inc  (Developmental Studies Hybridoma Bank)

Journal: Oncology Reports

Article Title: Saikosaponin D overcomes gemcitabine resistance in pancreatic cancer via AKT/mTOR pathway inhibition and synergistic induction of apoptosis and autophagy

doi: 10.3892/or.2025.9033

Figure Lengend Snippet: AKT/mTOR pathway inhibition by GEM and SSD combination therapy. (A) Immunocytochemistry staining of AKT/mTOR signaling proteins in MIA PaCa-2 and AsPC-1 cells treated with Ctrl, G0.25, S4 or the combination (G0.25 + S4; magnification, ×40; insets show phosphoprotein details). Scale bar, 50 µm. (B) Western blot analysis of p-AKT (60 kDa), AKT (56 kDa), p-mTOR (289 kDa), mTOR (289 kDa) and β-actin (42 kDa). Semi-quantification of phosphorylation ratios in (C) MIA PaCa-2 and (D) AsPC-1 cells. In the figure, GEM represents 0.25 µmol/l GEM and SSD represents 4 µmol/l SSD. Data are presented as the mean ± SD (n=5). Compared with GEM monotherapy, the G0.25 + S4 combination led to a greater reduction in both p-AKT/AKT and p-mTOR/mTOR ratios (both P adj <0.0001), with large effect sizes (η 2 =0.73 and 0.81, respectively). All P-values are Benjamini-Hochberg-adjusted. ****P<0.0001 for G0.25 + S4 vs. Ctrl; ## P<0.01, ### P<0.001 for G0.25 + S4 vs. G0.25. Statistical analysis was performed using one-way ANOVA with Tukey's post hoc test. Ctrl, control; GEM, gemcitabine; SSD, Saikosaponin D; p-, phosphorylated.

Article Snippet: The following antibodies were purchased from Proteintech Group, Inc.: Mouse monoclonal antibodies against mTOR (1:500; cat. no. 66888-1-Ig), phosphorylated (p-)mTOR (Ser2448) (1:2,000; cat. no. 67778-1-Ig), p-AKT (Ser473) (1:500; cat. no. 66444-1-Ig), caspase-3 (1:2,000; cat. no. 66470-2-Ig), cleaved caspase-3 (1:1,000; cat. no. 66470-2-Ig), Bax (1:500; cat. no. 60267-1-Ig), AKT (1:500; cat. no. 60203-2-Ig) and β-actin (1:2,000; cat. no. 66009-1-Ig); as well as rabbit antibodies, including polyclonal antibodies against Bcl-2 (1:1,000; cat. no. 12789-1-AP), Beclin 1 (1:1,000; cat. no. 11306-1-AP) and LC3 (1:1,000; cat. no. 14600-1-AP).

Techniques: Inhibition, Immunocytochemistry, Staining, Western Blot, Phospho-proteomics, Control

Journal: Oncology Reports

Article Title: Saikosaponin D overcomes gemcitabine resistance in pancreatic cancer via AKT/mTOR pathway inhibition and synergistic induction of apoptosis and autophagy

doi: 10.3892/or.2025.9033

Figure Lengend Snippet: Proposed mechanism of GEM and SSD combination therapy. Co-treatment with GEM and SSD synergistically inhibits the AKT/mTOR pathway, concurrently inducing apoptosis (via Bax/Bcl-2 modulation and caspase-3 activation) and autophagy (via Beclin 1 activation and LC3 conversion), thereby overcoming GEM resistance in pancreatic cancer cells. GEM, gemcitabine; P, phosphorylated; SSD, Saikosaponin D.

Article Snippet: The following antibodies were purchased from Proteintech Group, Inc.: Mouse monoclonal antibodies against mTOR (1:500; cat. no. 66888-1-Ig), phosphorylated (p-)mTOR (Ser2448) (1:2,000; cat. no. 67778-1-Ig), p-AKT (Ser473) (1:500; cat. no. 66444-1-Ig), caspase-3 (1:2,000; cat. no. 66470-2-Ig), cleaved caspase-3 (1:1,000; cat. no. 66470-2-Ig), Bax (1:500; cat. no. 60267-1-Ig), AKT (1:500; cat. no. 60203-2-Ig) and β-actin (1:2,000; cat. no. 66009-1-Ig); as well as rabbit antibodies, including polyclonal antibodies against Bcl-2 (1:1,000; cat. no. 12789-1-AP), Beclin 1 (1:1,000; cat. no. 11306-1-AP) and LC3 (1:1,000; cat. no. 14600-1-AP).

Techniques: Activation Assay