U.S. patent application number 11/350308 was filed with the patent office on 2006-12-07 for porphyrin-polyamine conjugates for cancer therapy.
Invention is credited to Hirak S. Basu, Subhra Bhattacharya, Andrei V. Blokhin, Linda Clifford, Benjamin Frydman, Laurence J. Marton, Venodhar K. Reddy, Aparajita Sarkar, Aldonia L. Valasinas, Yu Wang.
Application Number | 20060276444 11/350308 |
Document ID | / |
Family ID | 34192928 |
Filed Date | 2006-12-07 |
United States Patent
Application |
20060276444 |
Kind Code |
A1 |
Frydman; Benjamin ; et
al. |
December 7, 2006 |
Porphyrin-polyamine conjugates for cancer therapy
Abstract
Porphyrin-polyamine conjugate compounds are disclosed which have
anticancer and antitumor effects. The porphyrin moiety selectively
localizes in tumors, while the polyamine moiety serves as a
cytotoxic agent. Methods of making and using the
porphyrin-polyamine conjugate compounds are also disclosed.
Inventors: |
Frydman; Benjamin; (Madison,
WI) ; Clifford; Linda; (Madison, WI) ;
Valasinas; Aldonia L.; (Buenos Aires, AR) ; Reddy;
Venodhar K.; (Madison, WI) ; Basu; Hirak S.;
(Madison, WI) ; Sarkar; Aparajita; (Madison,
WI) ; Bhattacharya; Subhra; (Madison, WI) ;
Wang; Yu; (Madison, WI) ; Marton; Laurence J.;
(Palo Alto, CA) ; Blokhin; Andrei V.; (Madison,
WI) |
Correspondence
Address: |
MORRISON & FOERSTER LLP
755 PAGE MILL RD
PALO ALTO
CA
94304-1018
US
|
Family ID: |
34192928 |
Appl. No.: |
11/350308 |
Filed: |
February 7, 2006 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10606016 |
Jun 24, 2003 |
7026347 |
|
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11350308 |
Feb 7, 2006 |
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60392171 |
Jun 26, 2002 |
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Current U.S.
Class: |
514/185 ;
514/410 |
Current CPC
Class: |
C07D 487/22 20130101;
A61P 35/00 20180101 |
Class at
Publication: |
514/185 ;
514/410 |
International
Class: |
A61K 31/409 20060101
A61K031/409; A61K 31/555 20060101 A61K031/555 |
Claims
1. A composition comprising a compound according to the formula
##STR15## wherein at least one of J.sub.1, J.sub.2, J.sub.3,
J.sub.4, J.sub.5, J.sub.6, J.sub.7 and J.sub.8 is independently
selected from the group consisting of
--(B-A-B).sub.x-G-(B-A-B).sub.m--(N(P)--B-A-B).sub.n--K. wherein
each A is independently selected from the group consisting of: a
nonentity, C.sub.1-C.sub.12 alkyl, C.sub.2-C.sub.12 alkenyl,
C.sub.2-C.sub.12 alkynyl, C.sub.3-C.sub.12 cycloalkyl,
C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.12 cycloalkenyl,
C.sub.3-C.sub.12 cycloalkynyl, C.sub.1-C.sub.12 alkanol,
C.sub.3-C.sub.12 cycloalkanol, and C.sub.3-C.sub.8 hydroxyaryl;
each B is independently selected from the group consisting of: a
nonentity, C.sub.1-C.sub.12 alkyl, C.sub.2-C.sub.12 alkenyl,
C.sub.2-C.sub.12 alkynyl, C.sub.3-C.sub.12 cycloalkyl,
C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.12 cycloalkenyl,
C.sub.3-C.sub.12 cycloalkynyl, C.sub.1-C.sub.12 alkanol,
C.sub.3-C.sub.12 cycloalkanol, and C.sub.3-C.sub.8 hydroxyaryl; and
with the proviso that each --B-A-B-- unit contain at least one
carbon atom; wherein G is independently selected from the group
consisting of --N(P)--, --(C.dbd.O)--N(P)--, --N(P)--(C.dbd.O)--,
and a nonentity; x is independently 0 or 1; m is independently 0 or
1; n is independently an integer from 0 to 20; each P is
independently selected from the group consisting of H and
C.sub.1-C.sub.12 alkyl; each K is independently selected from the
group consisting of H, C.sub.1-C.sub.12 alkyl, C.sub.2-C.sub.12
alkenyl, C.sub.2-C.sub.12 alkynyl, C.sub.3-C.sub.12 cycloalkyl,
C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.12 cycloalkenyl,
C.sub.3-C.sub.12 cycloalkynyl, C.sub.1-C.sub.12 alkanol,
C.sub.3-C.sub.12 cycloalkanol, and C.sub.3-C.sub.8 hydroxyaryl, and
Q; where each Q is independently selected from the group consisting
of ##STR16## where each P is independently selected from the group
consisting of H and C.sub.1-C.sub.12 alkyl, each D is selected from
the group consisting of H and C.sub.1-C.sub.32 alkyl, y is an
integer from 1 to 8, and z is an integer from 0 to 5, and where the
Q moiety is attached to the remainder of the molecule at any C or N
atom in the Q moiety (including C atoms in the D or P moieties) by
removing a hydrogen atom, a P substituent, or a D substituent of
the Q moiety to form an open valence for attachment to the
remainder of the molecule; and where the remaining members or
member of J.sub.1, J.sub.2, J.sub.3, J.sub.4, J.sub.5, J.sub.6,
J.sub.7 and J.sub.8 are each independently selected from the group
consisting of H, --B-A-B, --COOH, --SO.sub.3H, --B-A-B--COOH, or
--B-A-B--SO.sub.3H, where each A and each B are independently
selected as defined above and with the proviso that each --B-A-B--
unit has at least one carbon atom.
2. A composition comprising a compound according to the formula
##STR17## wherein at least one of J.sub.1, J.sub.2, J.sub.3,
J.sub.4, J.sub.5, J.sub.6, J.sub.7 and J.sub.8 is independently M,
where each M is independently selected from the group consisting of
--(B-A-B).sub.x-G-(B-A-B).sub.m--(N(P)--B-A-B).sub.n--K wherein
each A is independently selected from the group consisting of: a
nonentity, C.sub.1-C.sub.12 alkyl, C.sub.2-C.sub.12 alkenyl,
C.sub.2-C.sub.12 alkynyl, C.sub.3-C.sub.12 cycloalkyl,
C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.12 cycloalkenyl,
C.sub.3-C.sub.12 cycloalkynyl, C.sub.1-C.sub.12 alkanol,
C.sub.3-C.sub.12 cycloalkanol, and C.sub.3-C.sub.8 hydroxyaryl;
each B is independently selected from the group consisting of: a
nonentity, C.sub.1-C.sub.12 alkyl, C.sub.2-C.sub.12 alkenyl,
C.sub.2-C.sub.12 alkynyl, C.sub.3-C.sub.12 cycloalkyl,
C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.12 cycloalkenyl,
C.sub.3-C.sub.12 cycloalkynyl, C.sub.1-C.sub.12 alkanol,
C.sub.3-C.sub.12 cycloalkanol, and C.sub.3-C.sub.8 hydroxyaryl; and
with the proviso that each --B-A-B-- unit contain at least one
carbon atom; wherein G is independently selected from the group
consisting of --N(P)--, --(C.dbd.O)--N(P)--, --N(P)--(C.dbd.O)--,
and a nonentity; x is independently 0 or 1; m is independently 0 or
1; n is independently an integer from 0 to 20; each P is
independently selected from the group consisting of H and
C.sub.1-C.sub.12 alkyl; each K is independently selected from the
group consisting of H, C.sub.1-C.sub.12 alkyl, C.sub.2-C.sub.12
alkenyl, C.sub.2-C.sub.12 alkynyl, C.sub.3-C.sub.12 cycloalkyl,
C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.12 cycloalkenyl,
C.sub.3-C.sub.12 cycloalkynyl, C.sub.1-C.sub.12 alkanol,
C.sub.3-C.sub.12 cycloalkanol, and C.sub.3-C.sub.8 hydroxyaryl, and
Q; where each Q is independently selected from the group consisting
of ##STR18## where each P is independently selected from the group
consisting of H and C.sub.1-C.sub.12 alkyl, each D is selected from
the group consisting of H and C.sub.1-C.sub.32 alkyl, y is an
integer from 1 to 8, and z is an integer from 0 to 5, and where the
Q moiety is attached to the remainder of the molecule at any C or N
atom in the Q moiety (including C atoms in the D or P moieties) by
removing a hydrogen atom, a P substituent, or a D substituent of
the Q moiety to form an open valence for attachment to the
remainder of the molecule; and where the remaining members or
member of J.sub.1, J.sub.2, J.sub.3, J.sub.4, J.sub.5, J.sub.6,
J.sub.7 and J.sub.8 are each independently selected from the group
consisting of H, --B-A-B, --COOH, --SO.sub.3H, --B-A-B--COOH, or
--B-A-B--SO.sub.3H, where each A and each B are independently
selected as defined above and with the proviso that each --B-A-B--
unit has at least one carbon atom; with the proviso that M excludes
moieties of the form
--K.sub.1-G.sub.5-L.sub.5-(N(P.sub.5)-A.sub.5).sub.n-K.sub.2 where
K.sub.1 is independently selected from the group consisting of
C.sub.1-C.sub.8 alkyl and where the valence to the left of K.sub.1
attaches to the porphyrin ring; G.sub.5 is --O--, --(C.dbd.O)--,
--C(.dbd.O)--O--, --O--(C.dbd.O)--, --O--(C.dbd.O)--O--,
--O--(C.dbd.O)--N--, --N--(C.dbd.O)--O--, or a nonentity; L.sub.5
is C.sub.1-C.sub.8 alkyl, C.sub.3-C.sub.8 cycloalkyl,
C.sub.3-C.sub.8 cycloaryl, C.sub.1-C.sub.8 alkoxy, C.sub.1-C.sub.8
alkyl-C.sub.3-C.sub.8 cycloalkyl, C.sub.1-C.sub.8
alkyl-C.sub.3-C.sub.8 cycloaryl, C.sub.1-C.sub.8
alkoxy-C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.8
cycloalkyl-C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.8
cycloalkyl-C.sub.1-C.sub.8 alkyl, C.sub.3-C.sub.8
cycloaryl-C.sub.1-C.sub.8 alkyl, C.sub.3-C.sub.8
cycloaryl-C.sub.1-C.sub.8 alkoxy, C.sub.3-C.sub.8
cycloaryl-C.sub.3-C.sub.8 cycloalkyl, or a nonentity; each A.sub.5
is independently selected from the group consisting of
C.sub.1-C.sub.8 alkyl, C.sub.2-C.sub.8 alkenyl, C.sub.2-C.sub.8
alkynyl, C.sub.3-C.sub.8 cycloalkyl, C.sub.3-C.sub.8 cycloaryl,
C.sub.3-C.sub.8 cycloalkenyl, and C.sub.3-C.sub.8 cycloalkynyl;
P.sub.5 is selected from the group consisting of H and
C.sub.1-C.sub.8 alkyl; n is an integer from 2 to 8; and K.sub.2 is
independently selected from the group consisting of H,
C.sub.1-C.sub.8 alkyl, C.sub.2-C.sub.8 alkenyl, C.sub.2-C.sub.8
alkynyl, C.sub.3-C.sub.8 cycloalkyl, C.sub.3-C.sub.8 cycloaryl,
C.sub.3-C.sub.8 cycloalkenyl, C.sub.3-C.sub.8 cycloalkynyl,
C.sub.1-C.sub.8 alkanol, C.sub.3-C.sub.8 cycloalkanol, and
C.sub.3-C.sub.8 hydroxyaryl.
3. The composition of claim 2, where G is independently selected
from --(C.dbd.O)--N(P)-- and --N(P)--(C.dbd.O)--.
4. The composition of claim 2, where the Q moiety is attached to
the remainder of the molecule at any N atom in the Q moiety by
removing a P substituent of the Q moiety to form an open valence
for attachment to the remainder of the molecule.
5. The composition of claim 2, wherein each A and B substituent, if
present, is selected from C.sub.1-C.sub.12 alkyl.
6. The composition of claim 2, wherein at least one A substituent
comprises a cyclopropane group.
7. The composition of claim 2 comprising a compound of the formula
##STR19## where J.sub.1 and J.sub.2 are independently M and each M
is independently selected from the group consisting of
--(B-A-B).sub.x-G-(B-A-B).sub.m--(N(P)--B-A-B).sub.n--K; J.sub.3,
J.sub.4, J.sub.6 and J.sub.8 are independently selected from methyl
and ethyl; and J.sub.5 and J.sub.7 are independently selected from
methyl, ethyl, and --SO.sub.3H.
8. The composition of claim 7, where J.sub.1 and J.sub.2 are
independently M and each M is independently selected from the group
consisting of --(B-A-B)-G-(B-A-B)--(N(P)--B-A-B).sub.n--K.
9. The composition of claim 8, wherein at least one B-A-B unit
comprises a cycloalkyl moiety.
10. The composition of claim 9, wherein at least one B-A-B unit
comprises a cyclopropyl moiety.
11. The composition of claim 7, where J.sub.1 and J.sub.2 are
independently M and each M is independently selected from the group
consisting of --C.sub.1-C.sub.12 alkyl-G-C.sub.1-C.sub.12
alkyl-(N(P)--B-A-B).sub.n--K.
12. The composition of claim 7, where J.sub.1 and J.sub.2 are
independently M and each M is independently selected from the group
consisting of --C.sub.1-C.sub.12
alkyl-(C.dbd.O)--N(P)--C.sub.1-C.sub.12
alkyl-(N(P)--B-A-B).sub.n--K.
13. The composition of claim 7, where J.sub.1 and J.sub.2 are
independently M and each M is independently selected from the group
consisting of
--(CH.sub.2).sub.2C(.dbd.O)N(P.sub.2)--C.sub.1-C.sub.4
alkyl-[NH(CH.sub.2CH.sub.2CH.sub.2CH.sub.2)].sub.fC.sub.1-C.sub.12
alkyl, where P.sub.2 is H, methyl, or ethyl, and f is an integer
from 1 to 10.
14. The composition of claim 10, where J.sub.1 and J.sub.2 are
identical.
15. The composition of claim 11, where J.sub.1 and J.sub.2 are
identical.
16. The composition of claim 1, wherein each --K is independently
Q; where each Q is independently selected from the group consisting
of ##STR20## wherein only one D moiety is selected from the group
consisting of C.sub.1-C.sub.32 alkyl and all remaining D moieties
are H; wherein three P groups are selected from the group
consisting of --H and --CH.sub.3; wherein the fourth P group is
absent and the Q moiety is attached to the remainder of the
molecule at that valence; and wherein y is 2, 3, or 4 and z is 0,
1; or 2.
17. The composition of claim 2, wherein each --K is independently
Q; where each Q is independently selected from the group consisting
of ##STR21## wherein only one D moiety is selected from the group
consisting of C.sub.1-C.sub.32 alkyl and all remaining D moieties
are H; wherein three P groups are selected from the group
consisting of --H and --CH.sub.3; wherein the fourth P group is
absent and the Q moiety is attached to the remainder of the
molecule at that valence; and wherein y is 2, 3, or 4 and z is 0,
1, or 2.
18. The composition of claim 6, wherein each --K is independently
Q; where each Q is independently selected from the group consisting
of ##STR22## wherein only one D moiety is selected from the group
consisting of C.sub.1-C.sub.32 alkyl and all remaining D moieties
are H; wherein three P groups are selected from the group
consisting of --H and --CH.sub.3; wherein the fourth P group is
absent and the Q moiety is attached to the remainder of the
molecule at that valence; and wherein y is 2, 3, or 4 and z is 0,
1, or 2.
19. The composition of claim 1, wherein --K is ##STR23##
20. The composition of claim 2, wherein --K is ##STR24##
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority benefit of U.S. Provisional
Patent Application No. 60/392,171, filed Jun. 26, 2002. The content
of that application is hereby incorporated by reference herein in
its entirety.
STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT
[0002] Not applicable.
REFERENCE TO AN APPENDIX
[0003] Not applicable.
TECHNICAL FIELD
[0004] This invention relates to porphyrin-polyamine conjugate
compounds used for treatment of cancer and other diseases.
BACKGROUND OF THE INVENTION
[0005] Cancer is the third most common cause of death in the world
according to the World Health Organization, after heart disease and
infectious disease. Cancer is the second most common cause of death
(after heart disease) in the developed world. Accordingly,
discovery of new and effective treatments for cancer is a high
priority for health care researchers.
[0006] Cancer is often treated by using chemotherapy to selectively
kill or hinder the growth of cancer cells, while having a less
deleterious effect on normal cells. Chemotherapeutic agents often
kill rapidly dividing cells, such as cancer cells; non-malignant
cells which are dividing less rapidly are affected to a lesser
degree. Other agents, such as antibodies attached to toxic agents,
have been evaluated for use against cancers. These agents target
the cancer cells by making use of a characteristic specific to the
cancer, for example, higher-than-normal rates of cell division, or
unique antigens expressed on the cancer cell surface.
[0007] As toxic agents specifically targeted against cancer cells
can enhance therapeutic efficacy, reduce undesirable side effects,
or both, many efforts have been made to achieve selective
localization of well-defined chemical materials in malignant
tumors. A significant advance in the field occurred with the
introduction of tetraphenylporphine sulfonates (TPPS), which are
non-naturally occurring porphyrins (Winkelman J. (1962) Cancer Res.
22:589). A hematoporphyrin derivative (HPD) was also found to
localize in tumors (Lipson R L, Baldes, E J, & Gray M S (1967)
Cancer 20: 2255). HPD is a complex mixture of porphyrins currently
used as a sensitizer derivative that concentrates in tumor cells
and destroys them after the tumor is irradiated with light or a
laser beam (Dougherty T J, (1987) Photochem. Photobiol. 45:879). A
wide variety of porphyrins and porphyrin analogues have been found
to be selectively taken up by tumors, such as the naturally
occurring porphyrins; for example, the octacarboxylic
uroporphyrins, the tetracarboxylic coproporphyrins, and the
dicarboxylic protoporphyrin. Synthetic porphyrins are also
selectively taken up by tumors; among them are the meso-tetraphenyl
porphyrins and the different porphyrin sulfonates TPPS.sub.4,
TPPS.sub.3, TPPS.sub.2a and TPPS.sub.1, which are listed in order
of decreasing number of sulfonic acid substituents and decreasing
hydrophilicity. Many factors determine the uptake and concentration
of porphyrins in the tumors; one important factor is the structure
(hydrophobicity, size, polarity) of the drug; another important
factor is the formulation in which it is delivered (Sternberg E and
Dolphin D (1996) Current Med Chemistry 3, 239). The mechanism(s) of
porphyrin localization in tumors is still not entirely clear; the
more hydrophobic porphyrins are preferentially incorporated in the
lipid core of lipoproteins. Tightly aggregated porphyrins circulate
as unbound pseudomicellar structures which can be entrapped in the
interstitial regions of the tumor, can be localized in macrophages,
or can enter neoplastic cells via pinocytotic processes. Low
density lipoproteins (LDL), which are endocytosed by neoplastic
cells through a specific receptor-mediated pathway, display the
most selective release of porphyrins into the tumors (Jori G (1989)
Photosensitizing Compounds, Ciba Foundation Symp 146, pp
78-94).
[0008] The present invention describes the synthesis and cytotoxic
actions of porphyrin-polyamine conjugates. They are taken up by the
tumor cells due to their porphyrin moiety, while the polyamine
moiety provides the cytotoxic effects (see International Patent
Application Nos. WO 00/66587 and WO 02/10142, and U.S. Pat. Nos.
6,392,098, 5,889,061, and 5,677,350).
SUMMARY OF THE INVENTION
[0009] The invention provides porphyrin-polyamine conjugate
compounds and compositions comprising such compounds.
[0010] In one embodiment, the invention embraces a composition
comprising a compound according to the formula ##STR1##
[0011] wherein at least one of J.sub.1, J.sub.2, J.sub.3, J.sub.4,
J.sub.5, J.sub.6, J.sub.7 and J.sub.8 is independently M, where M
is selected from the group consisting of
--(B-A-B).sub.x-G-(B-A-B).sub.m--(N(P)--B-A-B).sub.n--K
[0012] wherein each A is independently selected from the group
consisting of: a nonentity, C.sub.1-C.sub.12 alkyl,
C.sub.2-C.sub.12 alkenyl, C.sub.2-C.sub.12 alkynyl,
C.sub.3-C.sub.12 cycloalkyl, C.sub.3-C.sub.8 cycloaryl,
C.sub.3-C.sub.12 cycloalkenyl, C.sub.3-C.sub.12 cycloalkynyl,
C.sub.1-C.sub.12 alkanol, C.sub.3-C.sub.12 cycloalkanol, and
C.sub.3-C.sub.8 hydroxyaryl;
[0013] each B is independently selected from the group consisting
of: a nonentity, C.sub.1-C.sub.12 alkyl, C.sub.2-C.sub.12 alkenyl,
C.sub.2-C.sub.12 alkynyl, C.sub.3-C.sub.12 cycloalkyl,
C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.12 cycloalkenyl,
C.sub.3-C.sub.12 cycloalkynyl, C.sub.1-C.sub.12 alkanol,
C.sub.3-C.sub.12 cycloalkanol, and C.sub.3-C.sub.8 hydroxyaryl;
[0014] and with the proviso that each --B-A-B-- unit contain at
least one carbon atom;
[0015] wherein G is independently selected from the group
consisting of --N(P)--, --(C.dbd.O)--N(P)--, --N(P)--(C.dbd.O)--,
and a nonentity;
[0016] x is independently 0 or 1;
[0017] m is independently 0 or 1;
[0018] n is independently an integer from 0 to 20;
[0019] each P is independently selected from the group consisting
of H and C.sub.1-C.sub.12 alkyl;
[0020] K is independently selected from the group consisting of H,
C.sub.1-C.sub.12 alkyl, C.sub.2-C.sub.12 alkenyl, C.sub.2-C.sub.12
alkynyl, C.sub.3-C.sub.12 cycloalkyl, C.sub.3-C.sub.8 cycloaryl,
C.sub.3-C.sub.12 cycloalkenyl, C.sub.3-C.sub.12 cycloalkynyl,
C.sub.1-C.sub.12 alkanol, C.sub.3-C.sub.12 cycloalkanol, and
C.sub.3-C.sub.8 hydroxyaryl, and Q;
[0021] where each Q is independently selected from the group
consisting of ##STR2## where each P is independently selected from
the group consisting of H and C.sub.1-C.sub.12 alkyl, each D is
selected from the group consisting of H and C.sub.1-C.sub.32 alkyl,
y is an integer from 1 to 8, and z is an integer from 0 to 5, and
where the Q moiety is attached to the remainder of the molecule at
any C or N atom in the Q moiety (including C atoms in the D or P
moieties) by removing a hydrogen atom, a P substituent, or a D
substituent of the Q moiety to form an open valence for attachment
to the remainder of the molecule;
[0022] and where the remaining members or member of J.sub.1,
J.sub.2, J.sub.3, J.sub.4, J.sub.5, J.sub.6, J.sub.7 and J.sub.8
are each independently selected from the group consisting of H,
--B-A-B, --COOH, --SO.sub.3H, --B-A-B--COOH, or --B-A-B--SO.sub.3H,
where each A and each B are independently selected as defined above
and with the proviso that each --B-A-B-- unit has at least one
carbon atom.
[0023] In another embodiment, M excludes moieties of the form
--K.sub.1-G.sub.5-L.sub.5-(N(P.sub.5)-A.sub.5).sub.n-K.sub.2 where
K.sub.1 is independently selected from the group consisting of
C.sub.1-C.sub.8 alkyl and where the valence to the left of K,
attaches to the porphyrin ring; G.sub.5 is --O--, --(C.dbd.O)--,
--C(.dbd.O)--O--, --O--(C.dbd.O)--, --O--(C.dbd.O)--O--,
--O--(C.dbd.O)--N--, --N--(C.dbd.O)--O--, or a nonentity; L.sub.5
is C.sub.1-C.sub.8 alkyl, C.sub.3-C.sub.8 cycloalkyl,
C.sub.3-C.sub.8 cycloaryl, C.sub.1-C.sub.8 alkoxy, C.sub.1-C.sub.8
alkyl-C.sub.3-C.sub.8 cycloalkyl, C.sub.1-C.sub.8
alkyl-C.sub.3-C.sub.8 cycloaryl, C.sub.1-C.sub.8
alkoxy-C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.8
cycloalkyl-C.sub.3-C.sub.8 cycloaryl, C.sub.3-C.sub.8
cycloalkyl-C.sub.1-C.sub.8 alkyl, C.sub.3-C.sub.8
cycloaryl-C.sub.1-C.sub.8 alkyl, C.sub.3-C.sub.8
cycloaryl-C.sub.1-C.sub.8 alkoxy, C.sub.3-C.sub.8
cycloaryl-C.sub.3-C.sub.8 cycloalkyl, or a nonentity; each A.sub.5
is independently selected from the group consisting of
C.sub.1-C.sub.8 alkyl, C.sub.2-C.sub.8 alkenyl, C.sub.2-C.sub.8
alkynyl, C.sub.3-C.sub.8 cycloalkyl, C.sub.3-C.sub.8 cycloaryl,
C.sub.3-C.sub.8 cycloalkenyl, and C.sub.3-C.sub.8 cycloalkynyl;
P.sub.5 is selected from the group consisting of H and
C.sub.1-C.sub.8 alkyl; n is an integer from 2 to 8; and K.sub.2 is
independently selected from the group consisting of H,
C.sub.8-C.sub.8 alkyl, C.sub.2-C.sub.8 alkenyl, C.sub.2-C.sub.8
alkynyl, C.sub.3-C.sub.8 cycloalkyl, C.sub.3-C.sub.8 cycloaryl,
C.sub.3-C.sub.8 cycloalkenyl, C.sub.3-C.sub.8 cycloalkynyl,
C.sub.1-C.sub.8 alkanol, C.sub.3-C.sub.8 cycloalkanol, and
C.sub.3-C.sub.8 hydroxyaryl.
[0024] In another embodiment, G is independently selected from
--(C.dbd.O)--N(P)-- and --N(P)--(C.dbd.O)--. In another embodiment,
the Q moiety is attached to the remainder of the molecule at any N
atom in the Q moiety by removing a P substituent of the Q moiety to
form an open valence for attachment to the remainder of the
molecule. In another embodiment, each A and B substituent, if
present, is selected from C.sub.1-C.sub.12 alkyl. In another
embodiment, at least one A substituent comprises a cyclopropane
group.
[0025] In another embodiment, the invention embraces a composition
comprising a compound according to the formula ##STR3## where
J.sub.1 and J.sub.2 are independently
--(B-A-B).sub.x-G-(B-A-B).sub.m--(N(P)--B-A-B).sub.n--K; J.sub.3,
J.sub.4, J.sub.6 and J.sub.8 are independently selected from methyl
and ethyl; and J.sub.5 and J.sub.7 are independently selected from
methyl, ethyl, and --SO.sub.3H. In another embodiment, J.sub.1 and
J.sub.2 are independently
--(B-A-B)-G-(B-A-B)--(N(P)--B-A-B).sub.n--K. In another embodiment,
at least one B-A-B unit comprises a cycloalkyl moiety, such as a
cyclopropyl moiety. In another embodiment, J.sub.1 and J.sub.2 are
independently --C.sub.1-C.sub.12 alkyl-G-C.sub.1-C.sub.12
alkyl-(N(P)--B-A-B).sub.n--K. In another embodiment, J.sub.1 and
J.sub.2 are independently --C.sub.1-C.sub.12
alkyl-(C.dbd.O)--N(P)--C.sub.1-C.sub.12
alkyl-(N(P)--B-A-B).sub.n--K. In another embodiment, J.sub.1 and
J.sub.2 are independently
--(CH.sub.2).sub.2C(.dbd.O)N(P.sub.2)--C.sub.1-C.sub.4
alkyl-[NH(CH.sub.2CH.sub.2CH.sub.2CH.sub.2)].sub.fC.sub.1-C.sub.12
alkyl, where P.sub.2 is H, methyl, or ethyl, and f is an integer
from 1 to 10.
[0026] In still further embodiments, J.sub.1 and J.sub.2 are
identical.
[0027] In still further embodiments, whenever any embodiment
comprises a Q moiety (that is, whenever any K is Q), only one D
moiety is selected from the group consisting of C.sub.1-C.sub.32
alkyl and all remaining D moieties are H; three P groups are
selected from the group consisting of --H and --CH.sub.3 and the
fourth P group is absent and the Q moiety is attached to the
remainder of the molecule at that valence; and y is 2, 3, or 4 and
z is 0, 1, or 2.
[0028] In further embodiments, whenever any embodiment comprises a
Q moiety, Q can be ##STR4##
BRIEF DESCRIPTION OF THE DRAWING(S)
[0029] FIG. 1. is a graph depicting the in vitro effects of
increasing concentrations of SL-11209 on the growth of cultured
human prostate cancer cells DUPRO.
[0030] FIG. 2. is a graph depicting the in vitro effects of
increasing concentrations of SL-11211 on the growth of cultured
human prostate cancer cells DUPRO.
[0031] FIG. 3. is a graph depicting the in vitro effects of
increasing concentrations of SL-11209 on the survival of cultured
human prostate cancer cells DUPRO after 5 days of treatment.
[0032] FIG. 4. is a graph depicting the in vitro effects of
increasing concentrations of SL-11211 on the survival of cultured
human prostate cancer cells DUPRO after 3 days of treatment.
[0033] FIG. 5. is a graph depicting the in vitro effects of
increasing concentrations of SL-11211 on the survival of cultured
human prostate cancer cells DUPRO after 5 days of treatment.
[0034] FIG. 6. is a graph depicting the in vitro effects of
increasing concentrations of SL-11217 on the survival of cultured
human prostate cancer cells DUPRO after 3 and 5 days of
treatment.
[0035] FIG. 7. is a graph depicting the in vitro effects of
increasing concentrations of SL-11211 on the survival of cultured
human prostate cancer cells PC3 after 5 days of treatment.
[0036] FIG. 8. is a graph depicting the in vitro effects of
increasing concentrations of SL-11217 on the survival of cultured
human prostate cancer cells PC3 after 5 days of treatment.
[0037] FIG. 9. is a graph depicting the in vitro effects of
increasing concentrations of SL-11237 on the survival of cultured
human prostate cancer cells PC3 after 5 days of treatment.
[0038] FIG. 10. is a graph depicting the in vitro effects of 10
.mu.M SL-11217 and SL-11237 on the growth of cultured human
pancreatic cancer cells BxPC3.
[0039] FIG. 11. is a graph depicting the in vitro effects of 10
.mu.M SL-11217 and SL-11237 on the growth of cultured human
pancreatic cancer cells Panc1.
[0040] FIG. 12. is a graph depicting the in vitro effects of
increasing concentrations of SL-11217 on the survival of cultured
human brain tumor cells U251MG NCI after 3 days of treatment.
[0041] FIG. 13. is a graph depicting the in vitro effects of
increasing concentrations of SL-11237 on the survival of cultured
human brain tumor cells U251MG NCI after 3 days of treatment.
[0042] FIG. 14 depicts the effects of SL-11237 via oral
administration. Male athymic nude mice were given subcutaneous
injections of 0.75.times.10.sup.6 DU145 cells on Day 0. Beginning
on Day 10, mice were treated once weekly for 3 weeks with acidified
water, 100 mg/kg, or 500 mg/kg of SL-11237 via oral gavage at 10
ml/kg dosing volume (the third treatment was actually 400 mg/kg in
the high dose group). The top panel depicts average tumor volume in
the mice. The bottom panel depicts average body weight of the
mice.
DETAILED DESCRIPTION OF THE INVENTION
[0043] The invention is directed to various novel
porphyrin-polyamine conjugate compounds and compositions containing
them as described herein. The invention includes all salts of the
compounds described herein. Particularly preferred are
pharmaceutically acceptable salts. Pharmaceutically acceptable
salts are those salts which retain the biological activity of the
free bases and which are not biologically or otherwise undesirable.
The desired salt may be prepared by methods known to those of skill
in the art by treating the compound with an acid. Examples of
inorganic acids include, but are not limited to, hydrochloric acid,
hydrobromic acid, sulfuric acid, nitric acid, and phosphoric acid.
Examples of organic acids include, but are not limited to, formic
acid, acetic acid, propionic acid, glycolic acid, pyruvic acid,
oxalic acid, maleic acid, malonic acid, succinic acid, fumaric
acid, tartaric acid, citric acid, benzoic acid, cinnamic acid,
mandelic acid, sulfonic acids, and salicylic acid. Salts of the
compounds with amino acids, such as aspartate salts and glutamate
salts, can also be prepared.
[0044] The invention also includes all stereoisomers of the
compounds, including diastereomers and enantiomers, as well as
mixtures of stereoisomers, including, but not limited to, racemic
mixtures. Unless stereochemistry is explicitly indicated in a
structure, the structure is intended to embrace all possible
stereoisomers of the compound depicted.
[0045] The term "alkyl" refers to saturated aliphatic groups
including straight-chain, branched-chain, cyclic groups, and
combinations thereof, having the number of carbon atoms specified,
or if no number is specified, having up to 12 carbon atoms.
"Straight-chain alkyl" or "linear alkyl" groups refers to alkyl
groups that are neither cyclic nor branched, commonly designated as
"n-alkyl" groups. Examples of alkyl groups include, but are not
limited to, groups such as methyl, ethyl, n-propyl, isopropyl,
butyl, n-butyl, isobutyl, sec-butyl, t-butyl, pentyl, n-pentyl,
hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, neopentyl,
cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and adamantyl.
Cyclic groups can consist of one ring, including, but not limited
to, groups such as cycloheptyl, or multiple fused rings, including,
but not limited to, groups such as adamantyl or norbornyl.
Preferred subsets of alkyl groups include C.sub.1-C.sub.12,
C.sub.1-C.sub.10, C.sub.1-C.sub.8, C.sub.1-C.sub.6,
C.sub.1-C.sub.4, C.sub.1-C.sub.2, C.sub.3-C.sub.4, C.sub.3, and
C.sub.4 alkyl groups.
[0046] "Substituted alkyl" refers to alkyl groups substituted with
one or more substituents including, but not limited to, groups such
as halogen (fluoro, chloro, bromo, and iodo), alkoxy, acyloxy,
amino, hydroxyl, mercapto, carboxy, benzyloxy, phenyl, benzyl,
cyano, nitro, thioalkoxy, carboxaldehyde, carboalkoxy and
carboxamide, or a functionality that can be suitably blocked, if
necessary for purposes of the invention, with a protecting group.
Examples of substituted alkyl groups include, but are not limited
to, --CF.sub.3, --CF.sub.2--CF.sub.3, and other perfluoro and
perhalo groups.
[0047] "Hydroxyalkyl" specifically refers to alkyl groups having
the number of carbon atoms specified substituted with one --OH
group. Thus, "C.sub.3 linear hydroxyalkyl" refers to
--CH.sub.2CH.sub.2CHOH--, --CH.sub.2CHOHCH.sub.2--, and
--CHOHCH.sub.2CH.sub.2--.
[0048] The term "alkenyl" refers to unsaturated aliphatic groups
including straight-chain (linear), branched-chain, cyclic groups,
and combinations thereof, having the number of carbon atoms
specified, or if no number is specified, having up to 12 carbon
atoms, which contain at least one double bond (--C.dbd.C--).
Examples of alkenyl groups include, but are not limited to,
--CH.sub.2--CH.dbd.CH--CH.sub.3; and
--CH.sub.2--CH.sub.2-cyclohexenyl, where the ethyl group can be
attached to the cyclohexenyl moiety at any available carbon
valence. The term "alkynyl" refers to unsaturated aliphatic groups
including straight-chain (linear), branched-chain, cyclic groups,
and combinations thereof, having the number of carbon atoms
specified, or if no number is specified, having up to 12 carbon
atoms, which contain at least one triple bond (--C.ident.C--).
"Hydrocarbon chain" or "hydrocarbyl" refers to any combination of
straight-chain, branched-chain, or cyclic alkyl, alkenyl, or
alkynyl groups, and any combination thereof. "Substituted alkenyl,"
"substituted alkynyl," and "substituted hydrocarbon chain" or
"substituted hydrocarbyl" refer to the respective group substituted
with one or more substituents, including, but not limited to,
groups such as halogen, alkoxy, acyloxy, amino, hydroxyl, mercapto,
carboxy, benzyloxy, phenyl, benzyl, cyano, nitro, thioalkoxy,
carboxaldehyde, carboalkoxy and carboxamide, or a functionality
that can be suitably blocked, if necessary for purposes of the
invention, with a protecting group.
[0049] For all of the foregoing definitions, preferred subsets of
the groups include C.sub.1-C.sub.12, C.sub.1-C.sub.10,
C.sub.1-C.sub.8, C.sub.1-C.sub.6, C.sub.1-C.sub.4, C.sub.1-C.sub.2
(when chemically possible), C.sub.3-C.sub.4, C.sub.3, and C.sub.4
groups.
[0050] "Aryl" or "Ar" refers to an aromatic carbocyclic group
having a single ring (including, but not limited to, groups such as
phenyl) or multiple condensed rings (including, but not limited to,
groups such as naphthyl or anthryl), and includes both
unsubstituted and substituted aryl groups. "Substituted aryls"
refers to aryls substituted with one or more substituents,
including, but not limited to, groups such as alkyl, alkenyl,
alkynyl, hydrocarbon chains, halogen, alkoxy, acyloxy, amino,
hydroxyl, mercapto, carboxy, benzyloxy, phenyl, benzyl, cyano,
nitro, thioalkoxy, carboxaldehyde, carboalkoxy and carboxamide, or
a functionality that can be suitably blocked, if necessary for
purposes of the invention, with a protecting group.
[0051] "Heteroalkyl," "heteroalkenyl," and "heteroalkynyl" refer to
alkyl, alkenyl, and alkynyl groups, respectively, that contain the
number of carbon atoms specified (or if no number is specified,
having up to 12 carbon atoms) which contain one or more heteroatoms
as part of the main, branched, or cyclic chains in the group.
Heteroatoms include, but are not limited to, N, S, O, and P; N and
O are preferred. Heteroalkyl, heteroalkenyl, and heteroalkynyl
groups may be attached to the remainder of the molecule either at a
heteroatom (if a valence is available) or at a carbon atom.
Examples of heteroalkyl groups include, but are not limited to,
groups such as --O--CH.sub.3, --CH.sub.2--O--CH.sub.3,
--CH.sub.2--CH.sub.2--O--CH.sub.3,
--S--CH.sub.2--CH.sub.2--CH.sub.3,
--CH.sub.2--CH(CH.sub.3)--S--CH.sub.3,
--CH.sub.2--CH.sub.2--NH--CH.sub.2--CH.sub.2--,
1-ethyl-6-propylpiperidino, 2-ethylthiophenyl, and morpholino.
Examples of heteroalkenyl groups include, but are not limited to,
groups such as --CH.dbd.CH--NH--CH(CH.sub.3)--CH.sub.2--.
"Heteroaryl" or "HetAr" refers to an aromatic carbocyclic group
having a single ring (including, but not limited to, examples such
as pyridyl, thiophene, or furyl) or multiple condensed rings
(including, but not limited to, examples such as imidazolyl,
indolizinyl or benzothienyl) and having at least one hetero atom,
including, but not limited to, heteroatoms such as N, O, P, or S,
within the ring. Unless otherwise specified, heteroalkyl,
heteroalkenyl, heteroalkynyl, and heteroaryl groups have between
one and five heteroatoms and between one and twelve carbon atoms.
"Substituted heteroalkyl," "substituted heteroalkenyl,"
"substituted heteroalkynyl," and "substituted heteroaryl" groups
refer to heteroalkyl, heteroalkenyl, heteroalkynyl, and heteroaryl
groups substituted with one or more substituents, including, but
not limited to, groups such as alkyl, alkenyl, alkynyl, benzyl,
hydrocarbon chains, halogen, alkoxy, acyloxy, amino, hydroxyl,
mercapto, carboxy, benzyloxy, phenyl, benzyl, cyano, nitro,
thioalkoxy, carboxaldehyde, carboalkoxy and carboxamide, or a
functionality that can be suitably blocked, if necessary for
purposes of the invention, with a protecting group. Examples of
such substituted heteroalkyl groups include, but are not limited
to, piperazine, substituted at a nitrogen or carbon by a phenyl or
benzyl group, and attached to the remainder of the molecule by any
available valence on a carbon or nitrogen, --NH--SO.sub.2-phenyl,
--NH--(C.dbd.O)O-alkyl, --NH--(C.dbd.O)O-alkyl-aryl, and
--NH--(C.dbd.O)-alkyl. If chemically possible, the heteroatom(s) as
well as the carbon atoms of the group can be substituted. The
heteroatom(s) can also be in oxidized form, if chemically
possible.
[0052] The term "alkylaryl" refers to an alkyl group having the
number of carbon atoms designated, appended to one, two, or three
aryl groups.
[0053] The term "alkoxy" as used herein refers to an alkyl,
alkenyl, alkynyl, or hydrocarbon chain linked to an oxygen atom and
having the number of carbon atoms specified, or if no number is
specified, having up to 12 carbon atoms. Examples of alkoxy groups
include, but are not limited to, groups such as methoxy, ethoxy,
and t-butoxy.
[0054] The term "alkanoate" as used herein refers to an ionized
carboxylic acid group, such as acetate
(CH.sub.3C(.dbd.O)--O.sup.(-1)), propionate
(CH.sub.3CH.sub.2C(.dbd.O)--O.sup.(-1)), and the like. "Alkyl
alkanoate" refers to a carboxylic acid esterified with an alkoxy
group, such as ethyl acetate
(CH.sub.3C(.dbd.O)--O--CH.sub.2CH.sub.3). "(O-haloalkyl alkanoate"
refers to an alkyl alkanoate bearing a halogen atom on the
alkanoate carbon atom furthest from the carboxyl group; thus, ethyl
.omega.-bromo propionate refers to ethyl 3-bromopropionate, methyl
.omega.-chloro n-butanoate refers to methyl 4-chloro n-butanoate,
etc.
[0055] The terms "halo" and "halogen" as used herein refer to Cl,
Br, F or I substituents.
[0056] "Protecting group" refers to a chemical group that exhibits
the following characteristics: 1) reacts selectively with the
desired functionality in good yield to give a protected substrate
that is stable to the projected reactions for which protection is
desired; 2) is selectively removable from the protected substrate
to yield the desired functionality; and 3) is removable in good
yield by reagents compatible with the other functional group(s)
present or generated in such projected reactions. Examples of
suitable protecting groups can be found in Greene et al. (1991)
Protective Groups in Organic Synthesis, 2nd Ed. (John Wiley &
Sons, Inc., New York). Amino protecting groups include, but are not
limited to, mesitylenesulfonyl (Mes), benzyloxycarbonyl (CBz or Z),
t-butyloxycarbonyl (Boc), t-butyldimethylsilyl (TBDIMS or TBDMS),
9-fluorenylmethyloxycarbonyl (Fmoc), tosyl, benzenesulfonyl,
2-pyridyl sulfonyl, or suitable photolabile protecting groups such
as 6-nitroveratryloxy carbonyl (Nvoc), nitropiperonyl,
pyrenylmethoxycarbonyl, nitrobenzyl, dimethyl dimethoxybenzil,
5-bromo-7-nitroindolinyl, and the like. Hydroxylprotecting groups
include, but are not limited to, Fmoc, TBDIMS, photolabile
protecting groups (such as nitroveratryl oxymethyl ether (Nvom)),
Mom (methoxy methyl ether), and Mem (methoxy ethoxy methyl ether),
NPEOC (4-nitrophenethyloxycarbonyl) and NPEOM
(4-nitrophenethyloxymethyloxycarbonyl).
[0057] Synthesis of Porphyrin-Polyamine Conjugates: Overview
[0058] Syntheses are described with reference to the schemes below.
The synthesis of SL-11211 (Scheme 1) started with acetal 1 that was
protected by mesitylene sulfonation to give 2. Alkylation of the
known triamide 3 with 4-bromobutyronitrile following the procedure
described previously (see Examples), reduction of the resulting
nitrile and mesitylene sulfonation of the free amine gave octaamide
4. Treatment of 4 with 1,4-dibromobutane gave 5, that was then
condensed with 2 to afford 6. Cleavage of the acetal residue of 6
resulted in the aldehyde 7, that was subjected to reductive
amination with ethylamine to give 8. The amine was then condensed
with mesoporhyrin dihdrochloride to the porphyrin diamide 9;
deprotection of the amino residues of 9 gave SL-11211.
[0059] The synthesis of SL-11233 (Scheme 2) started with the known
triamide 10 that was alkylated with 1,4-dibromobutane to give 11.
The latter is condensed with 2 to give 12, the acetal cleaved to
aldehyde 13, and the latter reductively aminated to 14.
Condensation of 14 with deuteroporphyrin IX 2,4-disulfonic acid
gave 15. Cleavage of the protecting groups in 15 allowed the
synthesis of SL-11233.
[0060] The synthesis of SL-11235 started with the condensation of 8
and deuteroporphyrin IX-2,4-disulfonate to give 16 (Scheme 3).
Deprotection of the amino residues gave SL-11235
eicosahydrobromide.
[0061] The synthesis of SL-11236 started with the condensation of 8
and N-methylmesoporphyrin IX to give 17, that was then deprotected
to give SL-11236 eicosahydrobromide (Scheme 3).
[0062] The synthesis of SL-11237 started with the previously
described cyclic amine 18 (patent cyclic polyamines) that was
condensed with mesoporphyrin IX. dihydrochloride to give SL-11237
(Scheme 3).
[0063] The synthesis of SL-11217 (Scheme 4) started with the known
cyclopropyl derivative 19, that was hydrolyzed to the acid and the
latter transformed into its chloride 20. Condensation of 20 with a
protected N-ethyl 1,4-diaminobutane gave 21, that was reduced with
diborane and then acylated with mesitylenesulfonyl chloride to give
22. Alkylation of 22 with dibromobutane in the presence of sodium
iodide gave 23, that was condensed with ethylamine to give 24.
[0064] Condensation of 24 with mesoporphyrin IX dihydrochloride
gave 25, deprotection of the amino residues gave SL-11217
hydrobromide.
[0065] The synthesis of SL-11209 started with the known amine 26
that was alkylated with the benzyl ether of 4-bromobutanol to give
27 (Scheme 5). Hydrolysis of the benzyl ether gave the alcohol 28,
the alcohol was protected by reaction with t-butyloxycarbonyl
anhydride to give 29, and the latter oxidized to the aldehyde 30.
Reductive amination of 30 gave 31. In tandem, reduction of the
diester of mesoporphyrin gave the dialdehyde 32. Condensation of 32
with 31, followed by acid deprotection of the amino residues gave
SL-11209.
[0066] The synthesis of SL-11210 started with the known nitrile 33
that was reduced to the amine and the latter condensed with 32
following a reductive amination procedure (Scheme 6). Deprotection
of the amino residues gave SL-11210.
[0067] Reductive amination procedures allowed the condensation of
18 and aldehyde 32 that gave SL-11257 (Scheme 7) ##STR5## ##STR6##
##STR7## ##STR8## ##STR9## ##STR10## ##STR11## ##STR12## ##STR13##
##STR14##
[0068] Therapeutic Use of Porphyrin-Polyamine Conjugate
Compounds
[0069] Porphyrin-polyamine conjugate compounds of the present
invention are useful for treatment of a variety of diseases caused
by uncontrolled proliferation of cells, including cancer,
particularly prostate cancer. The compounds are used to treat
mammals, preferably humans. "Treating" a disease using a
porphyrin-polyamine conjugate compound of the invention is defined
as administering one or more porphyrin-polyamine conjugate
compounds of the invention, with or without additional therapeutic
agents, in order to prevent, reduce, or eliminate either the
disease or the symptoms of the disease, or to retard the
progression of the disease or of symptoms of the disease.
"Therapeutic use" of the porphyrin-polyamine conjugate compounds of
the invention is defined as using one or more porphyrin-polyamine
conjugate compounds of the invention to treat a disease, as defined
above.
[0070] In order to evaluate the efficacy of a particular
porphyrin-polyamine conjugate compound for a particular medicinal
application, the compounds can be first tested against
appropriately chosen test cells in vitro. In a non-limiting
example, porphyrin-polyamine conjugate compounds can be tested
against tumor cells, for example, prostate tumor cells. Exemplary
experiments can utilize cell lines capable of growing in culture as
well as in vivo in athymic nude mice, such as LNCaP. Horoszewicz et
al. (1983) Cancer Res. 43:1809-1818. Culturing and treatment of
carcinoma cell lines, cell cycle and cell death determinations
based on flow cytometry; enzyme assays including ODC, SAMDC and
SSAT activities; and high pressure liquid chromatography detection
and quantitation of natural polyamines and polyamine analogs are
described in the art, for example, Mi et al. (1998) Prostate
34:51-60; Kramer et al. (1997) Cancer Res. 57:5521-27; and Kramer
et al. (1995) J. Biol. Chem. 270:2124-2132. Evaluations can also be
made of the effects of the porphyrin-polyamine conjugate compound
on cell growth and metabolism.
[0071] Analysis begins with IC.sub.50 determinations based on
dose-response curves ranging from 0.1 to 1000 .mu.M performed at 72
hr. From these studies, conditions can be defined which produce
about 50% growth inhibition and used to: (a) follow time-dependence
of growth inhibition for up to 6 days, with particular attention to
decreases in cell number, which may indicate drug-induced cell
death; (b) characterize porphyrin-polyamine conjugate compound
effects on cell cycle progression and cell death using flow
cytometry (analysis to be performed on attached and detached
cells); (c) examine porphyrin-polyamine conjugate compound effects
on cellular metabolic parameters. Porphyrin-polyamine conjugate
compound effects can be normalized to intracellular concentrations
(by HPLC analysis), which also provide an indication of their
relative ability to penetrate cells. Marked differences in
porphyrin-polyamine conjugate compound uptake can be further
characterized by studying the compound's ability to utilize and
regulate the polyamine transporter, as assessed by competition
studies using radiolabeled spermidine, as previously described in
Mi et al. (1998). Porphyrin-polyamine conjugate compounds could
also enter the cells by a diffusion mechanism.
[0072] In Vivo Testing of Porphyrin-Polyamine Conjugate
Compounds
[0073] Porphyrin-polyamine conjugate compounds found to have potent
anti-proliferative activity in vitro towards cultured carcinoma
cells can be evaluated in in vivo model systems. The first goal is
to determine the relative toxicity of the compounds in
non-tumor-bearing animals, such as DBA/2 mice. Groups of three
animals each can be injected intraperitoneally with increasing
concentrations of a porphyrin-polyamine conjugate compound,
beginning at, for example, 10 mg/kg. Toxicity as indicated by
morbidity is closely monitored over the first 24 hr. A
well-characterized polyamine analog compound, such as BE-333, can
be used as an internal standard in these studies, since a data base
has already been established regarding acute toxicity via a single
dose treatment relative to chronic toxicity via a daily.times.5 d
schedule. Thus, in the case of porphyrin-polyamine conjugate
compounds, single dose toxicity relative to BE-333 is used to
project the range of doses to be used on a daily.times.5 d
schedule. The toxicity of the porphyrin-polyamine conjugate
compound can also be tested versus the free polyamine compound,
that is, versus the same polyamine which is present in the
porphyrin-polyamine conjugate compound but without a conjugated
porphyrin.
[0074] After the highest tolerated dosage on a daily.times.5 d
schedule is deduced, antitumor activity is determined. Typically,
tumors can be subcutaneously implanted into nude athymic mice by
trocar and allowed to reach 100-200 mm.sup.3 before initiating
treatment by intraperitoneal injection daily.times.5 d. Most
porphyrin-polyamine conjugate compounds can be given in a range
between 10 and 200 mg/kg. Porphyrin-polyamine conjugate compounds
can be evaluated at three treatment dosages with 10-15 animals per
group (a minimum of three from each can be used for pharmacodynamic
studies, described below). Mice can be monitored and weighed twice
weekly to determine tumor size and toxicity. Tumor size is
determined by multi-directional measurement from which volume in
mm.sup.3 is calculated. Tumors can be followed until median tumor
volume of each group reaches 1500 mm.sup.3 (i.e., 20% of body
weight), at which time the animals can be sacrificed. Although the
initial anti-tumor studies focuses on a daily.times.5 d schedule,
constant infusion can be performed via Alzet pump delivery for 5
days since this schedule dramatically improves the anti-tumor
activity of BE-333 against A549 human large cell hung carcinoma.
Sharma et al. (1997) Clin. Cancer Res. 3:1239-1244. In addition to
assessing anti-tumor activity, free porphyrin-polyamine conjugate
compound levels and free polyamine levels in tumor and normal
tissues can be determined in test animals.
[0075] Methods of Administration of Porphyrin-Polyamine Conjugate
Compounds
[0076] The porphyrin-polyamine conjugate compounds of the present
invention can be administered to a mammalian, preferably human,
subject via any route known in the art, including, but not limited
to, those disclosed herein. Methods of administration include but
are not limited to, oral, intravenous, intraarterial, intratumoral,
intramuscular, topical, inhalation, subcutaneous, intraperitoneal,
gastrointestinal, and directly to a specific or affected organ. The
porphyrin-polyamine conjugate compounds described herein are
administratable in the form of tablets, pills, powder mixtures,
capsules, granules, injectables, creams, solutions, suppositories,
emulsions, dispersions, food premixes, and in other suitable forms.
The compounds can also be administered in liposome formulations.
The compounds can also be administered as prodrugs, where the
prodrug undergoes transformation in the treated subject to a form
which is therapeutically effective. Additional methods of
administration are known in the art.
[0077] The pharmaceutical dosage form which contains the compounds
described herein is conveniently admixed with a non-toxic
pharmaceutical organic carrier or a non-toxic pharmaceutical
inorganic carrier. Typical pharmaceutically-acceptable carriers
include, for example, mannitol, urea, dextrans, lactose, potato and
maize starches, magnesium stearate, talc, vegetable oils,
polyalkylene glycols, ethyl cellulose, poly(vinylpyrrolidone),
calcium carbonate, ethyl oleate, isopropyl myristate, benzyl
benzoate, sodium carbonate, gelatin, potassium carbonate, silicic
acid, and other conventionally employed acceptable carriers. The
pharmaceutical dosage form can also contain non-toxic auxiliary
substances such as emulsifying, preserving, or wetting agents, and
the like. A suitable carrier is one which does not cause an
intolerable side effect, but which allows the novel
porphyrin-polyamine conjugate compound(s) to retain its
pharmacological activity in the body. Formulations for parenteral
and nonparenteral drug delivery are known in the art and are set
forth in Remington's Pharmaceutical Sciences, 18th Edition, Mack
Publishing (1990). Solid forms, such as tablets, capsules and
powders, can be fabricated using conventional tableting and
capsule-filling machinery, which is well known in the art. Solid
dosage forms, including tablets and capsules for oral
administration in unit dose presentation form, can contain any
number of additional non-active ingredients known to the art,
including such conventional additives as excipients; desiccants;
colorants; binding agents, for example syrup, acacia, gelatin,
sorbitol, tragacanth, or polyvinylpyrollidone; fillers, for example
lactose, sugar, maize-starch, calcium phosphate, sorbitol or
glycine; tableting lubricants, for example magnesium stearate,
talc, polyethylene glycol or silica; disintegrants, for example
potato starch; or acceptable wetting agents such as sodium lauryl
sulfate. The tablets can be coated according to methods well known
in standard pharmaceutical practice. Liquid forms for ingestion can
be formulated using known liquid carriers, including aqueous and
non-aqueous carriers, suspensions, oil-in-water and/or water-in-oil
emulsions, and the like. Liquid formulations can also contain any
number of additional non-active ingredients, including colorants,
fragrance, flavorings, viscosity modifiers, preservatives,
stabilizers, and the like. For parenteral administration,
porphyrin-polyamine conjugate compounds can be administered as
injectable dosages of a solution or suspension of the compound in a
physiologically acceptable diluent or sterile liquid carrier such
as water or oil, with or without additional surfactants or
adjuvants. An illustrative list of carrier oils would include
animal and vegetable oils (e.g., peanut oil, soy bean oil),
petroleum-derived oils (e.g., mineral oil), and synthetic oils. In
general, for injectable unit doses, water, saline, aqueous dextrose
and related sugar solutions, and ethanol and glycol solutions such
as propylene glycol or polyethylene glycol are preferred liquid
carriers. The pharmaceutical unit dosage chosen is preferably
fabricated and administered to provide a final concentration of
drug at the point of contact with the cancer cell of from 1 .mu.M
to 10 mM. More preferred is a concentration of from 1 to 100 .mu.M.
The optimal effective concentration of porphyrin-polyamine
conjugate compounds can be determined empirically and will depend
on the type and severity of the disease, route of administration,
disease progression and health and mass or body area of the
patient. Such determinations are within the skill of one in the
art. Porphyrin-polyamine conjugate compounds can be administered as
the sole active ingredient, or can be administered in combination
with another active ingredient, including, but not limited to,
cytotoxic agents, antibiotics, antimetabolites, nitrosourea, vinca
alkaloids, polypeptides, antibodies, cytokines, etc.
EXAMPLES
[0078] The following examples are provided to illustrate the
invention, and are not intended to limit the invention in any
manner.
Synthesis of SL11211
Example 1
N-- Mesitylenesulfonyl 4-aminobutyraldehyde diethyl acetal 2
[0079] Amine 1 (Aldrich) (3.5 g, 21.7 mol) was dissolved in a
mixture of chloroform (30 ml) and 1N sodium hydroxide (24 ml) and
15 ml of mesitylenesulfonyl chloride dissolved in 15 ml of
chloroform were added at 5.degree. C. The mixture was stirred for 2
h, the reaction mixture was then diluted with chloroform (50 ml),
the organic layer was separated, washed with a saturated solution
of ammonium chloride, dried (Na.sub.2SO.sub.4), and evaporated to
dryness. The residual oil crystallized after drying and was used in
the next step without further purification; 7.0 g (95%) of 2 were
obtained; .sup.1HNMR (CDCl.sub.3): ppm 1.15(t,6H), 1.55 (m,4H),
2.30 (s, 3H), 2.65 (s, 6H), 2.95 (q, 2H), 3.40-3.55 (m, 4H), 4.40
(t, 1H), 4.90 (t, 1H), 6.95 (s, 2H); .sup.13CNMR (CDCl.sub.3): ppm
15.19, 20.80, 22.85, 24.55, 30.83, 42.39, 61.40, 102.41, 131.84,
133.82, 138.99, 141.92.
Example 2
[0080]
.sup.1N,.sup.6N,.sup.11N,.sup.16N,.sup.21N,.sup.26N,.sup.31N,.sup.-
36N-Octakis(mesitylensulfonyl)-1,6,11,16,21,26,31,36-octaazaoctatriacontan-
e 4 was obtained starting with compound 3 (U.S. PAT APPL.
60/329,982) following the homologation procedure described in WO
00/66587; namely, alkylation with 4-bromobutyronitrile, followed by
reduction of the nitrile and protection of the free amino residue
with mesitylenesulfonyl chloride. Starting with 7 g of 3, 5.6 g
(70%) of 4 were obtained over the aforementioned three synthetic
steps; .sup.1HNMR (CDCl.sub.3): 0.95 (t,3H), 1.30 (m, 28H), 2.30
(s, 24H), 2.55 (m, 48H), 2.75 (t, 1H), 3.0 (m, 30H), 6.95 (, 16H);
.sup.13CNMR (CDCl.sub.3): 12.71, 20.93, 22.78, 24.49, 24.79, 25.68,
40.07, 41.91, 44.59, 44.95, 131.98, 133.39, 138.94, 139.96,
142.06.
Example 3
.sup.3N,.sup.8N,
.sup.13N,.sup.18N,.sup.23N,.sup.28N,.sup.33N,.sup.38N-Octakis(mesitylenes-
ulfonyl)-3,8,13,18,23,28,33,38-octaaza-42-bromo-dotetracontane
5
[0081] To a solution of amide 4 (5.6 g, 2.8 mmol) and
1,4-dibromobutane (3.6 g, 16.8 mmol) in 45 ml of DMF kept at
5.degree. C. was added 135 mg (3.36 mmol) of NaH (60% dispersion in
mineral oil) with constant stirring. The mixture was kept at
22.degree. C. for 18 h; the solvent was then evaporated to dryness,
the residue dissolved in chloroform, washed twice with a saturated
solution of ammonium chloride, the organic layer separated, dried
(Na.sub.2SO.sub.4) and evaporated to dryness. The residue was
crystallized from ethyl acetate-hexane; 5.3 g (88%) of 5 were
obtained; mp 109.degree. C.; .sup.1HNMR (CDCl.sub.3): 0.95 (t, 3H),
1.40 (m, 32H), 2.30 (s, 24H), 2.50 (m, 48H), 3.00 (m,32H), 3.25 (t,
2H), 6.95 (s, 16H); .sup.13CNMR (CDCl.sub.3): 12.68, 20.89, 22.68,
24.42, 25.78, 29.61, 32.77, 40.03, 44.51, 44.89, 45.04, 131.95,
133.46, 139.91, 142.28.
Example 4
.sup.3N,.sup.8N,.sup.13N,.sup.18N,.sup.23N,.sup.28N,.sup.33N,.sup.38N,.sup-
.43N-Nonakis(mesitylenesulfonyl)-3,8,13,18,23,28,33,38,43-nonaaza-heptatet-
racontylaldehyde diethyl acetal 6
[0082] To a solution of amide 5 (5.19 g, 2.43 mmol) and acetal 2
(0.915 g, 2.67 mmol) in 50 ml of DMF kept at 5.degree. C. was added
128 mg (3.20 mmol) of NaH (60% dispersion in mineral oil) with
constant stirring. The mixture was kept at 22.degree. C. for 18 h
and the work up followed the procedure reported for 5; 5.0 g (86%)
of 6 were obtained; mp 102.4.degree. C.; .sup.1HNMR (CDCl.sub.3):
0.95 (t, 3H), 1.15 (t, 6H), 1.30 (m, 36H), 2.30 (s, 27H), 2.50 (s,
54H), 3.05 (m,36H), 3.45 (m,m, 4H), 6.95 (s,18H); .sup.13CNMR
(CDCl.sub.3): 12.68, 15.26, 20.89, 22.67, 24.57, 30.81, 40.04,
4456, 44.88, 45.23, 102.31, 131.88, 133.39, 139.91, 142.19.
Example 5
.sup.3N,.sup.8N,.sup.13N,.sup.18N,.sup.23N,.sup.28N,.sup.33N,.sup.38N,.sup-
.43N-Nonakis(mesitylenesulfonyl)-3,8,13,18,28,33,38,43-nonaaza-heptatetrac-
ontylaldehyde 7
[0083] Acetal 6 (5.0 g) was dissolved in acetone (140 ml) and water
(1.5 ml), Amberlyst-15 resin (600 mg) was added and the reaction
mixture was stirred for 1 h; the resin was filtered, the solvent
evaporated to dryness in vacuo, and the oily residue was used in
the next step without further purification; .sup.1HNMR
(CDCl.sub.3): 0.95 (t,3H), 1.30(m,36H), 1.72 (m,2H), 2.30 (s,27H),
2.52(s,s, 54H), 3.05 (m,36H), 6.95(s,18H), 9.60 (s, 1H);
.sup.13CNMR (CDCl.sub.3): 12.85, 19.89, 21.07, 22.92, 24.60, 24.92,
40.21, 40.79, 44.73, 45.06, 132.13, 133.54, 140.10, 142.47, 200.94;
MS (MALDI): 2345.2 (M+Na.sup.+), 2361.2 (M+K.sup.+).
Example 6
.sup.3N,.sup.8N,.sup.13N,.sup.18N,.sup.23N,.sup.28N,.sup.33N,.sup.38N,.sup-
.42N-Nonakis(mesitylenesulfonyl)-3,8,13,18,23,28,33,38,42,47-decaaza-nonat-
etracontane 8
[0084] To a solution of 4.2 g (1.7 mmol) of aldehyde 7 in 120 ml of
DCE, were added 7 ml (8 eq) of a 2M solution of ethylamine in THF.
The mixture was kept at 22.degree. C. for 18 h with constant
stirring, after which sodium triacetoxyborohydride (720 mg, 3.4
mol) was added. After 2 h at 22.degree. C., the mixture was washed
(2.times.20 ml) with a saturated solution of sodium bicarbonate,
dried, and evaporated to dryness. The residue was purified by flash
chromathography using Cl.sub.3CH/MeOH (5% to 10%) as eluant; 2.8 g
(68%) of 8 were recovered; .sup.1HNMR (Cl.sub.3CD): 0.95 (t,3H),
1.10 (t,3H), 1.30(m, 36H), 2.25 (s, 27H), 2.50(m, 58H), 3.05 (m,
36H), 2.25 (s, 27H), 2.50 (m, 58H), 3.05 (m, 36H), 6.95 (s, 18H);
.sup.13CNMR (CDCl.sub.3): 12.66, 14.53, 20.89, 22.74, 24.70, 39.99,
43.73, 44.50, 44.84, 45.17, 48.57, 131.93, 133.31, 139.90, 142.28;
MS (MALDI): 2351.92 (M+H.sup.+), 2373.10 (M+Na.sup.+), 2389.97
(M+K.sup.+).
Example 7
Mesoporphyrin
IX-bis[.sup.3N,.sup.8N,.sup.13N.sup.18N,.sup.23N,.sup.28N,.sup.33N,.sup.3-
8N,.sup.42N-nonakis(mesitylenesulfonyl)-3,8,13,18,23,28,33,38,42,47-decaaz-
anonatetracontyl amide]9
[0085] A mixture of amine 8 (750 mg, 0.3 mmol), mesoporphyrin IX
(102 mg, 1.4 mmol), and diisopropylethylamine (0.25 ml, 1.4 mmol)
in 30 ml of DMF were cooled to 5.degree. C. and kept under a
nitrogen atmosphere while 204 mg (0.54 mmol) of HBTU were added.
The reaction mixture was stirred for 2 h, the solvent evaporated to
dryness, the residue dissolved in chloroform, washed twice with a
saturated bicarbonate solution, the organic layer dried
(Na.sub.2SO.sub.4) and evaporated to dryness. The residue was
purified by chromathography on silica gel using ethyl
acetate:hexane (9:1) as eluant; 630 mg (75%) of 9 were recovered;
MS (ESI): 5239.3 (M+H.sup.+); 5261.3 (M+Na.sup.+).
Example 8
Porphyrin conjugate SL11211 eicosahydrobromide
[0086] Porphyrin amide 9 (630 mg) was dissolved in a mixture of
methylene chloride (12 ml). 30% hydrogen bromide in glacial acetic
acid (12 ml), and phenol (900 mg). The reaction mixture was kept at
22.degree. C. for 18 h with stirring, the reaction product was then
extracted into water (35 ml), the aqueous layer washed with
methylene chloride (3.times.12 ml), the aqueous solution evaporated
to dryness, and the residue crystallized from water/ethanol; 300 mg
of SL-11211 hydrobromide (86%) were obtained; mp. 250.degree. C.
(dec); MS (ESI): 1958.4 (M+H.sup.+, free base), 1980
(M+Na.sup.+).
Example 9
.sup.3N,.sup.8N,.sup.13N-Tri(mesitylenesulfonyl)-3,8,13-triaza-17-bromohep-
tadecane 11
[0087] Intermediate 11 was prepared starting with 10 (ref) and
following the procedure described for 5. Starting with 3.6 g of 10
were obtained 2.77 g (65%) of 11; .sup.1HNMR (CDCl.sub.3): 0.98
(t,3H), 1.40 (m, 8H), 1.65 (m,4H), 2.30 (s, 9H), 2.60 (s,18H), 3.10
(m,12H), 3.30 (t, 2H), 6.95 (s,6H); .sup.13CNMR (CDCl.sub.3):
12.75, 20.91, 22.72, 24.58, 25.85, 29.66, 32.79, 40.07, 44.59,
45.10, 131.90, 133.23, 140.03, 142.32.
Example 10
.sup.3N,.sup.8N,.sup.13N,.sup.18N-Tetrakis(mesitylenesulfonyl)-3,8,13,18-t-
etraaza-doeicosanylaldehyde diethyl acetal 12
[0088] Prepared from 11 following the procedure described for 6.
Starting with 2.8 g of 11 were obtained 3.5 g (97%) of acetal 12;
.sup.1HNMR (CDCl.sub.3): 1.00 (t, 3H), 1.15 (t, 6H), 1.35 (m, 16H),
2.35 (s, 12H), 2.55(s, 24H), 2.10(m,16H), 3.50 (m, 4H), 4.35
(t,1H), 6.95(s, 8H); .sup.13CNMR (CDCl.sub.3): 12.70, 15.26, 20.87,
22.44, 24.52, 30.81, 40.04, 44.55, 45.00, 45.23, 61.18, 102.31,
131.87, 133.35, 139.97, 142.18.
Example 111
.sup.3N,.sup.8N,.sup.13N,.sup.18N-Tetrakis(mesitylenesulfonyl)-3,8,13,18-t-
etraaza-doeicosanylaldehyde 13
[0089] The aldehyde was obtained from 12 following the procedure
described for 7. From 3.5 g of acetal 12, were obtained 2.9 g (90%)
of aldehyde 13; .sup.1HNMR (CDCl.sub.3): 0.95 (t, 3H), 1.35(m,
12H), 1.75 (m,2H), 2.30(m, 14H), 2.45(s, 24H), 3.05 (m, 2H), 2.30
(m, 14H), 2.45 (s, 24H), 3.05 (m, 16H), 6.95 (s, 8H), 9.60 (s, 1H);
.sup.13CNMR (CDCl.sub.3): 12.63, 19.68, 20.80, 22.67, 24.68, 26.32,
39.98, 40.54, 43.70, 44.50, 45.06, 131.88, 133.04, 140.32, 142.22,
200.71.
Example 12
.sup.3N,.sup.8N,.sup.13N,.sup.18N-Tetrakis(mesitylenesulfonyl)-3,8,13,18,2-
3-pentaazapentacosane 14
[0090] Amine 14 was prepared from 13 following the procedure
described for 8. Starting with 4.7 g of 13 were obtained 3.5 g
(72%) of amine 14; .sup.1HNMR (CDCl.sub.3): 1.00 (t, 3H), 1.15(t,
3H), 1.40 (m, 16H), 2.30(s, 12H), 2.60 (m, 28H), 3.10(m, 16H),
6.95(s, 8H); .sup.13CNMR(CDCl.sub.3): 11.70, 12.65, 20.83, 22.69,
23.46, 24.67, 39.96, 42.59, 44.47, 45.07, 46.60, 131.81, 133.26,
139.82, 142.36.
Example 13
2,4-Disulfonyl-Deuteroporphyrin
IX-bis[.sup.3N,.sup.8N,.sup.13N,.sup.18N-tetrakis(mesitylenesulfonyl)-3,8-
,13,18,23-pentaazapentacosyl amide] 15
[0091] Porphyrin conjugate 15 was prepared by condensation of 14
with deuteroporphyrin IX disulfonate following the procedure
described for 9. From 200 mg of 14 and 70 mg of the porphyrin, 144
mg (55%) of 15 were obtained; MS(MALDI): 2808.79 (M+H.sup.+),
2830.55 (M+Na.sup.+), 2853 (M+2Na.sup.+), 2875.52
(M+.sup.3Na.sup.+).
Example 14
Porphyrin conjugate SL11233 decahydrobromide
[0092] Conjugate SL-11233 was obtained from 15 following the
procedure described for SL-11211. From 144 mg of 15 were obtained
60 mg (55%) of SL-11233 decahydrobromide; MS (ESI): 1350
(M+H.sup.+, M=free base).
Example 15
2,4-Disulfonyl-Deuteroporphyrin IX-bis
[.sup.3N,.sup.8N,.sup.13N,.sup.18N,.sup.23N,.sup.28N,.sup.33N,.sup.38N,.s-
up.42N-nonakis(mesitylenesulfonyl)-3,8,13,18,23,28,33,38,42,47-decaaza-non-
atetracontyl amide] 16
[0093] Porphyrin conjugate 16 was prepared by condensation of amine
8 (216 mg) and deuteroporphyrin IX disulfonate (34 mg) following
the procedure described for 9; 140 mg (57%) of 16 were obtained;
MS(MALDI): 5342 (M+H.sup.+), 5363 (M+Na.sup.+).
Example 16
Porphyrin conjugate SL-11235 eicosahydrobromide
[0094] Conjugate SL-11235 was obtained from 140 mg of 16 following
the procedure described for the synthesis of SL-11211; 70 mg (72%)
of SL-11235 eicosahydrobromide were obtained; MS (MALDI):
2062.0(M+H.sup.+, M=free base), 1031 (M*/2), 688.0 (M*/3).
Example 17
N-Methyl mesoporphyrin
IX-bis[.sup.3N,.sup.8N,.sup.13N,.sup.18N,.sup.23N,.sup.28N,.sup.33N,.sup.-
38N,.sup.42N
nonakis(mesitylenesulfonyl)-3,8,13,18,23,28,33,38,42,47-decaaza-nonatetra-
contyl amide] 17
[0095] Amide 17 was prepared by condensation of amine 8 (404 mg)
with N-methyl mesoporphyrin IX (50 mg) following the procedure
described for 9; 226 mg (50%) of 17 were obtained; MS (MALDI): 5253
(M+H.sup.+).
Example 18
Porphyrin conjugate SL-11236 eicosahydrobromide
[0096] SL-11236 was prepared from 215 mg of 17 following the
procedure described for the synthesis of SL-11211; 75 mg (52%) of
SL-11236 eicosahydrobromide were obtained; MS(MALDI): 1972.0
(M+H.sup.+, M=free base), 1989.0 (M+NH.sub.4.sup.+), 986.6
(M.sup.+/2).
Example 19
Porphyrin conjugate S111237 decahydrochloride
[0097] SL-11237 was prepared by condensation of 424 mg (0.6 mmol)
of amine 18 and 191 mg (0.3 mmol) of mesoporphyrin IX following the
procedure described for 9. SL-11237 was purified by chromathography
on silica gel using chloroform/methanol/ammonium hydroxide: 8/2/0.1
as eluant; the eluted residue was further crystallized from
methanol/hydrogen chloride/ethyl acetate; 430 mg (73%) of SL-11237
decahydrochloride were obtained; MS(ESI): 1579.6 (M+H.sup.+, M=free
base), 1725.6 (M+4HCl), 1871.8 (M+8HCl), 790.23 (M.sup.+/2), 527.21
(M.sup.+/3), 790.23(M.sup.+/2).
Synthesis of SL-11217
Example 20
trans-2-Cyanocyclopropanecarbonyl chloride 20
[0098] 1N Sodium hydroxide (71.9 ml, 71.9 mmol) was added to a
solution of nitrile 19 (Payne G B, JOC (1967) 32, 3351) (10.0 g,
71.9 mmol) in 40 ml of methanol. The mixture was stirred during 1
h, the methanol was evaporated, conc.HCl was added to pH 2, the
solution extracted with ethyl ether (3.times.30 ml), the pooled
organic layers were dried (Na.sub.2SO.sub.4) and evaporated to
dryness. The residual solid (7.4 g, 93%) was used in the next step
without further purification. It was dissolved in thionyl chloride
(13 ml), the mixture was heated to 65.degree. C./4 h, the thionyl
chloride was then distilled off and 20 was purified by distillation
at 50.degree. C./0.5 mm; 4.3 g (54% over two steps) were obtained;
.sup.1HNMR (Cl.sub.3CD): 1.80 (m, 2H), 2.25 (m, 1H), 2.80 (m, 1H);
.sup.13CNMR (Cl.sub.3CD); 8.62, 1701, 30.20, 117.46, 171.34.
Example 21
trans 2-Nitrile-1-(N-ethyl-N-mesitylenesulfonyl-aminobutyl)
cyclopropanecarboxamide 21
[0099] A solution of acyl chloride 20 (4.36 g, 33.7 mmol) in THF
(43 ml) was added dropwise to a solution of
N-ethyl-N(mesitylenesulfonyl)-1,4-diamine (10.0 g, 33.7 mmol) (ref)
and triethylamine (2.9 ml) in 100 ml of THF while the mixture was
kept at 5.degree. C. under nitrogen. Triethylammonium chloride
precipitated; the mixture is further kept at 22.degree. C. during
18 h, then extracted with ethyl acetate (80 ml), the organic layer
washed with 2N HCl (10 ml), then with a saturated ammonium chloride
solution (10 ml), dried (Na.sub.2SO.sub.4), and evaporated to
dryness. The residue was purified by flash chromathography on
silica gel using hexane/ethyl acetate: 6/4 as eluant; 10.3 g (78%)
of 21 were obtained; .sup.1HNMR (CDCl.sub.3): 10.2 (t, 3H),
1.35(m,1H), 1.55 (m, 5H), 1.90 (m, 1H), 2.05 (m, 1H), 2.35 (s, 3H),
2.60 (s, 6H), 3.25 (m, 6H), 6.35 (t, 1H), 6.95 (s, 2H); .sup.13CNMR
(Cl.sub.3CD): 4.44, 12.59, 20.87, 22.62, 22.69, 25.00, 26.37,
39.42, 40.04, 44.63, 120.14, 131.91, 133.23, 140.00, 142.37,
168.17.
Example 22
trans
1N-(Mesitylenesulfonyl)-2N(mesitylenesulfonyl)-2N(1'-N,N-(mesitylene-
sulfonyl)ethylaminobutyl)1,2-diaminomethylcyclopropane 22
[0100] Amide 21 (8.5 g, 21.7 mmol) was dissolved in 40 ml of THF,
156 ml of THF.1M BH3 were added and the solution was heated at
70.degree. C. during 2 h. The solution was cooled to 5.degree. C.,
30 ml of 6N HCl was slowly added while stirring, and the mixture
was kept at 5.degree. C. during 18 h. The pH of the mixture was
then adjusted to pH 10 with 50% potassium hydroxide, the oil that
separated was extracted into chloroform (3.times.50 ml), the
organic extracts were dried (Na.sub.2SO.sub.4), and evaporated to
dryness. The residue was dissolved in 100 ml of chloroform, 50 ml
of 2N sodium hydroxide were added, the mixture cooled to 5.degree.
C., and mesitylenesulfonyl chloride (8.2 g, 386 mmol) dissolved in
10 ml of chloroform were added with efficient stirring. After 2 h,
the organic layer was separated, dried (Na.sub.2SO.sub.4), and
evaporated to dryness. The residue was purified by flash
chromathography on silica gel using hexane/ethyl acetate: 7/3 as
eluant; 11.33 g (69% over two steps) of 22 were obtained;
.sup.1HNMR (Cl.sub.3CD): 0.40 (t, 2H), 0.95 (m, 5H), 1.25 (m, 4H),
2.25 (s, 9H), 2.35-2.65 (m, s, s, 20H), 2.85-3.30 (m, 8H), 5.50 (t,
1H), 6.95 (s, 6H); .sup.13CNMR (CDCl.sub.3) 10.02, 12.63, 16.15,
17.57, 20.87, 22.63, 22.69, 22.89, 24.01, 24.61, 39.97, 44.49,
44.89, 46.52, 48.28, 131.85, 132.34, 133.39, 133.97, 139.01,
139.97, 140.30, 141.73, 142.22, 142.60; MS (TOF): 768.2
(M+Na.sup.+), 784.2 (M+K.sup.+).
Example 23
.sup.3N,.sup.8N,.sup.13N-Tris(mesitylenesulfonyl)-17-iodo-((E)-10,11-cyclo-
propane)-3,8,13-triaza-heptadecane 23
[0101] Triamide 22 (10.3 g, 13.8 mmol) was dissolved in 100 ml of
DMF, cooled to 5.degree. C., and sodium hydride (662 mg, 16.5 mmol)
was added. The reaction mixture reached 22.degree. C. when
1,4-dibromobutane (29.8 g, 138 mmol) and sodium iodide (20.7 g, 138
mmol) were added, and the mixture was heated at 75.degree. C. for
90 min. The solution was evaporated to dryness, the residue
dissolved in chloroform, the solution was washed with sodium
thiosulfate, dried (Na.sub.2SO.sub.4), and evaporated to dryness.
The residue was purified on a silica gel column using hexane/ethyl
acetate; from 8/2 to 7/3 as eluant; 10.8 g (84%) of 23 were
obtained; .sup.1HNMR (Cl.sub.3CD): 0.40 (m, 2H), 0.80 (m, 2H), 1.02
(t, 3H), 1.40(m, 4H), 1.60 (m, 4H), 2.30 (s, 9H), 2.60 (s, 18H),
2.80-3.30 (m, 14H), 6.95 (s, 6H); .sup.13CNMR (Cl.sub.3CD): 5.73,
11.01, 12.73, 16.07, 20.93, 22.74, 24.41, 25.65, 27.96, 29.62,
30.35, 32.92, 40.03, 44.40, 44.58, 45.24, 131.93, 140.09, 142.34,
142.48.
Example 24
.sup.3N,.sup.8N,.sup.13N,.sup.18N-Tris(mesitylenesulfonyl)-((E)-10,11-cycl-
opropane) 3,8,13,18-tetraazaeicosane 24
[0102] Triamide 23 (10.8 g, 11.6 mmol) was dissolved in 25 ml of
THF and a 2M ethylamine solution in methanol was added (150 ml).
The solution was heated at 65.degree. C. during 16 h, then
evaporated to dryness, the residue dissolved in chloroform, the
chloroform washed with a concentrated solution of ammonium
chloride, dried (Na.sub.2SO.sub.4), evaporated to dryness, and the
residue purified by column chromathography on silica gel using from
5% to 10% methanol in chloroform as an eluant; 9.3 g (94%) of 24
were obtained; .sup.1HNMR (Cl.sub.3CD): 0.40 (t, 2H), 0.80 (m, 3H),
1.03 (t, 3H), 1.20 (t, 3H), 1.35 (m, 4H), 1.55 (m,4H), 2.25 (s,
9H), 2.40-3.35 (s, m, 34H), 6.95 (s, 6H); .sup.13CNMR (Cl.sub.3CD):
11.02, 12.70, 13.70, 16.04, 20.90, 22.71, 24.37, 24.76, 25.57,
40.02, 43.43, 44.57, 45.18, 45.33, 48.02, 48.83, 131.91, 133.13,
140.04, 142.34; MS (ESI): 846 (M+H.sup.+).
Example 25
Mesoporphyrin IX-bis
[.sup.3N,.sup.8N,.sup.13N,.sup.18N-tris(mesitylenesulfonyl)-((E)-10,11-cy-
clopropane)-3,8,13,18-tetrazaeicosanylamide] 25
[0103] Porphyrin diamide 25 was prepared by the condensation of 8.9
g (10.5 mmol) of 24 and mesoporphyrin IX (3.2 g, 5 mmol) following
the procedure described for 9; 9.24 g (83%) of 25 were obtained; MS
(MALDI): 2241 (M+Na.sup.+).
Example 26
SL-11217 octahydrobromide
[0104] SL-11217 was prepared by cleavage of the protecting groups
of 4.6 g of 25 following the procedure described for the synthesis
of SL-11211; 3.4 g (96%) of SL-11217 octahydrobromide were
obtained; mp>250.degree. C. (dec), crystallized from
methanol/ethyl acetate; MS (ESI): 1128.2 (M+H.sup.+), 1150
(M+Na.sup.+), 1167 (M+K.sup.+), 564.6 (M+/2).
Synthesis of SL-11209 dodecahydrochloride
Example 27
Benzyl .sup.3N,.sup.8N,
.sup.13N,.sup.18N-Tetrakis(mesitylenesulfonyl)-3,8,13,18-tetraazauneicosa-
nyl alcohol 27
[0105] A suspension of NaH (60% in mineral oil, 440 mg, 14 mmol) in
DMF (50 ml) was slowly added to a stirred solution of
benzyl-4-bromobutyl ether (3.33 g, 13.7 mmol) and amide 26 (5.41 g,
5.48 mmol) (WO 00/66587) in DMF (100 ml) kept at 5.degree. C. The
reaction mixture was stirred for 10 h at 50.degree. C., quenched
with 5 ml of H.sub.2O at 0.degree. C., and evaporated to dryness in
vacuo. The residue was taken up in ethyl acetate, washed with
H.sub.2O, and purified on a silica gel column using ethyl
acetate/hexane: 3/7 as eluant; 5.1 g, (81%) of 27 were obatained;
.sup.1H-NMR (CDCl.sub.3): 0.97 (t, J=7.1 Hz, 3H), 1.2-1.5 (m, 16H),
2.27 (s, 3H), 2.29 (s, 9H), 2.55 (s, 24H), 2.9-3.2 (m, 16H), 3.31
(t, J=6.0 Hz), 4.41 (s, 2H), 6.9-7.0 (m, 8H), 7.2-7.4 (m, 5H).
Example 28
3,8,13,18-Tetrazauneicosanyl alcohol 28
[0106] A solution of 30% HBr in glacial acetic acid (90 ml) was
added to a stirred solution of 27 (4.50 g) and phenol (12.65 g) in
methylene chloride (45 ml) at 0.degree. C. The cooling bath was
removed and the reaction mixture was stirred for 24 h at 20.degree.
C. The reaction mixture was quenched with H.sub.2O (90 ml), washed
with methylene chloride, and concentrated to dryness in vacuo. The
residue was cooled to 0.degree. C., basified with 2N sodium
hydroxide (9 ml), followed by 50% potassium hydroxide (9 ml). The
product was extracted with chloroform (7.times.10 ml); 1.07 g (81%)
of 28 were obtained; .sup.1H-NMR (CDCl.sub.3): 1.10 (t, J=7 Hz,
3H), 1.40-1.75 (m, 16H), 2.55-2.75 (m, 16H), 3.57 (t, J=5.0 Hz);
.sup.13C-NMR (CDCl.sub.3): 15.23, 27.55, 27.92, 28.58, 32.35,
44.02, 49.35, 49.66, 49.80, 62.32.
Example 29
.sup.3N,.sup.8N,.sup.13N,.sup.18N--
Tetrakis(butyloxycarbonyl)-3,8,13,18-tetrazauneicosanyl alcohol
29
[0107] A solution of 10% sodium carbonate (26 ml) was added to a
solution of tetramine 28 (634 mg, 1.92 mmol) in dioxane (16 ml).
Di-tert-butyl dicarbonate (2.5 g, 11.5 mmol) in dioxane (16 ml) was
added into the reaction mixture at 0.degree. and stirred for 10 h
at 20.degree. C. The reaction mixture was diluted with chloroform
(200 ml), washed with water, then with brine, dried
(Na.sub.2SO.sub.4), evaporated to dryness, and purified by
chromathography on a silica gel column using ethyl
acetate/hexane:4/6 as eluant; 1.34 g, (96%) of 29 were obtained;
.sup.1H-NMR (CDCl.sub.3): 1.09 (t, J=7.1 Hz, 3H), 1.4-1.7 (m, 52H),
3.05-3.3 (m, 16H,), 3.67 (t, J=5.8 Hz, 2H).
Example 30
.sup.3N,.sup.8N,.sup.13N,.sup.18N-Tetrakis(butyloxycarbonyl)-3,813,18-tetr-
azauneicosanyl aldehyde 30
[0108] Oxalyl chloride (2N solution in methylene chloride, 0.821
.mu.l, 1.64 mmol) was diluted with anhydrous methylene chloride (6
ml) at -60.degree. C. DMSO (223 .mu.l, 2.59 mmol) in methylene
chloride (3 ml) was added to the mixture, the latter stirred for 5
min at -60.degree. C., and 29 (1.12 g, 1.53 mmol) dissolved in
methylene chloride (9 ml) was added to the reaction mixture. After
30 min of stirring at -60.degree. C., triethylamine (1.06 ml, 14.46
mmol) was added to the reaction mixture and the temperature was
allowed to rise to 20.degree. C. (ca. 1.5 h). The reaction mixture
was diluted with methylene chloride, washed with H.sub.2O,
saturated sodium bicarbonate, and brine. The organic layer was
concentrated to dryness in vacuo and purified by column
chromatography on silica gel using ethyl/acetate/hexane:3/7 as
eluant; 989 mg (89%) of 30 were obtained; .sup.1H-NMR (CDCl.sub.3):
1.09 (t, J=7.0 Hz, 3H), 1.4-1.6 (m, 48H), 1.84 (m, 2H), 2.45 (t,
J=6.8, 2H), 3.05-3.3 (m, 16H), 9.78 (s, 1H).
Example 31
.sup.3N,.sup.8N,.sup.13N,.sup.18N,.sup.23N-Tetrakis(butyloxyarbonyl)-3,8,1-
3,18,23-pentaza-pentaeicosane 31
[0109] Platinum oxide (100 mg) was reduced in methanol (30 ml) with
hydrogen at 30 psi. for 15 min. Aldehyde 30 (989 mg, 1.36 mmol)
dissolved in a 2M solution of ethylamine in ethanol (7 ml) was
added to the hydrogenation flask, and the mixture hydrogenated for
10 h at 50 psi. The catalyst was removed by filtration through
celite and the filtrate was concentrated to dryness in vacuo; 1.0 g
(99%) of 31 were obtained; .sup.1H-NMR (CDCl.sub.3): 1.09 (t, J=7.6
Hz, 3H), 1.12 (t, J=7.2 Hz, 3H), 1.3-1.65 (m, 50H, CH.sub.2), 1.66
(m, 2H), 2.71 (m, 2H), 3.1-3.3 (m, 18H). MS-MALDI (m/z):758.8
(M.sup.+, 100%), 744 (30%).
Example 32
1,3,5,8-Tetramethyl-Z 4-diethyl-6,7-di(propionaldehyde)porphyrin
32
[0110] Diisobutylaluminum hydride (1.16 ml of 1.5 M solution in
toluene, 1.74 mmol) was added to a solution of mesoporphyrin IX
dimethyl ester (500 mg, 0.84 mmol) in CH.sub.2Cl.sub.2 (10 ml) at
-78.degree. C., the mixture was stirred at this temperature for 1
h, then quenched with a saturated solution of NH.sub.4Cl (1 ml),
followed by a 3.7% solution of HCl (2 ml). The temperature of the
reaction mixture was allowed to rise to 20.degree. C., the product
was extracted with CH.sub.2Cl.sub.2, dried (Na.sub.2SO.sub.4), and
purified on a column of silica gel using ethyl acetatelhexane:3/7
as eluant, 330 mg (73%) of 32 were obtained; .sup.1HNMR
(CDCl.sub.3): 1.86 (t, J=7.6 Hz, 6H), 3.39 (t, J=7.4 Hz, 6H), 3.60
(s, 6H), 3.62 (s, 6H), 4.04.2 (m, 4H), 4.5-4.45 (m, 4H), 9.97 (s,
1H), 10.04 (s, 1H), 10.05 (s, 1H), 10.06 (s, 1H), 10.065 (s, 1H),
10.07 (s, 1H).
Example 33
SL-11209 dodecahydrochloride
[0111] Amine 31 (182 mg, 0.24 mmol) and dialdehyde 32 (58 mg, 0.11
mmol) were mixed in 1,2-dichloroetane (3 mL) and sodium
triacetylborohydride (60 mg, 0.28 mmol) was added at 22.degree. C.,
the mixture was stirred for 3.5 h and then quenched with a solution
of sodium bicarbonate. The reaction mixture was diluted 3 times
with chloroform, washed with H.sub.2O, dried (Na.sub.2SO.sub.4) and
concentrated to dryness in vacuo. The residue was dissolved in
methylene chloride, cooled to 0.degree. C. and trifluoroacetic acid
added. After stirring for 1.5 h, the cooling bath was removed, the
mixture was evaporated to dryness.
[0112] The residue was dissolved in 10% HCl, the aqueous layer
washed with chloroform, and the water removed in vacuo; 134 mg
(74%) of crude SL-11209 were obtained. The product was purified by
HPLC (Column: 21.5 mm.times.250 mm, C.sub.18 Dynamax, eluent A=0.1%
TFA, eluent B=0.088% TFA in 90% acetonitrile). The pure product was
dissolved in 10% HCl (5 mL), and evaporated to dryness in vacuo.
.sup.1H NMR (D20): 1.16 (t, J=7.0 Hz, 6H), 1.34 (t, J=7.3 Hz, 6H),
1.60-2.00 (m, 38H), 2.50-2.70 (m, 4H), 2.90-3.30 (m, 40H) 3.50-3.65
(m, 4H), 3.75 (s, 6H), 3.82 (s, 6H), 4.20-4.35 (m, 4H), 4.45-4.60
(m, 4H), 10.5 (bs, 4H). MS (MALDI), 1240.6 [M+Na].sup.+, 1218.4,
[M+1].sup.+.
SLIL-11210 dodecahydrochloride
Example 34
1,3,5,8-Tetramethyl-Z
7-diethyl-6,7-bis[.sup.3'N,.sup.8'N,.sup.13'N,.sup.18'N-tetrakis
(mesitylenesulfonyl)-3',8',13',18',23'-pentaazaheptaeicosane]porphyrin
34
[0113] A solution of nitrile 33 (1.6 g, 1.5 mmol) (U.S. PAT APPL.
60/329,982) in ethanol (90 ml) and chloroform (1.6 ml) was
hydrogenated in the presence of PtO.sub.2 (160 mg) under 50 psi for
10 h, the suspension filtered through a celite cake, evaporated to
dryness and dried in vacuo. The product was dissolved in
1,2-dichloroetane (10 mL), dialdehyde 32 (370 mg, 0.69 mmol) was
added followed by triethylamine (0.23 ml, 1.67 mmol). The reaction
was stirred for 20 h, after which sodium triacetylborohydride (352
mg, 1.66 mmol) was added and the mixture further stirred for 3.5 h.
The reaction mixture was quenched with a solution of sodium
bicarbonate, thrice its volume of chloroform was added; the organic
layer was washed with H.sub.2O, dried, and evaporated to dryness in
vacuo. The residue was dissolved in methylene chloride (20 mL),
cooled to 0.degree. C., made basic with 2N sodium hydroxide (5 mL)
and mesitylsulfonyl chloride (333 mg, 1.5 mmol) was added. After 10
h of stirring at 22.degree. C. and following the usual workup the
reaction product was purified by column chromatography on silica
gel using chloroform/ethyl acetate; 9/1 as eluant; 729 mg (35%) of
34 were obtained; .sup.1H NMR (CDCl.sub.3): 0.93 (t, J=7.14 Hz),
1.05-1.50 (m,) 1.50-1.70 (m), 1.90 (t, J=7.14 Hz), 2.05 (s), 2.07
(s), 2.08 (s), 2.13 (s), 2.16 (s), 2.21 (s), 2.24 (s), 2.29 (s),
2.41 (s), 2.45 (s), 2.49 (s), 2.52 (s), 2.70-3.10 (m), 3.10-3.25
(m), 3.40-3.52 (m), 3.53 (s), 3.54 (s), 3.66 (s), 3.85-4.00 (m),
4.0-4.2 (m), 5.97 (s), 6.02 (s), 6.75 (s), 6.79 (s), 6.84 (s), 6.87
(s), 6.92 (s), 9.69 (s), 10.08 (s), 10.14 (s); MS (MALDI), 3007.02
[M+Na].sup.+, 2985.05 [M+1].sup.+, 2983.95 [M].sup.+, 1493.58
[M].sup.2+.
Example 35
SL-11210 dodecahydrochloride
[0114] SL-11210 was prepared from 34 following the procedure
described for the synthesis of SL-11211. From 730 mg of 34 were
obtained 360 mg (69%) of the dodecahydrobromide; .sup.1HNMR (D20):
.delta. 1.34 (t, J=7;3 Hz, 6H), 1.70-2.00 (m, 38H), 2.50-2.70 (m,
4H), 3.05-3.35 (m, 36H), 3.40-3.55 (m, 4H), 3.78 (2s, 6H), 3.82
(2s, 6H), 4.20-4.40 (m, 4H), 4.40-4.60 (m, 4H), 10.40 (bs, 4H). MS
(free base, MALDI), 1161.95 [M].sup.+. The dodecahydrobromide was
converted into dodecahydrochloride after HPLC purification and
treatment of the eluate with 20% HCl. MS (free base, MALDI),
1162.02 [M].sup.+, 581.82 [M].sup.2+.
Example 36
SL-11257 dodecahydrochloride
[0115] Amine 18 (310 mg, 0.44 mmol), dialdehyde 32(118 mg, 0.22
mmol), and triethylamine (0.16 ml) were dissolved in 27 ml of
dichloroethane. The reaction was kept at 22.degree. C. during 18 h,
sodium triacetoxyborohydride (186 mg, 10.9 mmol) was then added,
the reaction mixture was kept for further 2 h, it was then diluted
with chloroform, the solution washed with saturated sodium
bicarbonate, dried (Na.sub.2SO.sub.4), and evaporated to dryness.
The residue was purified by column chromathography on silica gel
using chloroform/methanol/ammonium hydroxide; 8/2/0.3 as eluant;
190 mg of SL-11257 were obtained. After purification by HPLC, 90 mg
(20%) of pure material were obtained; MS (MALDI): 1551.8
(M+H.sup.+, M=free base), 311.18 (M.sup.+/2), 388.79(M.sup.+/4),
517.8 (M.sup.+/3), 776.03 (M.sup.+/2).
Example 37
MTT Assay
[0116] A conventional MTT assay was used to evaluate percent cell
survival. Exponentially growing monolayer cells were plated in
96-well plates at a density of 500 cells per well and allowed to
grow for 24 hours. Serial dilutions of the drugs were added to the
wells. Six days after drug treatment, 25 .mu.l of MTT solution (5
mg/ml) was added to each well and incubated for 4 hours at
37.degree. C. Then 100 .mu.l of lysis buffer (20% sodium dodecyl
sulfate, 50% DMF, and 0.8% acetic acid, pH 4.7) was added to each
well and incubated for an additional 22 hours. A microplate reader
("EMAX"-brand, Molecular Devices, Sunnyvale, Calif.) set at 570 nm
was used to determine the optical density of the cultures. Results
are expressed as a ratio of the optical density in drug-treated
wells to the optical density in wells treated with vehicle only.
Tables 1, 2, and 3 below describe the results of the assays on
various cell lines. FIGS. 1-13 also indicate the effects of the
compounds on various cell lines.
[0117] Other suitable assays for testing the compounds of the
invention are described in International Patent Application Nos. WO
00/66587 and WO 02/10142, and U.S. Pat. Nos. 6,392,098, 5,889,061,
and 5,677,350 TABLE-US-00001 TABLE 1 Effect of Porphyrin Polyamine
Analogues on Human Prostate Tumor Cell Growth by the MTT assay
ID.sub.50 (.mu.M) values for Human prostate tumor Cell Lines
Tsu-pr1- Compounds DuPro PC-3 DU145 LnCap Tsu-pr1 ADR SL-11209 1.4
1.7 -- -- -- SL-11211 0.46 1.7 -- -- 0.35 0.66 SL-11217 3.6 2.8 --
-- -- -- SL-11233 1.4 3.9 >31.25 >31.25 -- -- SL-11235 0.12
0.45 0.17 0.2 -- -- SL-11236 0.08 0.49 0.14 0.2 -- -- SL-11237 1.9
1.7 -- -- 1.87 12.62 -- Not done
[0118] TABLE-US-00002 TABLE 2 Effect of Porphyrin Polyamine Analogs
on Human Pancreatic Cancer Cell Growth by MTT Assay.
ID.sub.50(.mu.M) values for Human Pancreatic Cancer Cell Lines
Compounds BxPC-3 Panc-1 SL-11217 6.87 6.38 SL-11237 5.92 12.45
[0119] TABLE-US-00003 TABLE 3 Effect of Porphyrin Polyamine Analogs
on Human Brain Tumor Cell Growth by MTT Assay. ID.sub.50(.mu.M)
values for Human Brain Tumor Cells U251MG Compounds NCI SL-11217
5.65 SL-11237 2.30
Example 38
Oral Administration of SL-11237
[0120] Male athymic nude mice were given subcutaneous injections of
0.75.times.10.sup.6 DU145 cells on Day 0. Beginning on Day 10, mice
were treated once weekly for 3 weeks with acidified water, 100
mg/kg, or 500 mg/kg of SL-11237 via oral gavage at 10 ml/kg dosing
volume (the third treatment was actually 400 mg/kg in the high dose
group). The results are depicted in FIG. 14, where the top panel
depicts average tumor volume in the mice. The bottom panel of FIG.
14 depicts average body weight of the mice. Oral administration
thus provides an effective and convenient means of administering
the compounds of the invention.
[0121] All references, publications, patents and patent
applications mentioned herein are hereby incorporated by reference
herein in their entirety.
[0122] Although the foregoing invention has been described in some
detail by way of illustration and example for purposes of clarity
and understanding, it will be apparent to those skilled in the art
that certain changes and modifications may be practical. Therefore,
the description and examples should not be construed as limiting
the scope of the invention, which is delineated by the appended
claims.
* * * * *