The seroprevalence of toxocariasis and related risk factors in children in Ho Chi Minh City, Vietnam: results from a school-based cross-sectional study.

BACKGROUND: Children are especially vulnerable to Toxocara infection and its severe complications; however, there have not been any published data on the disease prevalence and treatment effectiveness in the population of Vietnamese children. This study was conducted to determine the prevalence of toxocariasis and explore factors associated with Toxocara infection in children aged 3-15 y in Ho Chi Minh City, Vietnam. METHODS: We conducted a cross-sectional study using a multistage cluster sampling approach in public schools. Blood samples were collected, and toxocariasis cases were confirmed, based on a history of contact with dogs/cats and positive anti-Toxocara antibody detection via ELISA. We calculated the percentage of seropositive children across gender, grade levels, districts and caregiver education. Multiple regression models were employed to identify potential risk factors. RESULTS: Anti-Toxocara antibodies were found in 14.2% of the 986 children studied. Significant variations in seropositivity were observed across grade levels, districts and caregiver education levels. Multivariable analysis identified caregiver education, contact with dogs/cats and improper handling of pet feces as seropositivity risk factors. CONCLUSION: This was the first community-based prevalence study of toxocariasis in a pediatric population in Vietnam. Implementation of preventive measures such as public education, routine fecal examinations and chemotherapeutic treatment of animals is highly recommended.

Light-Responsive Elastin-Like Peptide-Based Targeted Nanoparticles for Enhanced Spheroid Penetration.

We describe here a near infrared light-responsive elastin-like peptide (ELP)-based targeted nanoparticle (NP) that can rapidly switch its size from 120 to 25 nm upon photo-irradiation. Interestingly, the targeting function, which is crucial for effective cargo delivery, is preserved after transformation. The NPs are assembled from (targeted) diblock ELP micelles encapsulating photosensitizer TT1-monoblock ELP conjugates. Methionine residues in this monoblock are photo-oxidized by singlet oxygen generated from TT1, turning the ELPs hydrophilic and thus trigger NP dissociation. Phenylalanine residues from the diblocks then interact with TT1 via π-π stacking, inducing the re-formation of smaller NPs. Due to their small size and targeting function, the NPs penetrate deeper in spheroids and kill cancer cells more efficiently compared to the larger ones. This work could contribute to the design of "smart" nanomedicines with deeper penetration capacity for effective anticancer therapies.

Significant gaps in hepatitis B vaccination in adults in Viet Nam: Important targets toward hepatitis B elimination by 2030.

BACKGROUND: Gaps in adult hepatitis B vaccination were undefined in Vietnam, a lower-middle-income country. To address these gaps, this study defined hepatitis B vaccine coverage in adults and its associated factors in Ho Chi Minh City (HCMC), Viet Nam. We also proposed interventional strategies, prioritizing gap identification to facilitate hepatitis B elimination by 2030 and beyond. METHOD: During 2019-2020, a multi-stage cluster serosurvey with probability proportional to size was conducted to representatively invite 20,000 adults (18 years or older) throughout HCMC for hepatitis B screening (HBsAg, anti-HBs, and anti-HBc). Serologic results defined two dependent variables: vaccine-induced immunity (i.e., isolated anti-HBs) and susceptibility (i.e., HBV naive). Associations of dependent variables with surveyed demographics, socioeconomic statuses, behaviors, and medical history at risk for hepatitis B were evaluated using weighted Poisson regression. RESULTS: The prevalence was 18.5% (95%CI, 17.3-20.0%) for vaccine-induced immunity and 37.7% (35.6-39.8%) for susceptibility. Even though analyses in the general population revealed a falling trend in vaccine-induced immunity prevalence from younger to older age groups, sensitivity analyses in the non-infected population (i.e., those who were both negative for HBsAg and anti-HBc) showed that younger age groups, especially those aged 30 to 50 years, had the lowest prevalence. Social inequalities existed in different ethnicities, residence areas, education levels, house ownership, and health insurance statuses. There was no significant association between vaccine-induced immunity or susceptibility and risky behaviors and medical histories. CONCLUSION: This study depicts a significant unmet need for hepatitis B vaccination in the general adult population in HCMC, Viet Nam. Indeed, the lack of vaccination was unevenly distributed regarding age groups, geographical areas, and socioeconomic statuses, which reveals profound social disparities. Therefore, to achieve hepatitis B elimination goals, besides the current recommendations for infants and risk-based strategies, hepatitis B vaccination should be recommended for the broader population.

Establishing baseline framework for hepatitis B virus micro-elimination in Ho Chi Minh City, Vietnam - A community-based seroprevalence study.

Background: We conducted a community-based seroprevalence study using three HBV seromarkers (HBsAg, anti-HBs, anti-HBc) in Ho Chi Minh City (HCMC), Vietnam, to (1) determine the prevalence of HBV serologic profiles; (2) document factors associated with HBV infection or susceptibility; and (3) propose strategies toward HBV elimination by 2030. Methods: During 2019-2020, we deployed a multistage cluster design with probability proportionate to size, to recruit 20,000 adults for an HBV screening and linkage to care program citywide. Screening results with interpretation, recommendations, and health education materials were returned to participants. Post-study surveys were conducted within three months to identify gaps in linkage to care. Findings: Of the 17,600 adults invited, 15,275 (86.7%) participated in the study, 14,674 (96.1%) completing all data for final analyses. The prevalence of HBsAg (+) and HBV-naïve were 7.5% and 37.7%, respectively. HBV vaccination rates were 18.7% and about 50% of HCMC population had been exposed to HBV. Of the persons with HBsAg (+), 27.1% linked to care (76% used health insurance). There were wide variations in HBsAg (+) and HBV vaccination rates between districts, risk factors, and socio-economic statuses. Interpretation: The significant disease burden of and gaps in the continuum of care highlight the need and urgency to address the HBV public health problem in Vietnam. Using three screening seromarkers that tailor interventions to the needs of HBV micro-populations could be an effective strategy to pursue HBV elimination goals. Funding: Gilead Sciences Inc; Roche Diagnostic International Ltd; Roche Diagnostics-Vietnam; Abbott Diagnostics-Vietnam; Hepatitis B Foundation; Medic MedicalCenter, Vietnam; Center of Excellence for Liver Disease in Vietnam, Johns Hopkins University School of Medicine.

Demonstration of a population-based HCV serosurvey in Ho Chi Minh City, Viet Nam: Establishing baseline prevalence of and continuum of care for HCV micro-elimination by 2030.

Background: A baseline of hepatitis C virus (HCV) burden and other HCV epidemiological profiles is necessary for HCV micro-elimination in Ho Chi Minh City (HCMC), Viet Nam. This study aimed to determine HCV exposure and prevalence of HCV viremia as well as the proportion of HCV testing and treatment uptake among participants. Methods: From 2019 to 2020, the probability proportionate to size sampling method was deployed to representatively invite approximately 20,000 adults (18 or older) throughout HCMC to free screening and linkage to care for HCV. Findings: In HCMC, the weighted prevalence of anti-HCV was 1·3% (95% CI, 1·1%-1·6%). Individuals born from 1945 to 1964 had the anti-HCV prevalence of 3·6% (95% CI, 3·0%-4·2%) and represented 40·4% of all HCV cases. There were wide variations in anti-HCV prevalence in HCMC, including variations between districts, risk factors, and socioeconomic statuses. A baseline HCV continuum of care for the city demonstrated that only 28·5% (85/298, 95%CI 23·4-33·7%) of persons with anti-HCV (+) were aware of their HCV status, with 77.6% (66/85, 95%CI 68·8-86·5%) diagnosing HCV incidentally, 82·7% (62/75, 95%CI 74·1-91·2%) initiating anti-HCV therapy, and 53.6% (30/56, 95%CI 40·5-66·6%) achieving HCV cures. Interpretation: There remains a considerable disease burden of HCV in HCMC of which a significant proportion was in the age group born between 1945 to 1964. Additionally, there were significant gaps in HCV awareness, screening, and access to care in the community in Viet Nam. Thus, future interventions must have pragmatic targets, be tailored to the local needs, and emphasise screening. Funding: This work was supported by investigator-sponsored research grants from Gilead Sciences Inc. (Grant No: IN-US-987-5382); Roche Diagnostic International Ltd. (Grant No. SUB-000196); and in-kind donations from Abbott Diagnostic Viet Nam; Hepatitis B Foundation; Medic Medical Center, Viet Nam; Johns Hopkins University School of Medicine's Center of Excellence for Liver Disease in Viet Nam; and the Board of Directors, Viet Nam Viral Hepatitis Alliance (V-VHA).

Self-Assembly or Coassembly of Multiresponsive Histidine-Containing Elastin-Like Polypeptide Block Copolymers.

In this study a histidine containing elastin-like polypeptide (ELP) diblock copolymer is described with multiresponsive assembly behavior. Self-assembly into micelles is examined by two methods. First, the self-assembly is triggered by the addition of divalent metal ions, with Zn2+ being the most suitable one. Increasing the Zn2+ concentration stabilizes the nanoparticles over a large temperature window (4-45 °C). This diblock exhibits furthermore pH-responsiveness, and particles disassemble under mildly acidic conditions. Second, the coassembly of this ELP with a diblock ELP is examined, which is not responsive to pH and metal ions. Coassembly is triggered by heating the ELPs quickly above the transition temperature of the less hydrophobic block, which results in stable nanoparticles without the need to add metal ions. This novel ELP system offers a versatile modular nanocarrier platform that can respond to different stimuli and can be tuned effectively.

Pathway-Dependent Co-Assembly of Elastin-Like Polypeptides.

In natural systems, temperature-induced assembly of biomolecules can lead to the formation of distinct assembly states, created out of the same set of starting compounds, based on the heating trajectory followed. Until now it has been difficult to achieve similar behavior in synthetic polymer mixtures. Here, a novel pathway-dependent assembly based on stimulus-responsive polymers is shown. When a mixture of mono- and diblock copolymers, based on elastin-like polypeptides, is heated with a critical heating rate co-assembled particles are created that are monodisperse, stable, and have tunable hydrodynamic radii between 20 and 120 nm. Below this critical heating rate, the constituents separately form polymer assemblies. This process is kinetically driven and reversible in thermodynamically closed systems. Using the co-assembly pathway, fluorescent proteins and bioluminescent enzymes are encapsulated with high efficiency. Encapsulated cargo shows unperturbed function even after delivery into cells. The pathway-dependent co-assembly of elastin-like polypeptides is not only of fundamental interest from a materials science perspective, allowing the formation of multiple distinct assemblies from the same starting compounds, which can be interconverted by going back to the molecularly dissolved states. It also enables a versatile way for constructing highly effective vehicles for the cellular delivery of biomolecular cargo.

Bluetongue Virus Particles as Nanoreactors for Enzyme Delivery and Cancer Therapy.

The side effects of chemotherapy can be reduced by targeting tumor cells with an enzyme (or the corresponding gene) that converts a nontoxic prodrug into a toxic drug inside the tumor cells, also killing the surrounding tumor cells via the bystander effect. Viruses are the most efficient gene delivery vehicles because they have evolved to transfer their own nucleic acids into cells, but their efficiency must be balanced against the risks of infection, the immunogenicity of nucleic acids, and the potential for genomic integration. We therefore tested the effectiveness of genome-free virus-like particles (VLPs) for the delivery of Herpes simplex virus 1 thymidine kinase (HSV1-TK), the most common enzyme used in prodrug conversion therapy. HSV1-TK is typically delivered as a gene, but in the context of VLPs, it must be delivered as a protein. We constructed VLPs and smaller core-like particles (CLPs) based on Bluetongue virus, with HSV1-TK fused to the inner capsid protein VP3. TK-CLPs and TK-VLPs could be produced in large quantities in plants. The TK-VLPs killed human glioblastoma cells efficiently in the presence of ganciclovir, with an IC50 value of 14.8 µM. Conversely, CLPs were ineffective because they remained trapped in the endosomal compartment, in common with many synthetic nanoparticles. VLPs are advantageous because they can escape from endosomes and therefore allow HSV1-TK to access the cytosolic adenosine triphosphate (ATP) required for the phosphorylation of ganciclovir. The VLP delivery strategy of TK protein therefore offers a promising new modality for the treatment of cancer with systemic prodrugs such as ganciclovir.

Rheology of Dispersions of High-Aspect-Ratio Nanofibers Assembled from Elastin-Like Double-Hydrophobic Polypeptides.

Elastin-like polypeptides (ELPs) are promising candidates for fabricating tissue-engineering scaffolds that mimic the extracellular environment of elastic tissues. We have developed a "double-hydrophobic" block ELP, GPG, inspired by non-uniform distribution of two different hydrophobic domains in natural elastin. GPG has a block sequence of (VGGVG)5-(VPGXG)25-(VGGVG)5 that self-assembles to form nanofibers in water. Functional derivatives of GPG with appended amino acid motifs can also form nanofibers, a display of the block sequence's robust self-assembling properties. However, how the block length affects fiber formation has never been clarified. This study focuses on the synthesis and characterization of a novel ELP, GPPG, in which the central sequence (VPGVG)25 is repeated twice by a short linker sequence. The self-assembly behavior and the resultant nanostructures of GPG and GPPG were when compared through circular dichroism spectroscopy, atomic force microscopy, and transmission electron microscopy. Dynamic rheology measurements revealed that the nanofiber dispersions of both GPG and GPPG at an extremely low concentration (0.034 wt%) exhibited solid-like behavior with storage modulus G' > loss modulus G" over wide range of angular frequencies, which was most probably due to the high aspect ratio of the nanofibers that leads to the flocculation of nanofibers in the dispersion.

Presentation and Delivery of Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand via Elongated Plant Viral Nanoparticle Enhances Antitumor Efficacy.

Potato virus X (PVX) is a flexuous plant virus-based nanotechnology with promise in cancer therapy. As a high aspect ratio biologic (13 × 515 nm), PVX has excellent spatial control in structures and functions, offering high-precision nanoengineering for multivalent display of functional moieties. Herein, we demonstrate the preparation of the PVX-based nanocarrier for delivery of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a promising protein drug that induces apoptosis in cancer cells but not healthy cells. TRAIL bound to PVX by coordination bonds between nickel-coordinated nitrilotriacetic acid on PVX and His-tag on the protein could mimic the bioactive "membrane-bound" state in native TRAIL, resulting in an elongated nanoparticle displaying up 490 therapeutic protein molecules. Our data show that PVX-delivered TRAIL activates caspase-mediated apoptosis more efficiently compared to soluble TRAIL; also in vivo the therapeutic nanoparticle outperforms in delaying tumor growth in an athymic nude mouse model bearing human triple-negative breast cancer xenografts. This proof-of-concept work highlights the potential of filamentous plant virus nanotechnologies, particularly for targeting protein drug delivery for cancer therapy.

Biodistribution of Filamentous Plant Virus Nanoparticles: Pepino Mosaic Virus versus Potato Virus X.

Nanoparticles with high aspect ratios have favorable attributes for drug delivery and bioimaging applications based on their enhanced tissue penetration and tumor homing properties. Here, we investigated a novel filamentous viral nanoparticle (VNP) based on the Pepino mosaic virus (PepMV), a relative of the established platform Potato virus X (PVX). We studied the chemical reactivity of PepMV, produced fluorescent versions of PepMV and PVX, and then evaluated their biodistribution in mouse tumor models. We found that PepMV can be conjugated to various small chemical modifiers including fluorescent probes via the amine groups of surface-exposed lysine residues, yielding VNPs carrying payloads of up to 1600 modifiers per particle. Although PepMV and PVX share similarities in particle size and shape, PepMV achieved enhanced tumor homing and less nonspecific tissue distribution compared to PVX in mouse models of triple negative breast cancer and ovarian cancer. In conclusion, PepMV provides a novel tool for nanomedical research but more research is needed to fully exploit the potential of plant VNPs for health applications.

Combination of Plant Virus Nanoparticle-Based in Situ Vaccination with Chemotherapy Potentiates Antitumor Response.

Immunotherapeutics are gaining more traction in the armamentarium used to combat cancer. Specifically, in situ vaccination strategies have gained interest because of their ability to alter the tumor microenvironment to an antitumor state. Herein, we investigate whether flexuous plant virus-based nanoparticles formed by the potato virus X (PVX) can be used as an immunotherapeutic for in situ vaccine monotherapy. We further developed dual chemo-immunotherapeutics by incorporating doxorubicin (DOX) into PVX yielding a dual-functional nanoparticle (PVX-DOX) or by coadministration of the two therapeutic regimes, PVX immunotherapy and DOX chemotherapy (PVX+DOX). In the context of B16F10 melanoma, PVX was able to elicit delayed tumor progression when administered as an intratumoral in situ vaccine. Furthermore, the coadministration of DOX via PVX+DOX enhanced the response of the PVX monotherapy through increased survival, which was also represented in the enhanced antitumor cytokine/chemokine profile stimulated by PVX+DOX when compared to PVX or DOX alone. Importantly, coadministered PVX+DOX was better for in situ vaccination than PVX loaded with DOX (PVX-DOX). Whereas the nanomedicine field strives to design multifunctional nanoparticles that integrate several functions and therapeutic regimens into a single nanoparticle, our data suggest a paradigm shift; some therapeutics may need to be administered separately to synergize and achieve the most potent therapeutic outcome. Altogether, our studies show that development of plant viral nanoparticles for in situ vaccines for treatment is a possibility, and dual mechanistic therapeutics can increase efficacy. Nonetheless, combining immunotherapeutics with cytolytic chemotherapy requires detailed investigation to inform optimal integration of cytolytic and immunotherapies and maximize synergy and efficacy.

Chemical addressability of potato virus X for its applications in bio/nanotechnology.

Potato virus X (PVX), a type member of the plant virus potexvirus group, offers a unique nanotechnology platform based on its high aspect ratio and flexible filamentous shape. The PVX platform has already been engineered and studied for its uses in imaging, drug delivery, and immunotherapies. While genetic engineering procedures are well established for PVX, there is limited information about chemical conjugation strategies for functionalizing PVX, partly due to the lack of structural information of PVX at high resolution. To overcome these challenges, we built a structural model of the PVX particle based on the available structures from pepino mosaic virus (PepMV), a close cousin of PVX. Using the model and a series of chemical conjugation experiments, we identified and probed the addressability of cysteine side chains. Chemical reactivity of cysteines was confirmed using Michael-addition and thiol-selective probes, including fluorescent dyes and biotin tags. LC/MS/MS was used to map Cys 121 as having the highest selectivity for modification. Finally, building on the availability of two reactive groups, the newly identified Cys and previously established Lys side chains, we prepared multifunctional PVX nanoparticles by conjugating Gd-DOTA for magnetic resonance imaging (MRI) to lysines and fluorescent dyes for optical imaging to cysteines. The resulting functionalized nanofilament could have applications in dual-modal optical-MRI imaging applications. These results further extend the understanding of the chemical properties of PVX and enable development of novel multifunctional platforms in bio/nanotechnology.

Double-hydrophobic elastin-like polypeptides with added functional motifs: Self-assembly and cytocompatibility.

We have recently developed a novel double-hydrophobic elastin-like triblock polypeptide called GPG, designed after the uneven distribution of two different hydrophobic domains found in elastin, an extracellular matrix protein providing elasticity and resilience to tissues. Upon temperature trigger, GPG undergoes a sequential self-assembling process to form flexible beaded nanofibers with high homogeneity and excellent dispersibility in water. Given that GPG might be a potential elastin-mimetic material, we sought to explore the biological activities of this block polypeptide. Besides GPG, several functionalized derivatives were also constructed by fusing functional motifs such as KAAK or KAAKGRGDS at the C-terminal of GPG. Although the added motifs affected the kinetics of fiber formation and ß-sheet contents, all three GPGs assembled into beaded nanofibers at the physiological temperature. The resulting GPG nanofibers preserved their beaded structures in cell culture medium; therefore, they were coated on polystyrene substrates to study their cytocompatibility toward mouse embryonic fibroblasts, NIH-3T3. Among the three polypeptides, GPG having the cell-binding motif GRGDS derived from fibronectin showed excellent cell adhesion and cell proliferation properties compared to other conventional materials, suggesting its promising applications as extracellular matrices for mammalian cells. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2475-2484, 2017.

Potato virus X, a filamentous plant viral nanoparticle for doxorubicin delivery in cancer therapy.

Plant viral nanoparticles (VNPs) are a novel class of nanocarriers with implications for drug delivery in cancer therapy. VNPs are characterized by their highly symmetrical nanoscale structures. Furthermore, plant VNPs are biocompatible, biodegradable, and non-infectious in mammals. VNPs provide a proteinaceous platform technology that can be readily engineered to carry contrast agents and therapies using chemical and genetic modifications. Of particular interest are high aspect ratio, elongated filaments such as the ones formed by potato virus X (PVX, measuring 515 × 13 nm). PVX has demonstrated enhanced tumor homing and penetration properties compared to spherical counterparts. Here, we sought to investigate the potential of PVX as a drug carrier delivering doxorubicin (DOX), a commonly used cancer chemotherapy. We synthesized therapeutic PVX nanoparticles using a simple in-solution mixing protocol; after 5 days of mixing of DOX and PVX and ultra-centrifugal purification, ∼1000 DOX per PVX were stably associated with the carrier, most likely based on hydrophobic interaction. Efficacy and drug activity of PVX-DOX were confirmed using a panel of cancer cell lines including ovarian cancer, breast cancer, and cervical cancer. Lastly, we demonstrated treatment of athymic mice bearing human MDA-MB-231 breast cancer xenografts: PVX-DOX treatment resulted in reduced tumor growth in this model. Our results open the door for further development of PVX and other high aspect ratio plant VNPs for applications in cancer therapy.