scholarly journals Targeted Mass Spectrometry Enables Quantification of Novel Pharmacodynamic Biomarkers of ATM Kinase Inhibition

Cancers ◽  
2021 ◽  
Vol 13 (15) ◽  
pp. 3843
Author(s):  
Jeffrey R. Whiteaker ◽  
Tao Wang ◽  
Lei Zhao ◽  
Regine M. Schoenherr ◽  
Jacob J. Kennedy ◽  
...  
Keyword(s):  
Dna Damage ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Limit Of Quantification ◽  
Peripheral Blood Mononuclear ◽  
Atm Kinase ◽  
Support Mechanism ◽  
Radiation Induced ◽  
Pharmacodynamic Biomarkers ◽  
Preclinical And Clinical Studies

The ATM serine/threonine kinase (HGNC: ATM) is involved in initiation of repair of DNA double-stranded breaks, and ATM inhibitors are currently being tested as anti-cancer agents in clinical trials, where pharmacodynamic (PD) assays are crucial to help guide dose and scheduling and support mechanism of action studies. To identify and quantify PD biomarkers of ATM inhibition, we developed and analytically validated a 51-plex assay (DDR-2) quantifying protein expression and DNA damage-responsive phosphorylation. The median lower limit of quantification was 1.28 fmol, the linear range was over 3 orders of magnitude, the median inter-assay variability was 11% CV, and 86% of peptides were stable for storage prior to analysis. Use of the assay was demonstrated to quantify signaling following ionizing radiation-induced DNA damage in both immortalized lymphoblast cell lines and primary human peripheral blood mononuclear cells, identifying PD biomarkers for ATM inhibition to support preclinical and clinical studies.

scholarly journals Interaction of Deubiquitinase 2A-DUB/MYSM1 with DNA Repair and Replication Factors

2020 ◽  
Vol 21 (11) ◽  
pp. 3762
Author(s):  
Carsten Kroeger ◽  
Reinhild Roesler ◽  
Sebastian Wiese ◽  
Adelheid Hainzl ◽  
Martina Vanessa Gatzka
Keyword(s):  
Mass Spectrometry ◽  
Dna Damage ◽  
Dna Repair ◽  
Transcriptional Activation ◽  
Mononuclear Cells ◽  
Bone Marrow Cells ◽  
Human Peripheral Blood ◽  
Cell Cycle Control ◽  
Mass Spectrometry Data ◽  
Peripheral Blood Mononuclear

The deubiquitination of histone H2A on lysine 119 by 2A-DUB/MYSM1, BAP1, USP16, and other enzymes is required for key cellular processes, including transcriptional activation, apoptosis, and cell cycle control, during normal hematopoiesis and tissue development, and in tumor cells. Based on our finding that MYSM1 colocalizes with γH2AX foci in human peripheral blood mononuclear cells, leukemia cells, and melanoma cells upon induction of DNA double-strand breaks with topoisomerase inhibitor etoposide, we applied a mass spectrometry-based proteomics approach to identify novel 2A-DUB/MYSM1 interaction partners in DNA-damage responses. Differential display of MYSM1 binding proteins significantly enriched after exposure of 293T cells to etoposide revealed an interacting network of proteins involved in DNA damage and replication, including factors associated with poor melanoma outcome. In the context of increased DNA-damage in a variety of cell types in Mysm1-deficient mice, in bone marrow cells upon aging and in UV-exposed Mysm1-deficient skin, our current mass spectrometry data provide additional evidence for an interaction between MYSM1 and key DNA replication and repair factors, and indicate a potential function of 2A-DUB/MYSM1 in DNA repair processes.

DNA damage and methylation induced by organophosphate flame retardants: Tris(2-chloroethyl) phosphate and tris(1-chloro-2-propyl) phosphate in human peripheral blood mononuclear cells

2019 ◽  
Vol 38 (6) ◽  
pp. 724-733 ◽  
Author(s):  
Karol Bukowski ◽  
Daniel Wysokinski ◽  
Katarzyna Mokra ◽  
Katarzyna Wozniak
Keyword(s):  
Dna Methylation ◽  
Dna Damage ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Flame Retardants ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Human Organism ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

Phosphorus flame retardants are a group of chemicals that are used to slow or prevent the spread of fire. These compounds have been detected in different environments including human organism. In the present study, we have investigated DNA-damaging potential and effect on DNA methylation of tris(2-chloroethyl) phosphate (TCEP) and tris(1-chloro-2-propyl) phosphate (TCPP) in human peripheral blood mononuclear cells (PBMCs). In order to determine DNA damage and repair, the alkaline and neutral versions of the comet assay were used. The level of DNA methylation was determined with specific antibodies against methylated DNA. PBMCs were exposed to TCEP and TCPP at the concentrations in the range of 1–1000 µM for 24 h. We have observed that TCEP and TCPP induced DNA damage—DNA breaks and alkali-labile sites. All DNA damages were effectively repaired during 120-min repair incubation. The results have also shown that TCEP and TCPP decreased the level of DNA methylation in PBMCs. In the case of TCEP, this effect was observed at a very low concentration of 1 µM.

scholarly journals Cryptococcus neoformans and gattii promote DNA damage in human peripheral blood mononuclear cells

2017 ◽  
Vol 56 (3) ◽  
pp. 344-349 ◽  
Author(s):  
Jéssica Cristina Bilizario Noguerol Andrade ◽  
Mariana Gatto ◽  
Daniela Ramos Rodrigues ◽  
Ângela Maria Victoriano de Campos Soares ◽  
Sueli Aparecida Calvi
Keyword(s):  
Dna Damage ◽  
Cryptococcus Neoformans ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

DNA damage and methylation induced by glyphosate in human peripheral blood mononuclear cells ( in vitro study)

2017 ◽  
Vol 105 ◽  
pp. 93-98 ◽  
Author(s):  
Marta Kwiatkowska ◽  
Edyta Reszka ◽  
Katarzyna Woźniak ◽  
Ewa Jabłońska ◽  
Jaromir Michałowicz ◽  
...  
Keyword(s):  
Dna Damage ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
In Vitro Study ◽  
Vitro Study ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells
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