ARVO 2015 Annual Meeting Abstracts
206 Drug and gene delivery systems
Monday, May 04, 2015 8:30 AM–10:15 AM
1EF Mile High Blrm Paper Session
Program #/Board # Range: 1294–1299
Organizing Section: Physiology/Pharmacology
Program Number: 1294
Presentation Time: 8:30 AM–8:45 AM
Impact insertion of transfer-molded microneedle for localized
and minimally invasive ocular drug delivery
Hyun Beom Song1, 2, Kang Ju Lee3, Sang-Mok Lee4, Jin Hyoung Kim1,
WonHyoung Ryu3, Jeong Hun Kim1, 2. 1Fight against AngiogenesisRelated Blindness (FARB) Laboratory, Biomedical Research
Institute, Seoul National University Hospital, Seoul, Korea (the
Republic of); 2Department of Biomedical Sciences, Seoul National
University Graduate School, Seoul, Korea (the Republic of);
3
Department of Mechanical Engineering, YONSEI University, Seoul,
Korea (the Republic of); 4Department of Ophthalmology, Hallym
University Sacred Heart Hospital, Anyang, Korea (the Republic of).
Purpose: It has been challenging for microneedles to deliver
drugs effectively to thin tissues with little background support
such as cornea. The aim of this study is to demonstrate safety
and efficacy of newly designed microneedle pen (MNP) system,
a single microneedle (MN) with a spring-loaded applicator to
provide impact insertion.
Methods: Desired shape of master mold was fabricated by
KOH wet etching of silicon wafer. PDMS cavity molds were
fabricated from the master mold and used for transfer-molding
to attach MN to the tip of a spring-loaded applicator. A mixture
of drug and carrier materials was prepared and loaded on the
surface of MN tips using a dip coating method. Rhodamine B,
Evans blue, or sunitinib malate were used as model drugs in
this experiment. To investigate the localized delivery and the
safety of MNPs, MNs coated with Evans blue were applied to
mouse corneas and area of distribution and corneal thickness
were compared with methods using 30G hypodermic needles.
To investigate the efficacy of MNPs in delivering drugs, we
delivered sunitinib malate with MNPs and evaluated their
inhibitory effect on suture-induced corneal angiogenesis. Right
after the intrastromal suture, sunitinib malate was delivered and
corneal neovascularization was evaluated on the eighth day after
injection.
Results: MN made of SU-8 had the base area of 200 × 200
mm2 and the height of 140 mm. Application of MNPs on mouse
cornea successfully delivered rhodamine dye to the stroma
without perforation. When compared with delivery of Evans
blue using 30G needle syringe or 30G needle tip, MNPs achieved
the smallest area of distribution (886 ± 64 μm2) and left no
evidence of corneal edema on the next day. Delivery of sunitinib
malate with MNPs to suture-induced corneal angiogenesis
model effectively decreased vessel length and clock hour of
vascularization while delivery with 30G needle tip had no effect.
Conclusions: We designed a novel applicator that enables a
targeted and impact-insertion of a MN in order to deliver drugs
to thin tissues with little background support such as corneas.
To enhance applicability of MN to macro-applicator, we applied
transfer molding process. The fabricated MNPs could achieved a
localized, minimally invasive and effective drug delivery to tissues
without much background support.
Commercial Relationships: Hyun Beom Song, None; Kang
Ju Lee, None; Sang-Mok Lee, None; Jin Hyoung Kim, None;
WonHyoung Ryu, None; Jeong Hun Kim, None
Program Number: 1295
Presentation Time: 8:45 AM–9:00 AM
Assessing ocular hyaluronidase to develop a hyaluronan-based
polymer drug delivery platform
Randy C. Bowen1, Won Y. Lee2, David W. Grainger3, Barbara M.
Wirostko2. 1School of Medicine, University of Utah, Salt Lake
City, UT; 2Jade Therapeutics, Salt Lake City, UT; 3Departments of
Pharmaceutics and Bioengineering, University of Utah, Salt Lake
City, UT.
Purpose: Current treatments for ophthalmologic diseases require
frequent and long-term therapy causing inconvenience and financial
burden to patients. Cross-linked hyaluronan (CMHA) matrix such as
Hystem® enables us to design sustained release drug delivery system
potentially enhancing the efficacy of the treatment. Drug release
from CMHA matrix largely depends upon the degradation of CMHA
matrix by hyaluronidases (HAase) present in various ocular tissues.
However, the presence of ocular HAase is poorly understood. Our
study aims to address this issue by investigating the concentrations of
HAase (Hyal-1) in various ocular tissues.
Methods: Human aqueous humor (hAH), vitreous (hV), sclera (hS),
and cornea (hC), and monkey (RxGen, St. Kitts) aqueous humor
(mAH), vitreous (mV), sclera (mS), cornea (mC), and lens (mL)
were collected from cadaver eyes and prepared to prevent crosscontamination. HAase concentrations in all human ocular samples
(hAH=5, hV=8, hS=2, hC=2) and 24 monkey ocular samples
(mAH=7, mV=7, mS=3, mC=4, mL=3) were determined using
commercially available ELISA kits (R&D, MN, USA; Cusabio,
China).
Results: Hyal-1 was present in all parts of the ocular tissues tested
except in the lens. The concentration of Hyal-1 was homogeneous
in human samples but varied in monkey tissue samples. In human
tissues, Hyal-1 concentrations were 1.4–2.3 ng/mL in hAH; 3.2-4.7
ng/mL in hV; 1.9-2.3 ng/mL; 1.2-4.4 ng/mL in hC. In monkey tissues,
Hyal-1 were 1.8–15.2 ng/mL in mAH; 4.6-5.0 ng/mL in mV; 2.7-4.3
ng/mL in mS; 9.9-12.2 ng/mL in; but undetectable in mL.
Conclusions: Ocular Hyal-1 concentration varies depending on the
tissues in the eye. We provide quantitative data on HAase levels that
correlate to enzymatic activity within the eye. These data will support
our effort to optimize drug delivery system using CMHA matrix
for diseases occurring in various ocular tissues thereby reducing
treatment frequency and enhancing therapeutic efficacy.
Commercial Relationships: Randy C. Bowen, Jade Therapeutics
(F); Won Y. Lee, Jade Therapeutics (C), Jade Therapeutics (F);
David W. Grainger, Jade Therapeutics (C), Jade Therapeutics (F),
Jade Therapeutics (I); Barbara M. Wirostko, Jade Therapeutics (E),
Jade Therapeutics (F), Jade Therapeutics (I), Jade Therapeutics (P),
Jade Therapeutics (S)
Support: NSF Grant IIP-1430921
Program Number: 1296
Presentation Time: 9:00 AM–9:15 AM
Biodegradable Sustained-Release Drug Delivery Systems
Fabricated using a Dissolvable Hydrogel Template Technology
for the Treatment of Ocular Indications
Nikita Malavia, Laxma Reddy, Istvan Szinai, Norman Betty, Justin
Pi, Jeremiah Kanagaraj, Alisha Simonian, Robert Jennings, Glenn
Stoller. Ohr Pharmaceutical Inc., San Diego, CA.
Purpose: Biodegradable drug delivery systems can provide a
continuous drug supply to target sites for extended periods. These
systems dramatically improve compliance by decreasing the
burden of repeated injections in ocular diseases. A dissolvable
hydrogel template technology was used to produce biodegradable
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
microparticles custom designed in three dimensions for sustained
release drug delivery.
Methods: In this study OHR1031, a small molecule drug for
glaucoma, was incorporated into microparticles using a dissolvable
hydrogel template technology. The drug was dissolved into a PLGA
polymer-solvent mixture. The mixture was applied to templates with
wells of uniform size that define the resulting microparticle size
and shape. Templates were dissolved in water. Microparticles were
recovered by filtration and dried. Particle size analysis was performed
using a laser light scattering instrument. An in vitro release study was
conducted in phosphate buffered saline at 37oC using conditions to
ensure drug solubility did not impact drug release rates. Drug content
in the microparticles and release media was measured by HPLC.
Results: In this example, microparticles obtained using the
dissolvable template technology had a median particle size of 60 μm
with 80% of the particles within 10 μm (Figure 1). OHR1031 content
in the microparticles was 57% w/w with almost 100% incorporation
efficiency. Drug release rate was nearly zero-order for over three
months with virtually no initial burst (Figure 2).
Conclusions: The hydrogel template technology is amenable to small
molecules and biologics. It provides a means to achieve high drug
content in biodegradable materials with high incorporation efficiency.
Microparticles are recovered from readily soluble hydrogel templates.
Any water insoluble material can be chosen as the microparticle
matrix to achieve the desired release durations. The technology
allows for precise control of size and shape of template wells in
every dimension. The resulting microparticles have a narrow size
distribution reflecting the size and shape of template wells. These
attributes enable microparticle injection using narrow bore needles
into sensitive ocular spaces of long acting drug products.
Particle size distribution of microparticles recovered from a
dissolvable template
In vitro release of OHR1031 from PLGA microparticles
Commercial Relationships: Nikita Malavia, Ohr Pharmaceutical
Inc. (E); Laxma Reddy, Ohr Pharmaceutical Inc. (E); Istvan Szinai,
Ohr Pharmaceutical Inc. (E); Norman Betty, Ohr Pharmaceutical
Inc. (E); Justin Pi, Ohr Pharmaceutical Inc. (E); Jeremiah
Kanagaraj, Ohr Pharmaceutical Inc. (E); Alisha Simonian, Ohr
Pharmaceutical Inc. (E); Robert Jennings, Ohr Pharmaceutical Inc.
(E); Glenn Stoller, Ohr Pharmaceutical Inc. (C)
Program Number: 1297
Presentation Time: 9:15 AM–9:30 AM
Safety Assessment of a Novel, Cross-Linked, Bio-Absorbable
Carboxymethyl Hyaluronic Acid (CMHA) Polymer in a Rabbit
Ocular Surface Model
Glenwood G. Gum1, Barbara M. Wirostko2, 3, MaryJane Rafii2,
Vatsala Naageshwaran1, Matthew Lyulkin1, Ruebuen Merideth4,
Brenda Mann5. 1Absorption Systems, San Diego, CA; 2Jade
Therapeutics, Salt Lake City, UT; 3University of Utah, Moran Eye
Center, Salt Lake City, UT; 4Eye Care for Animals, San Diego, CA;
5
SentrX Animal Care, Salt Lake City, CA.
Purpose: The safety and tolerability of a proprietary, cross-linked,
biodegradable hyaluronic acid (CMHA)-based polymer being
developed as a dry eye and corneal repair therapy was evaluated in a
28-day rabbit study.
Methods: Male and Female New Zealand white rabbit eyes were
dosed via bilateral topical dosing six times per day using two
concentrations (0.4% and 0.75%) of the CMHA (SentrX, UT) for
28 days. A subset of animals was observed for an additional 14 days
after the final dose of the test article. Slit-lamp biomicroscopy and
indirect ophthalmoscopy were performed at baseline and at Days
7, 14, 21, 28, 35 and 42. The examinations utilized the McDonaldShadduck scoring system and were performed by a board certified
veterinary ophthalmologist. Representative photographs were
taken at various exam time points. IOP was performed at baseline,
immediately prior to dosing, and prior to termination. Complete
blood count, clinical chemistry, and coagulation factor testing was
performed for all animals in the study prior to dosing and prior to
necropsy. Urine was collected at the time of necropsy for urinalysis.
Blood was collected at baseline (pre-dose) and at 28 days prior to
termination for clinical chemistry. Eye globes were enucleated and
adnexa collected from each animal and preserved for histopathology
analysis. Gross necropsy was conducted and main organs collected
and stored for future histopathology analysis.
Results: The compound was safe and well tolerated. Animals
exhibited a lack of clinical anomalies, with the exception of some
mild conjunctival congestion in a minimal number of animals. No
effect on the IOP was observed during the course of the study. In
addition, clinical pathology values were within normal ranges.
Conclusions: Variations of this novel CMHA polymer have
previously been shown to be efficacious in promoting ocular wound
healing and to act as a potential ocular drug delivery vehicle for small
and large molecules. This study establishes the ocular tolerability and
safety of one version of the polymer.
Commercial Relationships: Glenwood G. Gum, Absorption
Systems (E); Barbara M. Wirostko, Jade Therapeutics (E), Jade
Therapeutics (F), Jade Therapeutics (I), Jade Therapeutics (P), Jade
Therapeutics (R), Jade Therapeutics (S); MaryJane Rafii, Jade
Therapeutics (E), Jade Therapeutics (F), Jade Therapeutics (I), Jade
Therapeutics (P), Jade Therapeutics (R), Jade Therapeutics (S);
Vatsala Naageshwaran, Absorption Systems (E); Matthew Lyulkin,
Absorption Systems (E); Ruebuen Merideth, Eye Care for Animals
(C); Brenda Mann, SentrX Animal Care (E)
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].
ARVO 2015 Annual Meeting Abstracts
Program Number: 1298
Presentation Time: 9:30 AM–9:45 AM
TRPV4 channels regulate the inflow pathway in the anterior eye
Andrew O. Jo1, Daniel A. Ryskamp1, Amber Frye1, Tam T. phuong1,
Bruce A. Berkowitz2, David Krizaj1. 1Ophthalmology & Visual
Sciences, University of Utah, South Jordan, UT; 2Anatomy, Wayne
State University, Detriot, MI.
Purpose: Aqueous humor dynamics is regulated by balanced
production in the ciliary body and removal through the primary
(“conventional”) and secondary outflow pathways. Fluid production
is highly influenced by osmoregulatory mechanisms. To determine
the molecular mechanism regulating inflow, we examined the
expression and function of the osmosensory/mechanosensitive
TRPV4 channels in the anterior eye.
Methods: The ciliary body was dissected from the mouse eye
and incubated with agmatine (AGB, a cation influx marker). The
tissue was exposed to selective TRP channel agonists, fixed and
immunostained for AGB. Alternatively, epithelial cells were loaded
with the calcium indicator dye fura-2 and stimulated with TRPeffective compounds; calcium concentration [Ca2+]i was determined
using high-resolution optical imaging. The effect of TRP channel
agonists in vivo was determined using high-resolution manganeseenhanced resonance imaging (MEMRI). Eye sections were processed
for hematoxylin and eosin, apoptosis markers, metabolic markers,
and EM ultrastructure.
Results: TRPV4 immunoreactivity was observed was expressed
in the pars plicata of the fluid producing, non-pigmented epithelial
(NPE) cells, but was absent from pigmented epithelial cells of the
ciliary body. Hypotonic stimuli induced dose-dependent increases in
NPE cell volume, which in turn was associated with entry of cations
into the ciliary body. Both swelling response and hypotonicityinduced Ca signals were TRPV4-dependent. Accordingly, selective
TRPV4 agonists such as GSK1016790A increased [Ca2+]i in NPE
cells, augmented AGB immunoreactivity, and increased manganese
uptake via calcium channels into the ciliary body in vivo. The extent
of hypoosmotic swelling was reduced by BAPTA. Importantly
HTS-induced changes in volume regulation, kinetics of hypotonic
induced [Ca2+]i signals, and calcium signal amplitude to GSK were
compromised in mice lacking aquaporin 4 (AQP4) channels.
Conclusions: Our data provide molecular and physiological evidence
that TRPV4 mediates osmosensation in the ciliary body, possibly as
part of a macromolecular complex that includes the water channel
AQP4. Thus, these findings suggest a novel mechanism that regulates
inflow pathway in the anterior eye by balancing the bidirectional
water transport and ion intake in NPE cells. By sensing NPE volume
and adjusting aqueous secretion, TRPV4 is proposed to represent a
linchpin in the regulation of intraocular pressure.
Commercial Relationships: Andrew O. Jo, None; Daniel A.
Ryskamp, None; Amber Frye, None; Tam T. phuong, None; Bruce
A. Berkowitz, None; David Krizaj, None
Support: EY13870, EY022076, P30 EY14800, Department of
Defense, Glaucoma Research Foundation, State of Utah TCIP and an
unrestricted award from Research to Prevent Blindness to the Moran
Eye Institute.
bioavailability in the treated eye. This study aims at studying the
decisive value of contralateral eye results in a case of comparative
pharmacokinetic (PK) study of a drug versus its prodrug.
Methods: The studied drug was 3H radiolabeled dexamethasone
(in 0.1 % preparations) as a base (MW=392, logP=1.77) or as a
lipophilic prodrug (MW=630, logP=7.93). In two separate PK’s, the
drug or its prodrug were determined in aqueous humor and vitreous
of both eyes, as well as in the systemic blood and cerebrospinal fluid
(CSF), at 15, 30, 45, 60, 120, 180 and 360 minutes after one single
topical application (50 mL) to the cul-de-sac of the treated albino
rabbit eye. The area under curve of drug concentration against time
(AUC.0-last) was calculated (min. x ng/gr) for each sampled site in
both PK’s. The relative bioavailability, of the drug with respect to the
prodrug, for each of the sampled sites was calculated as follows: F.rel
= AUC.drug / AUC.prodrug.
Results: F.rel values were 5.599, 1.533, 1.737, 1.359, 0.931 and
1.017 for treated eye aqueous, treated eye vitreous, blood, CSF,
contralateral eye aqueous and contralateral eye vitreous respectively.
Conclusions: In the present study the relative bioavailability
of dexamethasone (drug/prodrug) have surprisingly showed
bioequivalence, between the drug and the prodrug, in the aqueous
humor and vitreous of the contralateral eye. This bioequivalence was
contrary to the decisive F.rel in the treated eye aqueous humor and
vitreous, which was in favor of the drug over the prodrug in both
cases, and with no correlation to the F.rel in either the blood or the
CSF, where in both cases the results were again in favor of the drug
over the prodrug . This discrepancy could be related to the main site
of prodrug cleavage. Yet, the present results challenges the possibility
of a single or one principle mediating-compartment between both
eyes, including the traditional (systemic blood circulation) and the
alternative (CSF) mediating-compartment assumptions. Finally, the
traditional way of concluding the net bioavailability in the treated
eye with reference to the contralateral eye (net bioavailability =
AUC.treated - AUC.contralateral) should be questioned in cases of
comparisons between a drug and its prodrug.
Commercial Relationships: Muhammad Abdulrazik, None
Program Number: 1299
Presentation Time: 9:45 AM–10:00 AM
The Decisive Value of Contralateral Eye Results in Comparative
Ocular Pharmacokinetic Studies of a Drug Versus its Prodrug
Muhammad Abdulrazik. Ophthal/Innovative Interventions, East
Jerusalem Biomedical Institute, East Jerusalem, Palestine, State of.
Purpose: The extent of drug retention in the untreated contralateral
eye is being traditionally considered in concluding the net
©2015, Copyright by the Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Go to iovs.org to access the version of record. For permission
to reproduce any abstract, contact the ARVO Office at [email protected].