Fig 2: Immunohistochemical (IHC) staining for lipolysis-stimulated lipoprotein receptor (LSR) in gastric cancer (GC) patient samples(A) Primary GC tissue (signet cell carcinoma). (B) Normal tissues: gastric mucosa, lymph node, and liver. (C) Primary GC, lymph node metastasis, and distant metastasis (peritoneum and liver) of 7 patients with GC. (D) Typical weak and strong LSR staining in well- and poorly differentiated GC. Scale bars: black = 100 μm

Fig 3: Antitumor effect of the anti-human lipolysis-stimulated lipoprotein receptor (LSR) monoclonal antibody (mAb) (#1–25) in gastric cancer (GC) xenograft mouse models (MKN74 and NUGC-3; female ICR nu/nu mice 6–8 weeks of age were injected with 5 × 106 GC cells)When tumor volumes reached approximately 100 mm3, control IgG2a mAb or anti-hLSR mAb (#1–25) was injected intraperitoneally. (A) Tumor volumes were measured twice per week. Immunohistochemical analysis of LSR in subcutaneous tumors of GC cell lines in xenograft mouse models are shown in the insets; scale bar = 100 μm. Values shown represent the means ± standard errors of the means. (B) Tumor volumes were calculated after the end of treatment and compared using Student's t-tests (*P < 0.05, **P < 0.01). Values shown represent the means ± standard deviations (SDs). (C) Western blot analysis of p-STAT3/STAT3, pAKT(thr308), pAKT(ser473), and AKT in MKN74 and NUGC-3 cell-derived tissues from control IgG2a mAb or anti-hLSR mAb-injected animals. (D) Mice body weights were measured twice per week. Values shown represent the means ± SDs.

Fig 4: LSR overexpression renders MDA-MB-231 cells resistant to doxorubicin through inhibition of apoptosis.A) 231/LSR cells were more resistant to doxorubicin than 231/C cells. The paired cell lines were treated with doxorubicin with concentrations ranging from 0.03 to 10 µM for four days, followed by IC50 analysis based on CCK-8 assays. The statistical differences in IC50 values between the paired groups based on three independent experiments were analyzed using Welch’s t-test, with a significance level of p < 0.01. B) Clonogenic analysis of doxorubicin treated 231/C and 231/LSR cells. The cells were treated with doxorubicin at indicated concentrations for 10 d, followed by crystal violet staining. Colony numbers of each group were quantified and analyzed with Student’s t-test. C) LSR overexpression attenuates doxorubicin-induced activation of Caspase cascade in MDA-MB-231 cells. The paired cell lines were treated with doxorubicin at the indicated concentrations for 24 hours, followed by Western blot detection of PARP (both total and cleaved PARP (c-PARP)), total Caspase 3 (Casp3), and cleaved Caspase 3 (c-Casp3). The relative protein levels of c-PARP, Casp3, and c-Casp3 (signals from both 19 and 17 kD bands) were quantified and are indicated beneath the corresponding blots. D) The effect of LSR overexpression on doxorubicin-induced apoptosis in MDA-MB-231 cells was measured using an apoptosis ELISA assay. Data from clonogenic assays and apoptosis ELISA were analyzed with one-way ANOVA followed by Tukey’s post hoc test, *p < 0.05; **p < 0.01.