Size-selective purification of hepatitis B virus-like particle in flow-through chromatography: types of ion exchange adsorbent and grafted polymer architecture

Hepatitis B virus-like particles expressed in Escherichia coli were purified using anion exchange adsorbents grafted with polymer poly(oligo(ethylene glycol) methacrylate) in flow-through chromatography mode. The virus-like particles were selectively excluded, while the relatively smaller sized host cell proteins were absorbed. The exclusion of virus-like particles was governed by the accessibility of binding sites (the size of adsorbents and the charge of grafted dextran chains) as well as the architecture (branch-chain length) of the grafted polymer. The branch-chain length of grafted polymer was altered by changing the type of monomers used. The larger adsorbent (90 μm) had an approximately twofold increase in the flow-through recovery, as compared to the smaller adsorbent (30 μm). Generally, polymer-grafted adsorbents improved the exclusion of the virus-like particles. Overall, the middle branch-chain length polymer grafted on larger adsorbent showed optimal performance at 92% flow-through recovery with a purification factor of 1.53. A comparative study between the adsorbent with dextran grafts and the polymer-grafted adsorbent showed that a better exclusion of virus-like particles was achieved with the absorbent grafted with inert polymer. The grafted polymer was also shown to reduce strong interaction between binding sites and virus-like particles, which preserved the particles' structure

Targeted delivery of 5-fluorouracil-1-acetic acid (5-FA) to cancer cells overexpressing epithelial growth factor receptor (EGFR) using virus-like nanoparticles

Chemotherapy is widely used in cancer treatments. However, non-specific distribution of chemotherapeutic agents to healthy tissues and normal cells in the human body always leads to adverse side effects and disappointing therapeutic outcomes. Therefore, the main aim of this study was to develop a targeted drug delivery system based on the hepatitis B virus-like nanoparticle (VLNP) for specific delivery of 5-fluorouracil-1-acetic acid (5-FA) to cancer cells expressing epithelial growth factor receptor (EGFR). 5-FA was synthesized from 5-fluorouracil (5-FU), and it was found to be less toxic than the latter in cancer cells expressing different levels of EGFR. The cytotoxicity of 5-FA increased significantly after being conjugated on the VLNP. A cell penetrating peptide (CPP) of EGFR was displayed on the VLNP via the nanoglue concept, for targeted delivery of 5-FA to A431, HT29 and HeLa cells. The results showed that the VLNP displaying the CPP and harboring 5-FA internalized the cancer cells and killed them in an EGFR-dependent manner. This study demonstrated that the VLNP can be used to deliver chemically modified 5-FU derivatives to cancer cells overexpressing EGFR, expanding the applications of the VLNP in targeted delivery of chemotherapeutic agents to cancer cells overexpressing this transmembrane receptor

Isolation, identification and characterization of pentacholorophenol degrading bacterium

Pentachlorophenol (PCP) is a wide spectrum biocide applied in agriculture, industry and public health. PCP is lethal to all forms of life due to its nature to inhibit oxidative phosphorylation. Moreover, it is recalcitrant to biodegradation due to the stable aromatic ring and high chlorine content. Thus, the extensive use of PCP has led this chlorophenolic compound becoming a common environmental contaminant. The degradation of PCP is very important since it is highly toxic and carcinogenic. This study was conducted to isolate and characterize a potential PCP-degrading bacterium. The study also included the effectiveness of cell immobilization in enhancing PCP degradation by the bacterial cells as well as protecting the cells from the toxic effect of heavy metals which is the common co-contaminants at the sites of contamination. A potent PCP degrading bacterium has been isolated in this work. The bacterium was identified as Klebsiella sp. and assigned as Klebsiella sp. strain GBK1 based on 16S rDNA phylogenetic analysis. The 16S rDNA sequence has been submitted to genbank under the accession number FJ958193. The optimal PCP degradation conditions occurred at 30°C, pH 7 with 0.3 to 0.4 gL-1 ammonium chloride and 0.6% (w/v) glucose as supplemental carbon source. Klebsiella sp. strain GBK1 was able to degrade PCP with elevated concentration as high as 800 mgL-1. The efficiency of PCP degradation by Klebsiella sp. strain GBK1 was enhanced by immobilization with gellan gum as compared to freely suspended cells. Immobilized Klebsiella sp. strain GBK 1 best degraded PCP with gellan gum concentration of 0.75% (w/v), bead size of 5 mm diameter (estimated surface area of 78.54 mm2) and bead number of 500/100 ml medium. Degradation of PCP by both immobilized and free cells exhibited similar profile characteristics at lower concentrations of PCP. At higher concentration of PCP, the use of immobilized Klebsiella sp. strain GBK 1 resulted in rapid and extensive PCP degradation (degraded PCP up to 1750 mg/L) compared to free cells system whose PCP degrading ability is inhibited at concentration of 900 mg/L. Generally, the rate of PCP degradation was higher in immobilized Klebsiella sp. strain GBK1 and it is more tolerant to high concentrations of PCP (as high as 1750 mg/L). Also, the immobilized cells are more resistant towards heavy metals including As, Zn, Cr, Cd,Cu, Ni, Ag, Pb and Hg (as high as 20ppm) as compared to the free cells system whose activity were affected by 1ppm of heavy metals. In conclusion, Klebsiella sp. strain GBK1 is a potent candidate for bioremediation of PCP and its derivatives at chlorophenols contaminated sites and cell immobilization system was shown to be a more powerful system for biodegradation as compared to freely suspended cells system

Targeted 5-fluorouracil-1-acetic acid delivery utilising cell penetrating peptide conjugated hepatitis b virus-like nanoparticles to squamous cell carcinoma A431

Skin cancer is a prominent global public health problem with no signs of plateauing in its incidence. As the most common treatments for skin cancer, surgical resection inevitably damages a patient's appearance, and chemotherapy has many side effects. Thus, the main goal of the study was to screen for a cell penetrating peptide (CPP) for the development of a targeting drug delivery system applying truncated hepatitis B core antigen (tHBcAg) virus-like nanoparticles (VLNPs) for skin cancer. With the application of biopanning utilizing Ph.D.™-12 Phage Display Peptide Library, a CPP with the sequence NRPDSAQFWLHH that specifically targets the human squamous carcinoma A431 cells was identified. The uptake of the CPP by A431 cells is an energy dependent process, and the CPP was proven to enter the A431 cells via the interaction with epidermal growth factor receptor (EGFR), a transmembrane protein that is involved in cell signaling pathways that control cell division and survival. Methyl-β-cyclodextrin (MβCD) and chlorpromazine hydrochloride (CPZ) inhibited the internalisation of the CPP into the A431 cells, suggesting the peptide entered the cells via clathrin-dependent endocytosis. To conjugate the CPP to tHBcAg VLNPs, the CPP was cosynthesised with peptide SLLGRMKGA (the nanoglue) at its C-terminus, and these sequences were separated by a linker (GGG). The resulting 24-residue peptide, NRPDSAQFWLHHGGG-SLLGRMKGA, was directly coupled covalently to the spikes of the tHBcAg VLNPs via carboxylate groups using the zero length cross-linker EDC and Sulfo-NHS. The CPP displayed on hepatitis B virus-like nanoparticles (VLNPs) via the nanoglue successfully delivered the nanoparticles into A431 cells. The CPP conjugated tHBcAg VLNPs (CPP-tHBcAg VLNPs) were further conjugated with 5-fluorouracil-1-acetic acid (5-FA) via the primary amine groups on the surface of tHBcAg VLNPs using Sulfo-NHS/EDC as the targeting ligand for cancer chemotherapy. Transmission electron microscopy showed that the tHBcAg VLNPs maintained their integrity after the conjugation of the CPP and 5-FA on their surfaces. Approximately 833 5-FA molecules were conjugated covalently to each CPP-tHBcAg VLNP. 5-FA is the derivative of 5-fluorouracil (5-FU) which is less toxic but with enhanced cytotoxicity once it was coupled covalently to the CPP conjugated tHBcAg VLNPs. Furthermore, the targeting property of the CPP resulted in selective cytotoxicity of the 5-FA-CPP-tHBcAg VLNPs in EGFR-dependent manner. The MTT assay indicated that tHBcAg VLNPs conjugated with CPP and 5- FA significantly increased the cytotoxicity of 5-FA in A431 cells, which expressed the highest level of EGFR (2.14 folds) as compared with that of free 5-FU. The cytotoxicity of 5-FA-CPP-tHBcAg VLNPs was reduced in HT29 cells and HeLa cells, which have a lower number of EGFR per cell. This demonstrated that the CPP is a potential ligand for targeted delivery of VLNPs into skin cancer cells and other cancer cells, which overexpress epithelial growth factor receptor (EGFR). This paves the way to deliver drugs, nucleic acids and molecules into cells overexpressing EGFR. The application of this peptide is not restricted as a ligand to target and internalise VLNPs into cells, it can also be incorporated into other nanoparticles for a wider application in nanomedicine and targeting cancer imaging. In addition, the newly established drug delivery systems demonstrated that the tHBcAg VLNP is a potential nano-vehicle in chemotherapeutics to target various cells specifically by displaying different cell specific ligands at the tips of tHBcAg VLNPs

Next-generation viral nanoparticles for targeted delivery of therapeutics: Fundamentals, methods, biomedical applications and challenges

10.1080/17425247.2023.2228202Expert Opinion on Drug Delivery207955-97

Enhanced phenol degradation by immobilized Acinetobacter sp. strain AQ5NOL 1

A locally isolated Acinetobacter sp. Strain AQ5NOL 1 was encapsulated in gellan gum and its ability to degrade phenol was compared with the free cells. Optimal phenol degradation was achieved at gellan gum concentration of 0.75% (w/v), bead size of 3 mm diameter (estimated surface area of 28.26 mm2) and bead number of 300 per 100 ml medium. At phenol concentration of 100 mg l−1, both free and immobilized bacteria exhibited similar rates of phenol degradation but at higher phenol concentrations, the immobilized bacteria exhibited a higher rate of degradation of phenol. The immobilized cells completely degrade phenol within 108, 216 and 240 h at 1,100, 1,500 and 1,900 mg l−1 phenol, respectively, whereas free cells took 240 h to completely degrade phenol at 1,100 mg l−1. However, the free cells were unable to completely degrade phenol at higher concentrations. Overall, the rates of phenol degradation by both immobilized and free bacteria decreased gradually as the phenol concentration was increased. The immobilized cells showed no loss in phenol degrading activity after being used repeatedly for 45 cycles of 18 h cycle. However, phenol degrading activity of the immobilized bacteria experienced 10 and 38% losses after the 46 and 47th cycles, respectively. The study has shown an increased efficiency of phenol degradation when the cells are encapsulated in gellan gum

Targeted delivery of cell penetrating peptide virus-like nanoparticles to skin cancer cells

Skin cancer or cutaneous carcinoma, is a pre-eminent global public health problem with no signs of plateauing in its incidence. As the most common treatments for skin cancer, surgical resection inevitably damages a patient's appearance, and chemotherapy has many side effects. Thus, the main aim of this study was to screen for a cell penetrating peptide (CPP) for the development of a targeting vector for skin cancer. In this study, we identified a CPP with the sequence NRPDSAQFWLHH from a phage displayed peptide library. This CPP targeted the human squamous carcinoma A431 cells through an interaction with the epidermal growth factor receptor (EGFr). Methyl-β-cyclodextrin (MβCD) and chlorpromazine hydrochloride (CPZ) inhibited the internalisation of the CPP into the A431 cells, suggesting the peptide entered the cells via clathrin-dependent endocytosis. The CPP displayed on hepatitis B virus-like nanoparticles (VLNPs) via the nanoglue successfully delivered the nanoparticles into A431 cells. The present study demonstrated that the novel CPP can serve as a ligand to target and deliver VLNPs into skin cancer cells

Comparison of factors associated with disordered eating between male and female Malaysian university students

Disordered eating is prevalent among university students, especially females. Whilst literature suggests that factors associated with disordered eating may differ according to gender, such an association has not been studied in Malaysia. This cross-sectional study aims to compare factors associated with disordered eating between male and female university students. A total of 716 university students (male: 27.4%; female: 72.6%) were recruited in Kuala Lumpur and Selangor, Malaysia. All participants completed a set of self-administered questionnaires and their body weight and height were recorded. About one in five of the university students (20.3%) were found to have disordered eating. There were more female students (22.9%) disordered eating compared to males (13.3%, χ2 = 8.16, p < 0.05). In male students (β = 0.228, p < 0.01), depressive symptoms were the only significant predictor for disordered eating. In females, the strongest predictor was depressive symptoms (β = 0.214, p < 0.001), followed by body size satisfaction (β = −0.145, p < 0.01) and body appreciation (β = −0.101, p < 0.05). These findings suggest that there are gender differences in the factors associated with disordered eating among Malaysian university students. Intervention programmes that address disordered eating should take into account these sex differences and its contributing factors

Size-selective purification of hepatitis B virus-like particle in flow-through chromatography: Types of ion exchange adsorbent and grafted polymer architecture

Hepatitis B virus‐like particles expressed in Escherichia coli were purified using anion exchange adsorbents grafted with polymer poly(oligo(ethylene glycol) methacrylate) in flow‐through chromatography mode. The virus‐like particles were selectively excluded, while the relatively smaller sized host cell proteins were absorbed. The exclusion of virus‐like particles was governed by the accessibility of binding sites (the size of adsorbents and the charge of grafted dextran chains) as well as the architecture (branch‐chain length) of the grafted polymer. The branch‐chain length of grafted polymer was altered by changing the type of monomers used. The larger adsorbent (90 μm) had an approximately twofold increase in the flow‐through recovery, as compared to the smaller adsorbent (30 μm). Generally, polymer‐grafted adsorbents improved the exclusion of the virus‐like particles. Overall, the middle branch‐chain length polymer grafted on larger adsorbent showed optimal performance at 92% flow‐through recovery with a purification factor of 1.53. A comparative study between the adsorbent with dextran grafts and the polymer‐grafted adsorbent showed that a better exclusion of virus‐like particles was achieved with the absorbent grafted with inert polymer. The grafted polymer was also shown to reduce strong interaction between binding sites and virus‐like particles, which preserved the particles’ structure