BH3 interacting domain death agonist

• US 5,998,583 A
• Filed: 09/05/1997
• Issued: 12/07/1999
• Est. Priority Date: 09/09/1996
• Status: Expired due to Term

• Current Assignee: Washington University In St Louis
• Original Assignee: Washington University In St Louis
• Inventors: Korsmeyer, Stanley J.
• Primary Examiner(s): Hutzell, Paula K.
• Assistant Examiner(s): Worrall, Timothy A.
• Application Number:
  US08/924,695
• Time in Patent Office: 823 Days
• Field of Search: 424/94.1, 424/185.1, 424/192.1, 435/4, 435/69.4, 435/69.7, 435/69.8, 435/71.1, 435/69.1, 514/2, 530/300, 530/324, 530/325, 530/326, 530/327, 530/328, 530/350, 530/358
• US Class Current: 530/350
• CPC Class Codes: A61P 15/00 Drugs for genital or sexual disorders for disorders of sex hormones A61P5/24; Contraceptives
• CPC Class Codes: A61P 17/02 for treating wounds, ulcers, burns, scars, keloids, or the like
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/00 Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 9/10 for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: C07K 14/4747 Apoptosis related proteins
• CPC Class Codes: C07K 2319/00 Fusion polypeptide

Abstract

A novel death agonist, BID, which contains only a BH3 domain but not the BH1, BH2, BH4 or membrane anchoring domains found in related BCL-2 family members is provided along with derivatives of BID, bid polynucleotides and derivatives thereof and compositions and uses therefor.

Claims

1. An isolated and purified BID (BH3 Interacting domain Death agonist) polypeptide comprising an amino acid sequence which is at least 85 percent identical to the amino acid sequence set forth in SEQ ID NO:

4.

2. The isolated and purified BID polypeptide of claim 1 wherein said polypeptide is at least 95 percent identical to the amino acid sequence set forth in SEQ ID NO:

4.

3. An isolated and purified BID polypeptide which (a) lacks the carboxyl terminal signal-anchor sequence characteristic of the membrane bound members of the BCL-2 family, (b) lacks BH1 and BH2 domains, (c) has a BH3 domain, and (d) heterodimerizes with BAX, BCL-2 or BCL-X_L.

4. The isolated and purified BID polypeptide of claim 3 comprising an immunogenic peptide selected from the group consisting of:

(SEQ ID NO;

29);

(SEQ ID NO;

30);

(SEQ ID NO;

31);

(SEQ ID NO;

32);

(SEQ ID NO;

33);

(SEQ ID NO;

34);

(SEQ ID NO;

35);

(SEQ ID NO;

36); and

(SEQ ID NO;

37).

5. The isolated and purified BID polypeptide of claim 3 comprising a sequence which is at least 65% identical to SEQ ID NO:

4

(-MDCEVNNGSSLRDECITNLLVFGFLQSCSDNSFRRELDALGHELPVLAPQWEGYDELQTDGNRSSHSRLGRIEADSESQEDIIRNIARHLAQVGDSMDRSIPPGLVNGLALQLRNTSRSEEDRNRDLATALEQLLQAYPRDMEKEKTMLVLALLLAKKVASHTPSLLRDVFHTTVNFINQNLRTYVRSLARNGMD-).

6. The isolated and purified BID polypeptide of claim 5 as set forth in SEQ ID NO:

4

(MDCEVNNGSSLRDECITNLLVFGFLQSCSDNSFRRELDALGHELPVLAPQWEGYDELQTDGNRSSHSRLGRIEADSESQEDIIRNIARHLAQVGDSMDRSIPPGLVNGLALQLRNTSRSEEDRNRDLATALEQLLQAYPRDMEKEKTMLVLALLLAKKVASHTPSLLRDVFHTTVNFINQNLRTYVRSLARNGMD).

7. The isolated and purified BID polypeptide of claim 3 comprising a sequence which is at least 65% identical to SEQ ID NO:

5

(-MDCEVNNGSSLRDECITNLLVFGFLQSCSDNSFRRELDALGHELPVLAPQWEGYDELQTDGNRSSHSRLGRIEADSESQEDIIRNIARHLAQVGDSMDRSIPPGLVNGLALQLRNTSRSEEDRNRDLATALEQLLQAYPRDMEKEKTMLVLALLLAKKVASHTPSLLRDVFHTTVNFINQNLRTYVRSLARNVRTLEGMD-)

or at least 65% identical to SEQ ID NO;

6

(-MDSEVSNGSGLGAKHITDLLVFGFLQSSGCTRQELEVLGRELPVQAYWEADLEDELQTDGSQASRSFNQGRIEPDSESQEEIIHNIARHLAQIGDEMDHNIQPTLVRQLAAQFMNGSLSEEDKRNCLAKALDEVKTAFPRDMENDKAMLIMTMLLAKKVASHAPSLLRDVFHTTVNFINQNLFSYVRNLVRNEMD-).

8. The isolated and purified BID polypeptide of claim 7 as set forth in SEQ ID NO:

5

(MDCEVNNGSSLRDECITNLLVFGFLQSCSDNSFRRELDALGHELPVLAPQWEGYDELQTDGNRSSHSRLGRIEADSESQEDIIRNIARHLAQVGDSMDRSIPPGLVNGLALQLRNTSRSEEDRNRDLATALEQLLQAYPRDMEKEKTMLVLALLLAKKVASHTPSLLRDVFHTTVNFINQNLRTYVRSLARNVRTLEGMD)

or SEQ ID NO;

6

(MDSEVSNGSGLGAKHITDLLVFGFLQSSGCTRQELEVLGRELPVQAYWEADLEDELQTDGSQASRSFNQGRIEPDSESQEEIIHNIARHLAQIGDEMDHNIQPTLVRQLAAQFMNGSLSEEDKRNCLAKALDEVKTAFPRDMENDKAMLIMTMLLAKKVASHAPSLLRDVFHTTVNFINQNLFSYVRNLVRNEMD).

12. An isolated and purified fusion protein comprising a BID polypeptide according to claim 5 covalently linked to a heterologous polypeptide sequence which enhances intracellular delivery of the BID polypeptide.

9. An isolated and purified polypeptide comprising an immunogenic fragment of at least 10 contiguous amino acids of a BID polypeptide as set forth in SEQ ID NO:

4

(MDCEVNNGSSLRDECITNLLVFGFLQSCSDNSFRRELDALGHELPVLAPQWEGYDELQTDGNRSSHSRLGRIEADSESQEDIIRNIARHLAQVGDSMDRSIPPGLVNGLALQLRNTSRSEEDRNRDLATALEQLLQAYPRDMEKEKTMLVLALLLAKKVASHTPSLLRDVFHTTVNFINQNLRTYVRSLARNGMD),

SEQ ID NO;

5

(MDCEVNNGSSLRDECITNLLVFGFLQSCSDNSFRRELDALGHELPVLAPQWEGYDELQTDGNRSSHSRLGRIEADSESQEDIIRNIARHLAQVGDSMDRSIPPGLVNGLALQLRNTSRSEEDRNRDLATALEQLLQAYPRDMEKEKTMLVLALLLAKKVASHTPSLLRDVFHTTVNFINQNLRTYVRSLARNVRTLEGMD),

or SEQ ID NO;

6

(MDSEVSNGSGLGAKHITDLLVFGFLQSSGCTRQELEVLGRELPVQAYWEADLEDELQTDGSQASRSFNQGRIEPDSESQEEIIHNIARHLAQIGDEMDHNIQPTLVRQLAAQFMNGSLSEEDKRNCLAKALDEVKTAFPRDMENDKAMLIMTMLLAKKVASHAPSLLRDVFHTTVNFINQNLFSYVRNLVRNEMD).

10. The isolated and purified fragment of claim 9 which contains a BH3 domain as set forth in SEQ ID NO:

8 or SEQ ID NO;

41.

11. The isolated and purified fragment of claim 11 which has the sequence as set forth in SEQ ID NO:

51, SEQ ID NO;

52 or SEQ ID NO;

53.

14. The isolated and purified fusion protein according to claim 14 wherein the heterologous polypeptide sequence is a Tat peptide as set forth in SEQ ID NO:

54 (YGRKKRRQRRR).

13. The isolated and purified fusion protein according to claim 14 wherein the BID polypeptide is as set forth in SEQ ID NO:

55 (DSESQEEIIHNIARHLAQIGDEMDHNIQPTLV).

15. The isolated and purified fusion protein of claim 16 comprising the sequence as set forth in SEQ ID NO:

56 (YGRKKRRQRRRGDSESQEEIIHNIARHLAQIGDEMDHNIQPTLV).