Organic compounds -- part of the class 532-570 series – Organic compounds – Heterocyclic carbon compounds containing a hetero ring...
Reexamination Certificate
2002-03-26
2003-10-07
Davis, Zinna Northington (Department: 1625)
Organic compounds -- part of the class 532-570 series
Organic compounds
Heterocyclic carbon compounds containing a hetero ring...
C548S566000, C546S276400, C560S021000, C564S050000
Reexamination Certificate
active
06630589
ABSTRACT:
BACKGROUND OF THE INVENTION
The invention relates to the fields of screening assays and compounds and methods for treating cancer and other proliferative diseases.
The regulation of protein expression can occur at a number of levels: transcriptional, post-transcriptional, or post-translational. The modulation of protein expression is often critical for the treatment of disease. To this end, one area of research has been directed at developing small molecules that regulate transcription factors. Another approach to modulating protein expression is achieved by targeting the RNA encoding the target protein using antisense technology.
Another manner in which protein expression is regulated is through modulating translation efficiency. In eukaryotes, translational regulation usually occurs at the initiation step. Therefore, cis-elements in the 5′ untranslated region (UTR) are important for overall regulation of protein synthesis. There is also a growing body of evidence that supports the role of the poly(A) tail and elements in the 3′ UTR in regulating initiation of translation
HER2 overexpression occurs in a number of primary human tumors, including 25-30% of breast and ovarian carcinomas, and is associated with an adverse prognosis and rapid tumor growth. HER2 is regulated at both the transcriptional and post-transcriptional level. For example, it has been previously shown that HER2 mRNA is translated more efficiently in transformed cells than in primary cells. Therefore, therapeutics that decrease HER2 polypeptide levels within a cell would be valuable as drugs for the treatment of conditions such as cancer and other proliferative diseases.
In addition, there is a need for the identification of compounds that can be used to treat cancer and proliferative diseases, regardless of the mechanism of action.
SUMMARY OF THE INVENTION
The invention features screening assays for compounds that are potentially useful for treating or preventing a proliferative disease. In addition, the invention features compounds identified by the assays of the invention. The invention further features compounds structurally related to those identified by the screening assays. Finally, the invention features methods of treating or preventing a proliferative disease using the compounds of the invention.
Accordingly, in one aspect, the invention features a method of identifying a compound for treating or preventing a proliferative disease. This method includes the steps of (a) providing a cell, e.g., a mammary cell, a gastric cell, an ovarian cell, a bladder cell, a lung cell, a salivary gland cell, or a cancer cell, including a nucleic acid molecule containing a HER2 regulatory element operatively linked to a reporter coding sequence and stably integrated into a chromosome of the cell; (b) contacting the cell with a candidate compound; and (c) assaying for the effectiveness of said candidate compound for the treatment or prevention of said proliferative disease. In one embodiment, in step (a), the method further includes adding an RNA binding protein to the cell, under conditions that allow interaction between the nucleic acid molecule and the RNA binding protein. The interaction in step (a) between said nucleic acid molecule and said RNA binding protein may also be mediated by at least one polypeptide. The nucleic acid molecule including a HER2 regulatory sequence operatively linked to a reporter coding sequence may further include one or more additional regulatory elements. The HER2 regulatory element may, for example, include a HER2 5′ UTR, a HER2 3′ UTR, or a HER2 5′ UTR and a HER2 3′ UTR. The HER2 5′ UTR is, for example, SEQ ID NO: 1, and the HER2 3′ UTR is, for example, SEQ ID NO: 2.
In one embodiment, the assaying in step (c) is for a change in the level of translation efficiency of the reporter coding sequence relative to a cell not exposed to the candidate compound, and a modulation in the level of translation efficiency of said reporter coding sequence indicates a compound that modulates the translation efficiency of HER2. The modulation may be an increase or decrease in the level of translation efficiency of HER2. The reporter coding sequence is, for example, a luciferase, a &bgr;-galactosidase, or a green fluorescent protein reporter sequence. In one embodiment, the translation efficiency is assayed by polysomal distribution analysis, followed by detection of the level of reporter coding RNA expression. In another embodiment, the translation efficiency is assayed by detection of the level of reporter coding protein expression or activity.
In another embodiment, the assaying in step (c) is for toxicity of the candidate compound to the cell or the compounds ability to inhibit cell growth, relative to a cell not exposed to the compound. Step (c) may further include comparing the toxicity or growth inhibition of the candidate compound to the cell to the toxicity or growth inhibition of the compound to a reference cell. The toxicity or growth inhibition of a compound to a reference cell may be, for example, representative of the toxicity or growth inhibitory effect of the compound to the general cell population.
In another aspect, the invention features compounds of the formula:
or a pharmaceutically acceptable salt thereof, wherein U, V, Y, and Z are independently C, CH, N, O, or S; X is independently H, Hal, lower alkyl, OR
5
, SR
6
, or NR
7
R
8
, where R
5
, R
6
, R
7
, and R
8
are independently H or lower alkyl, and n is 1, 2, 3, or 4; R
1
and R
2
are independently H, Me, or Et, or R
1
and R
2
together are (CR
9
R
10
)
m
, where R
9
and R
10
are independently H, lower alkyl, CH
2
OR
11
, (CH
2
)
o
NR
12
R
13
, or OR
14
, where R
11
, R
12
, R
13
, and R
14
are H, lower alkyl, aryl, or alkaryl, m is 2, 3, 4, or 5, and o is 0 or 1; and Ar is an unsubstituted or substituted carbocyclic or heterocyclic aromatic ring or fused ring system. In one embodiment, U, V, Y, and Z are C or CH.
In another embodiment, the compound is of the formula:
where V, Y, and Z are independently N, CH, or C; B is N, CH, or CR
4
, where R
4
is F or lower alkyl; X is independently H, Hal, lower alkyl, OR
5
, SR
6
, or NR
7
R
8
, where R
5
, R
6
, R
7
, and R
8
are independently H or lower alkyl, and n is 1, 2, 3, or 4; R
1
and R
2
are independently H, Me, or Et, or R
1
and R
2
together are (CR
9
R
10
)
m
, where R
9
and R
10
are independently H, lower alkyl, CH
2
OR
11
, (CH
2
)
o
NR
12
R
13
, or OR
14
, where R
11
, R
12
, R
—, and R
14
are independently H, lower alkyl, aryl, or alkaryl, m is 2, 3, 4, or 5, and o is 0 or 1; and R
3
is H, lower alkyl, F, OR
15
, where R
15
is H, lower alkyl, or aralkyl, or a substituent of the structure:
where R
16
is NH, O, or S; R
17
is H, lower alkyl, or aralkyl; and W is lower alkyl, lower alkenyl, lower alkynyl, aryl, heteroaryl, alkaryl, OR
18
, or NR
19
R
20
, where R
18
, R
19
, and R
20
are independently H, lower alkyl, lower alkenyl, lower alkynyl, aryl, alkaryl, aralkyl, or (CH
2
)
k
CH
2
OR
21
, where R
21
is H or lower alkyl, and k is 1, 2, 3, or 4. In various embodiments, at least one of B, V, Y, and Z is N. In various other embodiments, when X
n
is H, V, Y, and Z are C or CH, R
1
and R
2
together are (CH
2
)
4
, B is CR
4
, and R
4
is F, R
3
is not F or H; when X
n
is H, V, Y, and Z are C or CH, R
1
and R
2
together are (CH
2
)
4
, B is CR
4
, and R
4
is Me, R
3
is not H; when X
n
is H, V, Y, and Z are C or CH, R
1
and R
2
together are (CH
2
)
4
, B is CR
4
, and R
4
is H, R
3
is not H or F; when X
n
is H, V, Y, and Z are C or CH, B is CR
4
, and R
1
, R
2
, and R
4
are Me, R
3
is not H; when X
n
is H, V, Y, and Z are C or CH, R
1
and R
2
are Me, B is CR
4
, and R
4
is F, R
3
is not F; when X
n
is H, V, Y, and Z are C or CH, R
1
and R
2
together are (CH
2
)
4
, B is CH, W is methyl, and R
16
is O, R
17
is not H; when X
n
is H, V, Y, and Z are C or CH, R
1
and R
2
are methyl, B is CH, W is methyl, and R
16
is O, R
17
is not H; when X
n
is H, V, Y, and Z are C or CH, B i
Giordano Anthony
Powers Gordon Donald
Sturgess Michael Alan
Yang Ke
Bieker-Brady Kristina
Clark & Elbing LLP
Davis Zinna Northington
Message Pharmaceuticals
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