US 12,419,913 B2
Modification of CAR-T cells
Yifang Shui, Boston, MA (US); Jeremy Minshull, Los Altos, CA (US); Maggie Lee, San Jose, CA (US); Feng Shi, Winchester, MA (US); and Mark Cobbold, Winchester, MA (US)
Assigned to DNA Twopointo, Inc., Newark, CA (US); and The General Hospital Corporation, Boston, MA (US)
Filed by DNA Twopointo, Inc., Newark, CA (US); and The General Hospital Corporation, Boston, MA (US)
Filed on Mar. 20, 2023, as Appl. No. 18/186,901.
Application 18/186,901 is a continuation in part of application No. 17/429,342, previously published as PCT/US2020/017283, filed on Feb. 7, 2020.
Claims priority of provisional application 63/491,171, filed on Mar. 20, 2023.
Claims priority of provisional application 63/321,262, filed on Mar. 18, 2022.
Claims priority of provisional application 62/803,142, filed on Feb. 8, 2019.
Prior Publication US 2023/0302054 A1, Sep. 28, 2023
Int. Cl. A61K 35/17 (2015.01); A61P 35/00 (2006.01); C07K 14/725 (2006.01)
CPC A61K 35/17 (2013.01) [A61P 35/00 (2018.01); C07K 14/7051 (2013.01)] 20 Claims
 
1. A modified T cell expressing a heterologous polynucleotide, the heterologous polynucleotide comprising:
(A) a first gene comprising a nucleotide sequence encoding a protein selected from the group consisting of Bcl-xL, Survivin, or CD28-D124E/T195P, wherein the first gene is operably linked to a first heterologous regulatory sequence effective for expression of the protein within a T cell, thereby enhancing survival of the T cell; and
(B) a second gene comprising a nucleotide sequence encoding a chimeric antigen receptor (CAR), wherein the CAR comprises an extracellular domain that specifically binds to a CD19 antigen, and wherein the second gene is operably linked to a second heterologous regulatory sequence effective for expression of the CAR in the T cell,
wherein the first heterologous regulatory sequence and the second heterologous regulatory sequence are separate regulatory sequences, and
wherein the heterologous polynucleotide is integrated into the genome of the modified T cell.
 
4. A method for preparing a modified T cell,
wherein the method comprises:
introducing a heterologous polynucleotide into a T cell ex-vivo, the heterologous polynucleotide comprising:
(A) a first gene comprising a nucleotide sequence encoding a protein selected from the group consisting of Bcl-xL, Survivin, or CD28-D124E/T195P, wherein the first gene is operably linked to a first heterologous regulatory sequence effective for expression of the protein within the T cell, thereby enhancing survival of the T cell; and
(B) a second gene comprising a nucleotide sequence encoding a chimeric antigen receptor (CAR), wherein the CAR comprises an extracellular domain that specifically binds to a CD19 antigen, and wherein the second gene is operably linked to a second heterologous regulatory sequence effective for expression of the CAR in the T cell,
wherein the first heterologous regulatory sequence and the second heterologous regulatory sequence are separate regulatory sequences.
 
15. A polynucleotide, comprising:
(A) a first gene comprising a nucleotide sequence encoding a protein selected from the group consisting of Bcl-xL, Survivin, or CD28-D124E/T195P, wherein the first gene is operably linked to a first regulatory sequence effective for expression of the protein within a T cell, thereby enhancing survival of the T cell; and
(B) a second gene comprising a nucleotide sequence encoding a chimeric antigen receptor (CAR), wherein the CAR comprises an extracellular domain that specifically binds to a CD19 antigen, and wherein the second gene is operably linked to a second regulatory sequence effective for expression of the CAR in the T cell,
wherein the first regulatory sequence and the second regulatory sequence are separate sequences.