Journal: Blood Cancer Journal

Article Title: Downregulation of PA28α induces proteasome remodeling and results in resistance to proteasome inhibitors in multiple myeloma

doi: 10.1038/s41408-020-00393-0

Figure Lengend Snippet: a Western blot analysis of ubiquitinated proteins in total protein lysates extracted from LP1 and RPMI8226 PA28α knockdown stable cells. β-actin as a loading control. b OPP pulse-chase assay of proteasome degradation and protein synthesis in LP1 PA28α knockdown stable cells. Western blot analysis of immunoglobulin lambda light chain (ƛ IgL) ( c ), eIF2α, p62/SQSTM1, and LC3B ( d ) in LP1 and RPMI8226 PA28α knockdown stable cells, β-actin as a loading control. ** P < 0.01, Student’s t test. e Working model of PA28α knockdown in MM.

Article Snippet: Antibodies used were as follows: PA28α (Cell Signaling), PSMA2 (Cell Signaling), S5a (Cell signaling), PA28β (Cell Signaling), Phospho-eIF2α (Ser51) (Cell signaling), eIF2α (Cell signaling), α-tubulin (Genetex), PA28γ (Genetex), PSMB5 (Genetex), PSMB6 (Enzo life science), PSMB7 (Genetex), PSMB8 (Genetex), PSMB9 (R&D systems), PSMB10 (R&D systems), Rpt5 (Enzo life science), Ubiquitin (Cell Signaling), β-actin (Santa Cruz Technology), TCF11/NRF1 (Cell Signaling), LC3B (Cell Signaling), p62/SQSTM1 (MBL International) human Ig lambda light chain (R&D systems), actin (Sigma). pLKO.1 empty vector, shRNA vector targeting human PA28α, NRF1 siRNA (ON-TARGETplus SMARTpool), PA28α siRNA (Accell SMARTpool), and control siRNA were purchased from Dharmacon.

Techniques: Western Blot, Knockdown, Control, Pulse Chase

Journal: Molecular Therapy

Article Title: Expression of Immunomodulatory Neutrophil-activating Protein of Helicobacter pylori Enhances the Antitumor Activity of Oncolytic Measles Virus

doi: 10.1038/mt.2012.4

Figure Lengend Snippet: Schematic representation of the recombinant measles virus (MV) strains used in the experiments: MV-lambda, MV-lambda-NAP, MV-s-NAP and MV-NIS. NAP, neutrophil-activating protein.

Article Snippet: Proteins were transferred to polyvinylidene fluoride membrane (Bio-Rad) and probed with the NAP-specific monoclonal antibody 16F4 or antibody against human lambda light chain as previously described.

Techniques: Recombinant, Virus

Journal: Molecular Therapy

Article Title: Expression of Immunomodulatory Neutrophil-activating Protein of Helicobacter pylori Enhances the Antitumor Activity of Oncolytic Measles Virus

doi: 10.1038/mt.2012.4

Figure Lengend Snippet: In vitro antitumor activity of engineered measles virus (MV) strains against breast cancer cells. (a) MCF-7 cell line and the highly tumorigenic MDA231-lu-P4 in vivo derivative of (b) MDA-MB-231 cells were infected with MV strains at an multiplicity of infection (MOI) = 1.0 and cell viability was determined by MTT assay. Both MV-lambda-NAP and MV-lambda viruses propagated rapidly causing complete destruction of the breast cancer monolayers 72 hours postinfection. MV-s-NAP and MV-NIS showed similar tumor cell-killing kinetics with complete eradication of MCF-7 cells or ~80% reduction of cell viability of MDA231-lu-P4 cells. (c) Vero cells were used as control. The data are presented as percent of untreated control cells ± SD. (d) NAP transgene induced significant interleukin (IL)-8 expression in MV vector-infected THP-1 cells. THP-1 monocytic cells were infected with MV-s-NAP or control MV-lambda virus at a MOI = 0.1 and incubated for 72 hours. Supernatants were collected and IL-8 concentration was measured by enzyme-linked immunosorbent assay (ELISA) specific for human IL-8. The effect of TNF-α (one of the known NAP-triggered inflammatory cytokines) on proliferation of MV-infected or uninfected MDA231-lu-P4 cells was examined by MTT assay (e,f). MDA231-lu-P4 cells were plated at 104 cell/well density and inoculated at MOI = 0.5 of MV-s-NAP in the presence or absence of 250 U/ml recombinant human TNF-α (e). The same experiment was repeated with lower cell density (2.5 × 103 per well) and MV-s-NAP at MOI of 0.25 (f).

Article Snippet: Proteins were transferred to polyvinylidene fluoride membrane (Bio-Rad) and probed with the NAP-specific monoclonal antibody 16F4 or antibody against human lambda light chain as previously described.

Techniques: In Vitro, Activity Assay, Virus, In Vivo, Infection, MTT Assay, Control, Expressing, Plasmid Preparation, Incubation, Concentration Assay, Enzyme-linked Immunosorbent Assay, Recombinant

Journal: Molecular Therapy

Article Title: Expression of Immunomodulatory Neutrophil-activating Protein of Helicobacter pylori Enhances the Antitumor Activity of Oncolytic Measles Virus

doi: 10.1038/mt.2012.4

Figure Lengend Snippet: Measles virus (MV) infection and neutrophil-activating protein (NAP) transgene expression in the malignant pleural effusion of mice-bearing MDA231-lu-P4 pleural xenografts. Mice were treated by a single transthoracic (t.t.). injection of MV-s-NAP or MV-lambda. NAP transgene expression in pleural fluid was demonstrated by immunoblotting using (a) NAP-specific monoclonal antibody (MAb) 16F4 or (b) lambda chain-specific antibody. The secretory form of the NAP transgene was detected in the pleural fluid of four of four MV-s-NAP-treated mice (lanes 1–4) but not in MV-lambda-injected (lanes 5, 6) control mice (a). Human lambda light chain was detected in three of four samples from MV-lambda-injected (lanes 3–6) mice but not in samples from MV-s-NAP-treated (lanes 1, 2) animals (b) using a lambda chain-specific detection antibody. MV-s-NAP (c) induced large multinucleated syncytia in infected tumor cells (Giemsa staining) in the pleural fluid of mice with MDA231-lu-P4 xenografts. Immunohistochemistry (IHC) staining for neutrophils in the pleural fluid of MV-s-NAP is shown in (d). MV-s-NAP was isolated from the pleural fluid by overlay on Vero cells. MV-s-NAP induced giant syncytia formation 24 hours after overlay (e). Uninfected control Vero cells (f).

Article Snippet: Proteins were transferred to polyvinylidene fluoride membrane (Bio-Rad) and probed with the NAP-specific monoclonal antibody 16F4 or antibody against human lambda light chain as previously described.

Techniques: Virus, Infection, Expressing, Injection, Western Blot, Control, Staining, Immunohistochemistry, Isolation

Journal: Molecular Therapy

Article Title: Expression of Immunomodulatory Neutrophil-activating Protein of Helicobacter pylori Enhances the Antitumor Activity of Oncolytic Measles Virus

doi: 10.1038/mt.2012.4

Figure Lengend Snippet: Therapeutic effect of neutrophil-activating protein (NAP)-expressing measles virus (MV) strains against lung metastatic breast cancer compared to control MV-NIS. (a) Engraftment of the systemically injected MDA231-lu-P3 or P4 cells was confirmed by bioluminescence imaging. (b,c) The animals with MDA231-lu-P3 lung tumors (8 per group) were treated with 10 repeat intravenous (i.v.) injections of 2 × 106 TCID50 of MV-lambda-NAP or heat-inactivated (HI-control) control. In a separate in vivo experiment MDA231-lu-P4 lung metastatic xenografts (9 mice per group) were treated by 7 i.v. injections of 106 TCID50 of MV-s-NAP, MV-NIS or the corresponding heat-inactivated controls (d–f). Both MV-lambda-NAP and MV-s-NAP improved the median survival with 10–12 days (P < 0.05) in this aggressive model of breast cancer metastatic to the lungs.

Article Snippet: Proteins were transferred to polyvinylidene fluoride membrane (Bio-Rad) and probed with the NAP-specific monoclonal antibody 16F4 or antibody against human lambda light chain as previously described.

Techniques: Expressing, Virus, Control, Injection, Imaging, In Vivo

Journal: Bmc Cancer [Electronic Resource]

Article Title: Upregulation of meiosis-specific genes in lymphoma cell lines following genotoxic insult and induction of mitotic catastrophe

doi: 10.1186/1471-2407-6-6

Figure Lengend Snippet: Antibody source and usage

Article Snippet: CREST serum against human kinetochore proteins (Acris Antibodies, Hiddenhausen, Germany) , Mouse anti-human IgG-FITC (Dr. MS Cragg).

Techniques: Plasmid Preparation