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- Table of Contents
2 Citations 16 Q&As
Facts about Methylated-DNA--protein-cysteine methyltransferase.
This is a suicide reaction: the enzyme is irreversibly inactivated. .
Human | |
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Gene Name: | MGMT |
Uniprot: | P16455 |
Entrez: | 4255 |
Belongs to: |
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MGMT family |
6-O-methylguanine-DNA methyltransferase; EC 2.1.1.63; methylated-DNA--protein-cysteine methyltransferase; methylguanine-DNA methyltransferase; MGMT; O-6-methylguanine-DNA methyltransferase; O6-methylguanine-DNA methyltransferase; O-6-methylguanine-DNA-alkyltransferase
Mass (kDA):
21.646 kDA
Human | |
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Location: | 10q26.3 |
Sequence: | 10; NC_000010.11 (129467241..129770983) |
Nucleus.
The MGMT gene is an enzyme involved in the repair of DNA that is that is implicated in the chemoresistance of alkylating agents. This DNA marker may be used to stain tissues and test for MGMT alleles. This article will discuss the most effective uses for this marker. Below are some of the most commonly used uses for this marker. There are many other uses for this gene.
MGMT is a direct DNA repair protein and is a ligand-induced degradeable protein found in human cell lines. As it breaks down, it is controlled by various independent UPP subpathways. The mechanism behind degradation isn't fully understood, but some data suggest that it functions through a suicide mechanism. Another evidence suggests that it is degraded in multiple ways and covalent targeting C145 could be required to stop the degradation of fusion.
In vitro studies have revealed that MGMT reduces the activity of HR. Treatment with CDDP and O6BG decreased the number of NPC cells with HR signaling. Furthermore the inhibition of MGMT inhibited the creation of genomic products caused by the HR pathway. The MGMT plays a role in the HR pathway and inhibition of it can have therapeutic effects against BRCA1-mutated cells.
MGMT has been demonstrated to be deregulated in cells that are exposed to temozolomide (a form of chemotherapy). To determine if lomeguatrib blocks MGMT researchers have employed the same method as above however with one difference: the mutated hMGMT protein contains an extra Cys residue which is necessary for its function. To introduce mutations using a Q5 Site Directed Mutagenesis Kit (Q5SMK) was employed. The fusion protein was amplified using PCR.
In addition to protecting cells from the harmful effects of alkylating mutagens, the MGMT shields the genome from carcinogenesis. Specifically, it repairs mutagenic adducts via the transfer of an alkyl group from the guanine to the cysteine residue. The enzyme can only make one turnover. This article is not meant to replace medical advice given by a professional.
A decrease in CAR-surface expression in cells that are not transduced also associated to a loss of MGMT. The loss of expression of MGMT is defined as a percentage of 25% or more of tumor DNAs that are altered. The interesting thing is that tumor DNAs that contain unmethylated MGMT promoters were also considered. One patient had an initial tumor that was not methylated and its metastatic counterpart modified.
Both the ALKBH pathway (and MGMT) are important for chemoresistance in the face of alkylated agents. Both pathways repair DNA damage and their deficiency could be used as biological markers of chemotherapy response. However the MMR pathway is more extensively studied than the ALKBH pathway. However, both pathways contribute to the resistance to chemotherapeutic agents that is why they require both types of drugs.
MGMT and MMR are essential for tumor cells to resist the damage caused by alkylating agents. Tumor cells are more susceptible to these agents if MGMT or MMR aren't working. Tumor cells can also suddenly silence the MGMT promoter, which limits their ability to repair DNA. This could increase their susceptibility to alkylating agents.
In addition to being the MGMT promoter, MGMT has many other functions in the body. Hypomethylation of the MGMT promoter is among the primary mechanisms behind resistance to TMZ. Genetic changes also contribute to tumor recurrence. In a recent study, the patient was diagnosed with a fusion of the MGMT gene was treated with TMZ but relapsed shortly after TMZ treatment. MGMT genome rearrangements might be relevant to alkylating agent resistance.
MGMT plays a role in DNA methylation, and is involved in the chemoresistance of alkylating agents. We extracted the reads from previous publications using RNA-sequencing info. We then analysed the fusions using the STAR-fusion program 1.5.032. Fusions that involve MGMT are not considered when the partners' genes are not well-studied or have two paralog genes. MGMT was found to be a fusion in a significant portion of tumors.
DNA alkylators damage DNA by alkylating the base of guanine. This leads to mispairing of bases DNA breakage, cell cycle arrest. MGMT is also involved in DNA alkylation and the chemoresistance (or oral chemotherapeutic agent) to TMZ. MGMT promoter hypermethylation is the sole known biomarker for TMZ response in patients with glioblastoma. A subset of the recurrent gliomas has genomic changes that cause MGMT overexpression.
Inhibiting MGMT activity reduces the toxic effects of TMZ. However, MGMT plays a role in DNA repair, and this was observed in fusion cells that expressed MGMT. O6-BG blocks MGMT and causes the accumulation of 53BP1 foci as well as gH2AX foci. This is why MGMT is involved in the chemoresistance of TMZ.
When MGMT is detected in tumor tissue, it is detected by immunohistochemistry. NeoMarkers Clone MT3.1 was used to detect MGMT. Non-tumour markers (lymphocytes and endothelial cell DNA) were also used. For internal positive controls lymphocytes were used. Semi-quantitative scoring was based on the immunoreactivity of MGMT, with an average score of 5% and 3+ for negative cells.
Detection of methylated MGMT is typically performed on paraffin-fixed formalin-fixed tumor specimens. The surgical site from which the sample was taken could influence MGMT expression. A pathologist must assess all available tissue and select the tissue block that contains the most viable tumor cells. The pathologist will then mark the location of the involvement of the tumor by using an H&E-stained portion of tissue.
The cells stained with the MGMT marker could also be positive for CD45 or CD68. These markers have been shown to be mutated in ovarian carcinoma. The third type of marker, CD45 may be useful in identifying lung tumor cells. The antibody used to identify MGMT is called CD68. This study used antibodies from Novocastra (and DakoCytomation) respectively.
Double-labelling immunohistochemistry can be used to determine the methylation status for MGMT. PSQ allows scientists determine whether cancer cell nuclei have been affected by methylation and can detect MGMT protein expression within cancerous tissues. This method is highly sensitive and specific and has excellent interlaboratory agreement as well as PPV. This makes it a very useful tool for cancer diagnosis.
Two types of MGMT-positive cases were identified. In the first case the tumor that was MGMT-positive was identified in 24% of the cases, while the other five (28 percent) cases were MGMT-negative. The second case did not have MGMT-negative melanomas. Three (27 percent) of the cases showed unambiguous MGMT immunostaining. However, in a third case in which the tumor cell was methylated was positive for MGMT.
The methylation status for MGMT was established in the third instance by determining whether there was a homogeneous MGMT promor. However, one-third of brain metastases were not methylated MGMT promoter which suggests that brain metastases are an ideal target for alkylating agents. A cancer cell's homogeneity MGMT expression is also determined by its status in methylation.
The MGMT gene is commonly modified in tumors. The second MGMT gene allele found in LGGs is methylated. This hinders MGMT's ability to repair DNA and confers sensitivity to TMZ treatment. However, it is unclear whether MGMT status in LGGs can provide any information about prognosis. MGMT status alone cannot help in selecting the most effective treatment.
The MGMT gene promoter is methylated in about 50 percent of gliomas with grade IV commonly referred to as GBMs. Although these tumors share similar mitotic activity and monosomy of chromosome 10 the modification of the remaining MGMT allele completely hinders MGMT-mediated DNA repair. Thus, methylation on the remaining MGMT allele is a hallmark of these tumors.
CpG islands in the MGMT promoter can influence the process of methylation. This promoter region spans 97 CpG dinucleotides. In addition, transcription is adversely affected by the methylation of CpG sites. DNA must be isolated for the methylation test, which requires high-quality DNA. The DNA must be tested for bisulfite conversion in order to make sure that false positive results are eliminated.
A molecular analysis of 99 GBMs from 73 patients showed that 27% of tumors contained methylated MGMT promoters. Additionally, the interlaboratory concordance between qMSP and MSP was 61%. Although qMSP isn't a conclusive method however, it is an excellent first tool in this field. It is inexpensive and easy to comprehend.
MGMT has significant clinical implications in clinical practice. It is also an important target in the treatment of cancer. A molecular test to determine methylation status for MGMT promoter may improve the prognosis. MGMT methylation is a key biomarker of tumor responses to temozolomide chemotherapy. Implementing this tool in clinical practice has been a major challenge.
To determine MGMT alleles Pyrosequencing tests are widely available. Pyrosequencing is demonstrated by the QIAGEN MGMT Pyro Kit. Illumina 850K (EPIC) BeadChip microarrays may be utilized in clinical diagnostics. Pyrosequencing, however offers a quantitative single-CpG readout.
PMID: 2405387 by Tano K., et al. Isolation and structural characterization of a cDNA clone encoding the human DNA repair protein for O6-alkylguanine.
PMID: 2188979 by Rydberg B., et al. cDNA cloning and chromosomal assignment of the human O6- methylguanine-DNA methyltransferase. cDNA expression in Escherichia coli and gene expression in human cells.
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