The Transition from Cancer "omics" to "epi-omics" through Next- and Third-Generation Sequencing

Abstract

Deciphering cancer etiopathogenesis has proven to be an especially challenging task since the mechanisms that drive tumor development and progression are far from simple. An astonishing amount of research has revealed a wide spectrum of defects, including genomic abnormalities, epigenomic alterations, disturbance of gene transcription, as well as post-translational protein modifications, which cooperatively promote carcinogenesis. These findings suggest that the adoption of a multidimensional approach can provide a much more precise and comprehensive picture of the tumor landscape, hence serving as a powerful tool in cancer research and precision oncology. The introduction of next- and third-generation sequencing technologies paved the way for the decoding of genetic information and the elucidation of cancer-related cellular compounds and mechanisms. In the present review, we discuss the current and emerging applications of both generations of sequencing technologies, also referred to as massive parallel sequencing (MPS), in the fields of cancer genomics, transcriptomics and proteomics, as well as in the progressing realms of epi-omics. Finally, we provide a brief insight into the expanding scope of sequencing applications in personalized cancer medicine and pharmacogenomics.

Keywords: DNA methylation; RNA sequencing; cancer genomics; epigenomics; epiproteomics; epitranscriptomics; massive parallel sequencing; pharmacogenomics.

Figure 1

The realm of “omics”. The current sequencing approaches allow the decoding of genetic information at multiple levels. Today, DNA sequencing can be performed to decipher the nucleotide sequence of our genes as well as their epigenetic marks (genomics and epigenomics), while RNA sequencing allows the exploration of transcriptomes and the detection of RNA modifications (transcriptomics and epitranscriptomics). Finally, developing sequencing approaches that aim to investigate proteins (proteomics and epiproteomics) will lay the groundwork for a deeper understanding of cellular function and structure.

Figure 2

Flowchart displaying the fundamental steps of NGS workflows. (a) For genomic and epigenomic studies, the library preparation includes the DNA extraction and fragmentation and the ligation of the adapters on the derived DNA fragments. After the size selection, the molecules are amplified via PCR and subsequently the generated NGS library is sequenced. The final step of this process is the analysis of the obtained sequencing data using appropriate algorithms. (b) For transcriptomic and epitranscriptomic studies, the construction of RNA libraries is similar to the DNA library preparation workflow, but they are distinguished due to an additional reverse transcription step prior to the PCR amplification. Notably, the identification of both DNA and RNA modifications is based on the usage of specific antibodies that capture the sequence of interest.

Figure 3

Different levels of genetic information and the related applications of MPS technology. DNA sequencing applications aim to detect DNA modifications, mutations, gene fusions and histone modifications, whereas sequencing-based RNA studies include the identification of alternative splicing events, transcript fusions and RNA modifications.

Figure 4

Pharmacogenomics is based on the sample collection from patients dealing with the same disease, the extraction of nucleic acids from each sample and the subsequent sequencing of them, following the appropriate workflow. After data analysis, the unique genetic profile of each patient is used as a reference point for the choice of the optimal drug treatment each one should receive.