Ocular barriers and drug delivery

MSc Course. 27 Nov 2017, S Brocchini

Learning points
• Basic knowledge of eye anatomy.
• Basic familiarity of diseases of the eye, specifically glaucoma and diseases
to the back of the eye (eg AMD).
• Be able to describe the corneal, precorneal and retinal barriers. Describe
the vitreous and aqueous.
• Describe the clearance and absorption pathways of eye drops including
the influence of the lacrimal system and the structure of tears, tear volume,
turnover and drainage features. Basic knowledge about the composition of
tears. Understand the effects of blinking. Be able to describe the structure
of the cornea.
• Understand the influence of pH, osmolality, surface tension in eye drops.
• Describe the clearance pathways for intravitreal injections and implants.
• Understand the issues about the administration of drugs to the back of the
eye. Be able to describe the use of steroids and therapeutic proteins to
treat blinding disease at the back of the eye.
Basic Anatomy
 eye). Eventually the aqueous humor drains from the eye via specialized tissues located

by University College London on 03/06/12. For p

Annu. Rev. Fluid Mech. 2012.44:347-372. Downloaded fro
of the anterior chamber, where the iris, cornea, and sclera (the white part of the eye
Physical and biochemical barriers also Figure 2 and Sections 2.1 and 2.2). These specialized tissues have a significant hyd
flow resistance, and the drainage of the aqueous humor out of the eye therefore require
be a positive pressure within the eye itself, the so-called intraocular pressure (IOP).
a Blood retinal barrier
b Arterioles Collector channels
Ciliary
Schlemm’s body
canal Vortex vein
Trabecular
meshwork Sclera
Iris Cornea

Cornea Choroid

Zonules
Iris RPE

Lens Retina

Schlemm’s canal Aqueous veins

Retinal vessels
Optic
Ciliary processes nerve
Ciliary muscle
Posterior
chamber Anterior
chamber
Trabecular
meshwork Ciliary
processes
Schlemm’s
canal Ciliary
Figure 1
muscle
Overview of a human eye with major anatomical structures identified. Abbreviation: RPE, retina
epithelium. Figure modified from Krey & Bräuer (1998), copyright ⃝ c MSD SHARP & DOHM
2
Aqueous flow-a physical barrier Germany with kind permission.
tion of aqueous humor flow patterns in the anterior chamber and key drainage tissues. (a) Cross-sectional view through the
r eye. The arrows show typical thermal convection patterns of aqueous humor in the anterior chamber for an upright subject in
bient temperature less than 37◦ C and also drainage pathways
348 from the anterior
Siggers ·
Ethier chamber into Schlemm’s canal and thence into
ueous veins/collector channels (bottom-most and top-most arrows). (b) Anterior-posterior view of Schlemm’s canal (thick green ring),
internet
RI 18 November 2011 14:2

The retina is complex

Membrane Transport Processes in the Eye 35
Direction of
light travel

Vitreous humor

Ganglion cell layer

Inner plexiform layer

Inner nuclear layer

Outer plexiform layer

Outer nuclear layer

Inner segments of photoreceptors
Outer segments of photoreceptors
Retinal pigment epithelium

Bruch’s membrane
Choroid

50 µm Sclera
Figure 9 Cellular organization of the retina.
ion through the retina and choroid within the macula (the part of the retina with the greatest
ion of rod and cone cells). The asterisk denotes the choriocapillaris, and the white arrowheads
ruch’s membrane. Reprinted from Curcio et al. (2009) with permission of Elsevier.
ability properties of inner limiting membrane are likely similar to the basal
hese have the effect of creating flow patterns within the vitreous humor, particularlylaminae,
if which retain particulate matter but allow the transport of mole-
uid characteristics, as explained in Section 4. A common cause of vision loss in elderly
cules up to the size of ferritin (400 kDa) (Wu, 1995). Among all the retinal
s age-related macular degeneration, which is thought to be caused by impaired transport
uch’s membrane, the membrane situated at the base of the retina, due to accumulation layers, the principal barrier to solute transport is the blood-retinal barrier
articles deposited within the membrane over a period of years (see Section 5).
(BRB), which includes retinal pigment epithelium and retinal vessels.
the notable exception of the lens, central cornea, and the vitreous humor, the eye is
pplied by a complex network of blood vessels, leading to many interesting physiological The BRB is located at two levels: the outer BRB, consisting of the
associated with the regulation of blood flow in the network. For example, the retinaretinal
has pigment epithelium (RPE), and the inner BRB, consisting of the
ably high metabolic rate and a correspondingly large need for blood, which was recently
ted by Liu et al. (2009) in a reconstructed network model to compute flow and mass endothelial membranes of the blood vessels of the retina. BRB plays a
. Moreover, because the ocular vasculature is contained within the eye globe, which critical
is role in the homeostatis of the neural retina by limiting the entry of
ssurized, the ocular veins can experience collapse, behaving as a Starling resistor. The
xenobiotics into the extravascular spaces of the retina and by preventing the
Some common ocular conditions

Periocular Diseases Intraocular Diseases

• Treated using topical formulations • Intraocular infections of the
• Periocular diseases include: aqueous humor, iris, vitreous humor
Blepharitis & retina
Conjunctivitis • Glaucoma:
Keratitis
Trachoma 2% of the population over 40 years;
Dry Eye • Increased pressure in the eye
• Define as IOP > 21 mm Hg
Diseases of the • Destruction of optic nerve
• Visual loss is irreversible
back of the eye • Second leading cause of blindness
• Cytomeaglovirus retinitis (CMVR)
• Proliferative vitreoretinopathy (PVR)
• Diabetic retinopathy
• Age-related macular degeneration
• Endophthalmitis
• Retinitis pigmentosa
 Topical administration. Eye drops
• Two pathways for ocular absorption: (1) The corneal route and (2) the
conjunctival / scleral route
• Conjunctival absorption is non –productive and is an additional drug loss
following topical dosing.
Ocular delivery

Nasolacrymal drainage Precorneal barrier Tear turnover

Drug metabolism
Protein drug binding

Ocular absorption

Nasal epithelium Conjuctival and scleral route Corneal route

Aqueous humor

Systemic circulation Ocular tissue (retina, iris, ciliary body)

Elimination
Expert Opion, Drug Deliv. (2012) 9(4)
The cornea
• Principal pathway for ocular absorption
• Paracellular or transcellular diffusion
• Little evidence of active transport
• Absorption largely governed by drug lipophilicity, although solubility, molecular
size and shape, charge and degree of ionisation are also factors.
The cornea
The Epithelium
• 10% of cornea thickness
• 5-6 cell layers
• 50-100 µm thick
• Hydrophobic tissue
• 90% of barrier to hydrophilic drugs, 10% to
hydrophobic drugs
The Bowman’s Membrane Descemet’s Membrane
• 6 µm thickness
• 8-14 µm thick homogeneous sheet • Strong, homogeneous, resistant membrane
• Does not regenerate if destroyed • In a state of tension
• Not a barrier to drug absorption • Regenerates if damaged
The Stroma
• 90% thickness of cornea The Endothelium
• Modified connective tissue • Single layer of flattened cells completely
• 70-80% wet weight is water covering the posterior surface of the cornea
• 20-25% dry weight is collagen, other • Gap junctions exist permitting permeation
proteins and mucopoly-saccharides of molecules
• Main barrier to hydrophobic drugs • Not rate-determining (permeability 200+
times that of the epithelium)
 Tears
• Precorneal loss of drug is due to tear turnover, drainage, systemic
absorption, and in situ metabolism.
• Tear turnover dilutes the drug, thus reducing the gradient of transport
through the cornea.
• Proteins in lacrimal fluid influence viscosity
• Viscosity 1.3-5.9 cps (mean 2.92 cps)
• Pseudoplastic
• High shear rate during blinking (10,000-40,000 s-1

The Tear Film

has structure
Constraints to topical ocular drug delivery
Loss of drug
– spillage
• normal tear volume=7 µl
• volume that can be accommodated without spillage=30 µl
• estimated drop volume =50 µl
• blinking > residual volume of ~10 µl
– removal by the naso-lacrimal apparatus
• takes place when reflex tearing causes
volume to exceed 7-10 µl
• eventually goes to GI tract: Potential
systemic effects, salty/bitter taste
• Major factor contributing to precorneal drug loss and poor ocular
bioavailability Max. volume held in lower eyelid sac is 25-30 µL. 3 µL
can be introduced into the tear film (7-9 µL) before it is destabilised
Eye drop characteristics that influence absorption
pH
• Normal tear pH is 7.4
• Reflex tears & drainage remove high & low pH solutions
• Ophthalmic solution pH 7.0-7.7
• If drug unstable in this range, then can formulate between 5-8 but with little or
no buffering capacity
Surface Tension
• Surface tension of tears is approx. 44-47 mN m-1. (Dry eye patients
approx. 50 mN m-1).
• Tear film is destabilised if surface tension falls well below the tear value.
This can occur with surface active drugs or excipients.
Osmolality
• Integrity of cornea is influenced by osmolality (tears 290-310 mOsm kg-1)
• Variations between 100-640 mOsm kg-1 are well tolerated
• Hypotonoic solutions can lead to better corneal absorption due to water
uptake by corneal tissues.
Clearance profile

Drug concentration –
time profile for eye drop
administration

Percent of topically
applied dose absorbed
into the systemic
circulation
 Table 1 Comparison of Anatomical and Physiological
Comparison of ineye
Factors the characteristics
Rabbit and Humanbetween
Eye rabbit and humans

Factor Rabbit Human

Tear volume (mL) 5–10 7–30

Tear turnover rate (mL/min) 0.6–0.8 0.5–2.2
Spontaneous blinking rate 4–5 times/h 6–15 times/min
Nictitating membrane Present Absent
Lacrimal punctum/puncta 1 2
pH of lacrimal fluids 7.3–7.7 7.3–7.7
Milliosmolarity of tears 305 305
Corneal thickness (mm) 0.40 0.52
Corneal diameter (mm) 15 12
Corneal surface area (cm2 ) 1.5–2.0 1.04
Ratio of conjunctival surface 9 17
and corneal surface
Aqueous humor volume (mL) 0.25–0.3 0.1–0.25
Aqueous humor turnover 3–4.7 2–3
rate (mL/min)

Source: Adapted from Refs. 8, 9.

Enhancement of Corneal Drug Permeability
• Transient modification of epithelium (e.g. use of chelators or
surfactants)
• Modify drug structure (lipophilicity, solubility & pKa) e.g. Prodrug
approach.
IOP lowering by Adrenaline and Dipivoyl Adrenaline
Other modes of administration
Systemic Delivery
• Some medicines can distribute into ocular tissues following systemic
administration
• Oral administration of carbonic anhydride inhibitors e.g. acetazolamide,
methazolamide & dichlorphenamide for glaucoma treatment reach the
ciliary processes to inhibit carbonic anhydrase isoenzyme II & hence
reduce the secretion of aqueous humor
• Steroids and antibiotics also by this route
• HIGH DOSES USUALLY REQUIRED

Intraocular Administration
• Intravitreal injections
• Intraocular implants
Intravitreal injection
Intravitreal (IVT) injection)-the only clinical route to give a high, ‘reproducible’ dose
• To be efficacious, a medicine needs to be in the posterior cavity.
Clearance routes from the posterior cavity
-Posterior (RCS; permeable molecules)
-Anterior (aqueous outflow; large, charged molecules)

retinal-choroid-
Anterior pathway
sclera (RCS)
pathway

Aqueous outflow is
responsible for most
mass transfer in the Posterior cavity
eye.

• Most large, charged molecules have a half life in the posterior cavity (~5-9 d).
• RCS permeable molecules (many low MWt) clear within hours once dissolved
in the vitreous solution.
Barriers for intravitreal drug delivery
Barrier Type of barrier Significance Molecule/particle-
(features) associated phenomena

Vitreous humour Physical (steric and Fiilled with viscous gel that limits
dynamic) particle diffusion
Demonstates different characteristics
in different regions
Flow mechanisms affect molcular
distribution around gel
Composition varies with age
Molecules and particles traverse more
freely around liquified model
Cationic particles aggregate with
negatively charged collagens and
glycosaminoglycans
Larger particle size or presence in the
mid-vitreous region results in sloe mobility
through gel

Inner limiting Physical (steric) Difficult to traverse for most Certain particulate carriers and
membrane molecules macromolecules have difficulty
Ability of molecules to associate with associating with and traversing this barrier
barrier is also linked with improved
therapeutic efficacy

Anterior clearance Biochemical Provides sink conditions ready for All substrates in this vitreous may be
pathway (elimination pathway) clearance of molecules from the cleared via this mechanism
vitreous Pathway most important for molecules
unable to be taken up by the retina
Favoured clearance pathway for
substrates present in anterior vitreous
regions

Blood-retinal barrier Biochemical Equipped with efflux pumps poised to Small lipophilic molecules most commonly
(elimination pathway) clear substrates from the retina cleared via this route
Substartes for efflux pumps are also prone
to elimination via this pathway

Expert Opin. Drug Deliv. (2014) 11(10)

Steroid implants
Much ongoing effort to decrease or avoid intravitreal injections. Ozurdex
is an implant for the posterior segment

• Ozurdex (Allergan), a biodegradable polymer delivery system that contains

dexamethasone, is the first FDA approved intravitreal steroid implant for the
treatment of macular edema secondary to retinal vein occlusion.
• Other implants available: Medidur, Retisert
• Triamcinolone acetonide long administered as a suspension
• Steroids are the main class of implants
The macula

Fundus photographs of the right eye (left

image) and the left eye (right image) seen from
the front so that the left in each image is to the
person’s right. The gaze is into the camera, so
in each picture the macula is in the centre of the
image, and the optic disk is located towards the
nose.
Wiki

High acuity visual function; central vision

Age related macular degeneration.

Normal Dry Macular Wet Macular

Macula Degeneration Degeneration

• Back of the eye is highly vascularise

• As we age, the back of the eye is
susceptible to vascular disease
• AMD is the main cause of blindness
in the elderly
neovascular lesions are well delineated
Neovascular (wet) AMD
and do not involve the centre of the fovea.
Photodynamic therapy (PDT), which uses the
intravenous laser-activated photosensitizing
Fovea
Blood vessels
Choroidal neovascularization
Figure 1 | Neovascular AMD. Neovascular
AMD involves the growth of abnormal new
blood vessels into the sub-retinal space and
retinal pigment epithelium. These weak vessels
Angiogenesis is implicated in many diseases
leak blood or serum, causing damage to the
macula, which is crucial for fine vision. Adapted
from REF. 3 © (2 0 0 5 ) Macmillan Magazines Ltd.
Disease
Diabetic retinopathy,
NATURE REVIEWS | DRUG
ageDISCOVERY
related macular degeneration (wet)
Rheumatoid arthritis
Atherosclerotic plaques
Endometreiosis
Crohn’s disease
Psoriasis
Uterine fibroids
Benign prostatic hypertrophy
Cancer
G1 κ isotype monoclonal antibody fragment for three consecutive doses, followed by a
designed for intraocular use7 –9 . It binds to
human VEGF and prevents interaction with
dose administered once every 3 months9 . The
primary efficacy endpoint was mean change
its receptors VEGFR1 and VEGFR2 on the in visual acuity at 1 2 months compared with
surface of endothelial cells, thereby reducing baseline9 . After an initial increase in visual
Neovascular AMD involves the growth of abnormal new
endothelial cell proliferation, vascular leakageacuity (following monthly dosing), on average
blood vessels into the sub-retinal space and retinal pigment
and the formation of new blood vessels7 –9 . patients dosed once every three months with
ranibizumab lost visual acuity, returning to
epithelium. These weak vessels leak blood or serum,
Clinical data baseline at month 1 2; almost all ranibizumab-
The efficacy and safety of ranibizumab were treated patients (9 0%) maintained their visual
causing damage to the macula, which is crucial for fine
evaluated in three randomized, double- acuity at month 1 29 .
vision.
masked, sham- or active-controlled trials On the basis of these trials, ranibizumab
Adapted from (2005) Macmillan Magazines Ltd.
involving ~1 ,3 00 patients with neovascular (0.5 mg) is recommended to be administered
AMD9 . by intravitreal injection once a month9 .
Current 1st line treatment is intravitreal injection of
In the first trial, patients with minimally Although less effective, treatment may be
ranibizumab. Other treatments in the past include
classic or occult (without classic) choroidal
neovascular (CNV) lesions received monthly
reduced to one injection every 3 months after
the first 4 injections if monthly injections
laser photodynamic therapy to occlude blood vessels.
ranibizumab 0.3 mg or 0.5 mg intravitreal are not feasible9 .
injections or monthly sham injections for 24
months9 . In the second trial, patients with Indications
predominantly classic CNV lesions received
either monthly ranibizumab (0.3 mg or 0.5 mg
Ranibizumab is approved by the FDA for
the treatment of patients with neovascular
intravitreal injections) and sham PDT, or sham (wet) AMD9 . ▶
Symptoms
Loss of vision
VOLUME 5 | O CTOBER 2006 | 815
Pain and immobility from destroyed cartilage
Chest pain, dyspnoea
Abdominal pain from intraperitoneal bleeding
Intestinal bleeding
Persistent severe ithing
Vaginal bleeding, abdominal pain
Urinary retention
Bleeding, thrombosis, anaemia, abdominal
ascites, bone pain, seizures
Judah Folkman, Nature Reviews Drug Discovery, 2007, 6, 273
Ranibizumab is a Fab

Joshua S. Klein et al. PNAS 2009;106:7385-7390

Ranibizumab Bevacizumab
(Fab) (monoclonal antibody)
Labelled to treat AMD
 Ranibizumab (Fab) vs Bevacizumab (IgG)
• Ranibizumab (Lucentis) is a Fab which also binds to
VEGF. Formulated for ocular injection~monthly to treat
AmD. Presented in a glass vial and is formulated for
intravitreal (IVT) injection to treat wet AMD
• Bevacizumab (Avastin) is much cheaper per dose. But is
not formulated for ocular injection. Presented in a vial
containing 16 mL at 25 mg/mL concentration of antibody
(total 400 mg/vial). Ranibizumab is dosed monthly as
needed.
• Until recently ranibizumab was presented as a solution in
a glass vial. Withdrawal of the liquid solution had to pass
a filter into the syringe. Ranibizumab is now also
presented in a ready to use glass syringe.
• During the ranibizumab trials it was found that bevacizumab
could also be used to treat wet AMD.
• Some hospital pharmacies fractionate bevacizumab into
plastic syringes for intravitreal injection. This means that
each bevacizumab IVT dose (0.05 mL; 50 μL) is much
cheaper than a ranibizumab IVT dose.
• What are the risks for using bevacizumab?
How was the dose of Bevacizumab determined?
• Bevacizumab was developed before ranibizumab to treat colon cancer. Both
medicines were developed by the same company.
• The antibody is formulated for infusion at 25 mg/mL
• When ranibizumab was being clinically trailed, an ophthalmologist thought to try
bevacizumab in an AMD patient.
• The dose was determined simply by removing an injection volume from the
bevacizumab vial.
• 0.05 mL × 25 mg/mL = 1.25 mg dose. The ranibizumab dose is 0.5 mg in 0.05 mL.
The bevacizumab dose is less on a molar basis than the dose for ranibizumab.
• Bevacizumab is 3× higher MWt than ranibizumab. The IgG is slower to diffuse
from the eye. IVT half-life is ~1-2 d longer.
• Subsequently IVAN and CATT trials have
shown that bevacizumab and
ranibizumab are clinically equivalent.
• However bevacizumab IVT dosage form
preparation is not controlled, which is a
problem.
 Aflibercept (VEGF-trap, Eylea)
Giving more…to exploit a therapeutic tail….(potency helps)
tap the
bubbles
• Aflibercept (Eylea) is a Fc-Fusion protein (~110 kDa).
• Initially presented in a glass vial. Now presented in a ready to use glass syringe
(single use).
• The dose of aflibercept is 2 mg (0.05 mL); monthly first 3 months and then every 2
months. This is the key point: Aflibercept is dosed less frequently. No one wants
IVT injections.
• Concentration of aflibercept is 40 mg/mL rather than 10 mg/mL for ranibizumab. So
aflibercept dose is 4× by mass compared to ranibizumab (0.5 mg dose).
• Aflibercept is dosed ~2× by
Figure 6
moles compared to
ranibizumab
e edge
opper

nd the

d to the
r being
~150 kDa ~50 kDa ~110 kDa

IVT injection
Figure 70.05 mL