Skip to main content
. 2023 Sep 20;34:102038. doi: 10.1016/j.omtn.2023.102038

Figure 4.

Figure 4

Selection of scaffolds for the expansion of the miRNA-based shRNA platform to a 4-plex

Starting with the sc106a-sc20b 2-plex as a basis, the platform was expanded into a 4-plex by the addition of two extra scaffolds. In a first cluster design, sc20a was added in third position, and sc93, sc106b, or sc17 in fourth position. In a second cluster design, sc93, sc106b, or sc17 were added in third position and sc20a in fourth position. Each position carried a different validated shGuide sequence (β2M, CD3ζ, CD28, and CD95). KD efficiency for each target was assessed by flow cytometry by monitoring the MFI of TCRa/b, HLA ABC, CD28, and CD95, respectively, in CAR T cells transduced with the 4-plex constructs or with a no-shRNA control. The different 4-plex combinations were tested in the context of both an anti-CD19 CAR (A) and an anti-BCMA CAR (B). MFIs are depicted as relative to the no-shRNA control. Bars represent mean ± SD for three independent biological replicates. Each symbol superimposed to the bars represents a different PBMC donor. All p values refer to the comparison with the no-shRNA control. The absolute MFI values for the no-shRNA control samples are reported in Table S2.