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. 2025 Feb 25;27(6):1443–1457. doi: 10.1093/neuonc/noaf055

Figure 5.

CRISPRoff targeting of MGMT in primary GBM.

CRISPRoff targeting of MGMT in primary GBM. (A) Western blots against MGMT protein in monoclonal (SF7996) or polyclonal (SF14259, SF14346, SF14590, SF14599) populations of patient-derived primary GBM cultures at the indicated number of days following delivery of CRISPRoff and the indicated sgRNA(s). (B) Crystal violet assay of cell viability 7 days following cell seeding and TMZ treatment (replenished every 3 days) at the indicated concentrations for SF7996 primary GBM. Quantification of cell growth (right). Scale bar = 1000 µm. (C) Propidium iodide cell cycle analysis of SF7996 cells treated with TMZ. P value = 2 tailed Student’s t-test for S-phase compared to sgScrambled 20 µM TMZ condition, n = 3 biological replicates. (D) Brightfield light micrographs of CRISPRoff-modified primary GBM cultures (SF14259, SF14346, SF14590, SF14599) 15–35 days following cell seeding, corresponding to the time at which vehicle-treated cells reached confluency. TMZ was replenished every 3 days during culture. Integrated intensity from live cells was quantified (right) for each cell type. (E) Apoptosis assay of primary GBM cultures 7 days following cell seeding and TMZ treatment. P value = 2 tailed Student’s t-test compared to cognate sgScrambled.