Journal: Neoplasia (New York, N.Y.)

Article Title: Oral bioavailable ITRI-148 degrades androgen receptor variants and overcomes antiandrogen resistance in advanced prostate cancer

doi: 10.1016/j.neo.2025.101253

Figure Lengend Snippet: ITRI-148 induces degradation of both AR and AR-V7 proteins in CRPC cells. (A) Chemical structures of ITRI-148 and its inactive analog. (B) Expression of Flag-tagged AR-V7 in 293T cells following ITRI-148 treatment. Cells transiently expressing Flag-AR-V7 were pretreated with 50 μg/mL cycloheximide for 1 hour, then exposed to ITRI-148 at the indicated concentrations for 8 hours. AR-V7 expression was detected using an anti-Flag antibody. (C) Representative immunoblots showing AR protein levels in VCaP and CWR22Rv1 cells after 48 hours of ITRI-148 treatment. An N-terminal AR antibody was used to detect both full-length AR and LBD-truncated variants (AR-V(ΔLBD)) in CWR22Rv1 cells, while AR-V7 was detected using an AR-V7-specific antibody. DC₅₀ values were calculated from normalized band intensities, and Dₘₐₓ represents the maximal degradation achieved at 10 μM ITRI-148. (D) Immunofluorescence staining of AR after 24 or 48 hours of ITRI-148 treatment, detected using an N-terminal AR antibody. (E) Western blot analysis of AR and AR-V7 expression in cells treated with DMSO (mock), 10 μM ITRI-148, or the inactive analog. (F) Western blot analysis of ectopically expressed wild-type (WT) and mutant Myc-tagged AR in DU145 cells following 24 hours of treatment with 10 μM ITRI-148. Quantified AR levels are as indicated.

Article Snippet: CWR22Rv1, VCaP, DU145, HEK293T, LNCaP, and PC3 were obtained from ATCC; PNT2 from Sigma-Aldrich.

Techniques: Expressing, Western Blot, Immunofluorescence, Staining, Mutagenesis

Journal: Neoplasia (New York, N.Y.)

Article Title: Oral bioavailable ITRI-148 degrades androgen receptor variants and overcomes antiandrogen resistance in advanced prostate cancer

doi: 10.1016/j.neo.2025.101253

Figure Lengend Snippet: ITRI-148 induces AR degradation through a ubiquitin-proteasome-dependent mechanism. (A) Proximity ligation assay (PLA) detecting the physical interaction between AR and CRBN upon ITRI-148 treatment. 293T cells transiently expressing Flag-AR or Flag-AR∆LBD were treated with DMSO or ITRI-148 for 8 hours, followed by PLA detection using anti-Flag and anti-CRBN antibodies. (B) Ubiquitination of AR and AR∆LBD proteins. 293T cells transfected with Flag-AR or Flag-AR∆LBD and HA-Ub were incubated with ITRI-148 and MG132. AR proteins were immunoprecipitated using anti-Flag, and ubiquitination of the AR proteins was detected by western blotting with anti-HA. (C) Western blot of Flag-AR and Flag-AR∆LBD in DU145 cells treated with ITRI-148 alone or in combination with MG132 (5 μM), MLN4924 (0.5 mM), or lenalidomide (80 μM). Inhibitors were added for 8 or 10 hours after the 24-hour ITRI-148 treatment. AR expression was detected using anti-Flag antibody.

Article Snippet: CWR22Rv1, VCaP, DU145, HEK293T, LNCaP, and PC3 were obtained from ATCC; PNT2 from Sigma-Aldrich.

Techniques: Ubiquitin Proteomics, Proximity Ligation Assay, Expressing, Transfection, Incubation, Immunoprecipitation, Western Blot

Journal: Cell reports

Article Title: APP-Mediated Signaling Prevents Memory Decline in Alzheimer’s Disease Mouse Model

doi: 10.1016/j.celrep.2019.03.087

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rabbit polyclonal anti-APP-CTM1 Dr. Gopal Thinakaran ( Vetrivel et al., 2009 ) N/A Rabbit polyclonal anti-APP-CT11 Dr. Gopal Thinakaran ( Vetrivel et al., 2009 ) N/A Rabbit polyclonal anti-APP-NT452 Dr. Gopal Thinakaran ( Vetrivel et al., 2009 ) N/A Mouse monoclonal anti-N-terminal Aβ antibody (clone 3D6) Dr. Dale Schenk (Elan Corporation PLC2) ( Andrew et al. 2017 ) N/A Mouse monoclonal anti-flag (clone M2) Sigma Aldrich Cat#F3165; RRID:AB_259529 Rabbit polyclonal MAP2 Sigma-Aldrich Cat#M3696; RRID:AB_1840999 Mouse monoclonal GAPDH Abeam Cat#ab8245; RRID:AB_2107448 Mouse monoclonal anti-HA (clone 6E2) Cell Signaling Cat#2367; RRID:AB_10691311 Goat anti-mouse Alexa 488 Invitrogen Cat#A-11029; RRID:AB_2534088 Goat anti-mouse Alexa 555 Invitrogen Cat#A-21424; RRID:AB_141780 Goat anti-rabbit Alexa 555 Invitrogen Cat#A-21429; RRID:AB_141761 IRDye 680LT donkey anti-mouse LI-COR Biosciences Cat# 926–68022; RRID:AB_10715072 IRDye 680 donkey anti-rabbit LI-COR Biosciences Cat# 926–32223; RRID:AB_621845 IRDye 800CW donkey anti-mouse LI-COR Biosciences Cat# 926–32212; RRID:AB_621847 IRDye 800CW goat anti-rabbit LI-COR Biosciences Cat#827–08365; RRID:AB_01796098 Aβ end-specific sandwich ELISA using end-specific antibodies (2.1.3: Aβ 42; 13.1.1: Aβ 40 for capture) and Ab5-HRP (pan Aβ) for detection Dr. Todd Golde ( Chakrabarty et al., 2015 ) N/A Mouse monoclonal Aβ anti-B608 for capture and anti-B436 for detection of sandwich ELISA Dr. Steven Wagner ( Vetrivel et al., 2009 ) N/A Bacterial and Virus Strains rAAV2/8 caps id packaging Dr. Todd Golde ( Chakrabarty et al., 2013 ) N/A Chemicals, Peptides, and Recombinant Proteins γ-secretase inhibitor Compound E Dr. Todd Golde Mayo Clinic core facility ( Seiffert et al., 2000 ) N/A Cis-N-(2-phenylcyclopentyl) azacyclotridec-1-en-2-amine (MDL-12,330A) Enzo Life Science Cat# BML-CN 145–0005 cOmplete EDTA-free - EDTA-free protease inhibitor cocktail Roche/Sigma-Aldrich Cat# 04693132001 PhosSTOP EasyPack - phosphatase inhibitor mixture Roche/Sigma-Aldrich Cat# 4906845001 Vectashield mounting medium Vector Laboratories Cat#H-1000 Lipofectamine 2000 Invitrogen/ThermoFisher Cat#11668019 LipoD293TM SignaGen Laboratories Cat# SL100668 Streptavidin-agarose beads Pierce/ThermoFisher Cat#20347 G-agarose beads Pierce/ThermoFisher Cat#20398 Premium sulfo NHS-SS-biotin Pierce/ThermoFisher Cat#PG82077 Experimental Models: Cell Lines HeLa cells expressing APP-FL containing the Swedish mutation (APPSwe) Dr.

Techniques: Sandwich ELISA, Virus, Recombinant, Protease Inhibitor, Plasmid Preparation, Expressing, Mutagenesis, Transgenic Assay, Knock-Out, Software