RNAi-Mediated Inhibition of Ocular Targets

Abstract

RNA interference is provided for inhibition of ocular hypertension target mRNA expression for lowering elevated intraocular pressure in patients with open-angle glaucoma or ocular hypertension. Ocular hypertension targets include carbonic anhydrase II, IV, and XII; β1- and β2 adrenergic receptors; acetylcholinesterase; Na + /K + -ATPase; and Na—K—2Cl cotransporter. Ocular hypertension is treated by administering interfering RNAs of the present invention.

1 . A method of attenuating expression of an ocular hypertension target mRNA of a subject, comprising:
 administering to the subject a composition comprising an effective amount of interfering RNA having a length of 19 to 49 nucleotides and a pharmaceutically acceptable carrier, the interfering RNA comprising:
 a sense nucleotide strand, an antisense nucleotide strand, and a region of at least near-perfect contiguous complementarity of at least 19 nucleotides; 
 wherein the antisense strand hybridizes under physiological conditions to a portion of mRNA corresponding to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:101, or SEQ ID NO:134, and has a region of at least near-perfect contiguous complementarity of at least 19 nucleotides with the hybridizing portion of mRNA corresponding to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:101, or SEQ ID NO:134, respectively, 
   
       wherein the expression of an ocular hypertension target mRNA is attenuated. 
     
     
         2 . The method of  claim 1  wherein the subject is a human and the human has ocular hypertension. 
     
     
         3 . The method of  claim 1  wherein the subject is a human and the human is at risk of developing ocular hypertension. 
     
     
         4 . The method of  claim 1  wherein the ocular hypertension target mRNA encodes carbonic anhydrase II, and the antisense strand hybridizes under physiological conditions to a portion of mRNA corresponding to SEQ ID NO:1 and has a region of at least near-perfect contiguous complementarity of at least 19 nucleotides with the hybridizing portion of mRNA corresponding to SEQ ID NO:1. 
     
     
         5 . The method of  claim 1  wherein the ocular hypertension target mRNA encodes carbonic anhydrase IV, and the antisense strand hybridizes under physiological conditions to a portion of mRNA corresponding to SEQ ID NO:2 and has a region of at least near-perfect contiguous complementarity of at least 19 nucleotides with the hybridizing portion of mRNA corresponding to SEQ ID NO:2. 
     
     
         6 . The method of  claim 1  wherein the ocular hypertension target mRNA encodes carbonic anhydrase XII, and the antisense strand hybridizes under physiological conditions to a portion of mRNA corresponding to SEQ ID NO:101 or SEQ ID NO:134 and has a region of at least near-perfect contiguous complementarity of at least 19 nucleotides with the hybridizing portion of mRNA corresponding to SEQ ID NO:101 or SEQ ID NO:134, respectively. 
     
     
         7 . The method of  claim 1  wherein the antisense strand is designed to target an mRNA corresponding to SEQ ID NO:1 comprising nucleotide 232, 527, 721, 728, 809, 810, 855, 856, 921, 1139, 506, 547, 548, 740, 911, 1009, 1140, 1149, 1150, 1151, 1188, 1194, 1195, 1223, 1239, 1456, 1457, 1458, 100, 158, 166, 247, 286, 318, 322, 328, 371, 412, 482, 504, 505, 541, 734, 772, 777, 814, 972, 998, 1232, 317, or 401. 
     
     
         8 . The method of  claim 1  wherein the antisense strand is designed to target an mRNA corresponding to SEQ ID NO:2 comprising nucleotide 213, 252, 258, 266, 399, 457, 463, 490, 595, 1064, 109, 112, 125, 126, 150, 261, 265, 280, 398, 453, 459, 462, 467, 492, 534, 785, 801, 825, 827, 876, 1003, or 1012. 
     
     
         9 . The method of  claim 1  wherein the antisense strand is designed to target an mRNA corresponding to SEQ ID NO:101 comprising nucleotide 191, 239, 274, 275, 341, 389, 412, 413, 423, 687, 689, 695, 710, 791, 792, 794, 983, 993, 994, 995, 691, 1039, 1568, 2326, 2332, 2425, 2433, 2844, 2845, 2880, 2884, 2891, 2954, 2955, 2956, 2957, 2964, 2965, 3006, 3007, 3012, or 3026. 
     
     
         10 . The method of  claim 1  wherein the antisense strand is designed to target an mRNA corresponding to SEQ ID NO:134 comprising nucleotide 687, 1535, 2293, 2299, 2392, 2400, 2811, 2812, 2847, 2851, 2858, 2921, 2922, 2923, 2924, 2931, 2932, 2973, 2974, 2979, or 2993. 
     
     
         11 . The method of  claim 1  further comprising administering to the subject a second interfering RNA having a length of 19 to 49 nucleotides, and comprising
 a sense nucleotide strand, an antisense nucleotide strand, and a region of at least near-perfect complementarity of at least 19 nucleotides;   wherein the antisense strand of the second interfering RNA hybridizes under physiological conditions to a second portion of mRNA corresponding to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:101, or SEQ ID NO:134, and the antisense strand has a region of at least near-perfect contiguous complementarity of at least 19 nucleotides with the second hybridizing portion of mRNA corresponding to SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:101, or SEQ ID NO:134, respectively.   
     
     
         12 . The method of  claim 1  wherein the sense nucleotide strand and the antisense nucleotide strand are connected by a loop nucleotide sequence. 
     
     
         13 . The method of  claim 1  wherein the composition is administered via a topical, intravitreal, transcleral, periocular, conjunctival, subtenon, intracameral, subretinal, subconjunctival, retrobulbar, or intracanalicular route. 
     
     
         14 . The method of  claim 1  wherein the composition is administered via in vivo expression from an expression vector capable of expressing the interfering RNA. 
     
     
         15 . A method of attenuating expression of an ocular hypertension target mRNA of a subject, comprising:
 administering to the subject a composition comprising an effective amount of single-stranded interfering RNA having a length of 19 to 49 nucleotides and a pharmaceutically acceptable carrier,
 wherein the single-stranded interfering RNA hybridizes under physiological conditions to a portion of mRNA corresponding to SEQ ID NO:1 comprising nucleotide 232, 527, 721, 728, 809, 810, 855, 856, 921, 1139, 506, 547, 548, 740, 911, 1009, 1140, 1149, 1150, 1151, 1188, 1194, 1195, 1223, 1239, 1456, 1457, 1458, 100, 158, 166, 247, 286, 322, 328, 371, 412, 482, 504, 505, 541, 734, 772, 777, 814, 972, 998, 1232, or 401, and the interfering RNA has a region of at least near-perfect contiguous complementarity with the hybridizing portion of mRNA corresponding to SEQ ID NO:1; 
 or 
 wherein the single-stranded interfering RNA hybridizes under physiological conditions to a portion of mRNA corresponding to SEQ ID NO:2 comprising nucleotide 213, 252, 258, 266, 399, 457, 463, 490, 595, 1064, 109, 112, 125, 126, 150, 261, 265, 280, 398, 453, 459, 462, 467, 492, 534, 785, 801, 825, 827, 876, 1003, or 1012, and the interfering RNA has a region of at least near-perfect contiguous complementarity of at least 19 nucleotides with the hybridizing portion of mRNA corresponding to SEQ ID NO:2; 
 or 
 wherein the single-stranded interfering RNA hybridizes under physiological conditions to a portion of mRNA corresponding to SEQ ID NO:101 comprising nucleotide 191, 239, 274, 275, 341, 389, 412, 413, 423, 687, 689, 695, 710, 791, 792, 794, 983, 993, 994, 995, 691, 1039, 1568, 2326, 2332, 2425, 2433, 2844, 2845, 2880, 2884, 2891, 2954, 2955, 2956, 2957, 2964, 2965, 3006, 3007, 3012, or 3026, and the interfering RNA has a region of at least near-perfect contiguous complementarity of at least 19 nucleotides with the hybridizing portion of mRNA corresponding to SEQ ID NO:101; 
 or 
 wherein the single-stranded interfering RNA hybridizes under physiological conditions to a portion of mRNA corresponding to SEQ ID NO:134 comprising nucleotide 687, 1535, 2293, 2299, 2392, 2400, 2811, 2812, 2847, 2851, 2858, 2921, 2922, 2923, 2924, 2931, 2932, 2973, 2974, 2979, or 2993, and the interfering RNA has a region of at least near-perfect contiguous complementarity of at least 19 nucleotides with the hybridizing portion of mRNA corresponding to SEQ ID NO:134; 
   
       wherein the expression of an ocular hypertension target mRNA is thereby attenuated. 
     
     
         16 . The method of  claim 15  wherein the composition is administered via a topical, intravitreal, transcleral, periocular, conjunctival, subtenon, intracameral, subretinal, subconjunctival, retrobulbar, or intracanalicular route. 
     
     
         17 . The method of  claim 15  wherein the composition is administered via in vivo expression from an expression vector capable of expressing the interfering RNA. 
     
     
         18 . A composition comprising interfering RNA having a length of 19 to 49 nucleotides and having a nucleotide sequence of any one of SEQ ID NO:8, SEQ ID NO:14-SEQ ID NO:32, SEQ ID NO:83-SEQ ID NO:100, SEQ ID NO:102-SEQ ID NO:122, SEQ ID NO:135-SEQ ID NO:139, SEQ ID NO:141-SEQ ID NO:219, and SEQ ID NO:721, or a complement thereof, and a pharmaceutically acceptable carrier. 
     
     
         19 . The composition of  claim 18  wherein the interfering RNA is an shRNA, siRNA, or miRNA.