Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Peptide containing doai
Reexamination Certificate
2000-04-07
2003-01-21
Low, Christopher S. F. (Department: 1653)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Peptide containing doai
C514S011400, C530S317000, C530S322000, C530S323000, C530S324000, C530S402000
Reexamination Certificate
active
06509315
ABSTRACT:
BACKGROUND OF THE INVENTION
Didemnin B is a macrocyclic depsipeptide isolated from a species of marine tunicate. Didemnin B exhibits potent anti-viral, immunosuppressive, and anti-tumor activities in vitro and in vivo, and was the first marine natural product to enter clinical testing against human cancers (Li et al., 1992, Studies in Natural Products Chemistry, 10:241-302; Sakai et al., 1996, J. Med. Chem. 39:2819-2834; Wipf, 1995, Chem. Rev. 95:2115-2134). Didemnin B is a didemnin, a family of compounds which potently inhibit protein synthesis and cell cycle progression, and induce more rapid apoptosis than any other natural products that has been isolated to date (Grubb et al., 1995, Biochem. Biophys, Res. Commun. 215:1130-1136; Johnson et al., 1996, FEBS Lett. 383:1-5; Johnson et al., 1999, Immunol. Cell Biol. 77:242-248; Johnson et al., 1999, J. Cell. Biochem. 72:269-278). Other members of this family of compounds, including didemnin M and dehydrodidemnin B, exhibit cytotoxic and cytostatic effects as well.
Tamandarin A (also designated {(2S)HIV
2
}didemnin B) is a naturally occurring didemnin analog which has recently been isolated from a marine tunicate. Tamandarin A exhibits biological activity which is analogous to the activities exhibited didemnin B. For example, tamandarin A is a potent inhibitor of protein synthesis, cell growth, and tumorigenesis. Tamandarin A exhibits greater in vitro activity against pancreatic carcinoma than does didemnin B (Liang et al., 1999, Org. Lett. 1: 1319-1322). A significant limitation on use of tamandarin A, either for research or for practical applications, is the limited supply of tamandarin A that is available from natural sources and the difficulty and expense of isolating this product. A need exists for a method of synthesizing tamandarin A and other didemnin analogs (including dehydrodidemnin analogs).
Despite the potency of didemnin B in isolated studies, its clinical effectiveness is hampered by side effects associated with therapeutic doses of the compound. As with many anti-proliferative agents, didemnin B exhibits a relatively narrow therapeutic window. Although didemnin M and dehydrodidemnin B exhibit improved therapeutic potential, relative to didemnin B, a need still exists for anti-proliferative agents which exhibit less toxicity at a therapeutic dose (i.e. didemnin analogs having a greater therapeutic index).
The present invention satisfies the needs set forth above.
BRIEF SUMMARY OF THE INVENTION
The invention relates to a composition comprising a didemnin analog having the structure of formula I
In formula I, R
1
is selected from the group consisting of
—H,
—(tert-butyloxycarbonyl),
—leucine,
—(N-methyl)leucine,
—(N-methyl)leucine-proline,
—(N-methyl)leucine-proline-lactate,
—(N-methyl)leucine-proline-pyruvate,
—(N-methyl)leucine-proline-lactate-(a first fluorophore),
—(N-methyl)leucine-proline-lactate-glutamine-pyroglutamate,
—(N-methyl)leucine-proline-lactate-glutamine-cyclopentanoate,
—(N-methyl)leucine-proline-alanine-leucine-pyroglutamate, and
—(N-methyl)leucine-proline-(N-methyl-alanine)leucine-pyroglutamate.
R
2
and R
3
in formula I, can be separate moieties or they can, together, be a single moiety. When R
2
and R
3
are separate moieties, R
3
is either a methyl group or a hydride radical and R
2
is selected from the group consisting of an isoleucine side chain, a valine side chain, an alanine side chain, a norleucine side chain, a norvaline side chain, leucine side chain, a histidine side chain, a tryptophan side chain, an arginine side chain, a lysine side chain, a second fluorophore, and a substituent having the structure of formula III
When R
2
and R
3
are, together, a single substituent, this substituent has the structure of formula IV
A In formulas III and IV, each of R
5
, R
6
, R
7
, R
8
, and R
9
is independently selected from the group consisting of —H, —OH, —OCH
3
, —CO(C
6
H
5
), —Br, —I, —F, —Cl, CH
3
, and —C
2
H
5
.
R
4
in formula I is either an isoleucine side chain or a valine side chain. Also, in formula I, X is either —O— or —(NH)—, Y is either a hydride radical or a hydroxyl protecting group, and R
10
is either a leucine side chain or a lysine side chain. The didemnin analog is an analog other than tamandarin A (i.e. {(2S)Hiv
2
}didemnin B). In one embodiment, every proline or lactate moiety that is present in R
1
exhibits (S) stereochemistry. In another, every moiety capable of exhibiting stereochemistry in R1 is present in its naturally occurring form (i.e. the (S) form for amino acid residues and lactate. It is believed that cyclopentanoate occurs naturally in an (S) stereochemistry.
Examples of didemnin analogs that are included in the invention are compound 103, compound 104, compound 105, compound 106, compound 107, compound 108, compound 109, compound 110, compound 115, compound 116, compound 117, compound 118, compound 119, compound 120, compound 121, compound 122, compound 123, compound 124, compound 125, compound 126, compound 127, compound 128, compound 129, compound 130, compound 133, compound 134, compound 136, compound 137, compound 139, compound 141, compound 142, compound 143.
In one embodiment, the didemnin analog has a photoreactive substituent, such as an R
2
moiety having the structure
In another embodiment, the didemnin analog has a fluorophore attached, such as an analog in which a fluorophore is attached at the omega amino moiety of a lysine side chain at R
2
or at R
10
. An example of the structure of such a fluorescent didemnin analog is show in FIG.
29
. Alternatively, the didemnin analog can be attached (e.g. covalently) with a support. In most embodiments, Y in formula I is preferably a hydride radical.
The invention includes an embodiment of a didemnin analog which can be activated (or the activity of which can be enhanced) by enzymatic cleavage of a moiety bound with the analog. For example, the invention includes compositions which comprise a didemnin analog having the structure of formula II
In formula II, R
2
, R
3
, R
4
, R
10
, X, and Y have the same identities described above for formula I. R
13
is an enzyme-cleavable moiety that is cleavable by an enzyme, such as one selected from the group consisting of a carboxypeptidase, a beta-lactamase, a beta-galactosidase, a penicillin V-amidase, a cytosine deaminase, a nitroreductase, a alkaline phosphatase, a beta-glucuronidase, and a catalytic antibody. By way of example, R
13
can have the structure of either of formulas V and VI
Examples of such didemnin analogs that include compound 131 and compound 132.
The invention also relates to compositions which comprise a didemnin fragment having the structure of formula VII
In formula VII, Y is either a hydride radical or a hydroxyl protecting group, X is either —O— or —(NH)—, R
4
is either an isoleucine side chain or a valine side chain, and APG is a amine protecting group. R
11
can be any of —OH, —NH
2
, —O(allyl), —O(pentafluorophenyl), and a substituent having the structure of formula VIII
In formula VIII, R
1
, R
2
, R
3
, and R
10
have the same identities described above for formula I, and R
12
can be either a hydride radical or a -2-(trimethylsilyl)ethoxycarbonyl moiety.
The didemnin analogs and fragments described herein can be formulated, together with one or more pharmaceutically acceptable carriers, to make pharmaceutical preparations. These preparations can be administered to a mammalian (e.g. human) cell (i.e. either in vitro or in vivo) in order to inhibit protein synthesis, inhibit growth, inhibit proliferation, inhibiting tumorigenesis, or enhance apoptosis in the cell or in one or more tissues of the mammal.
The invention further includes a method of making a didemnin fragment. This method comprises coupling a first reactant having the structure
and a second reactant having the structure
to yield a first didemnin fragment having the structure
In this structure, X is either —O— or —(NH)—, APG is an amine protecting group; Y is a hydroxyl protecting group (e.g. a -triisopropylsilyl group), and R
4
Joullie Madeleine M.
Liang Bo
Akin Gump Strauss Hauer & Feld L.L.P.
Low Christopher S. F.
Mohamed Abdel A.
The Trustees of the University of Pennsylvania
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