Intravitreal (Day 21)

Purpose

Precisely Engineered Biodegradable Intraocular Implants

for the Sustained Release of Dexamethasone Andres Garcia, Janet Tully, Benjamin Maynor, Benjamin Yerxa.

Liquidia Technologies, P.O. Box 110085, RTP, NC 27709

Corresponding Author: Andres Garcia, andres.garcia@liquidia.com, (919) 328-4388

Figure 1. PRINT Process. A liquid fluoropolymer (green) is added to the surface of a micropatterned

“master template” (grey) and photochemically crosslinked to generate a precise mold having micro- or

nanoscale cavities (upper middle). This mold is then filled with drug (top row, right). Particles can be

removed (bottom row, middle) from the mold and isolated as stable dispersions or free flowing powders

(bottom row, left). PRINT particles (red) maintain the dimensions of features on the master template.

Results

PRINT® Technology

Brings the precision and control of semiconductors to life sciences and other markets

Proprietary design and manufacturing platform to produce micro- and nano-particles

Monodisperse feature morphology designed into master template

Readily scalable using proven roll-to-roll manufacturing process

The ability to fabricate biodegradable intraocular implants with uniform size, shape and dose for

the sustained delivery of actives in multiple regions of the eye has proven elusive with current

technologies. The acceptance of intravitreal implants for the localized treatment of multiple back-of-

the-eye conditions have paved the way for the development of a new generation of smaller intraocular

implants in the anatomically and clinically desirable, yet “hard-to-manufacture” size range of 100μm to

1,000μm. The ability to reproducibly fabricate implants in this size range opens up a window of

opportunities for the injection and localization of implants against multiple target tissues of the inner

eye where greater spatial constraints may exist. We have previously described a novel particle

manufacturing technology, Particle Replication in Non-wetting Templates (PRINT®), for the production

of mono-disperse particles across multiple areas of drug delivery (1), as outlined in Figure 1. Using

the PRINT methodology, we report the ability to precisely fabricate 200μm x 200μm x 1,000μm

biodegradable implants for the sustained delivery of actives in the eye.

References: 1. Garcia et al. (2012), “Microfabricated engineered particle systems for respiratory drug delivery and other

pharmaceutical applications,” Journal of Drug Delivery.

Methods

Commercial Relationships Andres Garcia, Janet Tully, Benjamin Maynor and Benjamin Yerxa are all employees (E) of, and have personal

financial interest (I) in, Liquidia Technologies.

Using the PRINT technology four implant formulations comprised of a blend of 20% w/w dexamethasone

(DXM) and 80% of a biodegradable polymer (with varying degrees of molecular weights and lactide:glycolide

ratios) were prepared:

• Formulation 1: dexamethasone / Poly(D,L-lactide)

• Formulation 2: dexamethasone / Poly(D,L-lactide)

• Formulation 3: dexamethasone / Poly(D,L-lactide-co-glycolide)

• Formulation 4: dexamethasone / Poly(D,L-lactide-co-glycolide)

Physicochemical characterization of the implants was performed and dexamethasone release in-vitro was

evaluated:

• Physical morphology : implants were analyzed by scanning electron microscopy.

• Overall mass uniformity: implant mass of individual PRINT implants (n= 15) was measured using Mettler

MT5 microbalance for all formulations.

• Dexamethasone content uniformity: Dexamethasone content of individual PRINT implants at t=0 (n=15) was measured using a RP-HPLC method and Phenomenex Luna Phenyl-Hexyl, 3µm particle size, 4.6 x 100

mm analytical column. Mobile phase consisted of a gradient of 0.1% TFA in purified water and 0.1% TFA in

acetonitrile over 12 minutes at 1 mL/min. UV absorbance of dexamethasone was measured at 240 nm.

• In-vitro release of dexamethasone from the implants: The release profiles of individual PRINT implants

(n=20) were monitored at sink conditions for 141 days. Individual PRINT implants were incubated in 500μL of

1X PBS at 37°C (total possible dexamethasone concentration in release media = 20μg/mL). Supernatant of

each was sampled at 1, 3, 7, and 14 days, and at 4 week intervals thereafter, to measure cumulative

dexamethasone released from implants.

• Initial in-vivo implant injections: PRINT implants were placed in a 25G needle on a syringe prefilled with

viscoelastic solution (Viscoat® sodium hyaluronate – sodium chondroitin sulfate). Intravitreal injection s were

performed on anesthetized New Zealand white rabbits under operative microscope. Approximately 50µl of

Viscoat® were injected along with the implant. Fundus was observed using the slit lamp 15min after the

injection, on day 2, 7 and 14. Implant was retrieved from vitreous on day 21.Similarly, subconjuctival and

intracameral injections were performed and implants were evaluated over time.

Table 1. Summary of implant mass uniformity, measured dexamethasone loading in implant formulations

and % dexamethasone released from implants in-vitro after 141 days in 1X PBS at 37°C.

PRINT Formulation 1

Figure 2. Scanning electron micrographs of four different implant formulations consisting of a blend of

dexamethasone and a biodegradable polymer. Implant size for all formulations: 200µm x 200µm x 1000µm.

Figure 3. In-vitro release of dexamethasone (% DXM released) from 200µm x 200µm x 1000µm PRINT

implants over 141 days in 1X PBS at 37°C.

Intravitreal (Day 14)

Anterior part of the eye removed. Implant was visible and recovered from the center of the vitreous.

Preliminary in-vivo studies: subconjuctival, intravitreal and

intracameral injections of PRINT implants in rabbit eyes

Subconjuctival (Day 12)

Intracameral (Day 12)

Subconjuctival (Day 30)

Figure 4. Photographs showing placement of PRINT implants in various compartments of the eye. Implants

were inserted in the eye via subconjuctival, intravitreal and intracameral injections and were well-tolerated

for the duration of the study.

Conclusions The PRINT technology uniquely allows for the fabrication of intraocular implants with uniform size,

shape and dose. We demonstrated the ability to fabricate dexamethasone intraocular implants in

the desirable size range of 100μm to 1,000μm for sustained release applications where anatomical

constraints may call for uniquely engineered implants. PRINT implants are well-tolerated in-vivo and

offer a unique, new paradigm for the sustained delivery of actives in the eye.

No post-insertion adverse effects

observed

Subconjunctival Insertions: No suture needed.

No chemosis. No erythema.

Intravitreal injections: implants visible in the

middle of the vitreous, no inflammation.

Intracameral injections: Insertion through the

limbus rather than cornea. Implant localized at

site of injection, close to the angle at the 6

o’clock position. No iris inflammation.

New implant

Recovered after 21 days in vitreous

Implant morphological comparison after 21 days in vitreous

PRINT Formulation 2

PRINT Formulation 3 PRINT Formulation 4

PRINT implant in a 27 gauge, thin wall needle

Formulation Measured PRINT

implant mass at t = 0 (n=15)

Measured DXM mass in individual PRINT implants by HPLC at

t = 0(n=15)

Measured% DXM loading in PRINT implants at

t = 0

(API mass/implant mass)

141 day in-vitroevaluation

% DXM released from PRINT implants at

t = 141 days

(n=20)

AVE STDEV AVE STDEV AVE AVE

Formulation 1 52 μg 1 μg 10 μg 1 μg 19% 83%

Formulation 2 53 μg 1 μg 9 μg 1 μg 18% 1%

Formulation 3 54 μg 2 μg 10 μg 1 μg 19% 83%

Formulation 4 51 μg 1 μg 9 μg 1 μg 18% 93%