Note: Descriptions are shown in the official language in which they were submitted.
CA 02434831 2010-06-03
TITLE OF INVENTION
POLYPEPTIDE DRUG SUBSTANCES.
REFERENCE TO RELATED APPLICATIONS
[0001] This application claims benefit under Title 35, U.S.C. 119(e), of
United States Applications Serial Nos. 60/262,337, filed 17 January 2001;
60/332,636, filed 6 November 2001; 60/287,872, filed 1 May 2001; and
601287,886, also filed 1 May 2001.
FIELD OF THE INVENTION
[0002] This invention, in general, relates to compositions and methods
that permit oral and parenteral administration, and significantly enhance the
bioavailability and pharmacological effects of therapeutically active
polypeptides,
pseudo-peptides and peptide mimics, particularly those that are otherwise
poorly
orally absorbable or display only minimal bioavailability if administered
parenterally.
Background of the Invention
[0003] It has been observed in the literature that therapeutically effective
polypeptides (aaõ) with two or more amino acids (n >_ 2) are poorly absorbed
orally. Even a polypeptide of as few as two amino acids, or related
structures,
exhibits very narrow absorption windows and poor bioavailability. As an
example, the Physician's Desk Reference (PDR) reports that the angiotensin
converting enzyme (ACE) inhibitor Enalaprilat (R1-Ala-Pro; n = 2) is very
poorly
absorbed orally. Enalapril (R2-Ala-Pro), which is a pro-drug of Enalaprilat,
is
better absorbed orally, but the end result demonstrates only a 25% relative
bioavailability of the active moiety (Enalaprilat) released from in vivo
cleavage of
the prodrug. In comparison, Lisinopril (R3-Lys-Pro) has relatively good
solubility
in water, but only a moderate oral bioavailability (< 25%), with a Tm." (time
to
maximum serum levels in vivo) of more than seven hours. Thus, this class of
CA 02434831 2010-06-03
2
therapeutic species is preferably administered via a non-oral deliver method,
such as by injection. However, even delivered intravenously, the
therapeutically
active species has a relatively short serum half-life.
[0004] It is also known that some tri-peptides originating in food products
may be capable of effective oral absorption, but to an unknown extent.
Furthermore, no active tri- or longer peptide drug substances (n >_ 3)
displaying
oral absorption have been identified.
10005] In a currently pending U.S. patent application Serial No.
091844,426,
the present inventors disclosed a method permitting the oral absorption
of polypeptide drug substances (aa") and other poorly orally absorbed drugs.
We
have subsequently made the surprising discovering that additional carrier
systems can be effective to achieve similar enhancement of pharmacological
activity for poorly absorbed active drug species, and that an optional, more
sophisticated linker offers additional improvement in results. Furthermore, we
have discovered that, through practice of the methods of the present
invention,
the length of the polypeptide drug entity (n) can be increased, particularly
when
the composition is administered parenterally, such as by intravenous (i.v.
administration, with the result of drastically improved pharmacological and
therapeutic effects for the active drug moiety. Accordingly, through the
practice
of the present invention, it is possible to chemically modify a polypeptide
species
.(or, additionally, pseudo-peptides or peptide mimics) of known therapeutic
utility
to both permit the oral administration of the species and to drastically
improve
its pharmacological properties even when administered through a parenteral
route.
[0006] In the present disclosure, the word "peptide" corresponds to any
sequence of naturally occurring amino acids, as well as to pseudo-peptides and
to peptide mimics. By "pseudo-peptide," we mean a chemical modification of
one or more of the amino acid residues constituting the peptide or of their
bonds
such as, but not limited to, use of amino acids in their D-configuration, use
of N-
methyl amino acids, replacement of one or more peptidic bonds (-CO-NH) by a
reduced bond (-CH2NH) and/or by -NHCO, -CH2CH2, -COCH2, -CHOHCH2, -CH2O.
By "peptide mimic," we mean any amino acid sequence in which the -C-
CA 02434831 2003-07-14
WO 02/056916 PCT/IB02/00133
3
backbone has been replaced by an oligourea backbone or an oligocarbamate
backbone. We also include co-peptides in this definition.
SUMMARY OF THE INVENTION
[00071 In a first embodiment, the present invention provides a
pharmaceutical agent comprising a carrier moiety and a therapeutically active
peptide species, wherein the peptide is in the form aa, where n is the number
of
amino acid residues in the peptide. Preferably, the carrier moiety comprises
an
aryl or alkyl group of sufficient length or steric bulk to protect the active
peptide
species from enzymatic degradation in vivo. More preferably, the carrier is
selected from a group comprising cinnamoyl, benzoyl, phenylacetyl, 3,4-
methylenedioxycinnamoyl, 3,4,5-trimethoxycinnamoyl, t-butoxycarbonyl,
benzyloxycarbonyl, pivaloyl, N-9-fluorenylmethoxycarbonyl, and fumaroyl.
Furthermore the carrier moiety can be chemically linked to a therapeutically
active peptide species of the general formula aa, where n is an integer from 2
to
40. In addition, this embodiment of the present invention contemplates a
therapeutically active peptide species that is poorly absorbed orally.
Preferably,
n is an integer from 3 to 6. More preferably, n is 5. More preferably still,
the
therapeutically active peptide species comprises Tyr-Gly-Gly-Phe-Met.
[00081 In an alternative embodiment, the pharmaceutical agent of the
present invention further comprises a linker species linking the peptide to
the
carrier moiety. Preferably, the linker species is selected from the group
consisting of a natural peptide, a pseudo-peptide, and a peptide mimic, each
member of the group comprising 4 or fewer amino acid residues. In one aspect
of this embodiment of the present invention, the linker species is directly
bound
to the carrier. Alternatively, the linker species is bound to the carrier
through a
-C6 or -C8 acidic moiety. More preferably, the linker species is Gly-carba-
Gly, a
pseudo-peptide. More preferably still, the linker species is associated with a
-Cõ
chain, where n is an integer from 6 to 8.
[00091 In another embodiment, the present invention provides a
pharmaceutical composition for administration to a patient in need thereof
comprising the pharmaceutical agent described immediately above, and one or
more pharmaceutically acceptable adjuvants. Preferably, the composition is
CA 02434831 2003-07-14
WO 02/056916 PCT/IB02/00133
14
formulated for oral administration. Alternatively, the composition is
formulated
for parenteral administration. Preferably, the composition is formulated for
intravenous administration. This embodiment of the present invention also
contemplates a composition that releases a biologically active form. of the
pharmaceutical agent into the patient's system at physiologically effective
levels
over a period of time of up to twelve hours. Preferably, the composition
releases a biologically active form of the pharmaceutical agent into the
patient's
system at physiologically effective levels over a period of time of up to
twenty-
four hours. In this embodiment of the present invention, the peptide species
is
preferably an epitope or an immune sequence characteristic of an infectious,
viral or cancerous disease.
[0010] In yet another embodiment, the present invention contemplates a
method for the treatment of a physiological condition through administration
of a
therapeutically effective species comprising the steps of chemically linking a
therapeutic polypeptide of the general formula aan, where as is an amino acid,
and where n is an integer from 2 to 40, to an alkyl or aryl carrier moiety to
form
a pro-drug, and administering the pro-drug to a patient exhibiting the
physiological condition. Preferably, the therapeutic polypeptide used in the
practice of the invention is poorly absorbed orally, and the carrier moiety is
selected from the group comprising cinnamoyl, benzoyl, phenylacetyl, 3,4-
methylenedioxycinnamoyl, 3,4,5-trimethoxycinnamoyl, t-butoxycarbonyl,
benzyloxycarbonyl, pivaloyl, N-9-fluorenylmethoxycarbonyl, and fumaroyl
[00111 Alternatively, this embodiment of the present invention provides a
method wherein the pro-drug is administered orally or parenterally. In yet
another alternative of the present embodiment, the method contemplates the
use of a therapeutic polypeptide that is chemically linked to the carrier
moiety
through a linker species.
[0012] In still another alternative embodiment, the present invention
provides a method to enhance the absorption and bioavailability of an active
polypeptide drug substance of the form aaõ in a pharmaceutical formulation,
the
method comprising the steps of adding a polypeptide moiety X,,, where n = 1 -
3, and where a terminal amino acid is selected from the group consisting of
Pro,
CA 02434831 2003-07-14
WO 02/056916 PCT/IB02/00133
Met and Arg, to one end of the polypeptide drug substance, and adding a
protecting moiety to the opposite end of the polypeptide drug substance.
[0013] Alternatively, the invention of the instant application provides a
method to enhance the absorption and bioavailability of an active polypeptide
drug substance of the form aaõ in a pharmaceutical formulation, the method
comprising the step of formulating the active polypeptide drug substance with
a
terminal amino acid selected from the group consisting of Pro, Met and Arg,
and
with a protective moiety on the opposite terminus of the polypeptide
substance,
wherein the terminal amino acid (Pro, Met or Arg) is not blocked by the
protective moiety.
DETAILED DESCRIPTION OF THE INVENTION
[0014] In one embodiment, the present invention provides a
pharmaceutical composition for use in the treatment of physiological
conditions
comprising a carrier moiety and a therapeutically active peptide species as
defined above. The carrier comprises an aryl or alkyl group of sufficient
length
and/or steric bulk to inhibit rapid enzymatic degradation of the active drug
species in vivo. A preferred carrier is selected from a group comprising
cinnamoyl, benzoyl, phenylacetyl, 3,4-methylenedioxycinnamoyl, 3,4,5-
trimethoxycinnamoyl, t-butoxycarbonyl, benzyloxycarbonyl, pivaloyl, N-9-
fluorenylmethoxycarbonyl, and Fumaroyl. The carrier moiety is chemically
linked
to a therapeutic polypeptide of the general formula aan, where as is an amino
acid, or a chemical or structural variation thereof as defined above, where n
is an
integer from 2 to 40, and wherein the polypeptide is poorly absorbed orally.
Preferably, in the drug composition of the invention, n is an integer from 3
to 6.
More preferably, n is 5. In a particularly preferred embodiment, the
polypeptide
is Tyr-Gly-Gly-Phe-Met.'
[0015] In an alternative variation, the pro-drug of the present invention
further comprises a linker species linking the peptide to the carrier species.
Preferably, the linker species is a natural peptide, a pseudo-peptide, a
peptide
mimic of less than 4 residues, either directly bound to the carrier or through
a
'Tyr = Tyrosine; Gly = Glycine; Phe = Phenylalanine; Met = Methionine.
CA 02434831 2003-07-14
WO 02/056916 PCT/IB02/00133
,6
-Cr:, or -C8 acidic moiety, or a composition thereof. A preferred linker is
the
pseudo-peptide Gly-carba-Gly associated, or not, to a -C6 or a -C8 chain.
Thus,
the present invention can be viewed as a three-component entity: The first,
therapeutically active component is the peptide; the second is the linker
species,
and the third is the carrier moiety.
[0016] When delivered orally, the drug composition of the present
invention is capable of delivery a systemic dose of the active drug species to
a
patient ingesting the pro-drug. The active peptide, normally immediately
degraded in the gastrointestinal tract to non-therapeutic forms, survives due
to
the protective effect of the carrier component, and persists in the patient's
system for prolonged periods of time. Over time, the multi-component system is
slowly broken down, probably by enzymatic hydrolysis in the liver or the
plasma,
releasing the pharmacologically active component. An added benefit of the
present invention is that the kinetics of such breakdown to release the active
component are significantly slower than for the processes associated with
metabolic breakdown of the unmodified polypeptide drug species, effectively
permitting a sustained, controlled release of the active species into the
patient's
system, thus maintaining pharmacologically effective blood serum levels over
an
extended period of time.
[0017] In another embodiment, the present invention contemplates a
pharmaceutical composition comprising a similar multi-component entity which,
when administered through a parenteral route, makes use of protective activity
towards the enzymatic breakdown provided by association of the active drug
species with the carrier and/or linking components, increasing thereby the in
vivo
half-life of the therapeutic component and improving its pharmacological
properties. A preferred therapeutic moiety for use in this embodiment of the
present invention is an epitope or an immune sequence characteristic of an
infectious, viral or cancerous disease. This invention, therefore, provides a
delivery method for such immune competent peptides that enhances their
pharmacological efficacy.
[0018] In yet another embodiment, the present invention contemplates a
pharmaceutical composition as defined above comprising a carrier moiety
CA 02434831 2003-07-14
WO 02/056916 PCT/IB02/00133
i7
comprising an aryl or alkyl group, optionally a linker species, and a
therapeutic
polypeptide of the general formula aa, where as is an amino acid, or a
chemical
or structural variation thereof, where n is an integer from 2 to 40, and a
pharmaceutically effective adjuvant species.
[0019] As would be recognized by one of skill in the appropriate art area,
one or more of the amino acids of the therapeutically active polypeptides used
in
conjunction with the present invention may be modified chemically or
conformationally without significantly diminishing, or preferably enhancing,
the
pharmacological activity of the therapeutic entity . These modified
polypeptides
may be used in the practice of the present invention.
[0020] Ideally, the pro-drug of the present invention is formulated into a
pharmaceutical composition with pharmaceutically acceptable adjuvants known
to those of skill in the art of pharmaceutical formulation chemistry.
[0021] Known therapeutically active polypeptide species that have been
demonstrated to be pharmacologically ineffective when delivered through
typical
oral routes of administration can be modified through linkage to a carrier
species
to achieve effective bioavailability of the active entity, as well as
therapeutically
effective controlled release of the active species.
[0022] In another embodiment, the invention of the instant application
encompasses a method for the treatment of a physiological condition through
the oral or parenteral administration of a therapeutically effective species
comprising the steps of chemically linking a therapeutic polypeptide of the
general formula aa, where as is an amino acid, or a chemical or structural
variation thereof, where n is an integer from 2 to 40, and wherein the
polypeptide is poorly absorbed orally, to an alkyl or aryl carrier moiety
preferably
selected from the group comprising cinnamoyl, benzoyl, phenylacetyl, 3,4-
m ethyl enedioxycinnamoyl, 3,4,5-trimethoxycinnamoyl, t-butoxycarbonyl,
benzyloxycarbonyl, pivaloyl, N-9-fluorenylmethoxycarbonyl and fumaroyl to form
a pro-drug, and orally or parenterally administering the pro-drug to a patient
exhibiting the physiological condition. Alternatively, in the practice of the
CA 02434831 2003-07-14
WO 02/056916 PCT/IB02/00133
8
method of the present invention, the polypeptide is chemically linked to the
carrier moiety through a linker species.
[0023] Thus, utilizing the present invention, it is possible to treat
physiological conditions through oral administration of therapeutically active
polypeptides that would normally have to be administered through considerably
less desirable routes of administration or with less effectiveness.
[0024] The present invention provides that the absorption and
bioavailability of an active polypeptide drug substance can be greatly
enhanced
by application of either one of the following two strategies: addition of a
polypeptide moiety X,, (n = 1 - 3) ending with an amino acid selected from the
group consisting of Pro, Met and Arg to one end of the active polypeptide drug
substance, along with the addition of a protecting moiety to the opposite end
of
the active polypeptide; or, alternatively, through formulation of the active
polypeptide drug substance, or pro-drug entity, with a terminal amino acid
selected from the group consisting of Pro, Met and Arg, with the protective
moiety on the opposite terminus of the polypeptide substance, provided that
the
terminal amino acid (Pro, Met or Arg) is not blocked by the protective moiety.
[0025] In still another embodiment, the invention of the instant application
provides a method for the controlled release administration of a
therapeutically
effective polypeptide of the general formula aa,,, where as is an amino acid,
or a
chemical or structural variation thereof, where n is an integer from 2 to 40,
and
wherein the polypeptide is poorly absorbed orally, comprising the steps of
chemically linking the polypeptide to an aryl or alkyl carrier moiety
preferably
selected from the group comprising cinnamoyl; benzoyl, phenylacetyl, 3,4-
methylenedioxycinnamoyl, 3,4,5-trimethoxycinnamoyl, t-butoxycarbonyl,
benzyloxycarbonyl, pivaloyl, N-9-fluorenylmethoxycarbonyl and fumaroyl to
form a pro-drug, and orally administering the pro-drug to a patient. In a
preferred embodiment, the polypeptide is chemically linked to the carrier
moiety
through a linker species, and, more preferably still, the linker species is an
amino acid, a pseudo-peptide or a peptide mimic optionally bound to the
carrier
through a -C6 or -C8 acidic residue. Due to the kinetics of the presumed
enzymatic degradation of the pro-drug of the present invention,, the
CA 02434831 2003-07-14
WO 02/056916 PCT/IB02/00133
9
therapeutically active polypeptide species is released to the patient's system
over relatively long periods of time, in a dosage-dependent manner, for up to
twenty-four hours.
EXAMPLES
[00261 Met-Enkephalin (Tyr-Gly-Gly-Phe-Met) is a naturally occurring
pentapeptide (n = 5) belonging to the endorphin class. It is known to be
involved in the basic mechanisms of analgesia. It produces a transient
analgesic
effect when administered parenterally, but no effect has been observed when
given orally. Its mechanism of action is believed to involve binding to opioid
delta receptors in the brain. Met-Enkephalin is very rapidly degraded in vivo
into
a tetra-peptide that is subsequently metabolized. As for the pharmacokinetics
of
Met-Enkephalin, the plasma levels of the pro-drug, as well of those of the
metabolites, are barely measurable, even when administered parenterally.
Example 1 Analgesic Effects from Administration of CY5M, a Cinnamoyl-Met-
Enkephalin Pro-Drug of the Present Invention.
[00271 According to the present invention, a pro-drug, designated CY5M
for convenience of reference, comprising cinnamoyl-Met-Enkephalin (cinnamoyl-
Tyr-Gly-Gly-Phe-Met), of the general form carrier-aa5, demonstrated an
unexpectedly strong, long-lasting analgesia in a hot plate test with rats both
when administered orally, and when administered parenterally.
Methods and Materials
100281 Analgesic activities are classically demonstrated in a hot plate test
using rats as test animals. The time to first licking of the posterior foot by
the
rat is recorded after the rat has been place on a hot plate maintained at an
elevated temperature (40 C). This procedure provides accurate data on central
analgesic activities induced by various candidate drugs. Under placebo
conditions, the time to first licking of the posterior foot of the test animal
varies
between 30 and 50 seconds. A strong analgesia is demonstrated when this
time is more than doubled. In the experiments reported herein, a standard hot
plate test was used to assess analgesia and the time to first licking of the
test
animal's posterior foot was used as the triggering event for measurement of
CA 02434831 2003-07-14
WO 02/056916 PCT/IB02/00133
X10
elapsed time as indicative of the pharmacological effect of the administered
drug
species.
[0029] Seven groups of five male Wistar rats each were randomly
assigned to the following treatments: placebo, 1 mg/kg morphine (i.v.), 10
mg/kg morphine (oral), 10 mg/kg codeine (oral), 10 mg/kg ibuprofen (oral), 2.5
mg/kg CY5M (i.v. ), and 2.5 mg/kg CY5M (oral). The method was pre-validated
with two oral and L v. administrations of saline placebo and the results were
similar to those obtained with placebo in the experiment reported below.
Results
Time Oh 1h 2h 4h 6h 24h
Placebo 53.2 30.6 38.4 45.0 46.6 42.0
Morphine 51.8 84.8 81.2 58.8 48.8 42.0
Codeine 53.2 51.4 64.6 57.6 56.2 46.4
Ibuprofen 53.2 55.0 70.4 66.0 54.0 44.2
CY5M 53.6 46.2 78.8 78.2 82.6 98.8
Table 1: Time to first signal activity after oral administration
Time Oh 1h 2h 4h 6h 24h
Placebo 53.2 30.6 38.4 45.0 46.6 42.0
morphine 51.8 118.8 86.6 63.2 45.6 40.0
CY5M 51.0 57.0 114.0 88.2 106.0 86.6
Table 2: Time to first signal activity after i.v, administration
[0030] In a preliminary study (data not shown), Met-Enkephalin alone was
unable to demonstrate any effect after oral administration at a 5 mg/kg dose,
whereas a transient effect of about 15 minutes was observed after L v.
administration.
[00311 If one considers the area under the dose response curve a rough
estimate of the average effect, the results indicate that 1 mg/kg morphine
i.v. is
comparable to 10 mg/kg morphine oral. In comparison, CY5M, administered
CA 02434831 2003-07-14
WO 02/056916 PCT/IB02/00133
11
either orally or by i.v., is at least 8 times more effective than morphine by
the
same route of administration. Of further interest, the above data also
indicate
that in no case did morphine exhibit an analgesic effect lasting longer than
six
hours, whereas both oral and i.v. administrations of CY5M demonstrated a
significant analgesic effect for a period of time of 24 hours or longer.
[0032] It is also anticipated that an analog of CY5M comprising a linker
species in addition to the cinnamoyl carrier species, will demonstrate similar
or
greater effects than those provided above.
[0033] These results indicate that using a carrier such as disclosed herein
in association with a polypeptide drug species, permits the effective oral
absorption of peptides of at least 5 amino acids in length and allows a much
stronger pharmacological effect, with significantly enhanced pharmacokinetic
profiles, by both oral and i.v. routes of administration.
CA 02434831 2010-06-03
SEQUENCE LISTING
<110> Zimmer, Robert A.
<120> Compositions and methods for Enhanced Pharmacological Activity through
oral
and Parenteral Administration of Compositions Comprising Polypeptide Drug
substances and other Poorly Absorbed Active Ingredients
<130> 945505.019 PCT
<140> PCT /IB 02/00133
<141> 2002-01-16
<150> US 60/262,337
<151> 2001-01-17
<150> us 60/332,636
<151> 2001-11-06
<150> us 60/287,872
<151> 2001-05-01
<150> Us 60/287,886
<151> 2001-05-01
<160> 1
<170> Patentln version 3.1
<210> 1
<211> 5
<212> PRT
<213> Homo sapiens
<400> 1
Tyr Gly Gly Phe met
1 5
Page 12
