Microarray patches: scratching the surface of vaccine delivery

ABSTRACT Introduction Microneedles are emerging as a promising technology for vaccine delivery, with numerous advantages over traditional needle and syringe methods. Preclinical studies have demonstrated the effectiveness of MAPs in inducing robust immune responses over traditional needle and syringe methods, with extensive studies using vaccines targeted against different pathogens in various animal models. Critically, the clinical trials have demonstrated safety, immunogenicity, and patient acceptance for MAP-based vaccines against influenza, measles, rubella, and SARS-CoV-2. Areas covered This review provides a comprehensive overview of the different types of microarray patches (MAPs) and analyses of their applications in preclinical and clinical vaccine delivery settings. This review also covers additional considerations for microneedle-based vaccination, including adjuvants that are compatible with MAPs, patient safety and factors for global vaccination campaigns. Expert opinion MAP vaccine delivery can potentially be a game-changer for vaccine distribution and coverage in both high-income and low- and middle-income countries. For MAPs to reach this full potential, many critical hurdles must be overcome, such as large-scale production, regulatory compliance, and adoption by global health authorities. However, given the considerable strides made in recent years by MAP developers, it may be possible to see the first MAP-based vaccines in use within the next 5 years.


Introduction
The hypodermic needle and syringe have remained largely unchanged for the last 170 years.Currently, conventional needle and syringe (N&S) delivery methods such as intramuscular (IM) and subcutaneous (SC) injections are most effective and widely used for vaccination.While it is extremely effective, it is not without drawbacks surrounding vaccine supplies, transportation and storage, vaccine administration, waste disposal and needle phobia [1,2].To overcome these drawbacks, novel vaccination strategies have been developed, with needle-free technologies experiencing a significant increase in the recent years.Since then, microarray patches (MAPs) have emerged as a promising alternative vaccine delivery platform to the N&S.

Skin vaccination
The skin provides a dynamic and active barrier to the outside world.To provide this barrier function the skin has evolved complex protective immune and inflammatory responses to provocations including toxins, viral or bacterial pathogens and mechanical wounding.To protect the body from these insults the skin layers contain a relatively high density of antigen presenting cells that when targeted can procedure enhanced immune responses.Traditional needle-based vaccination results in vaccine being deposited into muscle or subcutaneous space.While these vaccination methods works, they bypass the layers of the skin which are rich in antigen presenting cells.As a result many technologies have been developed and are underdevelopment that seek to harness the potential of skin vaccination.The traditional intradermal (ID) injection, the Mantoux technique, can be technically challenging to perform and painful for the patient, resulting in the limited adoption of this method for vaccination.To overcome these challenges, multiple devices have been developed over the years as alternative ID delivery systems.Companies such as Becton Dickson and Nanopass Inc have designed microinjection devices that simplifies and increases the success rates of ID injection.Table 1 summarizes the advantages and disadvantages for the various ID delivery devices.

Needle-free vaccine delivery devices
Over the past 10 years, MAPs have been one of the most rapidly advancing needle-free technologies for vaccine delivery with several unique advantages, as such are the major focus of this review.MAPs, an alternative vaccine/drug delivery device, are designed to house an array of microprojections, measuring 100-1500 µm in length [50][51][52].Different types of MAPs have been developed (solid, coated, dissolvable, hollow, and hydrogel-forming) for various applications (vaccine, biologics, and small-molecule drug delivery) [53][54][55][56][57][58][59][60][61][62].Firstly, MAPs are designed to breach the tough outer layer of the skin, the stratum corneum, to deliver their respective payloads into the epidermal and dermal layers of the skin.Depending on the array density, MAPs are applied either by pressing onto the skin or with applicators that overcome the viscoelastic and mechanical properties of the skin.As vaccines are dry-coated onto or formulated in the microprojections often enhanced stability profiles of the vaccine payload can be achieved.As a result of the features of the MAP vaccine delivery technologies provide multiple programmatic benefits.Firstly, MAP vaccination eliminates the risk of needle-stick injuries, needle-phobias, and cross-contamination associated with the traditional N&S methods.It also removes the need of additional preparation steps where trained personnel are needed to perform both the initial reconstitution of the lyophilized vaccine and vaccination [63].Another advantage is improved thermal stability, which will potentially reduce the need for cold chain storage and lower logistical challenges associated with transport and storage, thus accelerating the introduction and reach of vaccines in resource limited settings [64][65][66][67][68].Other additional benefits of MAPs also incudes offering an alternative method of delivery to booster vaccines which avoids additional injections, improving vaccine uptake [69,70].
As a vaccine delivery platform, MAPs have great potential having been demonstrated to be compatible with a variety of vaccine modalities (DNA, live attenuated virus, mRNA, conjugate, subunit, inactivated virus, chimeric virus) and adjuvant types and have been evaluated both preclinically and clinically (Table 2).Combining these properties make MAPs the ideal vaccine delivery system for many pathogens in both the developing and developed world [51, [145][146][147].MAPs can be categorized into five main types: solid, coated, dissolvable,
The advantages and limitations of the different MAP design are summarized below (Table 3) and further expanded in the section after.

Solid MAPs
Solid MAPs are one of the initial designs manufactured by conventional microfabrication methods.Most commonly, they are used as a pre-treatment to penetrate the skin, forming micropores that allows enhanced uptake following the topical application of desired biological payload (typically drugs; Figure 1) [55,61,154].Other variations includes generating micro-abrasions [157][158][159][160] or rolling the solid MAP over the skin repeatedly to abrade the stratum corneum [159].Due to the mode of application, the materials selected must be able to tolerate the force applied at time of penetration without causing breakage of the microprojections.Solid MAPs were originally mainly fabricated from silicon, but silicon is prone to fracturing in the skin upon application.As such, other materials for manufacturing solid MAPs have been explored such as ceramics, various metals (titanium, nickel, and stainless steel), and different polymers [55,61,154,161].However, there are several drawbacks to this design, such as the reliability and reproducibility of vaccine delivery, the lack of precise dosing, as well as requirement for reformulation for many different types of vaccines [155].As vaccine delivery rate is often inconsistent as this method relies on the payload diffusing into the skin, the micropunctures generated by the microneedles as well as the concentration of the payload.
Although there have been no reports to date, infection of the micropunctures is a theoretical possibility due to the time for the reestablishment of the skins barrier function [162].These limitations of the solid MAPs have led to the development of other MAP designs [51,146].
As one of the first MAP concepts, the solid MAP has been trialed preclinically with many vaccine platforms, including subunit, DNA, and live attenuated vaccines.This approach has received mixed reports of success in preclinical models.Ding et al (2009) found pre-treatment with solid MAPs failed to elicit potent immune responses to subunit influenza vaccines [71].In contrast, other studies with hepatitis B and JEV vaccine observed improved antibody responses when delivered by to the skin prepared with MAP pretreatment as compared to groups without.The addition of adjuvants to the vaccine formulations, can further enhance antibody levels when applied to MAP treated skin [72].Likewise, non-human primates vaccinated with JEV by the solid MAP were observed to have a variable but elevated level of neutralizing antibodies when compared to the SC injection method [73].Although these results are promising, alternative MAP designs should be considered for vaccination studies due to the limitations and inconsistency of pre-treating the skin and dosing when using solid MAPs [145].

Coated MAPs
Coated MAPs are generally designed as a combination of solid MAPs with a liquid solution and coating methods.As the skin contains a relatively high concentration of antigen-presenting cells (APC), with skin-targeted immunizations using coated MAPs, minimal vaccine is needed to generate the required immune response, making coated MAPs a popular choice for vaccine delivery.Vaccines are dry coated to the tips of the microprojections at ambient temperatures before delivery into the skin (Figure 2).In order to maintain consistency and reproducibility of vaccine delivery, it is important that the microneedles have an uniform coating and consistently penetrate and engage the skin [192].As a result, several methods to increase coating efficiency have been explored.These methods range from dip coating [85,193], gas-jet coating [79,91,92,101,102,194,195], spray coating [196] to ink-jet coating [147].Another factor that contributes to coating uniformity are the excipients selected.Excipients selected for the coating solution also plays apart in vaccine stability.Most commonly, formulations include a viscous excipient (e.g.methylcellulose, carboxymethyl cellulose, polyethene glycol), stabilizers (e.g.trehalose, sucrose, cyclodextrin, protein carriers, and various amino acids) and adjuvants (e.g.Quil-A) Hollow/ porous MAPs • Serves as a reservoir to allow for passive diffusion • Allows greater volume of liquid formulations to be delivered into the skin • Potential of vaccine leakage, clogging of channel openings, and the collapse of microprojections due to their weak structure [60,141,148,[181][182][183][184][185][186][187][188] Hydrogelforming MAPs • Uses a reservoir to store the drug/vaccine, enabling a higher volume/dosage to be administered • Newly emerging technology that requires more research to understand the delivery efficacy, mechanism, safety, and other adverse effect • Have not been thoroughly investigated in pre-clinically in animal models for skin-targeted immunization against viral pathogens [54,174,[189][190][191] Figure 1.Schematic representation of the 'poke and patch' technique by the solid MAP.solid MAPs are commonly used to penetrate the skin before topical application of desired drug or vaccine, allowing them to diffuse through the holes created by the microneedles.Created with BioRender.com.[79,81,91,92,101,102,146,154,197].The coating formulation requires optimization for different vaccine modalities to ensure retention of antigen potency and stability through phase changes (from the drying process to reconstitution in the skin) [64][65][66][67][68].These factors, in turn, affect the delivery efficiency and delivered dose of the vaccine when the coated MAPs are applied to the skin.Therefore, selection of the appropriate excipients and coating method is important for the consistency and reproducibility of vaccine delivery when using coated MAPs [51,146].Coated MAPs are one of the more popular choices for vaccine delivery and have been evaluated with multiple vaccine platforms such as live attenuated, viral vector, inactivated virus, subunit, DNA, VLPs and chimeric viruses.Although most studies have been conducted with mice, research also continues with rats, guinea pigs, porcine and NHPs [74][75][76][77][79][80][81][82][83][84][85][91][92][93][94][95][97][98][99][100][101][102][103][104][105][106][163][164][165][166][167][168][169][170].Influenza vaccines (inactivated, DNA, subunit, VLP) delivered by coated MAPs were conducted by several groups including our own.All studies reported significantly higher induction of antigen-specific antibody responses and fully protected against lethal challenge when compared to traditional N&S method [74][75][76][77][82][83][84]106,[167][168][169][170]198].In particular, an influenza study by Fernando et al. [79] demonstrated 100-fold dose sparing when vaccinating with the Nanopatch (applied by a custom applicator) while yielding equivalent IgG responses to those of IM vaccination.Coated MAPs have also been evaluated with other viral vaccine candidates such as SARS-CoV-2, dengue, Ebola, poliovirus, measles, hepatitis C, HPV, and RSV [91-95,97-104].Similar results were observed for all studies with significantly higher antibody response observed when vaccinated using MAPs over the traditional N&S method.The inactivated polio vaccine (IPV), when delivered by the Nanopatch, demonstrated 40-fold dose sparing while eliciting protective levels of neutralizing antibodies from a single dose [101].Although not all MAP designs demonstrate dose-reduction effects, these results demonstrate promising potential of using coated MAPs for vaccination with higher induction of antibody responses.

Researchers at Stanford University and University of North
Carolina have also developed a 3D-printed MAP that has a greater surface area to allow for enhanced vaccine coating and has also designed a method that allows coating of multiple cargo in specific sections of the microarray.The combination of these two technologies could prove to be a potential improvement for MAP vaccination platforms [199].
Three phase I clinical trials for influenza using coated MAPs have been successfully conducted by Vaxxas Pty Ltd.The initial study evaluated the safety and tolerability of a silicon MAP in a randomized, partly blinded, placebo control trial (ACTRN12616000880448) [80].Volunteers were vaccinated with a monovalent inactivated influenza vaccine either: via a coated MAP, applied to the forearm or the upper arm using a spring-loaded applicator; or through IM injections into the deltoid.Volunteers receiving the MAP vaccinations preferred the MAP compared to IM injections based on past experiences [80].Overall, vaccination using MAPs was well tolerated with only mild or moderate AEs reported.Most importantly, antibody responses generated from the MAP vaccinated groups were similar to those vaccinated by IM injections, suggesting that MAP vaccinations were able to induce antibody levels equivalent to the traditional injection methods [80].
Recently, a clinical trial was conducted as a two-part randomized, partially double blind, placebo-control study using MAPs coated with a monovalent influenza vaccine (ACTRN12618000112268) [78,81].This study utilized a revised high-density MAP (HD-MAP) design delivered by a springloaded applicator, which measures 1 cm 2 with 3,136 microprojections per patch each 250 µm in length with a sharp point of <25 µm.The spring-loaded applicator works by pushing a button to release the spring which propels the HD-MAP into the skin [200].The applicator is designed for single use only and the device (which includes the HD-MAP) is discarded once triggered [201].Safety, tolerability, and cellular responses of the vaccine were assessed and evaluated in this clinical trial [78,81].Overall, HD-MAP delivery of the vaccine was well tolerated with only mild and moderate AEs reported.This study was the first to demonstrate the dose sparing potential of the HD-MAP in humans, with 2.5 µg of an influenza vaccine eliciting similar HAI titers as a 15 µg dose of influenza vaccine.Furthermore, this study also demonstrated the potential elimination of cold chain by thermostability of the vaccine at 40°C for 1 year [81].In response to the COVID-19 pandemic, Vaxxas conducted a Phase I clinical trial of a COVID-19 vaccine, Hexapro, delivered by the HD-MAP (ACTRN12622000597796) [86,87].This study was designed to assess the safety, tolerability and immunogenicity of the HD-MAP delivered COVID-19 vaccine in healthy adults aged 18-50 years old.This trial also consists of a dose ranging regime and will also analyze T cell and mucosal antibody responses.Interim results announced by the biotechnology company reported an 8-fold increase in antibody titers from samples taken 28 days post immunization.Funding for a Phase II clinical trial has also been obtained and will progress in 2024 [202,203].

Dissolvable MAPs
Dissolvable MAPs, in contrast to solid or coated MAPs, have been developed to dissolve in the skin completely and are usually made using natural, biocompatible materials.The desired drugs or vaccines are integrated together with the microprojections by casting them in a mold filled with a dissolvable solvents and then allowed to solidify [55,57,204].Depending on the release mechanics required for the drug/vaccine, the solvents must be carefully selected for.While the solvents selected must be biocompatible, biodegradable and mechanically strong enough to withstand the penetration of the skin, it also has to unalter the safety, potency and efficacy of the drug or vaccine.As such, watersoluble solvents, such as sugars are usually incorporated into vaccines as they rapidly dissolve into the skin upon application, while other solvents such as polymers are usually used with drugs as they slowly dissolve into the skin, controlling the release rate of the drug (Figure 3) [204].Other materials such as silk, polyvinyl acetate, hyaluronic acid and chitosan have also been reported in the preparation of vaccine containing dissolving microneedles [115,[175][176][177][178][179].Dissolvable MAPs also have a larger capacity to hold greater drug/vaccine payloads by adjusting the density of needles, needle size or patch size while ensuring a precise and consistent release of dosage.Despite these advantages, several factors remain to be considered without altering the original intentions of these dissolvable MAPs: vaccine compatibility and stability with solvent materials, effects of biodegradable materials on the mechanical strength of the MAPs.Furthermore, careful consideration must also be given regarding how they engage with the skin at the point of insertion and the effect of the solvent and biodegradable materials on the immune system [173,174].
The dissolvable MAPs have also been tested widely in preclinical trials for vaccinations against viruses such as influenza, Hepatitis B and C, HIV, measles, rubella, rabies, poliovirus, dengue, Ebola, Zika, MERS-CoV and SARS-CoV-2 [96, [106][107][108][109][110][111][112][113][114][117][118][119][120][121][122][123][124][125][126][127][129][130][131][132][133][134][135][136][137]180].These studies also cover different antigen platforms such as subunit, DNA, viral vectors, liveattenuated and inactivated viruses.An interesting feature of the dissolvable MAP is the time of dissolution and how it affects the type of immune response.Multiple studies have tested different fabrication methods to alter the dissolving time of the MAP to target the type of immune response required.Dissolvable MAPs by Vaxess Technologies have demonstrated thermostability of subunit vaccines when formulated with silk protein on a water-soluble poly(acrylic acid) base [205].These silk fabricated microneedles were able to regulate the kinetics of vaccine delivery skin to mimic patterns of natural infections [205].BALB/c mice vaccinated with a HIV envelope trimer vaccine delivered by the silk fibroin dissolvable MAP were observed to have a 1300-fold increase in antibody response as compared to intradermal injections [128].Similar to the coated MAPs, compact applicators have also been designed and tested for dissolving MAPs.The latch applicator designed by Kang et al is assembled using three plastic parts and is operated using thumb force by pressing on the trigger button [206].The overall length and diameter were kept within 20 mm and 40 mm respectively, making it easy for use and economical production [206].To further optimize skin vaccination using the dissolving MAPs, factors such as the long-term stability of vaccine formulated with biodegradable materials and reliability and reproducibility of vaccine delivery are currently being investigated [146].Ultimately, these dissolvable MAPs are an attractive vaccine delivery system that offers several advantages over other MAP technologies such as the payload capacity, ability to cast great high concentrations of vaccine stabilizing excipients, absence of biohazardous waste and high vaccine-dosage capacity.The biodegradability of the dissolvable MAPs makes them an attractive alternative for further development in the field of vaccine delivery systems.While dissolvable MAPs have been extensively studied in preclinical trials, relatively few human clinical studies have been conducted with vaccines [115,116,138,140].
The first clinical study evaluated the MicroHyala dissolvable hyaluronic acid MAPs with trivalent seasonal influenza hemagglutinin antigen.Participants in this trial received the vaccination via a handheld applicator and the MAP was left to dissolve for 6 h [116].No serious AEs were reported, and antigen-specific immune responses elicited in the MAP vaccinated groups were equal to, or more effective than the SC vaccinated groups.Surprisingly, no placebo control group was included, and more than half of the data had to be excluded due to unreproducible MAP application.Long-term stability at different temperatures (4, 25, 40°C) were also evaluated.Antigen content decreases after 6 months of storage at temperatures greater than 4°C suggesting that the vaccine formulation has to be further optimized [116].
The second dissolving MAP clinical trial (NCT02438423) conducted was a randomized, partly blinded, placebocontrolled study with Fluvirin (Novartis) [115].Participants were administered a single dose of the vaccine via the dissolvable MAP, either by trained healthcare personnel or via self-administration.IM delivery of the vaccine was included as a control.Immune responses elicited by MAP administration, were observed to be similar to the IM vaccinated group, with no significant differences in vaccine delivery efficiency observed between self-administration or application by a trained healthcare worker [115].Long-term stability was observed for 12 months at all temperatures tested (5, 25, 40°C).Delivery efficiencies of the MAP were also measured and revealed no significant differences between the two groups (healthcare personnel vs self-administration), supporting the self-administration potential of the MAP [115].
The most recent Phase I/II clinical trial was conducted by Micron Biomedical (NCT 04394689) in pediatric population, studying the delivery of measles and rubella vaccine delivered via a dissolvable MAP [138,139].This is also the first clinical study of any MAP-based technology in children.This clinical trial evaluated the safety, immunogenicity and acceptability of the MAP-based measles and rubella vaccine in adults, children and infant as young as 9 months old [138,139].93-100% seroprotection was observed in day 42 samples obtained from the study and the vaccine was safe and well tolerated without severe AEs observed.A survey from the clinical trial also revealed that majority of the parents of the children enrolled preferred the MAP vaccination over the traditional needle and syringe [138].These promising results from the clinical trials support further development of dissolving MAPs for other infectious pathogens as well as self-administration in the context of a pandemic or mass vaccination.

Hollow and porous MAPs
Hollow MAPs have been designed with channels that allows liquid to flow through the punctures created in the skin when administered (Figure 4).Like dissolvable MAPs, hollow MAPs are designed to directly deliver vaccines into the skin with a facilitated force-driven fluid flow [55,61,154,181].There are several methods in which the drugs/vaccines can be delivered into the skin using a hollow microneedle.Most commonly, drugs and vaccines are held in a reservoir before they passively delivered through the bore of the microneedle [183].Other methods include using an applicator (e.g., syringe, pump or pressurized gas) to apply pressure and actively deliver the vaccine through the bore of the microneedle into the skin [141,[184][185][186]. Microfluidic chips or micropumps can also be incorporated with the microneedle for a controlled release of the vaccine [207,208].As hollow MAPs have to be fabricated to ensure for consistent flow, extra attention on the mechanical strength is required when designing them.While increasing the bore of the microneedle may increase the flow of the liquid, it often compromises the mechanical strength of the MAP as well as the sharpness of the microneedle.Hence, researchers have designed hollow MAPs with the bore toward the side instead of at the tip of the microneedle [187,188].Hollow MAPs also allow a greater volume of liquid formulations to be delivered into the skin, but investigation must still be carried out to determine the optimal formulation for each drug/vaccine to maximize delivery.While numerous research on hollow MAP design and fabrication has been conducted, there are still multiple factors to overcome such as the potential of vaccine leakage, clogging of channel openings, and the collapse of microprojections due to their weak structure [60,148,181,182].To date, most of the research surrounding hollow MAPs have been directed toward the design and characterization aspects of fabrication and less on delivery efficacy into the skin [51,209].Therefore, fewer studies have been conducted using hollow MAPs as a vaccine delivery system as compared to solid and coated MAPs.Hollow MAPs have been evaluated for several viral vaccines including influenza, mumps, varicella, polio, and HPV.These studies have been conducted in animal models of mice, rat and guinea pigs [141][142][143]148,210]. Hollow MAPs delivering IPV using an applicator that allowed for controlled fractional dosing were observed to have ten times higher IgG titers than those of intradermal and IM vaccination methods [142].Another study delivering DNA-encoding ovalbumin using hollow MAPs observed the induction of higher cellular and humoral immune response when compared to SC injection [210].Results obtained from these pre-clinical studies have once again proved that antibody responses elicited when using MAPs as a delivery system can be attained using a fraction of a dose as compared to traditional N&S methods.
In a recent Phase I clinical trial study conducted by MyLife Technologies, vaccination using a nanoporous microneedle array with the Moderna mRNA-1273 vaccine have failed to induce an antibody response in the participants [144].While the microneedle platform has been found to be safe, however, no antibody or cellular immune response was produced presumably due inadequate vaccine delivery into the skin [144].There are several limitations to the study with regard to the microneedle length and depth penetration as well as the wear times of the microneedle array for a human skin.Vaccine coating procedure were also not optimized resulting in only 10% of the vaccine being coated on the needles while the remaining 90% were on the baseplate [144].Whilst this study has successfully shown that the delivery platform is safe for human use, much optimization is to be done before it can be in the markets.

Hydrogel-forming MAPs
Lastly, hydrogel-forming MAPs are emerging as a field of study for the delivery and extraction of biomolecules.This MAP is fabricated with polymers crosslinked with gelatin which rapidly swell upon insertion into the skin and eventually releases the drug into the surrounding microenvironment (Figure 5).This is due to the hydrophilic nature of the hydrogels, readily taking up water upon insertion into the skin [191].These hydrogel-forming MAPs makes use of a reservoir to store the drug, which enables a higher volume to be administered [54,174,189,190].However, the MAP must be left in the skin to deliver a larger dosage for an extended period.Unlike the dissolvable microneedles, the microprojections on the hydrogel-forming MAPs does not remain in the skin, leaving no polymer residues after application [174,189,190].Another advantage of this MAP is that it can be fabricated to many different geometries unlike the conventional solid, hollow or dissolvable MAPs.These MAPs can also be sterilized prior to insertion and are detachable more easily resulting in minimal damage to the skin and the MAP [190].Currently, the hydrogel forming MAP has been mostly used for extracting interstitial fluids from the skin [211][212][213][214], delivery of small molecular drugs [214][215][216][217] and antibodies [218], skin cancer treatment [219] and wound healing [220].Although this form of MAP has been increasingly researched on in the therapeutics field, more investigations are required to understand the delivery efficacy, mechanism, safety, and other adverse effects before utilizing them for skin vaccination.As hydrogel-forming MAPs are a relatively new concept, they have not been thoroughly investigated in pre-clinically in animal models for skin-targeted immunization against viral pathogens.

MAP compatible adjuvants
To further enhance the immune response of MAP-based vaccination, adjuvants have also been explored in preclinical studies.Both chemical and biological adjuvants have been widely evaluated with MAPs in preclinical models.From a practical standpoint, adjuvants must go through careful selection depending on the type of MAPs.For example, aluminum-based and oil emulsion-based adjuvants have typically been regarded as not suitable for coated MAPs due to their inability to coat evenly on the surface of the microprojections.Furthermore, care is needed in adjuvant selection and dose delivered with special consideration taken for the local skin environment.As MAPs deliver vaccine and adjuvant into the top layers of the skin, any local reactions will be visible.These reactions are likely similar to those that occur then adjuvants are delivered deep into the tissue via the N&S.However, the visibility of any immune reaction in the skin will likely introduce issues relating to reactogenicity and patient acceptance.Regardless, adjuvants such as imiquimod, poly(I:C), QS-21, Quil-A (QA), c-di-GMP, Flt3L and Matrix-M have been evaluated for coated MAPs; cationic liposomes, CpG-ODN, MPLA, QS-21, poly(I:C) for dissolvable MAPs; and GLA-AF for hollow MAPs [92,99,126,131,134,164,[221][222][223][224][225].Table 4 summarizes the adjuvants used for each MAP type, animal models trialed in and if the adjuvants are approved for human use.
Apart from relying on adjuvants to enhance the immune response of MAP-based vaccination, preclinical studies investigating the disruption of skin barrier function post MAPapplication and how it affects the immune response induced have been performed.These demonstrated that localized cell death in the skin caused by MAP application increases immunogenicity of the vaccine, thus acting as a 'physical immune enhancer' [78].These results suggests that with optimized MAP design and application condition, it is possible to generate mechanical stress, inducing controlled cell death to create a proinflammatory environment to generate enhanced immune responses [78,228].These promising results from preclinical studies supports the future of MAPs in skin-targeted vaccination.however, extensive work is still required to improve and standardize MAP platforms to allow for global immunization and effective response to future pandemics.The lack of standard protocols, animal models and efficacy measures result in a more challenges for preclinical studies to be conducted for different vaccine platforms [146].Much research is required in understanding the mechanism of MAPs and how it induces the skin immune response to successfully transition from traditional immunization routes to MAP-based immunization strategies.

MAP applicators and wear times
The highly effective barrier properties of the skin's stratum corneum results in challenges in delivering drugs or vaccines into the immune cell-rich layers of the epidermic ad upper dermis [229].To overcome these difficulties while ensuring a reproducible and repeatable delivery of the drugs and vaccines, MAPs have to penetrate the skin in a controlled manner.This has thus prompted the design and development of applicators to ensure that deliveries of the drugs and vaccines remain consistent.There are several factors to consider when designing these applicators which includes: 1) Uniformed and repeatable application that is not affected by variation of skin types; 2) Controlled depth penetration of the microprojections into the skin; 3) Sufficient mechanical strength to breach the initial layer of stratum corneum; 4) Ease of us and pain free; and 5) Retain physical stability and integrity of the applicator during storage and application [229][230][231].Applicator design ranges from manual (using pressure from the thumb) or mechanical (spring-loaded or electromagnets) applicators system either for single or multiple use [65,71,72,80,81,[86][87][88][89][90][91]101,102,116,138,139,165,176,201,232].Apart from applicators, different type of MAPs have different wear times ranging from instantaneous to 24 h.Coated and hollow MAPs have relatively short wear times from removing them instantly postapplication to about 2 min [65,80,81,[86][87][88][89][90][91]101,102,[141][142][143]148,165,201,232].While dissolving and hydrogel-forming MAPs have longer wear times of 20 min to 24 h to allow for the drugs or vaccines to fully dissolve into the skin over time [116,138,139,176,190,216,218].Wear time varies between different drug and vaccine types and are recommended to be refined accordingly to obtain optimal delivery [90].Companies like Vaxxas, Microhyala, Micron Biomedical and QuadMedicine have applicators designed to deliver their respective patches efficiently.Table 5 summarizes the type of MAPs and the different applicator designed used with them as well as their corresponding wear times.

Additional considerations for MAP-based immunizations
The concept of MAPs was first introduced in 1976 and since then, extensive research has been performed to improve Yes.Used in clinical trials for influenza H5N1 vaccine [221] and understand the mechanisms involved in the delivery of the payload into the skin [52].However, multiple factors must be established before the commercialization of MAPs is possible.The safety profile, thermostability, consistent and reproducible skin application and delivery of vaccine, disposal, cost, regulatory approval are some of the important factors that requires a full evaluation for each individual MAP vaccine candidate.The major challenge facing the MAP developers is how to produce vaccine MAPs at scale.There are many challenges to be overcome, associated with the development of vaccine MAP lines to fill and finish the final product.While of the challenges are common, unique challenges remain for each including mass production of sterile product, which cannot be terminally sterilized.For dissolving arrays, challenges remain around how to cast and set MAPs at scale while maintaining sterility.Likewise, coated MAPs face their own challenges associated with how to dispense vaccine to the tips of the projections so not to waste vaccine on the base of the patch.For both the dissolving and coated MAPs elegant engineering solutions are required to overcome the challenges associated with requirements to mass produce MAPs at a speed to be competitive with the N/S.Lastly, the criteria for regulatory approval of MAPs are not clearly defined and consultations will be required for each separate platform of MAPs.

Vaccine recipient safety
Vaccine recipient safety is by far the most important factor when considering widespread utilization of MAPs as a vaccine delivery system.Studies have shown that skin barrier recovery was observed 48 hours post MAP application with erythema clearing over the course of a 7-day period in healthy participants.However, more research is required to elucidate the relationship between the level of disruption to skin barrier and recovery and its correlation to skin infection especially between different skin types especially for the elderly and immunocompromised patients [146,235,236].Sterilization of MAPs, according to the Food and Drug Administration, must be considered in the early stages of the development process.Factors such as the target tissue, depth of penetration, MAP wear time, and target population determine whether the MAP requires sterilization [52].Aseptic manufacturing is complex and expensive, and some vaccine components are unsuitable to undergo current sterilizing methods.For such situations, fabrication and manufacturing of MAPs as well as vaccine coating will have to be done under aseptic conditions, increasing the overall cost considerably.

Global immunization campaigns
The emergence of the COVID-19 pandemic created an unprecedented need for accelerated vaccine development to combat SARS-CoV-2.Leading vaccine candidates were developed, tested, and granted approval for emergency use within 11 months.However, emergency use vaccines approved were administered through traditional needle and syringe methods and with several vaccines requiring ultra-low temperature storage with only limited stability in low temperatures (2 to 8°C) [237].This resulted in challenges in global distribution and transportation especially to low-income countries and requiring trained healthcare  [190,216,218] -Applied to the skin using manual application pressure for 30 s −24 h workers to administer the vaccine [238,239].These factors accompanied with vaccine noncompliance due to needle phobia, pain and inconvenience are obstacles for successful mass vaccination.With dose-sparing, vaccine stability with reduction in dependence on cold chain as well as its compatibility with a variety of antigen platforms, delivery of vaccines using MAPs could potentially eliminate the factors of concern found with N&S delivered vaccines [64][65][66][67][68]. MAP vaccination could also possibly increase vaccine accessibility especially in resource limited settings lacking appropriate cold chain infrastructure [64][65][66][67][68].The elimination of hazardous waste and trained personnel will also greatly facilitate routine outreach services and mass vaccination campaigns especially in areas where systems fail to routinely reach everyone [240].
To address these drawbacks and help accelerate MAP vaccine developments, a collaboration between the Gavi Secretariat, WHO, Bill & Melinda Gates Foundation, United Nations Children's Fund and PATH was formed to become the Vaccine Innovation Prioritization Strategy (VIPS) Alliance [241].VIPS concluded their meeting in May 2020 with a decision to prioritize 3 innovative approaches for advance development to help address immunization barriers and to increase immunization coverage [241].Of the three innovations, two are developing microarray patches and thermostable qualified vaccines.In their five-year action plan for MAPs, they have set to identify the requirements needed to accelerate MAP-based vaccine development for low-and middle-income countries as well as to advocate for MAP-based vaccine so as to attract the interest of global health partners and other industry funders [241].The COVID-19 pandemic has once again emphasized the need to implement innovations that will help overcome immunization barriers and ensure for vaccine equity across the globe.
There is a potential of self-administration of MAP-based vaccines in a mass pandemic scenario, however, for successful immunization, it is critical that the MAP is properly applied into the skin [242].Reliability and consistency in vaccine delivery is important for the induction of immune response in vaccinated individuals.MAPs, subjected to their design and type, can be self-administered or applied by trained medical personals.Vaxxas performed a study to observe the delivery characteristics of their HD-MAP when administered either by trained personnel or through selfapplication [232].Similar delivery characteristics of the HD-MAP was observed from both groups when the HD-MAP is applied to the upper deltoid of the arm.While the sample size for this study was small (n = 20), this study revealed the potential of self-administrating HD-MAP delivered vaccines to reduce the manpower strain on the healthcare workforce sector during a pandemic [232].Without the need of medical expertise, it is expected that the overall cost for MAP vaccine will be reduced.Apart from that, the elimination of cold chain for transport and storage and lack of sharps waste would also contribute to the cost reduction of MAP vaccines [115,243,244].Ultimately, more preclinical and clinical research is still needed to close the knowledge gaps for MAPs to be used in global immunization campaigns.

Conclusion
In conclusion, this review highlights the significant potential of MAPs for vaccine delivery and underscores their versatility, safety, and potential for large-scale implementation.The development and refinement of MAP-based vaccination approaches have the potential to transform the field of immunization, overcoming several barriers associated with traditional N&S injections.Future research and collaborative efforts should focus on addressing regulatory considerations, optimizing vaccine stability, and ensuring equitable access to MAP-based vaccines.With continued advancements, MAPs have the potential to revolutionize the way vaccines are administered, contributing to global health initiatives, and improving immunization outcomes.

Expert opinion
MAPs as vaccine delivery systems have shown precise and reproducible vaccine administration to the skin.Along with other advantages such as cost-effective fabrication, reduction in coldchain dependence and ease in application and disposal, suggest MAPs to be a more effective vaccine delivery platform than traditional needle-and-syringe.However, despite the large body of evidence acquired over nearly 2 decades or pre-clinical research, only a handful of microneedle system have been accelerated to clinical trials.Immunologically, the MAP as a vaccine delivery device has been repeatedly shown to be superior to traditional N&S delivery.However, there are other key factors that need to be addressed before MAPs can progress down the commercialization path.
As the most accessible organ in the body, the skin has been extensively studied.Skin variability with age is one issue that may impact consistency of MAP vaccination.Studies regarding the nature of the skin physiology and structure report varying evidence of whether age plays a part in changing the morphology and to what extent.While studies have generally reported similar dermal thickness across age and gender, there are insufficient information on pediatric and elderly skin characterization.Vaccines delivered by MAPs to these skin types may have to be optimized accordingly especially if they are found to be different.Consistent payload delivery, device penetration and shorter delivery time may also have to be addressed for MAPs to become broadly accepted.As the vaccines or microneedles are not visible upon delivery, robust data sets will need to be developed indicating that when administered, skin penetration is achieved to the required depth to ensure the correct delivery of the desired dose of vaccines.With self-administration being a potential factor when using the MAPs, companies will have to ensure that device performance is consistent whether applied by a trained healthworker, or self-applied.However, MAP-based vaccines will realistically have to be rolled out in stages.Starting with administration of the MAP-based vaccines with trained medical personnel, then to trained community volunteers before moving on to self-application.
A challenge facing all MAP developers is the skin reactogenicity associated with MAP application.When MAPs are applied to the skin, localized erythema occurs at the application site.It is possible that an identifying transient eryhtema could impact uptake of the technology.However, it also creates an opportunity for patient and user education, at a very simple level that this skin-response provides visual confirmation indicating the immune system responding to the vaccine.While MAP-based vaccination is promising, patient advocacy will be necessary as part of commercialization.
Currently, the manufacturing process are sufficient to produce MAPs for small-scale Phase I and II clinical trials.Large-scale Phase III clinical trials and commercialization will require manufacturing approaches that can allow for mass production of patches with appropriate industry investments and infrastructure.While regulatory knowledge has been built around the N&S vaccine delivery, new guidelines will need to be established by the regulatory authorities so that MAP developers can be certain of achieving quality standards and manufacturing requirements for registration of MAPs.Apart from regulatory considerations, product costs are also key to progressing MAP commercialization.While MAPs have been promoted as cost competitive with N&S methods, however, MAP manufacturing still requires significant funding from investors to build facilities capable for mass production.This consideration will also need to be extended to the feasibility of manufacturing MAPs in low-income countries to ensure equitable distribution.
Vaccination by MAPs could significantly improve immunization coverage and the continuous progress of MAPbased vaccines will be dependent on the investment from vaccine manufacturers and as well as funding by global health partners.There are still several hurdles to overcome, but with the recent advances by Vaxxas, Vaxess and Micron Biomedical in clinical trials and the manufacturing space and clinical trials, we may be able to see the first MAPbased vaccine in the market for use in the next 5 years.

Figure 2 .
Figure 2. Schematic representation of the coated MAP.vaccines are dry coated onto the tips of the microprojections before application to the skin.Created with BioRender.com.

Figure 3 .
Figure 3. Schematic representation of the dissolvable MAP. the desired drugs or vaccines are integrated together with the microprojections which dissolves into the skin over time after application.Created with BioRender.com.

Figure 4 .
Figure 4. Schematic representation of the hollow MAP.vaccines/drugs are loaded into channels of the microneedle that flows through and into the penetrated pores of the skin when administered.Created with BioRender.com.

Figure 5 .
Figure 5. Schematic representation of the hydrogel-forming MAP.hydrogel-forming MAPs are fabricated with polymers crosslinked with gelatin which rapidly swells upon penetration into the skin and eventually releases the drug/vaccines into the microenvironment.Created with BioRender.com.

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Summary of clinical trials (completed or in-progress) for MAP-based vaccine delivery• Additional considerations that are required for MAP-based vaccine delivery

Table 1 .
Advantages and disadvantages of ID delivery devices.

Table 2 .
Types of MAPs, antigen used and their development stages.

Table 3 .
Advantages and limitations of the different MAP designs.

Table 5 .
MAP applicators and wear time.