US2020239892A1PendingUtilityA1

Multi-conjugate of sirna and preparing method thereof

69
Assignee: KIP CO LTDPriority: Mar 13, 2009Filed: Feb 4, 2020Published: Jul 30, 2020
Est. expiryMar 13, 2029(~2.7 yrs left)· nominal 20-yr term from priority
Inventors:Tae Gwan ParkHye Jung MokSoo Hyeon Lee
C12N 15/10A61P 35/00C07H 21/02C12N 15/11C12N 15/111C12N 2310/51C12N 15/113C12N 15/1136C12N 2310/14
69
PatentIndex Score
0
Cited by
0
References
0
Claims

Abstract

The present invention relates to a multi-conjugate of small interfering RNA (siRNA) and a preparing method of the same, more precisely a multi-conjugate of siRNA prepared by direct binding of double stranded sense/antisense siRNA monomers or indirect covalent bonding mediated by a cross-linking agent or a polymer, and a preparing method of the same. The preparing method of a siRNA multi-conjugate of the present invention is characterized by simple and efficient reaction and thereby the prepared siRNA multi-conjugate of the present invention has high molecular weight multiple times the conventional siRNA, so that it has high negative charge density, suggesting that it has excellent ionic interaction with a cationic gene carrier and high gene delivery efficiency.

Claims

exact text as granted — not AI-modified
1 . A multi-conjugate of siRNA having the following structure of [Structure Formula I] or [Structure Formula II] prepared by direct or indirect covalent bonding of double-stranded sense/antisense siRNA monomers mediated by a cross-linking agent or a polymer:
   (—X-A-) n   [Structural Formula I]
   (Wherein,   X is double-stranded siRNA monomer;   A is either presented or not presented, a cross-linking agent or a polymer; and   n is the number of double-stranded siRNA monomer),
   x-A-(X-A-) n -x′  [Structural Formula II]
 
   (Wherein,   X is double-stranded siRNA monomer;   x or x′ is single-stranded siRNA monomer   A is either presented or not presented, a cross-linking agent or a polymer; and   n is the number of double-stranded siRNA monomer).   
     
     
         2 . The siRNA multi-conjugate according to  claim 1 , wherein the double-stranded sense/antisense siRNA monomer is composed of 15-29 nucleotides. 
     
     
         3 . The siRNA multi-conjugate according to  claim 1 , wherein the number n indicating the number of the double-stranded sense/antisense siRNA monomer is one of 1-100. 
     
     
         4 . A preparing method of a multi-conjugate of siRNA having the structure of [Structural Formula I] or [Structural Formula II] containing the step of direct or indirect covalent bonding of double-stranded sense/antisense siRNA monomers mediated by a cross-linking agent or a polymer
   (—X-A-) n   [Structural Formula I]
   (Wherein,   X is double-stranded siRNA monomer;   A is either presented or not presented, a cross-linking agent or a polymer; and   n is the number of double-stranded siRNA monomer),
   x-A-(X-A-) n -x′  [Structural Formula II]
 
   (Wherein,   X is double-stranded siRNA monomer;   x or x′ is single-stranded siRNA monomer   A is either presented or not presented, a cross-linking agent or a polymer; and   n is the number of double-stranded siRNA monomer).   
     
     
         5 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein the method is composed of the following steps:
 1) preparing yXy by complementary hydrogen bonding of single-stranded sense/antisense siRNA monomers (yx+x′y) having substitutions with same functional groups at the ends; and   2) inducing covalent bond of the yXy mediated by a cross-linking agent or a polymer:   (Herein,   X is double-stranded siRNA monomer;   x and x′ are single-stranded sense/antisense siRNA monomers; and   y is the functional group introduced in the end).   
     
     
         6 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein the method is composed of the following steps:
 1) preparing dimers of xyyx and x′yyx′ by covalent bonding of single-stranded sense/antisense siRNA monomers (yx and x′y) having substitutions with same functional groups at the ends; and   2) inducing complementary hydrogen bond between the dimers xyyx and x′yyx′:   (Herein,   X is double-stranded siRNA monomer;   x and x′ are single-stranded sense/antisense siRNA monomers; and   y is the functional group introduced in the end).   
     
     
         7 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein the method is composed of the following steps:
 1) preparing yXz by complementary hydrogen bonding of single-stranded sense/antisense siRNA monomers (yx+x′z) having substitutions with different functional groups at the ends; and   2) inducing covalent bond of the yXz mediated by a cross-linking agent or a polymer:   (Herein,   X is double-stranded siRNA monomer;   x and x′ are single-stranded sense/antisense siRNA monomers; and   y and z are the functional groups introduced in the ends).   
     
     
         8 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein the method is composed of the following steps:
 1) preparing dimers of xyzx and x′yzx′ by covalent bonding of single-stranded sense/antisense siRNA monomers (yx and xz, x′y and x′z) having substitutions with different functional groups at the ends; and   2) inducing complementary hydrogen bond between the dimers xyzx and x′yzx′:   (Herein,   X is double-stranded siRNA monomer;   x and x′ are single-stranded sense/antisense siRNA monomers; and   y and z are the functional groups introduced in the ends).   
     
     
         9 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein the covalent bond is selected from the group consisting of non-cleavable bonds (amide bond and urethane bond), acid cleavable bonds (ester bond, hydrazone bond and acethal bond), reductant cleavable bond (disulfide bond), bio-cleavable bonds and enzyme cleavable bonds. 
     
     
         10 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein the single stranded sense/antisense siRNA monomer has the substitution with a functional group at 3′end or at 5′end. 
     
     
         11 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein the polymer is one or more non-ionic hydrophilic polymers selected from the group consisting of PEG, Pluronic, polyvinylpyrolidone and polyoxazolin; or one or more biocleavable polyester polymers selected from the group consisting of poly-L-lactic acid, poly-glycolic acid, poly-D-lactic-co-glycolic acid, poly-L-lactic-co-glycolic acid, poly-D,L-lactic-co-glycolic acid, polycaprolactone, polyvalerolactone, polyhydroxybutyrate and polyhydroxyvalerate. 
     
     
         12 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein the cross-linking agent has the molecular weight of 1.00-10000 and is one or more compounds selected from the group consisting of DTME (Dithio-bis-maleimidoethane), BM(PEG) 2  (1,8-Bis-maleimidodiethyleneglycol), maleimide, NHS (N-hydroxysuccinimide), vinylsulfone, iodoacetyl, nitrophenyl azide, isocyanate, pyridyldisulfide, hydrazide and hydroxyphenyl azide. 
     
     
         13 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein a cell selective ligand is additionally added to the end of the siRNA multi-conjugate. 
     
     
         14 . The preparing method of a siRNA multi-conjugate according to  claim 13 , wherein the ligand is one or more compounds selected from the group consisting of cell specific antibody, cell selective peptide, cell growth factor, folic acid, galactose, mannose, RGD, and transferrin. 
     
     
         15 . The preparing method of a siRNA multi-conjugate according to  claim 4 , wherein the step of activating the said functional group is additionally included. 
     
     
         16 . An ionic complex formed by ionic interaction between the siRNA multi-conjugate of  claim 1 ; and one or more cationic gene carriers selected from the group consisting of cationic peptides, cationic lipids and cationic polymers. 
     
     
         17 . A method for treating cancer containing the step of administering the ionic complex of  claim 16  to a subject. 
     
     
         18 . A method for treating angiogenesis related disease containing the step of administering the ionic complex of  claim 16  to a subject. 
     
     
         19 . A use of the ionic complex of  claim 16  for the production of an anticancer agent. 
     
     
         20 . A use of the ionic complex of  claim 16  for the production of a therapeutic agent for angiogenesis related disease.

Cited by (0)

No later patents cite this yet.

References (0)

No backward citations on record.