	US 12,011,459 B2
	Methods for manufacturing T cells by direct sorting and compositions thereof
Amir Alpert, Houston, TX (US); Dominik Maurer, Moessingen (DE); Anastasiya Smith, Houston, TX (US); Claudia Wagner, Tuebingen (DE); and Ali Mohamed, Sugar Land, TX (US)
Assigned to IMMATICS US, INC., Houston, TX (US); and IMMATICS BIOTECHNOLOGIES GMBH, Tübingen (DE)
Filed by Immatics US, Inc., Houston, TX (US); and Immatics Biotechnologies GmbH, Tübingen (DE)
Filed on Jun. 4, 2020, as Appl. No. 16/893,246.
Claims priority of provisional application 62/858,167, filed on Jun. 6, 2019.
Claims priority of application No. 102019129341.3 (DE), filed on Oct. 30, 2019.
Prior Publication US 2021/0002610 A1, Jan. 7, 2021
Int. Cl. A61K 35/17 (2015.01); A61K 31/437 (2006.01); A61K 31/5383 (2006.01); A61K 35/76 (2015.01); C07K 14/47 (2006.01); C07K 14/725 (2006.01); C12N 5/0783 (2010.01); G01N 15/14 (2006.01); G01N 33/554 (2006.01); G01N 15/10 (2006.01); G01N 15/149 (2024.01)

CPC A61K 35/17 (2013.01) [A61K 31/437 (2013.01); A61K 31/5383 (2013.01); A61K 35/76 (2013.01); C07K 14/47 (2013.01); C07K 14/7051 (2013.01); C12N 5/0636 (2013.01); G01N 15/1459 (2013.01); G01N 33/554 (2013.01); C12N 2501/2307 (2013.01); C12N 2501/50 (2013.01); C12N 2523/00 (2013.01); G01N 2015/1006 (2013.01); G01N 2015/1402 (2013.01); G01N 15/149 (2024.01); G01N 2333/705 (2013.01); G01N 2333/7051 (2013.01); G01N 2500/10 (2013.01)]

14 Claims

1. A method for preparing T cells, comprising

(a) contacting CD8+ T cells with a first multimer comprising a target peptide in complex with an MHC molecule in the presence of a second multimer comprising an irrelevant peptide in complex with an MHC molecule and a third multimer comprising the target peptide in a complex with an MHC molecule,

wherein the first multimer is labelled with a first detectable agent, the second multimer is labelled with a second detectable agent, and the third multimer is labelled with a third detectable agent,

wherein the first, the second, and the third detectable agents are different detectable agents,

wherein the irrelevant peptide has less than 50% sequence identity to the target peptide,

wherein the contacting is performed at about 4° C.,

(b) sorting the contacted CD8+ T cells to collect a first sorted CD8+ T cells that are detected positive for the first and the third detectable agents,

(c) sorting the first sorted CD8+ T cells to collect a second sorted CD8+ T cells that are detected negative for the second detectable agent,

wherein the second sorted CD8+ T cells comprise at least 20-fold less false-positive CD8+ T cells that are detected positive for the first and the third detectable agents than that prepared by performing steps (a)-(c) in the absence of the second multimer, and

(d) expanding the second sorted CD8+ T cells prepared in the presence of the second multimer.