METHOD FOR IMPROVING GENETIC ENGINEERING OF ADOPTIVE CELL THERAPIES
The present disclosure provides methods for electroporating cells, in particular cells useful in cellular-based therapies, that increase cell viability, cell proliferation and transfection efficiency. The methods suitably utilize a post-electroporation supplement to improve the genetic engineering of the various cells.
1 . A method of electroporating a cell, comprising:
a. suspending the cell in an electroporation buffer to create a cell suspension;
b. subjecting the cell to one or more electroporation pulses; and
c. adding a post-electroporation supplement to the cell suspension, wherein the post-electroporation supplement comprises a lipid enriched, human serum.
2 . The method of claim 1 , wherein viability of the cell is at least 30% greater than a cell that has not received the post-electroporation supplement.
3 . The method of claim 1 or claim 2 , wherein a viable number of cells is at least 1.5-fold greater than a cell that has not received the post-electroporation supplement.
4 . The method of any one of claims 1 to 3 , wherein the cell is a T-cell.
5 . The method of any one of claims 1 to 3 , wherein the cell is a hematopoietic stem cell.
6 . The method of any one of claims 1 to 5 , wherein the post-electroporation supplement comprises lipid enriched, human AB serum.
7 . The method of any one of claims 1 to 6 , wherein the post-electroporation supplement is added at about 2% to about 10% by volume of the cell suspension.
8 . A method of transfecting a genetic construct into a cell, comprising:
a. suspending the cell in an electroporation buffer to create a cell suspension;
b. introducing the genetic construct into the cell suspension;
c. subjecting the cell to one or more electroporation pulses; and
d. adding a post-electroporation supplement to the cell suspension, wherein the post-electroporation supplement comprises a lipid enriched, human serum.
9 . The method of claim 8 , wherein viability of the cell is at least 30% greater than a cell that has not received the post-electroporation supplement.
10 . The method of claim 8 or claim 9 , wherein a viable number of cells is at least 1.5-fold greater than a cell that has not received the post-electroporation supplement.
11 . The method of any one of claims 8 to 10 , wherein a transfection efficiency of the cell is at least 15% greater than a cell that has not received the post-electroporation supplement.
12 . The method of any one of claims 8 to 11 , wherein the cell is a T-cell.
13 . The method of any one of claims 8 to 11 , wherein the cell is a hematopoietic stem cell.
14 . The method of any one of claims 8 to 13 , wherein the post-electroporation supplement comprises a lipid enriched, human AB serum.
15 . The method of any one of claims 8 to 14 , wherein the genetic construct is RNA or mRNA.
16 . The method of claim 15 , wherein the RNA is antisense RNA, siRNA or microRNA.
17 . The method of any one of claims 8 to 14 , wherein the genetic construct is DNA.
18 . The method of any one of claims 8 to 14 , wherein the genetic construct is a viral vector or a viral plasmid.
19 . The method of any one of claims 8 to 14 , wherein the genetic construct comprises a transposon.
20 . The method of any one of claims 8 to 14 , wherein the genetic construct comprises an endonuclease.
21 . The method of any one of claims 8 to 20 , wherein the post-electroporation supplement is added at about 2% to about 10% by volume of the cell suspension.
22 . A composition comprising a lipid enriched, human AB serum, wherein the composition increases cell viability of an electroporated cell population.
23 . The composition of claim 22 , prepared by heating human AB serum to a temperature of about 60° C. to about 80° C. for about 20-45 minutes.
24 . The composition of claim 22 , prepared by passing human AB serum through a silica column and recovering the lipid enriched, human AB serum.