Note: Descriptions are shown in the official language in which they were submitted.
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THERAPEUTIC POLYMERS AND METHODS OF USE
RELATED APPLICATIONS
[0001] This application relies for priority under 35 U.S.C. 119(e) on U.S.
Serial Nos.
60/687,570, filed June 3, 2005 and 60/759,179, filed January 13, 2006, each of
which is
incorporated herein by reference.
FIELD OF THE INVENTION
[0002] The invention relates, in general, to drug delivery systems and, in
particular, to
polymer delivery compositions that incorporate a therapeutic agent into a
biodegradable
polymer backbone.
BACKGROUND INFORMATION
[0003] The earliest drug delivery systems, first introduced in the 1970s, were
based on
polymers formed from lactic and glycolic acids. Today, polymeric materials
still provide
the most important avenues for research, primarily because of their ease of
processing and
the ability of researchers to readily control their chemical and physical
properties via
molecular synthesis. Basically, two broad categories of polymer systems, both
known as
"inicrospheres" because of their size and shape, have been studied: reservoir
devices and
niatrix devices. The former involves the encapsulation of a pharmaceutical
product within
a polymer shell, whereas the latter describes a system in which a drug is
physically
entrapped within a polymer network.
[0004] The release of medications from either category of polymer device
traditionally
has been diffusion-controlled. Currently, however, modem research is aimed at
investigating biodegradable polymer systems. These drug deliverers degrade
into
biologically acceptable compounds, often through the process of hydrolysis,
and leave
their incorporated medications behind. This erosion process occurs either in
bulk (wherein
the matrix degrades uniformly) or at the polymer's surface (whereby release
rates are
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related to the polymer's surface area). The degradation process itself
involves the
breakdown of these polymers into lactic and glycolic acids. These acids are
eventually
reduced by the Kreb's cycle to carbon dioxide and water, which the body can
easily expel.
[0005] Regular AA-BB type amino acid based bio-analogous poly(ester amides)
(PEAs) and poly(ester urethanes) (PEURs) consisting of nontoxic building
blocks, such as
hydrophobic a-amino acids, a,w-diols, and aliphatic dicarboxylic acids have
been
investigated as biomaterials for drug release and tissue engineering
applications (G.
Tsitlanadze et al. J. Biomater. Sci. Polymer Edfz, (2004) 15: 1-24). The
combination of
controlled enzymatic degradation and low nonspecific hydrolysis rates of PEAs
and
PEURs make them attractive for drug delivery applications. In particular,
these polymers
appear to be blood and tissue compatible witll advantageous properties for
cardiovascular
applications (K. DeFife et al. Transcatheter Cardiovascular Therapeutics - TCT
2004
Conference. Poster presentation. Washington DC. (2004)).
[0006] In most drug-eluting applications, the drug is physically matrixed by
dissolving
or melting with a polymer. Another approach has also been reported in which a
drug is
chenlically attached as a side group to a polymer.
[0007] If a drug or other therapeutic agent is covalently incorporated into a
biodegradable polymer, a tlierapeutic polymer is formed. Such compositions
represent
synthetic polymers that combine therapeutic or palliative bioactivity with
desirable
mechanical and physical properties, and degrade into useful therapeutic active
coinpounds.
In other words, the compositions have the activity of a drug, but have the
physical
properties of a material. Recently, new therapeutic polyesters, polyamides,
and poly(ester
anhydrides) were reported, wherein non-steroidal anti- inflammatory drugs
(NSAIDs)
were incorporated into a polymer backbone (R.C. Schmeltzer et al.
Biofraacromolecules.
(2005) 6(1):359-367). In such compositions, drug release is directly dependent
on the
hydrolytic or enzymatic cleavage of polymer-drug binding groups. One of the
advantages
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of a "backbone as a drug" polymer is that a high amount of drug or therapeutic
compound
can be incorporated into the structure.
[0008] Thus, there is a need in the art for more and better compositions and
methods
for incorporating therapeutic molecules, such as drugs and other bioactive
agents, into the
backbones of polymers for use in polymer delivery systems in which controlled
rate of
therapeutic release is combined with desirable mechanical and physical
properties.
[0009] Finally, recent research has shown that hydrogel-type materials can be
used to
shepherd various medications through the stomach and into the more allealine
intestine.
Hydrogels are cross-linked, hydrophilic, three-dimensional polymer networks
that are
highly permeable to entrap molecules, which can be released in vivo through
their weblilce
surfaces. Depending on the chemical composition of the gel, different internal
and
external stimuli (e.g., changes in pH, application of a magnetic or electric
field, variations
in temperature, and ultrasound irradiation) may be used to trigger the
swelling effect.
Once triggered, however, the rate of entrapped drug release is generally
determined by the
cross-linking level of the polymer network.
[0010] Thus, a need exists in the art for new and better compositions and
methods of
use for biodegradable polymer compositions for delivering therapeutic
molecules, such as
drugs and other bioactive agents. Particularly, the need exists for new and
better delivery
compositions that incorporate a therapeutic agent into the backbone of a
polymer for time
release of the therapeutic agent in a consistent and reliable manner.
SUMMARY OF THE INVENTION
[0011] The present invention is based on the premise that poly(ester amide)
(PEA),
poly(ester urethane) (PEUR), and poly(ester urea) (PEU) polymers, can be
formulated as
polymer delivery compositions that incorporate a therapeutic diol or di-acid
into the
backbone of the polymer for time release of the therapeutic agent in a
consistent and
reliable manner by biodegradation of the polymer.
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[0012] In one embodiment, the invention provides a biodegradable therapeutic
polymer
composition in which at least one therapeutic diol or di-acid is incorporated
into the
baclcbone of one or more biodegradable polymers. The biodegradable polymer of
the
composition contains or is a blend of at least one PEA having a structural
formula
described by structural formula (I),
O 0 H O O H
C-RI-C-N-6-6-O-R4-O-6-6-N
H R3 R3 H
n
Formula (I)
wherein n ranges from about 5 to about 150; R' is independently selected from
residues of
a,co-bis(4-carboxyphenoxy)-(C1-C8) alkane, 3,3'-(alkanedioyldioxy)dicinnamic
acid or
4,4'-(alkanedioyldioxy)dicinnamic acid, (C2 - C20) alkylene, (C2-C20)
alkenylene or
saturated or unsaturated residues of therapeutic di-acids; the R3s in
individual n monomers
are independently selected from the group consisting of hydrogen, (Cl-C6)
allcyl, (C2-C6)
alkenyl, (C2-C6) alkynyl, (C6-Clo) aryl (Cl-C6) allcyl, and -(CH2)2S(CH3); and
R4 is
independently selected from the group consisting of (CZ-C20) alkylene, (C2-
C20)
alkenylene, (C2-C8) alkyloxy (C2-C20) alkylene, bicyclic-fragments of 1,4:3,6-
dianhydrohexitols of structural formula (II), saturated or unsaturated
therapeutic diol
residues, and combinations thereof;
\
CH
HZC; CH2
O J'CH
Formula (II),
except that at least one of Rl and R4 is a therapeutic amount of the residue
of a therapeutic
di-acid and diol, respectively,
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[0013] or at least one PEA polymer having a chemical fonnula described by
structural
formula III:
O 0 H O 4 O H 0 1 0 H
C-R C-N-6-6-0-R -O-C-6-N C-R -C-N-C-(CHa)4 N
3 3 I
H R R H m H C-0-R2 H p
0 fn
Formula (III)
wherein n ranges from about 5 to about 150, m ranges about 0.1 to 0.9: p
ranges from
about 0.9 to 0.1; wherein Rl is independently selected from residues of a,co-
bis(4-
carboxyphenoxy)-(Ci-C8) alkane, 3,3'-(allcanedioyldioxy)dicinnamic acid or
4,4'-
(alkanedioyldioxy)diciimamic acid, (C2 - C20) alkylene, (C2-C20) alkenylene or
a saturated
or unsaturated residues of therapeutic di-acids; each R2 is independently
hydrogen, (Cl-
C12) alkyl or (CG-Clo) aryl or a protecting group; the R3s in individual m
monomers are
independently selected from the group consisting of hydrogen, (Cl-C6) alkyl,
(C2-C6)
alkenyl, (C2-C6) alkynyl, (C6-C10) aryl (C1-C6) alkyl, and
-(CH2)2S(CH3); and R4 is independently selected from the group consisting of
(C2-C20)
alkylene, (C2-CZO) alkenylene, (C2-C8) alkyloxy (C2-C20) alkylene, bicyclic-
fragments of
1,4:3,6-dianhydrohexitols of structural formula (II), residues of saturated or
unsaturated
therapeutic diols and combinations thereof, except that at least one of R' and
R4 in at least
one of the m monomers is the residue of a therapeutic di-acid or diol,
respectively,
[0014] or at least one PEUR having a chemical formula described by general
structural
formula (IV),
1 O O H O O H
C-O-R6-O-C-N-C-6-O-R4-O-C-C-N
H R3 R3 H
n
Formula (IV)
wherein n ranges from about 5 to about 150; wherein R3s in independently
selected from
the group consisting of hydrogen, (C1-C6) alkyl, (C2-C6) alkenyl, (C2-C6)
alkynyl, (C6-Clo)
aryl (C1-C6) alkyl, and -(CH2)2S(CH3) and; R4 is selected from the group
consisting of
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(C2-C20) alkylene, (CZ-C20) alkenylene or allcyloxy, and bicyclic-fragments of
1,4:3,6-
dianhydrohexitols of structural formula (II), or fragments of saturated or
unsaturated
therapeutic diols; and R6 is independently selected from (C2-C20) alkylene,
(C2-C20)
alkenylene or alkyloxy, bicyclic-fragments of 1,4:3,6-dianhydrohexitols of
general
formula (II), residues of saturated or unsaturated therapeutic diols, and
conibinations
thereof, except that the R4 and R6 within at least one of the n unit is the
residue of the
therapeutic diol;
[00151 or at least one PEUR polymer having a chemical structure described by
general
structural formula (V)
11O 0 H O O H 0 0 H
C-O-R6-O-C-N-C-C-0-R4-O-C-C-N ,,~C-O-R6-O-C-N-C-(CH2)4-N
H R3 R3 H H CO-R2 H p
O n
Formula (V)
wherein n ranges from about 5 to about 150, m ranges about 0.1 to about 0.9: p
ranges
from about 0.9 to about 0.1; R2 is independently selected from hydrogen, (C6-
C10) aryl
(C1-C6) alkyl, or a protecting group; the R3s in an individual m monomer are
independently selected from the group consisting of hydrogen, (C1-C6) alkyl,
(C2-C6)
alkenyl, (C2-C6) alkynyl, (C6-Clo) aryl(C1-C6) alkyl, and -(CH2)2S(CH3); R4 is
selected
from the group consisting of (C2-C20) alkylene, (C2-Cao) allcenylene or
alkyloxy, bicyclic-
fragments of 1,4:3,6-dianhydrohexitols of structural formula (II) or fragments
of saturated
or unsaturated therapeutic diols; and R6 is independently selected from (C2-
CZO) alkylene,
(C2-C20) alkenylene or alkyloxy, bicyclic-fragments of 1,4:3,6-
dianhydrohexitols of
general formula (II), a residue of a saturated or unsaturated therapeutic
diols, and
combinations thereof, except that the R4 or R6 within at least one of the m
unit is the
residue of a therapeutic diol,
[0016] or at least one PEU polymer having a chemical formula described by
general
structural formula (VI):
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1O H O O H
C-N-6-6-0-R4-0-C-C-N
H R3 R3 H n
Formula (VI),
wherein n is about 10 to about 150; each R3s within an individual n unit are
independently
selected from hydrogen, (C1-C6) alkyl, (C2-C6) alkenyl, (C2-C6) alkynyl, (C6 -
C10) aryl
(Cl-C6)alkyl, , and-(CH2)2S(CH3); R4 is independently selected from (C2-C20)
alkylene,
(C2-Cao) alkenylene, (C2-C8) alkyloxy (C2-C20) alkylene, a residue of a
saturated or
unsaturated therapeutic diol; or a bicyclic-fragment of a 1,4:3,6-
dianhydrohexitol of
structural formula (II), and combinations thereof, except that the R4 within
at least one of
the n units is the residue of a therapeutic diol;
[0017] or at least one PEU having a chemical formula described by structural
formula
(VII)
O H O O H 0 H
C-N-C-C-O-R4-O-C-C-N C-N-C-(CH2)4-N
H R3 R3H m H C-O-RZ H P "
O
Formula (VII),
wherein m is about 0.1 to about 1.0; p is about 0.9 to about 0.1; n is about
10 to about 150;
each R~ is independently hydrogen, (C1-C12) alkyl or (C6-Clo) aryl; the R3s
within an
individual m monomer are independently selected from hydrogen, (Cl-C6) alkyl,
(C2-C6)
alkenyl, (C2-C6) alkynyl, (C6 -Clo) aryl (C1-C6) alkyl, and -(CH2)2S(CH3);
each R4 is
independently selected from (C2-C2o) alkylene, (C2-C20) alkenylene, (C2-C8)
alkyloxy (C2-
C20) alkylene, a residue of a saturated or unsaturated therapeutic diol; or a
bicyclic-
fragment of a 1,4:3,6-dianhydrohexitol of structural formula (II), and
coinbinations
thereof, except that the R4 in at least one of the m units is the residue of a
therapeutic diol.
[0018] In another embodiment, the invention provides methods for administering
a
therapeutic diol or di-acid to a subject by administering to the subject an
invention
therapeutic polymer composition containing one or more polymers of formula(s)
(I) or
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(III)-(VII) in the form of a liquid dispersion, which composition biodegrades
by enzymatic
action to release the therapeutic diol or di-acid over time.
[0019] In yet another embodiment, the invention provides a bis-nucleophilic
compound
wherein the compound is a di(amino acid)-estradiol -3,17-(i-diester, or salt
thereof.
A BRIEF DESCRIPTION OF THE FIGURES
[0020] Fig. 1 is showing a 'H NMR (500 MHz, DMSO-d6) spectruni of 17p-
estradiol
based monomer (compound 5 of Example 1).
[0021] Fig. 2 is a trace of differential scanning calorimetry (DSC) of the
therapeutic
PEA polymer formed in Example 1, showing a first heating curve, with sharp
melting
endotherm.
[0022] Fig. 3 is showing a 'H NMR (500 MHz, DMSO-d6) spectrum of an invention
17(3-estradiol-based PEA copolymer (scheme 5) formed in Example 1.
DETAILED DESCRIPTION OF THE INVENTION
[0023] The invention is based on the discovery that biodegradable poly(ester
amide)
(PEA), and poly(ester urethane) (PEUR) polymers can be used to create a
therapeutic
polymer composition for in vivo delivery of at least one therapeutic diol or
di-acid
contained within a biodegradable polymer backbone. The therapeutic PEA, PEUR
and
PEU polymer compositions biodegrade in vivo by enzymatic action at the surface
so as to
release therapeutic diols or di-acids from the polymer backbone in a
controlled manner
over time. The invention compositions are stable, and can be lyophilized for
transportation and storage and can be redispersed for administration. Due to
structural
properties of the PEA and PEUR polymers used, the invention therapeutic
polymer
compositions provide for high loading of the therapeutic diol or di-acid, as
well as optional
bioactive agents.
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[0024] As used herein, a "therapeutic diol or di-acid" means any diol or di-
acid
molecule, whether synthetically produced, or naturally occurring (e.g.,
endogenously) that
affects a biological process in a inainmalian individual, such as a human, in
a therapeutic
or palliative manner when administered to the mammal.
[0025] As used herein, the terin "residue of a tlierapeutic di-acid" means a
portion of
such a therapeutic di-acid, as described herein, that excludes the two
carboxyl groups of
the di-acid. As used herein, the teml "residue of a therapeutic diol" means a
portion of a
therapeutic diol, as described herein, that excludes the two hydroxyl groups
of the diol.
The corresponding therapeutic di-acid or diol containing the "residue" thereof
is used in
synthesis of the polymer compositions. The residue of the therapeutic di-acid
or diol is
reconstituted in vivo (or under similar conditions of pH, aqueous media, and
the like) to
the corresponding diol or di-acid upon release from the backbone of the
polymer by
biodegradation in a controlled manner that depends upon the properties of the
particular
PEA, PEUR or PEU polymer selected to fabricate the composition, which
properties are
well known in the art and as described herein, for example in the Examples.
[0026] As used herein the term "bioactive agent" means a bioactive agent as
disclosed
herein that is not incorporated into the polymer backbone. One or more such
bioactive
agents may optionally be dispersed in the invention therapeutic polymer
compositions. As
used herein, the term "dispersed" is used to refer to bioactive agents (not
incorporated into
the polymer backbone) and means that the bioactive agent is dispersed, mixed,
dissolved,
homogenized, and/or covalently bound ("dispersed") in a polymer, for example
attached to
a functional group in the polymer of the composition or to the surface of a
polymer
particle, but not incorporated into the backbone of a PEA or PEUR polymer. To
distinguish backbone-incorporated therapeutic diols and di-acids from those
that are not
incorporated into the polymer backbone, (as a residue thereof), such dispersed
therapeutic
diols and di-acids are referred to herein as "bioactive agent(s)" and may be
contained
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within polymer conjugates or otherwise dispersed in the polymer composition in
the same
manner as other bioactive agents, as described below.
[0027] The term, "biodegradable" as used herein to describe the polymers used
in the
invention therapeutic polymer compositions means the polymer is capable of
being broken
down into innocuous and therapeutic products in the nonnal functioning of the
body. The
polymers in the invention therapeutic polymer compositions include
hydrolyzable ester
and enzymatically cleavable amide linkages that provide biodegradability, and
are
typically chain terminated predominantly with amino groups. Thus, in the case
of a
naturally occurring therapeutic diol or di-acid, the brealcdown product
delivered is the
naturally occurring molecule. Optionally, these amino tennini can be
acetylated or
otherwise capped by conjugation to any other acid-containing, bioconipatible
molecule, to
include without restriction organic acids, bioinactive biologics, and
bioactive agents as
described herein. In one embodiment, the entire therapeutic polymer
composition is
biodegradable.
[0028] More particularly, the invention therapeutic polymer composition
comprises a
biodegradable, biocompatible polymer with a residue of at least one
therapeutic diol or di-
acid incorporated into the backbone of the polymer. In one enlbodiment, the
invention
therapeutic polyiner composition comprises at least one PEA having a chemical
formula
described by structural formula (I),
1O 0 H O O H
C-RI-C-N-6-6-O-R4-0-6-C-N
H R3 R3 H
n
Formula (I)
wherein n ranges from about 5 to about 150; Rl is independently selected from
residues of
3,3'-(alkanedioyldioxy)dicinnamic acid or 4,4'-(alkanedioyldioxy)dicinnamic
acid or of
a,co-bis(4-carboxyphenoxy)-(Cl-C$) alkane, (C2 - C20) alkylene, (C2-CZO)
alkenylene or
residues of saturated or unsaturated therapeutic di-acids; the R3s in
individual n units are
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independently selected from the group consisting of hydrogen, (C1-C6) alkyl,
(C2-C6)
alkenyl, (C2-C6) alkynyl, (C6-Clo) aryl (C1-C6) allcyl, and -(CH2)2S(CH3); and
R4 is
independently selected from the group consisting of (CZ-C20) alkylene, (C2-
C20)
alleenylene, (Ca-C8) alkyloxy (Ca-C20) alkylene, bicyclic-fragments of 1,4:3,6-
dianhydrohexitols of structural formula (II), saturated or unsaturated
therapeutic diol
residues, and combinations thereof;
\
CH O
H2C'
3~ ~ H2
O CH
\
Formula (II),
except that at least one of Rl and R4 is a therapeutic amount of the residue
of a therapeutic
di-acid or diol, respectively;
[0029] or at least one PEA polyiner having a chemical formula described by
structural
formula (III):
O 0 H O O H 0 0 H
u 111 u 4 n i u ~ u
C-R CN-C-C-O-R -O-C-C-N C-R -C-N-C-(CH2)4 N
H R3 R3 H m H C-O-RZ H p
O n
Formula (III)
wherein n ranges from about 5 to about 150, m ranges about 0.1 to 0.9; p
ranges from
about 0.9 to 0.1; wherein Rl is independently selected from residues of a,w-
bis(4-
carboxyphenoxy)-(C1-Cg) alkane, 3,3'-(alkanedioyldioxy)dicinnamic acid or 4,4'-
(alkanedioyldioxy)dicinnamic acid, (C2 - CA alkylene, (C2-C20) alkenylene or a
saturated
or unsaturated residues of therapeutic di-acids; each R2 is independently
hydrogen, (C1-
C12) alkyl or (C6-Clo) aryl or a protecting group; the R3s in individual m
monomers are
independently selected from the group consisting of hydrogen, (C1-C6) alkyl,
(C2-C6)
alkenyl, (C2-C6) alkynyl, (C6-Clo) aryl (C1-C6) alkyl, and -(CH2)2S(CH3); and
R4 is
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independently selected from the group consisting of (C2-C20) allcylene, (C2-
C20)
alkenylene, (C2-C8) alkyloxy (C2-C20) alkylene, bicyclic-fragments of 1,4:3,6-
dianliydrohexitols of structural formula (II), residues of saturated or
unsaturated
therapeutic diols and combinations thereof; except that at least one of R' and
R4 in at least
one of the m units is the residue of a therapeutic di-acid or diol,
respectively;
[0030] or at least one PEUR having a chemical formula described by general
structural
formula (IV),
O 0 H O O H
C-O-R6-0-C-N-6-6-0-R4-0-C-C-N
H R3 R3 H
n
Formula (IV)
wherein n ranges from about 5 to about 150; wherein R3s in independently
selected from
the group consisting of hydrogen, (C1-C6) alkyl, (C2-C6) alkenyl, (C2-C6)
alkynyl, (C6-Clo)
aryl(C1-C6) alkyl, and -(CH2)2S(CH3) and; R4 is selected from the group
consisting of (C2-
C20) alkylene, (C2-CZO) alkenylene or alkyloxy, and bicyclic-fragments of
1,4:3,6-
dianhydrohexitols of structural formula (II), residues of saturated or
unsaturated
therapeutic diols and combinations thereof; and R6 is independently selected
from (C2-C2o)
alkylene, (Ca-CZO) alkenylene or alkyloxy, bicyclic-fragments of 1,4:3,6-
dianhydrohexitols
of general formula (II), residues of saturated or unsaturated therapeutic
diols, and
combinations thereof, except that the R4 and/or R6 within at least one of the
n units is the
residue of the therapeutic diol;
[0031] or at least one PEUR polymer having a chemical structure described by
general
structural formula (V)
11O 0 H O O H 0 0 H
C-0-R6-0-C-N-6-C-O-R4-0-C-6-N nlC-O-R6-O-C-N-C-(CH2)4-N
H R. R. H H C_O-R2 H
P
O n
Formula (V)
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wherein n ranges from about 5 to about 150, m ranges about 0.1 to about 0.9: p
ranges
from about 0.9 to about 0.1; Ra is independently selected from hydrogen, (C6-
C10) aryl
(Ci-C6) alkyl, or a protecting group; the R3s in an individual m units are
independently
selected from the group consisting of hydrogen, (Cl-C6) alkyl, (C2-C6)
allcenyl, (C2-C6)
alkynyl, (C6-C10) aryl(C1-C6) alkyl, and -(CH2)2S (CH3); R4 is selected from
the group
consisting of (C2-C20) alkylene, (C2-C20) alkenylene or allcyloxy, and
bicyclic-fragments of
1,4:3,6-dianhydrohexitols of structural formula (II); and R6 is independently
selected from
(C2-C20) allcylene, (C2-C20) allcenylene or allcyloxy, bicyclic-fraginents of
1,4:3,6-
dianhydrohexitols of general fonnula (II), a residue of a saturated or
unsaturated
therapeutic diol, and combinations thereof, except that the R6 within at least
one of the m
units is the residue of a therapeutic diol;
[0032] or at least one PEU polymer having a chemical formula described by
general
structural formula (VI):
1O H O O H
C-N-C-C-O-R4-O-C-C-N
H R3 R3 H n
Formula (VI),
wherein n is about 10 to about 150; each R3s within an individual n unit are
independently
selected from hydrogen, (C1-C6) alkyl, (C2-C6) alkenyl, (C2-C6) alkynyl, (C6 -
C10) aryl
(C1-C6)alkyl, and -(CH2)2S(CH3); R4 is independently selected from (C2-C20)
alkylene,
(C2-Cao) alkenylene, (C2-C8) alkyloxy (C2-C20) alkylene, a residue of a
saturated or
unsaturated therapeutic diol; or a bicyclic-fragment of a 1,4:3,6-
dianhydrohexitol of
structural formula (II), and combinations thereof, except that the R4 within
at least one of
the n units is the residue of a therapeutic diol;
[0033] or at least one PEU having a chemical formula described by structural
formula
(VII)
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14
O H O O H 0 H
C-N-C-C-O-R4-O-C-C-N C-N-C-(CH2)4-N
H R3 R3H m H C-O-R2 H p
O
Formula (VII),
wherein m is about 0.1 to about 1.0; p is about 0.9 to about 0.1; n is about
10 to about 150;
each W is independently hydrogen, (C1-C12) alkyl or (C6-CIO) aryl, or a
protecting group;
the R3s within an individual m unit are independently selected from hydrogen,
(C1-CG)
alkyl, (C2-C6) alkenyl, (C2-C6) alkynyl, (C6 -Clo) aryl (C1-C6)alkyl, and -
(CH2)2S(CH3);
each R4 is independently selected from (C2-C20) allcylene, (C2-C20)
alkenylene, (C2-C8)
alkyloxy (C2-CZO) alkylene, a residue of a saturated or unsaturated
therapeutic diol; or a
bicyclic-fragment of a 1,4:3,6-dianhydrohexitol of structural formula (II),
and
combinations thereof, except that the R4 in at least one of the m monomers is
the residue
of a therapeutic diol.
[00341 The bicyclic-fragments of such dianhydrohexitols can be derived from
sugar
alcohols, such as D-glucitol, D-mannitol and L-iditol. Dianhydrosorbitol is
the presently
preferred bicyclic fragment of a 1,4:3,6-dianhydrohexitol for use in the PEA,
PEUR and
PEU polymers used in fabrication of the invention therapeutic polymer
compositions.
[0035] The protecting group can be t-butyl or any other protecting group known
in the
art.
[0036] In one embodiment, the residue of the therapeutic diol or di-acid
incorporated
into the polymer backbone of the invention therapeutic polymer composition of
any one of
Formulas (I) and (III)-(VII) is a therapeutic amount of the therapeutic diol
or di-acid so
that, upon administration, the composition biodegrades to release a
therapeutic amount of
the tlierapeutic diol or di-acid to the subject.
[0037] The invention therapeutic polymer compositions in which a therapeutic
diol
and/or di-acid is used in the place of a diol and/or di-acid otherwise useful
in making PEA,
PEUR, or PEU polymers as described herein, can be fonnulated into particles to
provide a
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variety of properties. The particles can have a variety of sizes and
structures suitable to
meet differing therapeutic goals and routes of administration as described in
full in co-
pending U.S. provisional applications 60/654,715, filed February 17, 2005,
60/684,670,
filed May 25, 2005, and 60/737,401, filed November 14, 2005.
[0038] As used herein, the terms "amino acid" and "a-amino acid" mean a
chemical
compound containing an amino group, a carboxyl group and a pendent R group,
such as
the R3 groups defined herein. As used herein, the term "biological a-amino
acid" means
the amino acid(s) used in synthesis are selected from phenylalanine, leucine,
glycine,
alanine, valine, isoleucine, methionine, or a mixture thereof.
[0039] As used herein, a "therapeutic diol" or "therapeutic di-acid" means,
respectively, any diol or di-acid molecule, whether synthetically produced, or
naturally
occurring (e.g., endogenously) that affects a biological process in a
mammalian individual,
such as a human, in a therapeutic or palliative manner when administered to
the mammal.
[0040] As used herein, the term "residue of a therapeutic diol" means a
portion of a
therapeutic diol, as described herein, which portion excludes the two hydroxyl
groups of
the diol. As used herein, the term "residue of a therapeutic di-acid" means a
portion of a
therapeutic di-acid, as described herein, which portion excludes the two
carboxyl groups
of the di-acid. The corresponding therapeutic diol or di-acid containing the
"residue"
thereof is used in synthesis of the polymer compositions. The residue of the
therapeutic
di-acid or diol is reconstituted in vivo (or under similar conditions of pH,
aqueous media,
and the like) to the corresponding di-acid or diol upon release from the
backbone of the
polymer by biodegradation in a controlled manner that depends upon the
properties of the
PEA, PEUR or PEU polymer selected to fabricate the composition, which
properties are as
known in the art and as described herein.
[0041] As used herein the term "bioactive agent" means an active agent that
affects a
biological process in a mammalian individual, such as a human, in a
therapeutic or
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16
palliative manner when administered to the mammal and that is not incorporated
into the
polymer backbone. Bioactive agents may include, without limitation, small
molecule
drugs, peptides, proteins, DNA, eDNA, RNA, sugars, lipids and whole cells. One
or more
such bioactive agents may be dispersed in the invention therapeutic polymer
compositions.
[0042] As used herein, the term "dispersed" as used to refer to bioactive
agents means
that the bioactive agent is loaded into, mixed, dissolved, homogenized, and/or
covalently
bound ("dispersed") in a polymer, for example, attached to a functional group
in the
therapeutic polymer of the composition or to the surface of a polymer
particle, but not
incorporated into the backbone of a PEA, PEUR, or PEU polymer. To distinguish
dispersed therapeutic diols and di-acids from those that are incorporated into
the polymer
backbone, (as a residue thereof), such dispersed diols and di-acids are
referred to herein as
"bioactive agent(s)" and may be linked to the polymer, contained within
polymer
conjugates or otherwise dispersed in the invention therapeutic polymer
composition the
saine as other bioactive agents disclosed herein.
[0043] The term, "biodegradable" as used herein to describe the invention
therapeutic
polyiner compositions means the polymer used therein is capable of being
broken down
into innocuous products in the normal functioning of the body. This is
particularly true
when the amino acids used in fabrication of the therapeutic polymer
compositions are
biological L-a-amino acids. The polymers in the invention therapeutic polymer
compositions include hydrolyzable ester and enzymatically cleavable ainide
linkages that
provide biodegradability, and are typically chain terminated, predominantly
with amino
groups. Optionally, the amino termini of the polymers can be acetylated or
otherwise
capped by conjugation to any other acid-containing, biocompatible molecule, to
include
without restriction organic acids, bioinactive biologics, and bioactive agents
as described
herein. In one embodiment, the entire polymer composition, and any particles
made
thereof, is substantially biodegradable.
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17
[0044] In one alternative, at least one of the a-amino acids used in
fabrication of the
invention polymers is a biological a-amino acid. For example, when the R3s are
CH2Ph,
the biological a-amino acid used in synthesis is L-phenylalanine. In
alternatives wherein
the R3s are CHa-CH(CH3)2, the polymer contains the biological a-amino acid, L-
leucine.
By varying the R3s within monomers as described herein, other biological a-
amino acids
can also be used, e.g., glycine (when the R3s are H), alanine (when the R3s
are CH3),
valine (when the R3s are CH(CH3)2), isoleucine (when the R3s are CH(CH3)-CH2-
CH3),
phenylalanine (when the R3s are CH2-C6H5), or methionine (when the R3s are -
(CH2)2S-
CH3), and combinations thereof. In yet another alternative embodiment, all of
the various
a-amino acids contained in the polymers used in making the invention
therapeutic
polyiner compositions are biological a-amino acids, as described herein.
[0045] In yet a further embodiment wherein the polymer is a PEA, PEUR or PEU
of
any one of formulas (I) and (III)-(VII), at least one of the R3s further can
be -(CH2)3-,
wllich cyclizes to form the chemical structure described by structural formula
(XIII):
H O
N-C-C-O-
H2C,CCH2
H2
Formula (XIII)
When the R3s are -(CH2)3-, an a-imino acid analogous to pyrrolidine-2-
carboxylic acid
(proline) is used.
[0046] In certain embodiments, the polymer in the invention therapeutic
polymer
composition plays an active role in the treatment processes at the site of
local
administration, e.g., by injection, by holding the polymer in an agglomeration
or polymer
depot at the site of injection for a period of time sufficient to allow the
subject's
endogenous processes to slowly release particles or polymer molecules from the
agglomeration. Meanwhile, the subject's endogenous processes biodegrade the
polymer
backbone so as to release the incorporated therapeutic diol and/or di-acid
therapeutic
agents, as well as any bioactive agents dispersed in the polymer. The fragile
therapeutic
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18
diols and di-acids and optional bioactive agents are protected by the more
slowly
biodegrading polymer to increase half-life and persistence of the therapeutic
diol or di-
acid and bioactive agent(s) at the site of local administration.
[0047] In addition, the polymers disclosed herein (e.g., those having
structural formulas
(I) and (III)-(VII), upon enzymatic degradation, provide essential amino acids
and other
breakdown products that can be metabolized using patliways similar to those
used in
metabolizing fatty acids and sugars. Uptake of the invention therapeutic
polymer
compositions is safe: studies have shown that the subject can metabolize/clear
the polymer
degradation products. These polymers and the invention therapeutic polymer
compositions are, therefore, substantially non-inflammatory to the subject
both at the site
of injection and systemically, apart from any trauma caused by injection
itself.
[00481 The PEA, PEUR and PEU polymer molecules may also have the bioactive
agent attached thereto, optionally via a linker or incorporated into a
crosslinker between
molecules. For example, in one embodiment, the polymer is contained in a
polymer-
bioactive agent conjugate having structural formula VIII:
O O H H O O H H O O H 11 CRl-C-N-C-CO-R4-0-C-C-N C-R~-C-N-CH-(CH2)q-NH
R3 R3 4c
C=0 p J n
R5
R7
Formula (VIII)
wherein n, M. p, Rl, R3, and R4 are as above, R5 is selected from the group
consisting of -
O-,
-S-, and -NR8-, wherein R8 is H or (Ci-C8)alkyl; and R7 is the bioactive
agent.
[0049] In yet another embodiment, two molecules of the polymer of structural
formula
(IX) can be crosslinked to provide an -R5-R7-RS- conjugate. In another
embodiment, as
shown in structural formula (IX) below, the bioactive agent is covalently
linked to two
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19
parts of a single polymer molecule of structural fonnula (III) through the -RS-
R7-RS-
conjugate and R5 is independently selected from the group consisting of -0-, -
S-, and -
NR8-, wherein R8 is H or (C1-C8) allcyl; and R7 is the bioactive agent.
R3 R3
C-Rl-C-N-H-C-O-R4-O-C-CIH-N C-RI-C-N-C-(CH2)4-NH
0 0 0 o )Ir, o o~
Rs R7 Rs
R3 Ra
I , H
--(CH2)4-C-NH-C-R'-C NH-C-C-O-R4-O-C-CH-NC-R'=C
H ii n H is 11 11 u
0 O p O 0 O O m n
Formula (IX)
[0050] Alternatively still, as shown in structural formula (X) below, a
linker, -X-Y-,
can be inserted between RS and bioactive agent R7, in the molecule of
structural formula
(III), wherein X is selected from the group consisting of (C1-C18) alkylene,
substituted
alkylene, (C3-C8) cycloalkylene, substituted cycloalkylene, 5-6 membered
heterocyclic
system containing 1-3 heteroatoms selected from the group 0, N, and S,
substituted
heterocyclic, (C2-C18) alkenyl, substituted alkenyl, alkynyl, substituted
alkynyl, C6 and C1
aryl, substituted aryl, heteroaryl, substituted heteroaryl, alkylaryl,
substituted alkylaryl,
arylalkynyl, substituted arylalkynyl, arylalkenyl, substituted arylalkenyl,
arylalkynyl,
substituted arylalkynyl and wherein the substituents are selected from the
group H, F, Cl,
Br, I, (C1-C6) alkyl, -CN, -NO2, -OH, -O(C1-C4) alkyl, -S(C1-C6) alkyl, -
S[(=O)(C1-C6)
alkyl], -S[(02)(Ct-C6) alkyl], -C[(=O)(C1-C6) alkyl], CF3,-O[(CO)-( C1-C6)
alkyl], -
S(02)[N(R9R10)], -NH[(C=O)(Cl-CG) alkyl], -NH(C=O)N(WR10), -NWRlO); where R9
and R10 are independently H or (Cl-C6) alkyl; and Y is selected from the group
consisting
of -0-, -S-, -S-S-, -S(O)-,-S(02)-, -NR$-, -C(=O)-, -OC(=O)-, -C(=O)O-, -
OC(=O)NH-, -
NR$C(=O)-, -C(=O)NR$-, -NR8C(=O)NR8-, and -NR$C(=S)N R8-.
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O
H
11 O H H 0 0 11 H H O 0
C-R'-C-N-C-C-O-R4-O-C-C-N C-R'-C-N-CH-(CH2)4-NH
R3 R3 m C=0 )Pj n
R5
X
Y
I
R7
Formula (X)
[0051] In another embodiment, two parts of a single macromolecule are
covalently
linked to the bioactive agent through an -R5-R7-Y-X- RS- bridge (Formula XI):
R3 R3
H i I H H
C-RI-C-N-C-C-O-R4-O-C-CH-N C-R'-C-N-C-(CH2)4-NH
(\O O H O ~ m O 11 0 p n
R5 R7 Y X R510
O~/
'I( R3 R3
I H
--(CH2)4-C-NH-C-R'-C NH-C-C-O-R4-O-C-CH-N-C-R'-C
H H
0 op 0 0 0 omn
Formula (XI)
wherein, X is selected from the group consisting of (C1-Ci$) alkylene,
substituted alkylene,
(C3-C8) cycloalkylene, substituted cycloalkylene, 5-6 membered heterocyclic
system
containing 1-3 heteroatoms selected from the group 0, N, and S, substituted
heterocyclic,
(C2-C18) alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, (C6-C10)
aryl,
substituted aryl, heteroaryl, substituted heteroaryl, alkylaryl, substituted
alkylaryl,
arylalkynyl, substituted arylalkynyl, arylalkenyl, substituted arylalkenyl,
arylallcynyl,
substituted arylalkynyl, wherein the substituents are selected from the group
consisting of
H, F, Cl, Br, I, (C1-C6) alkyl, -CN,
-NO2,, -OH, -O(C1-C6) alkyl, -S(Cl-C6) alkyl, -S[(=O)(C1-C6) alkyl], -
S[(02)(C1-C6)
alkyl],
-C[(=O)(Cl-C6) alkyl], CF3,-O[(CO)-(Cl-C6) alkYl], -S(02)[N(RgR10)], -
NH[(C=O)(C1-C6)
alkyl], -NH(C=0)N(R9R10), wherein R9 and RI0 are independently H or (C1-C6)
alkyl, and
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21
-N(R11R1a), wherein Rll and R12 are independently selected from (Ca-C20)
allcylene and
(C2-C2o) alkenylene.
[0052] In yet another embodiment, the polymer particle delivery composition
contains
four molecules of the polymer, except that only two of the four molecules omit
R7 and are
crosslinked to provide a single RS-X-R5- conjugate.
[0053] The term "aryl" is used with reference to structural formulae herein to
denote a
phenyl radical or an ortho-fused bicyclic carbocyclic radical having about
nine to ten ring
atoms in which at least one ring is aromatic. In certain einbodiments, one or
more of the
ring atoms can be substituted with one or more of nitro, cyano, halo,
trifluoromethyl, or
trifluoromethoxy. Examples of aryl include, but are not limited to, phenyl,
naphthyl, and
nitrophenyl.
[0054] The term "alkenylene" is used with reference to structural formulae
herein to
mean a divalent branched or unbranched hydrocarbon chain containing at least
one
unsaturated bond in the main chain or in a side chain.
[0055] The molecular weights and polydispersities herein are determined by gel
permeation chromatography (GPC) using polystyrene standards. More
particularly,
number and weight average molecular weights (Mõ and M,) are determined, for
example,
using a Mode1510 gel permeation chromatography (Water Associates, Inc.,
Milford, MA)
equipped with a high-pressure liquid chromatographic pump, a Waters 486 UV
detector
and a Waters 2410 differential refractive index detector. Tetrahydrofuran
(THF) or N,N-
dimethylacetamide (DMAc) is used as the eluent (1.0 mL/min). Polystyrene or
poly(methyl methacrylate) standards having narrow molecular weight
distribution were
used for calibration.
[0056] Methods for making polymers containing a-amino acids in the backbone
are
well known in the art. For example, for the embodiment of the polymer of
structural
formula (I) wherein R4 is incorporated into an a-amino acid, for polymer
synthesis the a-
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22
amino acid with pendant R3 can be converted through esterification into a bis-
a,co-diamine,
for example, by condensing the a-amino acid containing pendant R3 with a diol
HO R4-
OH. As a result, di-ester monomers witli reactive a,co-ainino groups are
formed. Then,
the bis-a,co-diamine is entered into a polycondensation reaction with a di-
acid such as
sebacic acid, or bis-activated esters, or bis-acyl chlorides, to obtain the
final polymer
having both ester and amide bonds (PEA). Alternatively, for example, for
polymers of
structure (I), instead of the di-acid, an activated di-acid derivative, e.g.,
bis-4-nitrophenyl
diester, can be used as an activated di-acid. Additionally, a bis-di-
carbonate, such as
bis(4-nitrophenyl) dicarbonate, can be used as the activated species to obtain
polymers
containing a residue of a di-acid. In the case of PEUR polymers, a final
polymer is
obtained having both ester and urethane bonds.
[0057] More particularly, synthesis of the unsaturated poly(ester-amide)s
(UPEAs)
useful as biodegradable polymers of the structural formula (I) as disclosed
above will be
described,
O
0 0 ~J H
(a) 11 Rl is / \e C~/
H lol
Wherein and/or (b) R4 is -CHZ-CH=CH-CH2- . In cases where (a) is present and
(b) is not
present, R4 in (I) is -C4H8- or -C6H12-. In cases where (a) is not present and
(b) is present,
Rl in (I) is -C4H8- or -C8H16-.
[0058] The UPEAs can be prepared by solution polycondensation of either (1) di-
4-
toluene sulfonic acid salt of bis(a-amino acid) di-ester of unsaturated diol
and di-4-
nitrophenyl ester of saturated dicarboxylic acid or (2) di-4-toluene sulfonic
acid salt of bis
(a-amino acid) diester of saturated diol and di-4-nitrophenyl ester of
unsaturated
dicarboxylic acid or (3) di-4-toluene sulfonic acid salt of bis(a-anino acid)
diester of
unsaturated diol and di-4-nitrophenyl ester of unsaturated dicarboxylic acid.
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23
[0059] Salts of 4-toluene sulfonic acid are known for use in synthesizing
polymers
containing amino acid residues. The aryl sulfonic acid salts are used instead
of the free
base because the aryl sulfonic salts of bis (a-amino acid) diesters are easily
purified
through recrystallization and render the amino groups as unreactive animonium
tosylates
throughout workup. In the polycondensation reaction, the nucleophilic ainino
group is
readily revealed through the addition of an organic base, such as
triethylamine, so the
polymer product is obtained in high yield.
[0060] For polymers of structural formula (I), for example, the di-4-
nitrophenyl esters
of unsaturated dicarboxylic acid can be synthesized from 4-nitrophenyl and
unsaturated
dicarboxylic acid chloride, e.g., by dissolving triethylamine and 4-
nitrophenol in acetone
and adding unsaturated dicarboxylic acid chloride dropwise with stirring at -
78 C and
pouring into water to precipitate product. Suitable acid chlorides included f-
umaric,
maleic, mesaconic, citraconic, glutaconic, itaconic, ethenyl-butane dioic and
2-propenyl-
butanedioic acid chlorides. For polymers of structure (IV) and (V), bis-p-
nitrophenyl
dicarbonates of saturated or unsaturated diols are used as the activated
monomer.
Dicarbonate monomers of general structure (XII) are employed for polymers of
structural
formula (IV) and (V),
0
0
RS-O-C-O-R6-O-C-O-RS
Formula (XII)
wherein each R5 is independently (C6 -C1o) aryl optionally substituted with
one or more
nitro, cyano, halo, trifluoromethyl, or trifluoromethoxy; and R6 is
independently (C2 -C20)
alkylene, (C2 -C20) alkenylene or (C2 -C20) alkyloxy (C2 -C20) alkenylene,
fragnlents of
1,4:3,6-dianhydrohexitols of general formula (II), or a residue of a saturated
or unsaturated
therapeutic diol .
[0061] Suitable therapeutic diol compounds that can be used to prepare bis(oa-
amino
acid) diesters of therapeutic diol monomers, or bis(carbonate) of therapeutic
di-acid
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24
monomers, for introduction into the invention therapeutic polymer compositions
include
naturally occurring therapeutic diols, such as 17-(3-estradiol, a natural and
endogenous
hormone, useful in preventing restenosis and tumor growth (Yang, N.N., et al.
Identification of an estrogen response element activated by metabolites of 17-
(3-estradiol
and raloxifene. Science (1996) 273, 1222-1225; Parangi, S., et al.,
Inliibition of
angiogenesis and breast cancer in mice by the microtubule inhibitors 2-
methoxyestradiol
and taxol, Cancer Res. (1997) 57, 81-86; and Fotsis, T., et al., The
endogenous oestrogen
inetabolite 2-methoxyoestradiol inhibits angiogenesis and suppresses tumor
growth.
Nature (1994) 368, 237-239). The safety profiles of such endogenously
occurring
therapeutic diol molecules are believed to be superior to those of synthetic
and/or non-
endogenous molecules having a similar utility, such as sirolimus.
[0062] Incorporation of a therapeutic diol into the backbone of a PEA, PEUR or
PEU
polymer is illustrated in this application by Example 8, in which active
steroid hormone
17-(3-estradiol containing mixed hydroxyls - secondary and phenolic - is
introduced into
the backbone of a PEA polymer. When the PEA polymer is used to fabricate
particles and
the particles are implanted into a patient, for example, following
percutaneous
transluminal coronary angioplasty (PTCA), 17-0-estradiol released from the
particles in
vivo can help to prevent post-implant restenosis in the patient. 17-(3-
estradiol, however, is
only one example of a diol with therapeutic properties that can be
incorporated in the
backbone of a PEA, PEUR or PEU polymer in accordance with the invention. In
one
aspect, any bioactive steroid-diol containing primary, secondary or phenolic
hydroxyls can
be used for this purpose. Many steroid esters that can be made from bioactive
steroid diols
for use in the invention are disclosed in European application EP 0127 829 A2.
[0063] Due to the versatility of the PEA, PEUR and PEU polymers used in the
invention compositions, the amount of the therapeutic diol or di-acid
incorporated in the
polymer backbone can be controlled by varying the proportions of the building
blocks of
the polymer. For example, depending on the composition of the PEA, loading of
up to
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40% w/w of 17-(3-estradiol can be achieved. Two different regular, linear PEAs
with
various loading ratios of 17-(3-estradiol are illustrated in Scheme 1 below:
"homopoly"-bis-Leu-Estradiol-Adipate (40% w/w -estradiol on polymer)
CH(CH3)2 CH(CH3)2
H2C 0
CH~
- - O-C-C-NH-C-(CH2)4-C
N-C-CO ~ ~
H H 0 O H 0
n
Copolymer: Leu(ED)3Lys(OEt)Adip4 with 38% w/w estradiol loading
CH(CH3)2 CH(CH3)2
' H2C O
CH2 -C-C-NH-C-(CH2)4-C
N-C-C-O
LLHo o H 0
3n
O H H
C-(CH2)4-C-N-(CH2)q,-C-NH
0 COOEt
ln
Scheme 1
[0064] Siniilarly, the loading of the therapeutic diol into PEUR and PEU
polymer can
be varied by varying the amount of two or more building blocks of the polymer.
Synthesis
of a PEUR containing 17-beta-estradiol is illustrated in Example 9 below.
[0065] In addition, synthetic steroid based diols based on testosterone or
cholesterol,
such as 4-androstene-3, 17 diol (4-Androstenediol), 5-androstene-3, 17 diol (5-
Androstenediol), 19-nor5-androstene-3, 17 diol (19-Norandrostenediol) are
suitable for
incorporation into the backbone of PEA and PEUR polymers according to this
invention.
Moreover, therapeutic diol compounds suitable for use in preparation of the
invention
therapeutic polymer compositions include, for example, amikacin; amphotericin
B;
apicycline; apramycin; arbekacin; azidamfenicol; bambermycin(s); butirosin;
carbomycin;
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26
cefpiramide; chloramphenicol; chlortetracycline; clindamycin; clomocycline;
demeclocycline; diathymosulfone; dibekacin, dihydrostreptomycin;
dirithromycin;
doxycycline; erythromycin; fortimicin(s); gentamycin(s); glucosulfone
solasulfone;
guamecycline; isepamicin; josamycin; kanamycin(s); leucomycin(s); lincomycin;
lucensomycin; lymecycline; meclocycline; methacycline; micronomycin;
midecamycin(s);
minocycline; mupirocin; natamycin; neomycin; netilmicin; oleandomycin;
oxytetracycline; paromycin; pipacycline; podophyllinic acid 2-ethylhydrazine;
primycin;
ribostamycin; rifamide; rifampin; rafamycin SV; rifapentine; rifaximin;
ristocetin;
rokitamycin; rolitetracycline; rasaramycin; roxithromycin; sancycline;
sisomicin;
spectinomycin; spiramycin; streptomycin; teicoplanin; tetracycline;
thiamphenicol;
theiostrepton; tobramycin; trospectomycin; tuberactinomycin; vancomycin;
candicidin(s);
chlorphenesin; dermostatin(s); filipin; fungichromin; kanamycin(s);
leucomycins(s);
lincomycin; lvcensomycin; lymecycline; meclocycline; methacycline;
micronomycin;
midecamycin(s); minocycline; mupirocin; natamycin; neomycin; netilmicin;
oleandomycin; oxytetracycline; paramomycin; pipacycline; podopliyllinic acid 2-
ethylhydrazine; priycin; ribostamydin; rifamide; rifampin; rifamycin SV;
rifapentine;
rifaximin; ristocetin; rokitamycin; rolitetracycline; rosaramycin;
roxithroinycin;
sancycline; sisomicin; spectinomycin; spiramycin; strepton; otbrainycin;
trospectomycin;
tuberactinomycin; vancomycin; candicidin(s); chlorphenesin; dermostatin(s);
filipin;
fungichromin; meparticin; mystatin; oligomycin(s); erimycinA; tubercidin; 6-
azauridine;
aclacinomycin(s); ancitabine; anthramycin; azacitadine; bleomycin(s)
carubicin;
carzinophillin A; chlorozotocin; chromomcin(s); doxifluridine; enocitabine;
epirubicin;
gemcitabine; mannomustine; menogaril; atorvasi pravastatin; clarithromycin;
leuproline;
paclitaxel; mitobronitol; mitolactol; mopidamol; nogalamycin; olivomycin(s);
peplomycin; pirarubicin; prednimustine; puromycin; ranimustine; tubercidin;
vinesine;
zorubicin; coumetarol; dicoumarol; ethyl biscoumacetate; ethylidine
dicoumarol; iloprost;
taprostene; tioclomarol; amiprilose; romurtide; sirolimus (rapamycin);
tacrolimus; salicyl
alcohol; bromosaligenin; ditazol; fepradinol; gentisic acid; glucamethacin;
olsalazine; S-
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adenosylmethionine; azithromycin; salmeterol; budesonide; albuteal; indinavir;
fluvastatin; streptozocin; doxorubicin; daunorubicin; plicamycin; idarubicin;
pentostatin;
metoxantrone; cytarabine; fludarabine phosphate; floxuridine; cladriine;
capecitabien;
docetaxel; etoposide; topotecan; vinblastine; teniposide, and the like. The
therapeutic diol
can be selected to be either a saturated or an unsaturated diol.
[0066] Suitable naturally occurring and synthetic therapeutic di-acids that
can be used
to prepare an amide linkage in the PEA polymer compositions of the invention
include, for
example, bambermycin(s); benazepril; carbenicillin; carzinophillin A;
cefixime; cefininox
cefpimizole; cefodizime; cefonicid; ceforanide; cefotetan; ceftazidiine;
ceftibuten;
cephalosporin C; cilastatin; denopterin; edatrexate; enalapril; lisinopril;
methotrexate;
moxalactam; nifedipine; olsalazine; penicillin N; ramipril; quinacillin;
quinapril;
temocillin; ticarcillin; Tomudex (N-[[5-[[(1,4-Dihydro-2-methyl-4-oxo-6-
quinazolinyl)methyl] inethylamino]-2-thienyl]carbonyl]-L-glutamic acid), and
the like.
The safety profile of naturally occurring therapeutic di-acids is believed to
surpass that of
synthetic therapeutic di-acids. The therapeutic di-acid can be either a
saturated or an
unsaturated di-acid.
[0067] The chemical and therapeutic properties of the above described
therapeutic diols
and di-acids as tumor inhibitors, cytotoxic antimetabolites, antibiotics, and
the like, are
well known in the art and detailed descriptions thereof can be found, for
example, in
thel3th Edition of The Merck Index (Whitehouse Station, N.J., USA).
[0068] The di-aryl sulfonic acid salts of diesters of a-amino acid and
unsaturated diol
can be prepared by admixing a-amino acid, e.g., 4-aryl sulfonic acid
monohydrate and
saturated or unsaturated diol in toluene, heating to reflux temperature, until
water
evolution is minimal, then cooling. The unsaturated diols include, for
example, 2-butene-
1,3-diol and 1,18-octadec-9-en-diol.
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[0069] Saturated di-4-nitrophenyl esters of dicarboxylic acid and saturated di-
4-toluene
sulfonic acid salts of bis-a -amino acid esters can be prepared as described
in U.S. Patent
No. 6,503,538 Bl.
[0070] Synthesis of the unsaturated poly(ester-amide)s (UPEAs) useful as
biodegradable polymers of the structural formula (I) as disclosed above will
now be
described. UPEAs having the structural formula (I) can be made in similar
fashion to the
compound (VII) of U. S. Patent No. 6,503,538 Bl, except that R4 of (III) of
6,503,538
and/or Rl of (V) of 6,503,538 is (C2-C20) alkenylene as described above. The
reaction is
carried out, for example, by adding dry triethylamine to a mixture of said
(III) and (IV) of
6,503,538 and said (V) of 6,503,538 in dry N,N-dimethylacetamide, at room
temperature,
then increasing the temperature to 60 C and stirring for 16 hours, then
cooling the reaction
solution to room temperature, diluting with ethanol, pouring into water,
separating
polymer, washing separated polymer with water, drying to about 30 C under
reduced
pressure and then purifying up to negative test on p-nitrophenol and p-toluene
sulfonate.
A preferred reactant (IV) of 6,503,538 is p-toluene sulfonic acid salt of
Lysine benzyl
ester, the benzyl ester protecting group is preferably removed from (II) to
confer
biodegradability, but it should not be removed by hydrogenolysis as in Example
22 of U.S.
Patent No. 6,503,538 because hydrogenolysis would saturate the desired double
bonds;
rather the benzyl ester group should be converted to an acid group by a method
that would
preserve unsaturation. Alternatively, the lysine reactant (IV) of 6,503,538
can be
protected by a protecting group different from benzyl that can be readily
removed in the
finished product while preserving unsaturation, e.g., the lysine reactant can
be protected
with t-butyl (i.e., the reactant can be t-butyl ester of lysine) and the t-
butyl can be
converted to H while preserving unsaturation by treatment of the product (II)
with acid.
[0071] A working example of the compound having structural formula (I) is
provided
by substituting p-toluene sulfonic acid salt of bis(L-phenylalanine) 2-butene-
1,4-diester
for (III) in Example 1 of 6,503,538 or by substituting di-p-nitrophenyl
fumarate for (V) in
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Exainple 1 of 6,503,538 or by substituting the p-toluene sulfonic acid salt of
bis(L-
phenylalanine) 2-butene-1,4-diester for III in Example 1 of 6,503,538 and also
substituting
bis-p-nitrophenyl fumarate for (V) in Example 1 of 6,503,538.
[0072] In unsaturated compounds having either structural formula (I) or (IV),
the
following hold. An amino substituted aminoxyl (N-oxide) radical bearing group,
e.g., 4-
amino TEMPO, can be attached using carbonyldiimidazol, or suitable
carbodiiinide, as a
condensing agent. Bioactive agents, as described herein, can be attached via
the double
bond functionality. Hydrophilicity can be imparted by bonding to poly(ethylene
glycol)
diacrylate.
[0073] In yet another aspect, the PEA and PEUR polymers contemplated for use
in
forming the invention therapeutic polymer compositions include those set forth
in U.S.
Patent Nos. 5,516, 881; 6,476,204; 6,503,538; and in U.S. Application Nos.
10/096,435;
10/101,408; 10/143,572; and 10/194,965; the entire contents of each of which
is
incorporated herein by reference.
[0074] The biodegradable PEA, PEUR and PEU polymers can contain from one to
multiple different a-amino acids per polymer molecule and preferably have
weight
average molecular weights ranging from 10,000 to 125,000; these polymers and
copolymers typically have intrinsic viscosities at 25 C, determined by
standard
viscosimetric methods, ranging from 0.3 to 4.0, for example, ranging from 0.5
to 3.5.
[0075] PEA and PEUR polymers contemplated for use in the practice of the
invention
can be synthesized by a variety of methods well known in the art. For example,
tributyltin
(IV) catalysts are commonly used to form polyesters such as poly(s-
caprolactone),
poly(glycolide), poly(lactide), and the like. However, it is understood that a
wide variety
of catalysts can be used to form polymers suitable for use in the practice of
the invention.
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[0076] Such poly(caprolactones) conteniplated for use have an exemplary
structural
formula (XIV) as follows:
O
O-C-(CH2)s
Formula (XIV)
[0077] Poly(glycolides) contemplated for use have an exemplary structural
formula
(XV) as follows:
O H
O-C-C
H n
Formula (XV)
[0078] Poly(lactides) contemplated for use have an exemplary structural
fonnula (XVI)
as follows:
O Me
O-C-C
n
Formula (XVI)
[0079] An exemplary syntllesis of a suitable poly(lactide-co-s-caprolactone)
including
an aminoxyl moiety is set forth as follows. The first step involves the
copolymerization of
lactide and s-caprolactone in the presence of benzyl alcohol using stannous
octoate as the
catalyst to form a polymer of structural formula (XVII).
O O
Me
O-CH20HI + jt O -IA
~O + nz O >
Me
O
O H O
&CH2O C-C-O C-(CH2)5-O H
Me n In
Formula (XVII)
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[0080] The hydroxy terminated polymer chains can then be capped with maleic
anhydride to form polymer chains having structural formula (XVIII):
O H O O O
O-CH20 C-C-O C-(CH2)5-O C-C=C-C-OH
Me n m H H
Forniula (XVIII)
[0081] At this point, 4-amino-2,2,6,6-tetramethylpiperidine- 1 -oxy can be
reacted with
the carboxylic end group to covalently attach the aminoxyl moiety to the
copolymer via
the amide bond which results from the reaction between the 4-amino group and
the
carboxylic acid end group. Alternatively, the maleic acid capped copolymer can
be
grafted with polyacrylic acid to provide additional carboxylic acid moieties
for subsequent
attachment of further aminoxyl groups.
[0082] The description and methods of synthesis of PEA and PEUR polymers that
do
not have a therapeutic diol or di-acid incorporated into the backbone of the
polymer are set
forth in U.S. Patent Nos. 5,516, 881; 6,476,204; 6,503,538; and in U.S.
Application Nos.
10/096,435; 10/101,408; 10/143,572; 10/194,965; 10/362,848, 10/346,848,
10/788,747
and in provisional application 60/576,239, the entire content of each of which
is
incorporated herein by reference.
[0083] The invention bioactive PEA, PEUR and PEU polynler compositions useful
in
the invention methods biodegrade by enzymatic action at the surface.
Therefore, the
polymers, for example particles thereof, facilitate in vivo release of a
bioactive agent
incorporated into the backbone or dispersed in the polymer at a controlled
release rate,
which is specific and constant over a prolonged period. Additionally, PEA,
PEUR and
PEU polymers break down in vivo without production of adverse side products,
the
polymers in the compositions are substantially non-inflammatory.
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[0084] The biodegradable PEA, PEUR and PEU polymers can contain from one to
multiple different a-amino acids per polymer molecule and preferably have
weight
average molecular weights ranging from 10,000 to 125,000; these polymers and
copolymers typically have intrinsic viscosities at 25 C, determined by
standard
viscosimetric methods, ranging from 0.3 to 4.0, for example, ranging from 0.5
to 3.5.
[0085] The PEU polymers disclosed herein can be fabricated as higli molecular
weight
polymers useful for making the invention therapeutic polymer compositions for
delivery to
humans and other mammals of a variety of pharmaceutical and biologically
active agents.
The PEUs incorporate hydrolytically cleavable ester groups and non-toxic,
naturally
occurring monomers that contain a-amino acids in the polymer chains. The
ultimate
biodegradation products of PEUs will be amino acids, diols, and COz. In
contrast to the
PEAs and PEURs, the invention PEUs are crystalline or semi-crystalline and
possess
advantageous mechanical, chemical and biodegradation properties that allow
formulation
of completely synthetic, and hence easy to produce, crystalline and semi-
crystalline
polymer particles, for example nanoparticles. For example, the PEU polymers
used in the
invention therapeutic polymer compositions have high mechanical strength, and
surface
erosion of the PEU polymers can be catalyzed by enzymes present in
physiological
conditions, such as hydrolases.
[0086] In unsaturated compounds having structural formula (VII) for PEU, the
following hold: An amino substituted aminoxyl (N-oxide) radical bearing group
e.g., 4-
amino TEMPO, can be attached using carbonyldiimidazole, or suitable
carbodiimide, as a
condensing agent. Bioactive agents, and the like, as described herein,
optionally can be
attached via the double bond functionality provided that the therapeutic diol
residue in the
polymer composition does not contain a double or triple bond.
[0087] For example, the invention high molecular weight semi-crystalline PEUs
having
structural formula (VI) can be prepared inter-facially by using phosgene as a
bis-
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electrophilic monomer in a chloroform/water system, as shown in the reaction
scheme (2)
below:
1. Na2CO3 / H20
H 0 0 H 2. CICOCI / CHC13
HOTos.H2N-C-C-O-R4-O-C-C-NH2.TosOH ( VI )
R3 R3
Scheme (2)
[0088] Synthesis of copoly(ester ureas) (PEUs) containing L-Lysine esters and
having
structural formula (VII) can be carried out by a similar Scheme (3):
H O O H H
~2 HOTos.H2N-C-C-O-R4-O-C-C3 NH2.TosOH + p HOTos.H2N-C-(CH2)4-NH2.TosOH
R3 R C-O-R2
0
1. NaZCO3 / H20
2. CICOCI / CHCI3
(VII)
Scheme (3)
[0089] A 20% solution of phosgene (C1COCl) (highly toxic) in toluene, for
example
(commercially available (Flulca Chemie, GMBH, Buchs, Switzerland), can be
substituted
either by diphosgene (trichloromethylchloroformate) or triphosgene
(bis(trichloromethyl)carbonate). Less toxic carbonyldiimidazole can be also
used as a bis-
electrophilic monomer instead of phosgene, di-phosgene, or tri-phosgene.
General Procedure for Synthesis of PEUs
[0090] It is necessary to use cooled solutions of monomers to obtain PEUs of
high
molecular weight. For example, to a suspension of di-p-toluenesulfonic acid
salt of bis(a-
amino acid)-a,o)-alkylene diester in 150 mL of water, anhydrous sodium
carbonate is
added, stirred at room temperature for about 30 minutes and cooled to about 2 -
0 C,
forming a first solution. In parallel, a second solution of phosgene in
chloroform is cooled
to about 15 -10 C. The first solution is placed into a reactor for
interfacial
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polycondensation and the second solution is quickly added at once and stirred
briskly for
about 15 min. Then chloroform layer can be separated, dried over anhydrous
Na2SO4, and
filtered. The obtained solution can be stored for further use.
[0091] All the exemplary PEU polymers fabricated were obtained as solutions in
cliloroform and these solutions are stable during storage. However, some
polymers, for
example, 1-Phe-4, become insoluble in chloroform after separation. To overcome
this
problem, polyiners can be separated from chloroform solution by casting onto a
smooth
hydrophobic surface and allowing chloroform to evaporate to dryness. No
furtller
purification of obtained PEUs is needed. The yield and characteristics of
exemplary PEUs
obtained by this procedure are summarized in Table 2 herein.
General Procedure for Preparation of porous PEUs.
[0092] Methods for making the PEU polymers containing a-amino acids in the
general
fonnula will now be described. For example, for the embodiment of the polymer
of
formula (I) or (II), the a-amino acid can be converted into a bis(a-amino
acid)-a,co-diol-
diester monomer, for example, by condensing the a-amino acid with a diol HO-R'-
OH.
As a result, ester bonds are formed. Then, acid chloride of carbonic acid
(phosgene,
diphosgene, triphosgene) is entered into a polycondensation reaction with a di-
p-
toluenesulfonic acid salt of a bis(a-amino acid) -alkylene diester to obtain
the final
polymer having both ester and urea bonds. In the present invention, at least
one
therapeutic diol can be used in the polycondensation protocol.
[0093] The unsaturated PEUs can be prepared by interfacial solution
condensation of
di-p-toluenesulfonate salts of bis(a-amino acid)-alkylene diesters, comprising
at least one
double bond in R1. Unsaturated diols useful for this purpose include, for
example, 2-
butene-1,4-diol and 1,18-octadec-9-en-diol. Unsaturated monomer can be
dissolved prior
to the reaction in alkaline water solution, e.g. sodium liydroxide solution.
The water
solution can then be agitated intensely, under external cooling, with an
organic solvent
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layer, for example chloroform, which contains an equimolar amount of
monomeric,
dimeric or trimeric phosgene. An exothermic reaction proceeds rapidly, and
yields a
polymer that (in most cases) remains dissolved in the organic solvent. The
organic layer
can be washed several times with water, dried with anhydrous sodiunl sulfate,
filtered, and
evaporated. Unsaturated PEUs with a yield of about 75%-85% can be dried in
vacuum,
for example at about 45 C.
[0094] To obtain a porous, strong material, L-Leu based PEUs, such as 1-L-Leu-
4 and
1-L-Leu-6, can be fabricated using the general procedure described below. Such
procedure is less successful in formation of a porous, strong material wlien
applied to L-
Phe based PEUs.
[0095] The reaction solution or emulsion (about 100 mL) of PEU in chloroform,
as
obtained just after interfacial polycondensation, is added dropwise with
stirring to 1,000
mL of about 80 C -85 C water in a glass beaker, preferably a beaker made
hydrophobic
witlz dimethyldichlorsilane to reduce the adhesion of PEU to the beaker's
walls. The
polymer solution is broken in water into small drops and chloroform evaporates
rather
vigorously. Gradually, as chloroform is evaporated, small drops combine into a
compact
tar-like mass that is transformed into a sticky rubbery product. This rubbery
product is
removed from the beaker and put into hydrophobized cylindrical glass-test-
tube, which is
thermostatically controlled at about 80 C for about 24 hours. Then the test-
tube is
removed from the thermostat, cooled to room temperature, and broken to obtain
the
polymer. The obtained porous bar is placed into a vacuunl drier and dried
under reduced
pressure at about 80 C for about 24 hours. In addition, any procedure known
in the art for
obtaining porous polymeric materials can also be used.
[0096] Properties of high-molecular-weight porous PEUs made by the above
procedure
yielded results as summarized in Table 2.
Table 2. Properties of PEU Polymers of Formula (VI) and (VII)
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PEU* Yield yjred Mrv b) Mii ) M,v/Mu b) Tg e) Tm c)
[ /u] [dL/ ] [ C] C
1-L-Leu-4 80 0.49 84000 45000 1.90 67 103
1-L-Leu-6 82 0.59 96700 50000 1.90 64 126
1-L-Phe-6 77 0.43 60400 34500 1.75 - 167
[1-L-Leu-6]0,75- [1-L- 84 0.31 64400 43000 1.47 34 114
L s(OBn)]o.zs
1-L-Leu-DAS 57 0.28 55700 27700 2.1 ~ 56 165
*PEUs of general formula (VI), where,
1-L-Leu-4: R4 = (CH2)4, R3 = i-C4H9
1-L-Leu-6: R4 = (CH2)6a R3 = i-C4H9
1-L-Phe-6:.R4 = (CH2)6, R3 = -CH2-C6H5.
1-L-Leu-DAS: R4 = 1,4:3,6-dianhydrosorbitol, R3 = i-C4H
a) Reduced viscosities were measured in DMF at 25 C and a concentration 0.5
g/dL.
b) GPC Measurements were carried out in DMF, (PMMA).
'> Tg taken from second heating curve from DSC Measurements (heating rate10
C/min).
d) GPC Measurements were carried out in DMAc, (PS).
[0097] Tensile strength of illustrative synthesized PEUs was measured and
results are
summarized in Table 3. Tensile strength measurement was obtained using
dumbbell-
shaped PEU films (4 x 1.6 cm), which were cast from chloroform solution with
average
thickness of 0.125 mm and subjected to tensile testing on tensile strength
machine
(Chatillon TDC200) integrated with a PC using Nexygen FM software (Amtek,
Largo,
FL) at a crosshead speed of 60 nun/min. Examples illustrated herein can be
expected to
have the following mechanical properties:
[0098] 1. A glass transition temperature in the range from about 30 C to
about 90
C , for example, in the range from about 35 C to about 70 C ;
[0099] 2. A filni of the polymer with average thickness of about 1.6 cm will
have
tensile stress at yield of about 20 Mpa to about 150 Mpa, for example, about
25 Mpa to
about 60 Mpa;
[0100] 3. A film of the polymer with average thickness of about 1.6 cm will
have a
percent elongation of about 10 % to about 200%, for example about 50 % to
about 150%;
and
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[0101] 4. A film of the polymer with average thickness of about 1.6 cm will
have a
Young's modulus in the range from about 500 MPa to about 2000 MPa. Table 2
below
summarizes the properties of exemplary PEUs of this type.
Table 3. Mechanical Properties of PEUs
a> Tensile Stress Yotuig's
Polymer designation og at Yield Percent o Modulus
( C) (MPa) Elongation ( /o) (MPa)
1-L-Leu-6 64 21 114 622
[1-L-Leu-6]0,75- [1-L-Lys(OBn)]o,2534 25 159 915
[0102] The PEA, PEUR and PEU polymers described herein can be fabricated in a
variety of molecular weights and a variety of w/w% concentrations of the
therapeutic diol
or di-acid in the backbone of the polymer. The appropriate molecular weight
for use with
a given concentration of bioactive agent is readily determined by one of skill
in the art.
Thus, e.g., a suitable molecular weight will be on the order of about 5,000 to
about
300,000, for example about 5,000 to about 250,000, or about 75,000 to about
200,000, or
about 100,000 to about 150,000 and a suitable w/w% concentration of a residue
of a
bioactive agent incorporated into the backbone of the polymer will be on the
order of
about 5 w/w% to about 70 w/w%, for example about 10 w/w% to about 40 w/w%, or
about 20 w/w% to about 40 w/w%. The amount of bioactive agent incorporated
into the
backbone of the polymer will be highest in the case of a homopolymer (e.g.,
containing no
Lysine-based monomer) that incorporates both a therapeutic diol and a
therapeutic di-acid.
[0103] The molecular weights and polydispersities herein are determined by gel
permeation chromatography (GPC) using polystyrene standards. More
particularly,
number and weight average molecular weights (Mõ and M,,) are determined, for
example,
using a Model 510 gel permeation chromatography (Water Associates, Inc.,
Milford, MA)
equipped with a high-pressure liquid cliromatographic pump, a Waters 486 UV
detector
and a Waters 2410 differential refractive index detector. Tetrahydrofuran
(THF) or N,N-
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dimethylacetamide (DMAc) is used as the eluent (1.0 mL/min). The polystyrene
standards have a narrow molecular weight distribution.
[0104] While the optional bioactive agent(s) can be dispersed within the
polymer
matrix without chemical linkage to the polymer carrier, it is also
contemplated that one or
more bioactive agents or covering molecules can be covalently bound to the
biodegradable
polymers via a wide variety of suitable functional groups. For example, a free
carboxyl
group can be used to react with a complimentary moiety on a bioactive agent or
covering
molecule, such as an hydroxy, amino, or thio group, and the like. A wide
variety of
suitable reagents and reaction conditions are disclosed, e.g., in March's
Advanced Organic
Chemistry, Reactions, Mechanisms, and Structure, Fifth Edition, (2001); and
Comprehensive Organic Transformations, Second Edition, Larock (1999).
[0105] In other embodiments, one or more bioactive agent can be linked to any
of the
polymers of structures (I) and (III-VII) through an amide, ester, ether,
amino, ketone,
thioether, sulfinyl, sulfonyl, or disulfide linlcage. Such a linkage can be
formed from
suitably functionalized starting materials using synthetic procedures that are
known in the
art.
[0106] For example, in one embodiment a polymer can be linked to a bioactive
agent
via a free carboxyl group (e.g., COOH) of the polymer. Specifically, a
compound of
structures (I) and (III) can react with an amino functional group or a
hydroxyl functional
group of a bioactive agent to provide a biodegradable polymer having the
bioactive agent
attached via an amide linkage or ester linkage, respectively. In another
embodiment, the
carboxyl group of the polymer can be benzylated or transformed into an acyl
halide, acyl
anhydride/"mixed" anhydride, or active ester. In other embodiments, the free -
NH2 ends
of the polymer molecule can be acylated to assure that the bioactive agent
will attach only
via a carboxyl group of the polymer and not to the free ends of the polymer.
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[0107] Water soluble covering molecule(s), such as poly(ethylene glycol)
(PEG);
phosphatidylcholine (PC); glycosaminoglycans including heparin;
polysaccharides
including chitosan, alginates and polysialic acid; poly(ionizable or polar
amino acids)
including polyserine, polyglutamic acid, polyaspartic acid, polylysine and
polyarginine; as
described herein, and targeting molecules, such as antibodies, antigens and
ligands, are
bioactive agents that can also be conjugated to the polymer on the exterior of
particles
formed from the therapeutic polymer composition after production of the
particles to block
active sites not occupied by a bioactive agent or to target delivery of the
particles to a
specific body site as is known in the art. The molecular weights of PEG
molecules on a
single particle can be substantially any molecular weight in the range from
about 200 to
about 200,000, so that the molecular weights of the various PEG molecules
attached to the
particle can be varied.
[0108] Alternatively, a bioactive agent or covering molecule can be attached
to the
polymer via a linlcer molecule or by cross-linking two or more molecules of
the polymer
as described herein. Indeed, to improve surface hydrophobicity of the
biodegradable
polymer, to improve accessibility of the biodegradable polymer towards enzyme
activation, and to improve the release profile of the bioactive agents from
the
biodegradable polymer, a linker may be utilized to indirectly attach a
bioactive agent to
the biodegradable polymer. In certain embodiments, the linker compounds
include
poly(ethylene glycol) having a molecular weight (MW) of about 44 to about
10,000,
preferably 44 to 2000; amino acids, such as serine; polypeptides with repeat
number from
1 to 100; and any other suitable low molecular weight polymers. The linker
typically
separates the bioactive agent from the polymer by about 5 angstroms up to
about 200
angstroms.
[0109] In still further embodiments, the linker is a divalent radical of
formula W-A-Q,
wherein A is (C1-C24) alkyl, (C2-C24) alkenyl, (C2-C24) alkynyl, (C2-C20)
alkyloxy, (C3-C8)
cycloalkyl, or (C6-Clo) aryl, and W and Q are each independently N(R)C(=O)-, -
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C(=O)N(R)-, -OC(=O)-, -C(=O)O, -0-, -S-, -S(O), -S(0)2-, -S-S-, -N(R)-, -C(=0)-
,
wherein each R is independently H or (C1-C6) alkyl.
[0110] As used to describe the above linkers, the term "alkyl" refers to a
straight or
branched chain hydrocarbon group including methyl, ethyl, n-propyl, isopropyl,
n-butyl,
isobutyl, tert-butyl, n-hexyl, and the like.
[0111] As used herein used to describe the above linkers, "alkenyl" refers to
straight or
branched chain hydrocarbyl groups having one or more carbon-carbon double
bonds.
[0112] As used herein used to describe the above linkers, "allcynyl" refers to
straight or
branched chain hydrocarbyl groups having at least one carbon-carbon triple
bond.
[0113] As used herein used to describe the above linlcers, "aryl" refers to
aromatic
groups having in the range of 6 up to 14 carbon atoms.
[0114] In certain embodiments, the linker may be a polypeptide having from
about 2 up
to about 25 amino acids. Suitable peptides contemplated for use include poly-L-
glycine,
poly-L-lysine, poly-L-glutamic acid, poly-L-aspartic acid, poly-L-histidine,
poly-L-
ornithine, poly-L-serine, poly-L-threonine, poly-L-tyrosine, poly-L-leucine,
poly-L-lysine-
L-phenylalanine, poly-L-arginine, poly-L-lysine-L-tyrosine, and the like.
[0115] In one embodiment, a bioactive agent can covalently crosslink the
polymer, i.e.
the bioactive agent is bound to more than one polymer molecule, to form an
intermolecular bridge. This covalent crosslinking can be done with or without
a linker
containing a bioactive agent.
[0116] A bioactive agent molecule can also be incorporated into an
intramolecular
bridge by covalent attachment between two sites on the same polymer molecule.
[0117] A linear polymer polypeptide conjugate is made by protecting the
potential
nucleophiles on the polypeptide backbone and leaving only one reactive group
to be bound
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to the polymer or polymer linker construct. Deprotection is performed
according to
methods well known in the art for deprotection of peptides (Boc and Fmoc
chemistry for
example).
[0118] In one embodiment of the present invention, a bioactive agent is a
polypeptide
presented as a retro-inverso or partial f=etro-inverso peptide.
[0119] In other embodiments, a bioactive agent may be mixed with a
photocrosslinkable version of the polymer in a matrix, and, after
crosslinking, the material
is dispersed (ground) to form particles having an average diameter in the
range from about
0.1 to about 10 m.
[0120] The linker can be attached first to the polymer or to the bioactive
agent or
covering molecule. During synthesis, the linker can be either in unprotected
form or
protected from, using a variety of protecting groups well known to those
skilled in the art.
In the case of a protected linker, the unprotected end of the linker can first
be attached to
the polymer or the bioactive agent or covering molecule. The protecting group
can then
be de-protected using Pd/H2 hydrogenation for saturated polyiner backbones,
mild acid or
base hydrolysis for unsaturated polymers, or any other common de-protection
method that
is known in the art. The de-protected linker can then be attached to the
bioactive agent or
covering molecule, or to the polymer
[0121] An exemplary conjugate synthesis performed on a biodegradable polymer
according to the invention (wherein the molecule to be attached to the polymer
is an amino
substituted aminoxyl N-oxide radical) is set forth as follows. A biodegradable
polymer
herein can be reacted with an aminoxyl radical containing compound, e.g., 4-
amino-
2,2,6,6-tetramethylpiperidine-l-oxy, in the presence of N,N'-carbonyl
diimidazole or
suitable carbodiimide, to replace the hydroxyl moiety in the carboxyl group,
either on the
pendant carboxylic acids of the PEAs, PEURs or PEUs, or at the chain end of a
polyester
as described, with an amide linkage to the aminoxyl (N-oxide) radical
containing group.
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The amino moiety covalently bonds to the carbon of the carbonyl residue such
that an
amide bond is formed. The N,N'-carbonyldiimidazole or suitable carbodiimide
converts
the hydroxyl moiety in the carboxyl group at the chain end of the polyester
into an
intermediate activated moiety which will react with the amino group of the
aminoxyl (N
oxide) radical compound, e.g., the aniine at position 4 of 4-amino-2,2,6,6-
tetranlethylpiperidine-l-oxy. The aminoxyl reactant is typically used in a
mole ratio of
reactant to polyester ranging from 1:1 to 100:1. The mole ratio of N,N'-
carbonyldiimidazole or carbodiimide to aminoxyl is preferably about 1:1.
[0122] A typical reaction is as follows. A polyester is dissolved in a
reaction solvent
and reaction is readily carried out at the temperature utilized for the
dissolving. The
reaction solvent may be any in which the polyester will dissolve; this
information is
normally available from the manufacturer of the polyester. When the polyester
is a
polyglycolic acid or a poly(glycolide-L-lactide) (having a monomer mole ratio
of glycolic
acid to L-lactic acid greater than 50:50), highly refined (99.9+% pure)
dimethyl sulfoxide
at 115 C to 130 C or DMSO at room temperature suitably dissolves the
polyester. When
the polyester is a poly-L-lactic acid, a poly-DL-lactic acid or a
poly(glycolide-L-lactide)
(having a monomer mole ratio of glycolic acid to L-lactic acid 50:50 or less
than 50:50),
tetrahydrofuran, dichloromethane (DCM) and chloroform at room temperature to
40 -50
C suitably dissolve the polyester.
[0123] The product may be precipitated from the reaction mixture by adding
cold non-
solvent for the product. For example, aminoxyl-containing polyglycolic acid
and
aminoxyl-containing poly(glycolide-L-lactide) formed from glycolic acid-rich
monomer
mixture are readily precipitated from hot dimethylsulfoxide by adding cold
methanol or
cold acetone/methanol mixture and then recovered, e.g., by filtering. When the
product is
not readily precipitated by adding cold non-solvent for the product, the
product and
solvent may be separated by using vacuum techniques. For example, aminoxyl-
containing
poly-L-lactic acid is advantageously separated from solvent in this way. The
recovered
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product is readily further purified by washing away water and by-products
(e.g. urea) with
a solvent which does not dissolve the product, e.g., methanol in the case of
the modified
polyglycolic acid, polylactic acid and poly(glycolide-L-lactide) products
herein. Residual
solvent from such washing may be removed using vacuum drying.
Polymer - Bioactive agent Linkage
[0124] In one embodiment, the polymers used to make the invention therapeutic
polymer compositions as described herein have one or more bioactive agent
directly
linked to the polymer. The residues of the polymer can be linked to the
residues of the one
or more bioactive agents. For example, one residue of the polymer can be
directly linked
to one residue of a bioactive agent. The polymer and the bioactive agent can
each have
one open valence. Alternatively, more than one bioactive agent, multiple
bioactive agents,
or a mixture of bioactive agents having different therapeutic or palliative
activity can be
directly linked to the polyiner. However, since the residue of each bioactive
agent can be
linked to a corresponding residue of the polymer, the number of residues of
the one or
more bioactive agents can correspond to the number of open valences on the
residue of the
polymer having at least one diol or di-acid bioactive agent incorporated into
the baclcbone
of the polymer.
[0125] As used herein, a "residue of a polynier" refers to a radical of a
polymer having
one or more open valences. Any synthetically feasible atom, atoms, or
functional group of
the polymer (e.g., on the polymer backbone or pendant group) is substantially
retained
when the radical is attached to a residue of a bioactive agent. Additionally,
any
synthetically feasible functional group (e.g., carboxyl) can be created on the
polymer (e.g.,
on the polymer baclcbone as a pendant group or as chain termini) to provide
the open
valence, provided bioactivity of the backbone therapeutic agent is
substantially retained
when the radical is attached to a residue of a bioactive agent. Based on the
linkage that is
desired, those skilled in the art can select suitably functionalized starting
materials that can
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be used to derivatize the PEA and PEUR polymers used in the present invention
using
procedures that are known in the art.
[0126] As used herein, a "residue of a compound of structural formula (*)"
refers to a
radical of a compound of polymer formulas (I), (III)-(VII) as described herein
having one
or more open valences. Any synthetically feasible atom, atoms, or functional
group of the
compound (e.g., on the polymer baclcbone, pendant or end group) can be removed
to
provide the open valence, provided bioactivity of the backbone therapeutic
agent is
substantially retained when the radical is attached. Additionally, any
synthetically feasible
functional group (e.g., carboxyl) can be created on the compound of formulas
(I), (III)-
(VII) (e.g., on the polymer baclcbone or pendant group) to provide the open
valence,
provided bioactivity of the baclcbone therapeutic agent is substantially
retained when the
radical is attached to a residue of a bioactive agent. Based on the linkage
that is desired,
those skilled in the art can select suitably functionalized starting materials
that can be used
to derivatize the compound of formulas (I), (III)-(VII) using procedures that
are known in
the art.
[0127] For example, the residue of a bioactive agent can be linked to the
residue of a
compound of structural formula (I)-(III)-(VII) through an amide (e.g., -
N(R)C(=0)- or -
C(=O)N(R)-), ester (e.g., -OC(=O)- or -C(=O)O-), ether (e.g., -0-), amino
(e.g., -N(R)-),
ketone (e.g., -C(=O)-), thioether (e.g., -S-), sulfinyl (e.g., -S(O)-),
sulfonyl (e.g., -S(O)Z-),
disulfide (e.g., -S-S-), or a direct (e.g., C-C bond) linkage, wherein each R
is
independently H or (C1-C6) alkyl. Such a linkage can be formed from suitably
fiulctionalized starting materials using synthetic procedures that are known
in the art.
Based on the linkage that is desired, those skilled in the art can select
suitably functional
starting material to derivatize any residue of a compound of structural
formula (I) or (III) -
(VII) and thereby conjugate a given residue of a bioactive agent using
procedures that are
known in the art. The residue of the optional bioactive agent can be linlced
to any
synthetically feasible position on the residue of a compound of structural
formula (I) or
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(III) - (VII). Additionally, the invention also provides compounds having more
than one
residue of a bioactive agent directly linlced to a compound of structural
fonnula (I), (III)-
(VII).
[0128] The number of bioactive agents that can be linlced to the polymer
molecule can
typically depend upon the molecular weight of the polymer and the number of
baclcbone
therapeutic agents incorporated into the polymer. For example, for a compound
of
structural formula (I), wherein n is about 5 to about 150, preferably about 5
to about 70, up
to about 150 bioactive agent molecules (i.e., residues thereof) can be
directly linked to the
polymer (i.e., residue thereof) by reacting the bioactive agent with backbone,
pendant or
terminal groups of the polymer. The number of sites for linkage of a bioactive
agent in the
invention therapeutic polyiner compositions is accordingly reduced by the
number of
baclcbone therapeutic diol or di-acids incorporated into the polymer. In
unsaturated
polymers, bioactive agents can also be reacted with double (or triple) bonds
in the
polymer, provided that the therapeutic diol or di-acid residues incorporated
into the
polymer backbone do not contain any double (or triple) bonds themselves.
Hence, in the
case of estradiol incorporated into the baclcbone, linkage of a bioactive
agent at a double
bond in the polymer composition would not be recommended, to prevent bonding
of the
bioactive agent to a double bond in the backbone diol or di-acid residue
(i.e., the estradiol)
in a reaction.
[0129] In the therapeutic polymer composition, either in the fonn of particles
or not, a
bioactive agent can be covalently attached directly to the polymer, rather
than being
dispersed by "loading" into the polymer without chemical attachment, using any
of several
methods well known in the art and as described hereinbelow. The amount of
bioactive
agent is generally approximately 0.1 % to about 60% (w/w) bioactive agent to
polymer
composition, more preferably about 1% to about 25% (w/w) bioactive agent, and
even
more preferably about 2% to about 20% (w/w) bioactive agent. The percentage of
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bioactive agent will depend on the desired dose and the condition being
treated, as
discussed in more detail below.
[0130] In addition to serving as stand-alone delivery systems for therapeutic
diols and
di-acids (and optional bioactive agents) when directly administered in vivo,
for example,
in the form of inhalants, implants or local or systemic injectables, the
invention therapeutic
polymer compositions can be used in the fabrication of polymer coatings for
various types
of surgical devices. In this embodiment, the polymer coating on the surgical
device is
effective for controlled delivery to surrounding tissue of the therapeutic
diol or di-acid as
well as any bioactive agents dispersed in the polymer or covalently attached
to the surface
of a particle thereof.
[0131] In one embodiment, the invention therapeutic polymer composition can be
fabricated in the form of a pad, sheet or wrap of any desired surface area.
For example,
the polymer can be woven or formed as a thin sheet of randomly oriented
fibers. Such
pads, sheets and wraps can be used in a number of types of wound dressings for
treatment
of a variety of conditions, for example by promoting endogenous healing
processes at a
wound site. The polymer compositions in the wound dressing biodegrade over
time,
releasing the therapeutic diol or di-acid to be absorbed into a target cell in
a wound site
where it acts intracellularly, either within the cytosol, the nucleus, or
both, or the bioactive
agent can bind to a cell surface receptor molecule to elicit a cellular
response without
entering the cell. Alternatively, the therapeutic diol or di-acid released
from the polymer
composition, for example when used as the covering for a bioactive stent,
promotes
endogenous healing processes at the wound site by contact with the
surroundings into
which the wound dressing or implant is placed. A detailed description of wound
dressings, wound healing implants and surgical device coatings made using PEA
and
PEUR polymers is found in co-pending U.S. Patent Application Serial No.
11/128,903,
filed May 12, 2005.
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[0132] A detailed description of inetliods of making polymer particles using
PEA and
PEUR polynlers may be found in co-pending U.S. provisional applications
60/654,715,
filed February 17, 2005, and 60/674,670, May 25, 2005, each of which is
incorporated
herein in its entirety.
[0133] Bioactive agents contemplated for dispersion within the polymers used
in the
invention therapeutic polymer compositions include anti-proliferants,
rapamycin and any
of its analogs or derivatives, paclitaxel or any of its taxene analogs or
derivatives,
everolimus, sirolimus, tacrolimus, or any of its -limus named family of drugs,
and statins
such as simvastatin, atorvastatin, fluvastatin, pravastatin, lovastatin,
rosuvastatin,
geldanamycins, such as 17AAG (17-allylamino-17-demethoxygeldanamycin);
Epothilone
D and other epothilones, 17-dimethylaminoethylamino-17-demethoxy-geldanamycin
and
other polyketide inhibitors of heat shock protein 90 (Hsp90), cilostazol, and
the like.
[0134] Suitable bioactive agents for dispersion in the invention therapeutic
polymer
compositions and particles made therefrom also can be selected from those that
promote
endogenous production of a therapeutic natural wound healing agent, such as
nitric oxide,
which is endogenously produced by endothelial cells. Alternatively the
bioactive agents
released from the polymers during degradation may be directly active in
promoting natural
wound healing processes by endothelial cells. These bioactive agents can be
any agent
that donates, transfers, or releases nitric oxide, elevates endogenous levels
of nitric oxide,
stimulates endogenous synthesis of nitric oxide, or serves as a substrate for
nitric oxide
synthase or that inhibits proliferation of smooth muscle cells. Such agents
include, for
example, aminoxyls, furoxans, nitrosothiols, nitrates and anthocyanins;
nucleosides such
as adenosine and nucleotides such as adenosine diphosphate (ADP) and adenosine
triphosphate (ATP); neurotransmitter/neuromodulators such as acetylcholine and
5-
hydroxytryptamine (serotonin/5-HT); histamine and catecholamines such as
adrenalin and
noradrenalin; lipid molecules such as sphingosine- 1 -phosphate and
lysophosphatidic acid;
amino acids such as arginine and lysine; peptides such as the bradykinins,
substance P and
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calcium gene-related peptide (CGRP), and proteins such as insulin, vascular
endothelial
growtli factor (VEGF), and thrombin.
[0135] A variety of bioactive agents, coating molecules and ligands for
bioactive
agents can be attached, for example covalently, to the surface of the polymer
coatings or
particles. Bioactive agents, such as targeting antibodies, polypeptides (e.g.,
antigens) and
drugs can be covalently conjugated to the surface of the polymer coatings or
particles. In
addition, coating molecules, such as polyethylene glycol (PEG) as a ligand for
attachment
of antibodies or polypeptides or phosphatidylcholine (PC) as a means of
blocking
attachment sites on the surface of the particles, can be surface-conjugated to
the particles
to prevent the particles from sticking to non-target biological molecules and
surfaces in a
subject to which the particles are administered.
[0136] For example, small proteinaceous motifs, such as the B domain of
bacterial
Protein A and the functionally equivalent region of Protein G are known to
bind to, and
thereby capture, antibody molecules by the Fc region. Such proteinaceous
motifs can be
attached as bioactive agents to the invention therapeutic polymer
compositions, especially
to the surface of the polymer particles described herein. Such molecules will
act, for
example, as ligands to attach antibodies for use as targeting ligands or to
capture
antibodies to hold precursor cells or capture cells out of the blood stream.
Therefore, the
antibody types that can be attached to polymer coatings using a Protein A or
Protein G
functional region are those that contain an Fc region. The capture antibodies
will in turn
bind to and hold precursor cells, such as progenitor cells, near the polymer
surface while
the precursor cells, which are preferably bathed in a growth medium within the
polynier,
secrete various factors and interact with other cells of the subject. In
addition, one or more
bioactive agents dispersed in the polymer particles, such as the bradykinins,
may activate
the precursor cells.
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[0137] In addition, bioactive agents for attaching precursor cells or for
capturing
progenitor endothelial cells (PECs) from a blood stream in a subject to which
the polymer
compositions are administered are monoclonal antibodies directed against a
lcnown
precursor cell surface marker. For example, complementary determinants (CDs)
that have
been reported to decorate the surface of endothelial cells include CD3 1,
CD34, CD 102,
CD105, CD106, CD109, CDw130, CD141, CD142, CD143, CD144, CDw145, CD146,
CD147, and CD166. These cell surface markers can be of varying specificity and
the
degree of specificity for a particular cell/developmental type/stage is in
many cases not
fully characterized. Iii addition, these cell marker molecules against which
antibodies
have been raised will overlap (in terms of antibody recognition) especially
with CDs on
cells of the same lineage: monocytes in the case of endothelial cells.
Circulating
endothelial progenitor cells are some way along the developmental pathway from
(bone
marrow) monocytes to mature endothelial cells. CDs 106, 142 and 144 have been
reported to mark mature endothelial cells with some specificity. CD34 is
presently known
to be specific for progenitor endothelial cells and therefore is currently
preferred for
capturing progenitor endothelial cells out of blood in the site into which the
polymer
particles are implanted for local delivery of the active agents. Examples of
such
antibodies include single-chain antibodies, chimeric antibodies, monoclonal
antibodies,
polyclonal antibodies, antibody fragments, Fab fragments, IgA, IgG, IgM, IgD,
IgE and
humanized antibodies, and active fragments thereof.
[0138] The following bioactive agents and small molecule drugs will be
particularly
effective for dispersion within the invention therapeutic polymer
compositions, whether
sized to form a time release biodegradable polymer depot for local delivery of
the
bioactive agents, or sized for entry into systeinic circulation, as described
herein. The
bioactive agents that are dispersed in the invention therapeutic polymer
coinpositions and
methods of use will be selected for their suitable therapeutic or palliative
effect in
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treatment of a disease of interest, or symptoms thereof, or in experiments
designed for in
vitro testing of such effects in cells or tissue culture, or in vivo.
[0139] In one embodiment, the suitable bioactive agents are not limited to,
but include,
various classes of compounds that facilitate or contribute to wound healing
when
presented in a time-release fashion. Such bioactive agents include wound-
healing cells,
including certain precursor cells, which can be protected and delivered by the
biodegradable polymer in the invention compositions. Such wound healing cells
include,
for example, pericytes and endothelial cells, as well as inflammatory healing
cells. To
recruit such cells to the site of a polymer depot in vivo, the invention
therapeutic polymer
compositions and particles thereof used in the invention and methods of use
can include
ligands for such cells, such as antibodies and smaller molecule ligands, that
specifically
bind to "cellular adhesion molecules" (CAMs). Exemplary ligands for wound
healing
cells include those that specifically bind to Intercellular adhesion molecules
(ICAMs),
such as ICAM-1 (CD54 antigen); ICAM-2 (CD102 antigen); ICAM-3 (CD50 antigen);
ICAM-4 (CD242 antigen); and ICAM-5; Vascular cell adhesion molecules (VCAMs),
such as VCAM-1 (CD106 antigen); Neural cell adhesion molecules (NCAMs), such
as
NCAM-1 (CD56 antigen); or NCAM-2; Platelet endothelial cell adhesion molecules
PECAMs, such as PECAM-1 (CD31 antigen); Leukocyte-endothelial cell adhesion
molecules (ELAMs), such as LECAM-1; or LECAM-2 (CD62E antigen), and the like.
[0140] In another aspect, the suitable bioactive agents include extra cellular
matrix
proteins, macromolecules that can be dispersed into the polymer particles used
in the
invention therapeutic polymer compositions, e.g., attached either covalently
or non-
covalently. Examples of useful extra-cellular matrix proteins include, for
example,
glycosaminoglycans, usually linked to proteins (proteoglycans), and fibrous
proteins (e.g.,
collagen; elastin; fibronectins and laminin). Bio-mimics of extra-cellular
proteins can also
be used. These are usually non-human, but biocompatible, glycoproteins, such
as
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alginates and chitin derivatives. Wound healing peptides that are specific
fragments of
such extra-cellular matrix proteins and/or their bio-mimics can also be used.
[0141] Proteinaceous growth factors are another category of bioactive agents
suitable
for dispersion in the invention therapeutic polymer compositions and methods
of use
described herein. Such bioactive agents are effective in promoting wound
healing and
other disease states as is known in the art, for example, Platelet Derived
Growth Factor-
BB (PDGF-BB), Tuinor Necrosis Factor-a (TNF-a), Epidermal Growth Factor (EGF),
Keratinocyte Growth Factor (KGF), Thymosin B4; and, various angiogenic factors
such as
vascular Endothelial Growth Factors (VEGFs), Fibroblast Growth Factors (FGFs),
Tumor
Necrosis Factor-beta (TNF -beta), and Insulin-like Growth Factor-1 (IGF-1).
Many of
these proteinaceous growth factors are available commercially or can be
produced
recombinantly using techniques well known in the art.
[0142] Alternatively, expression systems comprising vectors, particularly
adenovirus
vectors, incorporating genes encoding a variety of biomolecules can be
dispersed in the
invention therapeutic polymer compositions and particles thereof for timed
release
delivery. Methods of preparing such expression systems and vectors are well
known in
the art. For example, proteinaceous growth factors can be dispersed into the
invention
therapeutic polymer compositions for administration of the growth factors
either to a
desired body site for local delivery, by selection of particles sized to form
a polymer
depot, or systemically, by selection of particles of a size that will enter
the circulation.
Growth factors, such as VEGFs, PDGFs, FGF, NGF, and evolutionary and
functionally
related biologics, and angiogenic enzymes, such as thrombin, may also be used
as
bioactive agents in the invention compositions.
[0143] Small molecule drugs are yet another category of bioactive agents
suitable for
dispersion in the invention therapeutic polymer compositions and methods of
use
described herein. Such drugs include, for example, antimicrobials and anti-
inflammatory
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agents as well as certain healing promoters, such as, for example, vitamin A
and synthetic
inhibitors of lipid peroxidation.
[0144] A variety of antibiotics can be dispersed as bioactive agents in the
invention
therapeutic polymer compositions to indirectly promote natural healing
processes by
preventing or controlling infection. Suitable antibiotics include many
classes, such as
aminoglycoside antibiotics or quinolones or beta-lactams, such as
cefalosporins, e.g.,
ciprofloxacin, gentamycin, tobramycin, erythromycin, vancomycin, oxacillin,
cloxacillin,
methicillin, lincomycin, ampicillin, and colistin. Suitable antibiotics have
been described
in the literature.
[0145] Suitable antimicrobials include, for example, Adriamycin PFS/RDF
(Pharmacia and Upjohn), Blenoxane (Bristol-Myers Squibb Oncology/Immunology),
Cerubidine (Bedford), Cosmegen (Merck), DaunoXome (NeXstar), Doxil
(Sequus), Doxorubicin Hydrochloride (Astra), Idamycin PFS (Pharmacia and
Upjohn),
Mithracin (Bayer), Mitamycin (Bristol-Myers Squibb Oncology/Immunology),
Nipen (SuperGen), Novantrone (Inununex) and Rubex (Bristol-Myers Squibb
Oncology/Immunology). In one embodiment, the peptide can be a glycopeptide.
"Glycopeptide" refers to oligopeptide (e.g. heptapeptide) antibiotics,
characterized by a
multi-ring peptide core optionally substituted with saccharide groups, such as
vancomycin.
[0146] Examples of glycopeptides included in this category of antimicrobials
may be
found in "Glycopeptides Classification, Occurrence, and Discovery," by Raymond
C. Rao
and Louise W. Crandall, ("Bioactive agents and the Pharmaceutical Sciences"
Volume 63,
edited by Ramakrishnan Nagarajan, published by Marcal Dekker, Inc.).
Additional
examples of glycopeptides are disclosed in U.S. Patent Nos. 4,639,433;
4,643,987;
4,497,802; 4,698,327, 5,591,714; 5,840,684; and 5,843,889; in EP 0 802 199; EP
0 801
075; EP 0 667 353; WO 97/28812; WO 97/38702; WO 98/52589; WO 98/52592; and in
J.
Amer. Chem. Soc.(1996)118: 13107-13108; J Amer. G'hem. Soc. (1997) 119:12041 -
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53
12047; and J. Amer. Claena. Soc. (1994) 116:4573-4590. Representative
glycopeptides
include those identified as A477, A35512, A40926, A41030, A42867, A47934,
A80407,
A82846, A83850, A84575, AB-65, Actaplanin, Actinoidin, Ardacin, Avoparcin,
Azureomycin, Balhimyein, Chloroorientiein, Chloropolysporin, Decaplanin, -
demethylvancomycin, Eremomycin, Galacardin, Helvecardin, Izupeptin, Kibdelin,
LL-AM374, Mannopeptin, MM45289, MM47756, MM47761, MM49721, MM47766,
MM55260, MM55266, MM55270, MM56597, MM56598, OA-7653, Orenticin,
Parvodicin, Ristocetin, Ristomycin, Synmonicin, Teicoplanin, UK-68597, UD-
69542,
UE'--72051, Vancomycin, and the like. The term "glycopeptide" or "glycopeptide
antibiotic" as used herein is also intended to include the general class of
glycopeptides
disclosed above on which the sugar moiety is absent, i.e. the aglycone series
of
glycopeptides. For example, removal of the disaccharide moiety appended to the
phenol
on vancomycin by mild hydrolysis gives vancomycin aglycone. Also included
within the
scope of the term "glycopeptide antibiotics" are synthetic derivatives of the
general class
of glycopeptides disclosed above, including alkylated and acylated
derivatives.
Additionally, within the scope of this term are glycopeptides that have been
further
appended with additional saccharide residues, especially aminoglycosides, in a
manner
similar to vancosamine.
[0147] The term "lipidated glycopeptide" refers specifically to those
glycopeptide
antibiotics that have been synthetically modified to contain a lipid
substituent. As used
herein, the term "lipid substituent" refers to any substituent contains 5 or
more carbon
atoms, preferably, 10 to 40 carbon atoms. The lipid substituent may optionally
contain
from 1 to 6 heteroatoms selected from halo, oxygen, nitrogen, sulfur, and
phosphorous.
Lipidated glycopeptide antibiotics are well known in the art. See, for
example, in U.S.
Patent Nos. 5,840,684, 5,843,889, 5,916,873, 5,919,756, 5,952,310, 5,977,062,
5,977,063,
EP 667, 353, WO 98/52589, WO 99/56760, WO 00/04044, WO 00/39156, the
disclosures
of which are incorporated herein by reference in their entirety.
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[0148] Anti-inflammatory bioactive agents are also useful for dispersion in
invention
therapeutic polymer compositions. Depending on the body site and disease to be
treated,
such anti-inflanunatory bioactive agents include, e.g. analgesics (e.g.,
NSAIDS and
salicyclates), steroids, antirheumatic agents, gastrointestinal agents, gout
preparations,
hormones (glucocorticoids), nasal preparations, ophthalmic preparations, otic
preparations
(e.g., antibiotic and steroid combinations), respiratory agents, and skin &
mucous
membrane agents. See, Physician's Desk Reference, 2005 Edition. Specifically,
the anti-
inflammatory agent can include dexamethasone, whicll is chemically designated
as (114,
16I)-9-fluro-11,17,21-trihydroxy-16-methylpregna-1,4-diene-3,20-dione.
Alternatively,
the anti-inflammatory bioactive agent can be or include sirolimus (rapamycin),
which is a
triene macrolide antibiotic isolated from Streptonzyces hygroscopicus.
[0149] The polypeptide bioactive agents included in the invention compositions
and
methods can also include "peptide mimetics." Such peptide analogs, referred to
herein as
"peptide mimetics" or "peptidomimetics," are commonly used in the
pharmaceutical
industry with properties analogous to those of the template peptide (Fauchere,
J. (1986)
Adv. Bioactive agent Res., 15:29; Veber and Freidinger (1985) TINS, p. 392;
and Evans et
al. (1987) J. Med. Cliem., 30:1229) and are usually developed with the aid of
computerized molecular modeling. Generally, peptidomimetics are structurally
similar to
a paradigm polypeptide (i.e., a polypeptide that has a biochemical property or
pharmacological activity), but have one or more peptide linkages optionally
replaced by a
linkage selected from the group consisting of.- --CHZNH--, --CH2S--, CH2-CH2--
, --
CH=CH-- (cis and trans), --COCH2--, --CH(OH)CH2--, and --CH2SO--, by methods
known in the art and further described in the following references: Spatola,
A.F. in
Claemistry and Biochemistry ofAmin.o Acids, Peptides, and Proteins, B.
Weinstein, eds.,
Marcel Dekker, New York, p. 267 (1983); Spatola, A.F., Vega Data (March 1983),
Vol. 1,
Issue 3, "Peptide Backbone Modifications" (general review); Morley, J.S.,
Trends. Pharm.
Sci., (1980) pp. 463-468 (general review); Hudson, D. et al., Int. J. Pept.
Prot. Res., (1979)
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14:177-185 (--CHa NH--, CH2CH2--); Spatola, A.F. et al., Life Sci., (1986)
38:1243-1249
(--CH2-S--); Harm, M. M., J. Claefn. Soc. Pef=kin Trans I(1982) 307-314 (--
CH=CH--,
cis and trans); Almquist, R.G. et al., J. Med. Claena., (1980) 23:2533 (--
COCH2--);
Jennings-Whie, C. et al., Tetrahedron Lett., (1982) 23:2533 (--COCH2--);
Szelke, M. et
al., European Appln., EP 45665 (1982) CA: 97:39405 (1982) (--CH(OH)CH2--);
Holladay,
M. W. et al., Tetrahedron Lett., (1983) 24:4401-4404 (--C(OH)CH2--); and
Hruby, V.J.,
Life Sci., (1982) 31:189-199 (--CHa-S--). Such peptide mimetics may have
significant
advantages over natural polypeptide embodiments, including, for example: more
economical production, greater chemical stability, enhanced pharmacological
properties
(half-life, absorption, potency, efficacy, etc.), altered specificity (e.g., a
broad-spectrum of
biological activities), reduced antigenicity, and others.
[0150] Additionally, substitution of one or more amino acids within a peptide
(e.g.,
with a D-Lysine in place of L-Lysine) may be used to generate more stable
peptides and
peptides resistant to endogenous peptidases. Alternatively, the synthetic
polypeptides
covalently bound to the biodegradable polymer, can also be prepared from D-
amino acids,
referred to as inverso peptides. When a peptide is assembled in the opposite
direction of
the native peptide sequence, it is referred to as a retro peptide. In general,
polypeptides
prepared from D-amino acids are very stable to enzymatic hydrolysis. Many
cases have
been reported of preserved biological activities for retro-inverso or partial
retro-inverso
polypeptides (US patent, 6,261,569 B1 and references therein; B. Fromme et al,
Endocrinology (2003)144:3262-3269.
[0151] It is readily apparent that the subject invention can be used to
prevent or treat a
wide variety of diseases or symptoms thereof.
[0152] Following preparation of the invention therapeutic polymer compositions
and
polymer particles thereof, optionally loaded with at least one bioactive
agent, the
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composition can be lyophilized and the dried composition suspended in an
appropriate
media prior to administration.
[0153] Any suitable and effective amount of the at least one bioactive agent
can be
released with time from the therapeutic polymer composition, including those
in a polymer
coating on a medical device, such as a stent or a depot formed from particles
thereof
introduced in vivo. The suitable and effective amount of the bioactive agent
will typically
depend, e.g., on the specific PEA or PEUR polymer and concentration of
therapeutic
backbone diol or di-acid incorporated therein, type of particle or
polymer/bioactive agent
linkage, if present. Typically, up to about 100% of the backbone diol(s) or di-
acid(s) and
optional bioactive agent(s) can be released from polymer particles sized to
avoid
circulation as described herein that form a polymer depot in vivo.
Specifically, up to about
90%, up to 75%, up to 50%, or up to 25% thereof can be released from the
polymer depot.
Factors that typically affect the release rate from the polymer depot are the
nature and
amount of the polymer/backbone therapeutic agent, the types of
polymer/bioactive agent
linkage, and the nature and amount of additional substances present in the
formulation.
[0154] Once the invention therapeutic polymer composition is made, as above,
the
composition is formulated for subsequent intrapulmonary, gastroenteral,
subcutaneous,
intramuscular, into the central nervous system, intraperitoneum or intraorgan
delivery.
The compositions will generally include one or more "pharmaceutically
acceptable
excipients or vehicles" appropriate for oral, mucosal or subcutaneous
delivery, such as
water, saline, glycerol, polyethylene glycol, hyaluronic acid, ethanol, etc.
Additionally,
auxiliary substances, such as wetting or emulsifying agents, pH buffering
substances,
flavorings, and the like, may be present in such vehicles.
[0155] For example, intranasal and pulmonary formulations will usually include
vehicles that neither cause irritation to the nasal mucosa nor significantly
disturb ciliary
function. Diluents such as water, aqueous saline or other known substances can
be
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enlployed with the subject invention. The intrapulmonary formulations may also
contain
preservatives such as, but not limited to, chlorobutanol and benzalkonium
chloride. A
surfactant may be present to enhance absorption by the nasal mucosa.
[0156] For rectal and urethral suppositories, the vehicle composition will
include
traditional binders and carriers, such as, cocoa butter (theobroma oil) or
other
triglycerides, vegetable oils modified by esterification, hydrogenation and/or
fractionation,
glycerinated gelatin, polyalkaline glycols, mixtures of polyethylene glycols
of various
molecular weights and fatty acid esters of polyethylene glycol.
[0157] For vaginal delivery, the invention therapeutic polymer compositions
can be
formulated in pessary bases, such as those including mixtures of polyethylene
triglycerides, or suspended in oils such as corn oil or sesame oil, optionally
containing
colloidal silica. See, e.g., Richardson et al., Int. J. Pharm. (1995) 115:9-
15.
[0158] For a further discussion of appropriate vehicles to use for particular
modes of
delivery, see, e.g., Remington: The Science and Practice of Pharmacy, Mack
Publishing
Company, Easton, Pa., 19th edition, 1995. One of skill in the art can readily
determine the
proper vehicle to use for the particular combination of PEA, PEUR or PEU
polymer with
backbone therapeutic agent or particles thereof and mode of administration.
[0159] In addition to humans, the invention therapeutic polymer compositions
are also
intended as delivery vehicles for use in veterinary administration of
bioactive agents to a
variety of mammalian patients, such as pets (for example, cats, dogs, rabbits,
and ferrets),
farm animals (for example, swine, horses, mules, dairy and meat cattle) and
race horses.
[0160] In one einbodiment, the therapeutic polymer compositions used in the
invention
methods of administration or delivery will comprise an "effective amount" of
one or more
backbone therapeutic diol or di-acid(s) and optional bioactive agents of
interest. That is,
an amount of a backbone diol or di-acid will be incorporated into the
composition that will
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produce a sufficient therapeutic or palliative response in order to prevent,
reduce or
eliminate symptoms. The exact amount necessary will vary, depending on the
subject to
which the composition is being administered; the age and general condition of
the subject;
the capacity of the subject's immune system, the degree of therapeutic or
palliative
response desired; the severity of the condition being treated or investigated;
the particular
therapeutic diol or di-acid selected and mode of administration of the
composition, anlong
other factors. An appropriate effective amount can be readily determined by
one of skill
in the art. Thus, an "effective amount" will fall in a relatively broad range
that can be
detemiined through routine trials. For exaniple, for purposes of the present
invention, an
effective amount will typically range from about 1 g to about 100 mg, for
example from
about 5 g to about 1 mg, or about 10 g to about 500 g of the active agent
delivered per
dose.
[0161] Once formulated, the invention therapeutic polymer compositions are
administered orally, mucosally, or by subcutaneously or intramuscular
injection, and the
like, using standard techniques. See, e.g., Remington: The Science and
Practice of
Pharnaac,y, Mack Publishing Company, Easton, Pa., 19th edition, 1995, for
mucosal
delivery techniques, including intranasal, pulmonary, vaginal and rectal
techniques, as
well as European Publication No. 517,565 and Illum et al., J. Controlled Rel.
(1994)
29:133-141, for techniques of intranasal adininistration.
[0162] Dosage treatment may be a single dose of the invention therapeutic
polymer
composition, or a multiple dose schedule as is known in the art. The dosage
regimen, at
least in part, will also be determined by the need of the subject and be
dependent on the
judgment of the practitioner. Furthermore, if prevention of disease is
desired, the
therapeutic polymer composition (in the form of particles, or not) is
generally
administered prior to primary disease manifestation, or symptoms of the
disease of
interest. If treatment is desired, e.g., the reduction of symptoms or
recurrences, the
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therapeutic polymer compositions are generally administered subsequent to
primary
disease manifestation.
[0163] The formulations can be tested in vivo in a number of animal models
developed
for the study of oral subcutaneous or inucosal delivery. For example, the
conscious sheep
model is an art-recognized model for testing nasal delivery of substances See,
e.g.,
Longenecker et al., J. Pharna. Sci. (1987) 76:351-355 and Illum et al., J.
Controlled Rel.
(1994) 29:133-141. The tlierapeutic polymer composition, generally in
powdered,
lyophilized form, is blown into the nasal cavity. Blood samples can be assayed
for active
agent using standard techniques, as known in the art.
[0164] The following examples are meant to illustrate, but not to limit the
invention.
EXAMPLE 1
[0165] Materials 17-(3-estradiol (estra-1,3,5(10)-triene-3,17(3-diol), L-
lysine, benzyl
alcohol, sebacoyl chloride, 1,6-Hexanediol, p-nitrophenol, triethylamine, 4-
N,N-
(dimethylamino)pyridine (DMAP), N,N'-dicyclohexylcarbodiimide (DCC), anhydrous
N,N-dimethylformamide (DMF), anhydrous dichloromethane (DCM), trifluoroacetic
acid
(TFA) , p-toluenesulfonic acid monohydrate (Aldrich Chemical Co., Milwaukee,
WI),
anhydrous toluene, Boc-L-leucine monohydrate (Calbiochem-Novabiochem, San
Diego,
CA) were used without further purification. Other solvents, ether and ethyl
acetate (Fisher
Chemical, Pittsburgh, PA).
[0166] Synthesis of Monomers and Polymers Synthesis of bioactive PEAs involved
three basic steps: (1) synthesis of bis(p-nitrophenyl) diesters of
dicarboxylic acid (of
sebacic acid, compound 1); (2) synthesis of di-p-toluenesulfonic acid salts
(or di-TFA salt)
of bis(L-leucine) diesters of diol (compounds 2 and 5) and of L-lysine benzyl
ester
(compound 2); and (3) solution polycondensation of the monomers obtained in
steps (1)
and (2).
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[0167] Synthesis of di-p-nitrophenyl esters of sebacic acid (compound 1) Di-p-
nitrophenyl ester of sebacic acid was prepared by reacting of sebacoyl
chloride with p-
nitrophenol as described previously (Katsarava et al. J. Polym. Sci. Part A:
Polyna. Cliem.
(1999) 37. 391-407) (Scheme 4):
O 0 _
CI TEA
cl~~~ + 02N OH ' 02N / \ O"O \ / NO2
O o
(1>
Scheme 4
[0168] Di-p-toluenesulfonic acid salt of L-lysine benzyl ester (2) was
prepared as
described earlier (US 6,503,538) by refluxing of benzyl alcohol,
toluenesulfonic acid
monohydrate and L-lysine monohydro-chloride in toluene, applying azeotropic
removal of
generated water (scheme 5).
Z n,_, ~~NHZ OH TosOH HOTos .H N NH2.TosOH
HCI.H2N --)0-
( 2 )
~
HO O + Toluene, O 11ii1III'L0 reflux
Scheme 5
[0169] Synthesis of acid salts of bis-(a-amino acid) diesters (3), (5) Di-p-
toluenesulfonic acid salt of bis-(L-leucine) hexane-1,6-diester (compound 3)
was prepared
by modified procedure of the previously published method as shown in scheme 6.
[0170] L-Leucine (0.132 mol), p-toluenesulfonic acid monohydrate (0.132 mol)
and
1,6-hexanediol (0.06 mol) in 250 mL of toluene were placed in a flask equipped
with a
Dean-Stark apparatus and overhead stirrer. The heterogeneous reaction mixture
was
heated to reflux for about 12 h unti14.3 mL (0.24 mol) of water evolved. The
reaction
mixture was then cooled to room temperature, filtered, washed with acetone,
and
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61
recrystallized twice from metlianol/toluene 2:1 mixture. Yields and Mp were
identical to
published data (Katsarava et al., supra) (see scheme 6).
O O O
H2N OH HO ::: HOTos.H 2N O NH2.TosOH
+ ~OH , (3)
reflux
Scheme 6
[0171] A di-TFA salt ofbis-L-leucine-(3-estradiol-diester (compound 5) was
prepared
by a two step reaction. 17-(3-Estradiol was first reacted with Boc-protected L-
Leucine,
applying carbodiimide mediated esterification, to form compound 4. In a second
step, Boc
groups were deprotected using TFA, converting at the same time into a di-TFA
salt of di-
amino monomer (compound 5) (see Schenle 7).
O
OH
O N O~
+ DCC,DMAP p H
2 Boc.N COOH DI'1F, RT
H 74% O
HO ~O~N O (4)
H O
TFA.H2N\ /O
(4) A O O NH2.TFA
DCM O
(5)
Scheme 7
[0172] Preparation of Bis(Boc-L-leucine)estradiol-3,17-(3-diester (5) 1.5 g
(5.51
mmol) of 17-(3-estradiol, 3.43 g (13.77 mmol) Boc-L-leucine monoliydrate and
0.055 g
(0.28 mmol) of p-toluenesulfonic acid monohydrate were dissolved into 20 mL of
dry
N,N-dimethylformamide at room temperature under a dry nitrogen atmosphere. To
this
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62
solution lOg of nlolecular sieves were added and stirring continued for 24 h.
Then, 0.067
g of DMAP and 5.4g of (26.17 mmol) DCC were introduced into the reaction
solution and
stirring was continued. After 6 h (no discoloration of the reaction was
observed), 1 mL of
acetic acid was added to destroy the excess of DCC. Precipitated urea and
sieves then
were filtered off and filtrate poured in 80 mL of water. Product was extracted
three times
with 30 mL of ethylacetate, dried over sodium sulfate, solvent evaporated, and
the product
was subjected to chromatography on a column (7:3 hexanes: ethylacetate). A
colorless
glassy solid of pure compound 4 obtained in a 2.85 g, 74 % yield and 100%
purity (TLC)
and was further converted to compound 5.
[0173] Di-TFA salt of bis(L-leucine)estradiol-3,17-(3-diester (compound 5). De-
protection of Boc-protected monomer (compound 4) was carried out substantially
quantitatively inlO mL of dry dichloromethane, by adding 4 mL of dry TFA.
After 2 h of
stirring at room temperature, a homogenous solution was diluted with 300 mL of
anhydrous ether and left in a cold room over night. Precipitated white
crystals were
collected, washed twice with ether, and dried in a vacuum oven at 45 C. Yield
2.67 g
(90%). Mp = 187.5 C. 1H NMR (see Fig. 1)
[0174] Polymer Synthesis. Synthesis of therapeutic PEA was carried out in DMF
in
mild conditions (60 C): activated di-acid monomer (compound 1) was reacted
with
combinations of the di-amino monomers 1.5 eq. (compound 2), 1.5 eq. (compound
5) and
1 eq. of (compound 3).
[0175] Triethylamine 1.46 mL (10.47 mmol) was added at once to the mixture of
monomers (compound 1) (4.986 mmol), (compound 2) (1.246 mmol), (compound 3)
(1.869 imnol), (compound 5) (1.869 mmol) in 3 mL of dry DMF and the solution
was
heated to 60 C while stirring. The reaction vial was kept at the same
temperature for 16h.
A yellow viscous solution was formed then was cooled down to room temperature,
diluted
with 9 mL of dry DMF, added 0.2 mL of acetic anhydride, and after 3 h
precipitated out
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three times: first in water, then from ethanol solution into ethylacetate and
lastly, from
chloroform in etlzyl acetate. A colorless hydrophobic polymer was cast as a
tough film
from chloroform : ethanol (1:1) mixture and dried in vacuuin. Yield: 1.74g
(70%)
[0176] Materials Characterization The chemical structure of monomers and
polymer
were characterized by standard chemical methods. NMR spectra were recorded by
a
Brulcer AMX-500 spectrometer (Numega R. Labs Inc. San Diego, CA) operating at
500
MHz for 'H NMR spectroscopy. Deuterated solvents CDC13 or DMSO-d6 (Cambridge
Isotope Laboratories, Inc., Andover, MA) were used with tetramethylsilane
(TMS) as
internal standard. The results are shown in Figs. 1 and 3.
[0177] Melting points of synthesized monomers were determined on an automatic
Mettler-Toledo FP62 Melting Point Apparatus (Columbus, OH). Thermal properties
of
synthesized monomers and polymers were characterized on Mettler-Toledo DSC
822e
differential scanning calorimeter. Samples were placed in aluminuin pans.
Measurements
were carried out at a scaiming rate of 10 C/min under nitrogen flow (Fig. 2).
[0178] The number and weight average molecular weights (Mw and Mn) and
molecular weight distribution of synthesized polymer was determined by
Mode1515 gel
permeation chromatography (Waters Associates Inc. Milford, MA) equipped with a
high
pressure liquid chromatographic pump, a Waters 2414 refractory index detector.
0.1 !0 of
LiCI solution in N,N-dimethylacetamide (DMAc) was used as eluent (1.0 mL/min).
Two
Styragel HR 5E DMF type columns (Waters) were coimected and calibrated with
polystyrene standards.
[0179] Tensile Properties: tensile strength, elongation at break and Young's
Modulus
were measured on a tensile strength instrunlent (Chatillon TCD200, integrated
with a PC
(NexygenTM FM software)(Chatillon, Largo, FL) at a crosshead speed of 100
mm/min.
The load capacity was 50 lbs. The film (4 x 1.6 cm) had a dumbbell shape and
thickness
of about. 0.125 mm.
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[0180] Results Four different monomers were copolymerized by polycondensation
of
activated monomers, affording copoly PEA containing 17 % w/w steroid load on a
total
polymer weight basis. Chemical structure of the product therapeutic polymer
composition
is shown in scheme 8.
0
NHO
O c O O NH
1.5 0
O
11 NH O (CH2)$ NNH
O H O
L'O-(CH2)s-O NH 4 O
O CH2C6H5
1.5
Scheme 8
[0181] Three monomers: bis-p-toluenesulfonic acid salts of L-lysine-benzyl
ester
(compound 2), bis(L-leucine) 1,6-hexane diester (compound 3), and bis(p-
nitrophenyl)
sebacate (compound 1) were prepared according to the literature and
characterized by
melting point (Fig. 1) and proton N1VIR spectroscopy (Fig. 3). Results were in
agreement
with those reported in literature.
[0182] In this example a PEA polymer containing a residue of j3-Estradiol in
the main
polymer backbone was prepared, where both hydroxyls of the diol steroid were
incorporated into monomer via ester bonds using a carbodiimide technique. The
final
monomer introduced into the polymerization reaction was a TFA salt. After
polycondensation, a high molecular weight copolymer was obtained. Gel
permeation
chromatography yielded an estimated weight average Mw = 82,000 and
polydispersity
PDI = 1.54. The product copolymer was partially soluble in etllanol (when
dry), well
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soluble in chloroform, chloroform:ethanol 1:1 mixture, dichloromethane, and in
polar
aprotic organic solvents: DMF, DMSO, DMAc.
[0183] Glass transition temperature was detected at Tg = 41 (midpoint, taken
from the
second heating curve) and a sharp melting endotherm was detected at 220 C by
Differential scanning calorimetry (DSC) analysis (Fig. 2). This result leads
to the
conclusion that the polymer has semi-crystalline properties.
[0184] The therapeutic polyiner formed a tough film when cast from chloroform
solution. Tensile characterization yielded the following results: Stress at
break 28.1 MPa,
Elongation 173%, Young's Modulus 715 MPa.
EXAMPLE 2
[0185] Synthesis of a therapeutic PEUR polymer composition (structural formula
IV)
containing a therapeutic diol in the polymer backbone is illustrated in this
example. A
first monomer used in the synthesis is a di-carbon0ate of a theorapeutic diol
with a general
-0-R is fomled using a
chemical structure illustrated by formula R5-0-C-0-R6-0-C 5
known procedure (compound (X) as described in U.S. Patent 6,503,538) wherein
RS is
independently (C6-Clo) aryl (e.g. 4-nitrophenol, in this example), optionally
substituted
with one or more nitro, cyano, halo, trifluoromethyl or trifluoromethoxy; and
at least some
of p-nitrophenol. At least some of R6 is a residue of a therapeutic diol as
described herein,
depending upon the desired drug load. In the case where all of R6 is not the
residue of a
therapeutic diol, each diol would first be prepared and purified as a separate
monomer.
For example, di-p-nitrophenyl-3,17-(3-estradiol-dicarbonate (compound 6) can
be prepared
by the inetllod of Schenie 9 below:
OH
O ~
O
2 OZN O-~-CI + / I Base _ OpN ~~ 0-~- O~~ O~~NC
HO ~ (6)
Scheme 9
CA 02610745 2007-12-03
WO 2006/132950 PCT/US2006/021395
66
[0186] Polycondensation of compound X from U.S. Pat. 6,503,538 (in our example
compound 6) with the monomers described above yields an estradiol-based co-
poly(ester
urethane) PEUR (compouild 11):
HOTos.H2N NH2.TosOH TFA.H2N 0
O NH2.TFA
O O O O
2
(5)
O
O2N ~ \\ O-ll-O 0-11--0 O NO2
(6)
wherein the reaction scheme is as follows
TEA
3eq. (compound 5) + leq. (compound 2) + 4eq. (compound 6) -------~ (compound
11 )
DMF
o
C-O -C NH O O _ O
Q O H C O ~~ OC-NH~H
O 3 O
CH2CsH5
Compound (11)
[0187] All publications, patents, and patent documents are incorporated by
reference
herein, as though individually incorporated by reference. The invention has
been
described with reference to various specific and preferred embodiments and
techniques.
However, it should be understood that many variations and modifications might
be made
while remaining within the spirit and scope of the invention.
[0188] Although the invention has been described with reference to the above
examples, it will be understood that modifications and variations are
encompassed within
the spirit and scope of the invention. Accordingly, the invention is limited
only by the
following claims.
