Oncogenic CD44 is essential for pancreatic cancer tumorigenesis: A novel targeted therapeutic strategy

Figure 1 Establishment and verification of CD44 knockout pancreatic cancer cell lines. Pancreatic cancer Patu-8988 and Panc-1 cells were stably transfected with Cas9-cDNA-vector. A: CD44 single-chain guide RNAs (sgRNAs) were cloned into a lentivirus vector, and the infectious sgRNAs-lentiviruses were prepared as described in the methods section. Cas9-positive pancreatic cancer Patu-8988 and Panc-1 cells were infected with CD44-sgRNA1 lentivirus, CD44-sgRNA2 lentivirus, or control sgRNA-lentivirus (CD44-NT). Further, Sanger sequencing confirmed the efficacy of CRISPR/Cas9 gene editing; B-E: CD44 knockout was verified by western blotting; F and G: Immunofluorescence staining, and confocal microscope analysis (600 ×), red: CD44, blue: DAPI, Merge: Both CD44 and DAPI. Ctrl: CD44-expressing cells, CD44-sgRNA1: CD44 gene single-chain guide RNA1, CD44-sgRNA2: CD44 gene single-chain guide RNA2. Notably, CD44 was almost completely knocked out by CD44-sgRNA2 in pancreatic cancer cells; accordingly, CD44-sgRNA2-mediated CD44 knockout pancreatic cells were named CD44-KO. Data in Figure 1B-E are shown as the mean ± SD of three independent replicates. ^bP < 0.01 vs CD44-NT in an unpaired t test.