U.S. patent application number 15/308220 was filed with the patent office on 2017-03-02 for non-invasive agent applicator.
The applicant listed for this patent is MuPharma Pty Ltd. Invention is credited to Sean Michael LANGELIER, Harry UNGER, Mark UNGER.
Application Number | 20170056637 15/308220 |
Document ID | / |
Family ID | 54391863 |
Filed Date | 2017-03-02 |
United States Patent
Application |
20170056637 |
Kind Code |
A1 |
UNGER; Harry ; et
al. |
March 2, 2017 |
NON-INVASIVE AGENT APPLICATOR
Abstract
There is disclosed systems and methods for non-invasive delivery
of an agent to biological tissues. Delivery of the agent to the
tissues can be by one or more modalities. In some embodiments the
systems and methods use agent carrier body including a tissue
contacting surface for non-invasively engaging tissues under
treatment. The tissue contacting surface can be at least partly
defined by a plurality of protrusions that are in fluid
communication with one or more reservoirs forming part of the agent
carrier body. The protrusions may extend outward from an inside of
a void and terminate at said tissue contacting surface.
Inventors: |
UNGER; Harry; (Victoria,
AU) ; LANGELIER; Sean Michael; (Victoria, AU)
; UNGER; Mark; (Victoria, AU) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
MuPharma Pty Ltd |
Victoria |
|
AU |
|
|
Family ID: |
54391863 |
Appl. No.: |
15/308220 |
Filed: |
May 6, 2015 |
PCT Filed: |
May 6, 2015 |
PCT NO: |
PCT/AU2015/050218 |
371 Date: |
November 1, 2016 |
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61M 2037/0023 20130101;
A61M 2037/003 20130101; A61N 1/0448 20130101; A61M 37/00 20130101;
A61M 2037/0007 20130101; A61N 1/303 20130101; A61M 37/0092
20130101 |
International
Class: |
A61M 37/00 20060101
A61M037/00; A61N 1/30 20060101 A61N001/30; A61N 1/04 20060101
A61N001/04 |
Foreign Application Data
Date |
Code |
Application Number |
May 6, 2014 |
AU |
PCT/AU2014/050027 |
Nov 12, 2014 |
AU |
2014904549 |
Claims
1.-52. (canceled)
53. A method of dispensing an agent from an agent carrier, the
method comprising: holding the agent within an agent carrier, said
agent carrier including a solid agent carrier body that terminates
at a tissue contacting surface in a plurality of protrusions,
wherein holding the agent within the agent carrier includes holding
at least some agent within the agent carrier body; engaging the
tissue contacting surface of the agent carrier body with a surface
of a biological tissue, wherein the plurality of protrusions do not
penetrate any layer of the tissue; and dispensing agent from the
agent carrier to the tissue surface by applying ultrasonic waves to
cause transportation of the agent through the agent carrier body to
the tissue surface wherein the method further includes applying the
ultrasonic waves to the tissue via the agent carrier to enhance or
permit penetration of the agent into the tissue.
54.-57. (canceled)
58. The method of claim 53, wherein the ultrasonic waves are
selected to deliver a chosen amount of agent to a chosen depth
within the tissue.
59. The method of claim 53, wherein operational parameters of the
agent carrier are used to enhance or cause delivery of said agent
to a selected depth within the tissue, and the operational
parameters are selected from one or more of: application pressure,
ultrasonic frequency, ultrasonic power level, ultrasonic waveform,
ultrasonic application duration, ultrasonic application duty cycle,
ultrasound direction.
60. The method of claim 53, wherein the plurality of protrusions is
in fluid communication with one or more agent reservoirs forming
part of the agent carrier body.
61. The method of claim 60, wherein each agent reservoir comprises
a void formed within the agent carrier body.
62. The method of claim 53, wherein the plurality of protrusions
extend outward from an inside of a void and terminate at the tissue
contacting surface.
63. The method of claim 62, wherein the void is formed by a
peripheral structure, wherein at least part of said peripheral
structure terminates at the tissue contacting surface.
64. The method of claim 63, wherein the peripheral structure
terminates in a common plane with the plurality of protrusions.
65. The method of claim 62, wherein at least some of the plurality
of protrusions extend outward from the void beyond the peripheral
structure.
66. The method of claim 65, wherein at least some of the plurality
of protrusions terminate in a plane and the peripheral structure
terminates short of the plane such that the protrusions extend
beyond the peripheral structure.
67. The method of claim 53, wherein the agent carrier body further
includes at least one micro channel extending at least partially
through the agent carrier body to the tissue contacting surface
enabling transportation of the agent to the tissue surface.
68. The method of claim 53, wherein the agent carrier body includes
a stack of layers including: a tissue-contacting layer which
includes the tissue contacting surface; and at least one other
layer; wherein a plurality of the layers have holes formed therein
to enable the agent to be transported from one said layer to the
next.
69. The method of claim 53, wherein the agent carrier body is able
to conduct a transmission stimulus to transport the agent within
the agent carrier body and to the tissue surface.
70. The method of claim 53, wherein the agent carrier is a
component of an applicator device.
71. The method of claim 70, wherein the agent carrier or agent
carrier body is coupled directly or indirectly to a handle unit of
the applicator device to facilitate handheld control of the
applicator device.
72. The method of claim 71, wherein the handle unit includes a
generator of ultrasound energy to generate the ultrasonic waves,
and the ultrasonic waves are transmitted to the agent carrier.
73. The method of claim 53, wherein the agent carrier is a
consumable applicator tip adapted for single use.
Description
FIELD OF THE INVENTION
[0001] The present invention relates to the application of an agent
to a target site. In a preferred form, the invention uses
ultrasonic energy to transport an agent contained within an agent
carrier body having a plurality of micro-scale structures within it
to the target site non-invasively. In this preferred form, at the
target site, penetration of the agent into the target site is
enabled or enhanced through sonophoretic mechanisms.
BACKGROUND OF THE INVENTION
[0002] WO 2007/143796 discloses a method of delivering a molecule
and/or particle to a target site using a device that includes
generating ultrasound for enhancing the penetration of a molecule
and/or particle into the target tissue.
[0003] The device of WO 2007/143796 includes an electro-conductive
polymeric gel material that is loaded with a molecule and/or
particle such as a pharmaceutical or ink etc. Application of an
electric field to the electro conductive polymer gel releases
substantially bound molecules or particles within the polymer gel
matrix and, ultimately, such molecules or particles are transported
through such polymer gel by ultrasound to the target tissue
surface. At the target tissue surface, penetration of the molecule
and/or particle into the tissue is enabled or enhanced through
sonophoretic mechanisms.
[0004] One difficulty relating to this delivery mechanism is that
the structure of the polymer gel can degrade over time, for example
due to loss of moisture, which results in reduced propagation of
the molecule and/or particle by ultrasound. Additionally, gels are
poor transmitters of ultrasound reducing the efficacy of the
sonophoretic process. Furthermore, it can be time consuming and
non-trivial to properly load an applicator with small volumes of
the molecule and/or particle loaded polymeric gel.
[0005] In light of these problems, an improved device and mechanism
for delivering an agent to a target tissue is sought.
[0006] Reference to any prior art in the specification is not, and
should not be taken as, an acknowledgment or any form of suggestion
that this prior art forms part of the common general knowledge in
Australia or any other jurisdiction, or that this prior art could
reasonably be expected to be ascertained, understood and regarded
as relevant, or combined with other prior art by a person skilled
in the art.
SUMMARY OF THE INVENTION
[0007] In one aspect of the invention there is provided an agent
carrier for non-invasive delivery of an agent to biological
tissues. Delivery of the agent to the tissues can be by one or more
modalities. The modality of delivery can be characterised by a
transportation stimulus or stimuli that causes transportation of
the agent through the agent carrier. In a preferred form, the
transportation stimulus also enhances or permits penetration of the
agent into the tissue. Preferred forms of the invention use only
ultrasound as the transportation stimulus.
[0008] In preferred forms the agent carrier includes an agent
carrier body configured to retain agent within the agent carrier
body. The agent carrier body has a tissue contacting surface for
engaging tissues under treatment, wherein application of the
transportation stimulus causes transportation of the agent through
the agent carrier body to the tissue contacting surface.
[0009] The agent to be delivered can include one or more molecules
or particles or one or more molecules and particles in any
combination. The agent can be a fluid or can be carried in a fluid
medium, e.g. by being dissolved, suspended or dispersed in a fluid
medium, such as water, oil, an emulsion, a gel or the like. To give
but a few examples, the agent can include, proteins, vaccines,
nucleic acids, monoclonal antibodies, nanoparticles or molecular
machines. In preferred embodiments the agent is a pharmaceutical or
pharmaceutical composition. The pharmaceutical or one or more
active pharmaceutical components of a pharmaceutical composition
may be, without limit, any one of: a synthesised compound; a
naturally occurring compound; or a biopharmaceutical. The purpose
of the delivery of the pharmaceutical or pharmaceutical composition
to the biological tissues can be for any desired clinical reason
including: treating, curing or mitigating a disease, condition, or
disorder; attenuating, ameliorating, or eliminating one or more
symptoms of a particular disease, condition, or disorder;
preventing or delaying the onset of one or more a disease,
condition, or disorder or a symptom thereof; diagnosing a disease,
condition, or disorder, or any agent intended to affect the
structure or any function of the body. In other embodiments the
agent can be an agent used for cosmetic purposes such as for
cleansing, beautifying, promoting attractiveness, or altering the
appearance of the body. The agent could also be a marker agent used
for creating human or machine perceptible makings, e.g. ink or
other. Other types of agents may also be used.
[0010] The transportation stimulus is the driving force for moving
the agent through the agent carrier to the tissue-contacting
surface, and may enhance and/or permit the penetration of the agent
from the tissue-contacting surface into the tissue.
[0011] In some embodiments the tissue can be any human or animal
biological tissue, including mucous membranes, skin and teeth.
Preferably the tissue is ocular tissue or oral mucosa. In some
embodiments the tissue is any plant tissue.
[0012] In an aspect there is provided an agent carrier body
including a tissue contacting surface for non-invasively engaging
tissues under treatment, the tissue contacting surface being at
least partly defined by a plurality of protrusions. The protrusions
may be in fluid communication with one or more reservoirs forming
part of the agent carrier body. Each agent reservoir may comprise a
void formed within the agent carrier body. The protrusions may
extend outward from an inside of a void and terminate at said
tissue contacting surface. The void may be formed by a peripheral
structure, where at least part of said peripheral structure may
terminate at the tissue contacting surface.
[0013] In some embodiments the peripheral structure terminates in a
common plane with the protrusions. In others at least some of said
protrusions defining the tissue contacting surface extend outward
from the void beyond the peripheral structure. In some embodiments,
the protrusions may terminate in a plane and the peripheral
structure may terminate short of the plane such that the
protrusions extend beyond the peripheral structure.
[0014] The agent carrier body may further include one or a
multiplicity of micro channels extending at least partially through
the agent carrier body to the tissue contacting surface enabling
transportation of the agent to a tissue surface. The micro channels
may extend through the agent carrier body to fluidly connect to an
agent reservoir.
[0015] The agent carrier body of these aspects can include a stack
of layers including a tissue-contacting layer, which includes the
tissue-contacting surface, and at least one other layer. The
tissue-contacting layer preferably has holes extending through it
to define at least a portion of the micro channels in the body. In
some embodiments a plurality of layers have holes formed therein to
enable agent to be transported from one layer to the next.
Preferably holes formed in one layer of the plurality of layers are
aligned with holes in an adjacent layer so that a plurality of
holes in a plurality of layers cooperate to form the micro
channels. In some embodiments the holes decrease in diameter and
increase in number from the first layer to the tissue-containing
layer. The micro channels may have a varying cross-section along
their length.
[0016] In some embodiments a reservoir for storing agent is at
least partly (and optionally fully) formed in the agent carrier
body.
[0017] The micro channels and/or agent reservoir(s) and/or
protrusions are defined by internal exposed surfaces within the
agent carrier body. Preferably these internal exposed surfaces are
configured to possess predetermined hydrophilic, hydrophobic,
and/or electro-conductive properties. In this case, at least part
of the internal exposed surfaces could be modified or treated to
configure their hydrophilic, hydrophobic, and/or electro-conductive
properties.
[0018] The agent carrier body may include a port to enable loading
of the agent carrier body and/or reservoir(s) with agent.
[0019] The agent carrier body can further include a stimulus
generator, operable to generate transportation stimulus. The
stimulus generator preferably includes an ultrasonic
transducer.
[0020] In some embodiments, the agent carrier preferably includes a
housing configured to mechanically support an agent carrier body,
of any type described herein. The housing can include a mounting
arrangement configured to be mounted to an applicator device. The
mounting arrangement preferably enables selective attachment and
removal of the agent carrier to and from the applicator device,
such that the agent carrier can be replaced.
[0021] The agent carrier housing also may include a recess or other
mounting formation formed therein for receiving the agent carrier
body. In some embodiments the agent carrier body can be selectively
attached to, or removed from, the recess or mounting formation such
that the agent carrier body can be replaced.
[0022] The agent carrier can include a port to enable loading of
the agent carrier body and/or reservoir(s) with agent.
[0023] The agent carrier can further include a stimulus generator,
operable to generate a transportation stimulus. The stimulus
generator preferably includes an ultrasonic transducer. At least
part of the stimulus generator can be formed as part of the agent
carrier body.
[0024] In a preferred embodiment the agent carrier or agent carrier
body is a consumable applicator tip adapted for one-time use as
part of an applicator device.
[0025] In another aspect of the invention there is provided a
non-invasive applicator device comprising an agent carrier and/or
an agent carrier body as described herein.
[0026] The agent carrier or agent carrier body can be coupled
directly or indirectly to a handle unit to facilitate hand held
operation of the applicator device. The handle unit preferably
includes a mounting arrangement configured to cooperate with a
complementary mounting arrangement of the agent carrier and/or
agent carrier body.
[0027] The handle unit may include an ultrasonic generator to
generate ultrasonic waves that are transmitted to the attached
agent carrier and/or agent carrier body.
[0028] Preferably the agent carrier is a consumable applicator tip
adapted for one-time use.
[0029] The agent carrier preferably includes an agent carrier body
including a tissue contacting surface for non-invasively engaging
tissues under treatment, the tissue contacting surface being at
least partly defined by a plurality of protrusions.
[0030] The agent carrier may include one or more agent reservoirs
for carrying said agent, wherein said protrusions are in fluid
communication with one or more reservoirs forming part of the agent
carrier. Each agent reservoir may at partly (or wholly) comprise a
void formed within the agent carrier body.
[0031] Also disclosed herein is a method of dispensing an agent
from an agent carrier described herein. The method comprises
holding the agent within an agent carrier, said agent carrier
including a solid agent carrier body. The method can further
comprise engaging a tissue contacting surface of the agent carrier
body with a tissue surface of the biological tissue. The method can
further comprise dispensing agent from the agent carrier to the
tissue surface by applying at least one transportation stimulus to
cause transportation of the agent through the agent carrier body to
the tissue surface.
[0032] In some forms the method further includes applying the
transportation stimulus to the tissue via the agent carrier to
enhance or permit penetration of the agent into the biological
tissue.
[0033] Holding the agent within an agent carrier can include
holding at least some agent within the carrier body.
[0034] In preferred embodiments the agent carrier body terminates
at its tissue contacting surface in a plurality of protrusions. In
this case, engaging a tissue contacting surface of the agent
carrier body with a tissue surface of the biological tissue,
includes engaging the tissue surface of the biological tissue with
the protrusions of the agent carrier body. Such engagement
preferably does not involve mechanically penetrating any layer of
biological tissue with the protrusions.
[0035] In another aspect of the invention there is provided a
method of dispensing an agent from an agent carrier, an agent
carrier body, or an applicator device as described previously, the
method including: contacting the tissue-contact surface of the
agent carrier with a tissue surface; and dispensing agent from the
agent carrier body to the tissue surface and into the target
tissue.
[0036] In some embodiments of any of the above methods the step of
dispensing the agent includes generating ultrasonic waves for agent
transport to the tissue contact surface. Even more preferably the
method includes propagating ultrasonic waves through the agent
carrier to the tissue. This aids the delivery of the agent through
the tissues via sonophoresis.
[0037] In some embodiments of any of the above methods the step of
dispensing the agent can include applying an electrical voltage
across the agent carrier body to cause agent transport to the
tissue contact surface. The electric voltage can also provide for
the transport of agent into and through the tissue via
iontophoresis. Even more preferably the method includes propagating
an electric current through the agent carrier to the tissue.
[0038] In yet another aspect of the present invention there is
provided a method of dispensing an agent from an agent carrier, an
agent carrier body or an agent applicator device as described
herein. The method including, contacting the tissue contacting
surface of the agent carrier body with a tissue surface; and
dispensing agent from the agent carrier to the tissue surface. The
step of dispensing the agent preferably includes generating
ultrasonic waves to cause or facilitate agent transportation to the
tissue-contacting surface. The method can include the application
of ultrasonic waves to the tissue surface to non-invasively cause
or facilitate agent penetration of the agent into and through the
tissue via sonophoresis.
[0039] The method further includes propagating ultrasonic waves
through the agent carrier or agent carrier body to the tissue.
[0040] In another aspect the present invention provides a method of
loading agent into any one of an agent carrier, agent carrier body,
an agent applicator device as described herein. The method
includes, exposing the agent carrier body to the agent to enable
filling either of both of, a reservoir or micro channels in fluid
communication with said reservoir, with said agent.
[0041] The method can include applying a negative pressure to the
agent carrier or agent carrier body to draw agent into the micro
channels or agent reservoirs in fluid communication with the micro
channels. The method can include applying a positive pressure to
the agent carrier or agent carrier body to inject the agent into
the micro channels or agent reservoirs in fluid communication with
the micro channels.
[0042] The step of filling the micro channels or agent reservoirs
with the agent can include the application of ultrasonic energy to
the agent carrier or agent carrier body to draw agent into the
agent carrier or agent carrier body.
[0043] In some embodiments, the voids and/or micro channels in the
agent carrier body are loaded by virtue of capillary forces when
the agent carrier is in contact with the agent.
[0044] In another aspect the present invention provides a method of
delivering an agent to a biological tissue, including: applying
said agent using an agent carrier, agent carrier body or applicator
of any one of the aspects or embodiments described herein, wherein
ultrasound is the transportation stimulus; and configuring the
operational parameters of the application to enhance or cause
delivery of said agent to a selected depth within such tissue. The
operational parameters configured may include (but are not limited
to) any one or more of: [0045] Application pressure; [0046]
Ultrasonic frequency; [0047] Ultrasonic power level; [0048]
Ultrasonic waveform; [0049] Ultrasonic application duration; [0050]
Ultrasonic application duty cycle; and [0051] Ultrasound
direction.
[0052] Preferably the operational parameters are selected to
deliver a chosen amount of agent to a chosen depth within
tissue.
[0053] The method may involve delivering the agent to or beyond any
one or more of the following tissues or tissue layers: [0054]
Mucous Membrane; [0055] Epithelium [0056] Sub-epithelium [0057]
Mucosa; [0058] Sub-mucosa [0059] Mucous membrane vasculature [0060]
Cornea; [0061] Corneal epithelium [0062] Bowman's membrane [0063]
Corneal stroma [0064] Corneal Endothelium [0065] Conjunctiva;
[0066] Tenon's Fascia; [0067] Episclera: [0068] Sclera; [0069]
Choroid; [0070] Choriocapillaris; [0071] Bruch's membrane; [0072]
Retinal Pigment Epithelium; [0073] Neural retina; [0074] Retinal
blood vessels; [0075] Internal Limiting Membrane: and [0076]
Vitreous.
[0077] As used herein, except where the context requires otherwise,
the term "comprise" and variations of such term, such as
"comprising", "comprises" and "comprised", are not intended to
exclude further things, additives, components, integers or steps.
Also, as used herein, except where there is express wording to the
contrary, specifying anything after the words `include` or `for
example` or similar expressions does not limit what else is
included.
BRIEF DESCRIPTION OF THE DRAWINGS
[0078] Further aspects of the present invention and further
embodiments of the aspects described in the preceding paragraphs
will become apparent from the following description, given by way
of example and with reference to the accompanying drawings. In the
drawings:
[0079] FIG. 1A shows a schematic cross-sectional block diagram of
an applicator device according to one embodiment, being applied to
a tissue surface and provides an illustration of the overall
components of one exemplary applicator device.
[0080] FIG. 1B shows a more detailed cross sectional view of the
agent carrier body of the embodiment shown in FIG. 1A and
previously described in the Applicant's Australian patent
application 2013901606.
[0081] FIG. 1C shows a similar agent carrier body to that of FIG.
1B that includes an ultrasonic transducer.
[0082] FIG. 2 provides a cross sectional block diagram of an
embodiment of a handle assembly of the applicator device and its
basic component parts.
[0083] FIG. 3 is a cross sectional view through an agent carrier
that takes the form of a single use applicator tip including an
agent carrier body of the type previously described in the
Applicant's Australian patent application 2013901606. As will be
appreciated, any agent carrier body as generally described herein,
and as exemplified in any one of FIGS. 8A to 10 or 23 to 30A may be
used as an alternative.
[0084] FIGS. 4A, 4B, and 4C provide illustrations of various
embodiments of a single layer agent carrier body with different
micro-channel, and or reservoir arrangements previously described
in the Applicant's Australian patent application 2013901606.
[0085] FIG. 4D provides an illustration of an embodiment of a first
surface and a tissue contact surface of a single layer agent
carrier body.
[0086] FIGS. 4E, 4F, 4G, and 4H provide illustrations of various
embodiments of a multiple layer agent carrier body with different
micro-channel and reservoir arrangements.
[0087] FIG. 4I provides an illustration of the embodiment shown in
FIG. 4H of a first surface and a second surface of a first layer of
the agent carrier body, and a first surface and a tissue contact
surface of the second layer of the agent carrier body.
[0088] FIG. 4J provides illustrations of further example
embodiments of agent reservoir contacting layer of an agent carrier
body that can store additional agent and replenish the
micro-channels as they are depleted of agent during the course of
usage.
[0089] FIGS. 5A, 5B and 5C provide illustrations of various
embodiments of the agent carrier body each of which has a
differently configured surface contact layer.
[0090] FIG. 5D provides an illustration of two exemplary types of
micro-protrusions that extend from the agent carriers shown in
FIGS. 5B and 5C.
[0091] FIG. 6 provides an illustration of an embodiment of an agent
carrier body having a stacked layer arrangement and an agent
filling port.
[0092] FIG. 7A and 7B provide illustrations of embodiments of the
holes, and the channels defined by the holes, in an agent carrier
body that has a stacked layer structure.
[0093] FIGS. 7C to 7E provides magnified images of the holes and
micro-channels created by the micro-manufacturing process.
[0094] FIGS. 8A and 8B are schematic representations of an
alternative embodiment of an agent carrier body, according to an
aspect of the present invention, and respectively illustrate plan
and perspective views thereof.
[0095] FIGS. 8C and 8D are schematic representations of an
alternative embodiment of an agent carrier body layer having micro
channels formed through it, and respectively illustrate plan and
perspective views thereof.
[0096] FIGS. 8E and 8F are schematic representations of an
alternative embodiment of an agent carrier body layer having a
reservoir formed therein, and respectively illustrate plan and
perspective views thereof.
[0097] FIGS. 8G and 8H are schematic representations of an agent
carrier body formed by the agent carrier body layer of FIGS. 8E and
8F stacked with the agent carrier body layer of FIGS. 8C and 8D,
and respectively illustrate the agent carrier body in unfilled and
filled configurations
[0098] FIG. 9A and is an electron micrograph of a portion of an
agent carrier body of any one of FIGS. 8A to 8H.
[0099] FIG. 9B and is an electron micrograph of a single protrusion
of an agent carrier body of any one of FIGS. 8A to 8H.
[0100] FIG. 10 illustrates a series of four mask designs, each
suitable for forming a respective agent carrier body (or layer
thereof) in embodiments of the present invention.
[0101] FIGS. 11A to 11C provide an illustration of a various
embodiments in which an agent reservoir is provided in an agent
carrier in a location external to the agent carrier body. As will
be appreciated any agent carrier body as generally described
herein, and as exemplified in any one of FIGS. 8A to 10 or 23 to
30A may be used with such an embodiment.
[0102] FIG. 12A to 12E illustrate steps in various embodiments of
charging or recharging methods that can be used in embodiments of
the present invention. As will be appreciated any agent carrier
body as generally described herein, and as exemplified in any one
of FIGS. 8A to 10 or 23 to 30A may be used with such an
embodiment.
[0103] FIGS. 13A and 13B illustrate an exploded view and cross
sectional view through agent carrier according to one embodiment.
The agent carrier can be used to carry any agent carrier body as
generally described herein, and as exemplified in any one of FIGS.
8A to 10 or 23 to 30A.
[0104] FIG. 14 shows plots for the evaluation of the uptake of
FPV-HIV-GFP vaccine 24 hours post lip delivery, illustrating I-Ad
APC MHC-Il cells containing the fluorescent GFP antigen of the
vaccine detected in the top right hand quadrant indicated by the
arrow. Each dot represents a single cell.
[0105] FIG. 15 illustrates plots for the evaluation of recruitment
of antigen uptake by different dendritic cell subsets to the
respective draining lymph nodes 24 hours post lip delivery. The
proportion of dendritic cells, identified as being MHC-11+, and
either CD11b+ (left two columns) or CD11c+ (right two columns) are
indicated in the top right hand quadrant (refer to arrows).
[0106] FIG. 16 illustrates plots for the evaluation of the uptake
of FPV-HIV-GFP vaccine 24 hours post lip delivery in cervical,
mediastinal and mesenteric nodes I-Ad APC MHC-II cells containing
the fluorescent GFP antigen of the vaccine are detected in the top
right hand quadrant indicated by the arrow.
[0107] FIG. 17 contains photographs sharing the following phases of
the experiments performed. The phases illustrated include: Loading
the microchips with the agent to be administered (top left),
Ultrasonic system settings (top right) and lip delivery to the mice
(bottom photos).
[0108] FIG. 18 illustrates plots enabling the evaluation of the
magnitude of HIV-specific splenic CD8 T cells using IFN-.gamma.
intracellular staining. The FACS data were analyzed using Cell
Quest Pro or FlowJo analysis. The box indicates the percentage of
HIV-specific splenic CD8 T cells expressing IFN-.gamma. following
Lip/i.m. (top 3 mice), i.n./i.m. (middle 2 mice) and booster only
(bottom 3 mice) vaccinations.
[0109] FIG. 19 illustrates plots enabling evaluation of
HIV-specific splenic CD8 T cells using tetramer staining. The FACS
data were analysed using Cell Quest Pro or FlowJo analysis. The box
indicates the percentage of HIV-specific splenic CD8 T cells
following different routes of vaccine delivery. Lip/i.m. (top three
mice), i.n./i.m. (middle two mice) and booster only (bottom two
mice).
[0110] FIG. 20 shows plots illustrating HIV-specific splenic CD8 T
cell responses observed with the four different microchips of FIG.
10. Data represent HIV-specific CD8 T cell numbers measured by
tetramer staining (data represent one mouse from each group).
[0111] FIG. 21 illustrates plots enabling evaluation of
HIV-specific splenic CD8 T cell responses using tetramer staining.
The FACS data were analyzed using Cell quest Pro software. Plots
represent three animals per group microchip 1 (top) & 2
(middle) prime-boost immunization data compared to oral delivery
(bottom). The upper right quadrants (arrows) indicate the % of
HIV-specific CD8 T cells observed following each vaccine
strategy.
[0112] FIG. 22 illustrates plots enabling evaluation of the
magnitude of HIV-specific CD8 T cell responses using IFN-.gamma.
intra cellular cytokine staining. The FACS data were analysed using
Cell quest Pro software. Plots represent three animals per group
microchip 1 (top) & 2 (middle) prime-boost immunization data
compared to oral delivery (bottom). The upper right quadrants (red
arrows) indicate the % of HIV-specific CD8 T cells expressing
IFN-.gamma..
[0113] FIG. 23 illustrates a series of mask designs for creation of
various agent carrier bodies (or layers thereof).
[0114] FIGS. 24 to 28 illustrate a series of electron micrographs
of agent carrier bodies and regions thereof of various
embodiments.
[0115] FIGS. 29 to 30A illustrate diagrammatically two hybrid agent
carrier bodies according to an embodiment of the present
invention.
[0116] FIG. 31A to 31D illustrates plots of the displacement (nm)
and velocity (m/s) during operation of five types of applicator
useable with embodiments of the present invention having different
tips.
[0117] FIG. 32 illustrates the tip displacement of the MP4 and AMO
1 applicators when driven at selected frequencies over a range of
drive voltages.
DETAILED DESCRIPTION OF THE EMBODIMENTS
[0118] In the following description, for the purposes of
explanation, numerous specific details are set forth in order to
provide a thorough understanding of the present invention. It will
be apparent, however, that the present invention may be practiced
without these specific details. In other instances, well-known
structures and devices are shown in block diagram form in order to
avoid unnecessary obscuring.
[0119] This description follows the following outline: [0120] 1.
Overview [0121] 2. General principles and Micro-Channel Embodiments
[0122] 3 Protrusion-based embodiments [0123] 4 Hybrid and
alternative embodiments [0124] 5. Loading and use examples [0125]
6. Trial results [0126] 7. References
1. Overview
Background to the Present Embodiments
[0127] The delivery of drugs, including macromolecules larger than
approximately 500 Daltons and hydrophilic drugs, to the body
without using hypodermic injections, ingestion or surgery has long
been a desired goal in medicine.
[0128] A myriad of drug delivery devices using a variety of
technologies have been developed to achieve this ("Drug delivery
devices"), however, these have been unable to non-invasively
deliver to the body a large range of drugs in a safe, practical,
predicable and effective way. Historically, the transdermal route,
has been the primary focus of non-invasive drug delivery
applications.
[0129] The advantages of delivering drugs to the body without
ingestion, includes bypassing the degradation of drugs by the acid
and or alkaline regions of the gastrointestinal tract and enzymes
in the gastro-intestinal tract and avoiding their metabolism by the
liver enzymes as well as removal of the dyspeptic side effects of
drugs. Advantages of delivering macromolecules or hydrophilic drugs
to the body without hypodermic injections include decreased or
elimination of pain, local trauma and side effects, increased
patient compliance, and lowering the incidence of needle
contamination, disease transmission and needle misuse. Delivering
drugs to the body without surgery that is required to introduce
implanted devices has advantages including decreased or elimination
of pain, local trauma and side effects and elimination of any
anaesthetic risk.
[0130] Drug delivery devices may be applied to skin for both
targeted applications and as a portal for systemic drug delivery.
The primary barrier for transdermal transport of hydrophilic
molecules and/or molecules larger than approximately 500 Daltons is
the outermost layer of the epidermis, the stratum corneum, which is
typically 10-20 .mu.m in thickness. The stratum corneum is a
nonviable cell layer that is comprised of highly-crosslinked
keratinocytes embedded in a continuous matrix of skin lipids. Drug
delivery devices are needed to overcome these natural semipermeable
barriers to deliver the drugs. Drug delivery devices for the skin
commonly use microneedles and/or iontophoresis as the primary means
of delivering drugs to such tissues.
[0131] Another application site for Drug Delivery devices, less
commonly used as a portal for systemic drug delivery, is mucosal
membranes. The primary barrier for trans-mucosal transport of
hydrophilic molecules and macromolecules is the epithelial layer.
Drug delivery devices for trans-mucosal delivery commonly use nasal
sprays, inhalants and/or iontophoresis as the primary means of
delivering drugs.
[0132] The following technology (either solely or in any
combination) is presently used in Drug delivery devices:
Iontophoresis
[0133] Drug delivery devices that deliver an agent to the body
using a process known as iontophoresis operate by generating an
electric current that results from the application of electrodes
which create and maintain a potential difference between the device
and the target tissue. Ionic forms of the drug to be delivered are
transported in the electric current and thereby gain access to the
target tissue. Devices that deliver an agent to the body using
iontophoresis commonly have a continuous layer of drug containing
fluid in which the electrode within the device is bathed. The
application time tends to be long, in many cases, hours.
[0134] Agents that can be delivered to the body using iontophoresis
must be both hydrophilic and have an electrical charge.
Iontophoresis is not capable of delivering neutral molecules and/or
particles including large proteins and vaccines.
Microneedles
[0135] Microneedles are discrete protrusions that function to
pierce one or more layers of tissue. Depending on their
application, microneedles can be partly or fully hollow or solid.
Microneedles used in Drug delivery devices commonly function as: 1)
structures that can increase permeability within tissue when
combined with certain external stimuli; 2) structures incorporating
an agent that dissolves into tissue; 3) hollow conduits for
injection of agent into tissue; and/or 4) structures designed to
scrape surface tissue or expose internal tissue. Microneedles are
commonly incorporated into patches that are applied to skin either
with adhesives or are mechanically engaged. They may also contain
compounds to enhance the penetration of agent through tissue or
applied to tissue after it has been pre-treated with permeation
enhancer compounds.
Sonophoresis
[0136] Drug delivery devices that deliver an agent to the body
using a process known as sonophoresis operate through applying
ultrasound to tissue that both increases the permeability of
tissues and provides kinetic energy to the agent. The increase in
permeability of tissues through ultrasound results from a number of
phenomena including any one or more of the following: 1) cavitation
though generation and oscillation of gas bubbles; 2) thermal
effects from an increase in temperature causing induction of
convective transport; or 3) mechanical effects through occurrence
of stresses due to pressure variation induced by ultrasound. Low
frequency ultrasound, generally in the range of 20-200 kHz, but
preferably below 100 kHz, has been found to be more effective for
sonophoresis than higher frequencies of ultrasound. The prime
method of sonophoretic transport through skin requires power
sufficient to create cavitation.
[0137] Drug delivery devices that deliver an agent to the body
using sonophoresis commonly have a layer of fluid containing the
agent in which the source of the ultrasound is bathed or is placed
in close proximity. These devices also sometimes include various
kinds of microneedles where the microneedles are bathed in such
fluid. In each of the aforementioned devices, because fluids
attenuate the power of ultrasound more than solid materials, and
the volume of fluid on which the ultrasound acts is large with
respect to solid structures within or around it, the ultrasound
energy is considerably attenuated by the time the wavefront
approaches the tissue surface. This ultrasonic wavefront is
partially reflected from the tissue surface back into the fluid
layer which further disrupts the efficiency of the ultrasound
resulting in the need for more power to be applied to the fluid.
These techniques have some potential drawbacks, for example,
ultrasound applied to tissue can, depending upon the magnitude of
power, cause localised damage from cavitation and thermal effects.
The threshold for damaging tissue from ultrasonic power depends on
a variety of factors including the type of tissue, the thickness of
tissue, the health of the tissue and whether the tissue is intact.
For example, the skin is capable of tolerating more ultrasonic
power being applied to it than mucous membranes on the eye.
Furthermore, ultrasound applied to an agent may, depending upon the
magnitude of power, cause the agent, or molecules within it, to
cleave or denature or otherwise be damaged from cavitation, thermal
or mechanical effects. Agents which are known to have a low
tolerance to ultrasonic cavitation, mechanical forces or
temperatures above 40 degrees centigrade include vaccines, proteins
and other biologics.
Overview of the Embodiments
[0138] In summary, preferred embodiments of the present invention
use low frequency ultrasound at low power to transport an agent,
contained within an agent carrier body having micro-scale
structures within it, for delivering the agent non-invasively to
tissues.
[0139] As will be appreciated, ultrasound will be applied over one
or more frequency bands or over a frequency spectrum having several
bands. Preferably the band(s) correspond to a resonant frequency of
the applicator device including the agent carrier body, and
optionally one or more harmonics of the resonant frequency. In some
forms of the present invention, the ultrasound applied is of a low
frequency, between 20 kHz to 100 kHz, most preferably the frequency
of the ultrasonic energy is between 20 kHz and 40 kHz. This is
particularly preferred for use with the mucous membranes, eyes and
other delicate tissues. However, in other embodiments the
applicator device may have a resonant frequency lower than this,
and the devices describe herein may be operated with a primary
resonant frequency at the tip of the agent carrier body of around
10 kHz. In testing, applicators suitable for use with embodiments
of the present invention have been operated at frequencies in any
one or more of the following frequency bands, a band centred at or
about 10 kHz; 20 kHz, 22 kHz, 28 kHz, 28.19 kHz, and 38 kHz and/or
frequency bands of 20-25 kHz, 25-30 kHz, 38-40 kHz, 40-45 kHz, 40
to 60 kHz, 40-80 kHz, 140-160 kHz.
[0140] For other tissues, such as skin, the ultrasonic frequency
may be outside these ranges.
[0141] In preferred forms of the present invention, the ultrasonic
power used is relatively low, typically in the range 0.05 to 3.5
Wcm.sup.-2. Higher intensity ultrasonic power may be needed in some
applications. In these cases it may be necessary to pulse or
otherwise control the duty cycle of the ultrasonic energy to
prevent tissue damage, (e.g. from thermal effects) and/or to
prevent damage to the agent.
[0142] The ultrasonic energy applied to the applicator causes
reciprocating motion of the tissue contacting surface. In typical
embodiments the displacement of the tissue contacting surface from
its mean position may be between about 100 nm and 2200 nm.
Embodiments may operate with a displacement more than 200 nm.
Embodiments may operate with a displacement less than 2100 nm.
Embodiments may operate with a displacement more than 400 nm. The
embodiments used in the experiments operated with a displacement
less than 500 nm, and more specifically less than 400 nm.
[0143] FIG. 31A illustrates plots of the displacement (nm) and
velocity (m/s) during operation of two types of applicator, MP1 and
MP4, with two types of tip assembly, e.g. agent carrier, "Tip
assembly 1" and "Tip assembly 2" at a range of frequencies between
0 and 200 kHz.
[0144] FIG. 31B illustrates plots of the displacement (nm) and
velocity (m/s) during operation of three types of handle unit
useable in an applicator according to an embodiment, AMO1 (Model
no. Sov37706); AMO2 (Model no. Sov39302) and ALCON1 (Model no.
Turbo Sonic-375), without a tip assembly, e.g. agent carrier, at a
range of frequencies between 0 and 200 kHz.
[0145] FIG. 31C illustrates plots of the displacement (nm) and
velocity (m/s) during operation of three handle units, AMO1; AMO2
and ALCON1, with a second type of tip assembly, e.g. agent carrier,
at a range of frequencies between 0 and 200 kHz.
[0146] FIG. 31D illustrates plots of the displacement (nm) and
velocity (m/s) during operation of three types of applicator, AMO1;
AMO2 and ALCON1, with a third type of tip assembly, e.g. agent
carrier, at a range of frequencies between 0 and 200 kHz.
[0147] FIG. 32 illustrates the tip displacement of the MP4 and AMO
1 applicators when driven at selected frequencies over a range of
drive voltages. The MP4 system was driven through an RF amplifier
at 22 kHz over a range of input voltages of between 50 to 400V peak
to peak. The AMO1 was driven at 28.19 kHz over the same voltage
range.
[0148] The studies of the devices were performed using a laser
doppler vibrometer model MSA400 from Polytec instruments in
Germany.
[0149] As can be seen by selecting the frequency (or frequency
band) of operation desired oscillation parameters can be chosen.
These parameters will vary depending on the particular agent
carrier used. In preferred embodiments these devices will be
operated in a frequency band that corresponds to one or more of the
peaks in motion as illustrated in the plots.
[0150] In typical embodiments the velocity of motion of the tissue
contacting surface may be between about 0.01 m/s and 0.4 m/s.
Embodiments may operate with a velocity more than 0.03 m/s.
Embodiments may operate with a velocity less than 0.36 m/s.
Embodiments may operate with a velocity more than 0.06 m/s. The
embodiments used in the experiments operated with a displacement
less than 0.05 m/s.
[0151] Embodiments of the present invention may advantageously be
used in the delivery of agent to delicate tissues, such as mucous
membranes (including the conjunctiva, buccal mucosa and labial
mucosa), the cornea and the external coats of the eye.
[0152] The ultrasonic power and/or frequency parameters in
embodiments may be increased or decreased for a variety of reasons
including to control the depth of penetration of the agent into
tissue. As an example, the ultrasonic power and/or frequency
parameters used for delivering agent to the epithelial surface
cells of a mucus membrane, may be lower than power and/or frequency
parameters used for delivering agent to the rich blood vessel
capillary beds and deeper connective tissue layers that lie below
the epithelial surface.
[0153] It is intended that the agent carrier body does not
penetrate any layer of the tissue surface. Although some
superficial cell damage may occur in using embodiments of the
present invention, it is not intended and is not relied upon in
order to achieve delivery of the agent to the target tissue.
Maintaining an intact tissue surface as much as possible may serve
to more accurately control the depth of penetration of the agent
into tissue layers.
[0154] The various micro-scale structures within the agent carrier
body described herein, amongst other things serves the purpose of
making direct contact between the agent carrier body and the tissue
surface to propagate ultrasonic energy, thereby minimizing the
extent of any continuous layer fluid within the agent carrier body
and between the agent carrier body and the target tissue (which
tend to attenuate ultrasonic waves).
[0155] One group of embodiments first described in the Applicant's
Australian patent application 2013901606, include an agent carrier
body having microstructures that form a plurality of micro channels
surrounded by rigid walls for delivery of various agents. The micro
channels are typically in the range of approximately 25 to 100
.mu.am across when measured transverse to the direction of
delivery, may have a length of between approximately 0.5 mm to 2
mm. Any suitable cross-sectional and/or longitudinal geometry can
be used.
[0156] In use, each channel contains the agent in a fluid column
within the channel and the ultrasonic energy is directly applied to
each fluid column and the walls surrounding the fluid column. The
ultrasonic wave is generated to be longitudinal in nature, i.e. it
propagates along the channel. In some embodiments, by using the
micron scale architecture of the microstructures, the wave front
that impacts the fluid column is concentrated within each micro
channel thus reducing attenuation of ultrasound. Reflection of
ultrasonic waves at tissue surface is minimized by having direct
contact of the device, and most preferably the agent carrier body,
with the tissue surface so as not to permit the presence of a
fluidic space between them. This further assists molecules to
efficiently move toward the target tissue under the influence of
ultrasound along the ultrasonic wavefront path. The ultrasonic
waves are also carried in the agent carrier body, and specifically
in the walls defining the micro channels. Since they do not
attenuate the ultrasonic energy as much as fluids do, they
efficiently transmit the sonophoretic power to the target tissue
directly.
[0157] In preferred embodiments, the tissue-contacting surface of
the device is not separated from the tissue by a continuous layer
of fluid. The tissue-contacting surface of the agent carrier body
presents a surface that has areas of solid body and liquid agent
(i.e. the openings of the micro channels), in some embodiments
approximating a solid-liquid "checker board"-like array. This
arrangement may facilitate the sonophoretic ability of the device
since the faces of the solid walls directly contact the tissue. In
such embodiments the device architecture might be conceptualized as
a large number of individual micron-scale sonophoretic delivery
devices tightly packed and joined together.
[0158] Another group of new embodiments include a plurality of
micro-scale structures that is formed by micron-scale protrusions
that together define the tissue contacting surface of the agent
carrier body. These protrusions contact the target tissue and the
agent to be delivered surrounds them. In preferred forms, the agent
carrier body has a peripheral structure, typically a wall, that
surrounds the protrusions and contains the agent in use. This
embodiment has a lower ratio of microstructures to fluid within the
agent carrier body compared to an agent carrier body comprised of
micro-channels. Preferably these embodiments maintain direct
contact between the ultrasonic source and the target tissue via the
protrusions, and possibly also the peripheral structure. The
longitudinally directed ultrasonic waves are conducted by the
protrusions and the fluid between. The protrusions act by
facilitating the transport of drugs toward the target tissue.
Waveform interference from fluid in adjacent spaces between
protrusions is minimised by the presence of the protrusions, which
serve to at least block propagation of waveforms.
[0159] Another group of new embodiments present a hybrid device,
having at least one region having multiple micron-scale protrusions
and at least one other region having micro channels surrounded by
rigid walls. Typically a region or regions having micro channels
surrounded by rigid walls will form part of a peripheral structure
bounding a region that has micron-scale protrusions.
[0160] Molecules that are known to the inventors to possibly be
delivered to the body using sonophoresis include 1) molecules
having any kind of electric charge or have a neutral (including
overall neutral) electrical charge and 2) small or large molecules
(including monoclonal antibodies of approximately 150,000 Daltons)
and 3) molecules that are hydrophilic or hydrophobic or
lipophilic.
[0161] Several exemplary embodiments of the various aspects of the
invention are described with reference to an exemplary applicator
device for delivering an agent non-invasively to a target tissue
surface site via a transportation modality, which preferably uses
only ultrasonic waves. In these exemplary embodiments, at the
target tissue surface site, penetration of the agent into the
target tissue surface site is enabled or enhanced through
sonophoretic mechanisms. Preferably, target tissue surface sites
are mucous membranes including, but not limited to, conjunctival,
vaginal, urethral, inner ear, tracheal and bronchial mucosa, anal,
oral, and nasal tissues. A target tissue surface can also include
the cornea.
2 General Principles and Micro-Channel Embodiments
[0162] The system comprises an applicator device that is preferably
hand-held and used for delivering an agent to a target tissue. The
preferred form of applicator device includes a handle coupled to an
applicator tip. The applicator tip includes an agent carrier body
that has micro channels formed in it through which the agent is
delivered from within the applicator tip to a target tissue
surface. The agent carrier body may be integrated within the
applicator tip, or may be a separate component (such as a
cartridge) that is attachable to the applicator tip.
[0163] The applicator tip may include a reservoir that holds an
agent. The reservoir may form part of the agent carrier body, or
may be a separate component that is in fluid communication with the
agent carrier body.
[0164] An ultrasonic transducer forming part of the handle or
applicator tip generates ultrasonic energy (waves) which causes the
agent to be moved through the micro channels in the agent carrier
body, egress through terminal pores of the micro channels at a
tissue contacting surface of the agent carrier body and onto the
target tissue surface. The ultrasonic waves also enhance and/or
permit agent uptake into the target tissue through
sonophoresis.
[0165] FIG. 1A is a highly schematic diagram illustrating a first
embodiment of an applicator device according to the present
invention that is useable with any agent carrier or agent carrier
body described herein. In this example, an applicator device 100
includes an applicator tip 102 coupled to an applicator handle 103
(entire device not shown). The applicator handle 103 includes an
ultrasonic generator 101. The applicator tip 102 is connected to
the handle 103 so that ultrasonic energy from the transducer 101 is
transmitted to it via a coupling rod 106. As will be appreciated
the application of ultrasound will be generally in accordance with
the parameters set out in the overview above. The tissue contact
surface of the applicator tip 102 is brought into contact with a
target tissue surface 108. The ultrasonic generator is then
activated, which results in the propagation of ultrasonic waves 110
via the coupling rod 106, through the applicator tip 102 and the
agent carrier body 104 and into the target tissue 108. In this
embodiment, agent is stored in the agent carrier body 104 and is
transported to the target tissue surface 108 via micro channels 112
that have been fabricated within the agent carrier body 104.
Ultrasonic waves assist in the transport of agent from the agent
carrier body 104 to the target tissue surface 108 via the micro
channels 112. Ultrasonic waves also enhance and/or permit the
penetration of the agent into the target tissue 108 via
sonophoretic effects on tissue ultrastructure.
[0166] In this example, the agent carrier body 104 may be of any
type described generally herein, and as exemplified in any one of
FIGS. 8A to 10 or 23 to 30A. However, to illustrate the principle
of operation of an agent carrier body FIG. 1B provides a more
detailed view of an agent carrier body 104 of the type previously
described in the Applicant's Australian patent application
2013901606, 1A applied to the tissue surface 108. The agent carrier
body 104 has a tissue-contacting surface 114. In this example it
includes with micro channels 112 fabricated within the agent
carrier body 104 that extend from within the interior of the agent
carrier body 104 to the tissue-contacting surface 114. The micro
channels 112 terminate as pores 116 at the tissue-contacting
surface 114. Agent is provided from the agent carrier body 104,
through the channels 112 where it egresses through the pores 116 in
the tissue-contacting surface 114, and on to the tissue surface
108. As an alternative the agent carrier body 104 may be of any
type described generally herein, and as exemplified in any one of
FIGS. 8A to 10 or 23 to 30A.
[0167] In this example, ultrasound 110 is generated and conducted
through the agent carrier body 104. This causes agent 118 stored
within the channels 112 to be released from the channels 112 and on
to the tissue surface 108. The penetration of agent into the tissue
108 is enhanced and/or permitted by the use of ultrasound, which
provides a sonophoretic effect on the tissue.
[0168] In the embodiment of FIG. 1A, the applicator handle 103 has
an ultrasonic transducer 101, which generates ultrasonic waves 110
that are transmitted through the applicator tip 102 to the agent
carrier body 104 via the coupling rod 106. However, in alternative
embodiments the applicator tip 102 can be fabricated to include
within its structure, a system that is capable of generating
ultrasonic waves itself without the need for an external ultrasonic
transducer. FIG. 10 illustrates an alternative embodiment in which
the agent carrier body 104 additionally includes an ultrasonic
transducer 124.
[0169] It is preferred that the inner surface(s) of the channel 112
are functionalised. The inner surface 113 of the channels 112 may
be functionalised with compounds or molecules having hydrophobic or
hydrophilic properties or a combination of both moieties.
Alternatively, the surface 113 of the channels 112 may be
functionalised by contacting the surface of the channels with small
molecules that are adsorbed to the surface of the channels,
exposing specific functional groups that have the desired physical
and/or chemical properties. The small molecules may be adsorbed
through chemisorption or physisorption to the internal surface of
the channels. Alternatively, or in addition to changing the
water/oil affinity, the inner surfaces of the micro-channels and/or
agent reservoirs may be functionalised by enabling them to become
electro-conductive.
[0170] FIG. 2 provides an illustration of an embodiment of the
handle assembly 200 of an applicator device, usable with an agent
carrier body of any type described generally herein, and as
exemplified in any one of FIGS. 8A to 10 or 23 to 30A. The handle
assembly 200 includes a main housing 202, which contains an
ultrasonic transducer 204. The transducer is powered by a battery
206 (or alternatively by an external power supply) and is
configured to generate ultrasonic waves and transmit them to a
coupling rod 208 that terminates in a connector 210. The connector
210 can be of any type for example a screw thread or bayonet
fitting or the like, that enables the handle assembly 200 to engage
with an agent carrier (through either direct or indirect
engagement).
[0171] FIG. 3 is a schematic cross section of an applicator tip 300
that may be used with the handle assembly 200 of FIG. 2. The
applicator tip 300 includes a housing 301 having a first end 302
and a second end 303. The first end 302 includes a mounting
mechanism 305 such as a bayonet fitting or screw thread or the
like, that makes a mechanical connection with a connector 210 of
the handle assembly 200. The applicator tip 300 further includes a
recess 304 at its second end 303 that is arranged to accept the
agent carrier body 104 or an agent carrier body of any type
described generally herein, and as exemplified in any one of FIGS.
8A to 10 or 23 to 30A. The applicator tip 300 is configured, in
use, to carry agent to the tissue-contacting surface 306 of the
agent carrier body 104 and deliver it as required to tissue being
treated by application of ultrasonic waves. In some embodiments the
applicator tip 300 can include an agent reservoir, which is
fluidically in contact with the micro channels formed in the agent
carrier body 104.
[0172] FIGS. 4A, 4B, 4C, and 4D provide illustrations of various
embodiments of single layer agent carrier bodies, and FIGS. 4E, 4F,
4G, 4H provide illustrations of various embodiments where an agent
carrier body is created from stacked agent carrier layers.
[0173] The agent carrier body 400 is formed of a layer(s) of solid
material and possesses a number or network of micro channels that
may be a variety of geometric shapes and sizes. These micro
channels can be used to store or retain an agent and also to
deliver agent from within the agent carrier body 400 to a
tissue-contacting surface 406 of the agent carrier body 400. The
micro channels can be created by a micro-fabrication technique. For
instance, in embodiments where the agent carrier body 400 is formed
from silicon, the micro channels can be formed by lithography,
etching and/or other processes. In embodiments made from metal,
plastics or polymers the micro channels can be created by other
techniques including the use of lasers of various types and
wavelengths and molding and extrusion technologies. The use of
these micro-fabrication techniques are particularly desirable as
they provide the advantages of retained agent volume accuracy, the
benefits of predicable micro-fluidics and further permits
refinements such as specialised surface chemical treatment to
either or both the exposed tissue-contacting surface and the
internal walls lining the micron-scale cavities 402 of the agent
carrier body 400. These benefits can be used, for example, to
further enhance agent loading, retention and delivery to a target
tissue.
[0174] The tissue-contacting surface 406 has a series of openings,
fenestrations or pores 404. A wide variety of shapes and sizes of
pores can be on the order of 10 to 100 .mu.m, but other embodiments
may have pore sizes up to 1000 .mu.m. The micro channels 402 extend
from the pores 404 in the tissue contact surface 406 at least
partially through the agent carrier body 400. The micro channels
402 can be used for both retention of the agent and transportation
of the agent to a tissue surface.
[0175] The pores 404 may have a patterned appearance and exhibit a
range of geometries, for example: close packed hexagon structures,
arrayed squares with assorted densities, mixed polygon mosaics,
spirals, lines etc. The desired geometries are physically etched
into the agent carrier body 400 so as to create arrays of micro
channels 402 for retention and/or transport of an agent. The micro
channels may be in a variety of shapes for example cylindrical,
conical etc.
[0176] The walls of the micro channels 402 and/or other internal
surfaces within the agent carrier body 400 may be treated such
that: they have hydrophilic or hydrophobic characteristics that may
be the same or opposite in nature to each other and/or the areas
between the pores 404 of the tissue-contacting surface 406. The
walls of the micro channels 402 and/or other internal surfaces
within the agent carrier body 400 may be treated such that they
conduct electric charge or can generate a local electric field that
may have the same or opposite polarity to each other and/or the
areas between the pores 404 of tissue contacting surface 406.
[0177] The agent carrier body 400 can be formed from a unitary
piece of material. However, in alternative embodiments the agent
carrier body may include a number of layers that are stacked. The
use of micro-fabricated solid material as single or multiple layers
to create an agent carrier body allows for improved acoustic
transmission and thus improved delivery of agent to a target tissue
site by ultrasound.
[0178] The dimensions and internal lining characteristics of the
micro channels 402 and/or other internal surfaces within the agent
carrier body 404, and the dimensions and number of layers
comprising the agent carrier, will be tailored to suit the agent
and the target tissues, and will vary as a consequence of agent
properties, dose and formulation requirements, ultrasonic power and
heat generation, and the duration of use.
[0179] FIG. 4B shows another embodiment, similar to that of FIG.
4A, except that the micro channels 402' are interconnected by
internal linking channels 408. Such a structure provides some level
of agent storage in addition to channels 402' alone.
[0180] FIG. 4C represents a further embodiment in which the single
layer agent carrier body 400'' has micro-channels 402'' which
terminate as pores 404'' in the tissue-contacting surface 406'' at
one end of the micro-channels 402'', and connect at their other end
to an agent reservoir 410.
[0181] FIG. 4D provides surface views of a single layer agent
carrier body shown in any one of FIGS. 4A to 4C. The agent carrier
body 400'' has a first surface 411 and a second surface 412 which
is the tissue-contacting surface. As previously discussed,
micro-channels extend from within the agent carrier body 400 (from
a reservoir 410 or linking channel 408 if present) and terminate as
pores 404 in the tissue-contacting surface 412.
[0182] In alternative embodiments, the agent carrier body has a
stacked layer structure and includes at least two layers. More
preferably, one or more layers have additional micro-reservoir
volumes formed within them and which are in fluid communication
with the micro-channels for holding agent prior to application to
the tissues being treated. The micro-reservoir volume may be a
single volume or a plurality of small volumes, e.g. each of which
is contiguous with one or a group of micro-channels. There may be a
single large reservoir volume in the layer furthest from the
tissue-contacting layer that is fluidically connected with the
channels. Alternatively, there may be multiple micro-reservoir
volumes, with each of the micro-reservoir volumes being in fluid
communication.
[0183] FIGS. 4E, 4F, and 4G correspond with FIGS. 4A, 4B, and 4C
respectively, except that the agent carrier body 413 includes a
first layer 414, 414', 414'' and second layer 416, 416', 416''. The
first layer 414, 414', 414'' is as generally described with respect
to the single layer embodiment of FIGS. 4A, 4B, and 4C, except
instead of having a tissue contacting surface 422, the first layer
has an interface surface 415 including pores or blind holes that
defines a portion of the micro channels that extend through the
first and second layers when the layers are stacked together. The
second layer 416, 416', 416'' includes a first surface 420 that
contacts the interface surface 415 of the first layer 414, 414',
414'' and a tissue-contacting surface 422 having pores 426 that are
formed by micro channels 424. As can be seen the micro channels 424
extend from within the first layer, through the second layer 416,
416', 416'', and terminate at the tissue-contacting surface 422 of
the second layer 416, 416', 416'' as pores 426. In this way, the
holes in the first layer 414, 414', 414'' and second layer 416,
416', 416'' are aligned to form the micro channels 424 so that the
first layer 414, 414', 414''and second layer 416, 416', 416'' are
connected permitting fluid continuity in the system.
[0184] FIG. 4H illustrates a further alternative embodiment of a
double stacked layer agent carrier body 413 in which the first
layer 414''' contains an open-ended agent reservoir 425 that
provides agent directly into the micro-channels of the second layer
416'''.
[0185] FIG. 4I provides surface views of the various layers of a
double layered agent 413'' carrier shown in FIG. 4H. The first
layer 414'' has a first surface 430 and a second surface 432. The
second layer 416'' has a first surface and a second surface (which
are the same and are generally represented as 434). The agent
reservoir 425, is formed by a recess formed in first layer 414''
that extends partially into it. The second surface 432 of the first
layer 414'' is aligned and placed over the interface surface of the
second layer 416'' such that substantially all of the
micro-channels 424 formed in the second layer are fluidically
connected with the agent reservoir 425 in the first layer
414''.
[0186] FIG. 4J provides illustrations of further embodiments of
agent reservoirs formed in an agent carrier body that can store
additional agent and replenish the micro-channels as they are
depleted of agent during the course of usage. The reservoirs may
connect to micro-channels in the same agent carrier body layer as
shown for example in FIG. 4G or connect to micro-channels in a
contiguous layer in the agent carrier body as shown for example in
FIG. 4H. Agent carrier body 438 includes a reservoir formed by two
annular ring shaped reservoir volumes 440 and 442 and includes a
conduit 444 extending through a port 446. When a vacuum is applied
to the port 446, or the port 446 is injected with agent, a negative
pressure or a positive pressure respectively is applied to the
reservoir 440, 442. A layer of this type is arranged in a stack of
layers to form the agent carrier body, the first layer overlies its
adjacent layer such that any holes in the adjacent layer
fluidically connect to the reservoir volumes to allow agent to
travel via micro channels through the layers and to the
tissue-contacting surface.
[0187] Agent carrier body 448 is another embodiment in which the
reservoir consists of a number of concentric rings each fluidically
connected to each other. It will be appreciated that other
arrangements of the agent reservoir volumes within a layer are
possible without departing from the invention.
[0188] Generally, the holes in a lower or intermediate layer of an
agent carrier body extend through the whole thickness of that layer
and in combination with subsequent fluidically connected holes in
other layers, form a micro channel that extends from the
tissue-contacting surface in the surface contact layer of the agent
carrier. It will be appreciated that in certain instances the holes
only extend partway into a particular layer; this can be the case
for the first layer as illustrated for example in FIGS. 4E-4G.
[0189] As stated previously, it is preferred that the inner
surface(s) of the micro channels and other internal surfaces of the
agent carrier, such as those of the agent reservoirs, may be
functionalised.
[0190] In the embodiments illustrated in FIGS. 5A, 5B and 5C the
agent carrier body includes six layers including a surface
tissue-contact layer and five layers stacked on top of one another
overlying the surface contact layer.
[0191] FIG. 5A shows an embodiment of an agent carrier body 500
having six stacked layers 501.1, 501.2, 501.3, 501.4, 501.5, 501.6
The first end of the agent carrier body is a surface tissue-contact
surface 502 on layer 501.6 that contacts the tissues being treated.
On top of this layer there are a plurality of additional layers and
a top most layer 501.1. In this embodiment the agent reservoirs and
micro channels for holding and delivering agent (not shown) may
extend through some or all of the layers 501.1-501.6 of the agent
carrier 500. In some embodiments, the channels extend from the
tissue-contact surface 502 in layer 501.6, through intermediate
layers 501.5 to 501.2, and terminate in the top-most layer
501.1.
[0192] FIG. 5B shows an alternative embodiment of a six stacked
layer agent carrier body 505 to that shown in FIG. 5A. In this
embodiment, the surface tissue-contact layer 501.6 includes a
number of micro-protrusions 506, which in this example are
micro-tubules. FIG. 5C shows a further alternative agent carrier
body 510 having a similar overall arrangement but in which the
micro-protrusions 506' are micro-needles. The micro-protrusions are
hollow, and included channels formed therein that form a part of
the system of minor channels for delivering the agent.
[0193] Micro protrusions, such as micro-needles and microtubules
can be created by secondary fabrication consisting of etching the
tissue contact surface 502 of a tissue-contacting layer 501.6 such
that the areas between the pores are largely removed. This leaves a
wall around each pore of the required protrusion to surround each
pore. The micro-needles and microtubules can be of any shape
desired. For example, FIG. 5D shows the micro-protrusions as having
a cylindrical shape (micro-tubules 510) and other micro-protrusions
as having a frustoconical shape (micro-needles 508). In other
embodiments, not illustrated, the surface 502 tissue contact layer
501.6 can be provided with other surface treatments, or surface
engaging structures, such as a saw tooth structure, ripples, rings
or the like to help the agent carrier body interface with the
target tissue.
[0194] In a preferred embodiment each layer is disc shaped or
cylindrical in shape. Preferably the layers have a thickness of
from about 0.3 mm to about 1.0 mm, and even more preferably each
layer has a thickness of about 0.5 mm. It is preferred that each
layer has a diameter of from about 3 mm to about 10 mm, and even
more preferably has a diameter of about 5 mm. The thickness
dimension and the diameter dimension may vary between layers. While
the layers and overall shape of the agent carrier body have been
described as being disc shaped or cylindrical in cross sectional
shape, as in FIG. 3, other shapes could be employed without
departing from the ambit of the invention, e.g. rectangular,
square, or other polygon, oval etc. Furthermore, while it is
preferred that the overall shape of the agent carrier body is of
constant cross section the overall shape of agent carrier body
could change along its length e.g. the agent carrier body could be
shaped as a frustum (whether conical or otherwise pyramidal), or a
prism etc. The overall shape and/or the shape of components of the
agent carrier and the agent carrier body can modified in order to
maximise the efficiency of the device which is dependent on the
transportation modality or combination of transportation modalities
employed.
[0195] FIG. 6 provides an illustration of an agent carrier 600
having an agent carrier body 601 with stacked arrangement. The
stack includes a bottom most layer 602, four intermediate layers
604, a top most layer 606. The bottom most layer 602 has
micro-tubules 608 extending to form the tissue-contact surface 610.
The agent carrier 600 additionally includes a port 612. In this
embodiment the port 612 is part of the first layer 606. The port
612 is connected with micro-channels formed in the agent carrier
body 601, preferably via an agent reservoir volume in the first
layer 606 so that fluid can flow between them. The port 612 is
configured to connect to a vacuum line or pressure injector so that
a negative or positive pressure respectively can be applied to the
port 612. This allows the agent to be loaded into the agent carrier
from an external source. On application of a vacuum to the port
612, agent is drawn through the pores in the microtubules in the
tissue contact surface 610, through micro-channels into the stack
of layers of the agent carrier body 601 to fill the micro channels
and the reservoir volumes. Alternatively, agent can be injected
into the agent carrier via the port 612. Using either method, the
agent carrier can be charged with an agent.
[0196] FIG. 6 also shows a closure or seal 614 applied to the port
612, and a closure or seal 616 applied over the surface contact
layer 610. The seal 616 seals the surface of the surface contact
layer 610 to maintain sterility and any vacuum that is created
within the micro channels. Similarly, seal 614 seals the port 612
for similar purposes. It is preferred that this seal layer is a
plastic film.
[0197] The embodiment of FIG. 6 also includes an additional layer
618 and an ultrasonic transducer 620. Layer 618 may be a simple
insulation layer that serves to cover the fenestrations in the top
layer (if the micro-channels extend the entire way through the top
layer) to prevent the egress of fluids and/or to prevent release of
a contained vacuum.
[0198] The transportation modality may use an electric field to
cause a charged agent to be transported. The electric field can be
provided by applying a voltage to an electrode in the agent carrier
using an internal battery in the applicator device or by an
external power supply. In a preferred form an electrode is located
within the applicator device, a second external electrode, also
connected to the applicator device power supply, can be located in
such a way that the target tissue effectively becomes an electrode
opposite in polarity to that of the internal electrode. The
polarity of the electrodes can be selected such that the internal
electrode is of the same polarity as the electric charge on the
agent. The voltage established between the two electrodes
transports an electrically charged agent through the agent carrier
to the tissue-contacting surface and can enhance and/or permit the
transport of the charged agent into the tissue via iontophoresis.
Embodiments of the invention can use multiple delivery modalities
using ultrasonic waves and electric current used in combination
either alternately or simultaneously. Accordingly, Layer 618 can
additionally be modified to include, or alternatively be, a
material that serves as an electrode. The electrode can be
positively or negatively charged and is used to generate a static
or dynamic electric field. In the case where the top surface of the
adjacent agent carrier layer does not have pores and the adjacent
agent carrier layer is made from a material that is not
electro-conductive, there is no direct contact between the
electrode and the ions or charged agents contained within the micro
channels or reservoirs however, ions and charged agents of the same
polarity as that existing on the electrode will be repelled. If the
adjacent agent carrier layer is made from a material that is
electro-conductive and the adjacent agent carrier layer does not
have holes, there is electrical conductivity established with the
ions or charged agents contained within the micro channels or
reservoirs. This scenario is functionally equivalent to the case
where the surface of the adjacent agent carrier layer does have
pores (and is not dependent on the electro-conductivity of the
adjacent agent carrier layer) and the electrode is in direct
contact with the ions or charged agents contained within the micro
channels or reservoirs, where a further electrode, opposite in
polarity to layer 618 can be placed on, or adjacent to, the target
tissue. To complete the electric circuit, the electrode placed on
or adjacent to the target tissue may be connected to the agent
carrier; applicator handle; or other component of the application
device (not shown). An applied voltage can provide the energy
required to cause an electrically charged agent of the same
polarity as the electrode of layer 618, to flow in the fluid
contained in the micro channels of an agent carrier body 601 to
migrate through the agent carrier, out of the pores to the tissue
surface to be delivered into the tissue by iontophoresis.
[0199] This provides an alternative embodiment whereby the agent
carrier is able to generate an electric voltage to facilitate the
flow of an electric current to transport electrically charged
agents through the agent carrier and out of the pores to the
tissue.
[0200] In some embodiments the agent carrier body includes (as with
layer 618), or is itself an electrode to facilitate the transport
of a charged agent through the agent carrier and out of the pores
to the target tissue. The electrode may be located adjacent to the
stack of layers, or may be an electrode layer that is integrated
within the stack of layers (as with layer 618).
[0201] In the above embodiment, ultrasonic energy and/or electrical
voltage provide the energy required to move the agent through the
agent carrier to its tissue contact surface where sonophoresis
and/or iontophoresis enable the agent to be delivered into the
target tissue.
[0202] As will be appreciated in the above embodiments, a layer
including the tissue contacting surface e.g. 422, 422', 422'' 502,
610 can be a layer including a tissue contacting surface being at
least partly defined by a plurality of protrusions, such as those
described in any one of FIGS. 8A to 10 and 23 to 28.
[0203] FIGS. 7A and 7B provide an illustration of an embodiment of
the holes, and the channels defined by the holes, in a stack of
layers forming the agent carrier body according to an embodiment of
the present invention. FIG. 7A provides an illustration of a stack
of layers 700 that includes two layers, 702 and 704. Layer 702 is a
layer that is further from the tissue-contacting surface than layer
704. The layer 702 includes a plurality of holes 706; the layer 704
includes a plurality of holes arranged as a cluster of holes 708.
These layers 704, 702 are arranged adjacent to each other in the
stack of layers 700 such that each cluster of holes 708 in layer
704 is aligned with a hole 706 in layer 702. The holes in the layer
704 are more numerous and smaller than the holes in layer 702. To
facilitate alignment in the layers during device fabrication each
layer 702, 704 can be provided with a datum point or structure 707,
707' which define the alignment of the layer. Layers can then be
aligned with their respective datum points 707, 707' arranged in a
predetermined fashion (e.g. aligned with each other) to achieve
correct alignment of holes in respective layers 702, 704, thereby
forming micro-channels that extend through multiple layers of a
stack 700.
[0204] FIG. 7B provides a further illustration of the variation and
alignment between holes of different sizes in different stack
layers of the agent carrier body. Hole 706' is a magnified version
of hole 706. The hole 706' overlies a first cluster of holes 708
(shown in dotted lines) in the next adjacent stack layer. Hole 708'
is a magnified version of hole 708. The hole 708' overlies a
corresponding cluster of holes 710 (shown in dotted lines) in the
next adjacent stack layer. Similarly Hole 710' is a magnified
version of hole 710. The hole 710' overlies a corresponding cluster
of holes 712 (shown in dotted lines) in the next adjacent stack
layer. Hole 712' is a magnified version of hole 712 and so on until
the final layer.
[0205] Multiple layers can be arranged such that progressing from
the top most layer, through the intermediate layers, to the surface
contact layer, the diameter of the holes decreases and the number
of holes may be increased. Each subsequent layer includes a cluster
of holes that is in alignment with a hole in the adjacent
subsequent layer. For example, a first layer (which may be the top
most layer or an upper one of the intermediate layers) has a number
of holes. This first layer overlies a second layer, wherein the
second layer has clusters of holes that are arranged beneath the
holes in the first layer. This second layer may overlie a third
layer and each hole in each of the cluster of holes in the second
layer overlies a further cluster of smaller holes in the third
layer (additional layers may also be provided in this manner).
[0206] The channels define a flow path for the agent through the
agent carrier body to the tissue surface. The channels are defined
initially by the diameter of the holes in the first hole possessing
layer. Subsequent layers have clusters of holes that are aligned
with the holes in this first hole possessing layer. Therefore,
progressing from the first hole possessing layer through subsequent
layers, the channels become multi-furcated into numerous branches.
It will be understood that these numerous branches all form a part
of the channel.
[0207] FIGS. 7C, 7D, and 7E show magnified images detailing
examples of micro-channels created by a micro-manufacturing
process. FIG. 7C and 7D (7D showing a higher magnification of 7C)
shows a layer in which the holes have square cross-sections. FIG.
7E shows a layer that includes holes having square and hexagonal
cross-sections.
3. Protrusion-Based Embodiments
[0208] The following series of embodiments include a plurality of
microstructures formed from micron-scale protrusions that together
define the tissue contacting surface of the agent carrier body.
These micro-scale structures contact the target tissue and the
agent to be delivered surrounds them. In preferred forms the agent
carrier body has a peripheral structure, typically a wall, that
surrounds the protrusions and contains the agent in use. This
embodiment has a lower ratio of microstructures to fluid within the
agent carrier body compared to an agent carrier body comprised of
micro-channels. Preferably these embodiments maintain direct
contact between the ultrasonic source and the target tissue via the
protrusions, and possibly also the peripheral structure. As will be
appreciated the application of ultrasound will be generally in
accordance with the parameters set out in the overview above.
[0209] More specifically FIGS. 8A to 10 and 23 to 30A illustrate
several embodiments that employ an agent carrier body including a
tissue contacting surface for engaging tissues under treatment, the
tissue contacting surface being at least partly defined by a
plurality of protrusions. The protrusions preferably extend outward
from an inside of a void and terminate at said tissue contacting
surface. The void may be formed by a peripheral structure, where at
least part of said peripheral structure may terminate at the tissue
contacting surface.
[0210] In some embodiments the peripheral structure terminates in a
common plane with the protrusions. In others at least some of said
protrusions defining the tissue contacting surface extend outward
from the void beyond the peripheral structure. In some embodiments,
the protrusions may terminate in a plane and the peripheral
structure may terminate short of the plane such that the
protrusions extend beyond the peripheral structure.
[0211] In such embodiments is should be noted that the protrusions
of the preferred embodiments do not act as microneedles. Unlike
microneedles, the protrusions of the preferred embodiments are not
intended to penetrate any layer of tissue. The function of
protrusions includes engaging the target tissue by applying
pressure resulting in a frictional force on the surface. This aids
the positioning of the device (e.g. on the slippery surface of
mucous membranes) and enhances the sonophoretic effect.
[0212] As will be appreciated, the agent carrier bodies exemplified
in these figures, can be used in place of any agent carrier body
illustrated herein e.g. agent carrier body 104, 810, 902, 903,
903', 903'', 903''', 903'''' and 1302, or with any of the agent
carriers described herein, e.g. agent carriers 300, 800, 800' and
900, 900', 900'', 900''', 900''''.
[0213] In the preferred embodiments, protrusions include the
following properties: [0214] They do not have a needle-like tip,
that is, they do not narrow to a point such that their width does
not decrease to near zero at the tip. [0215] The cross-section is
relatively constant, at least near their tip, and most preferably
along their whole length. In most cases the width will not narrow
by more than 20%, and preferably less than 10% towards its tip.
[0216] they typically have a tip width greater than 10 .mu.m. Thus
the scale of the protrusions also differs generally from that of
microneedles. [0217] They do not enter an intact epithelial surface
of the target tissue. [0218] They aid in stabilizing the device by
the frictional force they apply when the device is placed in
contact with the tissue. This is particularly advantageous on
mucous membranes that tend to have a low friction surface due to
local mucous secretions. [0219] They generally have a height to
width aspect ratio (across their shortest cross sectional width) of
between 1:1 to 10:1. Whilst higher aspect ratios may be used it is
difficult to achieve acceptable strength that they can withstand
handling, loading and application of ultrasonic energy without
damage. As will be appreciated cross sectional shape will greatly
affect the strength of them and will be chosen accordingly. [0220]
In preferred embodiments the protrusions occupy more than 5% of the
volume surrounding them in which agent is carried. This percentage
needs to be high enough so that the capillary force or other forces
retain the agent within the agent carrier body against gravity or
other forces caused by normal handling. In embodiments used with
water-like agents, will typically have a density of projections of
greater than 5% and most preferably greater than 10%. It should
also be appreciated that as the agents become thicker, e.g. protein
rich agents, the density of protrusions, or their size and/or wall
surface area, can be lowered.
[0221] FIGS. 8A to 8G illustrate schematic representations of
alternative embodiments of an agent carrier body and agent carrier
body layers which include multiple protrusions that together define
the tissue contacting surface of the agent carrier body.
[0222] In this embodiment the agent carrier body 750, can be used
for delivery of an agent to a tissue via a transportation stimulus.
The agent carrier body 750 includes a tissue contacting surface 752
for engaging tissues under treatment. In this example the tissue
contacting surface is defined, at least partly by a plurality of
protrusions 754.
[0223] The protrusions 754 may be of any shape, but in the present
example are generally cylindrical. Preferably the protrusions have
a constant cross sectional shape along their height. The
protrusions 754 extend outward from an inside of a void 756 that is
formed within the agent carrier body 750. The outward ends 758 at
least partly define the define the tissue contacting surface of the
agent carrier body 750,
[0224] The void 756 is formed by a peripheral structure 760, which
in this case takes the form or a peripheral wall or rim. The rim
760 also defines part of the tissue contacting surface 752.
[0225] The peripheral structure 760 in this embodiment terminates
in a common plane with the protrusions, to define a planar tissue
contacting surface 752. However, in other embodiments the at least
some of said protrusions 754 can extend beyond, and/or stop short
of the peripheral structure so that tissue contacting surface 752
is not planar. In some embodiments the protrusions 754 may all
extend beyond the peripheral structure 760.
[0226] The void 754 acts as a reservoir to hold agent within the
agent carrier body 750. However unlike previous embodiments this
reservoir is located on the tissue contacting surface side of the
agent carrier body.
[0227] The protrusions 754 are located within the reservoir so that
they are in fluid communication with the agent in the reservoir.
This allows the protrusions 754 to act on the agent within the
agent carrier body 750 and transmit the transportation stimulus
into the agent, whereas in the embodiments above the walls of the
micro channels acted on the agent within the agent carrier
body.
[0228] Embodiments of this type generally have more volume for
holding agent than embodiments described above. By having a larger
filling volume, the possibility of air entrapment may also be
reduced. These improved filling properties may give certain
embodiments improved filling accuracy and repeatability, which
contributes to an increase in dose accuracy, that may be important
in medical applications. Furthermore the improved filling may lead
to better ultrasonic energy transmission as dampening by retained
air spaces is reduced.
[0229] It is preferred that the inner surface(s) of the void 754
are functionalised. The inner surface of the void 754 and the
protrusions 752 may be functionalised with compounds or molecules
having hydrophobic or hydrophilic properties or a combination of
both moieties. Alternatively, the surface of the void 754 and the
protrusions 752 may be functionalised by contacting the surface of
the channels with small molecules that are adsorbed to the surface
of the channels, exposing specific functional groups that have the
desired physical and/or chemical properties. The small molecules
may be adsorbed through chemisorption or physisorption to the
internal surface of the channels. Alternatively, or in addition to
changing the water/oil affinity, the inner surfaces of the
micro-channels and/or agent reservoirs may be functionalised by
enabling them to become electro-conductive. In a preferred form
loading of the agent carrier body is performed by virtue of
capillary forces when the agent carrier is in contact with the
agent.
[0230] FIGS. 8C and 8D show an agent carrier body layer 750'. In
general the agent carrier body layer 750' is the same as the agent
carrier body 750 and like features are like numbered. However the
agent carrier body layer 750' additionally includes one or more
micro channels 762 extending through it. The micro channels 762
extend through the agent carrier body layer so that the reservoir
756 may be fluidly connected to an adjacent agent carrier body
layer as in previous embodiments. In this example, four micro
channels are used.
[0231] FIGS. 8E and 8F are schematic representations of an agent
carrier body layer having a reservoir formed therein. The agent
carrier body layer 764 is generally cylindrical in form and
includes a peripheral wall 766 that defines a reservoir volume 770
within it. In use the agent carrier body layer 764 is stacked on
the agent carrier body layer 750' such that the outer rim 768 of
the wall 766 contacts the back of the agent carrier body layer 750'
such that a reservoir volume 770 closed. The micro channels 762 in
the agent carrier body layer 750' allow agent within the reservoir
volume 770 to pass into the reservoir 756 for dispensing.
[0232] FIGS. 8G and 8H are schematic representations of an agent
carrier body formed by the agent carrier body layer of FIGS. 8E and
8F stacked with the agent carrier body layer of FIGS. 8C and 8D to
form an agent carrier body 780. The agent carrier body 780 includes
a stack of layers including the tissue-contacting layer 750' which
includes the tissue contacting surface 752 and one other layer 764.
More layers could also be stacked to form an agent carrier
body.
[0233] In FIG. 8H the agent carrier body 780 is shown filled with
agent. In this configuration the agent is filled to the tissue
contacting surface 752.
[0234] FIG. 9A and is an electron micrograph showing a portion of
an agent carrier body (or layer thereof) of the type schematically
illustrated in FIGS. 8A to 8H. FIG. 9A shows part of three pillars
782 that operate as protrusions 754. The surface 784 is the base of
the void 756 from which the pillars 782 extend. FIG. 9B and is an
electron micrograph showing a close up portion of another pillar
786. As can be seen these embodiments from their respective scales,
the pillars 782 and 786 are around 200 micrometres wide and a
similar height. However in other embodiments different heights and
widths may be used.
[0235] FIG. 10 illustrates a series four mask designs, each
suitable for forming a respective agent carrier body (or layer
thereof). The masks are used in a micromachining process for
forming the protrusions and peripheral structure of a tissue
contacting surface of an agent carrier. The protrusions are to be
arranged in a pattern, in this example in a regular array.
[0236] In FIG. 10 the mask for each device (Devices 1 to 4)
includes a first mask section 792 for defining a square peripheral
wall. Device 1 includes an array of 25 mask sections 794 arranged
in a 5.times.5 array to create a 5.times.5 array of protrusions.
Device 2 includes an array of 16 mask sections 794 arranged in a
4.times.4 array to create a 4.times.4 array of protrusions. Device
3 includes an array of 9 mask sections 794 arranged in a 3.times.3
array to create a 3.times.3 array of protrusions. Device 4 also
includes an array of 16 mask sections 794 arranged in a 4.times.4
array to create a 4.times.4 array of protrusions. As can be seen,
the protrusions of Device 4 are spaced more widely than that of
Device 2. The height of the protrusions in embodiments of the
present invention can be chosen to create a void having a desired
volume. In some embodiments the protrusions can be greater than 200
.mu.m in height. In some forms they can be less than 1 mm in
height. In some embodiments the protrusions can be greater than 300
.mu.m in height. In some forms they can be less than 800 .mu.m in
height. In some embodiments the protrusions can be greater than 400
.mu.m in height. In some forms they can be less than 700 .mu.m in
height. In some embodiments the protrusions can be greater than 500
.mu.m in height. In some forms they can be less than 600 .mu.m in
height. In some embodiments heights greater than 1 mm or less than
200 .mu.m could be used.
[0237] FIG. 23 illustrates a series (a) through (g) of mask designs
for creation of various agent carrier bodies (or layers thereof).
Figures (a) to (e) are embodiments with round protrusions 794,
whereas figures (f) and (g) have protrusions 794 that are
cross-shaped in plan-view. The embodiments are summarised in the
following table.
TABLE-US-00001 Protrusion Protrusion width separation Example Array
.mu.m .mu.m Protrusion shape a 7 .times. 7 100 200 round b 6
.times. 6 100 300 round c 5 .times. 5 100 400 round d 5 .times. 5
200 200 round e 4 .times. 4 200 400 round f 4 .times. 4 450 100
cross 200 .mu.m arm length, 50 .mu.m arm thickness g 3 .times. 3
650 100 cross 300 .mu.m arm length, 50 .mu.m arm thickness
[0238] It will be appreciated that these embodiments are not
exhaustive in any way and many alternative embodiments, having
different protrusion dimensions, separations, cross sectional
shapes can be devised. It should also be noted that, whilst these
embodiments are contained within a square rim designated by
reference numeral 792, other shapes can be used. Furthermore, the
array of protrusions need not be a regular array or have even
density or distribution across the chip. All protrusions 794 used
in an embodiment need not have the same cross sectional shape.
[0239] Examples (f) and (g) have cross shaped protrusions 794. The
cross shaped protrusions have the advantage that they have an
increased wall surface area compared to round protrusions, but a
reduced cross sectional area, thus maximising agent storage volume.
The geometry of cross shaped protrusions also have relatively good
mechanical properties, insofar as each arm acts as a buttresses to
support the transversely extending arm.
[0240] FIGS. 24 to 28 illustrate a series of electron micrographs
of agent carrier bodies and regions thereof. FIGS. 24(a) to (c)
show a first embodiment. This embodiment has a 4.times.4 array of
cross shaped protrusions 782 extending upward from a void 784. The
void 784 is surrounded by a peripheral wall 785, as in previous
embodiments. In use agent to be delivered is retained in the agent
carrier body by using the void 784 as a reservoir. As can be seen
the protrusions 782 are cross shaped in cross sectional shape over
their whole height although their width changes. The changes,
particularly near their tip are relatively small, such that the
topmost surface, which forms the tissue contacting surface of the
agent carrier body, is substantially flat. In this embodiment the
peripheral wall 785 is around half a millimetre high, and most
specifically 484.89 .mu.m. The protrusions are substantially the
same length. The width of the cross profile in this embodiment is
31.11 .mu.m. This is compared with a nominal cross section, as
defined by the mask of 50 .mu.m and represents almost a 40%
tapering of the protrusion. However, as can be seen the protrusions
are not sharp like microneedles despite the small thinning towards
the top. This flat surface and the plurality of densely packed
protrusions prevent mechanical damage or penetration of the surface
by these embodiments.
[0241] FIGS. 24(d) is a further embodiment also having cross shaped
protrusions 782 surrounded by a generally square peripheral rim
785. In this example the protrusions are arranged in a 3.times.3
pattern.
[0242] FIGS. 25(a) to (e) are electron micrographs of several
embodiments with circular cross section. Like features are like
numbered and will not be described in detail. As will be observed
however, despite a narrowing of the protrusions 782 towards their
tip, the tip is flat and not pointed like a microneedle. Again in
this embodiment the protrusions are about half a millimetre high
(525.25 .mu.m, as shown in picture (b). The diameter of the tip of
a protrusion, as illustrated in FIG. 25 is 184.02 .mu.m.
[0243] FIGS. 26(a) to (d) are electron micrographs of several
additional embodiments having cross shaped (a) and circular cross
section (b) to (d). In this example the sides of the projections
782 are more vertical than previous embodiments. That is, their
sides taper less than the previous embodiments. This is most
noticeable in picture (a) to (c).
[0244] FIGS. 27(a) to (d) are electron micrographs of several
additional embodiments having protrusions with a circular cross
section. Again in these examples the sides of the projections 782
are more vertical than previous embodiments. The height of the
peripheral wall and protrusions are again around half a millimetre.
Two final embodiments are shown in FIGS. 28(a) and (b). These
example are a 4.times.4 array and 5.times.5 array respectively.
[0245] In this micrograph the protrusions also taper at their
bottom. The tapering is a result of the etching process used during
manufacturing, and is largely unintentional. However, in some
embodiments, this tapering can be used to advantage as it increases
the volume of the void in which agent is held.
4 Hybrid and Alternative Embodiments
[0246] FIGS. 29 and 30 illustrate two hybrid embodiments of the
agent carrier body. In FIG. 29 the agent carrier body 104 includes
a plurality of micro channels 112 arranged around its peripheral
edge in the rim 785. It also has an array of protrusions 782 formed
within a central void 784.
[0247] FIGS. 30 and 30A illustrates an alternative embodiment,
which can be viewed as a protrusion-based embodiment, but with a
textured, or profiled rim. In this case the agent carrier body has
a peripheral rim 785 which is castellated. The rim 785 has a series
of channels or fenestrations 785A that extend through the rim 785
from the peripheral edge to the void 784. The void 784 also
contains protrusions 782 in a 3.times.3 array. To illustrate the
arrangement better a cross section along line 30A-30A is provided
in FIG. 30A.
[0248] An agent carrier having a plurality of agent carrier bodies,
perhaps arranged in a pattern such as an array, could also be
provided.
5. Loading and Use Examples
[0249] FIGS. 11A, 11B, and 11C illustrate an embodiment in which
the agent reservoir is provided within the agent carrier as a
separate component to the agent carrier body.
[0250] FIG. 11A illustrates a portion of an applicator according to
a further embodiment of the present invention. In this figure there
is illustrated an embodiment of an applicator tip 800 attached to a
coupling rod 802, for coupling the applicator tip 800 to a handle
portion of a hand-held agent applicator device. The applicator tip
800 includes an agent reservoir 804 formed within the tip's housing
803. The housing 803 also includes a recess area 806 for receiving
an agent carrier body. The agent reservoir 804 includes a port 808.
The port 808 may be configured for a number of different uses. In
certain embodiments the port 808 may be used to inject the agent
reservoir 804 with an agent. In other embodiments the port 808 may
be used to apply a vacuum to the agent reservoir 804 to draw agent
into the reservoir 804.
[0251] FIG. 11B provides applicator tip 800' with an agent carrier
body 810 inserted into the recess area 806 (not shown due to the
presence of the agent carrier body 810). As will be appreciated
from the description in FIG. 11A, the agent reservoir 804' may be
filled with an agent by suction applied to the port 808' whereby
the agent is drawn through the agent carrier body 810 via its micro
channels for storage/holding in the reservoir 804. Alternatively,
port 808' may be used to directly inject the agent reservoir 804'
with an agent which then fills both the reservoir 804' and the
micro channels in the agent carrier 810 with the agent.
[0252] FIG. 11C provides a further embodiment of an applicator tip
800'' as generally described above, and accordingly corresponding
features have been like numbered with the addition of double prime
to indicate the change of embodiment. The applicator tip 800'' is
connected to coupling rod 802''. It includes an agent reservoir
804'' and a stacked agent carrier body 810''. In other respects it
is the same as the previous examples.
[0253] FIGS. 12A, 12B, 12C, 12D, and 12E provide illustrations of
mechanisms, modifications and methods of charging an agent carrier
with agent and/or other substances that assist in the loading,
retention and delivery of agent by the system.
[0254] The loading mechanisms, generally illustrated in FIGS. 12A
to 12E, may also be used alone, or in combination, as methods for
lining the surface of the agent carrier or its cavities with
hydrophilic or hydrophobic moieties prior to loading an agent, or
with moieties that can conduct electric charges and/or participate
in generating or propagating electric fields prior to loading an
agent.
[0255] FIG. 12A provides an illustration of an embodiment of a
method for charging an agent carrier with an agent. In this
embodiment, the applicator tip 900 containing the agent carrier
body 902 is connected to a hand-held applicator device (not shown)
via its coupling rod 908. The agent carrier body 902 is at least
partially immersed in a container 904 containing an agent 906.
Ultrasonic vibration created by an ultrasonic transducer of the
applicator device is coupled, via the coupling rod 908 to the
applicator tip 900, and through it, to the agent carrier body 902.
The vibration expels air from the micro channels and at least
partially fills the micro channels and/or agent reservoirs within
the agent carrier body 902 with agent 906.
[0256] FIGS. 12B provides an illustration of another embodiment of
a method for charging an agent carrier with an agent. In this
embodiment, the agent carrier is a removable applicator tip 900'.
The applicator tip 900' and/or a separate agent carrier body 903
are at least partially immersed in a container 904' containing an
agent 906'. Ultrasonic vibration created by an external source 910
is applied to the container 904', which expels air from the micro
channels and/or agent reservoirs of the agent carrier contained in
the applicator tip 900' (not shown) and/or the separated agent
carrier body 903 and at least partially fills the micro channels
and/or agent reservoirs of the agent carrier within the applicator
tip 900'and/or the separated agent carrier body 903 with agent
906'. In other embodiments loading may be performed by simple
immersion of the agent carrier or agent carrier body without
application of ultrasonic vibration.
[0257] FIG. 12C provides an illustration of a vacuum chamber 912.
Vacuum is applied at the port 914 to remove air from the chamber
912 and the air within the micro channels and/or agent reservoirs
of an agent carrier held within an applicator tip 900'' or a
separated agent carrier body 903'. When the vacuum is complete, a
valve controlling the agent entry port 916 is opened so that agent
stored in chamber 917 is drawn into the chamber 912 through the
agent entry port 916 and into the micro channels and/or agent
reservoirs in the agent carrier body 902'' in the applicator tip
900'' and/or the separated agent carrier body 903'. Ingress of
agent occurs via the pores in the tissue-contact surface of the
agent carrier(s). Once charged with agent, the applicator tip 900''
and/or the separated agent carrier body 903' is removed from the
agent containing fluid and a seal layer may be applied over exposed
surfaces.
[0258] FIG. 12D provides another embodiment of a method in which a
vacuum is used to charge an agent carrier body 903''' with agent
906'''. Agent 906''' is held within a container 904'''. The agent
carrier 903''' is placed within the container 904''' and at least
partially submerged so that the pores of the tissue contact surface
920 of the agent carrier body 903'' are in the agent solution
906'''. A vacuum is applied to port 918 to draw agent solution up
through the micro channels in the agent carrier 903''' so that the
micro channels and/or agent reservoirs are at least partially
filled with the agent solution 906'''.
[0259] In an alternative embodiment of a method for charging an
agent carrier body with agent, an agent can be directly injected
into the port so that the air in the agent carrier (i.e. in the
micro channels and/or agent reservoirs) is expelled and replaced by
the agent.
[0260] FIG. 12E provides a similar method to that in FIG. 12D
except an applicator tip 900'''' having an agent carrier body
902''' is to be charged with agent. The applicator tip 900'''' is
illustrated in cross section to illustrate that the applicator tip
includes a reservoir 921 within its housing that is separate from
any reservoir formed within the agent carrier body 902'''. The
applicator tip 900'''' includes a vacuum port 922 that provides
access to the reservoir 921. As above, a vacuum is applied at the
vacuum port 922 which draws agent solution up through the micro
channels in the agent carrier body 902''' so that the micro
channels and/or agent reservoirs in either the agent carrier body
902''' or applicator tip's 900'''' housing are at least partially
filled with the agent solution 906''''.
[0261] In an alternative embodiment of a method for charging an
agent carrier or applicator tip having an agent carrier with agent,
agent can be directly injected into a port so that the air in the
agent carrier (e.g. in the micro channels and/or agent reservoirs)
is expelled and replaced by the agent.
[0262] As will be appreciated, the loading techniques described
above can be used with suitable micro-channel, hybrid or protrusion
based agent carrier bodies described herein or devised. However,
agent carrier bodies or agent carriers which permit direct access
to an agent reservoir may be loaded by directly placing agent into
the reservoir, e.g. by pipetting the agent onto the reservoir. One
example of such a mechanism was used in the experiments described
below. In this example the agent was pipetted into the void on the
tissue contacting surface of the agent carrier body of a
protrusion-based agent carrier body. In a similar manner, agent may
be pipetted to a reservoir on the back of the agent carrier body
for delivery via micro channels to the tissue contacting
surface.
[0263] The agent carrier may be provided as either empty agent
carriers or as charged agent carriers that are filled with an
agent. Where empty agent carriers are provided, an end user will
need to charge the agent carrier with agent prior to use.
[0264] The invention also relates to a method of charging the agent
carrier with an agent and discharging agent from the agent
carrier.
[0265] The method of discharging agent from the agent carrier or
dispensing agent to a tissue surface includes applying the agent
carrier to a tissue surface and dispensing agent from the agent
carrier to the tissue surface. Preferably the process of dispensing
the agent includes applying ultrasonic waves to the tissue surface
to facilitate penetration of the agent into the tissue through
sonophoresis.
[0266] As will be appreciated from the foregoing the agent carrier
or an agent carrier body itself can be an item separable from the
applicator device. In a preferred form the agent carrier or agent
carrier body is a single use item that is removable or
interchangeable. This aids in the sterility required for medical
usage and facilitates among other things cleaning and sterilising
of the hand-held applicator device between patients. The solid
physical nature of the preferred embodiments facilitates mounting
and handling of the agent carrier in circumstances where they are
replaceable. Moreover, the use of a solid material for the agent
carrier body to contain the agent facilitates loading of an agent
into an agent carrier, packaging, handling of agent carrier bodies
pre-loaded with agent. Importantly, the use of solid materials for
the agent carrier body facilitate the propagation of ultrasonic
waves that are used to move an agent through the agent carrier and
enhances and/or permits the entry of an agent into the target
tissue by sonophoresis.
[0267] FIGS. 13A and 13B illustrate one embodiment agent carrier.
The applicator tip 1300 is generally speaking equivalent to the
agent carrier tip 102 shown in FIG. 1. In this example the agent
carrier 1300 takes the form of an applicator tip with a removable
and interchangeable agent carrier body.
[0268] The agent carrier 1300 includes the following main
components: An agent carrier body 1302, and a tip housing 1303 that
includes a tip body 1304 and an agent carrier body retaining cap
1306.
[0269] The agent carrier body 1302 is generally rectilinear in plan
view, and in this example it is square. The agent carrier body 1302
may be made in accordance with any one of the examples given above
or aspects described herein. The agent carrier body 1302 has a
tissue contacting surface 1304.
[0270] The tip body 1304 serves to both connect the agent carrier
1300 to an agent applicator device and conduct transmission
stimulus, in the form of ultrasonic energy to the agent carrier
body 1302. To achieve this, the tip body 1304 is provided, on a
first end thereof, with a mounting mechanism 1305 in the form of a
screw thread. The mounting mechanism 1305 is used to make a
mechanical connection with a corresponding connector of a handle
assembly. The second end of the tip body 1304 is shaped to operate
as a horn to conduct ultrasonic energy, via mating surface 1307, to
the agent carrier body 1302.
[0271] The agent carrier body retaining cap 1306 serves to retain
the agent carrier body 1302 and hold it in contact with the mating
surface 1307. The agent carrier body retaining cap 1306 has an
aperture 1310 formed in it, through which the tissue contacting
surface 1308 of the agent carrier body 1302 is exposed in use. The
agent carrier body retaining cap 1306 is mounted to the tip body
1304 using a screw thread.
[0272] As will be appreciated there are many morphological and
mechanical variations can be made in such a system. For example the
shape of the components, including the agent carrier body, and its
associated tissue contacting surface may be varied. The present
square embodiment is particularly convenient when the agent carrier
body is made from a semiconductor material and its manufacturing
process most conveniently outputs square components. The shape of
the tip body can be varied to optimise transmission of ultrasonic
energy if ultrasonic energy is used as a transportation stimulus.
The shape of the aperture thorough which the tissue contacting
surface of the agent carrier body is exposed can be varied. In some
cases it may differ from the shape of the tissue contacting surface
of the agent carrier body.
[0273] The method of engagement of the agent carrier retaining cap
with the tip body can be varied widely to use any convenient type
of mechanism. In this example engagement is by screw thread,
however the agent carrier retaining cap could be press fit onto the
tip body, or engaged with snap fasteners, or a bayonet fitting, to
give a non-exhaustive list or alternatives. Similarly the mounting
mechanism of the agent carrier body can be varied to use any known
coupling mechanism.
[0274] An agent carrier having a plurality of agent carrier bodies,
perhaps arranged in a pattern such as an array, could also be
provided.
6. Trial Results and Treatment Methodologies
[0275] In a further aspect of the present invention there is
provided methods for delivering an agent to a living tissue, e.g.
animal, plant or human. It is considered that by selectively
choosing the operational parameters of the non-invasive agent
applicator presently described, the amount of agent delivered to a
selected depth within tissue may be controlled.
[0276] The controlled parameters may include one or more of: [0277]
Application pressure; [0278] Ultrasonic frequency; [0279]
Ultrasonic waveform; [0280] Ultrasound direction; [0281] Ultrasonic
power level; [0282] Ultrasonic application duration; and [0283]
Ultrasound duty cycle.
[0284] In particularly preferred forms, the system parameters can
be chosen to control the delivery of an agent to a desired depth in
the target tissue. An example of this would be setting system
parameters such that transepithelial delivery of an agent
predominantly into the stroma of the cornea would occur.
Advantageously, this presents the opportunity to deliver a drug,
vaccine or other agent to a selected tissue depth where it is known
to be most efficacious.
[0285] In this regard, the present invention in one form provides a
method of controlling the amount of agent delivered to a selected
depth within tissue using an agent carrier, agent carrier body or
applicator of any one of the aspects or embodiments described
herein.
Experimental Testing
[0286] A series of experiments were conducted using mice to
determine if an agent can be successfully delivered to tissues
using embodiments of the present invention. The transportation
stimulus in each case was ultrasonic energy only. In the present
experiments a vaccine virus was administered using an embodiment of
the present invention, using ultrasonic energy only, applied to the
inside of the lip to determine whether the agent was presented to
the immune system, and may induce an immune response. Researchers
noted that no damage occurred to the mucous membrane of the lip by
the application of the device for the period required to achieve a
systemic immune response in Experiment 2.
6.1 Experimental Summary
Experiment 1
[0287] Mice were vaccinated with an embodiment of the present
invention illustrated in FIG. 7c using two agent carrier bodies
(termed "microchips" in this discussion) totalling around
2-5.times.106 plaque forming units (pfu) of the fluorescent
labelled recombinant poxviral vector-based HIV vaccine per
mouse.
[0288] The proportion of antigen presenting cells taken up the
vaccine antigen (0.025-0. 068 vs 0.025-0.022), and the proportion
of dendritic cells recruited to the draining lymph nodes (0.25-0.54
vs 0.22-0.49) were similar in immunised and unimmunised mice,
respectively (FIGS. 1 and 2). The key conclusion was that an immune
response was not instigated using only two microchips.
Experiment 2
[0289] A full heterologous prime-boost vaccination using
recombinant poxviruses expressing HIV antigens was conducted using
three microchips per mouse prime, and the responses were compared
to mice primed intranasally (i.n.) (positive control), and to mice
not primed with any vaccine (negative control). All mice were given
an intramuscular (i.m.) booster vaccination two weeks after the
priming vaccination.
[0290] The magnitude of the systemic immune responses (responses in
the blood compartment) induced by different vaccination routes were
evaluated by determining the percent of HIV-specific CD8 T cells in
spleen. One of the mice vaccinated using an embodiment of the
present invention generated an immune response that exceeded the
positive control thus demonstrating proof of concept.
Experiment 3
[0291] In a further experiment a preliminary prime-boost
vaccination experiment was conducted using embodiments of the
present invention illustrated in FIG. 10. Mice were primed with the
lip delivery system using three microchips according to each
embodiment (around 2-5.times.10.sup.6 pfu) of FPV-HIV per mouse,
followed by an intramuscular booster vaccination. The percent of
HIV-specific CD8 T cells was used to assess the magnitude of the
immune responses induced. Data indicated that microchips 1 (1% of
cells) and 2 (0.6%) performed slightly better than microchips 3 and
4 (0.5%). It was also noted that during loading and delivery the
microchips 1 and 2 performed much more effectively than microchips
3 and 4.
Experiment 4
[0292] Full prime-boost vaccination experiment was performed using
the microchips 1 and 2 of FIG. 10. In this experiment one of the
mice in each of the groups vaccinated generated an immune response
that exceeded the intranasal positive control, whereas the other
two mice in each group had responses similar to the oral vaccine
negative control group.
[0293] Table 1 summarises the experimental parameters and outcomes
of each of the experiments.
TABLE-US-00002 TABLE 1 Summary of the prime-boost vaccination
experiments conducted on the original microchip and microchips 1
and 2. Chip % HIV-specific Magnitude of HIV- identification
Priming: Booster: CD8+ T cells Specific CD8+ T cell where route
dose route dose (tetramer test).sup.d response (ICS test).sup.d
relevant.sup.a FPV-HIV.sup.b VV-HIV.sup.c M#1 M#2 M#3 M#1 M#2 M#3
Original Lip i.m. 15.1 1.03 1.06 10.5 0.73 0.78 Mc (x3) -2.5
.times. 10.sup.6 pfu 1 .times. 10.sup.7 pfu Test group Positive
i.n. i.m. 8.94 9.33 6.85 6.14 control 1 .times. 10.sup.7 pfu 1
.times. 10.sup.7 pfu Negative i.m. 1.36 1.40 1.03 0.76 control 1
.times. 10.sup.7 pfu Mc1 (x3) Lip i.m. 0.38 15.5 0.67 0.06 1.5 0.08
Test group -2.5 .times. 10.sup.6 pfu 1 .times. 10.sup.7 pfu Mc2
(x3) Lip i.m. 0.81 0.73 9.45 0.12 0.08 2.0 Test group -2.5 .times.
10.sup.6 pfu 1 .times. 10.sup.7 pfu Negative Oral i.m. 1.17 0.45
2.87 0.8 0.05 0.35 control 5 .times. 10.sup.6 pfu 1 .times.
10.sup.7 pfu .sup.a(x3)--refers to the number of microchips of
vaccine administered to each mouse, thus "x3" means that three
microchips were applied; Mc--is an abbreviation of "microchip" and
us used to designate which type was used in each test; .sup.bDose,
is represented in plaque forming units (pfu) of the priming
vaccine, fowl pox virus expressing HIV antigens (FPV-HIV) are
provided. The route of vaccination delivery; is indicated as
follows: "Lip" designates that administration was made using an
embodiment of the present invention applied to the tissues of the
lip of the subject; "i.n." designates intranasal delivery; "oral"
designates delivery directly into the mouth .sup.cThe booster
vaccine is vaccinia virus expressing HIV antigens (VV-HIV); and in
all cases this was delivered using intramuscular (i.m.) route
.sup.dIn both cases, systemic immune response was investigated. M#
represents mouse number.
6.2 Experimental Detail
Experiment 1
[0294] Aims: To determine whether the lip delivery system using the
embodiment of FIG. 7c induced antigen uptake in the draining lymph
nodes (LN), the antigen antigen presentation and immune cell
recruitment was monitored 24 hours post vaccination as follows:
[0295] 1. Uptake of the vaccine antigens was monitored in cervical,
mediastinal and/or mesenteric lymph nodes following administration
of a number of microchips of a fluorescently labelled vaccine,
recombinant fowl pox virus expressing HIV antigens together with
green fluorescent protein (FPV-HIV-GFP);
[0296] 2. Evaluate whether antigen presenting cells (APC) are
recruited to these LN the relative number of dendritic cells (DCs)
and macrophages at these sites were identified by the staining for
characteristic cell surface markers
Methods
[0297] 1. Mice were immunised with FPV-HIV-GFP and responses were
evaluated 24 hours post vaccination. In these experiments, mice
were also kept as either [0298] a) unimmunised controls (FIGS. 14
and 15), or [0299] b) controls vaccinated with only FPV-HIV (i.e.
no GFP fluorescent antigen, FIG. 16).
[0300] Mice were given the vaccination with two microchips, one to
the left and one to the right lip (around 5.times.10.sup.6 pfu per
mouse).
[0301] 2. At 24 h the different draining lymph nodes were
harvested, pooled, and single cell suspensions were prepared in
complete medium (Ranasinghe et al., 2011, Ranasinghe et al., 2006,
Ranasinghe et al., 2007, Ranasinghe et al., 2013) 3.
1.times.10.sup.6 cells were aliquoted and stained with the
different cell surface markers. [Antigen presenting MHC-II cells
were stained with antibody to the I-A.sup.d APC cell surface
marker
[0302] Antibodies to cell surface markers CD11b-PE and CD11c-PerCP
were used to identify DCs, (FIG. 15) and antibody to cell surface
marker F4/80-PE Cy7 was used to identify macrophages (data not
shown)] (Ranasinghe et al 2103)
[0303] 4. Different cell subsets were analysed based on the
fluorescent-labelled cell surface marker expressed on the cell
surface using flow cytometry analysis (FACS). These experiments
were repeated three times, combined results are presented in FIGS.
14 to 16.
[0304] 5. In these experiments singe colour controls (SS) and
fluorescent minus one (FMO) controls were also used to set up the
gating and perform the correct analysis of the different cell
subsets.
Results and Conclusions
[0305] FIGS. 14 to 16 illustrate graphically the outcomes of the
experiments. In this regard, FIG. 14 shows plots for the evaluation
of the uptake of FPV-HIV-GFP vaccine 24 h post lip delivery,
illustrating I-A.sup.d APC MHC-II cells containing the fluorescent
GFP antigen of the vaccine detected in the top right hand quadrant
indicated by the arrow. Note in this and other FACS plots, each dot
represents a single cell.
[0306] FIG. 15 illustrates plots for the evaluation of recruitment
of antigen uptake by different dendritic cell subsets to the
respective draining lymph nodes 24 h post lip delivery. The
proportion of dendritic cells, identified as being MHC-11+, and
either CD11b+ (left two columns) or CD11c+ (right two columns) are
indicated in the top right hand quadrant (refer to arrows).
[0307] FIG. 16 illustrates plots for the evaluation of the uptake
of FPV-HIV-GFP vaccine 24 h post lip delivery in cervical,
mediastinal and mesenteric nodes (repeat experiment 3) I-Ad APC
MHC-II cells containing the fluorescent GFP antigen of the vaccine
are detected in the top right hand quadrant indicated by the arrow.
(Note that the top three graphs show the gating strategy).
[0308] As can be seen, no differences in the antigen uptake and
presentation (FIGS. 14 & 16) or the DC subsets recruited to the
draining lymph nodes (FIG. 15) were detected between the mice that
received the FPVHIV-GFP vaccine and the controls. The data
indicated that;
[0309] i) Vaccine delivery applied at a dose of two microchips per
mouse (dose .about.2-5.times.10.sup.6 pfu) was not effective.
[0310] ii) Thus, to obtain any immune outcomes, a minimum of 3
chips or more per mouse were used in the subsequent prime-boost
vaccination experiments.
Experiment 2
[0311] In this next experiment an evaluation of the efficacy of lip
delivery with the same microchip as experiment 1, using prime-boost
vaccination was performed.
[0312] Aims: To test whether lip prime followed by intramuscular
(i.m.) booster vaccination can induce effective HIV-specific CD8 T
cell immunity compared to intranasal prime (i.n.)/i.m. booster
vaccination strategy using:
[0313] 1. HIV gag-specific tetramer staining.
[0314] 2. Intracellular cytokine staining (ICS) of IFN-.gamma. in
HIV-specific CD8 T cells.
Methods
[0315] FIG. 17 are photographs showing the following phases of the
experiments performed. The phases illustrated include: Loading the
microchips (top left), Ultrasonic system settings (top right) and
lip delivery to the mice (bottom photos). The experimental method
was performed as follows.
1) Priming Vaccination with FPV-HIV
[0316] a. Vaccine (.about.600-800 .mu.l of the stock) was sonicated
(i.e. output: 30%; 3 cycles for 10 seconds per cycle) as for
routine i.n. delivery. 300-400 .mu.l/well of the sonicated virus
was added into two wells of a 48 well plate.
[0317] b. Microchips were soaked in FPV-HIV (5.times.108 PFU/ml) in
a 48 well plate (FIG. 17 top left). It was assumed that each
microchip could absorb and expel 5 .mu.l, thus the dose per
microchip was calculated to be 2.5.times.10.sup.6 pfu.
[0318] c. Six microchips per well were submerged in liquid without
any overlap and incubated for 30 minutes on ice (FIG. 17 top
left).
[0319] d. One microchip was taken out to test whether the chips
were loaded with virus, by placing the loaded microchip in a well
containing PBS. If the microchip floated it meant the chip was not
loaded, but if it sank it was considered to be loaded.
[0320] e. Controls: positive control two mice were immunised i.n.
(20 .quadrature.l/mouse 1.times.10.sup.7 pfu) and two mice were
vaccinated with i.m. booster (1.times.10.sup.7 pfu) only to solely
test its effect.
[0321] f. Test group: three mice were immunised for the lip/i.m.
group as follows. The microchip was mounted to the delivery
applicator, similar to that illustrated in FIG. 1, that was
connected to the power source. Ultrasonic gel was used between the
arm and the microchip for better contact). Power was switched
on.
[0322] g. Microchip was pressed firmly onto the inner lip region of
an anesthetised mouse. (FIG. 17 bottom)
[0323] h. Output switch was turned on an ultrasonic energy was
applied for 30 seconds, to deliver the virus into the lip region.
At this time point the instrument settings were transducer drive
voltage V=95-160; P=2800-3200 mW.
[0324] i. To check whether the virus has been delivered from the
microchip, the chip was placed in PBS as before. If the chip
floated it suggested that the virus was successfully expelled from
the chip. 80% of the time the chip floated suggesting that the
vaccine was expelled. If two microchips failed to deliver the
vaccine correctly, the mouse was discarded and a new mouse was
immunised.
[0325] j. This was repeated for 3 microchips per mouse, using one
new chip each time.
2) Intramuscular Booster Vaccination Using 10.sup.7 PFU VV-HIV
[0326] a. Booster vaccination was performed two weeks post FPV-HIV
priming vaccination
[0327] b. Booster vaccine was prepared for 9 mice total
9.times.10.sup.7 PFU in 900 .mu.l of PBS.
[0328] c. Virus was sonicated exactly as done for the FPV-HIV.
[0329] d. Mice were anesthetized with isoflurane using a nose cone
and 50 .mu.l of VV-HIV per quadriceps muscle was delivered i.m.
3) Preparation of Spleen Samples for Analysis
[0330] 7-14 days post booster vaccination spleens were harvested
from each mouse, and single cell suspensions were prepared as
described in Ranasinghe et al (2006).
[0331] The magnitude of the HIV-specific CD8 T cell responses was
assessed with tetramer staining and intracellular cytokine
staining, using 4.times.106 spleen cells from each mouse according
to the plate scheme in Tables 2 and 3 as follows:
[0332] a. Tetramer staining was performed as described in
(Ranasinghe et al., 2011, Ranasinghe et al., 2006, Ranasinghe et
al., 2007, Ranasinghe et al., 2013) [0333] Cells were stained for
45 min at room temperature with K.sup.dGag197-205-APC tetramer and
anti-CD8a FITC in FACS buffer. [0334] Cells were washed and fixed
in 0.5% PFA prior to analysis using FACS. [0335] b. Intra cellular
cytokine staining (ICS) for IFN-.gamma. was also performed as
described (Ranasinghe et al., 2011, Ranasinghe et al., 2006,
Ranasinghe et al., 2007, Ranasinghe et al., 2013) [0336] Cells were
stimulated over night with K.sup.dGag197-205 peptide for 1 h at
37.degree. C. +5% CO2 [0337] Brefeldin A was added to each well and
incubated for further 5 hours at 37.degree. C. [0338] Cells were
surface stained for 25 mins at 4.degree. C. with anti-CD8a FITC in
FACS buffer. [0339] Cells were fixed/permeabilized using IC/fix and
IC/perm from eBioscience [0340] Cells were then intracellular
stained with anti-IFN-.gamma., for 25 mins at 4.degree. C. (Table
2) [0341] Positive stain--anti IFN-.gamma. APC in in IC Perm [0342]
Single colour controls and FMO's.
TABLE-US-00003 [0342] TABLE 2 Plate Scheme for Tetramer Staining. 1
2 3 4 5 6 7 8 9 A ss Unstain FITC APC cont B LIP 1 LIP 2 LIP 3 i.n.
i.n. Boost Boost FMO FMO 1 2 only only CD8 tetramer 1 2
TABLE-US-00004 TABLE 3 Plate Scheme for ICS. 1 2 3 4 5 6 7 8 11 A
ss Unstain FITC APC cont B LIP 1 LIP 2 LIP 3 i.n. i.n. Boost Boost
FMO FMO stimulated 1 2 only only CD8 IFN-g 1 2 C LIP 1 LIP 2 LIP 3
i.n. i.n. Boost Boost Unstim 1 2 only only 1 2 SS = Single colour
control, FMO = Fluorescent minus one
Results and Conclusions
[0343] FIG. 18 shows plots illustrating the evaluation of the
magnitude of HIV-specific splenic CD8 T cells using IFN-.gamma.
intracellular staining. The FACS data were analyzed using Cell
Quest Pro or FlowJo analysis. The box indicates the percentage of
HIV-specific splenic CD8 T cells expressing IFN-.gamma. following
Lip/i.m. (top 3 mice), i.n./i.m. (middle 2 mice) and booster only
(bottom 3 mice) vaccinations. FIG. 19 illustrates plots enabling
evaluation of HIV-specific splenic CD8 T cells using tetramer
staining. Cells were stained as described in materials and methods.
The FACS data were analysed using Cell Quest Pro or FlowJo
analysis. The box indicates the percentage of HIV-specific splenic
CD8 T cells following different routes of vaccine delivery.
Lip/i.m. (top three mice), i.n./i.m. (middle two mice) and booster
only (bottom two mice).
[0344] The HIV-specific tetramer (FIG. 18) and staining (FIG. 19)
data indicated that unlike the i.n./i.m. delivery strategy that
gave highly consistent results (FIG. 18--range 8.94-9.33%), the
lip/i.m. delivery strategy did not yield consistent outcomes (FIG.
19--range 1.03-15.1%). Whilst it appears that this is due to the
inconsistency of the priming of the mice during lip delivery (Note:
see also lip/i.m. compared to i.m. booster only), one mouse (mouse
1) showed an immune response that exceeded that of the i.n./i.m.
delivery strategy, indicating that a response is possible using
embodiments of the present invention.
[0345] Data also revealed that 3.times. lip or 4.times. lip
microchip delivery was more effective than 5.times. lip microchip
delivery (data not shown). These experiments were performed twice
and data were found to be very similar between the experiments
(Experiments 4 & 5). Data are representative of one
experiment.
Experiments 3
[0346] A further experiment was performed to test prime-boost
vaccination strategy to assess the efficacy of lip delivery using
four protrusion-based embodiments of the present invention
illustrated in FIG. 10.
[0347] Aims: To test whether these microchips can load and deliver
the vaccine more effectively to the lip compared to microchips of
FIG. 7c using HIV gag-specific tetramer staining (FIG. 20).
1) Priming Vaccination with FPV-HIV
[0348] a. Vaccine was sonicated and 300-400 ml per well was added
into a 48 well plate as before.
[0349] b. The microchips were connected to the device, then 5-7
.mu.l of vaccine was loaded onto the tissue contacting surface of
the microchip using a pipette and immediately delivered to the lip
of the mouse. Unlike the microchip of FIG. 7c, these improved
microchips were NOT soaked in FPV-HIV for 30 min.
[0350] c. Controls: for the positive control, two mice were
immunised i.n. (20 ml/mouse); for the negative controls, two mice
were immunised orally and two mice were kept as controls for the
i.m. booster only to test the effect of i.m. vaccination only.
(similar to FIG. 5)
2) i.m Booster Vaccination and Evaluation of Immune Responses Using
Tetramer Staining
[0351] a) These were performed exactly as described in experiment
2.
Results and Conclusions
[0352] 1) Unlike the microchip of FIG. 7c, direct pipetting of the
vaccine onto the chips made it extremely easy to determine whether
the new microchips were properly loaded. Similarly, once the
vaccination was performed, the microchip was placed on a piece of
tissue to determine whether the vaccine had been properly expelled.
If the microchip was dry it meant the vaccine was delivered. We
also tested the above loading by visualising the empty, loaded and
used microchips under a microscope.
[0353] 2) It was observed that microchips 1 & 2 (FIG. 10 top)
loaded and discharged the vaccine much more effectively (without
leakage) compared to microchips 3 and 4 (FIG. 10 bottom). Even
though loading was much more effective, the vaccine leaked out of
microchip 3 (in particular) and 4 as soon as the device was held
against the lip, prior to turning on the output switch, making it
more of an oral delivery.
[0354] 3) The preliminary HIV-specific tetramer data further
confirmed that microchip 1 performed better than 3 & 4. Hence,
it was decided to repeat the prime-boost vaccination experiments
with microchips 1 and 2 of FIG. 10, including an oral prime/i.m.
booster immunization strategy as a control to validate the data in
experiment 4, below.
Experiment 4
[0355] In this experiment vaccination using a 3.times. lip/i.m.
vaccination strategy using microchips 1 & 2 was tested in a
similar manner to previous experiments.
[0356] Aim: Test the efficacy 3.times. lip/i.m, vaccination
strategy compared to lx oral/i.m. prime-booster vaccination using:
[0357] a) HIV gag-specific tetramer staining (FIG. 21) and [0358]
b) Intracellular cytokine staining (ICS) of IFN-.gamma. (FIG.
22)
Methods
[0359] Vaccination and analysis were performed exactly as in
experiment 3 with 3 mice per group. 1.times. oral prime/i.m.
booster vaccination was also performed as an additional control to
assess whether the priming was related to oral delivery or lip
delivery (oral dose=5.times.106 FPV-HIV). The HIV-specific CD8 T
cell responses were measured in the spleen 14 days post booster
vaccination using tetramer staining and intracellular IFN-.gamma.
staining. The experiments were performed two times.
Results and Conclusion
[0360] FIG. 21 illustrates plots enabling evaluation of
HIV-specific splenic CD8 T cell responses using tetramer staining.
The FACS data were analysed using Cell quest Pro software. Plots
represent three animals per group microchip 1 (top) & 2
(middle) prime-boost immunization data compared to oral delivery
(bottom). The upper right quadrants (red arrows) indicate the % of
HIV-specific CD8 T cells observed following each vaccine
strategy.
[0361] FIG. 22 illustrates plots enabling evaluation of the
magnitude of HIV-specific CD8 T cell responses using IFN-.gamma.
intra cellular cytokine staining. The FACS data were analysed using
Cell quest Pro software. Plots represent three animals per group
microchip 1 (top) & 2 (middle) prime-boost immunization data
compared to oral delivery (bottom). The upper right quadrants (red
arrows) indicate the % of HIV-specific CD8 T cells expressing
IFN-.gamma..
[0362] As can be seen the HIV-specific splenic CD8 T cell responses
observed with microchip 1--mouse 2 and microchip 2--mouse 3 (red
arrows) were greatly elevated compared to oral delivery (bottom 3
mice FIGS. 21 & 22), these results clearly indicated that the
responses observed were due to lip uptake not oral uptake.
[0363] Data indicated that if the delivery was uniform/consistent
the microchip 1 and 2 could induce good HIV-specific CD8 T cell
immunity in the blood compartment.
[0364] The positive responses detected with the microchips made in
accordance with FIG. 10 were very much similar to the positive
responses detected with the microchip of FIG. 7c used in
experiments 1 and 2). However, they present greater ease of
loading.
[0365] Data from experiments, suggest that if
uniformity/consistency could be attained, lip delivery could be
more effective than oral or intranasal delivery.
Discussion
[0366] Molecules that are known to the inventors to possibly be
delivered to the body using sonophoresis include 1) molecules of
any kind of electric charge or have a neutral (including overall
neutral) electrical charge and 2) small or large molecules
(including monoclonal antibodies of approximately 149,000 Daltons)
3) molecules that are hydrophilic or hydrophobic or lipophilic.
7. REFERENCES
[0367] References: RANASINGHE, C., EYERS, F., STAMBAS, J., BOYLE,
D. B., RAMSHAW, I. A. & RAMSAY, A. J
[0368] 2011. A comparative analysis of HIV-specific
mucosal/systemic T cell immunity and avidity following rDNA/rFPV
and poxvirus-poxvirus prime boost immunisations. Vaccine, 29,
3008-20
[0369] RANASINGHE, C., MEDVECZKY, J. C., WOLTRING, D., GAO, K.,
THOMSON, S., COUPAR, B
[0370] E. H., BOYLE, D. B., RAMSAY, A. J. & I.A., R. 2006.
Evaluation of fowlpox-vaccinia virus prime-boost vaccine strategies
for high-level mucosal and systemic Immunity against HIV-1.
Vaccine, 24, 5881-5895
[0371] RANASINGHE, C., TRIVEDI, S., STAMBAS, J. & JACKSON, R.
J. 2013. Unique IL-13Ralpha2-based HIV-1 vaccine strategy to
enhance mucosal immunity, CD8(+) T-cell avidity and protective
immunity. Mucosal Immunol, 6, 1068-80
[0372] RANASINGHE, C., TURNER, S. J., MCARTHUR, C., SUTHERLAND, D.
B., KIM, J. H., DOHERTY, P. C. & RAMSHAW, I. A. 2007. Mucosal
HIV-1 pox virus prime-boost immunization Induces high-avidity CD8+T
cells with regime-dependent cytokine/granzyme B profiles. J
Immunol., 178, 2370-9
[0373] It will be understood that the invention disclosed and
defined in this specification extends to all alternative
combinations of two or more of the individual features mentioned or
evident from the text or drawings. All of these different
combinations constitute various alternative aspects of the
invention.
* * * * *