	US 12,270,027 B2
	Methods and kits for identifying cancer treatment targets
Jennifer A. Doudna, Berkeley, CA (US); Chun-Hao Huang, Berkeley, CA (US); Spencer C. Knight, Berkeley, CA (US); and Nami Saghaei, Berkeley, CA (US)
Assigned to The Regents of the University of California, Oakland, CA (US)
Filed by The Regents of the University of California, Oakland, CA (US)
Filed on Feb. 1, 2023, as Appl. No. 18/104,640.
Application 18/104,640 is a continuation of application No. 17/717,594, filed on Apr. 11, 2022, granted, now 11,584,930.
Application 17/717,594 is a continuation of application No. 17/284,660, previously published as PCT/US2019/058853, filed on Oct. 30, 2019.
Claims priority of provisional application 62/753,631, filed on Oct. 31, 2018.
Prior Publication US 2024/0209352 A1, Jun. 27, 2024
This patent is subject to a terminal disclaimer.
Int. Cl. C12N 15/10 (2006.01); C12N 9/22 (2006.01); C12N 15/11 (2006.01); C12N 15/90 (2006.01); G16B 30/10 (2019.01); G16B 40/00 (2019.01)

CPC C12N 15/1079 (2013.01) [C12N 9/22 (2013.01); C12N 15/11 (2013.01); C12N 15/907 (2013.01); G16B 30/10 (2019.02); G16B 40/00 (2019.02); C12N 2310/20 (2017.05); C12N 2320/10 (2013.01); C12N 2800/80 (2013.01)]

25 Claims

1. A method comprising:

(a) introducing into a cancer cell and into a corresponding non-cancer cell of a same cell type as the cancer cell, a clustered regularly interspaced short palindromic repeat (CRISPR)/Cas system comprising:

i) a CRISPR/Cas effector polypeptide; and

ii) a CRISPR/Cas guide ribonucleic acid (RNA), or a nucleotide sequence encoding the CRISPR/Cas guide RNA, wherein the CRISPR/Cas guide RNA targets a target gene, thereby producing a modified cancer cell and a modified non-cancer cell, both having reduced expression of the target gene;

(b) sequencing RNA obtained from the modified cancer cell and the modified non-cancer cell to determine a molecular feature or a phenotype in the modified cancer cell and the modified non-cancer cell, thereby generating RNA sequencing data associated with the molecular feature or phenotype; and

(c) computer processing the RNA sequencing data associated with the molecular feature or phenotype to determine that the molecular feature or phenotype is indicative of a reduction of cancerous state of the modified cancer cell,

wherein the determining comprises assessing an extent of transcriptional reversion toward a non-cancer cell expression level as a result of having reduced expression of the target gene.