**Abstract**

Tumor viruses cause cancer; thus, extensive studies are being conducted on them. In this article, we will review the basic medical research on the current clinical genetic tests for tumor viruses such as human papilloma virus, hepatitis B virus, and T-cell leukemia virus. Recently, clinical genetic tests have been developed for quick diagnosis of the tumor virus infection. Additionally, we will review the mutagenesis of murine leukemia retrovirus. In particular, we will focus on the insertional mutagenesis. This will help in deciding the direction of future virus research by combining clinical and basic research.

**Keywords:** tumor virus, gene analysis, clinical examination, cancer diseases

## **1. Introduction**

#### **1.1 History of tumor virus research**

Tumor virus research is an important subject to understand the molecular biology of tumor viruses and carcinogenesis. Dr. Katsusaburo Yamagiwa, a Japanese pathologist, is famous for the experiment in which coal tar was applied to rabbit ears to induce carcinogenesis in 1915. He was also a candidate for the Nobel Prize in Medicine. The following year, his student Dr. Akira Fujinami was appointed to study carcinogenesis in Germany for 4 years. After returning to Japan, in 1900, he became the first professor at the Department of Pathology, Kyoto Imperial University Medical College. In 1905, he started studying chicken tumors, and as early as 1910, he established a chicken breast tumor transplant system. He had suggested the possibility of carcinogenesis by filterable pathogens, which were later called Fujinami sarcoma virus (FSV). He is also well known for the discovery of Japan schistosomiasis.

Interestingly, in the same year, Peyton Rous, who was studying at the Rockefeller laboratory in the United States, also established the avian sarcoma transplant system. In 1966, Rous received the Nobel Prize in Physiology and Medicine for the discovery of carcinogenic viruses. Hanafusa et al. elucidated the FSV gene structure and identified the similarity with the Rous sarcoma virus gene structure [1]. Both FSV and Rous sarcoma virus belong to the same alpha-retrovirus. FSV has the oncogene v-fps in its genome [2]. It also has long terminal repeats (LTRs) at the ends of the genome that stabilizes as parts of the host cell genome when inserted into host mesenchymal cells after reverse transcription. In addition to the host cell division, the viral genome also replicates during the DNA replication cycle. Tyrosine-protein

kinase transforming protein Fps encoded by v-fps has the ability to induce transformation by promoting tyrosine phosphorylation of the PDGFbeta receptor [3, 4]. Thus, the Japanese scientists have greatly contributed to the field during the early days of oncology research.

#### **1.2 Viral genome integration**

Retroviral integration is a type of insertional mutagenesis. Integration processes are widely studied from biochemistry, and their molecular mechanisms are elucidated. Retrovirus-induced tumorigenesis is divided into two types. First, the retroviral genome carries the oncogene such as *v-onc*, and the inserted viral element may induce malignant tumors. Second, the inserted retroviral element has the promoter or enhancer activity of the host proto-oncogene. The genome of human papilloma virus (HPV), hepatitis B virus (HBV), human T-cell adult leukemia virus (HTLV) [5], avian sarcoma virus (ASV) [6], feline leukemia virus (FeLV) [7], murine leukemia virus (MLV) [8–13], and mouse mammary tumor virus (MMTV) can be integrated in the host genome. As a survival strategy, the inserted retroviral genome (provirus) is thereafter replicated during host cell division.

The retroviral promoter and enhancer elements in LTRs originally promote the expression of retroviral genes such as *gag*, *env*, and *pol* [14] (**Figure 1**). However, once integrated in the host genome, LTR elements can promote host gene expression in place of the host genome promoters and enhancers. After infection, the retroviral RNA genome is reverse-transcribed into DNA, which forms a pre-insertion complex (PIC) with the oligomeric integrase enzyme [15], which catalyzes the insertion reaction. The PIC crosses the nuclear envelope through the nuclear pore complex to access host cell DNA. The retroviral DNA ends then attack and nick the host DNA with target nucleotides. These ends are ligated to the truncated host DNA to complete the insertion process involving replication of the target nucleotides.

To date, extensive research has been conducted on target motifs for retroviral insertion to better understand tumorigenesis, neuronal damage, aging, and immunodeficiency including acquired immunodeficiency syndrome (AIDS). Retroviralhost DNA interactions support specific host DNA structures [16–21] but are less likely to be inserted into particular base sequences. Previous studies on human immunodeficiency virus (HIV)-1 insertion have shown that the insertion sites are distributed in 10-bp cycles on the nucleosome surface of the outward DNA major groove of chromatin [16, 22].

#### **Figure 1.**

*A common structure of the retroviral genome. Plus strand RNA is shown. R, terminal redundant sequence; U5, unique 5*′ *sequence; PBS, tRNA primer-binding site; gag, group-specific antigen encoding capsid protein MA (matrix), CA, capsid protein; NC, nucleocapsid; pro, protease; pol, polymerase including reverse transcriptase (RT) and integrase (IN) catalyzing integration; env, envelope glycoproteins; U3, unique 3*′ *sequence. U3, R, and U5 are duplicated at both ends of the DNA provirus genome in the host genome sequence to form the long terminal repeat (LTR). LTR is essential for the initiation of viral DNA syntheses by reverse transcription as well as the integration of proviral DNA and the regulation of viral gene expression.*

**37**

*Basic Research and Clinical Examination of Tumor Virus*

HPV is one of the most common viruses that infect more than half women once in life through sexual intercourse. It is a small DNA virus with approximately 8000 bp circular double-stranded DNA in its genome. The viral particle has icosahedral capsid structure as its outer shell. The virus has been known to be involved in the development of cervical, anal, and vaginal cancer and condyloma acuminatum. Many types of HPV are known, and high-risk HPV based on this cancer risk is concentrated in specific subgroups epidemiologically. In particular, types 16 and 18 have high carcinogenic risks of cervical intraepithelial neoplasia (CIN) and cancer. Type 16 has been detected in the Japanese 5-year-old girls [23]. High-risk group (types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, and 68) causes bladder cancer and pharyngeal cancer in addition to vulvar and cervical cancer. Low-risk group (HPV2,

27, and 57) causes verruca vulgaris that develops in the hands and feet [24].

HPV genome has early genes (E1, E2, E4, E5, E6, and E7) and late genes encoding viral capsid proteins (L1 and L2). E6 and E7 are considered to be involved in carcinogenesis. E6 binds to a tumor-suppressor gene product p53 and degrades p53 through promoting the ubiquitination. It also contributes to carcinogenesis by reactivating telomerase reverse transcriptase (hTERT) and degrading proteins with PDZ domain [shared by—post-synaptic density protein (PSD95), Drosophila disc large tumor suppressor (Dlg1), and Zonula occludens-1 protein (zo-1)]. E7 binds to the tumor suppressor gene pRb and contributes to carcinogenesis by releasing and activating the transcription factor E2F bound to pRb. It is also involved in carcinogenesis by interacting with cyclin-dependent kinase (Cdk) inhibitors p21 and p27 to evade cell cycle checkpoint mechanisms and apoptosis. However, in many cases, immortalization does not occur only by the actions of human cervix mucosal cells E6 and E7. In the well-known HeLa cell line, telomerase is activated, preventing the

High-risk HPV can be detected in more than 90% patients suffering from cervical cancer and dysplasia, which is a precancerous lesion, and in the vulva of 5% normal women. Although HPV is also integrated into the genome of host cells, its integration pattern has unique features. The E6 and E7 genes are retained in the provirus with frequent deletion mutations in E2 gene. Thus, the viral genome is also found to be truncated in the complex reaction of integration, but its carcinogenicity is rather enhanced. Low-grade dysplasia of the cervix and cervical intraepithelial neoplasia (CIN) is well known. These pathological changes are reversible and are known to spontaneously disappear within about 3 years. However, about 10% infected patients sustain the infection for more than 3 years, and cervical cancer occurs after several months to several decades in some of them. HPV genome is integrated into the host cell genome during the progression of CIN, and the pro-

HPV-DNA typing tests have been conducted using real-time PCR, loopmediated isothermal amplification (LAMP) [25, 26], invader, and hybridization methods. In recent years, the Clinichip method [27] combining LAMP is broadly available in advanced countries including Japan due to the ability to amplify genes at a constant temperature. The LAMP method is very specific because it uses four primers to recognize the six regions of the HPV genome. Nonspecific amplification does not occur because the amplification reaction depends on the order of the six regions of the HPV gene. In addition, since the amplification product constitutes

*DOI: http://dx.doi.org/10.5772/intechopen.89331*

gradual shortening of telomeres in cell death.

gression becomes irreversible.

**2.1 Clinical test for HPV detection**

**2. HPV**

*Basic Research and Clinical Examination of Tumor Virus DOI: http://dx.doi.org/10.5772/intechopen.89331*
