Abstract

Epigenetics is the study of the phenotypic variation caused by external factors (e.g. diet, nicotine use, and carcinogenic chemical exposure) that influence the mechanisms through which cells read and interpret genes. Epigenetic modifications are independent of genetic mutations. Currently, epigenetics offers significant promise in novel noninvasive cancer therapies and early diagnostic tools. Key epigenetic processes including DNA methylation and histone modification are reversible, unlike most genetic mutations. Reversing these processes in tumor suppressor genes can restore normal behavior in tumor cells. This review discusses the biological basis and treatment potential of these processes and provides a brief analysis of their potential application in cancer treatment.

Introduction

DNA is a double-helical structure packed into the nuclei of eukaryotic cells in the form of chromatin. The basic unit of DNA packing in chromatin is the nucleosome, a complex of eight histone proteins and approximately 140 DNA base pairs. Further coiling of repeating nucleosome fibers eventually yields a chromatid, a key component of a chromosome.

Gene expression is epigenetically regulated through alterations in chromatin structure and the double helix of DNA by addition of functional groups. DNA methylation and histone modification are the most understood mechanisms of such epigenetic modulation. Errant modification of functional group patterns in DNA or histone tails could result in unintended silencing of critical regulatory genes, a process which contributes to the development of several diseases, including cancer. Understanding these complex mechanisms is essential for the development of cancer therapies that reverse the inhibition of tumor-suppressor genes. This review seeks to outline the biochemical basis of the gene silencing effected by DNA methylation and histone modification, as well as the current developments in reversing these mechanisms for the prevention of cancer.

Histone Modification & Its Implications on Cancer

Histone modifications are reversible modifications at the N-terminals of histones in a nucleosome. These include, but are not limited to, acetylation, methylation, phosphorylation, and ubiquitination. Depending on the group added or removed from a given histone, local gene transcription can be upregulated or downregulated. Histone acetylation, for instance, modifies chromatin into a less-condensed conformation that is more transcriptionally active. Conversely, histone deacetylases (HDACs) remove acetyl groups, increasing the ionic attractions between positively charged histones and negatively charged DNA and tightening chromatin structure. In order to further condense chromatin, HDACs can also recruit K9 histone methyltransferases (HMTs) to methylate H3K9 histone residues, providing the condensing agent heterochromatin protein 1 (HP1) with a binding site. As a result of the condensed heterochromatin conformation, the cell has limited transcriptional access to the genome.

Recent research has established histone modifications as useful biomarkers in cancer diagnosis. HDACs, specifically HDAC1, can often be identified in elevated quantities in prostate and gastric cancer patients. Prostate cancer tumors have been characterized by general hypomethylation of histone residues H4K20me1 and H4K20me2, as well as hypermethylation of residue H3K27 and increased activity of its specific methyltransferase. These conditions are now associated with progression and metastasis of prostate cancer. Histone modifications play a multifaceted role in cancer diagnosis and treatment. In addition to serving as diagnostic biomarkers, they are also potential therapeutic agents. Drugs that specifically inhibit HDACs are currently being manufactured and assessed for clinical application after recent FDA approval.5

DNA Methylation & Its Implications on Cancer

In DNA methylation, a methyl group is covalently added to a cytosine ring in the DNA sequence, forming a 5-methylcytosine. Molecules of 5-methylcytosine are found primarily at cytosine-guanine dinucleotides (CpGs). Some areas of high CpG density, such as CpG islands, are characteristically unmethylated and are located in or near the promoter regions of genes, allowing them to play a role in gene expression. Enzymes termed DNA methyltransferases (DNMTs) facilitate the methylation of CpG residues. Three key members of this family include Dnmt1, Dnmt3a, and Dnmt3b. Dnmt1 is labeled as the “maintenance” DNMT due to its methylation of DNA near replication forks, which preserves the epigenetic inheritance of methylation patterns across cellular generations. The other two DNMTs serve to directly methylate other CpG residues. These enzymes can either be recruited by a transcription factor bound to the promoter region of a given gene to methylate a specific CpG island or simply methylate all CpG sites across a genome not protected by a transcription factor. Several families of proteins, such as the UHRF proteins, the zinc-finger proteins, and most notably, the MBD proteins, are responsible for the interpretation of these methylation patterns. For example, MBD proteins possess transcriptional repression domains that allow them to bind to methylated DNA and silence nearby genes.

For a malignant cancer to develop, critical tumor-suppressor genes such as p53 must be knocked out. This process can be accomplished through hypermethylation of CpG islands in certain promoter regions and genes. For example, when the DNA repair gene BRCA1 becomes hypermethylated, it is rendered inactive, which can lead to the development of breast cancer. The reversibility of methylation implies that these tumor-suppressor genes could be reactivated to effectively treat tumors. Infiltration of a cell by pharmaceutical agents that inhibit methylation-mediated suppression could restore the function of a silenced tumor-suppressor gene.6

One recent study demonstrated that azacitidine, an agent that can be incorporated into DNA, is capable of inducing hypomethylation. The chemical modification of this agent’s diphosphate form by a ribonucleotide reductase and subsequent phosphorylation creates a triphosphate form that displaces cytosine bases in DNA. As a result, DNMTs are limited in methylation functionality since they are isolated on a substituted DNA strand.7 Such manipulation of naturally occurring DNA methylation processes in cancer cells appears to be a promising method of restoring the normal function of tumor-suppressor genes.

Co-Application of DNA Methylation & Histone Modification

The discovery of the protein MeCP2 has validated the hypothesis that DNA methylation regulates histone acetylation patterns. MeCP2 recruits histone HDACs to the promoter regions of methylated CpG islands. The resulting hypoacetylated histones yield a highly condensed heterochromatin structure that represses transcription of nearby genes. Clinical studies have thus emphasized the utilization of both demethylating agents and histone deacetylase inhibitors for transcriptional activation of tumor-suppressor genes.8

Conclusion

The field of epigenetics has opened a door to novel, promising cancer therapies unimaginable just a decade ago. Manipulating DNA methylation and histone modification can reverse tumor-suppressor gene silencing. Reactivating tumor-suppressor genes through epigenetic modifications can halt and reverse cancer proliferation.

References

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