Key words tumor chemotherapy thymidylate synthase (TS polymorphism resistant

    The molecular characteristics of the tumor has a high degree of heterogeneity and individual tumor lesions malignant characteristics of the tumor. Genomics and proteomics as a new platform for cancer treatment through gene polymorphism and its expression in tumor molecular typing individualized treatment purposes. Ancient medical same disease with different treatment, diagnosis and treatment basically the same point. Thymidylate synthase (thymidylate synthase, TS) is a key enzyme in DNA biosynthesis reaction, and its encoding gene into the target gene of the treatment of tumors of many chemicals. TS and its gene polymorphism has important guiding significance for cancer prevention, individualized treatment and prognosis.

    1 TS, TS gene polymorphism and tumor genomic instability

    1.1 TS

    TS is a dimeric protein composed of two identical subunits, and the relative molecular mass of 72 kDa, a key enzyme for DNA synthesis and methylation of it the catalytic Deoxyuridine acid (dUMP) into deoxythymidine acid ( dTMP). TS has a very important role in DNA synthesis and repair, cell proliferation and differentiation, and is an important target enzyme in cancer chemotherapy drug 5  Fu and methotrexate.

    1.2 TS gene polymorphism

    TS gene is located on chromosome 18 (18p11.32), long 16KB major transcription initiation site is located in the ATG initiation codon upstream 160 ~ 180bp, enhanced promoter is located about 400 bp upstream of the initiation codon ATG. Gene polymorphism is present on two or more alleles in the same loci in the normal population, and each allele are having a relatively high frequency (the frequency is greater than 1%). It has now been found that the non-translated region of the TS gene (untranslated region, UTR) polymorphism. Polymorphism can be divided into two categories, namely length polymorphism (length polymorphism) and polymorphism (site polymorphism).

    The 1.2.1 TS gene length polymorphism length polymorphism can be divided into two categories: the number of a class of tandem repeat sequence (variable number of tandem repeats, VNTRS), such as small satellites; another type of length polymorphism gene the due to the deletion or insertion of a particular fragment, such as microsatellites.

    (1) 5 ‘ UTR tandem repeat sequence the different numbers promoter transcription process starting epitope, the regulation of gene expression has indispensable. Enhanced promoter is a DNA sequence to enhance the transcription activity of the promoter. TS enhancer region of 28bp tandem repeat polymorphism exists divided containing VNTRs repeated two more times for the 2R, repeated 3 times for 3R, see Figure 1.

    More repetitions allele (4R, 5R, 9R) occurrence frequency rarely. Therefore, the common three genotypes 3R/3R, 2R/3R, 2R/2R.

    (2) TS gene 3 ‘ UTR of polymorphism in 2000, Ulrich found The TS mRNA 3’  UTR of 1494bp Department there is a six-base deletion / insertion polymorphism form a +6 bp / +6 bp in bp/-6bp +6 genotypes, -6bp/-6bp. Ulrich detected by 95 American Caucasian TS 3 ‘ UTR of genotype distribution may  6bp allele frequency distribution of low-6bp the allele called a variant allele. In fact, the dominant white British, Chinese people a lot of race-6bp allele type. Therefore, we believe-6bp the allele called a variant allele is inaccurate [2].

    1.2.2 TS gene polymorphism loci polymorphism the different bases scattered in the genome, including point mutations, single base substitution (also known as single nucleotide polymorphisms, SNP), missing and insertion. The Mandola found the first repeat sequences in the 5 ‘ UTR 2R allele and repeated twice before the 3R allele sequence USF family E  the box (CACTTG). The 3R allele second repeat sequence of 12 nucleotides, etc. in presence of G → C SNP, as shown in Figure 1 [1]. Thus 2R, 3R, respectively, 2G, 2C, 3G, 3C allele type. 2R/2R, 2R/3G, 2R/3C, 3G/3G, 3G/3C, 3C/3C common genotype. SNP is the most extensive polymorphisms in the human genome, the genetic causes of individual differences, attracting much attention in the study of the human genome.

    1.2.3 TS gene polymorphism distribution frequency there are racial differences in different ethnic populations in the TS gene polymorphism distribution of frequency are very different. (1) 2R, 3R allele frequency distribution of the Turks for 42%

    Figure 1 TS gene 5 ‘ UTR tandem repeat polymorphism and single nucleotide polymorphisms [1]

    Fig 1 The TS gene has a variable number of tandem repeats and single nucleotide polymorphism in the 5 ‘ untranslated region

    And 58% [3], in the Chinese population of 18.82% and 81.00%, respectively, while the Africans. (2) the American whites TS 3 ‘ UTR of +6 bp allele frequency was 71%, North China, South China, the Chinese population with respectively 32% and 30.9% [4]. Gene polymorphic frequency distribution of racial differences, and shows the diversity and complexity of the genetic background, may be an adaptive performance to withstand adverse environmental factors in human evolutionary process, the maintenance of the survival of the population continues to have important biological significance.

    1.3 tumor genomic instability

    Many tumor genomic instability, allelic imbalance characterized. Heterozygous deletion (LOH) is a common form of allelic imbalance. LOH is the lack of an allele from the father or the mother. The tumor suppressor gene inactivation mutation in one allele and the other allele and occurrence of LOH. Kawakami [5] by 151 cases of colorectal cancer detection 5 ‘ UTR repeat sequence genotype in 31 cases out of 50 patients with heterozygous 2R, 3R allele imbalance, that of LOH. Loss of heterozygosity (LOH) and microsatellite instability (MSI), the two main phenotypic characteristics of genomic instability. The deletion of the tumor suppressor gene is one of the key steps in many tumors; replication errors is an important part of many tumorigenic process can increase the mutation of the tumor suppressor gene. Thus the tumor tissues TS genotyping, genomic instability and other factors should also be considered.

    1.4 TS gene polymorphism and TS expression levels of relationship

    TS gene polymorphism affects the TS gene mRNA stability and translation efficiency, resulting in different genotypes of TS expression efficiency. Therefore, the TS gene polymorphism TS expression of the main factors for the level of its subtypes may exist. Morganti [6] TS mRNA expression levels show 3R/3R genotypes cancer tissue by 48 cases of colorectal cancer patients was significantly higher than 2R/2R, 2R/3R genotype. TS3 ‘ TUR6bp missing Mandola colorectal cancer patients is associated with reduced in vitro TSmRNA stability associated with lower TS expression in vivo tumor may increase the risk of cancer patients; +6 bp allele view consistent low risk of breast cancer [4].

    Some scholars believe that the TS gene polymorphism and TS expression levels of correlation does not exist. 2001 Kawakami [7] found that the TS gene 5 ‘ UTR polymorphism has nothing to do with the TS mRNA expression levels.

    2 TS, TS gene polymorphism and cancer relationship

    2.1 TS gene polymorphism and cancer susceptibility

    Susceptible individuals the 2.1.1 TS gene polymorphism and cancer susceptibility crowd there is because some tumor gene polymorphism. Therefore, many scholars believe, is closely related to the TS gene polymorphism with breast cancer risk. Adleff et al [8] reported the 3R alleles and the risk of colorectal cancer risk associated with elevated, this may be the 3R allele sake of TS expression levels, promote excessive cell proliferation, escape senescence and apoptosis.

    2.1.2 TS gene polymorphism and tumor susceptibility factors interaction between tumor occurrence and development of joint action by both genetic and environmental factors of multifactorial diseases, gene  gene, gene  environment interactions in the pathogenesis of important effects . (1) gene the  gene interactions. Zhengdong Zhang and other studies have shown: the TSER 2R TS3’UTR 6 bp allele, in southern China in the etiology of gastric cancer may play a synergistic role [9], there is a positive interaction. (2) gene  environment interactions. Gao Changming [2] found that the TS-6bp/-6bp genotype with smoking, alcohol consumption, and non-tea drinking habits increase the risk of gastric cancer in significant synergies. (3) affect folate levels. Human red cell folate levels <140ng/ml or plasma folate levels <3ng/ml will induce chromosomal damage and increase the risk of cancer. TS5 ‘ UTR of the 2R allele and low folate intake of colorectal cancer risk significantly increased, there was an interaction between the two [10]. TS gene polymorphism and cancer susceptibility correlation, it is possible to make a genetic diagnosis before the onset of symptoms, tumor prediction, prevention, early diagnosis of inestimable value.

    2.2 TS with cancer chemotherapy

    TS expression is regulated depends on the state of the cell cycle and cell proliferation. In synchronous cell, when cells from G0 into S phase, the TS increase in the level of activity of about 20 times. 5  Fu play its anticancer activity through inhibition of TS. 5  Fu enters the body into 5  fluorouracil nucleoside (FdUMP), exist in the cofactor 5,10  Methylentetrahydrofolate (CH2FH4), and TS combine to form the equivalent three-composite matter TS  FdUMP  to CH2FH4 affect the combination of TS with dUMP inhibited dTMP synthesis, DNA synthesis constrained to achieve the eradication of cancer cells purposes. ZOU K [11] and other recently reported esophageal cancer cell lines and colon cancer cell lines, insulin can increase the number of cells in S phase, change the complex of the three-level enhancements 5  Fu anticancer effects.

    Many scholars believe that there is a negative correlation between TS levels and the efficacy of chemotherapy. Banerjee et al [12] reported that TS cRNA transfected tumor cells, the TS high expression, cell 5  Fu and FdUrd resistant. DLD  1 cells transfected with TS antisense RNA was significantly inhibited TS expression and vitality, 5  Fu sensitivity significantly enhanced [13]. TS gene expression levels in patients with more than 4.0 × 10-3 is usually not sensitive to chemotherapy. These studies have shown that TS levels can predict tumor chemotherapy.

    But there are also many scholars do not agree with this view. Derenzini chemotherapy largely powered by the tumor cells [14] think: better rapidly growing tumor effect. The target enzyme TS high expression of the tumor to 5  Fu-based chemotherapy improve the prognosis better. About the relationship between the TS and the resistance, despite their conclusions differ, the correlation between the two has been most scientists agree. Some scholars TS expression levels and improve the prognosis in patients draw a negative conclusion, ignored efficacy prognosis is influenced by multiple factors, so not enough to negate the correlation between TS levels and the efficacy of chemotherapy does not have a statistically significant difference.

    In the TS inhibitors chemotherapy, the tumor cells, there may be increased expression of TS protein called TS-induced. TS TS protein expression induced causes excessive catalytic activity increased, which led the resistance of tumor cells. Phase Ⅱ clinical trials, new TS inhibitors nolatrexed, confirmed that its has a good therapeutic effect of a variety of tumors, but in the short-term uses of TS expression induced increases in tumor cells in their resistance. Li Yilei [15] study to reduce the TS induced to reduce the resistance of tumor cells.

    2.3 TS gene polymorphism and cancer chemotherapy

    Genetic pharmacological studies found that patients on the drug sensitivity largely depends on the patient’s genetic structure. Gene polymorphism can be caused by different individuals to produce different effects in the administration of Pharmacology and Toxicology, causing a difference in effects of the drug treatment. TS gene mRNA stability and translation efficiency differences, and can lead to cancer patients of different TS genotype 5  Fu based chemotherapy. Most scholars believe that TS genotype as a predictor of tumor therapy. Yawata et al [16] susceptibility studies show: 2R/2R and 2R/3R cell lines, than the of 3R/3R groups on FudR sensitivity higher; 2R/2R, 2R/3Rc 3Rc / 3Rc group (low TS expression group) The cell lines FudR higher sensitivity than 3Rg/3Rg group (high TS expression group), indicating that the repeat sequence 5 ‘ UTR genotyping helps guide individuals fluorouracil chemotherapy. Gao Changming et al [17] found that carrying TS  6 /  6bp  6 / +6 bp genotype in patients with chemotherapy significantly higher than +6 / +6 bp genotype patients. Villafranca F, et al [18] also believe that TS can be used as a predictor of tumor downstaging repeat polymorphism. TS gene polymorphism may formulate chemotherapy and prognosis assessment provides a theoretical basis and guidance.

    2.4 TS gene polymorphisms predict tumor chemotherapy limitations

    Many scholars believe that TS expression levels and TS genotype testing can predict the sensitivity of cancer patients on chemotherapy drugs and chemotherapy. However, data show the the TS expression fluorouracil validity negatively related conclusion does not hold. Lecomte [19] 5 ‘ UTR of polymorphism and 3’  UTR of polymorphism has nothing to do with the 5  Fu efficacy and survival study of 90 patients with colorectal cancer.

    Such opposite view, on the one hand, may be provided during the experiment, the electrophoresis conditions differences and digested completely and produce false negative, influence the detection result of the TS genotype, resulting Conclusion inconsistent; the other hand, affect tumor to 5  Fu sensitivity to a variety of factors. 5  Fu metabolism closely related to the other enzymes should be detected in addition to detect the TS thymidine phosphate cyclase (TP), thymidine kinase (TK), dihydropyrimidine dehydrogenase (DPD) and is not detected, and thus has certain limitations. p53, multidrug resistance gene and multidrug resistance-associated gene and other genes are also influencing factors of the efficacy of chemotherapy.

    2.5 TS resistance reversal

    Clinical failure of chemotherapy because the tumor cells become resistant to find resistance reversal agents is one important strategy for anticancer drugs. Reversal TS high expression of resistance used ECTA program, 5  Fu combined with leucovorin (LV) or IFN  γ. ECTA program NB1011 enzyme-catalyzed non-toxic reagents. The non-toxic reagents catalyzed by the TS have toxic, so high expression of TS deadly drug-resistant cancer cells, while leaving normal cells lethality weak, the TS-induced resistance effect is remarkable, and is an ideal reversal agent. Associated with LV or interferon  γ can reduce the TS induced and avoid secondary resistance occurs. Guided tumor cell apoptosis is the ultimate way of chemotherapy, 5  Fu inhibit TS, possibly through the tumor cell surface Fas receptor and Fas ligand binding guide apoptosis [20]. This involves the process of signal transduction, and thus have been proposed immunotherapy.

    The TS inhibitors chemotherapy, tumor cells can be the TS induced resistance. Ferguson [21] was designed with the antisense oligonucleotide complementary TSmRNA3 ‘ UTR nucleotide the TSmRNA level was inhibited by 70%, cell proliferation was inhibited more than 40%. Both combination therapy may overcome the TS upward due to drug resistance. Peters and other studies have shown that The TS activity guide tumor resistance, significantly enhanced in the combined presence of high DPD activity; advanced colon cancer, low TSmRAN / the low DPDmRNA by 5  FU and leucovorin, high TSmRNA / high DPDmRNA The use of oxaliplatin and CPT  Ⅱ, achieved good clinical efficacy [22].

    2.6 gene therapy

    Suicide gene therapy tumor gene therapy, especially attracting much attention, the principle is the suicide gene into tumor cells, the special of the gene encoding the enzyme, the original non-toxic prodrug metabolism in tumor cells to toxic products, causing these cell suicide. The tumor suicide gene therapy for clinical, the key is to make specific expression of target gene in the tumor cells to avoid introducing the normal tissue cells, to ensure the security of gene therapy. To this end, in front of the suicide gene connected to a tumor-specific promoter, showed a tumor-specific expression of the suicide gene. Waves, etc. [23] TS gene promoter, p16 gene promoter recombinant plasmid vector into drug HR  8348 cells results show that TS and p16 double promoter bootable TK gene targeted killing 5  Fu resistant tumor cells protect the body’s normal cells, to improve the safety of suicide gene therapy. Sue Wong [24] by retroviral vector-mediated RNA interference (RNAi) technology can indirectly inhibit expression of TSmRNA, new ideas and tools for high expression of TS gene therapy of cancer.

    2.7 TS, TS gene polymorphism with tumor invasiveness and prognosis

    Many studies have shown that tumor TS expression levels and the patient’s prognosis was negatively correlated with the expression of TS can be used as indicators to determine the prognosis of cancer patients. Yu Gang, [25] detected 164 cases of gastric cancer specimens showed that TS expression levels correlated with the degree of tumor differentiation, depth of invasion, lymph node metastasis or not and clinical pathological stage is closely related. TS high expression indicates that the tumor has strong invasive the TS gene products with low expression patients with local recurrence and distant metastases were significantly lower than patients with high expression. Its role in DNA synthesis, overexpression TS tumor cell population might have the potential advantages of high TS expression with poor prognosis may be a reflection of the state of cell proliferation. Scholars have come to different conclusions, the Board Chih-Ming et al [26] 51 patients with esophageal cancer specimens TS expression detection results show that TS expression levels in patients with lymph node metastasis and clinical pathological stage.

    The molecular characteristics of the tumor lesions decided a malignant tumor characteristics, transfer characteristics, characteristics of recurrence, the fundamental basis of tumor prognosis. Chih-Ming Dong et al [26] studies suggest that TS 5 ‘ UTR of polymorphism may be used as molecular indicators predict the ability of lymph node metastasis in esophageal squamous cell carcinoma. Visible population genotype may be associated with tumor grade, stage, predict tumor prognosis; molecular typing of tumors, doctors prognosis of the condition and trends may cause more detailed, more correctly, the monitoring of the efficacy The situation is more clear.

    3 Conclusion and Outlook

    Chemotherapy has important value in comprehensive cancer treatment, drug resistance is still a major problem in cancer chemotherapy. Different scholars reported conclusions differ, which may vary with the study population, age, geographical, end point and method. Organization of large randomized controlled study, system assessment, further study of the TS gene polymorphism with tumor relationship, will help to reveal the mechanism of the resistance of tumor cells. According to TS, TS genotyping to guide individual chemotherapy blindness, can improve the relevance and effectiveness of medication, reduce medication and its side effects. Many scholars believe, that TS genotyping is expected to predict tumor, chemotherapy assessment, prognosis indicators for Cancer Prevention and bring about a better future. Individualized medicine in the future, I believe that will be built on the basis of evidence-based medicine, the individualized tumor genomics and proteomics relying polyclinics mode. At home and abroad of the TS gene 3 ‘ UTR of polymorphism, SNP and tumor genomic instability reported, TS related drug resistance reversal and little TS gene therapy research, pending further study.