Production and utilization of a novel anti-cancer drug in therapy

• US 9,422,559 B2
• Filed: 08/12/2013
• Issued: 08/23/2016
• Est. Priority Date: 06/02/2010
• Status: Active Grant

• Current Assignee: David Ts Wu, Shi-Lung Lin
• Original Assignee: David Ts Wu, Shi-Lung Lin
• Inventors: Lin, Shi-Lung, Wu, David T S
• Primary Examiner(s): Schultz, Doug
• Application Number:
  US13/964,705
• Publication Number:
  US 20130324590A1
• Time in Patent Office: 1,107 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• 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 15/1137 against enzymes viral enzymes C12N15/1131; receptors C12N15/1138
• CPC Class Codes: C12N 2310/141 MicroRNAs, miRNAs
• CPC Class Codes: C12N 2330/50 Biochemical production, i.e. in a transformed host cell
• CPC Class Codes: C12N 2330/51 Specially adapted vectors

Abstract

This invention generally relates to a design and method for developing novel anti-tumor and/or anti-cancer drugs, vaccines and therapies, using microRNA and/or its shRNA homologs/mimics/derivatives. More specifically, the present invention relates to an use of a prokaryote-produced miRNA precursor (pro-miRNA) composition capable of being delivered into human cells and processed by the cells into mature miRNA effectors to elicit specific silencing effects on mir-302-targeted genes, subsequently leading to a beneficial result of tumor suppression and cancer therapy. The prokaryotic cells do not naturally express or process eukaryotic miRNA precursors (pre-miRNA); meanwhile, the present invention also teaches an inducible method for expressing pre-miRNAs, particularly mir-302 precursors by using the prokaryotic transcription system. Since mir-302 is a known tumor suppressor in human, this novel finding advances the design and method for developing new anti-cancer drugs, vaccines and/or therapies directed against multiple kinds of human tumors and cancers.

Claims

1. A method for inducing a gene silencing effect on BMI-1 and then leading to an increase of p16Ink4a or p14Arf tumor suppressor expression in vivo, comprising:

delivering a pro-mir-302 capable of being processed into at least one hairpin-like microRNA precursor in a human cell substrate to an amount, wherein said at least one hairpin-like microRNA precursor contains a seed sequence SEQ ID NO;

3 and said human cell substrate contains at least a cancerous cell type of human liver cancer, and wherein the amount of said hairpin-like microRNA precursor is higher than 200 micrograms per delivery in vivo.

2. The method of claim 1, wherein said step of delivering said pro-mir-302 further comprises providing said pro-mir-302 from prokaryotic competent cells.

3. The method of claim 2, wherein said pro-mir-302 is produced by said prokaryotic competent cells and is a precursor of mir-302.

4. The method of claim 2, wherein said step of providing said pro-mir-302 further comprises isolating said pro-mir-302 from an extract or a lysate of said prokaryotic competent cells.

5. The method of claim 1, wherein said step of delivering said pro-mir-302 capable of being processed into at least one hairpin-like microRNA precursor further comprises letting said pro-mir-302 be processed into mir-302a, mir-302b, mir-302c, and/or mir-302d in said human cell substrate.

6. The method of claim 1, wherein said gene silencing effect further comprises an effect leading to reprogram high-grade malignant cancers to a relatively benign low-grade state.

7. The method of claim 1, wherein said pro-mir-302 is delivered into said human cell substrate by a gene delivery method.

8. The method of claim 1, wherein said pro-mir-302 is used as a drug for treating human liver cancer.

9. The method of claim 1, wherein said pro-mir-302 is useful for pharmaceutical and/or therapeutic applications.