USE OF GLUCOCORTICOID ANALOGS TO ENHANCE RECOMBINANT ADENO-ASSOCIATED VIRUS YIELD
The invention provides methods for the production of recombinant adeno-associated virus vectors (rAAV), comprising contacting a host cell with a solution comprising a glucocorticoid analog, such as dexamethasone. Also provided are methods for increasing the production of rAAV by a host cell, comprising contacting a host cell with a solution comprising a glucocorticoid analog, such as dexamethasone.
1 . A method for producing a recombinant adeno-associated virus vector (rAAV) comprising contacting a host cell with a solution comprising a glucocorticoid analog.
2 . A method for increasing the amount of recombinant adeno-associated virus vector (rAAV) produced by a host cell, comprising contacting the host cell with a solution comprising a glucocorticoid analog.
3 . The method of claim 1 or 2 wherein the glucocorticoid analog is selected from the group consisting of dexamethasone, hydrocortisone, prednisolone, methylprednisolone, betamethasone, cortisone, prednisone, budesonide, and triamcinolone.
4 . The method of claim 3 , wherein the glucocorticoid analog is dexamethasone.
5 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in the solution is greater than or equal to 1 μM.
6 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in the solution is greater than or equal to 0.1 μM.
7 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in the solution is greater than or equal to 0.01 μM.
8 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in the solution is between 0 and 1 μM.
9 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in the solution is between 0 and 0.1 μM.
10 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in the solution is between 0 and 0.01 μM.
11 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in the solution is between 0.01 and 1 μM.
12 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in the solution is between 0.01 and 0.1 μM.
13 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in the solution is sufficient to produce at least 1.5-fold greater quantities of secreted rAAV compared to that produced by a host cell not contacted with a solution comprising a glucocorticoid analog.
14 . The method of any one of claims 1 - 4 , wherein the final concentration of the glucocorticoid analog in solution is sufficient to produce at least 1.5-fold greater quantities of total rAAV compared to that produced by a host cell not contacted with a solution comprising a glucocorticoid analog.
15 . The method of any one of claims 1 - 14 , wherein the host cell is contacted with the solution comprising the glucocorticoid analog for at least 2 days.
16 . The method of any one of claims 1 - 15 , further comprising the steps of harvesting and purifying the rAAV.
17 . The method of any one of claims 1 - 16 , wherein the host cell is a mammalian cell.
18 . The method of claim 17 , wherein the host cell is selected from the group consisting of HeLa, HEK293, COS, A549, BHK and Vero cells.
19 . The method of claim 18 , wherein the host cell is a HeLa cell.
20 . The method of any one of claims 1 - 16 , wherein the host cell is an insect cell.
21 . The method of claim 20 wherein the host cell is selected from the group consisting of Sf9, Sf-21, Tn-368, and BTI-Tn-5B1-4 (High-Five) cells.
22 . The method of any one of claims 1 - 21 , wherein the host cell comprises a heterologous nucleotide sequence flanked by AAV inverted terminal repeats.
23 . The method of any one of claims 1 - 22 , wherein the host cell comprises rep and cap genes.
24 . The method of any one of claims 1 - 23 , wherein the host cell comprises helper virus genes.
25 . The method of any one of claims 1 - 24 , wherein the host cell comprises a heterologous nucleotide sequence flanked by AAV inverted terminal repeats, rep and cap genes, and helper virus genes.
26 . The method of any one of claims 1 - 25 , wherein the host cell produces at least 1.5-fold greater quantities of secreted rAAV compared to that produced by a host cell not contacted with a solution comprising a glucocorticoid analog.
27 . The method of any one of claims 1 - 25 , wherein the host cell produces at least 1.5-fold greater quantities of total rAAV compared to that produced by a host cell not contacted with a solution comprising a glucocorticoid analog.
28 . A rAAV produced by the method of any one of claims 1 - 27 .
29 . A composition comprising the rAAV of claim 28 .
30 . A composition comprising a host cell and a glucocorticoid analog.