scholarly journals Expression of HDACs 1, 3 and 8 Is Upregulated in the Presence of Infiltrating Lymphocytes in Uveal Melanoma

Cancers ◽  
2021 ◽  
Vol 13 (16) ◽  
pp. 4146
Author(s):  
Zahra Souri ◽  
Aart G. Jochemsen ◽  
Annemijn P. A. Wierenga ◽  
Wilma G. M. Kroes ◽  
Rob M. Verdijk ◽  
...  
Keyword(s):  
Cell Lines ◽  
Uveal Melanoma ◽  
Histone Deacetylases ◽  
Cultured Cells ◽  
Chromosome 3 ◽  
Inflammatory Phenotype ◽  
Positive Correlations ◽  
Infiltrating Lymphocytes ◽  
Lymphocyte Fraction ◽  
Expression Loss

In Uveal Melanoma (UM), an inflammatory phenotype is strongly associated with the development of metastases and with chromosome 3/BAP1 expression loss. As an increased expression of several Histone Deacetylases (HDACs) was associated with loss of chromosome 3, this suggested that HDAC expression might also be related to inflammation. We analyzed HDAC expression and the presence of leukocytes by mRNA expression in two sets of UM (Leiden and TCGA) and determined the T lymphocyte fraction through ddPCR. Four UM cell lines were treated with IFNγ (50IU, 200IU). Quantitative PCR (qPCR) was used for mRNA measurement of HDACs in cultured cells. In both cohorts (Leiden and TCGA), a positive correlation occurred between expression of HDACs 1, 3 and 8 and the presence of a T-cell infiltrate, while expression of HDACs 2 and 11 was negatively correlated with the presence of tumor-infiltrating macrophages. Stimulation of UM cell lines with IFNγ induced an increase in HDACs 1, 4, 5, 7 and 8 in two out of four UM cell lines. We conclude that the observed positive correlations between HDAC expression and chromosome 3/BAP1 loss may be related to the presence of infiltrating T cells.

scholarly journals Monosomy 3 Is Linked to Resistance to MEK Inhibitors in Uveal Melanoma

2021 ◽  
Vol 22 (13) ◽  
pp. 6727
Author(s):  
Svenja Mergener ◽  
Jens T. Siveke ◽  
Samuel Peña-Llopis
Keyword(s):  
Cell Lines ◽  
Uveal Melanoma ◽  
Survival Data ◽  
Translation Initiation Factor ◽  
The Cancer Genome Atlas ◽  
Initiation Factor ◽  
Chromosome 3 ◽  
Mek Inhibitors ◽  
Mek Inhibition ◽  
Insight Into

The use of MEK inhibitors in the therapy of uveal melanoma (UM) has been investigated widely but has failed to show benefits in clinical trials due to fast acquisition of resistance. In this study, we investigated a variety of therapeutic compounds in primary-derived uveal melanoma cell lines and found monosomy of chromosome 3 (M3) and mutations in BAP1 to be associated with higher resistance to MEK inhibition. However, reconstitution of BAP1 in a BAP1-deficient UM cell line was unable to restore sensitivity to MEK inhibition. We then compared UM tumors from The Cancer Genome Atlas (TCGA) with mutations in BAP1 with tumors with wild-type BAP1. Principal component analysis (PCA) clearly differentiated both groups of tumors, which displayed disparate overall and progression-free survival data. Further analysis provided insight into differential expression of genes involved in signaling pathways, suggesting that the downregulation of the eukaryotic translation initiation factor 2A (EIF2A) observed in UM tumors with BAP1 mutations and M3 UM cell lines might lead to a decrease in ribosome biogenesis while inducing an adaptive response to stress. Taken together, our study links loss of chromosome 3 with decreased sensitivity to MEK inhibition and gives insight into possible related mechanisms, whose understanding is fundamental to overcome resistance in this aggressive tumor.

Effect of Heterogeneous Distribution of Monosomy 3 on Prognosis in Uveal Melanoma

2011 ◽  
Vol 135 (8) ◽  
pp. 1042-1047 ◽  
Author(s):  
Inge HG Bronkhorst ◽  
Willem Maat ◽  
Ekaterina S Jordanova ◽  
Wilma GM Kroes ◽  
Nicoline E Schalij-Delfos ◽  
...  
Keyword(s):  
Uveal Melanoma ◽  
Survival Data ◽  
Cultured Cells ◽  
Cox Proportional Hazards ◽  
Chromosome 3 ◽  
Likelihood Ratios ◽  
Heterogeneous Distribution ◽  
Isolated Nuclei ◽  
Monosomy 3 ◽  
Significant Difference

Context.—Fluorescence in situ hybridization (FISH) analyses on tumor sections and on isolated nuclei showed that even low numbers of cells with monosomy of chromosome 3 adversely affected survival. Objective.—To determine what percentage of uveal melanoma cells with monosomy of chromosome 3 influences patient mortality. Design.—To determine the presence of monosomy 3, karyotyping and FISH on cultured cells and FISH on isolated nuclei were performed on 50 primary uveal melanomas. Clinical and pathologic prognostic factors were assessed and compared with 5-year survival data. Analyses were performed using Cox proportional hazards test, log-rank analysis, sensitivity, specificity, and positive and negative likelihood ratios. Results.—Combined karyotyping and FISH on cultured cells showed monosomy 3 in 19 of 50 cases (38%), whereas determination of the monosomy 3 status by FISH on isolated nuclei with a threshold of 5% assigned 31 of 50 cases (62%) to the monosomy-3 category. When monosomy 3 on isolated nuclei with a cutoff value of 5% was used, a significant difference in 5-year survival was present (hazard ratio, 15.5; P  =  .007), indicating that monosomy 3 in greater than 5% of tumor cells is related to death due to metastases. Conclusion.—In uveal melanoma, the presence of greater than 5% of cells with monosomy 3, as determined by FISH on isolated nuclei, is associated with the development of metastases within 5 years after enucleation.

Monosomy of Chromosome 3 and an Inflammatory Phenotype Occur Together in Uveal Melanoma

2008 ◽  
Vol 49 (2) ◽  
pp. 505 ◽  
Author(s):  
Willem Maat ◽  
Long V. Ly ◽  
Ekaterina S. Jordanova ◽  
Didi de Wolff-Rouendaal ◽  
Nicoline E. Schalij-Delfos ◽  
...  
Keyword(s):  
Uveal Melanoma ◽  
Chromosome 3 ◽  
Inflammatory Phenotype

Establishment and characterization of two uveal melanoma cell lines derived from tumors with loss of one chromosome 3

2006 ◽  
Vol 83 (4) ◽  
pp. 858-864 ◽  
Author(s):  
Gordon Nareyeck ◽  
Michael Zeschnigk ◽  
Gabriele Prescher ◽  
Dietmar R. Lohmann ◽  
Gerasimos Anastassiou
Keyword(s):  
Cell Lines ◽  
Uveal Melanoma ◽  
Melanoma Cell ◽  
Chromosome 3 ◽  
Uveal Melanoma Cell ◽  
Melanoma Cell Lines

scholarly journals MiRNAs Correlate with HLA Expression in Uveal Melanoma: Both Up- and Downregulation Are Related to Monosomy 3

Cancers ◽  
2021 ◽  
Vol 13 (16) ◽  
pp. 4020
Author(s):  
Zahra Souri ◽  
Annemijn P. A. Wierenga ◽  
Emine Kiliç ◽  
Erwin Brosens ◽  
Stefan Böhringer ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Uveal Melanoma ◽  
Immune Cell ◽  
Hla Class I ◽  
Class I ◽  
Chromosome 3 ◽  
Data Set ◽  
Monosomy 3 ◽  
Genetic Status ◽  
Infiltrating Lymphocytes

MicroRNAs are known to play a role in the regulation of inflammation. As a high HLA Class I expression is associated with a bad prognosis in UM, we set out to determine whether any miRNAs were related to a high HLA Class I expression and inflammation. We also determined whether such miRNAs were related to the UM’s genetic status. The expression of 125 miRNAs was determined in 64 primary UM from Leiden. Similarly, the mRNA expression of HLA-A, HLA-B, TAP1, BAP1, and immune cell markers was obtained. Expression levels of 24 of the 125 miRNAs correlated with expression of at least three out of four HLA Class I probes. Four miRNAs showed a positive correlation with HLA expression and infiltration with leukocytes, 20 a negative pattern. In the first group, high miRNA levels correlated with chromosome 3 loss/reduced BAP1 mRNA expression, in the second group low miRNA levels. The positive associations between miRNA-22 and miRNA-155 with HLA Class I were confirmed in the TCGA study and Rotterdam cohort, and with TAP1 in the Rotterdam data set; the negative associations between miRNA-125b2 and miRNA-211 and HLA-A, TAP1, and CD4 were confirmed in the Rotterdam set. We demonstrate two patterns: miRNAs can either be related to a high or a low HLA Class I/TAP1 expression and the presence of infiltrating lymphocytes and macrophages. However, both patterns were associated with chromosome 3/BAP1 status, which suggests a role for BAP1 loss in the regulation of HLA expression and inflammation in UM through miRNAs.

Apoptotic Effects of N-(2-hydroxyphenyl)-2-propylpentanamide on U87-MG and U-2 OS Cells and Antiangiogenic Properties

2020 ◽  
Vol 20 ◽  
Author(s):  
Paola Castillo-Juárez ◽  
Sebastián C. Sanchez ◽  
Alma D. Chávez-Blanco ◽  
Humberto Mendoza-Figueroa ◽  
José Correa-Basurto
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Cancer Cell ◽  
Histone Deacetylases ◽  
Biochemical Mechanism ◽  
Antiproliferative Effects ◽  
Human Osteosarcoma ◽  
Antiproliferative Agent ◽  
Induced Apoptosis ◽  
Apoptotic Effects

Background and Objective: Histone deacetylases (HDACs) are important therapeutic targets for many types of human cancers. A derivative of valproic acid, N-(2-hydroxyphenyl)-2-propylpentanamide (HO-AAVPA), has antiproliferative properties on some cancer cell lines and inhibits the HDAC1 isoform. Materials and Methods: In this work, HO-AAVPA was tested as an antiproliferative agent in U87-MG (human glioblastoma) and U-2 OS cells (human osteosarcoma), which are types of cancer that are difficult to treat, and its antiangiogenic properties were explored. Results: HO-AAVPA had antiproliferative effects at 48 h with an IC50 = 0.655 mM in U87-MG cells and an IC50 = 0.453 mM in U-2 OS cells. Additionally, in the colony formation assay, HO-AAVPA decreased the number of colonies by approximately 99% in both cell lines and induced apoptosis by 31.3% in the U-2 OS cell line and by 78.2% in the U87-MG cell line. Additionally, HO-AAVPA reduced the number of vessels in chorioallantoid membranes (CAMs) by approximately 67.74% and IL-6 levels in both cell lines suggesting that the biochemical mechanism on cancer cell of HO-AAVPA is different compared to VPA. Conclusion: HO-AAVPA has antiproliferative effects on glioblastoma and osteosarcoma and antiangiogenic properties.

Direct protein delivery into intact plant cells using polyhistidine peptides

2021 ◽  
Author(s):  
Yoshino Tanaka ◽  
Yoshihiko Nanasato ◽  
Kousei Omura ◽  
Keita Endoh ◽  
Tsuyoshi Kawano ◽  
...  
Keyword(s):  
Cell Lines ◽  
Fusion Proteins ◽  
Protein Delivery ◽  
Fluorescent Protein ◽  
Cultured Cells ◽  
Plant Cells ◽  
Adenosine Monophosphate ◽  
Cell Penetrating Peptides ◽  
Intracellular Space ◽  
Intracellular Fluorescence

Abstract Polyhistidine peptides (PHPs), sequences comprising only histidine residues (>His8), are effective cell-penetrating peptides for plant cells. Using PHP-fusion proteins, we aimed to deliver proteins into cultured plant cells from Nicotiana tabacum, Oryza sativa, and Cryptomeria japonica. Co-cultivation of cultured cells with fusion proteins combining maltose-binding protein (MBP), red fluorescent protein (RFP), and various PHPs (MBP-RFP-His8–His20) in one polypeptide showed the cellular uptake of fusion proteins in all plant cell lines. Maximum intracellular fluorescence was shown in MBP-RFP-His20. Further, adenylate cyclase (CyaA), a synthase of cyclic adenosine monophosphate (cAMP) activated by cytosolic calmodulin, was used as a reporter for protein delivery in living cells. A fusion protein combining MBP, RFP, CyaA, and His20 (MBP-RFP-CyaA-His20) was delivered into plant cells and increased intracellular fluorescence and cAMP production in all cell lines. The present study demonstrates that PHPs are effective carriers of proteins into the intracellular space of various cultured plant cells.

scholarly journals Structure-guided selection of puromycin N-acetyltransferase mutants with enhanced selection stringency for deriving mammalian cell lines expressing recombinant proteins

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Alessandro T. Caputo ◽  
Oliver M. Eder ◽  
Hana Bereznakova ◽  
Heleen Pothuis ◽  
Albert Ardevol ◽  
...  
Keyword(s):  
Cell Lines ◽  
Mammalian Cell ◽  
Recombinant Proteins ◽  
Cultured Cells ◽  
Hek293 Cells ◽  
Reaction Products ◽  
Enzyme Form ◽  
X Ray Crystallography ◽  
Mammalian Cell Lines ◽  
Apparent Loss

AbstractPuromycin and the Streptomyces alboniger-derived puromycin N-acetyltransferase (PAC) enzyme form a commonly used system for selecting stably transfected cultured cells. The crystal structure of PAC has been solved using X-ray crystallography, revealing it to be a member of the GCN5-related N-acetyltransferase (GNAT) family of acetyltransferases. Based on structures in complex with acetyl-CoA or the reaction products CoA and acetylated puromycin, four classes of mutations in and around the catalytic site were designed and tested for activity. Single-residue mutations were identified that displayed a range of enzymatic activities, from complete ablation to enhanced activity relative to wild-type (WT) PAC. Cell pools of stably transfected HEK293 cells derived using two PAC mutants with attenuated activity, Y30F and A142D, were found to secrete up to three-fold higher levels of a soluble, recombinant target protein than corresponding pools derived with the WT enzyme. A third mutant, Y171F, appeared to stabilise the intracellular turnover of PAC, resulting in an apparent loss of selection stringency. Our results indicate that the structure-guided manipulation of PAC function can be utilised to enhance selection stringency for the derivation of mammalian cell lines secreting elevated levels of recombinant proteins.

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