Effect of scaffold combination on KD efficiency in a 2-plex miRNA platform
Results of two different approaches followed to build a chimeric cluster: combining two different scaffolds or replicating the same scaffold multiple times. Combinations of sc106b and sc20b as well as repetitions of either sc106a or sc20b were tested in a 2-plex with validated shGuide sequences against CD3ζ, β2M, and CD95. Different configurations of the 2-plex were tested, alternating the position of the scaffolds and of the shGuide sequences. KD efficiencies were evaluated by flow cytometry by comparing the MFI of TCRa/b, HLA ABC, and CD95, respectively, in CAR T cells transduced with the 2-plex and with a no-shRNA control. 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.