The Activity of Red Nigerian Propolis and Some of Its Components against Trypanosoma brucei and Trypanosoma congolense.

Propolis is a resin that is gathered by bees from exudates produced by various plants. Its exact chemical composition depends on the plants available near the hive. Bees use propolis to coat the surfaces of the hive, where it acts as an anti-infective. Regardless of the chemical composition of propolis, it is always anti-protozoal, probably because protozoan parasites, particularly Lotmarium passim, are widespread in bee populations. The protozoa Trypanosoma brucei and T. congolense cause disease in humans and/or animals. The existing drugs for treating these diseases are old and resistance is an increasingly severe problem. The many types of propolis present a rich source of anti-trypanosomal compounds-from a material gathered by bees in an environmentally friendly way. In the current work, red Nigerian propolis from Rivers State, Nigeria was tested against T. brucei and T. congolense and found to be highly active (EC50 1.66 and 4.00 µg/mL, respectively). Four isoflavonoids, vestitol, neovestitol, 7-methylvestitol and medicarpin, were isolated from the propolis. The isolated compounds were also tested against T. brucei and T. congolense, and vestitol displayed the highest activity at 3.86 and 4.36 µg/mL, respectively. Activities against drug-resistant forms of T. brucei and T. congolense were similar to those against wild type.

Chitin and Chitosan Binding to the α-Chitin Crystal: A Molecular Dynamics Study.

Understanding the binding of chitosan oligomers to the surface of a chitin nanocrystal is important for improving the enzymatic deacetylation of chitin and for the design of chitin/chitosan composite films. Here, we study the binding of several chito-oligomers to the (100) surface of an α-chitin crystal using molecular dynamics (MD), steered MD, and umbrella sampling. The convergence of the free energy was carefully considered and yielded a binding energies of -12.5 and -2 kcal mol-1 for 6-monomer-long chitin and uncharged chitosan oligomers, respectively. We also found that the results for the umbrella sampling were consistent with the force profile from the steered MD and with classical MD simulations of the adsorption process. Our results give insight into the molecular-scale interactions, which can be helpful for the design of new chitin composite films. Furthermore, the free energy curves we present can be used to validate coarse-grained models for chitin and chitosan, which are necessary to study the self-assembly of chitin crystals due to the long time scale of the process.

Characterisation of niosome nanoparticles prepared by microfluidic mixing for drug delivery.

Lipid nanoparticles have gained much attention due to their potential as drug delivery systems. They are safe, effective, and be targeted to particular tissues to deliver their payload. Niosomes are one type of lipid nanoparticles that comprise non-ionic surfactants which have proven to be effective due to their stability and biocompatibility. Different manufacturing processes have been reported for niosome preparation, but many of them are not scalable or reproducible for pharmaceutical use. In this study, microfluidic mixing was used to prepare niosomes with different lipid compositions by changing the type of non-ionic surfactant. Niosomes were evaluated for their physicochemical characteristics, morphology, encapsulation efficacy, release profiles of atenolol as a model hydrophilic compound, and cytotoxic activities. Microfluidic mixing allows for particle self-assembly and drug loading in a single step, without the need for post-preparation size reduction. Depending on the lipid composition, the empty particles were <90 nm in size with a uniform distribution. A slight but not significant increase in these values was observed when loading atenolol in most of the prepared formulations. All formulations were spherical and achieved variable levels of atenolol encapsulation. Atenolol release was slow and followed the Korsmeyer-Peppas model regardless of the surfactant type or the percentage of cholesterol used.

The Role of Antibodies in the Treatment of SARS-CoV-2 Virus Infection, and Evaluating Their Contribution to Antibody-Dependent Enhancement of Infection.

Antibodies play a crucial role in the immune response, in fighting off pathogens as well as helping create strong immunological memory. Antibody-dependent enhancement (ADE) occurs when non-neutralising antibodies recognise and bind to a pathogen, but are unable to prevent infection, and is widely known and is reported as occurring in infection caused by several viruses. This narrative review explores the ADE phenomenon, its occurrence in viral infections and evaluates its role in infection by SARS-CoV-2 virus, which causes coronavirus disease 2019 (COVID-19). As of yet, there is no clear evidence of ADE in SARS-CoV-2, though this area is still subject to further study.

Functionalisation of Inorganic Material Surfaces with Staphylococcus Protein A: A Molecular Dynamics Study.

Staphylococcus protein A (SpA) is found in the cell wall of Staphylococcus aureus bacteria. Its ability to bind to the constant Fc regions of antibodies means it is useful for antibody extraction, and further integration with inorganic materials can lead to the development of diagnostics and therapeutics. We have investigated the adsorption of SpA on inorganic surface models such as experimentally relevant negatively charged silica, as well as positively charged and neutral surfaces, by use of fully atomistic molecular dynamics simulations. We have found that SpA, which is itself negatively charged at pH7, is able to adsorb on all our surface models. However, adsorption on charged surfaces is more specific in terms of protein orientation compared to a neutral Au (111) surface, while the protein structure is generally well maintained in all cases. The results indicate that SpA adsorption is optimal on the siloxide-rich silica surface, which is negative at pH7 since this keeps the Fc binding regions free to interact with other species in solution. Due to the dominant role of electrostatics, the results are transferable to other inorganic materials and pave the way for new diagnostic and therapeutic designs where SpA might be used to conjugate antibodies to nanoparticles.

The Antiprotozoal Activity of Papua New Guinea Propolis and Its Triterpenes.

Profiling a propolis sample from Papua New Guinea (PNG) using high-resolution mass spectrometry indicated that it contained several triterpenoids. Further fractionation by column chromatography and medium-pressure liquid chromatography (MPLC) followed by nuclear magnetic resonance spectroscopy (NMR) identified 12 triterpenoids. Five of these were obtained pure and the others as mixtures of two or three compounds. The compounds identified were: mangiferonic acid, ambonic acid, isomangiferolic acid, ambolic acid, 27-hydroxyisomangiferolic acid, cycloartenol, cycloeucalenol, 24-methylenecycloartenol, 20-hydroxybetulin, betulin, betulinic acid and madecassic acid. The fractions from the propolis and the purified compounds were tested in vitro against Crithidia fasciculata, Trypanosoma congolense, drug-resistant Trypanosoma congolense, Trypanosoma b. brucei and multidrug-resistant Trypanosoma b. brucei (B48). They were also assayed for their toxicity against U947 cells. The compounds and fractions displayed moderate to high activity against parasitic protozoa but only low cytotoxicity against the mammalian cells. The most active isolated compound, 20-hydroxybetulin, was found to be trypanostatic when different concentrations were tested against T. b. brucei growth.

Limited Impact of the Protein Corona on the Cellular Uptake of PEGylated Zein Micelles by Melanoma Cancer Cells.

The formation of a protein layer "corona" on the nanoparticle surface upon entry into a biological environment was shown to strongly influence the interactions with cells, especially affecting the uptake of nanomedicines. In this work, we present the impact of the protein corona on the uptake of PEGylated zein micelles by cancer cells, macrophages, and dendritic cells. Zein was successfully conjugated with poly(ethylene glycol) (PEG) of varying chain lengths (5K and 10K) and assembled into micelles. Our results demonstrate that PEGylation conferred stealth effects to the zein micelles. The presence of human plasma did not impact the uptake levels of the micelles by melanoma cancer cells, regardless of the PEG chain length used. In contrast, it decreased the uptake by macrophages and dendritic cells. These results therefore make PEGylated zein micelles promising as potential drug delivery systems for cancer therapy.

Mucosal and systemic immune responses following mucosal immunisation of tetanus toxoid entrapped in lipid nanoparticles prepared by microwave reactor.

In this study, the use of a microwave reactor, which allowed high input of energy into a pressurised system in a short period of time, was investigated for preparation of lipid nanoparticles (LNPs). The aim was to optimise the formulation process by reducing manufacturing time. Two types of LNPs were prepared; non-ionic surfactant vesicles (NISV) and bilosomes (modified NISV incorporating bile salts), with a model antigen (tetanus toxoid, TT) and the immune response induced after mucosal (nasal and oral, respectively) administration was assessed. The TT loaded LNPs were characterised in terms of particle size, size distribution, morphology, and entrapment efficiency. Immunisation was evaluated by lethal challenge with tetanus toxin in an animal model. The efficiency of vaccination was evaluated by measuring the anti-TT IgG antibody levels in the vaccinated animals. Bilosomes formed by this method showed an immunogen entrapment efficiency of ∼30% which was significantly (p < 0.05) higher than entrapment efficiency in the NISV. The percentage of animals that survived when challenged with tetanus toxin correlated with the level of IgG determined in the serum of mice immunised with LNPs by the mucosal route. Moreover, there were significant (p < 0.05) differences between orally and nasally immunised groups. Animal groups immunised bilosomes via the oral route showed the highest level of IgG (1.2 ± 0.13) compared to the positive control, LN + Xn, and no immunised group. Similarly, groups immunised via the nasal route showed significantly (p < 0.0001) higher titres compared with the control group. Mucosal TT was capable of inducing systemic specific IgG anti-TT responses that were higher than the parenteral vaccine.

Potentiality of Melittin-Loaded Niosomal Vesicles Against Vancomycin-Intermediate Staphylococcus aureus and Staphylococcal Skin Infection.

BACKGROUND: Staphylococcus aureus is an important human pathogen, especially causing skin and soft tissue infections (SSTIs). Over the decades, the infections caused by antibiotic-resistant strains have often become life-threatening. Consequently, exploration and development of competent approaches to combat these serious circumstances are urgently required. METHODS: The antibacterial activity of melittin (Mel) on S. aureus, methicillin-resistant S. aureus (MRSA) and clinical isolates of vancomycin-intermediate S. aureus (VISA) was investigated by minimum inhibitory concentration (MIC) and time-killing assays. The localization of Mel on the bacterial cell was visualized by confocal laser scanning microscopy and its effect on the membrane was indicated based on propidium iodide uptake. The non-ionic surfactant vesicle (NISV) or niosome nanocarrier was established for Mel loading (Mel-loaded NISV) by the thin-film hydration method. Physicochemical and in vitro biological properties of Mel-loaded NISVs were characterized. The cellular uptake of Mel-loaded NISVs was evaluated by holotomography analysis. In addition, an ex vivo study was conducted on a porcine ear skin model to assess the permeation ability of Mel-loaded NISVs and their potential to inhibit bacterial skin infection. RESULTS: The effective inhibitory activity of Mel on skin pathogens was demonstrated. Among the tested strains, VISA was most susceptible to Mel. Regarding to its function, Mel targeted the bacterial cell envelope and disrupted cell membrane integrity. Mel-loaded NISVs were successfully fabricated with a nano-size of 120-200 nm and entrapment efficiency of greater than 90%. Moreover, Mel-loaded NISVs were taken up and accumulated in the intracellular space. Meanwhile, Mel was released and distributed throughout the cytosol and nucleus. Mel-loaded NISVs efficiently inhibited the growth of bacteria, particularly MRSA and VISA. Importantly, they not only penetrated epidermal and dermal skin layers, but also reduced the bacterial growth in infected skin. CONCLUSION: Mel-loaded NISVs have a great potential to exhibit antibacterial activity. Therapeutic application of Mel-loaded NISVs could be further developed as an alternative platform for the treatment of skin infection via dermal and transdermal delivery.

Multiple myeloma: therapeutic delivery of antibodies and aptamers.

Multiple myeloma is the second most common hematological malignancy in adults, accounting for 2% of all cancer-related deaths in the UK. Current chemotherapy-based regimes are insufficient, as most patients relapse and develop therapy resistance. This review focuses on current novel antibody- and aptamer-based therapies aiming to overcome current therapy limitations, as well as their respective limitations and areas of improvement. The use of computer modeling methods, as a tool to study and improve ligand-receptor alignments for the use of novel therapy development will also be discussed, as it has become a rapid, reliable and comparatively inexpensive method of investigation.

Structural Analysis of Anti-Hapten Antibodies to Identify Long-Range Structural Movements Induced by Hapten Binding.

Antibodies are well known for their high specificity that has enabled them to be of significant use in both therapeutic and diagnostic applications. Antibodies can recognize different antigens, including proteins, carbohydrates, peptides, nucleic acids, lipids, and small molecular weight haptens that are abundantly available as hormones, pharmaceuticals, and pesticides. Here we focus on a structural analysis of hapten-antibody couples and identify potential structural movements originating from the hapten binding by comparison with unbound antibody, utilizing 40 crystal structures from the Protein Data Bank. Our analysis reveals three binding surface trends; S1 where a pocket forms to accommodate the hapten, S2 where a pocket is removed when the hapten binds, and S3 where no pockets changes are found. S1 and S2 are expected for induced-fit binding, whereas S3 indicates that a pre-existing population of optimal binding antibody conformation exists. The structural analysis reveals four classifications of structural reorganization, some of which correlate to S2 but not to the other binding surface changes. These observations demonstrate the complexity of the antibody-antigen interaction, where structural changes can be restricted to the binding sites, or extend through the constant domains to propagate structural changes. This highlights the importance of structural analysis to ensure successful and compatible transformation of small antibody fragments at the early discovery stage into full antibodies during the subsequent development stages, where long-range structural changes are required for an Fc effector response.

Biomolecular interactions with nanoparticles: applications for coronavirus disease 2019.

Nanoparticles are small particles sized 1-100 nm, which have a large surface-to-volume ratio, allowing efficient adsorption of drugs, proteins, and other chemical compounds. Consequently, functionalized nanoparticles have potential diagnostic and therapeutic applications. A variety of nanoparticles have been studied, including those constructed from inorganic materials, biopolymers, and lipids. In this review, we focus on recent work targeting the severe acute respiratory syndrome coronavirus 2 virus that causes coronavirus disease (COVID-19). Understanding the interactions between coronavirus-specific proteins (such as the spike protein and its host cell receptor angiotensin-converting enzyme 2) with different nanoparticles paves the way to the development of new therapeutics and diagnostics that are urgently needed for the fight against COVID-19, and indeed for related future viral threats that may emerge.

siRNA Delivery to Melanoma Cells with Cationic Niosomes.

RNA interference (RNAi) is a posttranscriptional regulatory mechanism that employs siRNA. It typically results in the degradation of a target mRNA that encodes a particular protein. Treatment with siRNA therapeutics requires the use of an effective drug delivery system to assist in delivering these therapeutics into the cytoplasm of the transfected cells. Here we describe the transfection of melanoma cancer cells with siRNA using cationic niosome nanoparticles as a delivery system. The method of niosome preparation is first introduced and is followed by complex formation with siRNA and the transfection method.

Use of Nanoparticles in Delivery of Nucleic Acids for Melanoma Treatment.

Melanoma accounts for 4% of all skin cancer malignancies, with only 14% of diagnosed patients surviving for more than 5 years after diagnosis. Until now, there is no clear understanding of the detailed molecular contributors of melanoma pathogenesis. Accordingly, more research is needed to understand melanoma development and prognosis.All the treatment approaches that are currently applied have several significant limitations that prevent effective use in melanoma. One major limitation in the treatment of cancer is the acquisition of multidrug resistance (MDR). The MDR results in significant treatment failure and poor clinical outcomes in several cancers, including skin cancer. Treatment of melanoma is especially retarded by MDR. Despite the current advances in targeted and immune-mediated therapy, treatment arms of melanoma are severely limited and stand as a significant clinical challenge. Further, the poor pharmacokinetic profile of currently used chemotherapeutic agents is another reason for treatment failure. Therefore, more research is needed to develop novel drugs and carrier tools for more effective and targeted treatment.Nucleic acid therapy is based on nucleic acids or chemical compounds that are closely related, such as antisense oligonucleotides, aptamers, and small-interfering RNAs that are usually used in situations when a specific gene implicated in a disorder is deemed a therapeutically beneficial target for inhibition. However, the proper application for nucleic acid therapies is hampered by the development of an effective delivery system that can maintain their stability in the systemic circulation and enhance their uptake by the target cells. In this chapter, the prognosis of the different types of melanoma along with the currently used medications is highlighted, and the different types of nucleic acids along with the currently available nanoparticle systems for delivering these nucleic acids into melanoma cells are discussed. We also discuss recently conducted research on the use of different types of nanoparticles for nucleic acid delivery into melanoma cells and highlight the most significant outcomes.

Antitrypanosomal and Antileishmanial Activity of Chalcones and Flavanones from Polygonum salicifolium.

Trypanosomiasis and leishmaniasis are a group of neglected parasitic diseases caused by several species of parasites belonging to the family Trypansomatida. The present study investigated the antitrypanosomal and antileishmanial activity of chalcones and flavanones from Polygonum salicifolium, which grows in the wetlands of Iraq. The phytochemical evaluation of the plant yielded two chalcones, 2',4'-dimethoxy-6'-hydroxychalcone and 2',5'-dimethoxy-4',6'-dihydroxychalcone, and two flavanones, 5,7-dimethoxyflavanone and 5,8-dimethoxy-7-hydroxyflavanone. The chalcones showed a good antitrypanosomal and antileishmanial activity while the flavanones were inactive. The EC50 values for 2',4'-dimethoxy-6'-hydroxychalcone against Trypanosoma brucei brucei (0.5 µg/mL), T. congolense (2.5 µg/mL), and Leishmania mexicana (5.2 µg/mL) indicated it was the most active of the compounds. None of the compounds displayed any toxicity against a human cell line, even at 100 µg/mL, or cross-resistance with first line clinical trypanocides, such as diamidines and melaminophenyl arsenicals. Taken together, our study provides significant data in relation to the activity of chalcones and flavanones from P. salicifolium against both parasites in vitro. Further future research is suggested in order to investigate the mode of action of the extracted chalcones against the parasites.

Examination of the effect of niosome preparation methods in encapsulating model antigens on the vesicle characteristics and their ability to induce immune responses.

Niosome nanoparticles can be prepared using different methods, each of which can affect the size and homogeneity of the prepared particles. The aim of this study was to establish if the method of preparation impacted on the prepared vesicles when loaded with a model protein and the type of immune responses induced to the vaccine antigen. Niosomes were prepared using both the traditional thin film hydration (TFH) technique and the microfluidic mixing (MM) technique. Influenza antigen was then entrapped in the niosomes and formulations tested for their ability to induce in vivo immune responses in immunised BALB/c mice. Niosomes prepared by MM had a mean size of 157 ± 1.8 nm and were significantly more uniform compared with the niosomes prepared using TFH (mean size 388 ± 10 nm). Niosomes play a key role as an adjuvant to help raise high antibody immune responses. This was confirmed in this study since animals treated with both types of niosomes and antigen were more responsive than unentrapped (free) antigen. Cytokine analysis showed that the TFH niosomes induced a Th1 immune response by raising IgG2a and high levels of IFN-É£, while the MM niosomes induced a Th2 immune response by inducing IgG1 (p < .05). These results confirmed that the method of preparation of the niosomes nanoparticles induced different immune responses and the average particle size of the niosomes differed depending on the method of manufacture. This indicates that particle size and uniformity are of importance and should be taken into consideration when designing an oral based delivery system for vaccine delivery.

Antiparasitic and Cytotoxic Activity of Bokkosin, A Novel Diterpene-Substituted Chromanyl Benzoquinone From Calliandra portoricensis.

Calliandra portoricensis is a medicinal plant growing freely in Nigeria. It is used traditionally to treat tuberculosis, as an anthelmintic and an abortifacient. Phytochemical fractionation and screening of its root extracts has yielded a novel (5-hydroxy-7-methoxy-4-oxo-1-chromanyl)-4-methoxy-p-benzoquinone (breverin)-substituted cassane diterpene, which was designated bokkosin. It was obtained from column chromatography of the ethyl acetate extract of the roots. The compound was characterized using IR, NMR (1D and 2D) and mass spectral data. Promising antiparasitic activity was observed against the kinetoplastid parasite Trypanosoma brucei brucei, as well as moderate activity against Trypanosoma congolense and Leishmania mexicana and low toxicity in mammalian cells, with the best in vitro EC50 values against T. b. brucei (0.69 µg/mL against a standard laboratory strain, and its multi-drug resistant clone (0.33 µg/mL). The effect on T. b. brucei in culture was rapid and dose-dependent, leading to apparently irreversible growth arrest and cell death after an exposure of just 2 h at 2 × or 4 × EC50. The identification of bokkosin constitutes the first isolation of this class of compound from any natural source and establishes the compound as a potential trypanocide that, considering its novelty, should now be tested for activity against other microorganisms as well.

Antibody-protein binding and conformational changes: identifying allosteric signalling pathways to engineer a better effector response.

Numerous monoclonal antibodies have been developed successfully for the treatment of various diseases. Nevertheless, the development of biotherapeutic antibodies is complex, expensive, and time-consuming, and to facilitate this process, careful structural analysis beyond the antibody binding site is required to develop a more efficacious antibody. In this work, we focused on protein antigens, since they induce the largest antibody changes, and provide interesting cases to compare and contrast. The structures of 15 anti-protein antibodies were analysed to compare the antigen-bound/unbound forms. Surprisingly, three different classes of binding-induced changes were identified. In class (B1), the antigen binding fragment distorted significantly, and we found changes in the loop region of the heavy chain's constant domain; this corresponds well with expected allosteric movements. In class (B2), we found changes in the same loop region without the overall distortion. In class (B3), these changes did not present, and only local changes at the complementarity determining regions were found. Consequently, structural analysis of antibodies is crucial for therapeutic development. Careful evaluation of allosteric movements must be undertaken to develop better effector responses, especially during the transformation of these antibodies from small fragments at the discovery stage to full antibodies at the subsequent development stages.

Niosome-encapsulated balanocarpol: compound isolation, characterisation, and cytotoxicity evaluation against human breast and ovarian cancer cell lines.

Natural products have been successfully used to treat various ailments since ancient times and currently several anticancer agents based on natural products are used as the main therapy to treat cancer patients, or as a complimentary treatment to chemotherapy or radiation. Balanocarpol, which is a promising natural product that has been isolated from Hopea dryobalanoides, has been studied as a potential anticancer agent but its application is limited due to its high toxicity, low water solubility, and poor bioavailability. Therefore, the aim of this study is to improve the characteristics of balanocarpol and increase its anticancer activity through its encapsulation in a bilayer structure of a lipid-based nanoparticle drug delivery system where the application of nanotechnology can help improve the limitations of balanocarpol. The compound was first extracted and isolated from H. dryobalanoides. Niosome nanoparticles composed of Span 80 (SP80) and cholesterol were formulated through an innovative microfluidic mixing method for the encapsulation and delivery of balanocarpol. The prepared particles were spherical, small, and uniform with an average particles size and polydispersity index ∼175 nm and 0.088, respectively. The encapsulation of balanocarpol into the SP80 niosomes resulted in an encapsulation efficiency of ∼40%. The niosomes formulation loaded with balanocarpol showed a superior anticancer effect over the free compound when tested in vitro on human ovarian carcinoma (A2780) and human breast carcinoma (ZR-75-1). This is the first study to report the use of SP80 niosomes for the successful encapsulation and delivery of balanocarpol into cancer cells.