(12) United States Patent ao) Patent No.: us 6,348,311 Bi
Kastan et al. (45) Date of Patent: Feb. 19,2002
(54) ATM KINASE MODULATION FOR SCREENING AND THERAPIES
(75) Inventors: Michael Kastan; Christine Canman;
Seong-Tae Kim; Dae-Sik Lim, all of
Cordova, TN (US)
(73) Assignees: St. Jude Childre's Research Hospital, Memphis, TN (US); Johns-Hopkins University, Baltimore, MD (US)
( * ) Notice: Subject to any disclaimer, the term of this patent is extended or adjusted under 35 U.S.C. 154(b) by 0 days.
(21) Appl. No.: 09/400,653
(22) Filed: Sep. 21, 1999
Related U.S. Application Data
(63) Continuation of application No. 09/248,061, filed on Feb. 10, 1999.
(51) Int. CI.7 C12Q 1/70; C12Q 1/48
(52) U.S. CI 435/5; 435/15
(58) Field of Search 435/15, 5
(56) References Cited
U.S. PATENT DOCUMENTS
5,728,807 A 3/1998 Shiloh et al 530/350
5,756,288 A 5/1998 Shiloh 435/6
5,777,093 A 7/1998 Shiloh et al 536/235
FOREIGN PATENT DOCUMENTS
GB 2327498 1/1999
WO 97/18323 5/1997
WO 98/55602 12/1998
WO 98/56391 12/1998
Salles-Passadore et al., Life Sciences, 322:113-120, 1999.
Canman et al., Science 281:1677, 1998.
Cliby et 2l\.,emboj, 17:159, 1998.
The present invention relates to identification of the consensus sequence phosphorylated by ATM kinase. This, in turn, permitted identification of ATM kinase target proteins, and development of a convenient assay system for ATM kinase phosphorylation using fusion polypeptides as substrates. The assay system is adaptable to screening for ATM modulators, particularly inhibitors. In a specific embodiment, the substrate recognition sequence and mutagenized variants of this sequence were incorporated in a GST fusion protein and assayed for phosphorylation by ATM kinase. This assay system is useful in screening for ATM inhibitors. ATM function assays were validated using an ATM-kinase dead dominant-negative mutant.
2 Claims, 4 Drawing Sheets