Small molecule MMRi62 was synthesized in house at Chemler’s lab. The Betti reaction was for MMRi62 synthesis as described in literatures (19 , 20 ). The structure and purity of the synthesized MMRi62 was determined by NMR and the purity was determined by high resolution mass spectra obtained at SUNY Buffalo’s mass spec facility on a ThermoFinnigan MAT XL spectrometer. Bafilomycin A1 was obtained from Sigma (Cat# 19–148) and Carfilzomib was purchased from Cayman Chemical (Cat# 17554). All the compounds were dissolved in DMSO as 10 mM stocks and stored at −20 °C.
Carfilzomib
Manufactured by Cayman Chemical
Carfilzomib is a proteasome inhibitor laboratory reagent manufactured by Cayman Chemical. It functions by inhibiting the proteasome, a large protein complex responsible for the degradation of unwanted or damaged proteins within cells.
Automatically generated - may contain errors
Lab products found in correlation
Bortezomib, by Cayman Chemical (2 mentions)
Bafilomycin a1, by Merck Group (1 mentions)
Ym155, by Selleck Chemicals (1 mentions)
4 hydroxytamoxifen, by Merck Group (1 mentions)
Antimycin, by Merck Group (1 mentions)
Sulforaphane, by Merck Group (1 mentions)
Oligomycin, by Merck Group (1 mentions)
Anti sting, by Cell Signaling Technology (1 mentions)
Anti cgas, by Cell Signaling Technology (1 mentions)
B 5 1 2, by Merck Group (1 mentions)
Phospho histone h2a x ser139 antibody, by Cell Signaling Technology (1 mentions)
Ab56416, by Abcam (1 mentions)
Ab18256, by Abcam (1 mentions)
Bx795, by InvivoGen (1 mentions)
Anti rig 1, by Cell Signaling Technology (1 mentions)
Phospho sting ser365 d8f4w rabbit mab, by Cell Signaling Technology (1 mentions)
Ruxolitinib, by Selleck Chemicals (1 mentions)
Bay11 7082, by Selleck Chemicals (1 mentions)
Actinomycin d, by Merck Group (1 mentions)
Cycloheximide (chx), by Merck Group (1 mentions)
4 protocols using carfilzomib
1
Synthesis and Characterization of MMRi62
2
Compound Sourcing for QMAP-Seq and Validation
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YM155 was obtained from Selleckchem (#S1130). Compounds for QMAP-Seq were obtained from the FDA-Approved Drug Library (MedChemExpress, #HY-L022), the Clinical Compound Library (MedChemExpress, #HY-L026), or from the following vendors: 4-Hydroxytamoxifen was obtained from Sigma-Aldrich (#H7904), and Bortezomib was obtained from Cayman Chemical (#10008822). For validation experiments, Carfilzomib was obtained from a different vendor (Cayman Chemical, #17554) than where it was sourced for QMAP-Seq.
3
Antibody Treatments for Immune Signaling
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BX795 was purchased from Invivogen. CAY10576, BAY 11–7082, and Ruxolitinib were from Selleck Chemicals. Bortezomib and carfilzomib were from Cayman Chemical. Sulforaphane, cycloheximide, actinomycin D, oligomycin, and antimycin were obtained from Sigma-Aldrich. Mouse monoclonal Ngly1 antibody was a gift from A. Zuberi (Jackson Laboratories). Other antibodies used as follows: anti-STING (13647; Cell Signaling, 1:1,000 dilution), phospho-STING (Ser365; D8F4W) Rabbit mAb (72971; Cell Signaling; 1:1,000 dilution), anti-cGAS (31659; Cell Signaling; 1:1,000 dilution), anti–RIG-I (3743; Cell Signaling; 1:1,000 dilution), anti-MDA5 (5321; Cell Signaling; 1:1,000 dilution), anti-tubulin (B-5-1-2; Sigma-Aldrich; 1:20,000 dilution), anti-HMGB1 (ab18256; Abcam; 1:2,000 dilution), anti-GAPDH (2118; Cell Signaling; 1:2,000 dilution), anti-SQSTM1/p62 (ab56416; Abcam; 1:1,000 dilution), anti-phosphorylated p70 S6K (Thr389; 9205; Cell Signaling; 1:1,000 dilution), anti-p70 S6K (2708; Cell Signaling; 1:1,000 dilution), anti–phospho-ATM (Ser1981) antibody (clone 10H11.E12; EMD Millipore; 1:1,000 dilution), phospho-Histone H2A.X (Ser139) antibody (2577; Cell Signaling; 1:1,000 dilution), Nrf1 (8052; Cell Signaling; 1:1,000 dilution), Nrf2 (12721; Cell Signaling; 1:1,000 dilution).
4
Measuring Proteasome Peptidase Activity
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Proteasome peptidase activity in sciatic nerve lysates was measured as described previously (VerPlank et al, 2018 (link)). For chymotrypsin-like peptidase activity, 20 μM Suc-LLVY-amc (Enzo) was used, and for caspase-like peptidase activity, 20 μM AC-nLPnLD-amc (Enzo) was used. Proteasome-specific cleavage was calculated by subtracting the overall rate of peptide hydrolysis in the lysates by the rate of hydrolysis measured in the presence of 5 μM Carfilzomib (Cayman Chemical Company).
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