Adipocyte-specific constructs and methods for inhibiting platelet-type 12 lipoxygenase expression

Abstract

The present invention is directed to constructs, compositions and methods for modulating platelet-type 12 lipoxygenase (12-LO) in adipose tissue in vivo. Specifically, the invention provides constructs encoding expression-inhibiting oligonucleic acids, e.g. antisense and RNA interfering (RNAi) molecules, targeted to a platelet-type 12-LO gene or a transcript thereof, which are capable of reducing or silencing platelet-type 12-LO expression specifically in adipocytes and pre-adipocytes. Vectors comprising these constructs (including, but not limited to, viral vectors), compositions comprising them and methods of using same for the treatment and amelioration of conditions associated with excess fat cell mass and obesity are also provided.

1 .- 19 . (canceled) 
     
     
         20 . A method for reducing obesity in a subject in need thereof, comprising administering to the subject an effective amount of a recombinant nucleic acid construct, the construct comprising at least one adipose-specific transcription regulating sequence operably linked to at least one nucleic acid sequence, wherein the at least one nucleic acid sequence encodes an oligonucleic acid comprising at least one sequence substantially complementary to at least a part of the platelet-type lipoxygenase (platelet-type 12-LO) gene or transcript thereof, and wherein the oligonucleic acid inhibits or reduces the expression of platelet-type 12-LO. 
     
     
         21 . The method of  claim 20 , further comprising preventing or ameliorating an obesity-associated condition or a symptom associated therewith in said subject. 
     
     
         22 . The method of  claim 20 , wherein the obesity-associated condition is selected from the group consisting of: cardiovascular diseases, type 2 diabetes, hypertension, cancer, osteoarthritis and stroke. 
     
     
         23 . The method of  claim 20 , wherein said oligonucleic acid is selected from the group consisting of: an antisense molecule, a RNA interference (RNAi) molecule and an enzymatic nucleic acid molecule. 
     
     
         24 . The method of  claim 23 , wherein the oligonucleic acid is an antisense molecule having a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 1, 2, and 3, and analogs and fragments thereof comprising at least one sequence fully complementary to a target sequence of about 20 to about 30 nucleotides of the platelet-type 12-LO transcript. 
     
     
         25 . The method of  claim 20 , wherein the at least one adipose-specific transcription regulating sequence is selected from: human leptin promoter, human leptin enhancer, human adiponectin promoter, human adiponectin enhancer, and combinations thereof. 
     
     
         26 . The method according to  claim 20 , wherein said construct expresses said oligonucleic acid in large cell adipocytes. 
     
     
         27 . The method of  claim 20 , wherein the construct is administered locally to specific fat depots or wherein the construct is administered systemically. 
     
     
         28 . A method for reducing fat cell mass in a subject in need thereof, comprising administering to the subject an effective amount of a recombinant nucleic acid construct, the construct comprising at least one adipose-specific transcription regulating sequence operably linked to at least one nucleic acid sequence, wherein the at least one nucleic acid sequence encodes an oligonucleic acid comprising at least one sequence substantially complementary to at least a part of the platelet-type 12-LO gene or transcript thereof, and wherein the oligonucleic acid inhibits or reduces the expression of platelet-type 12-LO. 
     
     
         29 . The method of  claim 28 , wherein the oligonucleic acid is an antisense molecule having a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 1, 2, and 3, and analogs and fragments thereof comprising at least one sequence fully complementary to a target sequence of about 20 to about 30 nucleotides of the platelet-type 12-LO transcript, and the at least one adipose-specific transcription regulating sequence is selected from: human leptin promoter, human leptin enhancer, human adiponectin promoter, human adiponectin enhancer, and combinations thereof. 
     
     
         30 . The method according to  claim 28 , wherein said construct expresses said oligonucleic acid in large cell adipocytes. 
     
     
         31 . The method of  claim 28 , wherein the construct is administered locally to specific fat depots or wherein the construct is administered systemically. 
     
     
         32 . A method for inducing apoptosis specifically in a cell selected from the group consisting of adipocytes and pre-adipocytes, or for inhibiting or reducing the expression of platelet-type 12-LO in a cell selected from adipocytes and pre-adipocytes, comprising contacting the cell with a recombinant nucleic acid construct, the construct comprising at least one adipose-specific transcription regulating sequence operably linked to at least one nucleic acid sequence, wherein the at least one nucleic acid sequence encodes an oligonucleic acid comprising at least one sequence substantially complementary to at least a part of the platelet-type 12-LO gene or transcript thereof, and wherein the oligonucleic acid inhibits or reduces the expression of platelet-type 12-LO. 
     
     
         33 . The method of  claim 32  wherein the oligonucleic acid is an antisense molecule having a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 1, 2, and 3, and analogs and fragments thereof comprising at least one sequence fully complementary to a target sequence of about 20 to about 30 nucleotides of the platelet-type 12-LO transcript, and the at least one adipose-specific transcription regulating sequence is selected from: human leptin promoter, human leptin enhancer, human adiponectin promoter, human adiponectin enhancer, and combinations thereof. 
     
     
         34 . The method according to  claim 32 , wherein said construct expresses said oligonucleic acid in large cell adipocytes. 
     
     
         35 . The method according to  claim 32 , wherein the construct is administered to the subject in the form of a pharmaceutical composition further comprising a pharmaceutically acceptable carrier, excipient or diluent. 
     
     
         36 . The method according to  claim 35 , wherein the construct is administered locally to specific fat depots. 
     
     
         37 . The method according to  claim 35 , wherein the construct is administered systemically.