About InVivoMAb anti-human/mouse GRP78 The C107 monoclonal antibody reacts with human and mouse Glucose Regulated Protein 78 (GRP78) also known as BiP or HSPA5. GRP78 is a stress inducible, pro-survival, endoplasmic reticulum chaperone in the HSP70 family. Cell Surface GRP78 acts as a multifunctional receptor that affects both cell proliferation and viability. GRP78 is involved in many cellular processes, including translocating newly synthesized polypeptides across the ER membrane, facilitating the folding and assembly of proteins, targeting misfolded proteins for ER-associated degradation (ERAD), regulating calcium homeostasis, and serving as an ER stress sensor. Expression of cell surface GRP78 is associated with increased malignancy and resistance to chemotherapy and radiotherapy in various cancers, particularly prostate cancer. The C107 antibody binds to the C‐terminal domain of GRP78 and has been shown to have anti-tumoral potential in various in vitro and in vivo tumor models. InVivoMAb anti-human/mouse GRP78 Specifications IsotypeMouse IgG2b, κ Recommended Isotype Control(s)InVivoMAb mouse IgG2b isotype control, unknown specificity Recommended Dilution BufferInVivoPure pH 7.0 Dilution Buffer ImmunogenFull-length recombinant murine GRP78 protein Reported Applicationsin vivo GRP78 targeting in vitro GRP78 targeting Flow cytometry FormulationPBS, pH 7.0 Contains no stabilizers or preservatives Endotoxin<2EU/mg (<0.002EU/μg) Determined by LAL gel clotting assay Purity>95% Determined by SDS-PAGE Sterility0.2 μm filtered ProductionPurified from cell culture supernatant in an animal-free facility PurificationProtein A Molecular Weight150 kDa StorageThe antibody solution should be stored at the stock concentration at 4°C. Do not freeze. Application ReferencesInVivoMAb anti-human/mouse GRP78 (CLONE: C107)Mo L, Bachelder RE, Kennedy M, Chen PH, Chi JT, Berchuck A, Cianciolo G, Pizzo SV (2015). "Syngeneic Murine Ovarian Cancer Model Reveals That Ascites Enriches for Ovarian Cancer Stem-Like Cells Expressing Membrane GRP78" Mol Cancer Ther 14(3):747-56. PubMedPatients with ovarian cancer are generally diagnosed at FIGO (International Federation of Gynecology and Obstetrics) stage III/IV, when ascites is common. The volume of ascites correlates positively with the extent of metastasis and negatively with prognosis. Membrane GRP78, a stress-inducible endoplasmic reticulum chaperone that is also expressed on the plasma membrane ((mem)GRP78) of aggressive cancer cells, plays a crucial role in the embryonic stem cell maintenance. We studied the effects of ascites on ovarian cancer stem-like cells using a syngeneic mouse model. Our study demonstrates that ascites-derived tumor cells from mice injected intraperitoneally with murine ovarian cancer cells (ID8) express increased (mem)GRP78 levels compared with ID8 cells from normal culture. We hypothesized that these ascites-associated (mem)GRP78(+) cells are cancer stem-like cells (CSC). Supporting this hypothesis, we show that (mem)GRP78(+) cells isolated from murine ascites exhibit increased sphere forming and tumor initiating abilities compared with (mem)GRP78(-) cells. When the tumor microenvironment is recapitulated by adding ascites fluid to cell culture, ID8 cells express more (mem)GRP78 and increased self-renewing ability compared with those cultured in medium alone. Moreover, compared with their counterparts cultured in normal medium, ID8 cells cultured in ascites, or isolated from ascites, show increased stem cell marker expression. Antibodies directed against the carboxy-terminal domain of GRP78: (i) reduce self-renewing ability of murine and human ovarian cancer cells preincubated with ascites and (ii) suppress a GSK3α-AKT/SNAI1 signaling axis in these cells. Based on these data, we suggest that (mem)GRP78 is a logical therapeutic target for late-stage ovarian cancer.de Ridder GG, Ray R, Pizzo SV (2012). "A murine monoclonal antibody directed against the carboxyl-terminal domain of GRP78 suppresses melanoma growth in mice" Melanoma Res 22(3):225-35. PubMedThe HSP70 family member GRP78 is a selective tumor marker upregulated on the surface of many tumor cell types, including melanoma, where it acts as a growth factor receptor-like protein. Receptor-recognized forms of the proteinase inhibitor α2-macroglobulin (α2M*) are the best-characterized ligands for GRP78, but in melanoma and other cancer patients, autoantibodies arise against the NH2-terminal domain of GRP78 that react with tumor cell-surface GRP78. This causes the activation of signaling cascades that are proproliferative and antiapoptotic. Antibodies directed against the COOH-terminal domain of GRP78, however, upregulate p53-mediated proapoptotic signaling, leading to cell death. Here, we describe the binding characteristics, cell signaling properties, and downstream cellular effects of three novel murine monoclonal antibodies. The NH2-terminal domain-reactive antibody, N88, mimics α2M* as a ligand and drives PI 3-kinase-dependent activation of Akt and the subsequent stimulation of cellular proliferation in vitro. The COOH-terminal domain-reactive antibody, C38, acts as an antagonist of both α2M* and N88, whereas another, C107, directly induces apoptosis in vitro. In a murine B16F1 melanoma flank tumor model, we demonstrate the acceleration of tumor growth by treatment with N88, whereas C107 significantly slowed tumor growth whether administered before (P<0.005) or after (P<0.05) tumor implantation.
