Journal: Pharmaceutics

Article Title: Sulfonated Amphiphilic Poly(α)glutamate Amine—A Potential siRNA Nanocarrier for the Treatment of Both Chemo-Sensitive and Chemo-Resistant Glioblastoma Tumors

doi: 10.3390/pharmaceutics13122199

Figure Lengend Snippet: The development of resistance to TMZ enhances MGMT expression without altering Plk1 oncogene levels in GB. ( A ) Survival analysis performed on OSgbm database comparing MGMT expression (left panel) and Plk1 expression (right panel) of the top 25% longest GB survivors versus the bottom 25% shortest survivors. ( B ) Inhibitory concentration 50% (IC 50 ) plot of parental U251 glioblastoma cells vs. TMZR clone. ( C ) Table summarizing the IC 50 of wild-type U251 cells versus U251 TMZ-R cells. ( D ) TMZ resistance in U251 occurred via upregulation of MGMT, without changing Plk1 levels. ( E ) Intracranial U251 GB tumors show Plk1 expression (scale = 100 µm).

Article Snippet: Cryo-sections were immunostained for: Plk1- using rabbit anti-human/mouse Plk1 antibody (Cat. No. BS-3535R dilution 1:50, Bioss Antibodies, Woburn, MA, USA) and Alexa Fluor 488-goat anti-rabbit secondary antibody (Cat. No. ab150077, dilution 1:300, Abcam, Cambridge, UK); SELP- using mouse anti-human SELP antibody (Cat. No. BBA1, Clone BBIG-E, dilution 1:30, R&D systems, McKinley, MN, USA) and Alexa Fluor 568-goat anti-mouse secondary antibody (Cat. No. ab175473, dilution 1:300, Abcam, Cambridge, UK).

Techniques: Expressing, Concentration Assay

Journal: Pharmaceutics

Article Title: Sulfonated Amphiphilic Poly(α)glutamate Amine—A Potential siRNA Nanocarrier for the Treatment of Both Chemo-Sensitive and Chemo-Resistant Glioblastoma Tumors

doi: 10.3390/pharmaceutics13122199

Figure Lengend Snippet: SELP expression on U251 and U251 TMZ-R 3D spheroids. ( A ) Imaged by confocal microscopy (scale = 20 µm) and ( B ) analyzed by fluorescence activated cell sorting (FACS). ( C ) SELP expression on slices of U251 intracranial tumors and co-localization with Plk1 (scale = 100 µm).

Article Snippet: Cryo-sections were immunostained for: Plk1- using rabbit anti-human/mouse Plk1 antibody (Cat. No. BS-3535R dilution 1:50, Bioss Antibodies, Woburn, MA, USA) and Alexa Fluor 488-goat anti-rabbit secondary antibody (Cat. No. ab150077, dilution 1:300, Abcam, Cambridge, UK); SELP- using mouse anti-human SELP antibody (Cat. No. BBA1, Clone BBIG-E, dilution 1:30, R&D systems, McKinley, MN, USA) and Alexa Fluor 568-goat anti-mouse secondary antibody (Cat. No. ab175473, dilution 1:300, Abcam, Cambridge, UK).

Techniques: Expressing, Confocal Microscopy, Fluorescence, FACS

Journal: Pharmaceutics

Article Title: Sulfonated Amphiphilic Poly(α)glutamate Amine—A Potential siRNA Nanocarrier for the Treatment of Both Chemo-Sensitive and Chemo-Resistant Glioblastoma Tumors

doi: 10.3390/pharmaceutics13122199

Figure Lengend Snippet: Sulfonate modification maintained size and activity of APA complexes. ( A ) Electrophoretic shift assay (EMSA) of polyplexes of APA and APAS with siRNA at increasing N/P ratios. ( B ) Table summarizing the hydrodynamic diameter, polydispersity index (PDI), and zeta potential of the main population of APAS:siPlk1 polyplexes as obtained by Mobius and PALS instruments. ( C ) Specific Plk1 mRNA silencing obtained by RT-PCR, performed on U251 and U251 TMZ-R GB cells following treatment with APAS:siPlk1 polyplexes. ( D , E ) Specific MGMT mRNA silencing obtained by RT-PCR ( D ) and western Blot ( E ) performed on U251 and U251 TMZ-R GB cells following treatment with APAS:siMGMT polyplexes.

Article Snippet: Cryo-sections were immunostained for: Plk1- using rabbit anti-human/mouse Plk1 antibody (Cat. No. BS-3535R dilution 1:50, Bioss Antibodies, Woburn, MA, USA) and Alexa Fluor 488-goat anti-rabbit secondary antibody (Cat. No. ab150077, dilution 1:300, Abcam, Cambridge, UK); SELP- using mouse anti-human SELP antibody (Cat. No. BBA1, Clone BBIG-E, dilution 1:30, R&D systems, McKinley, MN, USA) and Alexa Fluor 568-goat anti-mouse secondary antibody (Cat. No. ab175473, dilution 1:300, Abcam, Cambridge, UK).

Techniques: Modification, Activity Assay, Shift Assay, Reverse Transcription Polymerase Chain Reaction, Western Blot

Journal: Oncotarget

Article Title: Disulfiram, a drug widely used to control alcoholism, suppresses self-renewal of glioblastoma and overrides resistance to temozolomide

doi:

Figure Lengend Snippet: (A) SF188 cells were exposed to DSF for 24 hrs using DMSO as a solvent control. Protein and transcript levels of PLK1 were assessed using immunoblotting or qRT-PCR. (B-C) PLK1 inhibition with siRNA inhibits SF188 cell growth and induced apoptosis based on PARP and caspase 3 cleavage. Scramble RNA oligo was transfected as a control. The efficacy of PLK1 inhibition on SF188 growth is exemplified in combination with 10 uM TMZ.

Article Snippet: PLK1 protein expression was evaluated using LSBio antibody (diluted 1:200, PLK1 rabbit anti-Human polyclonal Antibody LS-B4225- LSBio LifeSpan Bioscience, Seattle, WA).

Techniques: Solvent, Control, Western Blot, Quantitative RT-PCR, Inhibition, Transfection

Journal: Oncotarget

Article Title: Disulfiram, a drug widely used to control alcoholism, suppresses self-renewal of glioblastoma and overrides resistance to temozolomide

doi:

Figure Lengend Snippet: (A) U251 cells were exposed to DSF for 24 hrs using DMSO as a solvent control. Protein and transcript levels of PLK1 were assessed using immunoblotting or qRT-PCR. (B-C) Inhibiting PLK1 with BI-2536 or siRNA inhibits their growth. The efficacy of PLK1 inhibition on U251 growth is exemplified in combination with 10 uM TMZ. (D) Likewise, DSF inhibits the growth of U251 cells in a dose-dependent manner using DMSO as a solvent control.

Article Snippet: PLK1 protein expression was evaluated using LSBio antibody (diluted 1:200, PLK1 rabbit anti-Human polyclonal Antibody LS-B4225- LSBio LifeSpan Bioscience, Seattle, WA).

Techniques: Solvent, Control, Western Blot, Quantitative RT-PCR, Inhibition

Journal: Oncotarget

Article Title: Disulfiram, a drug widely used to control alcoholism, suppresses self-renewal of glioblastoma and overrides resistance to temozolomide

doi:

Figure Lengend Snippet: (A) U251 cells have lower PLK1 transcript expression than TMZ resistant SF188 cells. U251 cells were treated with 10 uM TMZ every 2 days for a total of 7 days and (B) partial TMZ sensitivity is demonstrated in a growth assay. (C) Immunoblot demonstrating an increase in PLK1 protein levels in TMZ treated U251 cells compared to untreated and DMSO after 24 hours. Actin is used as a loading control protein. (D) The surviving TMZ resistant cells were re-plated and treated with increasing concentrations of BI-2536 for 5 days.

Article Snippet: PLK1 protein expression was evaluated using LSBio antibody (diluted 1:200, PLK1 rabbit anti-Human polyclonal Antibody LS-B4225- LSBio LifeSpan Bioscience, Seattle, WA).

Techniques: Expressing, Growth Assay, Western Blot, Control

Journal: Oncotarget

Article Title: Disulfiram, a drug widely used to control alcoholism, suppresses self-renewal of glioblastoma and overrides resistance to temozolomide

doi:

Figure Lengend Snippet: (A) BT74 cells are resistant to TMZ yet sensitive to PLK1 inhibition with BI-2536. (B) BT241 cells are a second example to which the cells are TMZ resistant yet sensitive to PLK1 inhibition. Both models are maintained as primary isolates and only cultured as neurospheres. The combination of TMZ and BI-2536 did not further improve self-renewal inhibition. Scale bar = 500 um.

Article Snippet: PLK1 protein expression was evaluated using LSBio antibody (diluted 1:200, PLK1 rabbit anti-Human polyclonal Antibody LS-B4225- LSBio LifeSpan Bioscience, Seattle, WA).

Techniques: Inhibition, Cell Culture

Journal: Oncotarget

Article Title: Disulfiram, a drug widely used to control alcoholism, suppresses self-renewal of glioblastoma and overrides resistance to temozolomide

doi:

Figure Lengend Snippet: (A) PLK1 levels were assessed in aBT001 and aBT003 by immunostaining. Both cases express high levels of PLK1. (B-C) The PLK1 inhibitor BI-2536 suppressed the growth of aBT001 and induced cell death.

Article Snippet: PLK1 protein expression was evaluated using LSBio antibody (diluted 1:200, PLK1 rabbit anti-Human polyclonal Antibody LS-B4225- LSBio LifeSpan Bioscience, Seattle, WA).

Techniques: Immunostaining