AAV VARIANTS FOR THE TREATMENT OF DRY AMD

1 . A polynucleotide cassette for increasing the concentration of human CFI in the vitreous humor and/or retinal extracellular space of a mammalian eye, comprising:

a. an enhancer and promoter region;

b. a chimeric intron;

c. a Kozak sequence

d. a unique coding sequence optimized for increased expression and low CpG, operatively linked to the promoter region wherein the coding sequence is a human CFI gene,

e. a human scaffold attachment region; and

f. a polyadenylation site.

2 . The polynucleotide cassette of claim 1 , wherein the promoter region further comprises an enhancer element.

3 . The polynucleotide cassette of claim 1 , wherein the chimeric intron comprises at least one element selected from the group comprising: (a) an adenovirus tripartite leader sequence (TPL), (b) an enhancer element, and (c) an intron from mouse IgH.

4 . The polynucleotide cassette of claim 1 , further comprising AAV2 inverted terminal repeats (ITRs).

5 . The polynucleotide cassette of claim 1 , wherein the polyadenylation site is the human growth hormone polyadenylation site.

6 . The polynucleotide cassette of claim 1 , wherein the coding sequence has a nucleotide sequence having at least 90% identity to the nucleotide sequence set forth in SEQ ID NO:1.

7 . The polynucleotide cassette of claim 1 , wherein the coding sequence of the human CFI gene is selected from the group consisting of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, and SEQ ID NO:6.

8 . A method of increasing the concentration of human CFI in the vitreous humor and/or retinal extracellular space of a mammalian eye, comprising administering to a subject a recombinant adeno-associated virus (rAAV) vector at a dosage ranging from about 1E 9 to about 4E 12 vector genomes (vg)/eye, wherein the rAAV vector comprises an AAV2 capsid variant and wherein the rAAV vector comprises a polynucleotide cassette comprising:

a. An enhancer and promoter region;

b. a chimeric intron;

c. a Kozak sequence

d. a unique coding sequence optimized for increased expression and low CpG, operatively linked to the promoter region wherein the coding sequence is a human CFI gene,

e. a human scaffold attachment region; and

f. a polyadenylation site.

9 . The method of claim 8 wherein the polynucleotide cassette further comprises an enhancer element.

10 . The method of claim 8 , wherein the chimeric intron of the polynucleotide cassette comprises at least one element selected from the group comprising: (a) an adenovirus tripartite leader sequence (TPL), (b) an enhancer element, and (c) an intron from mouse IgH.

11 . The method of claim 8 , wherein the polynucleotide cassette further comprises AAV2 inverted terminal repeats (ITRs).

12 . The method of claim 8 , wherein the polyadenylation site of the polynucleotide cassette is the human growth hormone polyadenylation site.

13 . The method of claim 8 , wherein the polynucleotide cassette has a nucleotide sequence having at least 90% identity to the nucleotide sequence set forth in SEQ ID NO:1.

14 . The method of claim 8 , wherein the coding sequence of a human CFI gene is selected from the group consisting of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5 and SEQ ID NO:6.

15 . The method of claim 8 , wherein rAAV vector is administered intravitreally.

16 . The method of claim 8 , wherein the AAV2 capsid variant is selected from the group comprising AAV2.7m8 and AAV2.5T-LSV1.

17 . The method of claim 8 , wherein the rAAV vector is administered at a dosing range selected from the group of ranges consisting of from about 1E 9 to about 4E 12 vg/eye, from about 10E 10 to about 4E 11 vg/eye, from about 2E 10 to about 3E 11 vg/eye, from about 2E 10 to about 2E 11 vg/eye, from about 2.5E 10 to about 2E 11 vg/eye, from about 2E 10 to about 1E 11 vg/eye, from about 2E 10 to about 9E 10 vg/eye, from about 2E 10 to about 8E 10 vg/eye, and from about 3E 10 to about 7E 10 vg/eye.

18 . The method of claim 8 , wherein human CFI is detectable in the vitreous humor at least 4 weeks after intravitreal injection of said rAAV vector.

19 . The method of claim 8 , wherein the level of C3b-inactivating activity in an eye of said subject is increased.

20 . The method of claim 8 , wherein the level of iC3b-degradation activity in an eye of the subject is increased.

21 . The method of claim 8 , wherein the rAAV vector is administered as an intravitreal injection.

22 . The method of claim 8 , wherein the human CFI expressed from the rAAV vector is capable of degrading C3b into iC3b.

23 . A polynucleotide cassette for increasing the concentration of human CFI in a mammalian eye, comprising:

a. an enhancer and promoter region;

b. a chimeric intron;

c. a Kozak sequence

d. a unique coding sequence optimized for increased expression and low CpG, operatively linked to the promoter region wherein the coding sequence is a human CFI gene,

e. a human scaffold attachment region; and

f. a polyadenylation site.

24 . The polynucleotide cassette of claim 24 , wherein said polynucleotide cassette is for increasing the concentration of human CFI in the vitreous humor and/or retinal extracellular space of a mammalian eye.

25 . A method of treating dry acute macular degeneration (dry-AMD) in a human subject, comprising administering to a subject in need thereof a therapeutically effective amount of recombinant adeno-associated virus (rAAV) vector, wherein the rAAV vector comprises an AAV2 capsid variant and a polynucleotide cassette comprising:

a. An enhancer and promoter region;

b. a chimeric intron;

c. a Kozak sequence

d. a unique coding sequence optimized for increased expression and low CpG, operatively linked to the promoter region wherein the coding sequence is a human CFI gene,

e. a human scaffold attachment region; and

f. a polyadenylation site.

26 . The method of claim 26 , wherein the rAAV vector is administered intravitreally.

27 . The method of claim 26 , wherein the therapeutically effective amount is a dosage ranging from about 1E 9 to about 4E 12 vector genomes (vg)/eye.

28 . The method of claim 26 , wherein there is a reduction in the geographic atrophy area and/or in the rate of growth of the geographic atrophy area after administration of the rAAV vector.

29 . The method of claim 26 , wherein the therapeutically effective amount is a dosage ranging from about 2E 10 to about 9E 10 vector genomes (vg)/eye.

30 . An intravitreal dosage form, comprising a recombinant adeno-associated virus (rAAV) vector at a dosage ranging from about 1E 9 to about 4E 12 vg/eye, wherein the rAAV vector comprises a polynucleotide cassette according to claim 1 , and wherein the rAAV vector comprises an AAV2 capsid variant.

31 . The intravitreal dosage form of claim 30 , wherein rAAV vector comprises an AAV2_7m8 capsid variant and a dosage ranging from about 9E 9 to about 9E 10 vg/eye.