Treatment of PAR4 related diseases by inhibition of natural antisense transcript to PAR4

• US 10,100,315 B2
• Filed: 07/25/2017
• Issued: 10/16/2018
• Est. Priority Date: 05/14/2010
• Status: Active Grant

• Current Assignee: CURNA, Inc. (Opko Health, Inc.)
• Original Assignee: CURNA, Inc. (Opko Health, Inc.)
• Inventors: Collard, Joseph, Khorkova Sherman, Olga
• Primary Examiner(s): Vivlemore, Tracy
• Application Number:
  US15/659,122
• Publication Number:
  US 20170327828A1
• Time in Patent Office: 448 Days
• Field of Search: None
• CPC Class Codes: A61K 31/7088 Compounds having three or more nucleosides or nucleotides
• 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 35/00 Antineoplastic agents
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C12N 15/113 Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing when used in plants C12N15/8218
• CPC Class Codes: C12N 15/1138 against receptors or cell surface proteins
• CPC Class Codes: C12N 2310/111 spanning the whole gene, or a large part of it
• CPC Class Codes: C12N 2310/113 targeting other non-coding nucleic acids, e.g. antagomirs
• CPC Class Codes: C12N 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2310/321 2'-O-R Modification
• CPC Class Codes: C12N 2310/322 2'-R Modification
• CPC Class Codes: C12N 2310/323 modified ring structure
• CPC Class Codes: C12N 2310/3231 having an additional ring, e.g. LNA, ENA
• CPC Class Codes: C12N 2330/10 naturally occurring
• CPC Class Codes: C12Q 1/6811 Selection methods for production or design of target specific oligonucleotides or binding molecules
• CPC Class Codes: Y02A 50/30 Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

The present invention relates to antisense oligonucleotides that modulate the expression of and/or function of PAR4, in particular, by targeting natural antisense polynucleotides of PAR4. The invention also relates to the identification of these antisense oligonucleotides and their use in treating diseases and disorders associated with the expression of PAR4.

Claims

1. A method of identifying and selecting at least one active antisense oligonucleotide of 10 to 30 nucleotides in length which specifically hybridizes to a natural antisense polynucleotide of a PAR4 gene and modulates expression of said PAR4 gene comprising:

sequencing at least one natural antisense polynucleotide of the target PAR4 gene as a putative target wherein the PAR4 natural antisense polynucleotide comprises SEQ ID NO;

2;

identifying at least one antisense oligonucleotide which is complementary to the natural antisense polynucleotide target;

measuring the thermal melting point of a hybrid of said at least one antisense oligonucleotide and the target natural antisense polynucleotide under stringent hybridization conditions;

and measuring the modulatory activity of said at least one oligonucleotide and selecting at least one active antisense oligonucleotide.

2. The method according to claim 1 wherein the at least one antisense oligonucleotide is a siRNA.

3. The method according to claim 1 wherein the at least one antisense oligonucleotide is a single stranded, modified antisense oligonucleotide.

4. The method according to claim 3 wherein the modifications are selected from the group consisting of at least one modified sugar moiety;

at least one modified internucleotide linkage;

at least one modified nucleotide, and combinations thereof.

5. The method according to claim 4 wherein the at least one modified sugar moiety is selected from the group consisting of a 2′

-O-methoxyethyl modified sugar moiety, a 2′

-methoxy modified sugar moiety, a 2′

-O-alkyl modified sugar moiety, a bicyclic sugar moiety, and a combination thereof.

6. The method according to claim 4 wherein the at least one modified nucleotide is selected from the group consisting of peptide nucleic acids, locked nucleic acids (LNA), analogues, derivatives, and a combination thereof.

7. The method according to claim 4 wherein the least one modified internuceotide linkage is selected from the group consisting of phosphorothioate, alkylphosphonate, phosphorodithioate, alkylphosphonothioate, phosphoramidate, carbamate, carbonate, phosphate triester, acetamidate, carboxymethyl ester, and a combination thereof.

8. The method according to claim 1 wherein the antisense oligonucleotides are selected from the group consisting of SEQ ID NOS:

3 to 9.