Inhibitors of human EZH2, and methods of use thereof

• US 9,334,527 B2
• Filed: 05/29/2014
• Issued: 05/10/2016
• Est. Priority Date: 09/10/2010
• Status: Active Grant

• Current Assignee: Epizyme, Inc. (Ipsen Sa)
• Original Assignee: Epizyme, Inc. (Ipsen Sa)
• Inventors: Kuntz, Kevin Wayne, Knutson, Sarah Kathleen, Wigle, Timothy James Nelson
• Primary Examiner(s): Leith, Nancy J
• Application Number:
  US14/290,642
• Publication Number:
  US 20140275081A1
• Time in Patent Office: 712 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: A61K 31/4412 having oxo groups directly attached to the heterocyclic ring
• CPC Class Codes: A61K 31/4427 containing further heterocyclic ring systems
• CPC Class Codes: A61K 31/4439 containing a five-membered ring with nitrogen as a ring hetero atom, e.g. omeprazole nicotine A61K31/465
• CPC Class Codes: A61K 31/444 containing a six-membered ring with nitrogen as a ring heteroatom, e.g. amrinone
• CPC Class Codes: A61K 31/4545 containing a six-membered ring with nitrogen as a ring hetero atom, e.g. pipamperone, anabasine
• CPC Class Codes: A61K 31/496 Non-condensed piperazines containing further heterocyclic rings, e.g. rifampin, thiothixene or sparfloxacin
• CPC Class Codes: A61K 31/497 containing further heterocyclic rings
• CPC Class Codes: A61K 31/5377 not condensed and containing further heterocyclic rings, e.g. timolol
• CPC Class Codes: A61K 31/551 having two nitrogen atoms, e.g. dilazep
• CPC Class Codes: A61K 31/7076 containing purines, e.g. adenosine, adenylic acid
• CPC Class Codes: A61K 31/711 Natural deoxyribonucleic acids, i.e. containing only 2'-deoxyriboses attached to adenine, guanine, cytosine or thymine and having 3'-5' phosphodiester links
• CPC Class Codes: A61K 38/17 from animals; from humans enzyme inhibitors A61K38/005
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 35/02 specific for leukemia
• CPC Class Codes: C07D 405/12 linked by a chain containing hetero atoms as chain links
• CPC Class Codes: C07D 473/34 attached in position 6, e.g. adenine
• CPC Class Codes: C12Q 1/48 involving transferase
• CPC Class Codes: C12Q 1/68 involving nucleic acids
• CPC Class Codes: G01N 2333/91011 Methyltransferases (general) (2.1.1.)
• CPC Class Codes: G01N 2333/91017 with definite EC number (2.1.1.-)
• CPC Class Codes: G01N 2800/52 : Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis
• CPC Class Codes: G01N 33/5011 : for testing antineoplastic activity
• CPC Class Codes: G01N 33/57426 : leukemia
• CPC Class Codes: Y10T 436/143333 : Saccharide [e.g., DNA, etc.]

Abstract

The invention relates to inhibition of wild-type and certain mutant forms of human histone methyltransferase EZH2, the catalytic subunit of the PRC2 complex which catalyzes the mono- through tri-methylation of lysine 27 on histone H3 (H3-K27). In one embodiment the inhibition is selective for the mutant form of the EZH2, such that trimethylation of H3-K27, which is associated with certain cancers, is inhibited. The methods can be used to treat cancers including follicular lymphoma and diffuse large B-cell lymphoma (DLBCL). Also provided are methods for identifying small molecule selective inhibitors of the mutant forms of EZH2 and also methods for determining responsiveness to an EZH2 inhibitor in a subject.

Claims

1. A method of treating cancer in a subject in need thereof comprising

detecting the presence of a mutation in the EZH2 substrate pocket domain of SEQ ID NO:

7, in a sample from the subject, wherein said detecting comprises screening, at amino acid positions Y641, A677, and A687 of EZH2 of SEQ ID NO;

1, for the presence of at least one mutation, wherein the mutation at amino acid position Y641 is selected from the group consisting of a substitution of phenylalanine (F) for the wild type residue tyrosine (Y) at amino acid position 641 of SEQ ID NO;

1 (Y641F);

a substitution of histidine (H) for the wild type residue tyrosine (Y) at amino acid position 641 of SEQ ID NO;

1 (Y641H);

a substitution of asparagine (N) for the wild type residue tyrosine (Y) at amino acid position 641 of SEQ ID NO;

1 (Y641N);

a substitution of serine (S) for the wild type residue tyrosine (Y) at amino acid position 641 of SEQ ID NO;

1 (Y641S); and

a substitution of cysteine (C) for the wild type residue tyrosine (Y) at amino acid position 641 of SEQ ID NO;

1 (Y641C), wherein the mutation at amino acid position A677 is a substitution of glycine (G) for the wild type residue alanine (A) at amino acid position 677 of SEQ ID NO;

1 (A677G), wherein the mutation at amino acid position A687 is a substitution of valine (V) for the wild type residue alanine (A) at amino acid position 687 of SEQ ID NO;

1 (A687V), and wherein the mutation increases EZH2 trimethylation of Lys27 of histone H3 (H3-K27); and

identifying the subject as a candidate for treatment with an EZH2 inhibitor based on the presence of a detected EZH2 mutation;

administering a therapeutically effective amount of an EZH2 inhibitor to the identified subject, wherein the EZH2 inhibitor inhibits the conversion of H3-K27 to trimethylated H3-K27.

2. The method of claim 1, wherein the nucleic acid encoding SEQ ID NO:

7 is SEQ ID NO;

8.

3. The method of claim 1, wherein the EZH2 inhibitor is a small molecule.

4. The method of claim 1, wherein the cancer is lymphoma, selected from the group consisting of non-Hodgkin lymphoma, follicular lymphoma, diffuse large B-cell lymphoma and diffuse large B-cell lymphoma of germinal center B cell-like subtype.

5. The method of claim 1, wherein the EZH2 inhibitor is selected from the group consisting of

6. The method of claim 1 wherein the EZH2 inhibitor is

7. The method of claim 6, wherein the cancer is lymphoma.

8. The method of claim 7, wherein the lymphoma is non-Hodgkin lymphoma.

9. The method of claim 7, wherein the lymphoma is follicular lymphoma.

10. The method of claim 7, wherein the lymphoma is diffuse large B-cell lymphoma.

11. The method of claim 7, wherein the lymphoma is diffuse large B-cell lymphoma of germinal center B cell-like subtype.

12. The method of claim 2, wherein the EZH2 inhibitor) is

13. The method of claim 4, wherein the EZH2 inhibitor is