	US 11,851,676 B2
	Human neural precursor cells with inducible STIM1 knockdown
Renjitha Gopurappilly, Karnataka (IN); Gaiti Hasan, Karnataka (IN); and Bipan Kumar Deb, Karnataka (IN)
Assigned to NATIONAL CENTRE FOR BIOLOGICAL SCIENCES, Karnataka (IN)
Filed by NATIONAL CENTRE FOR BIOLOGICAL SCIENCES, Karnataka (IN)
Filed on Apr. 17, 2019, as Appl. No. 16/386,907.
Claims priority of application No. 201841014670 (IN), filed on Apr. 18, 2018.
Prior Publication US 2020/0017828 A1, Jan. 16, 2020
Int. Cl. C12N 15/113 (2010.01); C12N 15/86 (2006.01); C12N 5/0793 (2010.01); C12N 5/074 (2010.01)

CPC C12N 5/0619 (2013.01) [C12N 5/0696 (2013.01); C12N 15/113 (2013.01); C12N 15/86 (2013.01); C12N 2310/141 (2013.01); C12N 2740/15041 (2013.01)]

4 Claims

1. A process for transducing human neural precursor cells (hNPCs) or human neural precursor cell lines with lentiviral constructs for knockdown of STIM1 expression by shRNA comprising:

providing a lentiviral transfer vector containing sequences that package as a viral genome and encode for the shRNA for knockdown of STIM1 expression, wherein the sequences encoding the shRNA comprises TAATATTGCACCTCCACCTCAT (SEQ ID NO: 1), TTTATGATCTACATCATCCAGG (SEQ ID NO:2), and TCCAGTGAGTGGATGCCAGGGT (SEQ ID NO: 3);

co-transfecting the lentiviral transfer vector with lentivirus-based second generation packaging vectors encoding env, gag and pol protein into a packaging cell line for releasing viral particles therefrom;

harvesting the viral particles that contain the sequences that package as a viral genome and encode for the shRNA for knockdown of STIM1 expression from the supernatant of the packaging cell line;

providing human neural precursor cells (hNPCs) or human neural precursor cell lines; and

carrying out gene expression modulation of said human neural precursor cells (hNPCs) or human neural precursor cell lines by applying the harvested viral particles to said human neural precursor cells (hNPCs) or human neural precursor cell lines and inducing shRNA expression for knockdown of STIM1 expression, thereby regulating intracellular calcium signaling and decreasing Store Operated Calcium Entry (SOCE).