U.S. patent application number 15/245508 was filed with the patent office on 2016-12-15 for conjugates for treating diseases caused by psma expressing cells.
The applicant listed for this patent is ENDOCYTE, INC.. Invention is credited to Alicia BLOOMFIELD, Ryan DORTON, Christopher Paul LEAMON, Melissa NELSON, Joseph Anand REDDY, Marilynn VETZEL, Iontcho Radoslavov VLAHOV.
Application Number | 20160361432 15/245508 |
Document ID | / |
Family ID | 50731670 |
Filed Date | 2016-12-15 |
United States Patent
Application |
20160361432 |
Kind Code |
A1 |
VLAHOV; Iontcho Radoslavov ;
et al. |
December 15, 2016 |
CONJUGATES FOR TREATING DISEASES CAUSED BY PSMA EXPRESSING
CELLS
Abstract
The invention described herein pertains to the diagnosis,
imaging, and/or treatment of pathogenic cell populations. In
particular, the invention described herein pertains to the
diagnosis, imaging, and/or treatment of diseases caused by PSMA
expressing cells, such as prostate cancer cells, using compounds
capable of targeting PSMA expressing cells.
Inventors: |
VLAHOV; Iontcho Radoslavov;
(West Lafayette, IN) ; REDDY; Joseph Anand; (West
Lafayette, IN) ; BLOOMFIELD; Alicia; (Lafayette,
IN) ; DORTON; Ryan; (Lafayette, IN) ; NELSON;
Melissa; (Delphi, IN) ; VETZEL; Marilynn;
(Rossville, IN) ; LEAMON; Christopher Paul; (West
Lafayette, IN) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
ENDOCYTE, INC. |
West Lafayette |
|
IN |
|
|
Family ID: |
50731670 |
Appl. No.: |
15/245508 |
Filed: |
August 24, 2016 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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14443212 |
May 15, 2015 |
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PCT/US13/70007 |
Nov 14, 2013 |
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15245508 |
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61875971 |
Sep 10, 2013 |
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61788382 |
Mar 15, 2013 |
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61726991 |
Nov 15, 2012 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C07K 5/1019 20130101;
A61P 35/00 20180101; A61K 38/07 20130101; A61K 47/542 20170801;
A61K 51/04 20130101; C07K 5/10 20130101; A61K 38/06 20130101; A61K
38/05 20130101; A61K 47/64 20170801; C07K 5/06113 20130101; A61P
13/08 20180101; A61K 38/00 20130101; A61K 47/547 20170801; A61K
49/0002 20130101; A61K 38/08 20130101; A61P 13/10 20180101; C07K
5/021 20130101 |
International
Class: |
A61K 47/48 20060101
A61K047/48 |
Claims
1-29. (canceled)
30. A conjugate of formula B-L-(D).sub.n or a pharmaceutically
acceptable salt thereof; wherein B comprises a urea or thiourea of
lysine and an amino acid selected from the group consisting of
lysine, aspartic acid, glutamic acid, and homoglutamic acid,
wherein the urea or thiourea is capable of binding to PSMA; wherein
L is a polyvalent linker; wherein D is a radical of a drug; and
wherein n is 1; with a proviso that the conjugate is not
##STR00119##
31. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein L is a polyvalent linker comprising an
aminomethylphenylacetic acid diradical or an aminophenylacetic acid
diradical.
32. A conjugate of formula B-L-(D).sub.n or a pharmaceutically
acceptable salt thereof; wherein B is a radical of a PSMA binding
ligand wherein L is a polyvalent linker comprising at least one of
an aminomethylphenylacetic acid diradical or an aminophenylacetic
acid diradical; wherein D is a radical of a drug; and wherein n is
1; with a proviso that the conjugate is not ##STR00120##
33. The conjugate of claim 32, or a pharmaceutically acceptable
salt thereof, wherein B comprises a urea or thiourea of lysine and
an amino acid selected from the group consisting of lysine,
aspartic acid, glutamic acid, and homoglutamic acid.
34. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein B is selected from the group consisting of
##STR00121##
35. The conjugate of claim 34, or a pharmaceutically acceptable
salt thereof, wherein B is ##STR00122##
36. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein at least one drug is a tubulysin.
37. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein L forms a urea or thiourea with the
lysine.
38. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein L comprises a cysteine diradical.
39. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein L is a releasable linker.
40. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein L comprises a disulfide.
41. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein L comprises a diradical of the formula
##STR00123##
42. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein B-L comprises a diradical of the formula
##STR00124##
43. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein B-L comprises a diradical of the formula
##STR00125##
44. The conjugate of claim 30, or a pharmaceutically acceptable
salt thereof, wherein B-L comprises a diradical of the formula
##STR00126##
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority under 35 U.S.C.
.sctn.119(e) to U.S. Provisional Application Ser. No. 61/726,991,
filed Nov. 15, 2012, U.S. Provisional Application Ser. No.
61/788,382, filed Mar. 15, 2013, and U.S. Provisional Application
Ser. No. 61/875,971, filed Sep. 10, 2013, in which all of which are
incorporated herein by reference in their entirety.
TECHNICAL FIELD
[0002] The invention described herein pertains to the diagnosis,
imaging, and/or treatment of pathogenic cell populations. In
particular, the invention described herein pertains to the
diagnosis, imaging, and/or treatment of diseases caused by PSMA
expressing cells, such as prostate cancer cells, using compounds
capable of targeting PSMA expressing cells.
BACKGROUND AND SUMMARY OF THE INVENTION
[0003] The prostate is a male reproductive organ and functions to
produce and store seminal fluid that provides nutrients and fluids
for the survival of sperm introduced into the vagina during
reproduction. Like other tissues, the prostate gland may develop
either malignant (cancerous) or benign (non-cancerous) tumors. In
fact, prostate cancer is one of the most common male cancers in
western societies, and is the second leading form of malignancy
among American men. Current treatment methods for prostate cancer
include hormonal therapy, radiation therapy, surgery, chemotherapy,
photodynamic therapy, and combination therapy. However, many of
these treatments affect the quality of life of the patient,
especially for those men who are diagnosed with prostate cancer
over age 50. For example, the use of hormonal drugs is often
accompanied by side effects such as osteoporosis and liver damage.
Such side effects might be mitigated by the use of treatments that
are more selective or specific to the tissue being responsible for
the disease state, and avoid non-target tissues like the bones or
the liver.
[0004] Prostate-specific membrane antigen (PSMA) is a biomarker
that is overexpressed on prostate cancer. PSMA is over-expressed in
the malignant prostate tissues when compared to other organs in the
human body such as kidney, proximal small intestine, and salivary
glands. PSMA is also expressed on the neovasculature within many
non-prostate solid tumors, including lung, colon, breast, renal,
liver and pancreatic carcinomas, but not on normal vasculature.
PSMA is also expressed minimally in brain. PSMA is a type II cell
surface membrane-bound glycoprotein with .about.110 kD molecular
weight, including an intracellular segment (amino acids 1-18), a
transmembrane domain (amino acids 19-43), and an extensive
extracellular domain (amino acids 44-750). While the functions of
the intracellular segment and the transmembrane domains are
currently believed to be insignificant, the extracellular domain is
involved in several distinct activities. For example, PSMA plays a
role in the central nervous system, where it metabolizes
N-acetyl-aspartyl glutamate (NAAG) into glutamic and N-acetyl
aspartic acid. PSMA also plays a role in the proximal small
intestine where it removes .gamma.-linked glutamate from
poly-.gamma.-glutamated folate and .alpha.-linked glutamate from
peptides and small molecules. However, PSMA's particular function
on prostate cancer cells remains unresolved.
[0005] Unlike many other membrane-bound proteins, PSMA undergoes
rapid internalization into the cell in a similar fashion to cell
surface bound receptors like vitamin receptors. PSMA is
internalized through clathrin-coated pits and subsequently can
either recycle to the cell surface or go to lysosomes. Accordingly,
diagnostic, imaging, and therapeutic agents can be targeted to PSMA
for delivery into PSMA expressing cells, such as prostate cancer
cells.
[0006] Described herein are compounds capable of binding to PSMA.
Also described herein are compounds capable of targeting PSMA for
delivery of diagnostic, imaging, and therapeutic agents. Also
described herein are compounds and compositions, and methods and
uses thereof for diagnosing, imaging, and treating diseases caused
by pathogenic populations of cells that express, or overexpress,
PSMA.
[0007] It has been unexpectedly discovered that the conjugates
described herein exhibit high affinity for PSMA. It has also been
discovered that the compounds described herein are efficacious in
treating diseases caused by pathogenic cells that express PSMA,
such a prostate cancer cells.
[0008] In one illustrative embodiment of the invention, PSMA
binding drug delivery conjugates of the formula
B-L-(D).sub.n
or pharmaceutically acceptable salts thereof are described herein,
where B comprises a urea or thiourea of lysine and an amino acid,
or one or more carboxylic acid derivatives thereof, where the urea
or thiourea is capable of binding to PSMA, L is a polyvalent
linker, D is a radical of a drug, and n is an integer selected from
1, 2, 3, and 4. It is to be understood that as used herein, such
drugs, and the term drug, includes therapeutic agents, diagnostic
agents, imaging agents, and other compounds that are desirably
delivered to or targeted to PSMA and/or PSMA expressing cells.
[0009] In another illustrative embodiment, PSMA binding drug
delivery conjugates of the formula
B-L-(D).sub.n
or pharmaceutically acceptable salts thereof are described herein,
where B is a radical of a PSMA binding or targeting ligand, L is a
polyvalent linker comprising an aminomethylphenylacetic acid
diradical, or an amninophenylacetic acid diradical, or both, D is a
radical of a drug, and n is an integer selected from 1, 2, 3, and
4.
[0010] It is to be understood that every combination of the various
embodiments of each of B, L, D, and n described herein form
illustrative embodiments of the conjugates of the invention,
whether those various embodiments of each of B, L, D are species,
subgenera, or genera. It is to be further understood that each of
those additional illustrative embodiments of compounds may be used
in any of the compositions, unit doses, methods, and/or uses
described herein.
[0011] In another embodiment, pharmaceutical compositions
containing one or more of the compounds are also described herein.
In one aspect, the compositions are in bulk form and are suitable
for preparing unit doses, unit dosage forms, and the like that may
be included in the uses and/or methods described herein. In another
aspect, the compositions include a therapeutically effective amount
of the one or more compounds for diagnosis, imaging, and/or
treatment of diseases caused by PSMA expressing cells in a patient.
Illustrative compositions include unit doses, unit dosage forms,
and the like. It is to be understood that the compositions may
include other components and/or ingredients, including, but not
limited to, other therapeutically active compounds, and/or one or
more carriers, and/or one or more diluents, and/or one or more
excipients, and the like. In another embodiment, methods for using
the compounds and pharmaceutical compositions for diagnosis,
imaging, and/or treatment of diseases caused by PSMA expressing
cells in a patient are also described herein. In one aspect, the
methods include the step of administering one or more of the
compounds and/or compositions described herein to the patient. In
another embodiment, uses of the compounds and compositions in the
manufacture of a medicament for diagnosis, imaging, and/or
treatment of diseases caused by PSMA expressing cells in a patient
are also described herein. In one aspect, the medicaments include a
therapeutically effective amount of the one or more compounds
and/or compositions described herein.
[0012] It is appreciated herein that the compounds described herein
may be used alone or in combination with other compounds useful for
diagnosis, imaging, and/or treatment of diseases caused by PSMA
expressing cells in a patient, including those compounds that may
be therapeutically effective by the same or different modes of
action. In addition, it is appreciated herein that the compounds
described herein may be used in combination with other compounds
that are administered to treat other symptoms of the disease, such
as compounds administered to decrease pain, and the like.
BRIEF DESCRIPTION OF THE DRAWINGS
[0013] FIG. 1 shows the relative affinity of (.box-solid.) PMPA,
1.0 (normalized); ( ) DUPA, 0.05 (19-fold lower); (.smallcircle.)
EC1067, 30.times.; (.quadrature.) EC1069, 22.times.; and () EC1080,
6.times. in 10% serum/FDRPMI for PSMA.
[0014] FIG. 2 shows the relative affinity of (.box-solid.) PMPA,
1.0 (normalized); ( ) EC1100, 20.times.; () EC1168, 17.times.;
(.tangle-solidup.) EC1169, 7.times.; and (.quadrature.) EC1170,
7.times. in 10% serum/FDRPMI for PSMA.
[0015] FIG. 3 shows the dose response and IC50 for EC1169 against
LNCaP cells (2 h-72 h) as determined by .sup.3H-thymidine
incorporation cells in vitro.
[0016] FIG. 4 shows the dose response and IC50 for () EC1718,
(.diamond-solid.) EC11677, (.tangle-solidup.) EC1719, ( ) EC1720,
and (.box-solid.) EC1721 against LNCaP cells (2 h-72 h) as
determined by .sup.3H-thymidine incorporation cells in vitro.
[0017] FIG. 5 shows the in vivo efficacy of EC1169 (c), EC1550 ( ),
and EC1551 (.box-solid.), each at 2 .mu.mol/kg, TIW (three times
per week), 2 weeks, compared against vehicle-treated controls
(.diamond-solid.) in treating LNCaP tumor xenographs.
[0018] FIG. 6 shows that EC1169 (c), EC1550 ( ), and EC1551
(.box-solid.), each at 2 .mu.mol/kg, TIW, 2 weeks, compared against
vehicle-treated controls (.diamond-solid.) do not exhibit gross
animal toxicity.
[0019] FIG. 7 shows the in vivo efficacy of EC1584 () and EC1588
(.tangle-solidup.) each at 2 .mu.mol/kg, TIW, 2 weeks, compared
against vehicle-treated controls ( ) in treating LNCaP tumor
xenographs.
[0020] FIG. 8 shows that EC1584 () and EC1588 (.tangle-solidup.),
each at 2 .mu.mol/kg, TIW, 2 weeks, compared against
vehicle-treated controls ( ) do not exhibit gross animal
toxicity.
[0021] FIG. 9 shows the in vivo efficacy of EC1169 ( ) at 2
.mu.mol/kg, TIW, 2 weeks, compared to docetaxel, at 10 mg/kg, BIW,
2 weeks, MTD (), and each compared to vehicle-treated control
(.box-solid.) in treating LNCaP tumor xenographs.
[0022] FIG. 10 shows that of EC1169 ( ) administered at 2
.mu.mol/kg, TIW, 2 weeks, exhibits substantially less gross animal
toxicity compared to docetaxel, administered at 10 mg/kg, BIW, 2
weeks, MTD ().
[0023] FIG. 11 shows the in vivo efficacy of (.box-solid.) EC1718;
(.tangle-solidup.) EC1720; () EC1721; (.diamond-solid.) EC1719; and
(.smallcircle.) EC11677, each administered at 2 .mu.mol/kg, TIW, 2
weeks; compared to ( ) vehicle-treated control in treating LNCaP
tumor xenographs.
[0024] FIG. 12 shows that (.box-solid.) EC1718; (.tangle-solidup.)
EC1720; () EC1721; (.diamond-solid.) EC1719; and (.smallcircle.)
EC11677; compared to ( ) vehicle-treated control, do not exhibit
gross animal toxicity.
DETAILED DESCRIPTION
[0025] Several illustrative embodiments of the invention are
described by the following enumerated clauses:
[0026] 1. A conjugate of the formula
B-L-(D).sub.n
or a pharmaceutically acceptable salt thereof, wherein B comprises
a urea or thiourea of lysine and an amino acid, or one or more
carboxylic acid derivatives thereof, including, but not limited to
ureas or thioureas of lysine and aspartic acid, or glutamic acid,
or homoglutamic acid, where the urea or thiourea is capable of
binding to PSMA, L is a polyvalent linker, D is a radical of a
drug, and n is an integer selected from 1, 2, 3, and 4.
[0027] 2. A conjugate of the formula
B-L-(D).sub.n
or a pharmaceutically acceptable salt thereof, wherein B is a
radical of the formula
##STR00001##
L is a polyvalent linker, D is a radical of a drug, and n is an
integer selected from 1, 2, 3, and 4.
[0028] 3. The conjugate of clause 1 or 2 wherein L is a polyvalent
linker comprising an aminomethylphenylacetic acid diradical, or an
aminophenylacetic acid diradical, or both.
[0029] 4. A conjugate of the formula
B-L-(D).sub.n
or a pharmaceutically acceptable salt thereof, wherein B is a
radical of a PSMA binding ligand, L is a polyvalent linker
comprising an aminomethylphenylacetic acid diradical or an
aminophenylacetic acid diradical or both, D is a radical of a drug,
and n is an integer selected from 1, 2, 3, and 4.
[0030] 5. The conjugate of clause 3 wherein B comprises a urea or
thiourea of lysine and an amino acid, or one or more carboxylic
acid derivatives thereof, including, but not limited to ureas or
thioureas of lysine and aspartic acid, or glutamic acid, or
homoglutamic acid.
[0031] 6. The conjugate of any one of clauses 1 to 5 wherein B
comprises a urea or thiourea of lysine and glutamate, or one or
more carboxylic acid derivatives thereof.
[0032] 7. The conjugate of any one of clauses 1 to 5 wherein B
comprises a urea of lysine and glutamate.
[0033] 8. The conjugate of any one of clauses 1 to 5 wherein B
comprises a urea or thiourea of L-lysine and L-glutamate, or one or
more carboxylic acid derivatives thereof.
[0034] 9. The conjugate of any one of clauses 1 to 5 wherein B
comprises a urea of L-lysine and L-glutamate.
[0035] 10. The conjugate of any one of clauses 1 to 5 wherein B
comprises a urea or thiourea of lysine and glutamic acid.
[0036] 11. The conjugate of any one of clauses 1 to 5 wherein B
comprises a urea or thiourea of D-lysine and D-glutamic acid.
[0037] 12. The conjugate of any one of clauses 1 to 5 wherein B
comprises a urea or thiourea of D-lysine and one or the
following:
##STR00002##
[0038] 13. The conjugate of any one of clauses 1 to 5 wherein B
comprises a urea or thiourea of D-lysine and:
##STR00003##
[0039] 14. The conjugate of any one of clauses 1 to 5 wherein B is
a urea.
[0040] 15. The conjugate of any one of clauses 1 to 5 wherein B is
selected from the following
##STR00004##
[0041] 16. The conjugate of any one of clauses 1 to 5 wherein B is
selected from the following
##STR00005##
[0042] 17. The conjugate of any one of clauses 1 to 5 wherein B is
of the formula
##STR00006## [0043] The conjugate of any one of the preceding
clauses wherein n is 1, 2, or 3. [0044] The conjugate of any one of
the preceding clauses wherein n is 1 or 2. [0045] The conjugate of
any one of the preceding clauses wherein n is 1. [0046] The
conjugate of any one of the preceding clauses wherein at least one
drug is an imaging agent. [0047] The conjugate of any one of the
preceding clauses wherein at least one drug is a diagnostic agent.
[0048] The conjugate of any one of the preceding clauses wherein at
least one drug is a therapeutic agent. [0049] The conjugate of any
one of the preceding clauses wherein at least one drug is a
cytotoxic agent. [0050] The conjugate of any one of the preceding
clauses wherein at least one drug is a tubulysin. [0051] The
conjugate of any one of the preceding clauses wherein at least one
drug is a naturally occurring tubulysin. [0052] The conjugate of
any one of the preceding clauses wherein at least one drug is
tubulysin B. [0053] The conjugate of any one of the preceding
clauses wherein at least one drug is a tubulysin of the formula
##STR00007##
[0053] and pharmaceutical salts thereof are described, where
[0054] n is 1-3:
[0055] V is hydrogen, OR.sup.2, or halo, and W is hydrogen,
OR.sup.2, or alkyl, where R.sup.2 is independently selected in each
instance from hydrogen, alkyl, and C(O)R.sup.3, where R.sup.3 is
alkyl, cycloalkyl, alkenyl, aryl, or arylalkyl, each of which is
optionally substituted; providing that R.sup.2 is not H when both V
and W are OR.sup.2; or V and W are taken together with the attached
carbon to form a carbonyl;
[0056] X is hydrogen, alkyl, such as C.sub.1-6 alkyl, or C.sub.2-6
alkyl, C.sub.1-4 alkyl, or C.sub.2-4 alkyl, or alkenyl, such as
C.sub.2-6 alkenyl or C.sub.2-4 alkenyl, each of which is optionally
substituted;
[0057] Z is alkyl or C(O)R.sup.4, where R.sup.4 is alkyl, CF.sub.3,
or aryl;
[0058] Ar is aryl or heteroaryl, each of which is optionally
substituted; and
[0059] R is OH or R and the carbonyl to which it is attached is a
carboxylic acid derivative. [0060] The conjugate of any one of the
preceding clauses wherein Ar is optionally substituted phenyl.
[0061] The conjugate of any one of the preceding clauses wherein Ar
is phenyl substituted with one or more substituents selected from
the group consisting of halo, hydroxy, amino, thio, carboxylate or
a derivative thereof, sulfinyl or a derivative thereof, sulfonyl or
a derivative thereof, phosphinyl or a derivative thereof, or
phosphonyl or a derivative thereof, or alkyl, alkenyl, alkynyl,
cycloalkyl, cycloalkenyl, heteroalkyl, heteroalkenyl,
cycloheteroalkyl, cycloheteroalkenyl, aryl, heteroaryl, arylalkyl,
and heteroarylalkyl, each of which is optionally substituted.
[0062] The conjugate of any one of the preceding clauses wherein Ar
is phenyl. [0063] The conjugate of any one of the preceding clauses
wherein Ar is 4-hydroxyphenyl. [0064] The conjugate of any one of
the preceding clauses wherein X is CH.sub.2QR.sup.9, where Q is
--N--, --O--, or --S--; R.sup.9 is hydrogen or alkyl, alkenyl,
cycloalkyl, aryl, or arylalkyl, each of which is optionally
substituted, or C(O)R.sup.10. [0065] The conjugate of any one of
the preceding clauses wherein Q is O. [0066] The conjugate of any
one of the preceding clauses wherein R.sup.9 is optionally
substituted alkyl. [0067] The conjugate of any one of the preceding
clauses wherein R.sup.9 is alkyl. [0068] The conjugate of any one
of the preceding clauses wherein R.sup.10 is optionally substituted
alkyl. [0069] The conjugate of any one of the preceding clauses
wherein R.sup.10 is alkyl. [0070] The conjugate of any one of the
preceding clauses wherein at least one drug is selected from the
following:
[0070] ##STR00008## [0071] The conjugate of any one of the
preceding clauses wherein at least one drug is:
[0071] ##STR00009## [0072] The conjugate of any one of the
preceding clauses wherein at least one D is a radical of the
formula
[0072] ##STR00010## [0073] The conjugate of any one of the
preceding clauses wherein at least one D is a radical of the
formula
[0073] ##STR00011## [0074] The conjugate of any one of the
preceding clauses wherein at least one D is a radical of the
formula
[0074] ##STR00012## [0075] The conjugate of any one of the
preceding clauses wherein at least one D is a radical of the
formula
[0075] ##STR00013## [0076] The conjugate of any one of the
preceding clauses wherein at least one D is a radical of the
formula
[0076] ##STR00014## [0077] The conjugate of any one of the
preceding clauses wherein L comprises an aminomethylphenylacetic
acid diradical. [0078] The conjugate of any one of the preceding
clauses wherein L comprises an aminophenylacetic acid diradical
[0079] The conjugate of any one of the preceding clauses wherein L
forms a urea or thiourea with the lysine. [0080] The conjugate of
any one of the preceding clauses wherein L forms a urea with the
lysine. [0081] The conjugate of any one of the preceding clauses
wherein L forms an amide or thioamide with the lysine. [0082] The
conjugate of any one of the preceding clauses wherein L forms an
amide with the lysine. [0083] The conjugate of any one of the
preceding clauses wherein L comprises one or more aspartic acid
diradicals. [0084] The conjugate of any one of the preceding
clauses wherein L comprises two or more aspartic acid diradicals.
[0085] The conjugate of the preceding clauses wherein the aspartic
acid diradicals are L-aspartic acid diradicals. [0086] The
conjugate of any one of the preceding clauses wherein L comprises a
cysteine diradical. [0087] The conjugate of any one of the
preceding clauses wherein L comprises a L-cysteine diradical.
[0088] The conjugate of any one of the preceding clauses wherein L
comprises L-Asp-L-Asp-L-Cys. [0089] The conjugate of any one of the
preceding clauses wherein L is a releasable linker, such as a
releasable linker that is cleaved under conditions encountered at
or near, or inside of pathogenic cells expressing, preferentially
expressing, or overexpressing PSMA. [0090] The conjugate of any one
of the preceding clauses wherein L comprises a disulfide. [0091]
The conjugate of any one of the preceding clauses wherein L
comprises a cysteine disulfide diradical. [0092] The conjugate of
any one of the preceding clauses wherein L comprises a L-cysteine
disulfide diradical. [0093] The conjugate of any one of the
preceding clauses wherein L comprises L-Asp-L-Asp-L-Cys(S--S).
[0094] The conjugate of any one of the preceding clauses wherein L
comprises a diradical of the formula O--C(O)--N. [0095] The
conjugate of any one of the preceding clauses wherein L comprises a
diradical of the formula O--C(O)--NH. [0096] The conjugate of any
one of the preceding clauses wherein L and at least one D taken
together comprise a diradical of the formula O--C(O)--N. [0097] The
conjugate of any one of the preceding clauses wherein L and at
least one D taken together comprise a diradical of the formula
O--C(O)--NH. [0098] The conjugate of any one of the preceding
clauses wherein L comprises a diradical of the formula
S--(CH.sub.2).sub.m--O, where m is 2, 3, or 4. [0099] The conjugate
of any one of the preceding clauses wherein L comprises a diradical
of the formula S--(CH.sub.2).sub.m--O--C(O)--N, where m is 2, 3, or
4. [0100] The conjugate of any one of the preceding clauses wherein
L comprises a diradical of the formula
S--(CH.sub.2).sub.m--O--C(O)--NH, where m is 2, 3, or 4. [0101] The
conjugate of any one of the preceding clauses wherein L and at
least one D taken together comprise a diradical of the formula
S--(CH.sub.2).sub.m--O--C(O)--N, where m is 2, 3, or 4. [0102] The
conjugate of any one of the preceding clauses wherein L and at
least one D taken together comprise a diradical of the formula
S--(CH.sub.2).sub.m--O--C(O)--NH, where m is 2, 3, or 4. [0103] The
conjugate of any one of the preceding clauses wherein the terminal
sulfur atom forms a disulfide. [0104] The conjugate of any one of
the preceding clauses wherein m is 2. [0105] The conjugate of any
one of the preceding clauses wherein L comprises a chain of at
least about 7 atoms, at least about 8 atoms, at least about 9
atoms, at least about 10 atoms, at least about 11 atoms, at least
about 12 atoms, at least about 13 atoms, at least about 14 atoms,
or at least about 15 atoms. [0106] The conjugate of any one of the
preceding clauses wherein L comprises a chain of at least about 16
atoms, at least about 17 atoms, at least about 18 atoms, at least
about 19 atoms, at least about 20 atoms, at least about 21 atoms,
at least about 22 atoms, at least about 23 atoms, at least about 24
atoms, at least about 25 atoms, or at least about 26 atoms. [0107]
The conjugate of any one of the preceding clauses wherein L
comprises a chain of between about 7 and about 35 atoms, between
about 7 and about 30 atoms, or between about 7 and about 26 atoms.
[0108] The conjugate of any one of the preceding clauses wherein L
comprises a diradical of the formula
[0108] ##STR00015## [0109] The conjugate of any one of the
preceding clauses wherein L comprises a diradical of the
formula
[0109] ##STR00016## [0110] The conjugate of any one of the
preceding clauses wherein L comprises a diradical of the
formula
[0110] ##STR00017## [0111] The conjugate of any one of the
preceding clauses wherein L comprises a diradical of the
formula
[0111] ##STR00018## [0112] The conjugate of any one of the
preceding clauses where L comprises a diradical of the formula
[0112] ##STR00019## [0113] The conjugate of any one of the
preceding clauses where L comprises a diradical of the formula
[0113] ##STR00020## [0114] The conjugate of any one of the
preceding clauses wherein L comprises a diradical of the
formula
[0114] ##STR00021## [0115] The conjugate of any one of the
preceding clauses wherein L comprises a diradical of the
formula
[0115] ##STR00022## [0116] The conjugate of any one of the
preceding clauses wherein L comprises a diradical of the
formula
[0116] ##STR00023## [0117] The conjugate of any one of the
preceding clauses where L comprises a diradical of the formula
[0117] ##STR00024## [0118] The conjugate of any one of the
preceding clauses where L comprises a diradical of the formula
[0118] ##STR00025## [0119] The conjugate of any one of the
preceding clauses where L comprises a diradical of the formula
[0119] ##STR00026## [0120] The conjugate of any one of the
preceding clauses wherein L comprises a diradical of the
formula
[0120] ##STR00027## [0121] The conjugate of any one of the
preceding clauses wherein L comprises a diradical of the
formula
[0121] ##STR00028## [0122] The conjugate of any one of the
preceding clauses wherein B-L comprises a diradical of the
formula
[0122] ##STR00029## [0123] The conjugate of any one of the
preceding clauses wherein B-L comprises a diradical of the
formula
[0123] ##STR00030## [0124] The conjugate of any one of the
preceding clauses wherein B-L comprises a diradical of the
formula
[0124] ##STR00031## [0125] The conjugate of any one of the
preceding clauses wherein B-L comprises a diradical of the
formula
[0125] ##STR00032## [0126] A conjugate of the formula
##STR00033##
[0126] or a pharmaceutically acceptable salt thereof, and/or a
hydrate, and/or a solvate, and/or a co-crystal of the foregoing;
where D is radical of a drug. [0127] A conjugate of the formula
##STR00034##
[0127] or a pharmaceutically acceptable salt thereof, and/or a
hydrate, and/or a solvate, and/or a co-crystal of the foregoing;
where D is radical of a drug. [0128] A conjugate the formula
##STR00035##
[0128] or a pharmaceutically acceptable salt thereof, and/or a
hydrate, and/or a solvate, and/or a co-crystal of the foregoing;
where D is radical of a drug. [0129] A conjugate of the formula
##STR00036##
[0129] or a pharmaceutically acceptable salt thereof, and/or a
hydrate, and/or a solvate, and/or a co-crystal of the foregoing;
where D is radical of a drug. [0130] A pharmaceutical composition
comprising one or more of the compounds or conjugates of any one of
the preceding clauses. [0131] A pharmaceutical composition
comprising one or more of the compounds or conjugates of any one of
the preceding clauses for treating a disease in a host animal
caused by a pathogenic population of cells, said cells expressing
PSMA. [0132] A unit dose or unit dosage form in single or divided
form, the unit dose or unit dosage form comprising a
therapeutically effective amount of one or more of the compounds or
conjugates of any one of the preceding clauses for treating a
disease in a host animal caused by a pathogenic population of
cells, said cells expressing PSMA. [0133] The composition or unit
dose or unit dosage form of any one of the preceding clauses
further comprising one or more carriers, diluents, or excipients,
or a combination thereof. [0134] A method for treating a disease in
a host animal caused by a pathogenic population of cells, said
cells expressing PSMA, the method comprising the step of
administering to the patient a composition comprising a
therapeutically effective amount of one or more of the compounds or
conjugates or one or more of the compositions or unit doses or unit
dosage forms of any one of clauses 1 to 73 [0135] Use of one or
more of the compounds or conjugates, compositions, unit doses, or
unit dosage forms of any one of the preceding clauses in the
manufacture of a medicament for treating a disease in a host animal
caused by a pathogenic population of cells, said cells expressing
PSMA. [0136] The composition, unit doses or unit dosage form,
method, or use of any one of the preceding clauses wherein the
cells are prostate cancer cells. [0137] The composition, unit doses
or unit dosage form, method, or use of any one of the preceding
clauses wherein the disease is prostate cancer. [0138] The
composition, unit doses or unit dosage form, method, or use of any
one of the preceding clauses wherein the host animal is a
human.
[0139] In reciting the foregoing and following collection of
embodiments and clauses, it is to be understood that all possible
combinations of features, and all possible subgenera and
sub-combinations are described. For example, it is to be understood
that when B is limited to a binding ligand comprising urea of
L-lysine and L-glutamate, L may be limited to a linker comprising
one or more aspartic acid diradicals, or alternatively, to
comprising a cysteine diradical, or alternatively, comprising
L-Asp-L-Asp-L-Cys(S--S), and so forth. Similarly, when D is limited
to a naturally occurring tubulsyin, L may be limited to a linker
comprising diradical of the formula
S--(CH.sub.2).sub.m--O--C(O)--N, or alternatively, to comprising a
cysteine disulfide diradical, or alternatively, comprising an
aminophenylacetic acid diradical, and so forth. Similarly, when B
is limited to a binding ligand comprising a urea or thiourea of
lysine and glutamate, or one or more carboxylic acid derivatives
thereof, L may be limited to a linker comprising one or more
D-aspartic acid diradicals, and D may be limited to a tubulysin, or
alternatively, L may be limited to a linker comprising a diradical
of the formula O--C(O)--N, and D may be limited to an imaging
agent, or alternatively, L may be limited to a linker comprising a
diradical of the formula S--(CH.sub.2).sub.m--O--C(O)--NH, and D
may be limited to a therapeutic agent, and so forth. Other
combinations, subgenera and sub-combinations are also described by
the collection of clauses.
[0140] In another embodiment, at least one drug is an imaging
agent. Illustrative imaging agents for the conjugates described
herein include, but are not limited to, radioisotopes, such as a
radioactive isotope of a metal coordinated to a chelating group.
Illustrative radioactive metal isotopes include technetium,
rhenium, gallium, gadolinium, indium, copper, and the like,
including isotopes .sup.111In, .sup.99mTc, .sup.64Cu, .sup.67Cu,
.sup.67Ga, .sup.68Ga, and the like. Additional illustrative
examples of radionuclide imaging agents are described in U.S. Pat.
No. 7,128,893, the disclosure of which is incorporated herein by
reference. Additional illustrative chelating groups are tripeptide
or tetrapeptides, including but not limited to tripeptides having
the formula:
##STR00037##
wherein R is independently selected in each instance H, alkyl,
heteroalkyl, cycloalkyl, heterocyclyl, alkenyl, alkynyl, aryl,
heteroaryl, arylalkyl, heteroarylalkyl, and the like, each of which
is optionally substituted. It is to be understood that one R
includes a heteroatom, such as nitro, oxygen, or sulfur, and is the
point of attachment of linker L. Illustratively, the following
chelating groups are described:
##STR00038##
where X is oxygen, nitrogen, or sulfur, and where X is attached to
linker L, and n is an integer from 1 to about 5.
[0141] Illustrative imaging agents also include, but are not
limited to, fluorescent agents, such as Oregon Green fluorescent
agents, including but not limited to Oregon Green 488, Oregon Green
514, and the like, AlexaFluor fluorescent agents, including but not
limited to AlexaFluor 488, AlexaFluor 647, and the like,
fluorescein, and related analogs, BODIPY fluorescent agents,
including but not limited to BODIPY F1, BODIPY 505, and the like,
rhodamine fluorescent agents, including but not limited to
tetramethylrhodamine, and the like, DyLight fluorescent agents,
including but not limited to DyLight 680, DyLight 800, and the
like, CW 800, IRdye 800CW, Texas Red, phycoerythrin, and others.
Further illustrative fluorescent agents include compounds of the
following formula:
##STR00039##
where X is oxygen, nitrogen, or sulfur, and where X is attached to
linker L; Y is OR.sup.a, NR.sup.a.sub.2, or NR.sup.a.sub.3.sup.+;
and Y' is O, NR.sup.a, or NR.sup.a.sub.2.sup.+; where each R is
independently selected in each instance from H, fluoro, sulfonic
acid, sulfonate, and salts thereof, and the like; and R.sup.a is
hydrogen or alkyl. Further illustrative fluorescent agents include
compounds of the following formula:
##STR00040##
where X is oxygen, nitrogen, or sulfur, and where X is attached to
linker L; and each R is independently selected in each instance
from H, alkyl, heteroalkyl, and the like; and n is an integer from
0 to about 4.
[0142] Illustrative imaging agents also include, but are not
limited to, PET imaging agents, and FRET imaging agents.
Illustrative PET imaging agents include .sup.18F, .sup.11C,
.sup.64Cu, .sup.65Cu, and the like. Illustrative FRET imaging
agents include .sup.64Cu, .sup.65Cu, and the like. It is to be
understood that in the case of .sup.18F and .sup.11C, the imaging
isotope may be directly attached to the linker, or alternatively
may be present on a structure attached to the linker. For example
in the case of .sup.18F, fluoroaryl groups, such as fluorophenyl,
difluorophenyl, fluoronitrophenyl, and the like are described. For
example in the case of .sup.11C, alkyl and alkyl aryl are
described.
[0143] In another embodiment, the drug can be any molecule capable
of modulating or otherwise modifying cell function, including
pharmaceutically active compounds. Illustrative drugs include, but
are not limited to, peptides, oligopeptides, retro-inverso
oligopeptides, proteins, protein analogs in which at least one
non-peptide linkage replaces a peptide linkage, apoproteins,
glycoproteins, enzymes, coenzymes, enzyme inhibitors, amino acids
and their derivatives, receptors and other membrane proteins;
antigens and antibodies thereto; haptens and antibodies thereto;
hormones, lipids, phospholipids, liposomes; toxins; antibiotics;
analgesics; bronchodilators; beta-blockers; antimicrobial agents;
antihypertensive agents; cardiovascular agents including
antiarrhythmics, cardiac glycosides, antianginals and vasodilators;
central nervous system agents including stimulants, psychotropics,
antimanics, and depressants; antiviral agents; antihistamines;
cancer drugs including chemotherapeutic agents; tranquilizers;
anti-depressants; H-2 antagonists; anticonvulsants; antinauseants;
prostaglandins and prostaglandin analogs; muscle relaxants;
anti-inflammatory substances; immunosuppressants, stimulants;
decongestants; antiemetics; diuretics; antispasmodics;
antiasthmatics; anti-Parkinson agents; expectorants; cough
suppressants; mucolytics; and mineral and nutritional
additives.
[0144] Illustrative chemotherapeutic agents also include, but are
not limited to, compounds that are cytotoxic, enhance tumor
permeability, inhibit tumor cell proliferation, promote apoptosis,
decrease anti-apoptotic activity in target cells, used to treat
diseases caused by infectious agents, enhance an endogenous immune
response directed to the pathogenic cells, or are useful for
treating a disease state caused by the pathogenic cells. Such
chemotherapeutic agents may operate by any of a large variety of
mechanisms of action. For example, cytotoxic compounds may disrupt
any of a wide variety of cellular mechanisms that are important for
cell survival and/or cell proliferation and/or cause cell death or
apoptosis.
[0145] Illustrative chemotherapeutic agents also include, but are
not limited to, adrenocorticoids and corticosteroids, alkylating
agents, antiandrogens, antiestrogens, androgens, aclamycin and
aclamycin derivatives, estrogens, antimetabolites such as cytosine
arabinoside, purine analogs, pyrimidine analogs, and methotrexate,
busulfan, carboplatin, chlorambucil, cisplatin and other platinum
compounds, tamoxiphen, taxol, paclitaxel, paclitaxel derivatives.
Taxotere, cyclophosphamide, daunomycin, rhizoxin, T2 toxin, plant
alkaloids, prednisone, hydroxyurea, teniposide, mitomycins,
discodermolides, microtubule inhibitors, epothilones, tubulysins,
cyclopropyl benz[e]indolone, seco-cyclopropyl benz[e]indolone,
O-Ac-seco-cyclopropyl benz[e]indolone, bleomycin and any other
antibiotic, nitrogen mustards, nitrosureas, vinca alkaloids, such
as vincristine, vinblastine, vindesine, vinorelbine and analogs and
derivative thereof such as deacetylvinblastine monohydrazide
(DAVLBH), colchicine, colchicine derivatives, allocolchicine,
thiocolchicine, trityl cysteine, halicondrin B, dolastatins such as
dolastatin 10, amanitins such as .alpha.-amanitin, camptothecin,
irinotecan, and other camptothecin derivatives thereof,
geldanamycin and geldanamycin derivatives, estramustine,
nocodazole, MAP4, colcemid, inflammatory and proinflammatory
agents, peptide and peptidomimetic signal transduction inhibitors,
rapamycins, such as sirolimus and everolimus, and any other drug or
toxin.
[0146] In another embodiment, at least one drug is selected from
cryptophycins, bortezomib, thiobortezomib, tubulysins, aminopterin,
rapamycins, such as everolimus and sirolimus, paclitaxel,
docetaxel, doxorubicin, daunorubicin, .alpha.-amanatin, verucarin,
didemnin B, geldanomycin, purvalanol A, ispinesib, budesonide,
dasatinib, epothilones, maytansines, and tyrosine kinase
inhibitors, including analogs and derivatives of each of the
foregoing.
[0147] Other drugs that can be included in the conjugates described
herein include amphotericin B, acyclovir, trifluridine,
ganciclovir, zidovudine, amantadine, ribavirin, and the like.
[0148] In another embodiment, at least one drug is a tubulysin. As
used herein, the term "tubulysin" generally refers to the compounds
described herein and analogs and derivatives thereof. It is also to
be understood that any corresponding pharmaceutically acceptable
salt is also included in the illustrative embodiments described
herein. Illustrative derivatives of tubulysins include, but are not
limited to, those compounds that may be synthetically prepared from
the compounds described herein. It is to be understood that such
derivatives may include prodrugs of the compounds described herein,
compounds described herein that include one or more protection or
protecting groups, including compounds that are used in the
preparation of other compounds described herein.
[0149] As described herein, the tubulysin compounds may be
inhibitors of tubulin polymerization, and also may be
DNA-alkylators.
[0150] Illustrative tubulysins include, but are not limited to
compounds of the formula
##STR00041##
and pharmaceutical salts thereof are described, where
[0151] n is 1-3;
[0152] V is hydrogen, OR.sup.2, or halo, and W is hydrogen,
OR.sup.2, or alkyl, where R.sup.2 is independently selected in each
instance from hydrogen, alkyl, and C(O)R.sup.3, where R.sup.3 is
alkyl, cycloalkyl, alkenyl, aryl, or arylalkyl, each of which is
optionally substituted; providing that R.sup.2 is not H when both V
and W are OR.sup.2; or V and W are taken together with the attached
carbon to form a carbonyl:
[0153] X is hydrogen, alkyl, such as C.sub.1-4 alkyl, or alkenyl,
such as C.sub.2-4 alkenyl, each of which is optionally
substituted;
[0154] Z is alkyl or C(O)R.sup.4, where R.sup.4 is alkyl, CF.sub.3,
or aryl; or when Y is present, Z is alkyl; and Y is O:
[0155] Ar is aryl, such as phenyl, or heteroaryl, each of which is
optionally substituted; and
[0156] R is OH or R and the carbonyl to which it is attached is a
carboxylic acid derivative, such as an acylhydrazide.
[0157] In another embodiment, X is CH.sub.2QR.sup.9, where Q is
--N--, --O--, or --S--; R.sup.9 is hydrogen or alkyl, alkenyl,
cycloalkyl, aryl, or arylalkyl, each of which is optionally
substituted, or C(O)R.sup.10, where R.sup.10 is hydrogen or alkyl,
alkenyl, cycloalkyl, aryl, or arylalkyl In another embodiment,
R.sup.9 and Q are taken together to form S(O).sub.2R.sup.10,
P(O)(OR.sup.10a).sub.2, where R.sup.10 and OR.sup.10a are
independently selected in each instance from the group consisting
of hydrogen, and alkyl, alkenyl, cycloalkyl, aryl, heteroaryl, and
arylalkyl, each of which is optionally substituted, or R.sup.10a is
a metal cation.
[0158] In another embodiment, X is H. Illustrative examples of such
compounds, and their preparation are described in J. Med. Chem.
10.1021/jm701321p (2008), the disclosure of which is incorporated
herein by reference.
[0159] In another embodiment, X is a radical of the formula
##STR00042##
where R.sup.12 represents 1 or more substituents selected from
alkyl, alkenyl, cycloalkyl, aryl, and arylalkyl, each of which is
optionally substituted. It is to be understood that other olefins
may form by isomerization, depending on the conditions of the
reaction and the identity of R.sup.12. For example, when R.sup.12
is alkyl, it is appreciated that under the reaction conditions, the
double bond can migrate to other carbon atoms along the alkenyl
chain, including to form the terminal or .omega.-olefin.
[0160] In another embodiment, X is a radical of the formula
##STR00043##
where R.sup.13 is C(O)R.sup.10, C(O)OR.sup.10 or CN, where R.sup.10
is independently selected in each instance.
[0161] In another embodiment, X is CH.sub.2--OH.
[0162] In another embodiment, X is CH.sub.2--X.sup.A, where X.sup.A
is halogen, OS(O).sub.2R.sup.10, OP(O)(OR.sup.10a)R.sup.10, or
OP(O)(OR.sup.10a).sub.2; where R.sup.10 and R.sup.10a are
independently selected in each instance from the group consisting
of hydrogen, alkyl, alkenyl, cycloalkyl, aryl, and arylalkyl, each
of which is optionally substituted, or R.sup.10a is a metal
cation.
[0163] In another embodiment of any of the foregoing embodiments,
Ar is optionally substituted aryl. In another embodiment of any of
the foregoing embodiments, Ar is a radical of the formula
##STR00044##
where R.sup.1 is hydrogen, or R.sup.1 represents 1 to 3
substituents independently selected from the group consisting of
halo, nitro, carboxylate or a derivative thereof, cyano, hydroxyl,
alkyl, haloalkyl, alkoxy, haloalkoxy, and OR.sup.6, where R.sup.6
is hydrogen or optionally substituted alkyl, heteroalkyl, aryl, a
phenol protecting group, a prodrug moiety, C(O)R.sup.7,
P(O)(OR.sup.8).sub.2, or SO.sub.3R.sup.8, where R.sup.7 and R.sup.8
are independently selected in each instance from hydrogen, or
alkyl, alkenyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and
arylalkyl, each of which is optionally substituted, or R.sup.8 is a
metal cation are described.
[0164] In another embodiment of any of the foregoing embodiments, Z
is methyl. In another embodiment of any of the foregoing
embodiments, R.sup.1 is H. In another embodiment of any of the
foregoing embodiments, R.sup.1 is OR.sup.6 at C(4), where R.sup.6
is hydrogen, alkyl, or COR.sup.7. In another embodiment of any of
the foregoing embodiments, V is hydrogen, and W is OC(O)R.sup.3. In
another embodiment of any of the foregoing embodiments. V is
hydrogen, and W is acetyloxy.
[0165] In another embodiment of any of the foregoing embodiments,
the compounds of the various formulae have the following absolute
configuration:
##STR00045##
at each of the indicated asymmetric carbon atoms.
[0166] Additional illustrative tubulysins that are useable in the
conjugates described herein include the following:
##STR00046##
TABLE-US-00001 Tubulysin X.sup.B EC0313 --O--CH.sub.3 EC0346
--O--(CH.sub.2).sub.2--OH EC0356
--O--(CH.sub.2).sub.2CH(CH.sub.3).sub.2 EC0374
--S--(CH.sub.2).sub.2--SH EC0386 --OH EC0550
--(CH.sub.2).sub.2--CH.dbd.CH.sub.2 EC0560
--S--(CH.sub.2).sub.2--OH EC0575
--O--C(O)--(CH.dbd.CH)--CH.sub.2--Cl EC0585
--NH--C(O)--CH.sub.2CH(CH.sub.3).sub.2 EC0611
--O--(CH.sub.2).sub.2CH.sub.3 EC0623
--S--(CH.sub.2).sub.2CH.sub.3
and pharmaceutical salts thereof.
[0167] In another embodiment, the tubulysin is a naturally
occurring tubulysin. Natural tubulysins are generally linear
tetrapeptides consisting of N-methyl pipecolic acid (Mep),
isoleucine (Ile), an unnatural aminoacid called tubuvalin (Tuv),
and either an unnatural aminoacid called tubutyrosine (Tut, an
analog of tyrosine) or an unnatural amninoacid called
tubuphenylalanine (Tup, an analog of phenylalanine). In another
embodiment, naturally occurring tubulysins, and analogs and
derivatives thereof, of the following general formula are
described
##STR00047##
and pharmaceutical salts thereof, where Ar, R, and R.sup.10 are as
described in the various embodiments herein.
[0168] In another embodiment, the naturally occurring tubulysins of
the following general formula are described
##STR00048##
TABLE-US-00002 Factor R.sup.10 R.sup.1 A (CH.sub.3).sub.2CHCH.sub.2
OH B CH.sub.3(CH.sub.2).sub.2 OH C CH.sub.3CH.sub.2 OH D
(CH.sub.3).sub.2CHCH.sub.2 H E CH.sub.3(CH.sub.2).sub.2 H F
CH.sub.2CH.sub.3 H G (CH.sub.3).sub.2C.dbd.CH OH H CH.sub.3 H I
CH.sub.3 OH
and pharmaceutical salts thereof.
[0169] It is to be understood that the conjugate of the tubulysin
or analog or derivative thereof may be formed at any position.
Illustratively, conjugates of tubulysins are described where the
linker (L) is attached to any of the following positions:
##STR00049##
where the (*) symbol indicates optional attachment locations.
[0170] In another embodiment, compounds are described herein where
the conjugate is formed at the terminal carboxylic acid group or
the terminal acylhydrazine derivative group of each of the
tybulysins described herein.
[0171] Additional tubulysins useful in preparing the conjugates
described herein are described in US patent application publication
Nos. 2006/0128754 and 2005/0239713, the disclosures of which are
incorporated herein by reference. Additional tubulysins useful in
preparing the conjugates described herein are described in
co-pending U.S. patent application publication No. 2010/0240701 the
disclosure of which is incorporated herein by reference. Tubulysins
may also be prepared are described in Peltier et al., "The Total
Synthesis of Tubulysin D," J. Am. Chem. Soc. 128:16018-19 (2006),
the disclosure of which is incorporated herein by reference.
[0172] In another embodiment, at least one drug is a rapamycin. As
used herein, the term "a rapamycin" is understood to include
sirolimus (rapamycin), temsirolimus, everolimus, and ridaforolimus,
and related compounds, and compounds of the formula
##STR00050##
and pharmaceutically acceptable salts thereof, wherein
[0173] Y.sup.A is OR.sup.C or OCH.sub.2CH.sub.2OR.sup.C;
[0174] one of R.sup.A, R.sup.B, or R.sup.C is a bond connected to
L; and
[0175] the other two of R.sup.A, R.sup.B, and R.sup.C are
independently selected in each case from the group consisting of
hydrogen, optionally substituted heteroalkyl, prodrug forming
group, and C(O)R.sup.D, where R.sup.D is in each instance
independently selected from the group consisting of hydrogen, and
alkyl, alkenyl, heteroalkyl, cycloalkyl, cycloheteroalkyl, aryl,
arylalkyl, heteroaryl, and heteroarylalkyl, each of which is
optionally substituted is described.
[0176] In another embodiment, at least one drug is a vinca
alkaloids, such as vincristine, vinblastine, vindesine, vinorelbine
and analogs and derivative thereof such as deacetylvinblastine
monohydrazide (DAVLBH).
[0177] In another embodiment, at least one drug is a mitomycin, or
an analog or derivative thereof.
[0178] In another embodiment, the conjugates described herein
include at least two drugs, including those described herein, In
one variation, the drugs are the same. In another variation, at
least two of the drugs are different. In another variation, the two
or more drugs are selected from vinca alkaloids, cryptophycins,
bortezomib, thiobortezomib, tubulysins, aminopterin, rapamycins,
such as everolimus and sirolimus, paclitaxel, docetaxel,
doxorubicin, daunorubicin, .alpha.-amanatin, verucarin, didemnin B,
geldanomycin, purvalanol A, ispinesib, budesonide, dasatinib,
epothilones, maytansines, and tyrosine kinase inhibitors, including
analogs and derivatives of each of the foregoing.
[0179] As used herein, the term "linker" includes is a chain of
atoms that connects two or more functional parts of a molecule to
form a conjugate. Illustratively, the chain of atoms is selected
from C, N, O, S, Si, and P, or C, N, O, S, and P, or C, N, O, and
S. The chain of atoms covalently connects different functional
capabilities of the conjugate, such as binding ligands, drugs,
diagnostic agents, imaging agents, and the like. The linker may
have a wide variety of lengths, such as in the range from about 2
to about 100 atoms in the contiguous backbone. The atoms used in
forming the linker may be combined in all chemically relevant ways,
such as chains of carbon atoms forming alkylene, alkenylene, and
alkynylene groups, and the like; chains of carbon and oxygen atoms
forming ethers, polyoxyalkylene groups, or when combined with
carbonyl groups forming esters and carbonates, and the like; chains
of carbon and nitrogen atoms forming amines, imines, polyamines,
hydrazines, hydrazones, or when combined with carbonyl groups
forming amides, ureas, semicarbazides, carbazides, and the like;
chains of carbon, nitrogen, and oxygen atoms forming alkoxyamines,
alkoxylamines, or when combined with carbonyl groups forming
urethanes, amino acids, acyloxylamines, hydroxamic acids, and the
like; and many others. In addition, it is to be understood that the
atoms forming the chain in each of the foregoing illustrative
embodiments may be either saturated or unsaturated, thus forming
single, double, or triple bonds, such that for example, alkanes,
alkenes, alkynes, imines, and the like may be radicals that are
included in the linker. In addition, it is to be understood that
the atoms forming the linker may also be cyclized upon each other
or be part of cyclic structure to form divalent cyclic structures
that form the linker, including cyclo alkanes, cyclic ethers,
cyclic amines, and other heterocycles, arylenes, heteroarylenes,
and the like in the linker. In this latter arrangement, it is to be
understood that the linker length may be defined by any pathway
through the one or more cyclic structures. Illustratively, the
linker length is defined by the shortest pathway through the each
one of the cyclic structures. It is to be understood that the
linkers may be optionally substituted at any one or more of the
open valences along the chain of atoms, such as optional
substituents on any of the carbon, nitrogen, silicon, or phosphorus
atoms. It is also to be understood that the linker may connect the
two or more functional parts of a molecule to form a conjugate at
any open valence, and it is not necessary that any of the two or
more functional parts of a molecule forming the conjugate are
attached at any apparent end of the linker.
[0180] In another embodiment, the linker (L) comprises a radical of
the formula
##STR00051##
where m1, m2, m3, n, p, q, and r are integers that are each
independently selected from the range of 0 to about 8, providing
that at least one of m1, m2, m3, n, p, q, and r is not 0; AA is an
amino acid; and drugs are optionally attached at one or more of the
(*) atoms. It is to be understood that the drugs may be directly
attached, or attached through additional portions of the linker
(L). In another embodiment, AA is a naturally occurring amino acid
of either the natural or unnatural configuration. In another
embodiment, one or more of AA is a hydrophilic amino acid. In
another embodiment, one or more of AA is Asp and/or Arg. In another
embodiment, the integer n is 1 or greater. In another embodiment,
the integer n is 2 or greater. In another embodiment, the integer n
is 3 or greater. In another embodiment, the integer n is 4 or
greater. In another embodiment, the integer n is 5 or greater. In
another aspect, the integer q is 1 or greater. In another
embodiment, the integer m1 is 1 or greater. In another embodiment,
the integer m1 is 1. In another embodiment, the integer m2 is 1 or
greater. In another embodiment, the integer m2 is 1. In another
embodiment, the integer m3 is 1 or greater. In another embodiment,
the integer m3 is 1. In another embodiment, the integer p is 1 or
greater. In another embodiment, the integer p is 1. In another
embodiment, the integer p is 2. In another embodiment, the integer
q is 1 or greater. In another embodiment, the integer q is 1. In
another embodiment, the integer q is 2. In another embodiment, the
integer r is 1 or greater. In another embodiment, the integer r is
1. In another embodiment, the integer r is 2.
[0181] It is to be understood that all combinations of the
foregoing embodiments are described herein. For example, in another
embodiment, n is 1 or greater, and m1 is one or greater, or n is 1
or greater, m1 is 1, and q is 1; and so forth. For example, in
another embodiment, n is 1 or greater, and m2 is one or greater, or
n is 2 or greater, m2 is 1, and q is 1; or n is 2 or greater, m3 is
1, q is 1, and p is 1; and so forth. For example, in another
embodiment, n is 1 or greater, and m1 is one or greater; or n is 2
or greater, m3 is 1, and q is 1; or n is 2 or greater, m2 is 1, q
is 1, and p is 1; or n is 2 or greater, m1 is 1, q is 1, and r is
1; or n is 2 or greater, m3 is 1, q is 1, p is 1, and r is 1; and
so forth.
[0182] In another embodiment, the polyvalent linker includes one or
more divalent hydrophilic radicals, as described herein, which may
also be referred to as spacer linkers. It is appreciated that the
arrangement and/or orientation of the various hydrophilic linkers
may be in a linear or branched fashion, or both. For example, the
hydrophilic linkers may form the backbone of the linker forming the
conjugate between the ligand and the one or more drugs.
Alternatively, the hydrophilic portion of the linker may be pendant
to or attached to the backbone of the chain of atoms connecting the
binding ligand B to the one or more drugs D. In this latter
arrangement, the hydrophilic portion may be proximal or distal to
the backbone chain of atoms.
[0183] In another embodiment, the linker is generally linear, and
the hydrophilic groups are arranged generally in a series to form a
chain-like linker in the conjugate. Said another way, the
hydrophilic groups form some or all of the backbone of the linker
in such a linear linker embodiment.
[0184] In another embodiment, the linker is branched with
hydrophilic groups. In this branched embodiment, the hydrophilic
groups may be proximal to the backbone or distal to the backbone.
In each of these arrangements, the linker is generally more
spherical or cylindrical in shape. In another embodiment, the
linker is shaped like a bottle-brush. In another embodiment, the
backbone of the linker is formed by a linear series of amides, and
the hydrophilic portion of the linker is formed by a parallel
arrangement of branching side chains, such as by connecting
monosaccharides, sulfonates, and the like, and derivatives and
analogs thereof.
[0185] It is understood that the linker (L) may be neutral or
ionizable under certain conditions, such as physiological
conditions encountered in vivo. For ionizable linkers, under the
selected conditions, the linker may deprotonate to form a negative
ion, or alternatively become protonated to form a positive ion. It
is appreciated that more than one deprotonation or protonation
event may occur. In addition, it is understood that the same linker
may deprotonate and protonate to form inner salts or zwitterionic
compounds.
[0186] In another embodiment, the hydrophilic spacer linkers are
neutral, an in particular neutral under physiological conditions,
the linkers do not significantly protonate nor deprotonate. In
another embodiment, the hydrophilic spacer linkers may be
protonated to carry one or more positive charges. It is understood
that the protonation capability is condition dependent. In one
aspect, the conditions are physiological conditions, and the linker
is protonated in vivo. In another embodiment, the spacers include
both regions that are neutral and regions that may be protonated to
carry one or more positive charges. In another embodiment, the
spacers include both regions that may be deprotonated to carry one
or more negative charges and regions that may be protonated to
carry one or more positive charges. It is understood that in this
latter embodiment that zwitterions or inner salts may be
formed.
[0187] In another embodiment, the regions of the linkers that may
be deprotonated to carry a negative charge include carboxylic
acids, such as aspartic acid, glutamic acid, and longer chain
carboxylic acid groups, and sulfuric acid esters, such as alkyl
esters of sulfuric acid. In another embodiment, the regions of the
linkers that may be protonated to carry a positive charge include
amino groups, such as polyaminoalkylenes including ethylene
diamines, propylene diamines, butylene diamines and the like,
and/or heterocycles including pyrollidines, piperidines,
piperazines, and other amino groups, each of which is optionally
substituted. In another embodiment, the regions of the linkers that
are neutral include poly hydroxyl groups, such as sugars,
carbohydrates, saccharides, inositols, and the like, and/or
polyether groups, such as polyoxyalkylene groups including
polyoxyethylene, polyoxypropylene, and the like.
[0188] In another embodiment, the hydrophilic spacer linkers
described herein include are formed primarily from carbon,
hydrogen, and oxygen, and have a carbon/oxygen ratio of about 3:1
or less, or of about 2:1 or less. In another embodiment, the
hydrophilic linkers described herein include a plurality of ether
functional groups. In another embodiment, the hydrophilic linkers
described herein include a plurality of hydroxyl functional groups.
Illustrative fragments and radicals that may be used to form such
linkers include polyhydroxyl compounds such as carbohydrates,
polyether compounds such as polyethylene glycol units, and acid
groups such as carboxyl and alkyl sulfuric acids. In one variation,
oligoamide spacers, and the like may also be included in the
linker.
[0189] Illustrative divalent hydrophilic linkers include
carbohydrates such as saccharopeptides as described herein that
include both a peptide feature and sugar feature; glucuronides,
which may be incorporated via [2+3] Huisgen cyclization, also known
as click chemistry; .beta.-alkyl glycosides, such as of
2-deoxyhexapyranoses (2-deoxyglucose, 2-deoxyglucuronide, and the
like), and .beta.-alkyl mannopyranosides. Illustrative PEG groups
include those of a specific length range from about 4 to about 20
PEG groups. Illustrative alkyl sulfuric acid esters may also be
introduced with click chemistry directly into the backbone.
Illustrative oligoamide spacers include EDTA and DTPA spacers,
3-amino acids, and the like.
[0190] In another embodiment, the polyvalent linker L comprises one
or more polyethers, such as the linkers of the following
formulae:
##STR00052##
where m is an integer independently selected in each instance from
1 to about 8; p is an integer selected 1 to about 10; and n is an
integer independently selected in each instance from 1 to about 3.
In one aspect, m is independently in each instance 1 to about 3. In
another aspect, n is 1 in each instance. In another aspect, p is
independently in each instance about 4 to about 6. Illustratively,
the corresponding polypropylene polyethers corresponding to the
foregoing are contemplated herein and may be included in the
conjugates as hydrophilic spacer linkers. In addition, it is
appreciated that mixed polyethylene and polypropylene polyethers
may be included in the conjugates as hydrophilic spacer linkers.
Further, cyclic variations of the foregoing polyether compounds,
such as those that include tetrahydrofuranyl, 1,3-dioxanes,
1,4-dioxanes, and the like are contemplated herein.
[0191] In another embodiment, the polyvalent linker L comprises a
plurality of hydroxyl functional groups, such as linkers that
incorporate monosaccharides, oligosaccharides, polysaccharides, and
the like. It is to be understood that the polyhydroxyl containing
spacer linkers comprises a plurality of --(CROH)-- groups, where R
is hydrogen or alkyl.
[0192] In another embodiment, the polyvalent linker L comprises one
or more of the following fragments:
##STR00053## ##STR00054##
wherein R is H, alkyl, cycloalkyl, or arylalkyl; m is an integer
from 1 to about 3; n is an integer from 1 to about 5, or from 2 to
about 5, p is an integer from 1 to about 5, and r is an integer
selected from 1 to about 3. In one aspect, the integer n is 3 or 4.
In another aspect, the integer p is 3 or 4. In another aspect, the
integer r is 1.
[0193] In another embodiment, the polyvalent linker L comprises one
or more of the following fragments:
##STR00055##
wherein R is H, alkyl, cycloalkyl, or arylalkyl; m is an integer
from 1 to about 3; n is an integer from 1 to about 5, or from 2 to
about 5, p is an integer from 1 to about 5, and r is an integer
selected from 1 to about 3. In one aspect, the integer n is 3 or 4.
In another aspect, the integer p is 3 or 4. In another aspect, the
integer r is 1.
[0194] In another embodiment, the polyvalent linker L comprises one
or more of the following cyclic polyhydroxyl groups:
##STR00056## ##STR00057##
wherein n is an integer from 2 to about 5, p is an integer from 1
to about 5, and r is an integer from 1 to about 4. In one aspect,
the integer n is 3 or 4. In another aspect, the integer p is 3 or
4. In another aspect, the integer r is 2 or 3. It is understood
that all stereochemical forms of such sections of the linkers are
contemplated herein. For example, in the above formula, the section
may be derived from ribose, xylose, glucose, mannose, galactose, or
other sugar and retain the stereochemical arrangements of pendant
hydroxyl and alkyl groups present on those molecules. In addition,
it is to be understood that in the foregoing formulae, various
deoxy compounds are also contemplated. Illustratively, compounds of
the following formulae are contemplated:
##STR00058##
wherein n is equal to or less than r, such as when r is 2 or 3, n
is 1 or 2, or 1, 2, or 3, respectively.
[0195] In another embodiment, the polyvalent linker L comprises one
or more polyhydroxyl radicals of the following formula:
##STR00059##
wherein n and r are each an integer selected from 1 to about 3. In
one aspect, the linker includes one or more polyhydroxyl compounds
of the following formulae:
##STR00060##
It is understood that all stereochemical forms of such sections of
the linkers are contemplated herein. For example, in the above
formula, the section may be derived from ribose, xylose, glucose,
mannose, galactose, or other sugar and retain the stereochemical
arrangements of pendant hydroxyl and alkyl groups present on those
molecules.
[0196] In another embodiment, the polyvalent linker L comprises one
or more polyhydroxyl groups that are spaced away from the backbone
of the linker. In one embodiment, such carbohydrate groups or
polyhydroxyl groups are connected to the back bone by a triazole
group, forming triazole-linked hydrophilic spacer linkers.
Illustratively, the linker includes fragments of the following
formulae:
##STR00061##
wherein n, m, and r are integers and are each independently
selected in each instance from 1 to about 5. In one illustrative
aspect, m is independently 2 or 3 in each instance. In another
aspect, r is 1 in each instance. In another aspect, n is 1 in each
instance. In one variation, the group connecting the polyhydroxyl
group to the backbone of the linker is a different heteroaryl
group, including but not limited to, pyrrole, pyrazole,
1,2,4-triazole, furan, oxazole, isoxazole, thienyl, thiazole,
isothiazole, oxadiazole, and the like. Similarly, divalent
6-membered ring heteroaryl groups are contemplated. Other
variations of the foregoing illustrative hydrophilic spacer linkers
include oxyalkylene groups, such as the following formulae:
##STR00062##
wherein n and r are integers and are each independently selected in
each instance from 1 to about 5; and p is an integer selected from
1 to about 4.
[0197] In another embodiment, the polyvalent linker L comprises one
or more carbohydrate groups or polyhydroxyl groups connected to the
back bone by an amide group, forming amide-linked hydrophilic
spacer linkers. Illustratively, such linkers include fragments of
the following formulae:
##STR00063##
wherein n is an integer selected from 1 to about 3, and m is an
integer selected from 1 to about 22. In one illustrative aspect, n
is 1 or 2. In another illustrative aspect, m is selected from about
6 to about 10, illustratively 8. In one variation, the group
connecting the polyhydroxyl group to the backbone of the linker is
a different functional group, including but not limited to, esters,
ureas, carbamates, acylhydrazones, and the like. Similarly, cyclic
variations are contemplated. Other variations of the foregoing
illustrative hydrophilic spacer linkers include oxyalkylene groups,
such as the following formulae:
##STR00064##
wherein n and r are integers and are each independently selected in
each instance from 1 to about 5; and p is an integer selected from
1 to about 4.
[0198] In another embodiment, the polyvalent linker L comprises one
or more of the following fragments:
##STR00065## ##STR00066## ##STR00067##
wherein R is H, alkyl, cycloalkyl, or arylalkyl; m is an
independently selected integer from 1 to about 3; n is an integer
from 1 to about 6, p is an integer from 1 to about 5, and r is an
integer selected from 1 to about 3. In one variation, the integer n
is 3 or 4. In another variation, the integer p is 3 or 4. In
another variation, the integer r is 1.
[0199] In another embodiment, the polyvalent linker L comprises one
or more of the following fragments:
##STR00068##
wherein R is H, alkyl, cycloalkyl, or arylalkyl; m is an
independently selected integer from 1 to about 3; n is an integer
from 2 to about 6, p is an integer from 1 to about 5, and r is an
integer selected from 1 to about 3. In one variation, the integer n
is 3 or 4. In another variation, the integer p is 3 or 4. In
another variation, the integer r is 1.
[0200] In another embodiment, the polyvalent linker L comprises one
or more of the following fragments:
##STR00069## ##STR00070## ##STR00071##
wherein m is an independently selected integer from 1 to about 3; n
is an integer from 1 to about 6, p is an integer from 1 to about 5,
and r is an integer selected from 1 to about 3. In one variation,
the integer n is 3 or 4. In another variation, the integer p is 3
or 4. In another variation, the integer r is 1.
[0201] In another embodiment, the polyvalent linker L comprises one
or more of the following fragments:
##STR00072##
wherein m is an independently selected integer from 1 to about 3; n
is an integer from 2 to about 6, p is an integer from 1 to about 5,
and r is an integer selected from 1 to about 3. In one variation,
the integer n is 3 or 4. In another variation, the integer p is 3
or 4. In another variation, the integer r is 1.
[0202] In another embodiment, the polyvalent linker L comprises one
or more of the following fragments:
##STR00073## ##STR00074##
wherein m is an independently selected integer from 0 to about 3; n
is an integer from 1 to about 6, p is an integer from 1 to about 5,
and r is an integer selected from 1 to about 3. In one variation,
the integer n is 3 or 4. In another variation, the integer p is 3
or 4. In another variation, the integer r is 1.
[0203] In another embodiment, the polyvalent linker L comprises a
combination of backbone and branching side motifs such as is
illustrated by the following formulae
##STR00075##
wherein n is an integer independently selected in each instance
from 0 to about 3. The above formula are intended to represent 4,
5, 6, and even larger membered cyclic sugars. In addition, it is to
be understood that the above formula may be modified to represent
deoxy sugars, where one or more of the hydroxy groups present on
the formulae are replaced by hydrogen, alkyl, or amino. In
addition, it is to be understood that the corresponding carbonyl
compounds are contemplated by the above formulae, where one or more
of the hydroxyl groups is oxidized to the corresponding carbonyl.
In addition, in this illustrative embodiment, the pyranose includes
both carboxyl and amino functional groups and (a) can be inserted
into the backbone and (b) can provide synthetic handles for
branching side chains in variations of this embodiment. Any of the
pendant hydroxyl groups may be used to attach other chemical
fragments, including additional sugars to prepare the corresponding
oligosaccharides. Other variations of this embodiment are also
contemplated, including inserting the pyranose or other sugar into
the backbone at a single carbon, i.e. a spiro arrangement, at a
geminal pair of carbons, and like arrangements. For example, one or
two ends of the linker, or the drug D, or the binding ligand B may
be connected to the sugar to be inserted into the backbone in a
1,1; 1,2; 1,3; 1,4; 2,3, or other arrangement.
[0204] In another embodiment, the hydrophilic spacer linkers
described herein include are formed primarily from carbon,
hydrogen, and nitrogen, and have a carbon/nitrogen ratio of about
3:1 or less, or of about 2:1 or less. In one aspect, the
hydrophilic linkers described herein include a plurality of amino
functional groups.
[0205] In another embodiment, the polyvalent linker L comprises one
or more amino groups of the following formulae:
##STR00076##
where n is an integer independently selected in each instance from
1 to about 3. In one aspect, the integer n is independently 1 or 2
in each instance. In another aspect, the integer n is 1 in each
instance.
[0206] In another embodiment, the polyvalent linker L comprises one
or more sulfuric acid esters, such as an alkyl ester of sulfuric
acid. Illustratively, the linker includes the following
formula(e):
##STR00077##
where n is an integer independently selected in each instance from
1 to about 3. Illustratively, n is independently 1 or 2 in each
instance.
[0207] It is understood, that in such polyhydroxyl, polyamino,
carboxylic acid, sulfuric acid, and like linkers that include free
hydrogens bound to heteroatoms, one or more of those free hydrogen
atoms may be protected with the appropriate hydroxyl, amino, or
acid protecting group, respectively, or alternatively may be
blocked as the corresponding pro-drugs, the latter of which are
selected for the particular use, such as pro-drugs that release the
parent drug under general or specific physiological conditions.
[0208] In another embodiment, the polyvalent linker comprises one
or more of the following divalent radicals:
##STR00078##
wherein n is an integer from 2 to about 5, p is an integer from 1
to about 5, and r is an integer from 1 to about 4, as described
above.
[0209] It is to be further understood that in the foregoing
embodiments, open positions, such as (*) atoms are locations for
attachment of the binding ligand (B) or any drug (D) to be
delivered. In addition, it is to be understood that such attachment
of either or both of B and any D may be direct or through an
intervening linker comprising one or more of the radicals described
herein. In addition, (*) atoms may form releasable linkers with any
drug D, or other portion of the linker L.
[0210] In another embodiment, the hydrophilic spacer linker
comprises one or more carbohydrate containing or polyhydroxyl group
containing linkers. In another embodiment, the hydrophilic spacer
linker comprises at least three carbohydrate containing or
polyhydroxyl group containing linkers. In another embodiment, the
hydrophilic spacer linker comprises one or more carbohydrate
containing or polyhydroxyl group containing linkers, and one or
more aspartic acids. In another embodiment, the hydrophilic spacer
linker comprises one or more carbohydrate containing or
polyhydroxyl group containing linkers, and one or more glutamic
acids. In another embodiment, the hydrophilic spacer linker
comprises one or more carbohydrate containing or polyhydroxyl group
containing linkers, one or more glutamic acids, one or more
aspartic acids, and one or more beta amino alanines. In a series of
variations, in each of the foregoing embodiments, the hydrophilic
spacer linker also includes one or more cysteines. In another
series of variations, in each of the foregoing embodiments, the
hydrophilic spacer linker also includes at least one arginine.
[0211] In another embodiment, the polyvalent linker L includes a
hydrophilic spacer linker comprising one or more divalent
1,4-piperazines that are included in the chain of atoms connecting
at least one of the binding ligands (L) with at least one of the
drugs (D). In one variation, the hydrophilic spacer linker includes
one or more carbohydrate containing or polyhydroxyl group
containing linkers. In another variation, the hydrophilic spacer
linker includes one or more carbohydrate containing or polyhydroxyl
group containing linkers and one or more aspartic acids. In another
variation, the hydrophilic spacer linker includes one or more
carbohydrate containing or polyhydroxyl group containing linkers
and one or more glutamic acids. In a series of variations, in each
of the foregoing embodiments, the hydrophilic spacer linker also
includes one or more cysteines. In another series of variations, in
each of the foregoing embodiments, the hydrophilic spacer linker
also includes at least one arginine.
[0212] In another embodiment, the hydrophilic spacer linker
comprises one or more oligoamide hydrophilic spacers, such as but
not limited to aminoethylpiperazinylacetamide.
[0213] In another embodiment, the polyvalent linker L includes a
hydrophilic spacer linker comprising one or more triazole linked
carbohydrate containing or polyhydroxyl group containing linkers.
In another embodiment, the hydrophilic spacer linker comprises one
or more amide linked carbohydrate containing or polyhydroxyl group
containing linkers. In another embodiment, the hydrophilic spacer
linker comprises one or more PEG groups and one or more cysteines.
In another embodiment, the hydrophilic spacer linker comprises one
or more EDTE derivatives.
[0214] In another embodiment, the polyvalent linker L includes a
divalent radical of the formula
##STR00079##
wherein * indicates the point of attachment to a folate and **
indicates the point of attachment to a drug; and F and G are each
independently 1, 2, 3 or 4 are described.
[0215] In another embodiment, the polyvalent linker L includes a
trivalent radical of the formula
##STR00080##
wherein *, **, *** each indicate points of attachment to the folate
receptor binding moiety B, and the one or more drugs D. It is to be
understood that when there are fewer drugs, *, **, *** are
substituted with hydrogen or a heteroatom. F and G are each
independently 1, 2, 3 or 4; and W.sup.1 is NH or O is described. In
another aspect, m.sup.1 is 0 or 1.
[0216] In any of the embodiments described herein heteroatom
linkers can also be included in the polyvalent linker L, such as
--NR.sup.1R.sup.2--, oxygen, sulfur, and the formulae
--(NHR.sup.1NHR.sup.2)--, --SO--, --(SO.sub.2)--, and
--N(R.sup.3)O--, wherein R.sup.1, R.sup.2, and R.sup.3 are each
independently selected from hydrogen, alkyl, aryl, arylalkyl,
substituted aryl, substituted arylalkyl, heteroaryl, substituted
heteroaryl, and alkoxyalkyl. It is to be understood that the
heteroatom linkers may be used to covalently attach any of the
radicals described herein, including drug radicals D to the
polyvalent linker, ligand radicals B to the polyvalent linker, or
various di and polyvalent radicals that from the polyvalent linker
L
[0217] Illustrative additional bivalent radicals that can be used
to form parts of the linker are as follows.
##STR00081## ##STR00082## ##STR00083## ##STR00084##
##STR00085##
[0218] In another embodiment, the polyvalent linker L is a
releasable linker.
[0219] As used herein, the term "releasable linker" refers to a
linker that includes at least one bond that can be broken under
physiological conditions when the compounds described herein are
delivered to or inside of the target cell. The linker itself may
include one or more cleavable, scissile, or breakable bond, or form
one or more cleavable, scissile, or breakable bonds with the PSMA
binding ligand (B), and/or with one or more of the drugs (D).
However, it is appreciated that releasable linkers described herein
are advantageously not cleavable, scissile, or breakable until the
conjugate containing the releasable linker is at or near the
intended target site. Accordingly, releasable linkers described
herein do not generally include those linkers that have bonds that
are substantially cleavable, scissile, or breakable under
non-target conditions, or in non-target tissues. Similarly,
releasable linkers described herein do not include those linkers
that include bonds that are substantially only cleavable, scissile,
or breakable under non-physiological conditions.
[0220] The term releasable linker does not generally refer simply
to a bond that is labile in vivo, such as in serum, plasma, the
gastrointestinal tract, or liver, unless those systems are the
target for the cell surface receptor binding ligand. However, after
delivery and/or selective targeting, releasable linkers may be
cleaved by any process that includes at least one bond being broken
in the linker or at the covalent attachment of the linker to B or
any D under physiological conditions, such as by having one or more
pH-labile, acid-labile, base-labile, oxidatively labile,
metabolically labile, biochemically labile, and/or enzyme-labile
bonds. It is appreciated that such physiological conditions
resulting in bond breaking do not necessarily include a biological
or metabolic process, and instead may include a standard chemical
reaction, such as a hydrolysis reaction, for example, at
physiological pH, or as a result of compartmentalization into a
cellular organelle such as an endosome having a lower pH than
cytosolic pH.
[0221] It is understood that a cleavable bond can connect two
adjacent atoms within the releasable linker, and/or connect other
linkers with B, and/or any D, as described herein, at any ends of
the releasable linker. In the case where a cleavable bond connects
two adjacent atoms within the releasable linker, following breakage
of the bond, the releasable linker is broken into two or more
fragments. Alternatively, in the case where a cleavable bond is
between the releasable linker and another moiety, such as an
additional heteroatom, a spacer linker, another releasable portion
of the linker, any D, or B, following breakage of the bond, the
releasable linker is separated from the other moiety. It is to be
understood that a linker is a releasable linker when if forms a
cleavable, scissile, or breakable bond with the one or more of the
drugs (D) is capable of delivery of the one or more drugs (D) in a
traceless manner, where the one or more drugs (D) do not include
any residual part of the conjugate.
[0222] Illustrative radicals that themselves include a cleavable
bond, or form a cleavable bond with B and/or any D hemiacetals and
sulfur variations thereof, acetals and sulfur variations thereof,
hemiaminals, aminals, and the like, or which can be formed from
methylene fragments substituted with at least one heteroatom, such
as 1-alkoxyalkylene, 1-alkoxycycloalkylene,
1-alkoxyalkylenecarbonyl, 1-alkoxycycloalkylenecarbonyl, and the
like. Illustrative releasable linkers described herein include
polyvalent linkers that include carbonylarylcarbonyl,
carbonyl(carboxyaryl)carbonyl, carbonyl(biscarboxyaryl)carbonyl,
haloalkylenecarbonyl, and the like. Illustrative releasable linkers
described herein include polyvalent linkers that include
alkylene(dialkylsilyl), alkylene(alkylarylsilyl),
alkylene(diarylsilyl), (dialkylsilyl)aryl, (alkylarylsilyl)aryl,
(diarylsilyl)aryl, and the like. Illustrative releasable linkers
described herein include oxycarbonyloxy, oxycarbonyloxyalkyl,
sulfonyloxy, oxysulfonylalkyl, and the like. Illustrative
releasable linkers described herein include polyvalent linkers that
include iminoalkylidenyl, carbonylalkylideniminyl,
iminocycloalkylidenyl, carbonylcycloalkylideniminyl, and the like.
Illustrative releasable linkers described herein include polyvalent
linkers that include alkylenethio, alkylenearylthio, and
carbonylalkylthio, and the like. Each of the foregoing fragments is
optionally substituted with a substituent X.sup.2, as defined
herein.
[0223] The substituents X.sup.2 can be alkyl, alkoxy, alkoxyalkyl,
hydroxy, hydroxyalkyl, amino, aminoalkyl, alkylaminoalkyl,
dialkylaminoalkyl, halo, haloalkyl, sulfhydrylalkyl,
alkylthioalkyl, aryl, substituted aryl, arylalkyl, substituted
arylalkyl, heteroaryl, substituted heteroaryl, carboxy,
carboxyalkyl, alkyl carboxylate, alkyl alkanoate, guanidinoalkyl,
R.sup.4-carbonyl, R.sup.5-carbonylalkyl, R.sup.6-acylamino, and
R.sup.7-acylaminoalkyl, wherein R.sup.4 and R.sup.5 are each
independently selected from amino acids, amino acid derivatives,
and peptides, and wherein R.sup.6 and R.sup.7 are each
independently selected from amino acids, amino acid derivatives,
and peptides. In this embodiment the heteroatom linker can be
nitrogen, and the substituent X.sup.2 and the heteroatom linker can
be taken together with the releasable linker to which they are
bound to form an heterocycle.
[0224] The heterocycles can be pyrrolidines, piperidines,
oxazolidines, isoxazolidines, thiazolidines, isothiazolidines,
pyrrolidinones, piperidinones, oxazolidinones, isoxazolidinones,
thiazolidinones, isothiazolidinones, and succinimides.
[0225] Illustrative releasable linkers include ketals, acetals,
hemiaminals, and aminals formed from methylene, 1-alkoxyalkylene,
1-alkoxycycloalkylene, 1-alkoxyalkylenecarbonyl, and
1-alkoxycycloalkylenecarbonyl radicals, esters and amides formed
from carbonylarylcarbonyl, carbonyl(carboxyaryl)carbonyl,
carbonyl(biscarboxyaryl)carbonyl, and haloalkylenecarbonyl
radicals, oxysilanes and aminosilanes formed from
alkylene(dialkylsilyl), alkylene(alkylarylsilyl),
alkylene(diarylsilyl), (dialkylsilyl)aryl, (alkylarylsilyl)aryl,
and (diarylsilyl)aryl radicals, oxycarbonyloxy,
oxycarbonyloxyalkyl, sulfonyloxy, oxysulfonylalkyl,
iminoalkylidenyl, carbonylalkylideniminyl, iminocycloalkylidenyl,
carbonylcycloalkylideniminyl, alkylenethio, alkylenearylthio, and
carbonylalkylthio radicals, each of which is optionally
substituted.
[0226] Further illustrative releasable linkers include hydrazones,
acylhydrazones orthoformates, and carbamoyl derivatives.
[0227] Further illustrative releasable linkers include disulfides
and activated thioethers.
[0228] In any of the embodiments described herein, the releasable
linker may include oxygen bonded to methylene, 1-alkoxyalkylene,
1-alkoxycycloalkylene, 1-alkoxyalkylenecarbonyl, and
1-alkoxycycloalkylenecarbonyl to form an acetal or ketal, wherein
each of the fragments is optionally substituted with a substituent
X.sup.2, as defined herein. Alternatively, the methylene or
alkylene is substituted with an optionally-substituted aryl.
[0229] In any of the embodiments described herein, the releasable
linker may include nitrogen bonded to methylene, 1-alkoxyalkylene,
1-alkoxycycloalkylene, 1-alkoxyalkylenecarbonyl, and
1-alkoxycycloalkylenecarbonyl to form a hemiamninal ether or
aminal, wherein each of the fragments is optionally substituted
with a substituent X.sup.2, as defined herein. Alternatively, the
methylene or alkylene is substituted with an optionally-substituted
aryl.
[0230] In any of the embodiments described herein, the releasable
linker may include oxygen bonded to sulfonylalkyl to form an
alkylsulfonate.
[0231] In any of the embodiments described herein, the releasable
linker may include nitrogen bonded to iminoalkylidenyl,
carbonylalkylideniminyl, iminocycloalkylidenyl, and
carbonylcycloalkylideniminyl to form an hydrazone, each of which is
optionally substituted with a substituent X.sup.2, as defined
herein. In an alternate configuration, the hydrazone may be
acylated with a carboxylic acid derivative, an orthoformate
derivative, or a carbamoyl derivative to form releasable linkers
containing various acylhydrazones.
[0232] In any of the embodiments described herein, the releasable
linker may include oxygen bonded to alkylene(dialkylsilyl),
alkylene(alkylarylsilyl), alkylene(diarylsilyl),
(dialkylsilyl)aryl, (alkylarylsilyl)aryl, and (diarylsilyl)aryl to
form a silanol, each of which is optionally substituted with a
substituent X.sup.2, as defined herein.
[0233] In any of the embodiments described herein, the releasable
linker may include nitrogen bonded to carbonylarylcarbonyl,
carbonyl(carboxyaryl)carbonyl, carbonyl(biscarboxyaryl)carbonyl to
form an amide, or alternatively an amide with a drug nitrogen.
[0234] In any of the embodiments described herein, the releasable
linker may include oxygen bonded to carbonylarylcarbonyl,
carbonyl(carboxyaryl)carbonyl, carbonyl(biscarboxyaryl)carbonyl to
form an ester, or alternatively an ester with drug oxygen.
[0235] It is to be understood that the bivalent spacer linkers may
be combined in any chemically relevant way, either directly or via
an intervening heteroatom to construct the releasable linkers
described herein. It is further understood that the nature of the
arrangement of spacer and heteroatom linkers defines where the
releasable linker will cleave in vivo. For example, two spacer
linkers that terminate in a sulfur atom when combined form a
disulfide, which is the cleavable bond in the releasable linker
formed thereby.
[0236] For example, in another embodiment, the polyvalent linker
comprises a 3-thiosuccinimid-1-ylalkyloxymethyloxy moiety, where
the methyl is optionally substituted with alkyl or substituted
aryl.
[0237] In another embodiment, the polyvalent linker comprises a
3-thiosuccinimid-1-ylalkylcarbonyl, where the carbonyl forms an
acylaziridine with the drug.
[0238] In another embodiment, the polyvalent linker comprises a
1-alkoxycycloalkylenoxy moiety.
[0239] In another embodiment, the polyvalent linker comprises an
alkyleneaminocarbonyl(dicarboxylarylene)carboxylate.
[0240] In another embodiment, the polyvalent linker comprises a
dithioalkylcarbonylhydrazide, where the hydrazide forms an
hydrazone with the drug.
[0241] In another embodiment, the polyvalent linker comprises a
3-thiosuccinimid-1-ylalkylcarbonylhydrazide, where the hydrazide
forms a hydrazone with the drug.
[0242] In another embodiment, the polyvalent linker comprises a
3-thioalkylsulfonylalkyl(disubstituted silyl)oxy, where the
disubstituted silyl is substituted with alkyl or optionally
substituted aryl.
[0243] In another embodiment, the polyvalent linker comprises a
plurality of spacer linkers selected from the group consisting of
the naturally occurring amino acids and stereoisomers thereof.
[0244] In another embodiment, the polyvalent linker comprises a
2-dithioalkyloxycarbonyl, where the carbonyl forms a carbonate with
the drug.
[0245] In another embodiment, the polyvalent linker comprises a
2-dithioarylalkyloxycarbonyl, where the carbonyl forms a carbonate
with the drug and the aryl is optionally substituted.
[0246] In another embodiment, the polyvalent linker comprises a
4-dithioarylalkyloxycarbonyl, where the carbonyl forms a carbonate
with the drug, and the aryl is optionally substituted.
[0247] In another embodiment, the polyvalent linker comprises a
3-thiosuccinimid-1-ylalkyloxyalkyloxyalkylidene, where the
alkylidene forms an hydrazone with the drug, each alkyl is
independently selected, and the oxyalkyloxy is optionally
substituted with alkyl or optionally substituted aryl.
[0248] In another embodiment, the polyvalent linker comprises a
2-dithioalkyloxycarbonylhydrazide.
[0249] In another embodiment, the polyvalent linker comprises a 2-
or 3-dithioalkylamino, where the amino forms a vinylogous amide
with the drug.
[0250] In another embodiment, the polyvalent linker comprises a
2-dithioalkylamino, where the amino forms a vinylogous amide with
the drug, and the alkyl is ethyl.
[0251] In another embodiment, the polyvalent linker comprises a 2-
or 3-dithioalkylaminocarbonyl, where the carbonyl forms a carbamate
with the drug.
[0252] In another embodiment, the polyvalent linker comprises a
2-dithioalkylaminocarbonyl, where the carbonyl forms a carbamate
with the drug. In another aspect, the alkyl is ethyl.
[0253] In another embodiment, the polyvalent linker comprises a
2-dithioalkyloxycarbonyl, where the carbonyl forms a carbamate with
the drug. In another aspect, the alkyl is ethyl.
[0254] In another embodiment, the polyvalent linker comprises a
2-dithioarylalkyloxycarbonyl, where the carbonyl forms a carbamate
or a carbamoylaziridine with the drug.
[0255] In another embodiment, the polyvalent linker comprises a
4-dithioarylalkyloxycarbonyl, where the carbonyl forms a carbamate
or a carbamoylaziridine with the drug.
[0256] In another embodiment, the polyvalent linkers described
herein comprise divalent radicals of the formulae
##STR00086##
where n is an integer selected from 1 to about 4; R.sup.a and
R.sup.b are each independently selected from the group consisting
of hydrogen and alkyl, including lower alkyl such as
C.sub.1-C.sub.4 alkyl that are optionally branched; or R.sup.a and
R.sup.b are taken together with the attached carbon atom to form a
carbocyclic ring; R is an optionally substituted alkyl group, an
optionally substituted acyl group, or a suitably selected nitrogen
protecting group; and (*) indicates points of attachment for the
drug, vitamin, imaging agent, diagnostic agent, other bivalent
linkers, or other parts of the conjugate.
[0257] In another embodiment, the polyvalent linkers described
herein comprise divalent radicals of the formulae
##STR00087##
where m is an integer selected from 1 to about 4; R is an
optionally substituted alkyl group, an optionally substituted acyl
group, or a suitably selected nitrogen protecting group; and (*)
indicates points of attachment for the drug, vitamin, imaging
agent, diagnostic agent, other bivalent linkers, or other parts of
the conjugate.
[0258] In another embodiment, the polyvalent linkers described
herein comprise divalent radicals of the formulae
##STR00088##
where m is an integer selected from 1 to about 4; R is an
optionally substituted alkyl group, an optionally substituted acyl
group, or a suitably selected nitrogen protecting group; and (*)
indicates points of attachment for the drug, vitamin, imaging
agent, diagnostic agent, other divalent linkers, or other parts of
the conjugate.
[0259] In another embodiment, the compounds described herein
comprise one or more radicals linkers of selected from the
formulae:
##STR00089##
wherein X is NH, O, or S.
[0260] In another embodiment, the polyvalent linkers herein
described comprise a radical having the formula:
##STR00090##
[0261] Another embodiment, the polyvalent linkers described herein
comprise a radical of having the formula:
##STR00091##
where X is an heteroatom, such as nitrogen, oxygen, or sulfur, n is
an integer selected from 0, 1, 2, and 3, R is hydrogen, or a
substituent, including a substituent capable of stabilizing a
positive charge inductively or by resonance on the aryl ring, such
as alkoxy, and the like, and the symbol (*) indicates points of
attachment. It is appreciated that other substituents may be
present on the aryl ring, the benzyl carbon, the alkanoic acid, or
the methylene bridge, including but not limited to hydroxy, alkyl,
alkoxy, alkylthio, halo, and the like.
[0262] In another embodiment, the polyvalent linkers described
herein comprise radicalsf selected from carbonyl, thionocarbonyl,
alkylene, cycloalkylene, alkylenecycloalkyl, alkylenecarbonyl,
cycloalkylenecarbonyl, carbonylalkylcarbonyl, 1
alkylenesuccinimid-3-yl, 1 (carbonylalkyl)succinimid-3-yl,
alkylenesulfoxyl, sulfonylalkyl, alkylenesulfoxylalkyl,
alkylenesulfonylalkyl, carbonyltetrahydro-2H-pyranyl,
carbonyltetrahydrofuranyl,
1-(carbonyltetrahydro-2H-pyranyl)succinimid-3-yl, and
1-(carbonyltetrahydrofuranyl)succinimid-3-yl, wherein each of said
spacer linkers is optionally substituted with one or more
substituents X.sup.1;
[0263] wherein each substituent X.sup.1 is independently selected
from the group consisting of alkyl, alkoxy, alkoxyalkyl, hydroxy,
hydroxyalkyl, amino, aminoalkyl, alkylaminoalkyl,
dialkylaminoalkyl, halo, haloalkyl, sulfhydrylalkyl,
alkylthioalkyl, aryl, substituted aryl, arylalkyl, substituted
arylalkyl, heteroaryl, substituted heteroaryl, carboxy,
carboxyalkyl, alkyl carboxylate, alkyl alkanoate, guanidinoalkyl,
R.sup.4-carbonyl, R.sup.5-carbonylalkyl, R.sup.6-acylamino, and
R.sup.7-acylaminoalkyl, wherein R.sup.4 and R.sup.5 are each
independently selected from the group consisting of an amino acid,
an amino acid derivative, and a peptide, and wherein R.sup.6 and
R.sup.7 are each independently selected from the group consisting
of an amino acid, an amino acid derivative, and a peptide.
[0264] It is to be understood that the compounds described herein
may contain one or more chiral centers, or may otherwise be capable
of existing as multiple stereoisomers. It is to be understood that
in one embodiment, the invention described herein is not limited to
any particular stereochemical requirement, and that the compounds,
and compositions, methods, uses, and medicaments that include them
may be optically pure, or may be any of a variety of stereoisomeric
mixtures, including racemic and other mixtures of enantiomers,
other mixtures of diastereomers, and the like. It is also to be
understood that such mixtures of stereoisomers may include a single
stereochemical configuration at one or more chiral centers, while
including mixtures of stereochemical configuration at one or more
other chiral centers.
[0265] Similarly, the compounds described herein may include
geometric centers, such as cis, trans, E, and Z double bonds. It is
to be understood that in another embodiment, the invention
described herein is not limited to any particular geometric isomer
requirement, and that the compounds, and compositions, methods,
uses, and medicaments that include them may be pure, or may be any
of a variety of geometric isomer mixtures. It is also to be
understood that such mixtures of geometric isomers may include a
single configuration at one or more double bonds, while including
mixtures of geometry at one or more other double bonds.
[0266] In each of the foregoing and each of the following
embodiments, it is also to be understood that the formulae include
and represent not only all pharmaceutically acceptable salts of the
compounds, but also include any and all hydrates and/or solvates of
the compound formulae. It is appreciated that certain functional
groups, such as the hydroxy, amino, and like groups form complexes
and/or coordination compounds with water and/or various solvents,
in the various physical forms of the compounds. Accordingly, the
above formulae are to be understood to be a description of such
hydrates and/or solvates, including pharmaceutically acceptable
solvates.
[0267] In each of the foregoing and each of the following
embodiments, it is also to be understood that the formulae include
and represent each possible isomer, such as stereoisomers and
geometric isomers, both individually and in any and all possible
mixtures. In each of the foregoing and each of the following
embodiments, it is also to be understood that the formulae include
and represent any and all crystalline forms, partially crystalline
forms, and non crystalline and/or amorphous forms, and co-crystals
of the compounds.
[0268] In another embodiment, the compounds described herein can be
internalized into the targeted pathogenic cells by binding to PSMA.
In particular, PSMA selectively and/or specifically binds the
conjugate, and internalization can occur, for example, through
PSMA-mediated endocytosis. Once internalized, conjugates containing
a releasable linker can complete delivery of the drug to the
interior of the target cell. Without being bound by theory, it is
believed herein that in those cases where the drug is toxic to
normal cells or tissues, such a delivery system can decrease
toxicity against those non-target cells and tissues because the
releasable linker remains substantially or completely intact until
the compounds described herein are delivered to the target cells.
Accordingly, the compounds described herein act intracellularly by
delivering the drug to an intracellular biochemical process, which
in turn decreases the amount of unconjugated drug exposure to the
host animal's healthy cells and tissues.
[0269] The conjugates described herein can be used for both human
clinical medicine and veterinary applications. Thus, the host
animal harboring the population of pathogenic cells and treated
with the compounds described herein can be human or, in the case of
veterinary applications, can be a laboratory, agricultural,
domestic, or wild animal. The present invention can be applied to
host animals including, but not limited to, humans, laboratory
animals such rodents (e.g., mice, rats, hamsters, etc.), rabbits,
monkeys, chimpanzees, domestic animals such as dogs, cats, and
rabbits, agricultural animals such as cows, horses, pigs, sheep,
goats, and wild animals in captivity such as bears, pandas, lions,
tigers, leopards, elephants, zebras, giraffes, gorillas, dolphins,
and whales.
[0270] The drug delivery conjugate compounds described herein can
be administered in a combination therapy with any other known drug
whether or not the additional drug is targeted. Illustrative
additional drugs include, but are not limited to, peptides,
oligopeptides, retro-inverso oligopeptides, proteins, protein
analogs in which at least one non-peptide linkage replaces a
peptide linkage, apoproteins, glycoproteins, enzymes, coenzymes,
enzyme inhibitors, amino acids and their derivatives, receptors and
other membrane proteins, antigens and antibodies thereto, haptens
and antibodies thereto, hormones, lipids, phospholipids, liposomes,
toxins, antibiotics, analgesics, bronchodilators, beta-blockers,
antimicrobial agents, antihypertensive agents, cardiovascular
agents including antiarrhythmics, cardiac glycosides, antianginals,
vasodilators, central nervous system agents including stimulants,
psychotropics, antimanics, and depressants, antiviral agents,
antihistamines, cancer drugs including chemotherapeutic agents,
tranquilizers, anti-depressants, H-2 antagonists, anticonvulsants,
antinauseants, prostaglandins and prostaglandin analogs, muscle
relaxants, anti-inflammatory substances, stimulants, decongestants,
antiemetics, diuretics, antispasmodics, antiasthmatics,
anti-Parkinson agents, expectorants, cough suppressants,
mucolytics, and mineral and nutritional additives.
[0271] As used herein, the term "alkyl" includes a chain of carbon
atoms, which is optionally branched. As used herein, the term
"alkenyl" and "alkynyl" includes a chain of carbon atoms, which is
optionally branched, and includes at least one double bond or
triple bond, respectively. It is to be understood that alkynyl may
also include one or more double bonds. It is to be further
understood that in certain embodiments, alkyl is advantageously of
limited length, including C.sub.1-C.sub.24, C.sub.1-C.sub.12,
C.sub.1-C.sub.8, C.sub.1-C.sub.6, and C.sub.1-C.sub.4, and
C.sub.2-C.sub.24, C.sub.2-C.sub.12, C.sub.2-C.sub.8,
C.sub.2-C.sub.6, and C.sub.2-C.sub.4, and the like Illustratively,
such particularly limited length alkyl groups, including
C.sub.1-C.sub.8, C.sub.1-C.sub.6, and C.sub.1-C.sub.4, and
C.sub.2-C.sub.8, C.sub.2-C.sub.6, and C.sub.2-C.sub.4, and the like
may be referred to as lower alkyl. It is to be further understood
that in certain embodiments alkenyl and/or alkynyl may each be
advantageously of limited length, including C.sub.2-C.sub.24,
C.sub.2-C.sub.12, C.sub.2-C.sub.8, C.sub.2-C.sub.6, and
C.sub.2-C.sub.4, and C.sub.3-C.sub.24, C.sub.3-C.sub.12,
C.sub.3-C.sub.8, C.sub.3-C.sub.6, and C.sub.3-C.sub.4, and the
like. Illustratively, such particularly limited length alkenyl
and/or alkynyl groups, including C.sub.2-C.sub.8, C.sub.2-C.sub.6,
and C.sub.2-C.sub.4, and C.sub.3-C.sub.8, C.sub.3-C.sub.6, and
C.sub.3-C.sub.4, and the like may be referred to as lower alkenyl
and/or alkynyl. It is appreciated herein that shorter alkyl,
alkenyl, and/or alkynyl groups may add less lipophilicity to the
compound and accordingly will have different pharmacokinetic
behavior. In embodiments of the invention described herein, it is
to be understood, in each case, that the recitation of alkyl refers
to alkyl as defined herein, and optionally lower alkyl. In
embodiments of the invention described herein, it is to be
understood, in each case, that the recitation of alkenyl refers to
alkenyl as defined herein, and optionally lower alkenyl. In
embodiments of the invention described herein, it is to be
understood, in each case, that the recitation of alkynyl refers to
alkynyl as defined herein, and optionally lower alkynyl.
Illustrative alkyl, alkenyl, and alkynyl groups are, but not
limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl,
sec-butyl, tert-butyl, pentyl, 2-pentyl, 3-pentyl, neopentyl,
hexyl, heptyl, octyl, and the like, and the corresponding groups
containing one or more double and/or triple bonds, or a combination
thereof.
[0272] As used herein, the term "alkylene" includes a divalent
chain of carbon atoms, which is optionally branched. As used
herein, the term "alkenylene" and "alkynylene" includes a divalent
chain of carbon atoms, which is optionally branched, and includes
at least one double bond or triple bond, respectively. It is to be
understood that alkynylene may also include one or more double
bonds. It is to be further understood that in certain embodiments,
alkylene is advantageously of limited length, including
C.sub.1-C.sub.24, C.sub.1-C.sub.12, C.sub.1-C.sub.8,
C.sub.1-C.sub.6, and C.sub.1-C.sub.4, and C.sub.2-C.sub.24,
C.sub.2-C.sub.12, C.sub.2-C.sub.8, C.sub.2-C.sub.6, and
C.sub.2-C.sub.4, and the like. Illustratively, such particularly
limited length alkylene groups, including C.sub.1-C.sub.8,
C.sub.1-C.sub.6, and C.sub.1-C.sub.4, and C.sub.2-C.sub.8,
C.sub.2-C.sub.6, and C.sub.2-C.sub.4, and the like may be referred
to as lower alkylene. It is to be further understood that in
certain embodiments alkenylene and/or alkynylene may each be
advantageously of limited length, including C.sub.2-C.sub.24,
C.sub.2-C.sub.12, C.sub.1-C.sub.8, C.sub.2-C.sub.6, and
C.sub.2-C.sub.4, and C.sub.3-C.sub.24, C.sub.3-C.sub.12,
C.sub.3-C.sub.8, C.sub.3-C.sub.6, and C.sub.3-C.sub.4, and the
like. Illustratively, such particularly limited length alkenylene
and/or alkynylene groups, including C.sub.2-C.sub.8,
C.sub.2-C.sub.6, and C.sub.2-C.sub.4, and C.sub.3-C.sub.8,
C.sub.3-C.sub.6, and C.sub.3-C.sub.4, and the like may be referred
to as lower alkenylene and/or alkynylene. It is appreciated herein
that shorter alkylene, alkenylene, and/or alkynylene groups may add
less lipophilicity to the compound and accordingly will have
different pharmacokinetic behavior. In embodiments of the invention
described herein, it is to be understood, in each case, that the
recitation of alkylene, alkenylene, and alkynylene refers to
alkylene, alkenylene, and alkynylene as defined herein, and
optionally lower alkylene, alkenylene, and alkynylene. Illustrative
alkyl groups are, but not limited to, methylene, ethylene,
n-propylene, isopropylene, n-butylene, isobutylene, sec-butylene,
pentylene, 1,2-pentylene, 1,3-pentylene, hexylene, heptylene,
octylene, and the like.
[0273] As used herein, the term "cycloalkyl" includes a chain of
carbon atoms, which is optionally branched, where at least a
portion of the chain in cyclic. It is to be understood that
cycloalkylalkyl is a subset of cycloalkyl. It is to be understood
that cycloalkyl may be polycyclic. Illustrative cycloalkyl include,
but are not limited to, cyclopropyl, cyclopentyl, cyclohexyl,
2-methylcyclopropyl, cyclopentyleth-2-yl, adamantyl, and the like.
As used herein, the term "cycloalkenyl" includes a chain of carbon
atoms, which is optionally branched, and includes at least one
double bond, where at least a portion of the chain in cyclic. It is
to be understood that the one or more double bonds may be in the
cyclic portion of cycloalkenyl and/or the non-cyclic portion of
cycloalkenyl. It is to be understood that cycloalkenylalkyl and
cycloalkylalkenyl are each subsets of cycloalkenyl. It is to be
understood that cycloalkyl may be polycyclic. Illustrative
cycloalkenyl include, but are not limited to, cyclopentenyl,
cyclohexylethen-2-yl, cycloheptenylpropenyl, and the like. It is to
be further understood that chain forming cycloalkyl and/or
cycloalkenyl is advantageously of limited length, including
C.sub.3-C.sub.24, C.sub.3-C.sub.12, C.sub.3-C.sub.8,
C.sub.3-C.sub.6, and C.sub.5-C.sub.6. It is appreciated herein that
shorter alkyl and/or alkenyl chains forming cycloalkyl and/or
cycloalkenyl, respectively, may add less lipophilicity to the
compound and accordingly will have different pharmacokinetic
behavior.
[0274] As used herein, the term "heteroalkyl" includes a chain of
atoms that includes both carbon and at least one heteroatom, and is
optionally branched. Illustrative heteroatoms include nitrogen,
oxygen, and sulfur. In certain variations, illustrative heteroatoms
also include phosphorus, and selenium. As used herein, the term
"cycloheteroalkyl" including heterocyclyl and heterocycle, includes
a chain of atoms that includes both carbon and at least one
heteroatom, such as heteroalkyl, and is optionally branched, where
at least a portion of the chain is cyclic. Illustrative heteroatoms
include nitrogen, oxygen, and sulfur. In certain variations,
illustrative heteroatoms also include phosphorus, and selenium.
Illustrative cycloheteroalkyl include, but are not limited to,
tetrahydrofuryl, pyrrolidinyl, tetrahydropyranyl, piperidinyl,
morpholinyl, piperazinyl, homopiperazinyl, quinuclidinyl, and the
like.
[0275] As used herein, the term "aryl" includes monocyclic and
polycyclic aromatic carbocyclic groups, each of which may be
optionally substituted. Illustrative aromatic carbocyclic groups
described herein include, but are not limited to, phenyl, naphthyl,
and the like. As used herein, the term "heteroaryl" includes
aromatic heterocyclic groups, each of which may be optionally
substituted. Illustrative aromatic heterocyclic groups include, but
are not limited to, pyridinyl, pyrimidinyl, pyrazinyl, triazinyl,
tetrazinyl, quinolinyl, quinazolinyl, quinoxalinyl, thienyl,
pyrazolyl, imidazolyl, oxazolyl, thiazolyl, isoxazolyl,
isothiazolyl, oxadiazolyl, thiadiazolyl, triazolyl, benzimidazolyl,
benzoxazolyl, benzthiazolyl, benzisoxazolyl, benzisothiazolyl, and
the like.
[0276] As used herein, the term "amino" includes the group
NH.sub.2, alkylamino, and dialkylamino, where the two alkyl groups
in dialkylamino may be the same or different, i.e. alkylalkylamino.
Illustratively, amino includes methylamino, ethylamino,
dimethylamino, methylethylamino, and the like. In addition, it is
to be understood that when amino modifies or is modified by another
term, such as aminoalkyl, or acylamino, the above variations of the
term amino are included therein. Illustratively, aminoalkyl
includes H.sub.2N-alkyl, methylaminoalkyl, ethylaminoalkyl,
dimethylaminoalkyl, methylethylaminoalkyl, and the like.
Illustratively, acylamino includes acylmethylamino, acylethylamino,
and the like.
[0277] As used herein, the term "amino and derivatives thereof"
includes amino as described herein, and alkylamino, alkenylamino,
alkynylamino, heteroalkylamino, heteroalkenylamino,
heteroalkynylamino, cycloalkylamino, cycloalkenylamino,
cycloheteroalkylamino, cycloheteroalkenylamino, arylamino,
arylalkylamino, arylalkenylamino, arylalkynylamino,
heteroarylamino, heteroarylalkylamino, heteroarylalkenylamino,
heteroarylalkynylamino, acylamino, and the like, each of which is
optionally substituted. The term "amino derivative" also includes
urea, carbamate, and the like.
[0278] As used herein, the term "amino acid" refers generally to
beta, gamma, and longer amino acids, such as amino acids of the
formula:
--N(R)--(CR'R'').sub.q--C(O)--
where R is hydrogen, alkyl, acyl, or a suitable nitrogen protecting
group, R' and R'' are hydrogen or a substituent, each of which is
independently selected in each occurrence, and q is an integer such
as 1, 2, 3, 4, or 5. Illustratively, R' and/or R'' independently
correspond to, but are not limited to, hydrogen or the side chains
present on naturally occurring amino acids, such as methyl, benzyl,
hydroxymethyl, thiomethyl, carboxyl, carboxylmethyl,
guanidinopropyl, and the like, and derivatives and protected
derivatives thereof. The above described formula includes all
stereoisomeric variations. For example, the amino acid may be
selected from asparagine, aspartic acid, cysteine, glutamic acid,
lysine, glutamine, arginine, serine, ornitine, threonine, and the
like.
[0279] As used herein, the term "amino acid derivative" generally
refers to an amino acid as defined herein where either, or both,
the amino group and/or the side chain is substituted. Illustrative
amino acid derivatives include prodrugs and protecting groups of
the amino group and/or the side chain, such as amine, amide,
hydroxy, carboxylic acid, and thio prodrugs and protecting groups.
Additional Illustrative amino acid derivatives include substituted
variations of the amino acid as described herein, such as, but not
limited to, ethers and esters of hydroxy groups, amides,
carbamates, and ureas of amino groups, esters, amides, and cyano
derivatives of carboxylic acid groups, and the like.
[0280] As used herein, the term "hydroxy and derivatives thereof"
includes OH, and alkyloxy, alkenyloxy, alkynyloxy, heteroalkyloxy,
heteroalkenyloxy, heteroalkynyloxy, cycloalkyloxy, cycloalkenyloxy,
cycloheteroalkyloxy, cycloheteroalkenyloxy, aryloxy, arylalkyloxy,
arylalkenyloxy, arylalkynyloxy, heteroaryloxy, heteroarylalkyloxy,
heteroarylalkenyloxy, heteroarylalkynyloxy, acyloxy, and the like,
each of which is optionally substituted. The term "hydroxy
derivative" also includes carbamate, and the like.
[0281] As used herein, the term "thio and derivatives thereof"
includes SH, and alkylthio, alkenylthio, alkynylthio,
heteroalkylthio, heteroalkenylthio, heteroalkynylthio,
cycloalkylthio, cycloalkenylthio, cycloheteroalkylthio,
cycloheteroalkenylthio, arylthio, arylalkylthio, arylalkenylthio,
arylalkynylthio, heteroarylthio, heteroarylalkylthio,
heteroarylalkenylthio, heteroarylalkynylthio, acylthio, and the
like, each of which is optionally substituted. The term "thio
derivative" also includes thiocarbamate, and the like.
[0282] As used herein, the term "acyl" includes formyl, and
alkylcarbonyl, alkenylcarbonyl, alkynylcarbonyl,
heteroalkylcarbonyl, heteroalkenylcarbonyl, heteroalkynylcarbonyl,
cycloalkylcarbonyl, cycloalkenylcarbonyl, cycloheteroalkylcarbonyl,
cycloheteroalkenylcarbonyl, arylcarbonyl, arylalkylcarbonyl,
arylalkenylcarbonyl, arylalkynylcarbonyl, heteroarylcarbonyl,
heteroarylalkylcarbonyl, heteroarylalkenylcarbonyl,
heteroarylalkynylcarbonyl, acylcarbonyl, and the like, each of
which is optionally substituted.
[0283] As used herein, the term "carbonyl and derivatives thereof"
includes the group C(O), C(S). C(NH) and substituted amino
derivatives thereof.
[0284] As used herein, the term "carboxylic acid and derivatives
thereof" includes the group CO.sub.2H and salts thereof, and esters
and amides thereof, and CN.
[0285] As used herein, the term "sulfinic acid or a derivative
thereof" includes SO.sub.2H and salts thereof, and esters and
amides thereof.
[0286] As used herein, the term "sulfonic acid or a derivative
thereof" includes SO.sub.3H and salts thereof, and esters and
amides thereof.
[0287] As used herein, the term "sulfonyl" includes alkylsulfonyl,
alkenylsulfonyl, alkynylsulfonyl, heteroalkylsulfonyl,
heteroalkenylsulfonyl, heteroalkynylsulfonyl, cycloalkylsulfonyl,
cycloalkenylsulfonyl, cycloheteroalkylsulfonyl,
cycloheteroalkenylsulfonyl, arylsulfonyl, arylalkylsulfonyl,
arylalkenylsulfonyl, arylalkynylsulfonyl, heteroarylsulfonyl,
heteroarylalkylsulfonyl, heteroarylalkenylsulfonyl,
heteroarylalkynylsulfonyl, acylsulfonyl, and the like, each of
which is optionally substituted.
[0288] As used herein, the term "phosphinic acid or a derivative
thereof" includes P(R)O.sub.2H and salts thereof, and esters and
amides thereof, where R is alkyl, alkenyl, alkynyl, cycloalkyl,
cycloalkenyl, heteroalkyl, heteroalkenyl, cycloheteroalkyl,
cycloheteroalkenyl, aryl, heteroaryl, arylalkyl, or
heteroarylalkyl, each of which is optionally substituted.
[0289] As used herein, the term "phosphonic acid or a derivative
thereof" includes PO.sub.3H.sub.2 and salts thereof, and esters and
amides thereof.
[0290] As used herein, the term "hydroxylamino and derivatives
thereof" includes NHOH, and alkyloxylNH alkenyloxylNH alkynyloxylNH
heteroalkyloxylNH heteroalkenyloxylNH heteroalkynyloxylNH
cycloalkyloxylNH cycloalkenyloxylNH cycloheteroalkyloxylNH
cycloheteroalkenyloxylNH aryloxylNH arylalkyloxylNH
arylalkenyloxylNH arylalkynyloxylNH heteroaryloxylNH
heteroarylalkyloxylNH heteroarylalkenyloxylNH
heteroarylalkynyloxylNH acyloxy, and the like, each of which is
optionally substituted.
[0291] As used herein, the term "hydrazino and derivatives thereof"
includes alkylNHNH, alkenylNHNH, alkynylNHNH, heteroalkylNHNH,
heteroalkenylNHNH, heteroalkynylNHNH, cycloalkylNHNH,
cycloalkenylNHNH, cycloheteroalkylNHNH, cycloheteroalkenylNHNH,
arylNHNH, arylalkylNHNH, arylalkenylNHNH, arylalkynylNHNH,
heteroarylNHNH, heteroarylalkylNHNH, heteroarylalkenylNHNH,
heteroarylalkynylNHNH, acylNHNH, and the like, each of which is
optionally substituted.
[0292] The term "optionally substituted" as used herein includes
the replacement of hydrogen atoms with other functional groups on
the radical that is optionally substituted. Such other functional
groups illustratively include, but are not limited to, amino,
hydroxyl, halo, thiol, alkyl, haloalkyl, heteroalkyl, aryl,
arylalkyl, arylheteroalkyl, heteroaryl, heteroarylalkyl,
heteroarylheteroalkyl, nitro, sulfonic acids and derivatives
thereof, carboxylic acids and derivatives thereof, and the like.
Illustratively, any of amino, hydroxyl, thiol, alkyl, haloalkyl,
heteroalkyl, aryl, arylalkyl, arylheteroalkyl, heteroaryl,
heteroarylalkyl, heteroarylheteroalkyl, and/or sulfonic acid is
also optionally substituted.
[0293] As used herein, the terms "optionally substituted aryl" and
"optionally substituted heteroaryl" include the replacement of
hydrogen atoms with other functional groups on the aryl or
heteroaryl that is optionally substituted. Such other functional
groups illustratively include, but are not limited to, amino,
hydroxy, halo, thio, alkyl, haloalkyl, heteroalkyl, aryl,
arylalkyl, arylheteroalkyl, heteroaryl, heteroarylalkyl,
heteroarylheteroalkyl, nitro, sulfonic acids and derivatives
thereof, carboxylic acids and derivatives thereof, and the like.
Illustratively, any of amino, hydroxy, thio, alkyl, haloalkyl,
heteroalkyl, aryl, arylalkyl, arylheteroalkyl, heteroaryl,
heteroarylalkyl, heteroarylheteroalkyl, and/or sulfonic acid is
optionally substituted.
[0294] Illustrative substituents include, but are not limited to, a
radical --(CH.sub.2).sub.xZ.sup.X, where x is an integer from 0-6
and Z.sup.X is selected from halogen, hydroxy, alkanoyloxy,
including C.sub.1-C.sub.6 alkanoyloxy, optionally substituted
aroyloxy, alkyl, including C.sub.1-C.sub.6 alkyl, alkoxy, including
C.sub.1-C.sub.6 alkoxy, cycloalkyl, including C.sub.3-C.sub.8
cycloalkyl, cycloalkoxy, including C.sub.3-C.sub.8 cycloalkoxy,
alkenyl, including C.sub.2-C.sub.6 alkenyl, alkynyl, including
C.sub.2-C.sub.6 alkynyl, haloalkyl, including C.sub.1-C.sub.6
haloalkyl, haloalkoxy, including C.sub.1-C.sub.6 haloalkoxy,
halocycloalkyl, including C.sub.3-C.sub.8 halocycloalkyl,
halocycloalkoxy, including C.sub.3-C.sub.8 halocycloalkoxy, amino.
C.sub.1-C.sub.6 alkylamino, (C.sub.1-C.sub.6 alkyl)(C.sub.1-C.sub.6
alkyl)amino, alkylcarbonylamino, N--(C.sub.1-C.sub.6
alkyl)alkylcarbonylamino, aminoalkyl, C.sub.1-C.sub.6
alkylaminoalkyl, (C.sub.1-C.sub.6 alkyl)(C.sub.1-C.sub.6
alkyl)aminoalkyl, alkylcarbonylaminoalkyl, N--(C.sub.1-C.sub.6
alkyl)alkylcarbonylaminoalkyl, cyano, and nitro; or Z.sup.X is
selected from --CO.sub.2R.sup.4 and --CONR.sup.5R.sup.6, where
R.sup.4, R.sup.5, and R.sup.6 are each independently selected in
each occurrence from hydrogen, C.sub.1-C.sub.6 alkyl,
aryl-C.sub.1-C.sub.6 alkyl, and heteroaryl-C.sub.1-C.sub.6
alkyl.
[0295] As used herein, the term "leaving group" refers to a
reactive functional group that generates an electrophilic site on
the atom to which it is attached such that nucleophiles may be
added to the electrophilic site on the atom. Illustrative leaving
groups include, but are not limited to, halogens, optionally
substituted phenols, acyloxy groups, sulfonoxy groups, and the
like. It is to be understood that such leaving groups may be on
alkyl, acyl, and the like. Such leaving groups may also be referred
to herein as activating groups, such as when the leaving group is
present on acyl. In addition, conventional peptide, amide, and
ester coupling agents, such as but not limited to PyBop. BOP-Cl,
BOP, pentafluorophenol, isobutylchloroformate, and the like, form
various intermediates that include a leaving group, as defined
herein, on a carbonyl group.
[0296] As used herein the term "radical" with reference to, for
example, the PSMA binding or targeting ligand, and/or the
independently selected drug, refers to a PSMA binding or targeting
ligand, and/or an independently selected drug, as described herein,
where one or more atoms or groups, such as a hydrogen atom, or an
alkyl group on a heteroatom, and the like, is removed to provide a
radical for conjugation to the polyvalent linker L.
[0297] The term "prodrug" as used herein generally refers to any
compound that when administered to a biological system generates a
biologically active compound as a result of one or more spontaneous
chemical reaction(s), enzyme-catalyzed chemical reaction(s), and/or
metabolic chemical reaction(s), or a combination thereof. In vivo,
the prodrug is typically acted upon by an enzyme (such as
esterases, amidases, phosphatases, and the like), simple biological
chemistry, or other process in vivo to liberate or regenerate the
more pharmacologically active drug. This activation may occur
through the action of an endogenous host enzyme or a non-endogenous
enzyme that is administered to the host preceding, following, or
during administration of the prodrug. Additional details of prodrug
use are described in U.S. Pat. No. 5,627,165; and Pathalk et al.,
Enzymic protecting group techniques in organic synthesis,
Stereosel. Biocatal. 775-797 (2000). It is appreciated that the
prodrug is advantageously converted to the original drug as soon as
the goal, such as targeted delivery, safety, stability, and the
like is achieved, followed by the subsequent rapid elimination of
the released remains of the group forming the prodrug.
[0298] Prodrugs may be prepared from the compounds described herein
by attaching groups that ultimately cleave in vivo to one or more
functional groups present on the compound, such as --OH--, --SH,
--CO.sub.2H, --NR.sub.2. Illustrative prodrugs include but are not
limited to carboxylate esters where the group is alkyl, aryl,
arylalkyl, heteroaryl, heteroarylalkyl, acyloxyalkyl,
alkoxycarbonyloxyalkyl as well as esters of hydroxyl, thiol and
amines where the group attached is an acyl group, an
alkoxycarbonyl, aminocarbonyl, phosphate or sulfate. Illustrative
esters, also referred to as active esters, include but are not
limited to 1-indanyl, N-oxysuccinimide; acyloxyalkyl groups such as
acetoxymethyl, pivaloyloxymethyl, .beta.-acetoxyethyl,
.beta.-pivaloyloxyethyl, 1-(cyclohexylcarbonyloxy)prop-1-yl,
(1-aminoethyl)carbonyloxymethyl, and the like;
alkoxycarbonyloxyalkyl groups, such as ethoxycarbonyloxymethyl,
.alpha.-ethoxycarbonyloxyethyl, .beta.-ethoxycarbonyloxyethyl, and
the like; dialkylaminoalkyl groups, including di-lower alkylamino
alkyl groups, such as dimethylaminomethyl, dimethylaminoethyl,
diethylaminomethyl, diethylaminoethyl, and the like;
2-(alkoxycarbonyl)-2-alkenyl groups such as 2-(isobutoxycarbonyl)
pent-2-enyl, 2-(ethoxycarbonyl)but-2-enyl, and the like; and
lactone groups such as phthalidyl, dimethoxyphthalidyl, and the
like.
[0299] Further illustrative prodrugs contain a chemical moiety,
such as an amide or phosphorus group functioning to increase
solubility and/or stability of the compounds described herein.
Further illustrative prodrugs for amino groups include, but are not
limited to, (C.sub.3-C.sub.20)alkanoyl;
halo-(C.sub.3-C.sub.20)alkanoyl; (C.sub.3-C.sub.20)alkenoyl;
(C.sub.4-C.sub.7)cycloalkanoyl;
(C.sub.3-C.sub.6)-cycloalkyl(C.sub.2-C.sub.16)alkanoyl; optionally
substituted aroyl, such as unsubstituted aroyl or aroyl substituted
by 1 to 3 substituents selected from the group consisting of
halogen, cyano, trifluoromethanesulphonyloxy,
(C.sub.1-C.sub.3)alkyl and (C.sub.1-C.sub.3)alkoxy, each of which
is optionally further substituted with one or more of 1 to 3
halogen atoms; optionally substituted
aryl(C.sub.2-C.sub.16)alkanoyl and optionally substituted
heteroaryl(C.sub.2-C.sub.16)alkanoyl, such as the aryl or
heteroaryl radical being unsubstituted or substituted by 1 to 3
substituents selected from the group consisting of halogen,
(C.sub.1-C.sub.3)alkyl and (C.sub.1-C.sub.3)alkoxy, each of which
is optionally further substituted with 1 to 3 halogen atoms; and
optionally substituted heteroarylalkanoyl having one to three
heteroatoms selected from O, S and N in the heteroaryl moiety and 2
to 10 carbon atoms in the alkanoyl moiety, such as the heteroaryl
radical being unsubstituted or substituted by 1 to 3 substituents
selected from the group consisting of halogen, cyano,
trifluoromethanesulphonyloxy, (C.sub.1-C.sub.3)alkyl, and
(C.sub.1-C.sub.3)alkoxy, each of which is optionally further
substituted with 1 to 3 halogen atoms. The groups illustrated are
exemplary, not exhaustive, and may be prepared by conventional
processes.
[0300] It is understood that the prodrugs themselves may not
possess significant biological activity, but instead undergo one or
more spontaneous chemical reaction(s), enzyme-catalyzed chemical
reaction(s), and/or metabolic chemical reaction(s), or a
combination thereof after administration in vivo to produce the
compound described herein that is biologically active or is a
precursor of the biologically active compound. However, it is
appreciated that in some cases, the prodrug is biologically active.
It is also appreciated that prodrugs may often serves to improve
drug efficacy or safety through improved oral bioavailability,
pharmacodynamic half-life, and the like. Prodrugs also refer to
derivatives of the compounds described herein that include groups
that simply mask undesirable drug properties or improve drug
delivery. For example, one or more compounds described herein may
exhibit an undesirable property that is advantageously blocked or
minimized may become pharmacological, pharmaceutical, or
pharmacokinetic barriers in clinical drug application, such as low
oral drug absorption, lack of site specificity, chemical
instability, toxicity, and poor patient acceptance (bad taste,
odor, pain at injection site, and the like), and others. It is
appreciated herein that a prodrug, or other strategy using
reversible derivatives, can be useful in the optimization of the
clinical application of a drug.
[0301] It is to be understood that in every instance disclosed
herein, the recitation of a range of integers for any variable
describes the recited range, every individual member in the range,
and every possible subrange for that variable. For example, the
recitation that n is an integer from 0 to 8, describes that range,
the individual and selectable values of 0, 1, 2, 3, 4, 5, 6, 7, and
8, such as n is 0, or n is 1, or n is 2, etc. In addition, the
recitation that n is an integer from 0 to 8 also describes each and
every subrange, each of which may for the basis of a further
embodiment, such as n is an integer from 1 to 8, from 1 to 7, from
1 to 6, from 2 to 8, from 2 to 7, from 1 to 3, from 2 to 4,
etc.
[0302] As used herein, the term "composition" generally refers to
any product comprising the specified ingredients in the specified
amounts, as well as any product which results, directly or
indirectly, from combinations of the specified ingredients in the
specified amounts. It is to be understood that the compositions
described herein may be prepared from isolated compounds described
herein or from salts, solutions, hydrates, solvates, and other
forms of the compounds described herein. It is also to be
understood that the compositions may be prepared from various
amorphous, non-amorphous, partially crystalline, crystalline,
and/or other morphological forms of the compounds described herein.
It is also to be understood that the compositions may be prepared
from various hydrates and/or solvates of the compounds described
herein. Accordingly, such pharmaceutical compositions that recite
compounds described herein are to be understood to include each of,
or any combination of, the various morphological forms and/or
solvate or hydrate forms of the compounds described herein. In
addition, it is to be understood that the compositions may be
prepared from various co-crystals of the compounds described
herein.
[0303] Illustratively, compositions may include one or more
carriers, diluents, and/or excipients. The compounds described
herein, or compositions containing them, may be formulated in a
therapeutically effective amount in any conventional dosage forms
appropriate for the methods described herein. The compounds
described herein, or compositions containing them, including such
formulations, may be administered by a wide variety of conventional
routes for the methods described herein, and in a wide variety of
dosage formats, utilizing known procedures (see generally,
Remington: The Science and Practice of Pharmacy, (21.sup.st ed.,
2005)).
[0304] The term "therapeutically effective amount" as used herein,
refers to that amount of active compound or pharmaceutical agent
that elicits the biological or medicinal response in a tissue
system, animal or human that is being sought by a researcher,
veterinarian, medical doctor or other clinician, which includes
alleviation of the symptoms of the disease or disorder being
treated. In one aspect, the therapeutically effective amount is
that which may treat or alleviate the disease or symptoms of the
disease at a reasonable benefit/risk ratio applicable to any
medical treatment. However, it is to be understood that the total
daily usage of the compounds and compositions described herein may
be decided by the attending physician within the scope of sound
medical judgment. The specific therapeutically-effective dose level
for any particular patient will depend upon a variety of factors,
including the disorder being treated and the severity of the
disorder; activity of the specific compound employed; the specific
composition employed; the age, body weight, general health, gender
and diet of the patient: the time of administration, route of
administration, and rate of excretion of the specific compound
employed; the duration of the treatment; drugs used in combination
or coincidentally with the specific compound employed; and like
factors well known to the researcher, veterinarian, medical doctor
or other clinician of ordinary skill.
[0305] It is also appreciated that the therapeutically effective
amount, whether referring to monotherapy or combination therapy, is
advantageously selected with reference to any toxicity, or other
undesirable side effect, that might occur during administration of
one or more of the compounds described herein. Further, it is
appreciated that the co-therapies described herein may allow for
the administration of lower doses of compounds that show such
toxicity, or other undesirable side effect, where those lower doses
are below thresholds of toxicity or lower in the therapeutic window
than would otherwise be administered in the absence of a
cotherapy.
[0306] In addition to the illustrative dosages and dosing protocols
described herein, it is to be understood that an effective amount
of any one or a mixture of the compounds described herein can be
readily determined by the attending diagnostician or physician by
the use of known techniques and/or by observing results obtained
under analogous circumstances. In determining the effective amount
or dose, a number of factors are considered by the attending
diagnostician or physician, including, but not limited to the
species of mammal, including human, its size, age, and general
health, the specific disease or disorder involved, the degree of or
involvement or the severity of the disease or disorder, the
response of the individual patient, the particular compound
administered, the mode of administration, the bioavailability
characteristics of the preparation administered, the dose regimen
selected, the use of concomitant medication, and other relevant
circumstances.
[0307] The dosage of each compound of the claimed combinations
depends on several factors, including: the administration method,
the condition to be treated, the severity of the condition, whether
the condition is to be treated or prevented, and the age, weight,
and health of the person to be treated. Additionally,
pharmacogenomic (the effect of genotype on the pharmacokinetic,
pharmacodynamic or efficacy profile of a therapeutic) information
about a particular patient may affect the dosage used.
[0308] It is to be understood that in the methods described herein,
the individual components of a co-administration, or combination
can be administered by any suitable means, contemporaneously,
simultaneously, sequentially, separately or in a single
pharmaceutical formulation. Where the co-administered compounds or
compositions are administered in separate dosage forms, the number
of dosages administered per day for each compound may be the same
or different. The compounds or compositions may be administered via
the same or different routes of administration. The compounds or
compositions may be administered according to simultaneous or
alternating regimens, at the same or different times during the
course of the therapy, concurrently in divided or single forms.
[0309] The term "administering" as used herein includes all means
of introducing the compounds and compositions described herein to
the patient, including, but are not limited to, oral (po),
intravenous (iv), intramuscular (im), subcutaneous (sc),
transdermal, inhalation, buccal, ocular, sublingual, vaginal,
rectal, and the like. The compounds and compositions described
herein may be administered in unit dosage forms and/or formulations
containing conventional nontoxic pharmaceutically-acceptable
carriers, adjuvants, and/or vehicles.
[0310] Illustrative formats for oral administration include
tablets, capsules, elixirs, syrups, and the like.
[0311] Illustrative routes for parenteral administration include
intravenous, intraarterial, intraperitoneal, epidurial,
intraurethral, intrasternal, intramuscular and subcutaneous, as
well as any other art recognized route of parenteral
administration.
[0312] Illustratively, administering includes local use, such as
when administered locally to the site of disease, injury, or
defect, or to a particular organ or tissue system. Illustrative
local administration may be performed during open surgery, or other
procedures when the site of disease, injury, or defect is
accessible. Alternatively, local administration may be performed
using parenteral delivery where the compound or compositions
described herein are deposited locally to the site without general
distribution to multiple other non-target sites in the patient
being treated. It is further appreciated that local administration
may be directly in the injury site, or locally in the surrounding
tissue. Similar variations regarding local delivery to particular
tissue types, such as organs, and the like, are also described
herein. Illustratively, compounds may be administered directly to
the nervous system including, but not limited to, intracerebral,
intraventricular, intracerebroventricular, intrathecal,
intracisternal, intraspinal and/or peri-spinal routes of
administration by delivery via intracranial or intravertebral
needles and/or catheters with or without pump devices.
[0313] Depending upon the disease as described herein, the route of
administration and/or whether the compounds and/or compositions are
administered locally or systemically, a wide range of permissible
dosages are contemplated herein, including doses falling in the
range from about 1 .mu.g/kg to about 1 g/kg. The dosages may be
single or divided, and may administered according to a wide variety
of protocols, including q.d., b.i.d., t.i.d., or even every other
day, once a week, once a month, once a quarter, and the like. In
each of these cases it is understood that the therapeutically
effective amounts described herein correspond to the instance of
administration, or alternatively to the total daily, weekly, month,
or quarterly dose, as determined by the dosing protocol.
[0314] In making the pharmaceutical compositions of the compounds
described herein, a therapeutically effective amount of one or more
compounds in any of the various forms described herein may be mixed
with one or more excipients, diluted by one or more excipients, or
enclosed within such a carrier which can be in the form of a
capsule, sachet, paper, or other container. Excipients may serve as
a diluent, and can be solid, semi-solid, or liquid materials, which
act as a vehicle, carrier or medium for the active ingredient.
Thus, the formulation compositions can be in the form of tablets,
pills, powders, lozenges, sachets, cachets, elixirs, suspensions,
emulsions, solutions, syrups, aerosols (as a solid or in a liquid
medium), ointments, soft and hard gelatin capsules, suppositories,
sterile injectable solutions, and sterile packaged powders. The
compositions may contain anywhere from about 0.1% to about 99.9%
active ingredients, depending upon the selected dose and dosage
form.
[0315] The effective use of the compounds, compositions, and
methods described herein for treating or ameliorating diseases
caused by pathogenic cells expressing PSMA may be based upon animal
models, such as murine, canine, porcine, and non-human primate
animal models of disease. For example, it is understood that
prostate cancer in humans may be characterized by a loss of
function, and/or the development of symptoms, each of which may be
elicited in animals, such as mice, and other surrogate test
animals. In particular the mouse models described herein where
cancer cells, such as LNCaP cells are subcutaneously implanted may
be used to evaluate the compounds, the methods of treatment, and
the pharmaceutical compositions described herein to determine the
therapeutically effective amounts described herein.
[0316] The compounds, linkers, intermediates, and conjugates
described herein may be prepared using conventional processes,
including those described in International Patent Publication Nos.
WO 2009/002993, WO 2004/069159, WO 2007/022494, and WO 2006/012527,
and U.S. patent application Ser. No. 13/837,539 (filed Mar. 15,
2013). The disclosures of each of the foregoing are herein
incorporated by reference in their entirety.
[0317] Each of the publications cited herein is incorporated herein
by reference.
[0318] The following examples further illustrate specific
embodiments of the invention; however, the following illustrative
examples should not be interpreted in any way to limit the
invention.
Examples
##STR00092##
[0319] Example. Compound 104
[0320] In a 250 mL round-bottom flask, H-Glu(OtBu)-OtBu.HCl (1)
(4.83 g, 16.3 mmol) and 4-nitrophenyl chloroformate (102) (3.47 g,
17.2 mmol) were dissolved in dichloromethane (50 mL) and stirred in
an ice bath under argon. Diisopropylethylamine (6.28 mL, 36.1 mmol)
was added slowly, dropwise and the reaction mixture was stirred in
the ice bath for 5 min, then warmed to room temperature and stirred
for 30 min. H-Lys(Z)-OtBu.HCl (103) (7.01 g, 18.8 mmol) was added
portionwise, followed by dropwise addition of diisopropylethylamine
(6.54 mL, 37.5 mmol), and stirred at room temperature for 1 hr. The
reaction mixture was concentrated under reduced pressure, then
purified by silica gel chromatography in 10-100% ethyl
acetate/petroleum ether to yield 104 (8.76 g, 86%, ESI m/z=622.54
[M+H].sup.+).
Example. Compound 105
[0321] 104 (8.76 g, 14.1 mmol) was dissolved in anhydrous methanol
(100 mL) and added slowly along the walls of the 250 mL
round-bottom flask containing palladium on carbon, 10 wt. % (100
mg). A balloon containing hydrogen gas was attached to the flask
using a three-way stopcock adapter, and the atmosphere of the flask
was evacuated under reduced pressure, then replaced with hydrogen
gas (3.times.), then stirred at room temperature under hydrogen gas
for 1 hr. To the reaction mixture was added dry, untreated celite
(.about.20 g) and stirred for 5 min. The reaction mixture was
filtered and concentrated under reduced pressure to yield 105 (6.86
g, quantitative, ESI m/z=488.46 [M+H].sup.+).
##STR00093##
Example. Compound 107
[0322] Boc-4-aminomethylphenylacetic acid (106) (2.00 g, 7.5 mmol)
dissolved in a solution of trifluoroacetic acid (9.75 mL) and
triisopropylsilane (0.25 mL) and stirred at room temperature for 30
min, then concentrated under reduced pressure and coevaporated with
dichloromethane (3.times.), then placed under vacuum, to yield
4-aminomethylphenylacetic acid (107) (quantitative).
##STR00094##
Example. Compound 108
[0323] To a stirring solution of 4-nitrophenyl chloroformate (102)
(1.01 g, 5.0 mmol) in dry dimethylformamide (10 mL) was added
slowly dropwise a solution of 105 (2.45 g, 5.0 mmol) and
diisopropylethylamine (0.88 mL, 5.0 mmol) in dry dimethylformamide
(10 mL), and the reaction mixture was stirred at room temperature
for min under argon. The reaction mixture was cooled in an ice bath
and a suspension of 7 (.about.1.25 g, .about.7.5 mmol) and
diisopropylethylamine (1.76 mL, 10.1 mmol) in dry dimethylformamide
(10 mL) was added slowly dropwise to the reaction vessel, then the
reaction mixture was warmed to room temperature and stirred for 30
min under argon. The reaction mixture was purified by preparative
HPLC in 10-100% acetonitrile/0.1% formic acid to yield 8 (0.56 g,
16%, .sup.1H NMR consistent with structure of 108; ESI m/z=679.50
[M+H].sup.+).
##STR00095##
Example. Preparation of Protected Ligand 7, Including Coupling
Group
##STR00096##
[0324] Example. Peptide 109
TABLE-US-00003 [0325] TABLE 1 Reagents for peptide 109 synthesis
Molecular weight Reagent mmol Equivalents (g/mol) quantity H-Cys(4-
0.87 1.0 methoxytrityl)-2- chlorotrityl-Resin Fmoc-Asp(OtBu)--OH 2
.times. 1.74 2 .times. 2.0 411.5 716 mg PyBOP 2 .times. 1.73 2
.times. 2.0 520.39 900 mg diisopropylethylamine 2 .times. 3.48 2
.times. 4.0 129.25 606 .mu.L (d = 0.742 g/mL)
[0326] In a peptide synthesis vessel
H-Cys(4-methoxytrityl)-2-chlorotrityl-resin (0.87 mmol) was loaded
and washed with isopropyl alcohol (3.times.10 mL) followed by
dimethylformamide (3.times.10 mL). To the vessel was then
introduced Fmoc-Asp(OtBu)-OH (2.0 equiv) in dimethylformamide,
diisopropylethylamine (4.0 equiv), and PyBOP (2.0 equiv). Argon was
bubbled for 1 hr, the coupling solution was drained, and the resin
was washed with dimethylformamide (3.times.10 mL) and isopropyl
alcohol (3.times.10 mL). Kaiser tests were performed to assess
reaction completion. Fmoc deprotection was carried out using 20%
piperidine in dimethylformamide (3.times.10 mL) before each amino
acid coupling. The above sequence was repeated to complete 2
coupling steps. The resin was dried under argon for 30 min.
##STR00097##
Example. Peptide 110
TABLE-US-00004 [0327] TABLE 2 Reagents for peptide 110 synthesis
Molecular weight Reagent mmol Equivalents (g/mol) quantity
Fmoc-Asp(OtBu)- 0.18 1.0 Asp(OtBu)-Cys(Mmt)- 2-ClTrt-resin 108 0.22
1.2 678.81 150 mg PyBOP 0.37 2.0 520.39 191 mg
diisopropylethylamine 0.74 4.0 129.25 128 .mu.L (d = 0.742
g/mL)
[0328] In a peptide synthesis vessel 109 (0.18 mmol) was loaded and
washed with isopropyl alcohol (3.times.10 mL) followed by
dimethylformamide (3.times.10 mL). Fmoc deprotection was carried
out using 20% piperidine in dimethylformamide (3.times.10 mL).
Kaiser tests were performed to assess reaction completion. To the
vessel was then introduced 108 (1.2 equiv) in dimethylformamide,
diisopropylethylamine (4.0 equiv), and PyBOP (2.0 equiv). Argon was
bubbled for 1 hr, the coupling solution was drained, and the resin
was washed with dimethylformamide (3.times.10 mL) and isopropyl
alcohol (3.times.10 mL). Kaiser tests were performed to assess
reaction completion. Peptide was cleaved from the resin using a
cleavage mixture consisting of dithiothreitol (114 mg, 0.74 mmol)
dissolved in a solution of trifluoroacetic acid (19 mL). H.sub.2O
(0.5 mL), triisopropylsilane (0.5 mL). One-third of the cleavage
mixture was introduced and argon was bubbled for 30 min. The
cleavage mixture was drained into a clean flask. The resin was
bubbled 2 more times with more cleavage mixture, for 30 min each,
and drained into a clean flask. The drained cleavage mixture was
then concentrated and purified by preparative HPLC in 0-30%
acetonitrile/0.1% formic acid to yield 110 (66.9 mg, 43%, .sup.1H
NMR consistent with structure of 110; ESI m/z=844.57
[M+H].sup.+).
Example
[0329] Similarly, the following compounds are prepared as described
herein:
##STR00098## ##STR00099## ##STR00100##
Example. EC1169 (Compound 112)
[0330] In a 25 mL round bottom flask, 16 (47 mg, 0.04 mmol) was
dissolved in dimethylsulfoxide (2 mL). A solution of 110 (36 mg,
0.04 mmol) in 20 mM pH7 sodium phosphate buffer (2 mL) was added
dropwise, stirring at room temperature with Argon bubbling for 30
min. The reaction mixture was purified by preparative HPLC (10-100%
acetonitrile/50 mM NH.sub.4HCO.sub.3 pH7) to yield 112 (56.6 mg,
74%, .sup.1H NMR consistent with structure of EC1169; ESI
m/z=895.58 [M+2H].sup.2+).
Example. Synthesis of 3-nitro-2-disulfenylethanol 2
##STR00101##
[0332] A three-necked 500 mL flask was dried and argon purged, then
fitted with an addition funnel. 3-Nitro-2-sulfenyl chloride
pyridine 1 (5.44 g, 27.11 mmol, 1.4 equiv) was added to the flask
and dissolved in 200 mL of CH.sub.2Cl.sub.2. The solution was
cooled to 0.degree. C. Mercaptoethanol (1.33 mL, 18.98 mmol) was
diluted with 50 mL of CH.sub.2Cl.sub.2 and placed in the addition
funnel. The 2-mercaptoethanol solution was then added drop-wise
slowly over the course of 15 minutes. The reaction progress was
monitored by TLC (Rf 0.4 in 5% CH.sub.3OH/CH.sub.2Cl.sub.2).
Solvent was removed under reduced pressure and dried. The crude
product was purified over silica gel (5%
CH.sub.3OH/CH.sub.2Cl.sub.2). The fractions were collected and
solvent was removed by evaporating on a rotary evaporator and
dried. 3.4 g of 3-nitro-2-disulfenylethanol 2 was obtained (77%
yield).
Example. Synthesis of
4-nitrophenyl-(3'-nitropyridin-2'-yl)disulfenylethyl carbonate
3
##STR00102##
[0334] A 250 mL Round-Bottomed Flask was dried and argon purged.
3-Nitro-2-disulfenylethanol 2 (3.413 g, 14.69 mmol) was added and
dissolved in 45 mL of CH.sub.2Cl.sub.2. 4-Nitrophenylchloroformate
(3.663 g, 17.63 mmol, 1.2 equiv) was added, along with
triethylamine (2.9 mL, 20.57 mmol, 1.4 equiv), and the mixture
stirred under argon overnight. The mixture was concentrated under
reduced pressure and dried. The residue was purified by silica (30%
EtOAc/petroleum ether) and the fractions were collected, solvent
was removed under reduced pressure, and dried. 2.7 g of
4-nitrophenyl-(3'-nitropyridin-2'-yl)disulfenylethyl carbonate 3
was obtained (47% yield).
Example. Synthesis of 2-(Boc-tubutyrosine (Tut))hydrazinecarboxylic
acid (3'nitropyridyl-2'-yl)disulfanylethyl ester 6
##STR00103##
[0336] 10.67 g (33 mmol) of Boc-Tut-acid 4 was dissolved in 100 mL
anhydrous THF, 17.24 g (33 mmol) of PyBop, and 17.50 mL (99 mmol,
3.0 equiv) of DIPEA were added. The reaction mixture stirred for
few minutes, 1.0 mL (31.68 mmol, 0.96 equiv) of hydrazine was added
and stirred for 15 minutes. LC-MS analysis (X-Bridge shield RP18,
3.5 .quadrature.m column; gradient 10% to 100% acetonitrile in 6
min, pH 7.4 buffer) confirmed the hydrazide 5 formation. 14.47 g
(36.3 mmol, 1.1 equiv) of
4-nitrophenyl-(3'-nitropyridin-2'-yl)disulfenylethyl carbonate 2
was added. The resulting clear solution was stirred at room
temperature for 24 hours. LC-MS analysis (X-Bridge shield RPI8,
3.5.quadrature.m column; gradient 30% to 100% acetonitrile in 9
min, pH 7.4 buffer) indicated >98% conversion. The reaction
mixture was diluted with EtOAc (.about.1.0 L), washed with sat.
NH.sub.4Cl (400 mL), sat. NaHCO.sub.3 solution (3.times.300 mL),
and brine (300 mL). The organic layer was dried over
Na.sub.2SO.sub.4 (100 g), and concentrated under reduced pressure.
The crude product was loaded onto a Teledyne Redisep Gold Silica
Column and eluted with MeOH/CH.sub.2Cl.sub.2 (330 g column; 0 to
10% gradient) using a CombiFlash chromatography system. The
fractions were collected and solvent was removed under reduced
pressure and dried. 16.10 g of 2-(Boc-Tut)hydrazinecarboxylic acid
(3'nitropyridyl-2'-yl)disulfanylethyl ester 6 was obtained (82%
yield).
Example. Synthesis of azido methylbutyrate dipeptide 9
##STR00104##
[0338] Dipeptide 7 (10.83 g, 27.25 mmol) was dissolved in 100 mL
dichloromethane and imidazole (2.05 g, 1.1 eq.) was added. The
reaction mixture was stirred at room temperature to dissolve all
solids and cooled in the ice bath for 10 min. TESCl (4.8 mL, 1.05
eqiv.) was added drop-wise at 0.degree. C., stirred under argon,
and warmed to room temperature over 1.5 h. TLC (3:1 hexanes/EtOAc)
showed complete conversion. The reaction was filtered to remove the
imidazole HCl salt. 125 mL dichloromethane was added to the
filtrate, and the resulting solution was extracted with 250 mL
brine. The brine layer was extracted with 125 mL dichloromethane.
The combined organic phase was washed with 250 mL brine, separated,
dried over 45.2 g of Na.sub.2SO.sub.4, and filtered. The resulting
solution was concentrated under reduced pressure, co-evaporated
with toluene (2.times.5 mL) and dried over high-vacuum overnight to
give 14.96 g of crude product 8.
[0339] The crude product 8 was used without further purification.
TES protected dipeptide was dissolved in 100 mL THF (anhydrous,
inhibitor-free), cooled to -45.degree. C., and stirred at
-45.degree. C. for 15 minutes before adding KHMDS (0.5 M in
toluene, 61 mL, 1.05 equiv.), drop-wise. After the addition of
KHMDS was finished, the reaction was stirred at -45.degree. C. for
20 minutes, and chloromethyl butyrate (4.4 mL, 1.1 equiv.) was
added. The reaction mixture was stirred at -45.degree. C. for
another 20 minutes. The reaction was quenched with 25 mL MeOH and
warmed to room temperature. 250 mL EtOAc and 250 mL brine were
added to the reaction mixture, and the organic phase was separated.
The solvent was evaporated to reduce the volume of solution. The
solution was passed through 76.5 g silica in a 350 mL sintered
glass funnel. The silica plug was washed with 500 mL
EtOAc/petroleum ether (1:4). The filtrate and the wash were
concentrated to oily residue and dried under high vacuum to give
16.5 g product 9 as a light yellow wax.
Example. Synthesis of tripeptide methyl ester 10
##STR00105##
[0341] Based on 16.5 g of alkylated dipeptide 9 (26.97 mmol.),
N-methyl pipecolinate (MEP) (5.51 g, 1.4 equiv.) and
pentafluorophenol (7.63 g, 1.5 equiv.) were added to a 300 mL
hydrogenation flask. NMP (115 mL) was then added, followed by EDC
(7.78 g, 1.5 equiv.). The mixture was stirred at room temperature
for overnight. 16.5 g of alkylated dipeptide 9 was dissolved in
16.5 mL NMP, transferred the solution into the hydrogenation flask,
washed the residual 9 with 8 mL NMP, and transferred into the
hydrogenation flask. Dry 10% Pd/C (1.45, 0.05 eq.) was added. The
reaction mixture was vacuumed/back filled with hydrogen 3 times,
and the flask was shaken under hydrogen (.about.35 psi) for 3.5
hours. The reaction mixture was analyzed by HPLC. The reaction
mixture was filtered through 40 g of celite in a 350 mL sintered
glass funnel and washed with 250 mL of EtOAc. The filtrate and the
wash were transferred to a separatory funnel and washed with a 1%
NaHCO.sub.3/10% NaCl solution (200 mL.times.3). The organic layer
was isolated and dried over 45.2 g of Na.sub.2SO.sub.4. The
solution was filtered and rotovaped under reduced pressure. A
sticky amber residue was obtained and dried under high vacuum
overnight to give 19.3 g of crude product. The crude product was
dissolved in 10 mL of dichloromethane, split into two portions, and
purified with a 330 g Teledyne Redisep Silica Gold column. The
combined fractions of two purifications were evaporated and dried
under high vacuum to give 7.64 g of 10 as a pale yellow solid
(overall yield: 39% over 3 steps from compound 7).
Example. Synthesis of tripeptide acid 11
##STR00106##
[0343] Methyl ester 10 (6.9 g, 9.7 mmol) was dissolved in
1,2-dichloroethane (193 mL) and added to a round bottomed flask,
equipped with a stir bar and condenser. To this solution was added
trimethyltin hydroxide (24.6 g, 14 eq.). The mixture was heated at
70.degree. C. for 5 hours. LC-MS analysis indicated that the
desired product had been formed and <15% of starting methyl
ester 10 remained. The reaction was cooled in an ice bath for 30
minutes. The resulting precipitate was then removed by filtration.
The filtrate was stored overnight at -20.degree. C. The filtrate
was then divided into two portions and each was subjected the
chromatography procedure which follows.
[0344] Each portion was concentrated under reduced pressure and
then placed under high vacuum for 30 min. The concentrate was then
immediately dissolved in acetonitrile (95 mL). To this solution was
then added an ammonium bicarbonate solution (95 mL; 50 mM, pH=7).
This solution was loaded onto a Biotage SNAP C18 reverse phase
cartridge (400 g, KP-C18-HS) and eluted with 50 mM ammonium
bicarbonate and acetonitrile (1:1 to 100% ACN) using a Biotage
chromatography system. Fractions were analyzed by LC-MS. Pure
fractions were combined and ACN was removed under reduced pressure.
The resulting aqueous suspension was extracted with EtOAc
(3.times.). The combined organic layers were washed with brine,
dried over anhydrous Na.sub.2SO.sub.4, and concentrated under
reduced pressure. Purification of the two portions resulted in the
recovery of clean 11 (4.6 g, 65%).
Example. Synthesis of acetyl tripeptide acid 13
##STR00107##
[0346] In a round bottomed flask, tripeptide acid 11 (3.9 g, 5.6
mmol) was dissolved in anhydrous THF (23 mL). To this solution was
added 3 HF.TEA complex (1.8 mL, 2 eq.). The reaction was stirred at
room temperature for 1 hour. LC-MS analysis indicated complete
conversion to the desired des-TES product 12. The solvent was
removed under reduced pressure and the residue was placed on the
high vacuum for 40 minutes. The resulting residue was then
dissolved in pyridine (26 mL), and acetic anhydride (7.9 mL, 15
eq.) and DMAP (25 mg) were added. The reaction was stirred at room
temperature for 1 hour. LC-MS analysis indicated complete
conversion to the desired acetyl tripeptide acid 13. To the
reaction mixture was then added a 1:1 solution of 1,4-dioxane/water
(150 mL). The reaction was stirred for 1 hour at which point the
solvents were removed under high vacuum rotovap. To the residue was
added toluene and the solvent was removed under vacuum (80 mL,
3.times.). The resulting crude 13 was dried under high vacuum
overnight. The crude material was then dissolved in ACN (72 mL).
Sodium phosphate buffer (50 mM, pH=7.8, 288 mL) was then added, and
the pH of the resulting suspension was adjusted to neutral using
saturated sodium bicarbonate solution. This solution was loaded
onto a Biotage SNAP C18 reverse phase cartridge (400 g. KP-C18-HS)
and eluted with water and acetonitrile (20% ACN to 65% ACN) using a
Biotage chromatography system. Fractions were analyzed by LC-MS.
Clean fractions were combined, the ACN was removed, and the aqueous
solution was placed on the freeze dryer, resulting in purified
acetyl tripeptide 13 (2.5 g, 71%).
Example. Synthesis of 2-(tubulysin B)hydrazinecarboxylic acid
(3'nitropyridyl-2'-yl)disulfanylethyl ester 16
##STR00108##
[0348] The activated Boc-Tut-fragment 6 (2.63 g, 4.42 mmol, 1.1
equiv) was treated with TFA/CH.sub.2Cl.sub.2 (42 mL; 1:1) and
stirred for 30 minutes. LC-MS analysis (X-Bridge shield RP18, 3.5
.quadrature.m column; gradient 10% to 100% acetonitrile in 6 min,
pH 7.4 buffer) confirmed the product formation. TFA was removed
under reduced pressure, co-evaporated with CH.sub.2Cl.sub.2
(3.times.30 mL) and activated Tut-derivative 14 was dried under
high vacuum for 18 h. In another flask, the tripeptide acid 13
(2.51 g, 4.02 mmol) was dissolved in 70 mL CH.sub.2Cl.sub.2
(anhydrous) and 1.48 g (8.04 mmol, 2.0 equiv) of pentafluorophenol
in 5 mL of CH.sub.2Cl.sub.2 was added, followed by 8.74 g (20.1
mmol, 5.0 equiv) of DCC-resin. The resulting reaction mixture was
stirred at room temperature for 20 hours. LC-MS analysis (X-Bridge
shield RP18, 3.5 .quadrature.m column; gradient 10% to 100%
acetonitrile in 6 min, pH 7.4 buffer) indicated >99% conversion.
The DCC-resin was filtered off, the CH.sub.2Cl.sub.2 was removed
under reduced pressure, and the pentafluorophenol activated product
15 was dried under high vacuum for 10 minutes. The residue was
dissolved in 16.7 mL DMF, and DIPEA (12.6 mL, 72.36 mmol, 18.0
equiv) was added. Tut-fragment trifluoroacetic acid salt 14 in DMF
(8.5 mL) was added slowly over 5 min. The resulting clear solution
was stirred at room temperature for 1 h. LC-MS analysis (X-Bridge
shield RP18, 3.5 .quadrature.m column; gradient 10% to 100%
acetonitrile in 6 min. pH 7.4 buffer) confirmed the product
formation. The reaction mixture was diluted with EtOAc (700 mL),
washed with brine (300 mL, 2.times.100 mL), dried over
Na.sub.2SO.sub.4 (75 g), concentrated, and dried for 15 hours. The
crude product was dissolved in CH.sub.2Cl.sub.2 (25 mL) and loaded
onto a Teledyne Redisep Gold Silica Column and eluted with
MeOH/CH.sub.2Cl.sub.2 (330 g column; 0 to 5% gradient) using
Combiflash chromatographic system. The fractions were collected and
solvent was removed by evaporating on a rotary evaporator and
dried. 3.91 g of 2-(tubulysin B)hydrazinecarboxylic acid
(3'nitropyridyl-2'-yl)disulfanylethyl ester 16 was obtained (89%
yield).
Example. Preparation of 2-(tubulysin B)hydrazinecarboxylic acid
(pyrid-2-yl)disulfanylethyl ester 3
##STR00109##
[0349] Example
[0350] Similarly, the following compounds are prepared as described
herein:
##STR00110##
Example
[0351] Additional tubulysins described herein may be isolated from
natural sources, including but not limited to bacteria and other
fermentations. Alternatively, the tubulysins described herein may
be prepared according to conventional processes, including but not
limited to the processes described in PCT International Publication
Nos. WO 2009/055562, WO 2012/019123, and WO 2013/149185, and
co-pending U.S. application Ser. No. 13/841,078, the disclosures of
each of which are incorporated herein by reference in their
entirety.
Example. Alternative Preparation of EC1169 (Compound 112)
##STR00111##
[0352] Example
[0353] The following representative example compounds are described
to better illustrate the invention described herein and may be
prepared according to the synthetic methods described for the above
examples, and/or using conventional processes.
##STR00112## ##STR00113## ##STR00114## ##STR00115## ##STR00116##
##STR00117##
Method Example. PSMA Relative Affinity Assay
[0354] LNCaP cells are seeded in 12-well Corning Cell-BIND plates
and allowed to form adherent monolayers overnight in RPMI/HIFCS.
Spent incubation media is replaced with RPMI supplemented with 10%
HIFCS and containing a standard PSMA binding ligand, such as 100 nM
of 3H-PMPA or a competing compound, such as EC0652, Re-EC652, or
.sup.99mTc-EC0652, in the absence and presence of increasing
concentrations of test compound, such as unlabeled PMPA, or a
compound described herein, such as EC1169 or EC1568, a negative
control intermediate lacking a PSMA binding ligand which is used as
a negative control. Cells are incubated for 1 h at 37.degree. C.
and then rinsed three times with 0.5 mL of PBS. Five hundred
microliters of 1% sodium dodecylsulfate in PBS are added to each
well; after 5 min, cell lysates are collected, transferred to
individual tubes or to vials containing 5 mL of scintillation
cocktail, and then counted for radioactivity. Cells exposed to only
the standard PSMA binding ligand, such as 3H-PMPA, or competing
compound, such as .sup.99mTc-EC0652, in FFRPMI (no competitor) are
designated as negative controls, whereas cells exposed to the
standard PSMA binding ligand, such as 3H-PMPA, plus 1 mM unlabeled
PMPA or competing compound, such as .sup.99mTc-EC0652 plus
Re-EC0652, serve as positive controls. Disintegrations per minute
(DPMs) measured in the latter samples (representing nonspecific
binding of label) are subtracted from the DPM values from all
samples. Relative affinities are defined as the inverse molar ratio
of compound required to displace 50% of the standard PSMA binding
ligand, such as .sup.3H-PMPA, or the competing compound, such as
.sup.99mTc-EC0652, bound to PSMA on LNCaP cells, and the relative
affinity of the standard PSMA binding ligand, such as PMPA, or the
competing compound, such as Re-EC0652, for PSMA is set to 1.
Method Example. Dose Response Assay Against PSMA+ LNCaP Cells
[0355] LNCaP cells are seeded in 24-well Corning Cell-BIND plates
and allowed to form nearly confluent monolayers overnight in
RPMI/HIFCS. Thirty minutes prior to the addition of test compound,
such as a compound described herein, spent medium is aspirated from
all wells and replaced with fresh RPMI. Following one rinse with 1
mL of fresh RPMI/HIFCS, each well receives 1 mL of media containing
increasing concentrations of test compound (four wells per sample).
Test compound treated cells are pulsed for 2 h at 37.degree. C.,
rinsed four times with 0.5 mL of media, and then chased in 1 mL of
fresh media up to 70 h. Spent media is aspirated from all wells and
replaced with fresh media containing 5 .mu.Ci/mL .sup.3H-thymidine.
Following a further 4 h 37.degree. C. incubation, cells are washed
three times with 0.5 mL of PBS and then treated with 0.5 mL of
ice-cold 5% trichloroacetic acid per well. After 15 min, the
trichloroacetic acid is aspirated and the cells are solubilized by
the addition of 0.5 mL of 0.25 N sodium hydroxide for 15 min. Four
hundred and fifty microliters of each solubilized sample is
transferred to scintillation vials containing 3 mL of Ecolume
scintillation cocktail and then counted in a liquid scintillation
counter. Final tabulated results are expressed as the percentage of
.sup.3H-thymidine incorporation relative to untreated controls.
Method Example. Activity In Vivo Against PSMA+ Expressing Tumor
Implanted in Mice
[0356] Four to seven week-old male nu/nu mice (Harlan Sprague
Dawley, Inc., Indianapolis, Ind.) are maintained on a standard 12 h
light-dark cycle and fed ad libitum with rodent diet #2918 (Harlan
Teklad, Madison, Wis.) for the duration of the experiment. LNCaP
cells are grown in RPMI in 10% HIFCS at 37.degree. C. in a 5%
CO.sub.2/95% air-humidified atmosphere, harvested and resuspended
on ice in matrigel solution (50% RPMI+50% matrigel high
concentration. BD#354248) to a final concentration of
1.times.10.sup.6 cells/50 .mu.L. Cell solution and injection
needles (28 gauge) are kept on ice prior to injection and 50 .mu.L
of the cell solution injected in the subcutis of the dorsal medial
area. Mice are divided into groups of five, seven, or nine, and
freshly prepared test compound solutions are injected through the
lateral tail vein under sterile conditions in a volume of 200 .mu.L
of phosphate-buffered saline (PBS). Intravenous (i.v.) treatments
are typically initiated when the LNCaP tumors are approximately
100-150 mm.sup.3 in volume. The mice in the control groups do not
receive any treatment. Growth of each s.c. tumor is followed by
measuring the tumor three times per week during treatment and twice
per week thereafter, until a volume of 1500 mm.sup.3 is reached.
Tumors are measured in two perpendicular directions using Vernier
calipers, and their volumes are calculated as
0.5.times.L.times.W.sup.2, where L=measurement of longest axis in
mm and W=measurement of axis perpendicular to L in mm. As a general
measure of gross toxicity, changes in body weights are determined
on the same schedule as tumor volume measurements. Maximum % weight
loss on any given day due to treatment is determined for each
mouse. Survival of animals is monitored daily. Animals that are
moribund (or unable to reach food or water) are euthanized by
CO.sub.2 asphyxiation.
Example. Relative Affinity of Compounds Described Herein Compared
to PSMA Inhibitors DUPA and PMPA
[0357] PMPA is reportedly one of the highest affinity ligands, or
the highest affinity ligand, for PSMA. The data in FIG. 1 and FIG.
2 show that compounds described herein exhibit higher affinity for
PSMA than does PMPA.
##STR00118##
[0358] It was unexpectedly discovered that the ligands described
herein have a higher affinity for PSMA than the reportedly highest
affinity ligand PMPA. In addition, it was unexpectedly discovered
herein that conjugates of the ligands described herein had even
higher affinity for PSMA.
[0359] The binding data for additional illustrative compounds
described herein are shown in the following table
TABLE-US-00005 Relative PSMA Binding Affinity Example (fold over
PMPA = 1.0) EC1080 6 EC1067 30 EC1100 20 EC1167 11 EC1168 17 EC1170
7 EC1069 22 EC1183 9 EC1241 1.1 EC1303 7 EC1307 28 EC1308 20 EC1310
10 EC1584 6 EC1568 0 (negative control)
Example. Dose Response of Compounds Described Herein Against
PSMA+LNCaP Cells
[0360] Using a standard .sup.3H-thymidine incorporation assay as a
measure of cytotoxicity, the data in FIG. 3 show that EC1169
exhibits dose responsive cytotoxicity against cells in vitro with
an IC.sub.50 of 13 nM. The corresponding dose responsive
cytotoxicity and IC.sub.50 values for () EC718. IC.sub.50 17.9 nM;
(.diamond-solid.) EC11677, IC.sub.50 20.9 nM; (.tangle-solidup.)
EC1719, IC.sub.50 37.5 nM; ( ) EC1720. IC.sub.50 54.2 nM;
(.box-solid.) EC1721, IC.sub.50 65.6 nM are shown in FIG. 4
Example
[0361] Additional compounds described herein against LNCaP cells (2
h-72 h) as determined by .sup.3H-thymidine incorporation cells in
vitro are shown in the following table.
TABLE-US-00006 % .sup.3H-thymidine Example incorporation EC1069 13
nM EC1268 59.1 EC1385 184 EC1386 57 EC1387 24 EC1388 12 EC1437 30
EC1550 22 EC1551 20 EC1452 22 EC1584 33 EC1588 42
Example. Activity of Compounds Described Herein Against PSMA+
Tumors In Vivo
[0362] As shown in FIG. 5 treatment of nude mice bearing
PSMA-positive LNCaP human xenografts with EC1169 (c), EC1550 ( ),
and EC1551 (.box-solid.), each at 2 .mu.mol/kg, TIW, 2 weeks, leads
to complete responses in all tested animals. Each compound was
compared against vehicle-treated controls (.diamond-solid.). A
complete response is observed when the tumor does not appear to
have any net growth during the treatment period of 14 days (the
vertical dotted line indicates the last treatment day). As
described herein, it is to be understood that the implants comprise
the cancer cells in a matrix (100-150 mm.sup.3 total volume).
Because the matrix remains during the entire observation period, a
decrease in the size of the tumor cannot always be determined by
external measurement. It was also surprisingly found that,
treatment with compounds described herein leads to cure. For
example, EC1169 leads to cure in 2/7 tested animals. A cure is
observed when the tumor does not appear to grow during the entire
observation period of 85 days. The data shown in FIG. 5 are the
average of the measurements for each cohort. Therefore, it is to be
understood that the increase in tumor volume beginning at about day
40-45 represents regrowth in the remaining test animals.
Example
[0363] Gross toxicity of compounds described herein. As shown in
FIG. 6, the observed efficacy of EC1169 (c), EC1550 ( ), and EC1551
(.box-solid.), occurred in the absence of weight loss or major
organ tissue degeneration.
Example
[0364] Activity of compounds described herein against PSMA+ tumors
in vivo. Similarly, as shown in FIG. 7, treatment of nude mice
bearing PSMA-positive LNCaP human xenografts with EC1584 () and
EC1588 (.tangle-solidup.), each at 2 .mu.mol/kg, TIW, 2 weeks,
leads to complete responses in all tested animals. Each compound
was compared against vehicle-treated controls ( ). It was also
surprisingly found that treatment with EC1588 leads to cure in 3/7
tested animals.
Example
[0365] Gross toxicity of compounds described herein. As shown in
FIG. 8, the observed efficacy of EC1584 () and EC1588
(.tangle-solidup.) occurred in the absence of weight loss or major
organ tissue degeneration.
Example
[0366] Activity of compounds described herein against PSMA+ tumors
compared to conventional chemotherapeutic agents. As shown in FIG.
9, treatment of LNCaP-tumor bearing mice with docetaxel (the most
active chemotherapeutic agent approved for prostate cancer) at 10
mg/kg, BIW, 2 weeks, MTD (), was found to produce only modest
anti-tumor activity, and showed only 1/4 cures, even when
administered at its MTD. In addition, as shown in FIG. 10, that
modest observed docetaxel efficacy was accompanied by high gross
toxicity, as evidenced by severe weight loss (18%). EC1169,
administered at 2 .mu.mol/kg, TIW, 2 weeks ( ), is more active and
less toxic than docetaxel against PSMA+ LNCaP tumors. FIG. 9 shows
that treatment with EC1169 leads to a complete response in all test
animals, and resulted in cures. FIG. 10 also shows that the higher
efficacy displayed by EC1169 was not accompanied by substantially
lower toxicity than docetaxcl, providing a significantly wider
therapeutic window. The efficacy of each compound was compared to
vehicle-treated control (.box-solid.).
Example
[0367] The in vivo efficacy of (.box-solid.) EC1718;
(.tangle-solidup.) EC1720; () EC1721; (.diamond-solid.) EC1719; and
(.smallcircle.) EC11677; compared to ( ) untreated control is shown
in FIG. 11. All compounds were administered at 2 .mu.mol/kg, TIW
for 2 weeks, beginning on day 21 post tumor implant (PTI). The
dotted line indicates the final treatment day. The data indicate
that the compounds described herein are efficacious in decreasing
tumor growth in vivo compared to untreated animals. In addition,
(.box-solid.) EC1718 lead to 1/7 cures; () EC1721 lead to 1/7
cures; (.diamond-solid.) EC1719 lead to 2/7 cures; and
(.smallcircle.) EC11677 lead to 4/7 cures, where regrowth of the
tumor in those animals was not observed during the observation
period. In addition, the compounds described herein do not show
gross toxicity to the test animals, as shown in FIG. 12. Without
being bound by theory, it is believed herein that the weight change
observed in FIG. 12 for EC1718 at about day 81 is due to the
effects of the tumor size.
Example
[0368] Specificity of compounds described herein. PSMA-negative KB
tumors did not appreciably respond to EC1169 therapy, supporting
the conclusion that the compounds described herein exhibit target
specificity for PSMA-expressing cells.
Example
[0369] Hematological Toxicity. Conjugates described herein
demonstrate significantly improved hematological toxicity. EC1169,
EC1584, and EC1588 were administered to rats i.v. at 0.33 and 0.51
.mu.mol/kg, twice per week (BIW), for 2 weeks. The hematological
toxicity in red blood cells and white blood cells was significantly
lower than untreated controls.
* * * * *