CCCTC-binding factor (CTCF) is a multivalent 11-zinc finger (ZF) protein that binds tens of thousands of sites in the human genome [1]. Initially identified as transcriptional regulator of chicken MYC proto-oncogene (c-myc gene) in 1990 [2, 3], CTCF was later found to mediate gene imprinting at the H19 imprinted maternally expressed transcript/insulin-like growth factor 2 (H19/Igf2) locus [4]. Follow-up studies indicated that CTCF interferes with the interaction between enhancers and promoters because of its binding sites frequent presence in various chromosomal structural boundaries, such as the topologically associated domains (TADs) [5, 6]. Increasing evidence indicated that CTCF protein is highly conserved from Drosophila to humans [7]. CTCF is one of the key regulatory proteins in vertebrates, and its knockout leads high in utero embryonic lethality [8]. CTCF can participate in a variety of biological processes and have different functions, including insulator function [9], regulation of gene transcription [10], gene imprinting (Figure 1) [11], X chromosome inactivation [12, 13], affecting messenger RNA (mRNA) alternative splicing as well as nucleosome rearrangement and DNA replication [14, 15]. Recently, high-throughput chromosome conformation capture (Hi-C) indicated that CTCF has dynamic roles as regulator of imprinted loci and governing higher-order chromatin architecture other than enhancer blocking activity [5, 16]. The latest evidences revealed that CTCF is directly involved in the transcriptional modulation of various key factors of cellular cycle control, apoptosis, senescence, and differentiation [17, 18]. Indeed, ectopic expression of CTCF in several human tumoral cell lines inhabits cell division and clonogenicity [18]. Somatic mutations at CTCF-binding sites, which can abrogate the CTCF-mediated spatial folding of chromosomes [19], have been profoundly found in various human cancers [20]. Interestingly, CTCF also has a key role in the establishment of three-dimensional (3D) chromatin structure during human embryogenesis [21]. The basic framework of the hypothesis of cancer evolution and development (Cancer Evo-Dev) is shown in Figure 2. The complex interactions between host genetic susceptibility and environmental exposure [such as hepatitis B virus (HBV) infections] induced immune system disorder. The dysfunctional immune system activates and maintains chronic inflammation with the permanent existence of HBV in host. Under the inflammatory microenvironment, the imbalance of mutagenic factors and mutant repair factors, cooperating with HBV integration and epigenetic modulations, promotes HBV-related somatic mutations and HBV mutations. A majority of hepatocytes with genomic mutations and HBV variants are eliminated in the survival competition in inflammatory microenvironments. Only a small part of cells survive by alternating survival signaling pathways and exhibit “stemness” characteristics. The survivals gradually evolved to be cancer stem cells, which drives the development and evolution of cancer [22]. According to the Cancer Evo-Dev hypothesis, CTCF is considered as an essential regulator during cancer cells “backward evolution” and “retro-differentiation”.

There is a growing consensus that a hierarchical chromatin structure is established within the nucleus in the interphase of the cell cycle [23, 24]. The spatial folding of chromosomes and their organization, which is largely mediated by CTCF, have profound effects on gene expression [25, 26]. Genetically inducible CTCF deletion in several specific cell types, including oocytes, lymphocytes, and cardiomyocytes, leads to organ-specific failure [27, 28]. Knocking out the CTCF of the oocyte seriously disrupts the entry of the fertilized oocyte into the blastocyst stage [28]. Furthermore, homozygous deletion of CTCF at the whole embryo level results in embryo death [8], which is characterized by aberrant enhancer-promoter interactions and transcriptional dysregulation [29]. However, albeit numerous studies have explored the function of CTCF binding, the in vivo roles of CTCF during embryonic development are largely unknown.

Recent studies revealed that TAD structures are totally lost in human 2-cell embryos, are at a low level in 8-cell embryos, and then gradually established during embryonic development [21]. As an essential hierarchical structural feature of chromatin organization, A/B compartmentalization is absent in human 2-cell embryos and reformed during embryogenesis [21, 30, 31]. Unlike in mature mouse sperm, TAD structures are absent in human sperm. Previous studies have shown that depletion of CTCF can lead to the disruption of TADs, suggesting that the lack of CTCF may contribute to the loss of TAD structures in human sperm [21, 32]. Immature TAD boundaries gained at the 2-cell embryos tend to locate around housekeeping genes, which may promote the expression levels of housekeeping genes. In addition, both CTCF expression and TAD establishment in human embryos require human zygotic genome activation (ZGA). Further studies suggested that CTCF expression is required, but is not the only factor needed, for TAD establishment during human ZGA [21].

CTCF can induce both cell cycle progression or arrest to accomplish its function [17], which is dependent by cell-specific CTCF-binding DNA sequences (CTSs), protein partners and chromatin long-range interactions [33]. CTCF can repress cell growth and colony formation suggesting a suppressor role of CTCF [34, 35].

It is reported that CTCF depletion activates DNA damage response and increases the risk of chromosomal instability [36]. DNA damage signaling, Mre11 (MRE11 homolog, double strand break repair nuclease)-Rad50 (RAD50 double strand break repair protein)-Nbs1 (nijmegen breakage syndrome 1) complex and CTCF DNA-binding domain promote CTCF enrichment at DNA damage sites [36]. As the core DNA-binding protein of homologous recombination (HR), RAD51 is overexpressed in a variety of tumors [37]. CTCF participates in HR repair of DNA double strand breaks by interacting with RAD51 and promoting the formation of RAD51 repair foci [38]. CTCFL (CTCF like), otherwise known as Brother of the Regulator of Imprinted Sites (BORIS), is a male system-specific protein with the same 11-zinc finger structure as CTCF [38]. In non-small cell lung cancer (NSCLC), BORIS suppresses DNA damage and promotes cisplatin resistance by enhancing the mismatch repair system of cancer cells [39].

Gene methylation detection showed that DNA methylation at the CpG sites in the CTCF coding gene caused the CTCF binding sites to be blocked [40]. The function of CTCF in regulating gene imprinting suggested that the binding of CTCF to the target gene is methylation sensitive [4]. The aberrant DNA methylation of key regulate regions can hinder CTCF binding, which may lead to epigenetic silencing of tumor suppressor loci or lead to activation of oncogenes [41]. Large tumor suppressor (LATS) kinase can be activated by a variety of stress responses, such as glucose deficiency, and then phosphorylates downstream pathways to promote cell survival, while abnormal LATS activation promotes tumorigenesis. LATS kinase directly phosphorylates CTCF at the ZF junction, inhibiting its DNA binding activity [42]. Recent studies indicated that DNA methylation-regulated alternative cleavage and polyadenylation (APA) requires CTCF and the cohesin complex [43]. Epigenetic inactivation of Ras association domain family 1 isoform A (RASSF1A) and E-cadherin (CDH1) in breast cancer is associated with alternations in CTCF recognition sites [44]. Aberrant DNA methylation can inhibit CTCF-mediated silencing of BCL6 transcription repressor (BCL6) gene, thus increasing the expression of proto-oncogene BCL6 in lymphoma [45]. In addition, alternations in CTCF gene in endometrial cancer can promote tumorigenesis by promoting cell survival and changing cell polarity [46].

CTCF hemizygous deletions are frequently found in various human tumors [47]. Loss of a single CTCF allele (Ctcf ^+/−) significantly increases the risk of cancer and enhances malignant progression [40]. CTCF hemizygosity dysregulates cancer-related pathways, such as Ras, Ras-mitogen-activated protein kinase (Ras-MAPK) and extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathways [48]. Ctcf ^+/− mice remain a disorder methylation status, and even lead to a modest overall increase levels of genome-wide DNA methylation in lung [40]. Chromosome 16q22.1 is a common deletion region in various epithelial cancers, and CTCF is located on this region. In prostate cancer cells, knockdown of CTCF results in hypermethylation of the CTCF/cohesin-binding sites (CBSs). Prostate and breast cancers with insufficient CTCF copy number show increased DNA hypermethylation events in vivo [49]. In addition, high frequency mutations of CBSs also lead to disorders of specific sites hypermethylation and promote the occurrence and development of cancer [43].

CTCF has been shown to modulate the expression of various cancer-associated genes. c-myc plays vital roles in tumorigenesis, embryogenesis, and somatic cells reprogramming [50–52]. CTCF exerts divergent roles on c-myc regulation by binding various sequences at c-myc promoter [1]. In myeloid cells, overexpression of CTCF decreases c-myc levels and growth rate, which promotes myeloid differentiation and induces cell cycle arrest in numerous tumoral cells [34, 53]. In most malignant tumors, telomerase is activated to prevent telomere shortening, which confers high proliferative capacity to tumoral cells [54, 55]. As one of the telomerase components, human telomerase reverse transcriptase (hTERT) is only expressed in telomerase-positive cells, and depletion of CTCF induced hTERT transcription in TERT-negative cells [56, 57]. However, CTCF is not able to exert a transcriptional repressive function because of DNA-methylation or BORIS-binding in TERT-positive cells [58]. Moreover, in cancer cells, methylation of CTSs at hTERT exon inhibits CTCF-binding, which prevents hTERT repression [59]. Retinoblastoma (RB) is considered a tumor suppressor gene and CTCF maintains RB gene promoters at an active epigenetic status [60, 61]. The inactivated of RB gene family leads to the loss of cell cycle control and cancer. Other evidences showed that CTCF is also involved in an epigenetic balance at the cyclin-dependent kinase inhibitor 2A locus (CDKN2A) and tumor protein p53 (TP53) gene promoters [58, 62]. In sum, by binding various CTSs, CTCF can modulate c-myc, hTERT, RB, and other tumor-related genes, which emphasize the cell-type dependency of its tumor suppressor role.

Cancer development is characterized by an evolutionary process of “mutation-selection-adaptation”, some highly conserved genes are highly expressed in the embryo, not expressed or low expressed in normal adult tissues, but highly expressed in cancer tissues, leading to reverse cell differentiation, malignant proliferation and enhanced migration capacity [22]. Recent evidences revealed that in normal prostate tissue, CTCF expression was negative to low, while in prostate cancers, CTCF expression was seen in 7,726 of 12,555 (61.5%) tumors and was considered low in 44.6% and high in 17% of cancers [79]. CTCF expression is a feature of poor prognostic in prostate cancer, but CTCF is a dissatisfactory candidate biomarker because of its low predictive power [79]. Similarly, CTCF is frequently up-regulated in partial primary hepatocellular carcinoma (HCC) compared with non-neoplastic liver. Overexpression of CTCF is associated with shorter disease-free survival in patients and the absence of CTCF can lead to decreased motility and invasiveness of HCC cells [80]. In addition, chromosomal ring anchors bound by CTCF and cohesin are prone to continuous DNA breakage, and regions of translocation break points in various cancers are enriched in these anchors [67]. Continuous DNA break and repair provide opportunities for genetic mutations, and in the context of numerous mutations, cancer is constantly selecting and adapting. Furthermore, multiple cancer types accumulate CBS mutations and CBSs are major mutational hotspots in the noncoding cancer genome, which emphasize the significant role of CTCF in cancer evolution and development. In summary, CTCF hemizygotic mutations and CBSs mutations disrupt genomic stability and, in combination with epigenetic modifications such as methylation, promote the mutation and adaptive selection of cancer cells, thereby promoting the evolution and development of cancer.

CTCF, a highly conserved and multifunctional protein, contributes to formation of multi-dimensions genome and control of central signals to transcriptional networks [5]. CTCF plays an essential role in embryonic development and cancer development, but its molecular mechanism has not been fully elucidated. CTCF exerts a tumor suppressor role by regulating several key factors related to growth and development and regulating the higher-order structure of chromosomes, but its high expression in various cancer cells may promote the malignant proliferation and migration, and often predicts a poor prognosis [33, 79]. Although roles of CTCF in carcinogenesis have been intensively explored, more researches are needed to better understand the significant functions and mechanisms of CTCF in embryonic development and cancer development.

With the continuous development of modern bioinformatics and biotechnology, the combined use of a variety of high-throughput analysis techniques such as chromatin immunoprecipitation-chip (ChIP-chip), ChIP-qPCR, and so on can constantly find out the corresponding binding sites and regulatory models of CTCF with DNA, protein and RNA, which can further clarify the action mechanism of CTCF. The developed highly specific epigenome editing technology based on clustered regularly interspersed short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) as a broad prospect because it can modify the genome of specific regulatory elements [81]. For example, targeted editing of the DNA methylation status of CTCF binding sites can alter the expression of CTCF, thereby altering the expression of target genes by affecting the structure of advanced chromatin [82]. Therefore, the highly specific epigenome editing technology based on CRISPR/Cas9 may become an attractive epigenome-based cancer therapy in the next few years to regulate the occurrence and development of tumors.