Qiang Lv,Yuan-Hong Zhao,Yu Lu,Rui-Xue Yang,Yuan-Tao Zhong,Jie Wang,Zhi-Li Zhao,Long Zhang

1Department of oncology,the First Teaching Hospital of Tianjin University of Traditional Chinese Medicine,Tianjin,300385,China.

Abstract

Objective:To investigate the expression of serum thymidine kinase 1 (TK1) in malignant tumors and its relationship with common tumor markers.Methods:195 cancer patients in the First Teaching Hospital of Tianjin University of traditional Chinese medicine were selected as the subjects.The serum TK1 concentration was measured by enzyme-linked immune-electrotransfer blot and the correlation between TK1 concentration and some biological parameters of tumor patients were analyzed retrospectively.Results:The positive rate of serum TK1 in patients with tumor stage IV was higher than that in other stages (29.9% ＞16.4%, P = 0.029＜0.05).The positive rate of serum TK1 in patients with carbohydrate antigen 199 (CA199) positive was higher than the negative(43.5%＞19.6%, P=0.036＜0.05).There was a positive correlation between serum TK1 and CA199 (r=0.244,P=0.043＜0.05).Conclusion:The expression of TK1 in serum is related to the proliferation of tumor cells.It is consistent with the concentration of CA199,which can be used as a combined choice for the study of digestive tract tumors,especially pancreatic cancer markers.

Keywords:Serum thymidine kinase 1,Tumor marker,Tumor,Clinical,Digestive system

Background

Thymidine kinase 1 (TK1),also known as cytoplasmic thymidine kinase,is the isoenzyme of mitochondrial thymidine kinase 2 (TK2),which are two forms of thymidine kinase in cells.As a key enzyme closely related to cell proliferation and the salvage pathway of DNA,it catalyzes the transformation of deoxythymidine to deoxythymidine monophosphate[1].In the cell cycle,TK1 began to increase in the G1 phase,and gradually increased in the S phase until the G2 phase.The concentration of TK1 was consistent with the increase of DeoxyriboNucleic Acid (DNA)synthesis so that it can be used as a cell cycle-dependent marker.Tumor cells are the abnormal proliferation of cells.TK1 will be released into blood and tissues after its cleavage,so the detection of serum TK1 can effectively reflect the proliferation activity of tumor cell proliferation.And its normal range is 0.0-2.0 pmol/L.At present,the detection of serum TK1 is considered to have clinical application value in screening tumor,diagnosis and treatment evaluation.The literature of its research has been found in the diagnosis and treatment evaluation of breast cancer,lung cancer,digestive tract tumor and hematologic tumors [1,2].To further clarify its application value in tumor diagnosis and treatment,this study detected the expression of serum TK1 in tumor patients,and explored the relationship between TK1 and common biological parameters.

Materials and methods

Clinical data

We retrospectively analyzed 195 cases of patients with a malignant tumor in the oncology department of the first teaching hospital of Tianjin University of traditional Chinese medicine from December 2015 to January 2020.There were 89 males and 106 females.And their ages range from 29 to 90,with a median age of 65.There were 7 head and neck tumors,69 digestive tract tumors,61 lung tumors,20 breast tumors,18 gynecological tumors,9 urinary system tumors and 11 other kinds of tumors.

Materials and testing methods

Serum TK1 was detected by enzyme-linked immune-electrotransfer blot.Venous blood collected from patients who fasted for more than 12h was stored at -20℃ for cryopreservation.Enzyme linked immune-electrotransfer blot was used for the detection,and the experimental operation was strictly carried out according to the instructions.Serum TK1＞2.0 pmol/L was treated as positive.

Detection of tumor markers by chemiluminescent immunoassay: taking 3 mL of fasting venous blood,centrifugation it for 10 min with 3000 R/min,and then the serum was used for the test.Carbohydrate antigen 199 (CA199),carbohydrate antigen 125 (CA125),carbohydrate antigen 153 (CA153) and cytokeratin 19 fragments in serum were detected by automatic microparticle chemiluminescence immunoassay.The experiment was carried out in strict accordance with the instructions.CA199＞37 U/mL was positive,cytokeratin 19＞2.08 ng/mL was positive,CA153＞30 U/mL was positive,CA125＞35 U/mL was positive.

Statistical methods

Statistical Product and Service Solutions (SPSS) 26.0 software was used for statistical analysis.The Chi-square test or Fisher exact probability method was used to deal with counting data Spearman rank correlation was used to analyze the correlation between the two variables.P＜0.05 was defined as statistically significant.

Results

Relationship between serum TK1 and the general condition of tumor patients

Among 195 patients,41 were TK1 positive,the total positive rate was 21.0%.There was no statistical difference in the positive rate of serum TK1 whatever different ages and genders(Table1).

Relationship between serum TK1 and tumor stage and metastasis

There was no significant difference in serum TK1 positive rate in tumor metastasis (P= 0.899＞0.05)and tumor type (P= 0.108＞0.05).The positive rate of serum TK1 was significantly different in the tumor stage (P= 0.029＜0.05).The positive rate of serum TK1 in stage IV was 29.9%,which was higher than that in other stages(16.4%)(Table2).

Relationship between serum TK1 and tumor serum markers

The positive rates of serum CA199,cytokeratin 19 fragments,CA125,CA153 and TK1 were analyzed according to tumor types.The results showed that there were statistical differences between the positive rates of serum CA199 and TK1(P = 0.036＜0.05).The positive rates of serum CA199 and TK1 were consistent.There was no significant statistical difference in the rest(Table3).

The correlation analysis between serum TK1 concentration and CA199 in patients with digestive system tumor showed that there was a positive correlation between serum TK1 concentration and CA199 in patients with digestive tract tumor(r=0.244,P= 0.043＜0.05),Figure1 shows that the trend of serum TK1 concentration is consistent with that of CA199.

Table1 The relationship between age,Gender and TK1 positive rate

Table2 The relationship between tumor characteristics and TK1 positive rate

Figure1 Diagram of the broken line change between TK1 and CA199

Table3 The relationship between the positive rate of tumor markers and TK1 in different tumor types

Discussion

As a kind of cell proliferation marker,serum TK1 has been studied in many aspects such as tumor screening,diagnosis and prognosis monitoring.A retrospective study of 56,286 people's health screening shows that serum TK1 is a reliable biomarker in the screening of malignant tumors,and a meta-analysis shows that serum TK1 combined with other tumor markers has a certain value in the diagnosis of tumors [3,4].In this study,the total positive rate of serum TK1 was 21%.It is lower than other studies,which is related to the fact that the patients in this study have been treated by surgery,radiotherapy and chemotherapy [5,6].According to the trend reflected by the test data,the positive rate of TK1 in the serum of stage IV tumor and stage I-III tumor was compared,and it was found that in stage IV(29.9%＞16.4%,P= 0.02＜0.05) higher than other stage tumor patients.Tumor cells themselves have genetic instability.With the development of the disease,tumor cells mutate and overlap,resulting in obvious heterogeneity of late tumor cells,which will lead to the emergence of tumor cells with stronger invasiveness and faster proliferation.During the accelerated or unregulated proliferation,tumor cells often disintegrate [1].Besides,patients with advanced tumors have a larger tumor load.As a result,the positive rate of serum TK1 in patients with stage IV increased.However,there is no statistical difference between the distribution of metastasis and serum TK1 positive rate.The way of grouping metastasis or not undoubtedly increases the annexation of the tumor stage,which makes the patients in stage IV may be mixed with the patients in non-stage IV.Therefore,the positive rate appears a balance between the two groups.

The positive rate of serum TK1 in different tumor types was compared horizontally at the same time,and there was no statistical significance found.The value of serum TK1 in the diagnosis and prognosis of many cancer types such as breast cancer,lung cancer and liver cancer has been reported now.Some scholars measured the serum TK1 concentration of patients who received iodine-125 particle implantation treatment with non-small cell lung cancer,and the scholars found that the serum TK1 concentration decreased after treatment.It is suggested that serum TK1 can be used as a reference for evaluation [7].The difference in serum TK1 positive rate can not reflect the difference in tumor category.It indicates the serum TK1 can reflect the same value as serum TK1 studies of breast cancer,lung cancer and liver cancer in the study of other cancers,which may provide evidence for expanding the quantitative study of serum TK1.

In this study,we tried to explore the relationship between tumor marker positive rate and serum TK1 positive rate.We found that there was a statistical difference between serum CA199 positive rate and serum TK1 positive rate in patients with digestive system tumors.The positive rate of serum TK1 was higher in CA199 positive patients (43.5%＞19.6%,P=0.036＜0.05).The correlation analysis between serum TK1 and CA199 showed that there was a positive correlation between serum TK1 and CA199 (r= 0.244,P= 0.043＜0.05).CA199 is a kind of glycoprotein on the cell membrane,which is distributed in the pancreas,bile duct epithelium and other places.The rise of CA199 is common in diseases such as liver cirrhosis,gallstone,type 2 diabetes,hepatobiliary pancreatic tumor.It is widely used in the screening and evaluation of clinical gastrointestinal tumors.

In this study,patients with serum CA199 positive had higher serum TK1 positive rate,which provided a basis for the combined detection of serum TK1 in the diagnosis and screening of digestive tract tumors.Wang Wei and other researchers found that the detection of serum TK1 combined with CEA and CA199 can improve the sensitivity of pancreatic cancer diagnosis [8].It has also been shown that the combined detection of serum TK1,carcinoembryonic antigen (CEA),CA199,carbohydrate antigen 724(CA724) has better performance in the diagnosis of gastric cancer and colorectal cancer[9].Serum CA199 is an essential index for the diagnosis and prognosis of pancreatic cancer.Felix K and other researchers compared 404 cases of pancreatic cancer and 136 cases of non-pancreatic cancer,found that the serum TK1 content of pancreatic cancer patients was significantly higher than that of non-pancreatic cancer patients.The follow-up found that the serum TK1 concentration was closely related to the total survival time,which could be used as one of the indicators of pancreatic cancer detection and prognosis evaluation [10].Zhu showed that knockdown of the TK1 gene could inhibit the proliferation of pancreatic tumor cells by inducing p21 up-regulated and mediating S phase arrest [11].The expression of CA199 is restricted to the population with negative expression of le-a and le-b genes,and the distribution of such a population in China is between 5.9% and 23.1%,which makes the detection of serum CA199 in pancreatic cancer patients appear higher false-negative [12,13].Given the positive correlation between serum TK1 concentration and serum CA199 concentration,further study on the relationship between serum TK1 concentration and the Lewis gene is a direction for the diagnosis of pancreatic cancer.

Serum TK1 is a reflection of tumor cell proliferation.Chemotherapy drugs related to cell cycle block will have a significant impact on its concentration.Ying-Zi Xia measured the serum TK1 concentration of patients with stage IV colon cancer before and after chemotherapy.It was found that the serum TK1 concentration decreased significantly after chemotherapy,which can be used as an indicator of therapeutic effect evaluation[14].On the other hand,it shows that the treatment of tumor patients will lead to the misjudgment of the positive rate of serum TK1.Besides,some studies show that the mutation of the TK1 gene sequence has a certain distribution difference in some codon polymorphism between breast cancer and colorectal cancer [15],which contradicts the conclusion that the difference of serum TK1 positive rate between tumor categories is not apparent.It can be seen that exploring the treatment process and primary biochemical indexes of the patients are conducive to examine further the status of serum TK1 in different tumor patients.In contrast,the study on the mutation of the TK1 gene sequence and tumor-related genes in different tumor subtypes still need to be further explored and improved.We believe that with the deepening of research,the specific mechanism of serum TK1 changes will provide a more in-depth theory for revealing the pathogenesis of tumors,and it can make clinical diagnosis and treatment more timely and accurate.