Metformin directly binds the alarmin HMGB1 and inhibits its proinflammatory activity

Metformin is the first-line drug in the treatment of type 2 diabetes. In addition to its hypoglycemic effect, metformin has an anti-inflammatory function, but the precise mechanism promoting this activity remains unclear. High mobility group box 1 (HMGB1) is an alarmin that is released from necrotic cells and induces inflammatory responses by its cytokine-like activity and is, therefore, a target of anti-inflammatory therapies. Here we identified HMGB1 as a novel metformin-binding protein by affinity purification using a biotinylated metformin analogue. Metformin directly bound to the C-terminal acidic tail of HMGB1. Both in vitro and in vivo, metformin inhibited inflammatory responses induced by full-length HMGB1 but not by HMGB1 lacking the acidic tail. In an acetaminophen-induced acute liver injury model in which HMGB1 released from injured cells exacerbates the initial injury, metformin effectively reduced liver injury and had no additional inhibitory effects when the extracellular HMGB1 was blocked by anti-HMGB1-neutralizing antibody. In summary, we report for the first time that metformin suppresses inflammation by inhibiting the extracellular activity of HMGB1. Because HMGB1 plays a major role in inflammation, our results suggest possible new ways to manage HMGB1-induced inflammation

Munc13 - 4 ワ GTPガタ Rab27 ケツゴウ タンパクシツ デ アリ ケッショウバン ノウセン カリュウ ブンピ オ セイギョスル

京都大学0048新制・課程博士博士(医学)甲第12953号医博第3113号新制||医||950(附属図書館)UT51-2007-H226京都大学大学院医学研究科内科系専攻(主査)教授 前川 平, 教授 野間 昭典, 教授 長澤 丘司学位規則第4条第1項該当Doctor of Medical ScienceKyoto UniversityDA

Endogenous APOBEC3B Overexpression Constitutively Generates DNA Substitutions and Deletions in Myeloma Cells

多発性骨髄腫における遺伝子変異蓄積の分子メカニズムの一端を解明 --新たな治療標的としてのDNAシトシン脱アミノ化酵素の可能性--. 京都大学プレスリリース. 2019-05-17.Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) DNA cytosine deaminases have emerged as potential genomic mutators in various cancers. Multiple myeloma accumulates APOBEC signature mutations as it progresses; however, the mechanisms underlying APOBEC signature acquisition and its consequences remain elusive. In this study, we examined the significance and clinical impact of APOBEC3B (A3B) activity in multiple myeloma. Among APOBECs, only highly expressed A3B was associated with poor prognosis in myeloma patients, independent of other known poor prognostic factors. Quantitative PCR revealed that CD138-positive primary myeloma cells and myeloma cell lines exhibited remarkably high A3B expression levels. Interestingly, lentiviral A3B knockdown prevented the generation of deletion and loss-of-function mutations in exogenous DNA, whereas in control cells, these mutations accumulated with time. A3B knockdown also decreased the basal levels of γ-H2AX foci, suggesting that A3B promotes constitutive DNA double-strand breaks in myeloma cells. Importantly, among control shRNA-transduced cells, we observed the generation of clones that harboured diverse mutations in exogenous genes and several endogenous genes frequently mutated in myeloma, including TP53. Taken together, the results suggest that A3B constitutively mutates the tumour genome beyond the protection of the DNA repair system, which may lead to clonal evolution and genomic instability in myeloma

APOBEC3B reporter myeloma cell lines identify DNA damage response pathways leading to APOBEC3B expression

Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) DNA cytosine deaminase 3B (A3B) is a DNA editing enzyme which induces genomic DNA mutations in multiple myeloma and in various other cancers. APOBEC family proteins are highly homologous so it is especially difficult to investigate the biology of specifically A3B in cancer cells. To easily and comprehensively investigate A3B function in myeloma cells, we used CRISPR/Cas9 to generate A3B reporter cells that contain 3×FLAG tag and IRES-EGFP sequences integrated at the end of the A3B gene. These reporter cells stably express 3xFLAG tagged A3B and the reporter EGFP and this expression is enhanced by known stimuli, such as PMA. Conversely, shRNA knockdown of A3B decreased EGFP fluorescence and 3xFLAG tagged A3B protein levels. We screened a series of anticancer treatments using these cell lines and identified that most conventional therapies, such as antimetabolites or radiation, exacerbated endogenous A3B expression, but recent molecular targeted therapeutics, including bortezomib, lenalidomide and elotuzumab, did not. Furthermore, chemical inhibition of ATM, ATR and DNA-PK suppressed EGFP expression upon treatment with antimetabolites. These results suggest that DNA damage triggers A3B expression through ATM, ATR and DNA-PK signaling