Polyphenol-Enabled 2D Nanopatch for Enhanced Nasal Mucoadhesion and Immune Activation.

The advancement of effective nasal mucoadhesive delivery faces challenges due to rapid mucociliary clearance (MCC). Conventional studies have employed mucoadhesive materials, mainly forming spherical nanoparticles, but these offer limited adhesion to the nasal mucosa. This study hypothesizes that a 2D nanoscale structure utilizing adhesive polyphenols can provide a superior strategy for countering MCC, aligning with the planar mucosal layers. We explore the use of tannic acid (TA), a polyphenolic molecule known for its adhesive properties and ability to form complexes with biomolecules. Our study introduces an unprecedented 2D nanopatch, assembled through the interaction of TA with green fluorescent protein (GFP), and cell-penetrating peptide (CPP). This 2D nanopatch demonstrates robust adhesion to nasal mucosa and significantly enhances immunoglobulin A secretions, suggesting its potential for enhancing nasal vaccine delivery. The promise of a polyphenol-enabled adhesive 2D nanopatch signifies a pivotal shift from conventional spherical nanoparticles, opening new pathways for delivery strategies through respiratory mucoadhesion.

Benefits of topical indigo naturalis nanofibrous patch on psoriatic skin: A transdermal strategy for botanicals.

Indigo naturalis (IN) has been extensively used as a topical treatment for psoriasis. However, clinical applications of IN in ointment were hampered by its limited transdermal efficiency and dark stains. To address the aforementioned issues, nanopatches carrying IN were fabricated using poly(ε-caprolactone, PCL)/poly(ethylene oxide, PEO) and topically applied to psoriasiform skin. The ideal ratio of 5% PCL/PEO was established to be 80:20 (w/w), and 15% IN as payload was confirmed. Investigations on the three principal active components of IN release indicated that indirubin and tryptanthrin were released in bursts, while indigo was released in a limited and controlled manner. Further biological analyses confirmed a favorable biocompatibility of amphiphilic IN-PCL/PEO, which coincided with the intended therapeutic outcomes as measured by severity index scoring and pathological evaluations in vivo. The advantages of IN as nanopatches over ointment could be due to improved transdermal distribution of indirubin and tryptanthrin, resulting in effective management of epidermal hyperplasia and blood vessel remodeling. Meanwhile, due to the lower preservation of epidermal indigo, IN-PCL/PEO nanopatches caused no skin coloration. Similarly, during a 4-week topical treatment of IN-PCL/PEO nanopatches, the safety and anti-psoriatic benefits were obtained in an initial human test. The conversion of IN from topical cream to electrospun nanofibers opens up new avenues for bench-to-bedside translation of this herbal therapy and provides mechanistic insight into IN's roles in the management of psoriasis.

Ultrafast Lifetime and Bright Emission from Graphene Quantum Dots Using Plasmonic Nanogap Cavities.

Graphene quantum dots (GQDs) are quasi-zero-dimensional, carbon-based luminescent nanomaterials that possess desirable physical properties, such as high photostability, low cytotoxicity, good biocompatibility, and excellent water solubility; however, their long radiative lifetimes significantly limit their use in, e.g., light emitting devices where a fast spontaneous emission rate is essential. Despite a few reports on GQD fluorescence enhancements using metal nanostructures, studies of enhanced spontaneous emission rate remain outstanding. Here, we report fast and bright luminescence by coupling gap plasmon modes to nanoparticle emitters. Through precise control over the nanoparticle's local density of states (LDOS), we achieved a 220-fold increase in the PL intensity. The shortest radiative lifetime obtained was below 8.0 ps and limited by the instrument response, which is over 288-fold shorter than the lifetime of uncoupled GQDs. These findings may benefit the future development of rapid displays and open the possibility of constructing high-frequency classical or quantum telecommunication systems.

Hybrid cube-in-cup nanoantenna: towards ordered photonics.

The most significant goal of nanophotonics is the development of high-speed quantum emitting devices operating at ambient temperature. In this regard, plasmonic nanoparticles-on-mirror are potential candidates for designing high-speed photon sources. We introduce a novel hybrid nanoantenna (HNA) with CdSe/CdS colloidal quantum dots (QDs) based on a silver nanocube in a metal cup that presents a nanoparticle-in-cavity coupled with an emitters system. We use focused ion beam nanolithography to fabricate an ordered array of cups, which were then filled with colloidal nanoparticles using the most simple drop-casting and spin coating methods. The spectral and time-resolved studies of the samples with one or more nanocubes in the cup reveal a significant change in the radiation characteristics of QDs inside the nanoantenna. The Purcell effect causes an increase in the fluorescence decay rate (≥30) and an increase in the fluorescence intensity (≥3) of emitters in the HNA. Using the finite element method simulations, we have discovered that the proximity of the cups wall affects the oscillation modes of the gap plasmon, which, in turn, leads to changes in the electric field enhancement inside the nanoantenna gap. Additionally, substantial variations in the behavior of the gap plasmons at different polarizations of the exciting radiation have been revealed. The proposed nanoantenna can be useful in the development of plasmonic sensors, display pixels, and single-photon sources.

Formulation Development and Improved Stability of a Combination Measles and Rubella Live-Viral Vaccine Dried for Use in the NanopatchTM Microneedle Delivery System.

Measles (Me) and rubella (Ru) viral diseases are targeted for elimination by ensuring a high level of vaccination coverage worldwide. Less costly, more convenient MeRu vaccine delivery systems should improve global vaccine coverage, especially in low - and middle - income countries (LMICs). In this work, we examine formulating a live, attenuated Me and Ru combination viral vaccine with Nanopatch™, a solid polymer micro-projection array for intradermal delivery. First, high throughput, qPCR-based viral infectivity and genome assays were established to enable formulation development to stabilize Me and Ru in a scaled-down, custom-built evaporative drying system to mimic the Nanopatch™ vaccine coating process. Second, excipient screening and optimization studies identified virus stabilizers for use during the drying process and upon storage in the dried state. Finally, a series of real-time and accelerated stability studies identified eight candidate formulations that met a target thermal stability criterion for live vaccines (<1 log10 loss after 1 week storage at 37°C). Compared to -80°C control samples, the top candidate formulations resulted in minimal viral infectivity titer losses after storage at 2-8°C for 6 months (i.e., <0.1 log10 for Me, and ~0.4 log10 for Ru). After storage at 25°C over 6 months, ~0.3-0.5 and ~1.0-1.4 log10 titer losses were observed for Me and Ru, respectively, enabling the rank-ordering of the stability of candidate formulations. These results are discussed in the context of future formulation challenges for developing microneedle-based dosage forms containing stabilized live, attenuated viral vaccines for use in LMICs.

Efficient Delivery of Dengue Virus Subunit Vaccines to the Skin by Microprojection Arrays.

Dengue virus is the most important arbovirus impacting global human health, with an estimated 390 million infections annually, and over half the world's population at risk of infection. While significant efforts have been made to develop effective vaccines to mitigate this threat, the task has proven extremely challenging, with new approaches continually being sought. The majority of protective, neutralizing antibodies induced during infection are targeted by the envelope (E) protein, making it an ideal candidate for a subunit vaccine approach. Using truncated, recombinant, secreted E proteins (sE) of all 4 dengue virus serotypes, we have assessed their immunogenicity and protective efficacy in mice, with or without Quil-A as an adjuvant, and delivered via micropatch array (MPA) to the skin in comparison with more traditional routes of immunization. The micropatch contains an ultra-high density array (21,000/cm2) of 110 µm microprojections. Mice received 3 doses of 1 µg (nanopatch, intradermal, subcutaneous, or intra muscular injection) or 10 µg (intradermal, subcutaneous, or intra muscular injection) of tetravalent sE spaced 4 weeks apart. When adjuvanted with Quil-A, tetravalent sE vaccination delivered via MPA resulted in earlier induction of virus-neutralizing IgG antibodies for all four serotypes when compared with all of the other vaccination routes. Using the infectious dengue virus AG129 mouse infectious dengue model, these neutralizing antibodies protected all mice from lethal dengue virus type 2 D220 challenge, with protected animals showing no signs of disease or circulating virus. If these results can be translated to humans, MPA-delivered sE represents a promising approach to dengue virus vaccination.

Immune response and reactogenicity of an unadjuvanted intradermally delivered human papillomavirus vaccine using a first generation Nanopatch™ in rhesus macaques: An exploratory, pre-clinical feasibility assessment.

The human papillomavirus (HPV) 9-valent, recombinant vaccine (Gardasil™9) helps protect young adults (males and females) against anogenital cancers and genital warts caused by certain HPV genotypes (ref. Gardasil™9 insert). This vaccine is administered intramuscularly (IM). The aim of this study was to determine preclinically whether intradermal (ID) vaccination with an unadjuvanted 9-valent recombinant HPV vaccine using a first-generation ID delivery device, the Nanopatch™, could enhance vaccine immunogenicity compared with the traditional ID route (Mantoux technique). IM injection of HPV VLPs formulated with Merck & Co., Inc., Kenilworth, NJ, USA Alum Adjuvant (MAA) were included in the rhesus study for comparison. The Nanopatch™ prototype contains a high-density array comprised of 10,000 microprojections/cm2, each 250 µm long. It was hypothesized the higher density array with shallower ID delivery may be superior to the Mantoux technique. To test this hypothesis, HPV VLPs without adjuvant were coated on the Nanopatch™, stability of the Nanopatch™ with unadjuvanted HPV VLPs were evaluated under accelerated conditions, skin delivery was verified using radiolabelled VLPs or FluoSpheres®, and the immune response and skin site reaction with the Nanopatch™ was evaluated in rhesus macaques. The immune response induced by Nanopatch™ administration, measured as HPV-specific binding antibodies, was similar to that induced using the Mantoux technique. It was also observed that a lower dose of unadjuvanted HPV VLPs delivered with the first-generation Nanopatch™ and applicator or Mantoux technique resulted in an immune response that was significantly lower compared to a higher-dose of alum adjuvanted HPV VLPs delivered IM in rhesus macaques. The study also indicated unadjuvanted HPV VLPs could be delivered with the first-generation Nanopatch™ and applicator to the skin in 15 s with a transfer efficiency of approximately 20%. This study is the first demonstration of patch administration in non-human primates with a vaccine composed of HPV VLPs.

Nanotechnology Approaches in Tackling Cardiovascular Diseases.

Cardiovascular diseases have continued to remain a leading cause of mortality and morbidity worldwide. Poor proliferation capability of adult cardiomyocytes disables the heart from regenerating new myocardium after a myocardial ischaemia event and therefore weakens the heart in the long term, which may result in heart failure and death. Delivery of cardioprotective therapeutics soon after the event can help to protect the heart from further cell death and improve cardiac function, but delivery methods and potential side effects of these therapeutics may be an issue. Advances in nanotechnology, particularly nanoparticles for drug delivery, have enabled researchers to obtain better drug targeting capability, thus increasing the therapeutic outcome. Detailed study of nanoparticles in vivo is useful as it can provide insight for future treatments. Nanogel can help to create a more favourable environment, not only for a sustained delivery of therapeutics, but also for a better navigation of the therapeutics to the targeted sites. Finally, if the damage to the myocardium is too severe for drug treatment, nanopatch can help to improve cardiac function and healing by becoming a platform for pluripotent stem cell-derived cardiomyocytes to grow for the purpose of cell-based regenerative therapy.

End-user acceptability study of the nanopatch™; a microarray patch (MAP) for child immunization in low and middle-income countries.

A promising new delivery technology, the microarray patch (MAPs) consists of an array of small solid-coated or dissolvable needles, up to one mm in length, that administers a dry formulation of a vaccine or pharmaceutical. This study is not a real-life evaluation study but determines the anticipated acceptability of the Nanopatch™, a solid microarray patch device, in Benin, Nepal and Vietnam for vaccine delivery, and identifies factors that could improve the acceptability of the technology to increase measles immunization coverage. This study combined several evaluation methods, including simulation of vaccine administration on children and in-depth interviews with key stakeholders, healthcare workers, community health volunteers, caretakers, and community representatives. A total of 314 people participated in the study. The overall rate of total acceptability of the patch for child immunization was 92.7%. General opinions were very positive, providing clinical studies confirm that MAP administration is demonstrated to be painless, safe and effective for infectious disease prevention. The study participants were asked to consider the best strategy to introduce such vaccine delivery innovation. Firstly, delivery by skilled healthcare workers at the healthcare facilities will be preferred to establish the technology. Following this, administration by selected volunteers and outreach delivery may be possible, though under the supervision of skilled healthcare workers. This study's protocol received approval from the World Health Organization (WHO) Ethical Research Committee (ERC0002813) and the national IRB in Benin, Nepal and Vietnam.

Chitosan/silk fibroin modified nanofibrous patches with mesenchymal stem cells prevent heart remodeling post-myocardial infarction in rats.

Poor functional survival of the engrafted stem cells limits the therapeutic efficacy of stem-cell-based therapy for myocardial infarction (MI). Cardiac patch-based system for cardiac repair has emerged as a potential regenerative strategy for MI. This study aimed to design a cardiac patch to improve the retention of the engrafted stem cells and provide mechanical scaffold for preventing the ventricular remodeling post-MI. The patches were fabricated with electrospinning cellulose nanofibers modified with chitosan/silk fibroin (CS/SF) multilayers via layer-by-layer (LBL) coating technology. The patches engineered with adipose tissue-derived mesenchymal stem cells (AD-MSCs) (cell nano-patch) were adhered to the epicardium of the infarcted region in rat hearts. Bioluminescence imaging (BLI) revealed higher cell viability in the cell nano-patch group compared with the intra-myocardial injection group. Echocardiography demonstrated less ventricular remodeling in cell nano-patch group, with a decrease in the left ventricular end-diastolic volume and left ventricular end-systolic volume compared with the control group. Additionally, left ventricular ejection fraction and fractional shortening were elevated after cell nano-patch treatment compared with the control group. Histopathological staining demonstrated that cardiac fibrosis and apoptosis were attenuated, while local neovascularization was promoted in the cell nano-patch group. Western blot analysis illustrated that the expression of biomarkers for myocardial fibrosis (TGF-ß1, P-smad3 and Smad3) and ventricular remodeling (BNP, ß-MHC: α-MHC ratio) were decreased in cell nano patch-treated hearts. This study suggests that CS/SF-modified nanofibrous patches promote the functional survival of engrafted AD-MSCs and restrain ventricular remodeling post-MI through attenuating myocardial fibrosis. STATEMENT OF SIGNIFICANCE: First, the nanofibrous patches fabricated from the electrospun cellulose nanofibers could mimic the natural extracellular matrix (ECM) of hearts to improve the microenvironment post-MI and provide three dimensional (3D) scaffolds for the engrafted AD-MSCs. Second, CS and SF which have exhibited excellent properties in previous tissue engineering research, such as nontoxicity, biodegradability, anti-inflammatory, strong hydrophilic nature, high cohesive strength, and intrinsic antibacterial properties further optimized the biocompatibility of the nanofibrous patches via LBL modification. Finally, the study revealed that beneficial microenvironment and biomimetic ECM improve the retention and viability of the engrafted AD-MSCs and the mechanical action of the cell nano-patches for the expanding ventricular post-MI leads to suppression of HF progression by inhibition of ventricular remodeling.

Ultrabright Room-Temperature Sub-Nanosecond Emission from Single Nitrogen-Vacancy Centers Coupled to Nanopatch Antennas.

Solid-state quantum emitters are in high demand for emerging technologies such as advanced sensing and quantum information processing. Generally, these emitters are not sufficiently bright for practical applications, and a promising solution consists in coupling them to plasmonic nanostructures. Plasmonic nanostructures support broadband modes, making it possible to speed up the fluorescence emission in room-temperature emitters by several orders of magnitude. However, one has not yet achieved such a fluorescence lifetime shortening without a substantial loss in emission efficiency, largely because of strong absorption in metals and emitter bleaching. Here, we demonstrate ultrabright single-photon emission from photostable nitrogen-vacancy (NV) centers in nanodiamonds coupled to plasmonic nanocavities made of low-loss single-crystalline silver. We observe a 70-fold difference between the average fluorescence lifetimes and a 90-fold increase in the average detected saturated intensity. The nanocavity-coupled NVs produce up to 35 million photon counts per second, several times more than the previously reported rates from room-temperature quantum emitters.

Safety, tolerability, acceptability and immunogenicity of an influenza vaccine delivered to human skin by a novel high-density microprojection array patch (Nanopatch™).

BACKGROUND: Injection using needle and syringe (N&S) is the most widely used method for vaccination, but requires trained healthcare workers. Fear of needles, risk of needle-stick injury, and the need to reconstitute lyophilised vaccines, are also drawbacks. The Nanopatch (NP) is a microarray skin patch comprised of a high-density array of microprojections dry-coated with vaccine that is being developed to address these shortcomings. Here we report a randomised, partly-blinded, placebo-controlled trial that represents the first use in humans of the NP to deliver a vaccine. METHODS: Healthy volunteers were vaccinated once with one of the following: (1) NPs coated with split inactivated influenza virus (A/California/07/2009 [H1N1], 15 µg haemagglutinin (HA) per dose), applied to the volar forearm (NP-HA/FA), n = 15; (2) NPs coated with split inactivated influenza virus (A/California/07/2009 [H1N1], 15 µg HA per dose), applied to the upper arm (NP-HA/UA), n = 15; (3) Fluvax® 2016 containing 15 µg of the same H1N1 HA antigen injected intramuscularly (IM) into the deltoid (IM-HA/D), n = 15; (4) NPs coated with excipients only, applied to the volar forearm (NP-placebo/FA), n = 5; (5) NPs coated with excipients only applied to the upper arm (NP-placebo/UA), n = 5; or (6) Saline injected IM into the deltoid (IM-placebo/D), n = 5. Antibody responses at days 0, 7, and 21 were measured by haemagglutination inhibition (HAI) and microneutralisation (MN) assays. FINDINGS: NP vaccination was safe and acceptable; all adverse events were mild or moderate. Most subjects (55%) receiving patch vaccinations (HA or placebo) preferred the NP compared with their past experience of IM injection with N&S (preferred by 24%). The antigen-vaccinated groups had statistically higher HAI titres at day 7 and 21 compared with baseline (p < 0.0001), with no statistical differences between the treatment groups (p > 0.05), although the group sizes were small. The geometric mean HAI titres at day 21 for the NP-HA/FA, NP-HA/UA and IM-HA/D groups were: 335 (189-593 95% CI), 160 (74-345 95% CI), and 221 (129-380 95% CI) respectively. A similar pattern of responses was seen with the MN assays. Application site reactions were mild or moderate, and more marked with the influenza vaccine NPs than with the placebo or IM injection. INTERPRETATION: Influenza vaccination using the NP appeared to be safe, and acceptable in this first time in humans study, and induced similar immune responses to vaccination by IM injection.

Development of Stabilizing Formulations of a Trivalent Inactivated Poliovirus Vaccine in a Dried State for Delivery in the Nanopatch™ Microprojection Array.

The worldwide switch to inactivated polio vaccines (IPVs) is a key component of the overall strategy to achieve and maintain global polio eradication. To this end, new IPV vaccine delivery systems may enhance patient convenience and compliance. In this work, we examine Nanopatch™ (a solid, polymer microprojection array) which offers potential advantages over standard needle/syringe administration including intradermal delivery and reduced antigen doses. Using trivalent IPV (tIPV) and a purpose-built evaporative dry-down system, candidate tIPV formulations were developed to stabilize tIPV during the drying process and on storage. Identifying conditions to minimize tIPV potency losses during rehydration and potency testing was a critical first step. Various classes and types of pharmaceutical excipients (∼50 total) were then evaluated to mitigate potency losses (measured through D-antigen ELISAs for IPV1, IPV2, and IPV3) during drying and storage. Various concentrations and combinations of stabilizing additives were optimized in terms of tIPV potency retention, and 2 candidate tIPV formulations containing cyclodextrin and a reducing agent (e.g., glutathione), maintained ≥80% D-antigen potency during drying and subsequent storage for 4 weeks at 4°C, and ≥60% potency for 3 weeks at room temperature with the majority of losses occurring within the first day of storage.

Safety, acceptability and tolerability of uncoated and excipient-coated high density silicon micro-projection array patches in human subjects.

Most vaccinations are performed by intramuscular injection with a needle and syringe. However, this method is not ideal due to limitations, such as the risk of needle-stick injury, the requirement for trained personnel to give injections and the need to reconstitute lyophilized vaccines. Therefore, we tested an alternative delivery technology that overcomes the problems with needle and syringe. The Nanopatch™ is an array of 10,000 silicon micro-projections per cm2 that can be dry-coated with vaccine for skin delivery. The high number and density of micro-projections means that high velocity application is required to achieve consistent skin penetration. Before clinically testing a vaccine Nanopatch, this study tests the safety, tolerability and acceptability/utility of uncoated and excipient-coated Nanopatches in healthy adults. Nanopatches were applied to skin of the upper arm and volar forearm and left in contact with the skin for two minutes before removal. The application sites were assessed for local skin response over 28 days. Acceptability interviews were also performed. No unexpected adverse events directly related to the Nanopatch application were reported. All applications of the Nanopatch resulted in an expected erythema response which faded between days 3 and 7. In some subjects, some skin discolouration was visible for several days or up to 3 weeks after application. The majority (83%) of subjects reported a preference for the Nanopatch compared to the needle and syringe and found the application process to be simple and acceptable. On a pain scale from 0 to 10, 78% of applications were scored "0" (no pain) with the average scores for less than 1. The results from this study demonstrate the feasibility of the Nanopatch to improve vaccination by showing that application of the product without vaccine to human skin is safe, tolerable and preferred to needle and syringe administration. Clinical trial registry ID: ACTRN1261500083549.

On the injectability of free-standing magnetic nanofilms.

Free-standing films with sub-micrometric thickness, composed of soft polymers and functional nanostructures are promising candidates for many potential applications in the biomedical field, such as reduced port abdominal surgery. In this work, freely suspended poly(L-lactic acid) nanofilms with controlled morphology embedding superparamagnetic iron oxide nanoparticles were fabricated by spin-coating deposition. The mechanical properties of magnetic nanofilms were investigated by Strain-Induced Elastic Buckling Instability for Mechanical Measurements (SIEBIMM) test. Our results show that these freely suspended nanocomposite nanofilms are highly flexible and deformable, with Young's moduli of few GPa. Since they can be handled in liquid with syringes, a quantitative description of the nanofilms behavior during the manipulation with clinically applicable needles has been also provided. These magnetic nanofilms, remotely controllable by external electromagnetic fields, have potential applications in minimally invasive surgery as injectable nanopatches on inner organs wall. Graphical abstract ᅟ.

The hyperelastic and failure behaviors of skin in relation to the dynamic application of microscopic penetrators in a murine model.

In-depth understanding of skin elastic and rupture behavior is fundamental to enable next-generation biomedical devices to directly access areas rich in cells and biomolecules. However, the paucity of skin mechanical characterization and lack of established fracture models limits their rational design. We present an experimental and numerical study of skin mechanics during dynamic interaction with individual and arrays of micro-penetrators. Initially, micro-indentation of individual skin strata revealed hyperelastic moduli were dramatically rate-dependent, enabling extrapolation of stiffness properties at high velocity regimes (>1ms-1). A layered finite-element model satisfactorily predicted the penetration of micro-penetrators using characteristic fracture energies (∼10pJµm-2) significantly lower than previously reported (≫100pJµm-2). Interestingly, with our standard application conditions (∼2ms-1, 35gpistonmass), ∼95% of the application kinetic energy was transferred to the backing support rather than the skin ∼5% (murine ear model). At higher velocities (∼10ms-1) strain energy accumulated in the top skin layers, initiating fracture before stress waves transmitted deformation to the backing material, increasing energy transfer efficiency to 55%. Thus, the tools developed provide guidelines to rationally engineer skin penetrators to increase depth targeting consistency and payload delivery across patients whilst minimizing penetration energy to control skin inflammation, tolerability and acceptability. STATEMENT OF SIGNIFICANCE: The mechanics of skin penetration by dynamically-applied microscopic tips is investigated using a combined experimental-computational approach. A FE model of skin is parameterized using indentation tests and a ductile-failure implementation validated against penetration assays. The simulations shed light on skin elastic and fracture properties, and elucidate the interaction with microprojection arrays for vaccine delivery allowing rational design of next-generation devices.