Organic compounds -- part of the class 532-570 series – Organic compounds – Fatty compounds having an acid moiety which contains the...
Reexamination Certificate
1999-05-14
2002-03-19
Carr, Deborah D. (Department: 1621)
Organic compounds -- part of the class 532-570 series
Organic compounds
Fatty compounds having an acid moiety which contains the...
C554S213000, C554S224000
Reexamination Certificate
active
06359158
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to 16-HETE analogs which are agonists and antagonists of 16-HETE. The invention also relates to methods and products for inhibiting neutrophil adhesion and neutrophil aggregation using the 16-HETE agonists.
BACKGROUND OF THE INVENTION
The inflammatory response is an important element of a host's natural defense mechanism against pathogens. It also is involved in wound healing. Despite the beneficial role that the inflammatory response plays in host survival, excessive inflammation may have clinically adverse results in some medical conditions.
Leukocytes are a major cellular component of inflammatory and immune responses. This class of cells includes neutrophils, lymphocytes, monocytes, eosinophils, and basophils. Neutrophils, which play a key role in the inflammatory response, are generally present within the body in a resting unstimulated state. Once stimulated, the neutrophils migrate to the site of injury and release toxic factors.
The migratory capability of a neutrophil is dependent on the ability of the neutrophil to alter its adhesive properties. In a resting unstimulated state a neutrophil is not adhesive and cannot migrate. Once the neutrophil has been stimulated, however, it becomes more adhesive and is capable of migrating. The increase in neutrophil adhesiveness causes the stimulated neutrophil to aggregate and to adhere to endothelium. Stimulation of the neutrophil also causes the neutrophil to undergo diapedesis, which involves the migration of the neutrophil between post-capillary endothelial cells into the tissues.
In the tissues, an activated neutrophil releases enzymes such as collagenase and elastase, among others. Neutrophil stimulation may also initiate a burst of oxygen consumption, with concomminant activation of the hexose-monophosphate shunt and activation of nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase. Activation of these systems results in the formation and release of factors such as hydrogen peroxide and hydroxyl radicals, which are toxic to microorganisms and tumor cells, and thereby facilitating the destruction of the injury causing agent.
Several studies have focused on analyzing the control and regulation of the adhesive properties of neutrophils. Much of this research has centered on adhesion receptors and also on metabolites of arachidonic acid such as C20 carbon fatty acid found in every cell membrane. Arachidonic acid metabolism occurs by different mechanisms in stimulated versus unstimulated neutrophils and results in the production of a different spectrum of metabolites in stimulated versus unstimulated neutrophils.
In stimulated neutrophils, the cytochrome P450 mixed function oxidase system appears to be more active. Moreover, during neutrophil stimulation, 5-lipoxygenase is translocated to the membrane compartment fraction, where it produces 5-hydroperoxyeicosatetraenoic acid (5-HPETE). 5-HPETE is then either metabolized to 5-hydroxyeicosatetraenoic acid (5-HETE) by peroxidase or dehydrated to form leukotriene A
4
. Leukotriene A
4
is converted into leukotriene B
4
which is a potent chemotactic agent and promoter of neutrophil adhesion.
In unstimulated neutrophils, the metabolism of arachidonic acid is markedly different than that in stimulated neutrophils. The metabolism of arachidonic acid in unstimulated neutrophils is sensitive to cytochrome P450 inhibitors but not to cyclooxygenase or lipoxygenase inhibitors. Hatzelmann and Ullrich characterized the metabolites produced in unstimulated neutrophils, reporting the finding that arachidonic acid is metabolized to 20-HETE and 15-HETE.
Hatzelmann, Eur. J. Biochem.
173, 445-452 (1988). Another study, Kraemer et al., found that the arachidonic acid metabolic products formed in unstimulated neutrophils exhibited a potent anti-aggregatory activity toward human neutrophils, suggesting that the identified arachidonic acid metabolites may play some role in the regulation of neutrophil adhesion and aggregation properties.
Kraemer et al., Am. J. Pathol.
128, 446-454 (1987).
SUMMARY OF THE INVENTION
It was recently discovered in co-pending U.S. patent application Ser. No. 08/652,327, filed May 22, 1996 and issued as U.S. Pat. No. 5,753,702 on May 19, 1998 and PCT Patent Application No. PCT/US97/08865, and its related national Stage U.S. patent application Ser. No. 09/194,166, the entire contents of which are hereby incorporated by reference, that 16-HETE (16-hydroxyeicosatetraenoic acid) is a component of arachidonic acid metabolism in neutrophils and that 16-HETE is a potent inhibitor of neutrophil adhesion and neutrophil aggregation. It was also disclosed in these applications that 16-HETE when administered alone actually reduces the size of brain infarcts resulting from acute stroke relative to the size of brain infarcts which occur in the absence of a therapeutic. When 16-HETE is administered in combination with clot lysing thrombolytic agents such as tPA, the therapeutic combination actually reduces the size of brain infarcts resulting from acute stroke relative to the size of brain infarcts which occur in a subject suffering from an acute stroke who has been treated only with a thrombolytic agent such as tPA.
The present invention relates to novel analogs of 16-HETE. Some of the analogs of 16-HETE are agonists which maintain the biological activity of 16-HETE but which are more stable and have longer half-lives. The 16-HETE analogs also include 16-HETE antagonists which inhibit the activity of 16-HETE. These antagonists are useful when it is desirable to prevent inhibition of neutrophil activity.
According to one aspect of the invention, compositions are provided. These compositions include the following 16-HETE analog:
wherein R is selected from the group consisting of —C(O)—X—SO
2
—R
1
, —C(O)—X—CO—R
1
, —C(O)—X—C(OH)
2
—R
1
, —C(O)—X—C(NH)
2
—R
1
, —C(O)—X—C(NH
2
)
2
—R
1
, piperonyl, —CN, —OR′, —SR′, —NO
2
, —NR′R′, amino acid, —C(O)R′, —C(S)R′, —C(O)OR′, —C(S)OR′, —C(O)SR, —C(S)SR′, —C(O)N(R′)
2
, —C(O)C(O)R′, —C(S)C(O)R′, —C(O)C(S)R′, —C(S)C(S)R′, —C(O)C(O)OR′, —C(S)C(O)OR′, —C(O)C(S)OR′, —C(O)C(O)SR′, —C(S)C(S)OR′, —C(S)C(O)SR′, —C(O)C(S)SR′, —C(S)C(S)SR′, —C(O)C(O)N(R′)
2
, —C(S)C(O)N(R′)
2
, —C(O)C(S)N(R′)
2
, or —C(S)C(S)N(R′)
2
; wherein X is selected from the group consisting of O, N, and a bond; wherein R
1
, R
2
, and R
3
each independently is selected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl, alkoxy, aryl, and heteroaryl; wherein each R′ is (CH
2
)
z
—NR″R″ and wherein R″ is independently selected from the group consisting of (C
1
-C
6
) alkyl, (C
1
-C
6
) alkenyl, (C
1
-C
6
) alkoxy, (C
1
-C
6
) alkynyl, (C
6
-C
20
) aryl, (C
6
-C
20
) substituted aryl, (C
6
-C
26
) alkaryl, substituted (C
6
-C
26
) alkaryl, and (C
5
-C
7
) heteroaryl.
16-HETE analogs include both agonists and antagonists. In some embodiments the 16-HETE agonists have the following general structure:
wherein either R
1
or R
2
is a C
3
alkyl and the other is a hydrogen.
In one embodiment R
1
, R
2
, and R
3
each independently is selected from the group consisting of hydrogen, (C
1
-C
6
) alkyl, (C
1
-C
6
) alkenyl, (C
1
-C
6
) alkynyl, and (C
1
-C
6
) alkoxy.
In another embodiment the 16-HETE analog has the following structure:
In a preferred embodiment X is NH and R
2
and/or R
3
is hydrogen. In another preferred embodiment R
3
is a C
3
alkyl. Preferably, the 16-HETE analog has the following structure:
In a preferred embodiment X is O and R
2
and/or R
3
is hydrogen. In another preferred embodiment R
3
is a C
3
alkyl.
In other preferred embodiments the 16-HETE analog is one of the following structures:
In another embodiment the 16-HETE analog is a 16-HETE antagonist having the following general structure:
wherein R
4
and R
5
each independently is selected from the group consisting of hydrogen, C
1-2
alkyl, C
4-6
Balazy Michael
Bednar Martin M.
Falck John R.
Gross Cordell E.
Carr Deborah D.
University of Vermont and State Agricultural College
Wolf Greenfield & Sacks P.C.
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