactivating SARS-CoV-2 using Prep Buffer A. GuHCl and Hexa-DTMC individually reduced the viral copy numbers to the same degree as Prep Buffer A. Although both components inhibited RNase activity, Hexa-DTMC, but not GuHCl, directly destroyed naked viral RNA. Our findings suggest that samples collected in Prep Buffer A should be stored at 4 °C when RT-PCR will not be performed for several days.

Natural guanidines, molecules that contain the guanidine moiety, are structurally unique and often exhibit potent biological activities. A phytochemical investigation of the leaves of Alchornea rugosa (Lour.) Müll.Arg. by MS/MS-based molecular networking revealed eight undescribed guanidine-flavanol conjugates named rugonines A-H. The chemical structures of the isolated compounds were comprehensively elucidated by NMR spectroscopy, HRESIMS, and circular dichroism (CD) analysis. All isolated compounds were tested for autophagosome formation in HEK293 cells stably expressing GFP-LC3. The results revealed that compounds rugonines D-G showed potential autophagy inhibitory activity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Supramolecules have been drawing increasing attention recently in addressing healthcare challenges caused by infectious pathogens. We herein report a novel class of guanidinium-perfunctionalized polyhedral oligomeric silsesquioxane (Gua-POSS) supramolecules with highly potent antimicrobial activities. The modular structure of Gua-POSS T m -C n consists of an inorganic T10 or T8 core ( m = 10 or 8), flexible linear linkers of varying lengths ( n = 1 or 3), and peripherally aligned cationic guanidinium groups as the membrane-binding units. Such Gua-POSS supramolecules with spherically arrayed guanidinium cations display high antimicrobial potency against Gram-positive ( Staphylococcus aureus ) and Gram-negative ( Escherichia coli ) bacteria, as well as fungus ( Candida albicans ), with the best showing excellently low minimal inhibitory concentrations (MICs) of 1.7-6.8 μM in media, yet with negligible hemolytic activity and low in vitro cytotoxicity to mammalian cells. More significantly, they can inhibit biofilm formation at around their MICs and near-completely break down preestablished difficult-to-break biofilms at 250 μg mL -1 (∼50 μM). Their strong antiviral efficacy was also experimentally demonstrated against the enveloped murine hepatitis coronavirus as a surrogate of the SARS-CoV species. Overall, this study provides a new design approach to novel classes of sphere-shaped organic-inorganic hybrid supramolecular materials, especially for potent antimicrobial, anti-biofilm, and antiviral applications.