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Archives of Dermatological Research Oct 2017SAHA (vorinostat) is a histone deacetylase inhibitor approved by the USA Food and Drug Administration (FDA) for treating advanced refractory cutaneous T cell lymphomas....
SAHA (vorinostat) is a histone deacetylase inhibitor approved by the USA Food and Drug Administration (FDA) for treating advanced refractory cutaneous T cell lymphomas. As SAHA alters the expression of many genes under control of the Sp1 transcription factor, we examined the effect of its association with the FDA-approved anticancer antibiotic Mithramycin A (MTR, plicamycin), a competitive inhibitor of Sp1 binding to DNA. Sézary syndrome (SS) cells, expanded ex vivo from peripheral blood mononuclear cells of 4 patients, were tested for their sensitivity to the drugs regarding cytotoxicity and differential responsive gene expression. Multivariate statistical methods were used to identify genes whose expression is altered by SAHA, MTR, and the synergist effect of the two drugs. MTR, like SAHA, induced the apoptosis of SS cells, while the two drugs in combination showed clear synergy or potentiation. Expression data stressed a likely important role of additive or synergistic epigenetic modifications in the combined effect of the two drugs, while direct inhibition of Sp1-dependent transcription seemed to have only limited impact. Ontological analysis of modified gene expression suggested that the two drugs, either independently or synergistically, counteracted many intertwined pro-survival pathways deregulated in SS cells, resistance of these tumors to intrinsic and extrinsic apoptosis, abnormal adhesion migration, and invasive properties, as well as immunosuppressive behavior. Our findings provide preliminary clues on the individual and combined effects of SAHA and MTR in SS cells and highlight a potential therapeutic interest of this novel pair of drugs for treatment of SS patients.
Topics: Antineoplastic Agents; Cell Line, Tumor; Drug Therapy, Combination; Gene Expression Regulation, Neoplastic; Humans; Hydroxamic Acids; Plicamycin; Sezary Syndrome; Skin Neoplasms; Transcriptome; Vorinostat
PubMed: 28695331
DOI: 10.1007/s00403-017-1761-0 -
Cytometry Jul 1980The feasibility of flow cytometry for measurements on bacteria has been demonstrated by measurements of DNA-associated fluorescence of Escherichia coli K-12 in various...
The feasibility of flow cytometry for measurements on bacteria has been demonstrated by measurements of DNA-associated fluorescence of Escherichia coli K-12 in various phases of cell growth. Bacteria were stained with a combination of ethidium bromide and mithramycin after fixation in 70% ethanol. Cultures grown to stationary phase accumulated in two peaks representing cells with two and four chromosomes. Qualitatively similar histograms were obtained with cells grown in the presence of chloramphenicol, whereas cells of the temperature sensitive strain E 177 (dnaA) ended up with only one chromosome per cell at the restrictive temperature. The fluorescence intensity of cells with one chromosome was about 10(3) times smaller than that of human diploid cells. Instrumental resolution at this level of intensity was CV = 5%, whereas peak widths corresponded to CV = 7-8%. Dyes bound to RNA did not appear to contribute significantly to the fluorescence.
Topics: Bacteriological Techniques; Chloramphenicol; Chromosomes, Bacterial; DNA Replication; DNA, Bacterial; Escherichia coli; Flow Cytometry; Interphase; Mutation; Temperature
PubMed: 7023880
DOI: 10.1002/cyto.990010108 -
British Journal of Cancer Nov 1973One hundred and seven untreated patients with acute myelogenous leukaemia (AML) were admitted to St Bartholomew's Hospital between 10 October 1970 and 31 January 1973.... (Clinical Trial)
Clinical Trial Randomized Controlled Trial
One hundred and seven untreated patients with acute myelogenous leukaemia (AML) were admitted to St Bartholomew's Hospital between 10 October 1970 and 31 January 1973. Before receiving drugs to induce remission they were allocated alternatively into 2 groups to decide their remission treatment-a group to receive chemotherapy alone and a group to receive the same chemotherapy with immunotherapy. The patients were then given induction chemotherapy and 45 of them attained complete remission. All patients in remission then received chemotherapy consisting of 5 days treatment every 28 days. Patients receiving immunotherapy were also given multiple weekly intradermal injections of irradiated stored AML cells and Glaxo B.C.G. using a Heaf gun. There were 19 patients in the group which received only chemotherapy during remission; 7 of these patients remain alive (median survival after attaining remission 303 days) and only 5 are still in their first remission (median remission length 188 days). Twenty-three patients were allocated to receive immunotherapy during remission in addition to chemotherapy and 16 remain alive (median 545 days) and 8 are in their first remission (median 312 days). The difference in survival of the two groups is significant with a P value of 0·003.
Topics: Adolescent; Adult; BCG Vaccine; Cytarabine; Daunorubicin; Female; Histocompatibility Testing; Humans; Immunity; Immunotherapy; Injections, Intradermal; Leukemia, Myeloid, Acute; Lymphocyte Activation; Lymphocytes; Male; Middle Aged; Plicamycin; Remission, Spontaneous; Statistics as Topic; Time Factors
PubMed: 4271320
DOI: 10.1038/bjc.1973.162 -
Experimental Cell Research Dec 2023The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is causative of the ongoing coronavirus disease 2019 (COVID-19) pandemic. The SARS-CoV-2 Spike protein...
The anti-SARS-CoV-2 BNT162b2 vaccine suppresses mithramycin-induced erythroid differentiation and expression of embryo-fetal globin genes in human erythroleukemia K562 cells.
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is causative of the ongoing coronavirus disease 2019 (COVID-19) pandemic. The SARS-CoV-2 Spike protein (S-protein) plays an important role in the early phase of SARS-CoV-2 infection through efficient interaction with ACE2. The S-protein is produced by RNA-based COVID-19 vaccines, that were fundamental for the reduction of the viral spread within the population and the clinical severity of COVID-19. However, the S-protein has been hypothesized to be responsible for damaging cells of several tissues and for some important side effects of RNA-based COVID-19 vaccines. Considering the impact of COVID-19 and SARS-CoV-2 infection on the hematopoietic system, the aim of this study was to verify the effect of the BNT162b2 vaccine on erythroid differentiation of the human K562 cell line, that has been in the past intensively studied as a model system mimicking some steps of erythropoiesis. In this context, we focused on hemoglobin production and induced expression of embryo-fetal globin genes, that are among the most important features of K562 erythroid differentiation. We found that the BNT162b2 vaccine suppresses mithramycin-induced erythroid differentiation of K562 cells. Reverse-transcription-qPCR and Western blotting assays demonstrated that suppression of erythroid differentiation was associated with sharp inhibition of the expression of α-globin and γ-globin mRNA accumulation. Inhibition of accumulation of ζ-globin and ε-globin mRNAs was also observed. In addition, we provide in silico studies suggesting a direct interaction between SARS-CoV-2 Spike protein and Hb Portland, that is the major hemoglobin produced by K562 cells. This study thus provides information suggesting the need of great attention on possible alteration of hematopoietic parameters following SARS-CoV-2 infection and/or COVID-19 vaccination.
Topics: Humans; K562 Cells; Plicamycin; COVID-19 Vaccines; BNT162 Vaccine; Leukemia, Erythroblastic, Acute; COVID-19; SARS-CoV-2; Hemoglobins; RNA, Messenger; Erythroid Cells
PubMed: 37944576
DOI: 10.1016/j.yexcr.2023.113853 -
Cancer Research Feb 2010Mithramycin (MIT) and tolfenamic acid (TA) inhibit the activity of the transcription factor Sp1. In the present study, we investigated whether pancreatic cancer...
Mithramycin (MIT) and tolfenamic acid (TA) inhibit the activity of the transcription factor Sp1. In the present study, we investigated whether pancreatic cancer treatment with a combination of these compounds has a synergistic effect on Sp1 activity, tumor growth, and their underlying response mechanisms. Treatment of pancreatic tumor xenografts with MIT and TA produced dose-dependent antitumor activity, and significant antitumor activity of either compound alone was directly associated with systemic side effects. Combination treatment with nontoxic doses of both compounds produced synergistic antitumor activity, whereas treatment with a nontoxic dose of either compound alone lacked a discernible antitumor effect. Synergistic therapeutic effects correlated directly with synergistic antiproliferation and antiangiogenesis in vitro. Moreover, combination treatment resulted in Sp1 protein degradation, drastically downregulating expression of Sp1 and vascular endothelial growth factor. Our findings established that Sp1 is a critical target of TA and MIT in human pancreatic cancer therapy, rationalizing clinical studies to determine the effect of existing pancreatic cancer therapy regimens on Sp1 signaling in tumors and normal pancreatic tissue, and the ability of Sp1-targeting strategies to modify cancer responses.
Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Blotting, Western; Body Weight; Cell Line, Tumor; Chromatin Immunoprecipitation; Dose-Response Relationship, Drug; Drug Synergism; Female; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Pancreatic Neoplasms; Plicamycin; Promoter Regions, Genetic; Protein Binding; Sp1 Transcription Factor; Tumor Burden; Vascular Endothelial Growth Factor A; Xenograft Model Antitumor Assays; ortho-Aminobenzoates
PubMed: 20086170
DOI: 10.1158/0008-5472.CAN-09-3282 -
Tumour Biology : the Journal of the... Jun 2017Pancreatic cancer is one of the most aggressive and difficult to treat cancers. Experimental and clinical evidence suggests that high basal state autophagy in pancreatic...
Pancreatic cancer is one of the most aggressive and difficult to treat cancers. Experimental and clinical evidence suggests that high basal state autophagy in pancreatic tumors could induce resistance to chemotherapy. Recently, we have demonstrated that penfluridol suppresses pancreatic tumor growth by autophagy-mediated apoptosis both in vitro and in vivo; however, the mechanism of autophagy induction by penfluridol was not clear. Several studies have established that endoplasmic reticulum stress could lead to autophagy and inhibit tumor progression. In this study, we demonstrated that penfluridol induced endoplasmic reticulum stress in BxPC-3, AsPC-1, and Panc-1 pancreatic cancer cell lines as indicated by upregulation of endoplasmic reticulum stress markers such as binding protein (BIP), C/EBP homologous protein (CHOP) and inositol requiring 1α (IRE1α) after treatment with penfluridol in a concentration-dependent manner. Inhibiting endoplasmic reticulum stress by pretreatment with pharmacological inhibitors such as sodium phenylbutyrate and mithramycin or by silencing CHOP using CHOP small interfering RNA, blocked penfluridol-induced autophagy. These results clearly indicate that penfluridol-induced endoplasmic reticulum stress lead to autophagy in our model. Western blot analysis of subcutaneously implanted AsPC-1 and BxPC-3 tumors as well as orthotopically implanted Panc-1 tumors demonstrated upregulation of BIP, CHOP, and IRE1α expression in the tumor lysates from penfluridol-treated mice as compared to tumors from control mice. Altogether, our study establishes that penfluridol-induced endoplasmic reticulum stress leads to autophagy resulting in reduced pancreatic tumor growth. Our study opens a new therapeutic target for advanced chemotherapies against pancreatic cancer.
Topics: Animals; Apoptosis; Autophagy; Cell Line, Tumor; Cell Proliferation; Endoplasmic Reticulum; Endoplasmic Reticulum Chaperone BiP; Endoplasmic Reticulum Stress; Endoribonucleases; Gene Expression Regulation, Neoplastic; Heat-Shock Proteins; Humans; Mice; Pancreatic Neoplasms; Penfluridol; Protein Serine-Threonine Kinases; RNA, Small Interfering; Transcription Factor CHOP; Xenograft Model Antitumor Assays
PubMed: 28618969
DOI: 10.1177/1010428317705517 -
Chemico-biological Interactions Aug 2014DIG-MSK (demycarosyl-3D-β-D-digitoxosyl-mithramycin SK) is a recently isolated compound of the mithramycin family of antitumor antibiotics, which includes mithramycin A...
DIG-MSK (demycarosyl-3D-β-D-digitoxosyl-mithramycin SK) is a recently isolated compound of the mithramycin family of antitumor antibiotics, which includes mithramycin A (MTA) and mithramycin SK (MSK). Here, we present evidence that the binding of DIG-MSK to DNA shares the general features of other mithramycins such as the preference for C/G-rich tracts, but there are some differences in the strength of binding and the DNA sequence preferentially recognized by DIG-MSK. We aimed at gaining further insights into the DIG-MSK mechanism of action by direct comparison with the effects of the parental MTA. Similar to MTA, MSK and DIG-MSK accumulated rapidly in A2780, IGROV1 and OVCAR3 human ovarian cancer cell lines, and DIG-MSK was a potent inhibitor of both basal and induced expression of an Sp1-driven luciferase vector. This inhibitory activity was confirmed for the endogenous Sp1 gene and a set of Sp-responsive genes, and compared to that of MTA and MSK. Furthermore, DIG-MSK was stronger than MTA as inhibitor of Sp3-driven transcription and endogenous Sp3 gene expression. Differences in the effects of MTA, MSK and DIG-MSK on gene expression may have a large influence on their biological activities.
Topics: Antibiotics, Antineoplastic; Binding Sites; Cell Line, Tumor; Female; GPI-Linked Proteins; Gene Expression Regulation, Neoplastic; Humans; Intracellular Signaling Peptides and Proteins; Kinetics; Ovarian Neoplasms; Plicamycin; RNA; Real-Time Polymerase Chain Reaction; Sp1 Transcription Factor; Spectrometry, Fluorescence; Transcription, Genetic
PubMed: 24907531
DOI: 10.1016/j.cbi.2014.05.019 -
International Journal of Radiation... Feb 2018Specificity protein 1 (SP1) is involved in the transcription of several genes implicated in tumor maintenance. We investigated the effects of mithramycin A (MTA), an...
PURPOSE
Specificity protein 1 (SP1) is involved in the transcription of several genes implicated in tumor maintenance. We investigated the effects of mithramycin A (MTA), an inhibitor of SP1 DNA binding, on radiation response.
METHODS AND MATERIALS
Clonogenic survival after irradiation was assessed in 2 tumor cell lines (A549, UM-UC-3) and 1 human fibroblast line (BJ) after SP1 knockdown or MTA treatment. DNA damage repair was evaluated using γH2AX foci formation, and mitotic catastrophe was assessed using nuclear morphology. Gene expression was evaluated using polymerase chain reaction arrays. In vivo tumor growth delay was used to evaluate the effects of MTA on radiosensitivity.
RESULTS
Targeting of SP1 with small interfering RNA or MTA sensitized A549 and UM-UC-3 to irradiation, with no effect on the BJ radiation response. MTA did not alter γH2AX foci formation after irradiation in tumor cells but did enhance mitotic catastrophe. Treatment with MTA suppressed transcription of genes involved in cell death. MTA administration to mice bearing A549 and UM-UC-3 xenografts enhanced radiation-induced tumor growth delay.
CONCLUSIONS
These results support SP1 as a target for radiation sensitization and confirm MTA as a radiation sensitizer in human tumor models.
Topics: Animals; Apoptosis; Cell Cycle; Cell Line, Tumor; DNA Damage; DNA Repair; Humans; Mice; Mitosis; Neoplasms, Experimental; Plicamycin; Radiation-Sensitizing Agents; Sp1 Transcription Factor
PubMed: 29157749
DOI: 10.1016/j.ijrobp.2017.09.049 -
PloS One 2015Expression and regulation of microRNAs is an emerging issue in erythroid differentiation and globin gene expression in hemoglobin disorders. In the first part of this...
Increase of microRNA-210, decrease of raptor gene expression and alteration of mammalian target of rapamycin regulated proteins following mithramycin treatment of human erythroid cells.
Expression and regulation of microRNAs is an emerging issue in erythroid differentiation and globin gene expression in hemoglobin disorders. In the first part of this study microarray analysis was performed both in mithramycin-induced K562 cells and erythroid precursors from healthy subjects or β-thalassemia patients producing low or high levels of fetal hemoglobin. We demonstrated that: (a) microRNA-210 expression is higher in erythroid precursors from β-thalassemia patients with high production of fetal hemoglobin; (b) microRNA-210 increases as a consequence of mithramycin treatment of K562 cells and human erythroid progenitors both from healthy and β-thalassemia subjects; (c) this increase is associated with erythroid induction and elevated expression of γ-globin genes; (d) an anti-microRNA against microRNA-210 interferes with the mithramycin-induced changes of gene expression. In the second part of the study we have obtained convergent evidences suggesting raptor mRNA as a putative target of microRNA-210. Indeed, microRNA-210 binding sites of its 3'-UTR region were involved in expression and are targets of microRNA-210-mediated modulation in a luciferase reporter assays. Furthermore, (i) raptor mRNA and protein are down-regulated upon mithramycin-induction both in K562 cells and erythroid progenitors from healthy and β-thalassemia subjects. In addition, (ii) administration of anti-microRNA-210 to K562 cells decreased endogenous microRNA-210 and increased raptor mRNA and protein expression. Finally, (iii) treatment of K562 cells with premicroRNA-210 led to a decrease of raptor mRNA and protein. In conclusion, microRNA-210 and raptor are involved in mithramycin-mediated erythroid differentiation of K562 cells and participate to the fine-tuning and control of γ-globin gene expression in erythroid precursor cells.
Topics: Adaptor Proteins, Signal Transducing; Cell Differentiation; Erythroid Cells; Female; Gene Expression Regulation; Humans; K562 Cells; Male; MicroRNAs; Plicamycin; Regulatory-Associated Protein of mTOR; TOR Serine-Threonine Kinases; beta-Thalassemia; gamma-Globins
PubMed: 25849663
DOI: 10.1371/journal.pone.0121567 -
Translational Vision Science &... Dec 2023To investigate the function and mechanism of tumor protein p53 in pathological scarring after glaucoma filtration surgery (GFS) using human Tenon's fibroblasts (HTFs)...
PURPOSE
To investigate the function and mechanism of tumor protein p53 in pathological scarring after glaucoma filtration surgery (GFS) using human Tenon's fibroblasts (HTFs) and a rabbit GFS model.
METHODS
The expression of p53 in bleb scarring after GFS and transforming growth factor-β (TGF-β)-induced HTFs (myofibroblasts [MFs]) was examined by western blot and immunochemical analysis. The interaction between p53 and specificity protein 1 (Sp1) was investigated by immunoprecipitation. The role of p53 and Sp1 in the accumulation of collagen type I alpha 1 chain (COL1A1) and the migration of MFs was evaluated by western blot, quantitative real-time polymerase chain reaction (qRT-PCR), wound healing, and Transwell assay. The regulatory mechanisms among p53/Sp1 and miR-29b were detected via qRT-PCR, western blot, luciferase reporter assay, and chromatin immunoprecipitation assay. The therapeutic effect of mithramycin A, a specific inhibitor of Sp1, on scarring formation was evaluated in a rabbit GFS model.
RESULTS
p53 was upregulated in bleb scar tissue and MFs. p53 and Sp1 form a transcription factor complex that induces the accumulation of COL1A1 and promotes the migration of MFs through downregulation of miR-29b, a known suppressor of COL1A1. The p53/Sp1 axis inhibits miR-29b expression by the direct binding promoter of the miR-29b gene. Mithramycin A treatment attenuated bleb scar formation in vivo.
CONCLUSIONS
The p53/Sp1/miR-29b signaling pathway plays a critical role in bleb scar formation after GFS. This pathway could be targeted for therapeutic intervention of pathological scarring after GFS.
TRANSLATIONAL RELEVANCE
Our research indicates that inhibition of p53/Sp1/miR-29b is a promising therapeutic strategy for preventing post-GFS pathological scarring.
Topics: Animals; Humans; Rabbits; Cicatrix; Down-Regulation; MicroRNAs; Tumor Suppressor Protein p53; Glaucoma; Filtering Surgery; Sp1 Transcription Factor
PubMed: 38051266
DOI: 10.1167/tvst.12.12.5