Rapidly – Dissolvable Microneedles for Transdermal Delivery via a Highly Reproducible Soft Lithography Approach

Katherine A. Moga1, Lissett R. Bickford1, Robert D. Geil1, Stuart S. Dunn1, Ashish A. Pandya1, Yapei Wang1, John H. Fain1,

Christine F. Archuleta1, Adrian T. O’Neill1, and Joseph M. DeSimone1,2,3

1The University of North Carolina at Chapel Hill, 131 South Rd, Chapel Hill, NC 27599, USA; 2North Carolina State

University, Raleigh, NC 27695 3Sloan-Kettering Institute for Cancer Research, New York, New York, 10021

kmoga@email.unc.edu

ABSTRACT SUMMARY Microneedle devices are an attractive method to

overcome the epidermis and effectively transport therapeutics transdermally. The fabrication of highly

reproducible polymer microneedles is described. These

completely dissolvable microneedle arrays are made on

flexible substrates using PRINT. Devices showed efficacy

in piercing the skin and delivering a fluorescent drug

surrogate to both ex vivo murine and human samples.

INTRODUCTION

Transdermal drug delivery is an attractive non-

traditional route of administration; it is non-invasive and

avoids first pass metabolism.1 One attractive method to

overcome the skin employs microneedle patches, arrays

of micron-sized projections for minimally-invasive drug delivery. Like hypodermic needles, these devices pierce

the skin but avoid nerve endings, causing no pain.1,2

Microneedles can transport therapeutics of virtually any

size through the skin, from small molecules to

nanoparticles.3 The low complexity of microneedle

devices may enable patient self-administration and

inexpensive fabrication. Thus, optimized microneedle

devices may offer the efficacy of a hypodermic needle with the benefits of transdermal delivery.

To overcome the barriers in fabrication of

microneedles seen previously, we have created

microneedle arrays using the Particle Replication In Non-

wetting Templates (PRINT®) technique.4 PRINT

combines a “top-down” method of soft lithography with

polymerization to create features on the nano- and micro-

scale with precise control of size, shape, and chemical composition. A wide range of materials including

polymers and pure drug could be used, and the mild

conditions required allow biologic cargo to maintain its

function throughout the process. PRINT can be adapted

on any scale, allowing affordable and quick fabrication.

Here, we demonstrate the fabrication of 100% water-

soluble microneedles on flexible substrates and their

ability to deliver a drug surrogate to ex vivo skin specimens. Array of discrete microneedles have been

manufactured via PRINT and collected on a flexible,

water-soluble substrates. This flexibility allows the array

of highly-dense microprojections to avoid this effect and

break the epidermis more efficiently. After application to

ex vivo murine and human skin specimins, the needle

patch remained long enough to allow the polymer to

dissolve and release a fluroescent drug surrogate. The substrate was then dissolved, leaving the entire

microneedle array (and drug payload) in the skin.

EXPERIMENTAL METHODS To fabricate PRINT microneedle patches, master

templates were first prepared using a tilted-rotated photolithography approach.5 First, a polished silicon

wafer was coated with thin an anti-reflective layer. A

thick layer of negative photoresist (SU-8) was

administered via spin coating and a mask (200 µm x 200

µm squares wtih 200 µm base-to-base spacing) was

applied. The complex was exposed to UV light at

incidence angles of 18-25°; the wafer was then rotated

90° about the surface normal for a total of four exposures. The resulting square pyramidal cavities were 360 µm

deep and had tip radii of curvature under 10 µm, seen via

Environmental Scanning Electron Microscopy (ESEM).

A positive replica of the master template was made

using polydimethylsiloxane (PDMS) as an intermediate.

A thick PDMS layer was cast upon the master,

centrifuged at 3000g, and cured overnight at 25°C. The

positive replica was then used to make PRINT-compatible molds from a photocurable perfluoropolyether (PFPE)

elastomer. A 0.2 wt% solution of 2,2-

diethoxyacetophenone in PFPE was cast onto the replica

and cured in a UV oven. The resulting molds are

consistent with the dimensions of the replicas,

reproducibly mimicking the SU-8 master templates.

Microneedles were fabricated using an adapted

PRINT process.4 Films of polyvinylpyrrolidone (PVP) were loaded with 0.1% rhodamine B fluorescent dye. A

film was mated to the PFPE mold and passed through a

heated nip at 105°C, filling the mold with discrete

microneedles. The filled mold was mated to a flexible,

water-soluble substrate (made from a blend of Plasdone, a

polyvinylpyrrolidone/polyvinylacetate blend, and triethyl

citrate) and passed through a heated nip at 65°C. The

mold was then removed, leaving a 100% water soluble microneedle patch. Microneedle morphology was

confirmed via ESEM and brightfield macroscopy.

Microneedle patches were tested on ex vivo nude

murine skin and human skin (obtained via the

Cooperative Human Tissue Network). Flexible PRINT

microneedle patches were “rolled” on with gentle force of

thumb and remained in the skin for a duration of either

10s or 10min. For the 10s tests, the patch backing was removed and the skin was exposed to green tissue-

marking dye for 5min. For the 10min tests, the patch

backing was then dissolved with <200 µL of tap water.

All skin samples were fixed for 2h in 2%

paraformaldehyde and left overnight in 15% sucrose in

1X PBS at 4°C. Control skin samples were also prepared;

these samples were not exposed to microneedles.

Tested murine and human skin samples were

embedded in Optimum Cutting Temperature medium and

cryosectioned. Sections (12 µm) were taken at -25°C.

Half of the sections imaged via fluorescent microscopy. The remaining sections were hematoxalin and eosin

(H&E) stained for brightfield microscopy imaging.

Staining was done using the procedure outlined by Cancer

Diagnostics for their CRYO-KIT prior to coverslipping.

RESULTS AND DISCUSSION Master templates, replicas, molds and PRINT

microneedles are shown in Figure 1A-D. The

microneedles retained the dimensions of the master with remarkable reproducibility. The flexibility of the array

can be seen in Figure 1E-F. The rigid microneedles

remained intact after the gentle bending of the array by

hand. Both the microneedles and the substrate were seen

to dissolve rapidly (~5min) in the presence of a few drops

of water. Therefore, novel 100% water-soluble

microneedle patches on flexible substrates can be made

quickly and reproducibly via PRINT processing.

Figure 1. ESEM images (A-D)of master template (A),

replica (B) and PFPE mold (C) and PVP microneedles

(D). Scale bar is 500 µm. Brightfield macroscopy (E-F) of microneedle arrays, showing flexibility. Scale bar is 1 cm.

The optimized PRINT microneedle arrays were first

tested in ex vivo murine skin samples (see Experimental

Methods). After H&E staining and brightfield imaging,

the control samples did not show any epidermal breach as

expected (Figure 2A); the skin was consistently smooth.

Evidence of epidermal breach was seen in skin sections

from both testing conditions (Figure 2B-C). Images of the unstained skin via fluorescent microscopy showed the

efficiency of the drug surrogate delivery. Seen in Figure

2D-F, a large qualitative difference in fluorescence

intensity was observed among the three samples. While

the control showed no fluorescence (Figure 2D), an

observable fluorescence was seen in the 10s test in

selective areas of the skin (Figure 2E). Comparatively,

considerably higher fluorescence intensity within the skin was seen for the 10min time period throughout the whole

skin section (Figure 2F). This confirms that the drug

surrogate was released from the needles and diffused

beneath the stratum corneum throughout the duration of

the patch application.

Figure 2. Images of ex vivo murine skin after testing with

PRINT microneedles. (A-C) Brightfield microscopic

images of skin sections after sectioning and H&E staining

[(A) control (B) 10s application (C) 10min application].

(D-F) Fluorescent microscopy images of skin after

sectioning. [(D) control (E) 10s application (F) 10min application]. Scale bar on all images is 35 µm.

In addition, pilot studies to determine the efficacy of

PRINT microneedles on human skin were also conducted.

Results from these experiments indicate that epidermal

breach and subsequent drug surrogate release. Figure 3A-

B shows a site of microneedle penetration and

corresponding rhodamine fluorescence in human skin.

While further optimization needs to be done, these findings support that PRINT microneedles may be used to

penetrate human skin and deliver loaded cargo.

Figure 3. Brightfield image (A) of a skin after

microneedle insertion for 10s. (B) Fluorescence image

after microneedle insertion for 10min. Scale bar is 70 µm.

CONCLUSION PRINT microneedle arrays (100% water-soluble)

were fabricated on flexible substrates. These arrays were

efficacious in piercing skin and delivering drug surrogate.

REFERENCES 1. Sullivan, S. P.; Murthy, N.; Prausnitz, M. R. Adv.

Mater. 2008, 20, 933-938.

2. Escobar-Chávez, J. J.; et al. J. Clin. Pharmacol.

2011, 51, 964-977. 3. Coulman, S. A.; et. al. Int. J. Pharm. 2009, 366, 190-

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4. Rolland, J. P.; et. al. J. Am. Chem. Soc. 2005, 127,

10096-10100.

5. Han, M.; Lee, W.; Lee, S. K.; Lee, S. S. Sensors and

Actuators A: Physical 2004, 111, 14-20.

ACKNOWLEDGMENTS Financial support was provided by the NIH Pioneer

Award, the University Cancer Research Fund at UNC

Chapel Hill, and the Charles N. Reilley Graduate

Fellowship through the UNC Department of Chemistry.

PRINT is a registered trademark of Liquidia

Technologies, Inc, and we thank them for their support.