| Abstract |
The main role of the DNA repair mechanism is to protect genetic material from destabilization.
The loss or a low expression of DNA mismatch repair (MMR) can lead to the accumulation of a
significant number of genetic alterations, including some in protooncogenes or genes regulating
the cell cycle. The dysregulation of MMR gene expression and specific small regulatory
molecules, like miRNAs, has been previously described in upper and lower aerodigestive tract
malignancies, such as lung and head and neck cancer. Although tobacco smoking is a known risk
factor that has been associated with the development and progression of upper aerodigestive tract
malignancies, the exact mechanism by which tobacco smoke components promote defects in the
DNA MMR mechanism associated with upper aerodigestive tract cancers remains unclear.
The aim of this thesis was to investigate whether exposure to Tobacco Smoke
Components (TSC), such as N-nitrosamines, can alter the MMR gene expression in squamous
cancer cells of the upper and lower aerodigestive tract and identify the role of specific miRNA
deregulation in this process.
This investigation showed that the in vitro exposure of human lung and head and neck
cancer cell lines (NCI and FaDu, respectively) to tobacco-specific nitrosamine 4-
(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) could alter the expression of MSH2
and MLH1, key MMR components, by promoting specific miRNA deregulation. The same in
vitro model revealed that either low (1 μM) or high (2 μM) dose of NNK induced significant
upregulation of “oncomirs” miR-21 and miR-155 and downregulation of “tumor suppressor”
miR-422a, as well as the reduction of MMR protein and mRNA expression, in Squamous
Cell Carcinoma (SCC) cells, compared to controls, promoting cancer cell progression. This
model also showed that inhibition of miR-21 restored the NNK-related, reduced MSH2
phenotype in both NCI and FaDu, suggesting that miR-21 might contribute to MSH2 regulation.
The data from in vivo experiments documented for the first time that the chronic exposure
(12–14 weeks) of murine (C57Bl/6J) hypopharyngeal epithelium to TSC [N-nitrosamines; 4-(NMethyl-
N-Nitrosamino)-1-(3-pyridyl)-1-butanone (0.2 mmol/L), N-nitrosodiethylamine (0.004
mmol/L), with or without nicotine (0.02 μmol/L)], can induce premalignant lesions. Molecular
analysis of the lesion revealed activation of NF-κB and its associated oncogenic pathways, as
well as by dysregulation of specific miRNA and MMR genes, similar to the in vitro data.
Specifically, dysplastic lesions caused by TSC exposure demonstrated a significant
downregulation of MSH2 and MLH1 gene expression and deregulation of oncomirs miR-21, miR-155, tumor suppressor
miR-34a, and miR-451a. Analysis of human specimens from tobacco smokers and their
corresponding controls provided a significant reduction in hMSH2 and hMLH1 mRNAs in
hypopharyngeal squamous cell carcinoma (HSCC) that had previously showed significant
deregulation of specific miRNA markers, such as miR-21, miR-155 miR-34a, and miR-451a.
In summary, deregulation of the MMR mechanism and miRNAs is caused by chronic
exposure to TS-related N-Nitrosamines with or without nicotine in the early stages of upper
aerodigestive tract carcinogenesis and can also be detected in human HSCC. Deregulated
miR-21, miR-155, and miR-422a and MMR gene expression patterns may be valuable
biomarkers for lung and head and neck squamous cell cancer progression in smokers. The
data of this thesis contributed to a better understanding of the biological role of MMR genes
in the development and progression of these types of cancer, and provided the foundation for
their future use in diagnostic, prognostic, and therapeutic approaches of upper aerodigestive
malignancies.
|
Collections
