Use of nuclear membrane rupture for transduction with recombinant AAV gene therapy vectors.
Ownerpeter wardinventor
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Abstract
A method for transducing cells with rAAV vectors that results in more rapid transduction than standard infection is described. The method involves induced nuclear membrane rupture. It requires less vector to achieve the same transduction level as standard infection and transduces individual cells with multiple vectors, even at low percentages of total transduced cells. It may be useful in treating people bitten by venomous snakes or who have been exposed to rabies. In one iteration it can target cells that have endogenous rupture of their nuclear membranes.
Claims14
An rAAV gene therapy method that results in an increased number of expressing recombinant adeno-associated virus vectors in cells infected with said vector and in a decreased length of time between infection of the cell by the vector and expression of the transgene in comparison with standard methods of infection, which method comprises:
- infecting the cells;
- incubating the cells until vector has accumulated around the nuclear envelope;
- squeezing the infected cells individually to an extent that is mild enough to leave the cell membrane unruptured, so that the cells survive, but that is sufficiently robust to rupture the nuclear membrane so that vector particles enter the nucleus in a passive manner.
A method of gene therapy with rAAV vectors in which cells to be targeted have endogenous nuclear membrane rupture;
- which method can consist of infections at very low levels of vector so as to provoke a minimal adaptive or innate immune response, which low vector level is made feasible because the passive uptake allows productive therapy by particles that would not be able to gain nuclear entry by an active mechanism;
- followed by multiple rounds of low level infections separated by intervals of waiting.
Dependencies are inferred from the claim text.