Tumor Markers for Diagnosis , Monitoring of Recurrence and Prognosis in Patients with Upper Gastrointestinal Tract Cancer

EC, Cardia cancer and GC are three of the most commonly seen malignant tumors of GIT cancer. EC is one of the most aggressive neoplasms. One characteristic of EC is its incidence diversity, with high indices in Asian countries and a milder incidence in European and American continents (Zheng et al., 2010; Lin et al., 2013). China is one of the high incidence areas of EC, particular in Henan, Hebei, and Shanxi in Central North China, which have the highest incidence rates in the world. GC is a desease with high morbidity and mortality. Two-thirds of the GC cases occur in developing country. Among them, more than 40% of cases are located in china. Although GC has shown a significant decline in morbidity in recent years, but it still ranks second among


Introduction
EC, Cardia cancer and GC are three of the most commonly seen malignant tumors of GIT cancer.
EC is one of the most aggressive neoplasms.One characteristic of EC is its incidence diversity, with high indices in Asian countries and a milder incidence in European and American continents (Zheng et al., 2010; RESEARCH ARTICLE

Tumor Markers for Diagnosis, Monitoring of Recurrence and Prognosis in Patients with Upper Gastrointestinal Tract Cancer
Jie-Xian Jing*, Yan Wang, Xiao-Qin Xu, Ting Sun, Bao-Guo Tian, Li-Li Du, Xian-Wen Zhao, Cun-Zhi Han all malignant tumors in china and is younger.In contrast with GC, cardia cancer has shown a relatively increased in incidence.Due to the anatomical characteristics of cardiac, cardia cancer are diagnosed already at advanced stages of the disease.Most of treatment outcomes of patients have been poor because the disease has already progressed to an advanced stage by the time it is diagnosed.Consequently, various tumor markers have been used to detect cancer at an early stage and monitor cancers.
However, the sensitivity of one TM is low, but the combination of these TMs has not been used to evaluating curative effect and prognosis of upper GIT cancers, especially all the EC, GC and cardia cancer.We conducted the present study to explore the relationship between the clinical characteristics of patients with upper GIT cancer and TMs, and to evaluate the predictive and prognostic value of preoperative serum levels of TMs for upper GIT cancer.

Patients and blood samples
The study included 573 patients who underwent surgical resection for primary upper GIT cancer between January 1, 2004 and December 30, 2007.Of the 573 patients, 463 underwent radical surgery and 110 underwent palliative surgery, with a median age of 58.4 years (range 29 ~ 81 years).Among them, 127 cases were EC, 264 were GC and 182 were cardia carcinoma.In 573 patients, 129 underwent comprehensive treatment after operation, which included stage I (n=3), II (n=20), III (n=82) and IV (n=24), according to the International Tumor Node Metastasis (TNM) staging system.The TNM staging for EC was performed according to NCCN (2002); cardia cancer and GC according to the AJCC (2003).All cases were confirmed by pathological histology and cytology examination.No chemotherapy and radiotherapy was accepted preoperation.No main organ dysfunction was found in these patients and normal bone marrow, liver and renal functions were assessed for inclusion in the study.Patients characteristics are presented in Table 1.

Follow-up
Overall, 360 patients were followed-up at the outpatient clinic after hospital discharge.213 cases were lost to follow-up.The follow-up system consisted of measurement of serum TMs routinely at 3-month intervals for the first year, and at 6-month intervals thereafter.The follow-up program included: clinical examination, hematological analyses and TM assay at each checkup; abdominal ultrasound and chest x-rays were scheduled every 6 months.Criteria for the establishment of recurrent disease included histological confirmation, palpable disease, or disease evident radiologically with subsequent clinical progression and supportive biochemical data.The follow-up end-date was Mar 1, 2010.All survival patients were followed-up for at least 36 months.138 patients died during the follow-up period.

Tumor marker assay
Peripheral blood from patients was obtained at the preoperative workup and three months after comprehensive treatment.And the samples were collected and then centrifuged for 10 minutes at 3000 rpm; serum was then immediately separated for examination.Serum CEA, CA19-9, CA24-2, AFP, SCC, CA72-4, TPA and TPS were measured by ELISA using TECAN and reagent kits (Sweden and IDL Biotech), with CEA>3μg/L, CA19-9>20 U/ml, CA24-2>12 U/ml, AFP>15μg/L, SCC>1 ng/ ml, CA72-4 >5 ng/ml, TPA>2 ng/ml and TPS>150 U/L being regarded as elevated status.The cut-off values of these TMs were previously established by our laboratory by taking into account several factors (as diet, living conditions and patient selection).

Statistical analyses
Statistical analysis was carried out using SPSS 16.0 software.The TMs levels of two groups were compared using Wilcoxon test; differences in several serum markers' levels were evaluated by Kruskal-Wallis test.Correlation   between CEA, CA19-9, CA24-2, CA72-4, TPS, TPA and SCC was used Spearman analysis.The two groups were compared by cross-table analysis using Pearson's chi-square test.Overall survival rates were calculated by the Kaplan-Meier method, and the differences in survival rates were analyzed by the log-rank test.P-value of less than 0.05 was considered to be statistically significant.
The follow-up time for this study was from 6 months to 38months.All survival patients were followed-up for at least 36 months.Overall 138 patients died during the follow-up period.The levels of TMs generally increased during follow-up while metastasis and/or deterioration increasingly occurred.The longest time interval between the increasing levels of the TMs and the appearance of

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the imaging features was 124 days.Elevated marker levels indicated poorer survival.Multivariate analysis indicated that preoperatively CA72-4, CA24-2 and SCC were prognostic factors of cardiac carcinoma, GC and EC, respectively.Median survival time for cardiac carcinoma with negative preoperative CA72-4 was 33.6 months, and was 16.3 months for patients with preoperative CA72-4 positive levels (p<0.005).Similarly, the median survival time for GC with negative preoperative CA24-2 was 18.1 months, and was 10.9 months for patients with preoperative CA24-2 positive levels (p<0.001), while the median survival time for esophagus cancer was 27.4 months and 16.5 months in patients with SCC negative levels and positive levels (p<0.005)(Figure 2-4).

Discussion
EC, cardia cancer and GC are common malignant upper GIT cancer in the world.In China, the incidence and mortality rates are more than twice of the world average.Shanxi is a high incidence area of upper GIT cancer.Among these, 90% of EC are esophageal squamous carcinoma, whereas most of cardiac cancer and GC are adenocarcinoma.As the cancers occured mostly in remote areas, most patients are diagnosed already at an advanced stage.
Surgery is the main approach for upper GIT cancer, and the most important prognostic factor of upper GIT cancer is tumor node metastasis (TNM) classification (Edge et al., 2010).However, it is difficult to obtain complete data preoperatively.For this reason, it may be important to find some other preoperative prognostic factors for evaluating the outcome of upper GIT cancer patients.TMs have been used for the early screening, diagnosis, evaluating prognosis, monitoring curative effect, and detecting relapse in patients of cancer (Bates et al., 1991;Nishimaki et al., 1999;Nishimaki et al., 1999;Zhang et al., 2009).However, each TM has its limitation in terms of diagnostic value, especially for the early diagnosis (Marrelli et al., 2004;Patriti et al., 2007).It is therefore necessary to find new molecules and combination of several of these TMs at the same time (Wang et al., 2005;Gao et al., 2007;Gupta et al., 2007;Park et al., 2007;Gao et al., 2007).
In the study, we detected serum levels of CEA, CA19-9, CA24-2, AFP, CA72-4, SCC, TPA and TPS    (Tian et al., 2014).In the present study, we found the most sensitive combinations of tumor markers were CEA+CA199+CA242+SCC in EC; while CEA+CA199+CA242+CA724 proved to be a better evaluation indicator to cardiac cancer and GC.Therefore, these tumor markers can be used for auxiliary diagnosis of patients with upper GIT cancer and preliminary judgment for pathological types.
Our results showed that CEA and CA72-4 levels were significantly higher in male patients than in female, whereas CA19-9 and CA24-2 levels in female were significantly higher than in male, which was similar to our previous finding.It needs to be further discussed.
Spearman analysis showed that preoperative level of CA19-9 significantly correlated with CA24-2 (r=0.810,p<0.001), as there may be similarities in the mechanism of generation of the two markers, suggesting that a combination test of CA19-9 and CA24-2 can improve the possibilities for the screening or monitoring disease recurrence and response to treatment.
Compared with preoperative levels, postoperative TMs levels significantly decreased.When metastasis and recurrence occurred, the CEA, CA19-9, CA24-2, SCC, and CA72-4 levels increased again in compared with postoperative concentrations.Our findings are similar to those of Takahashi et al. (2003) who reported that in most patients with high preoperative TMs levels, these TMs increased again at recurrence or metastasis.The finding indicated that patients, who with high preoperative tumor markers levels, and/or at stage II/III/ IV need comprehensive treatment.The levels of TMs generally increased during follow-up while metastasis and deterioration increasingly occurred and were related to the burden of the tumor.The elevated levels of TMs appeared earlier than the sensitive imaging results when the tumor recurred, with the longest interval being of 124 days.Therefore, these TMs could play an important role in the monitoring of tumor recurrence and metastasis, thus allowing identifying symptomless patient with tumor recurrence by routine postoperative serum TMs checkups.

Figure
Figure 1.Sensitivity of TMs in Upper Gastrointestinal Cancer I n c o n c l u s i o n , c o m b i n e d d e t e c t i o n o f CEA+CA199+CA242+SCC proved to be the most economic and practical strategy in diagnosis of EC; CEA+CA199+CA242+CA724 proved to be a better evaluation indicator to cardiac cancer and GC.The CEA and CA19-9, CA24-2, CA72-4 and SCC, examined postoperatively during follow-up, were useful to find early tumor recurrence and metastasis, and evaluate prognosis.AFP, TPA and TPS have no significant value in diagnosis patients with upper GIT cancer.

Table 2 . Sensitivity and Specificity of Combination of TMs in Detecting EC, Cardiac Cancer and GC
.7314/APJCP.2014.15.23.10267Tumor Markers for Diagnosis, Monitoring of Recurrence and Prognosis in Patients with Upper Gastrointestinal Tract Cancer *bCompared with a, p<0.05; b Compared with c, p>0.05; e Compared with d, p<0.05; e Compared with f, p>0.05 Asian Pacific Journal of Cancer Prevention, Vol 15, 2014 10269 DOI:http://dx.doi.org/10

Table 3 . Relationship between Positive Rate of TMs and the Pathological Types, TNM Staging in Upper Gastrointestinal Cancer Patients (%)
*compared with squamous cell carcinomas; **compared with I+II and IV; ***compared with I+II and III

Table 4 . Correlation between Serum Levels of TMs in Upper Gastrointestinal Cancer Patients and Sex and Age (M+Q)
*M (Median); Q (Interquartile range)

Table 5 . Serum Levels of TMs Pre-and Post-operation (M+Q)
DOI:http://dx.doi.org/10.7314/APJCP.2014.15.23.10267Tumor Markers for Diagnosis, Monitoring of Recurrence and Prognosis in Patients with Upper Gastrointestinal Tract Cancer in 127 patients with EC, 182 with Cardiac cancer and 264 with GC.Different studies have reported different rates of these tumor markers.Whatever Combination of multiple markers can improve overall detection sensitivity.Tian et al. indicated CEA, CA19-9, CA24-2 and CA50 maybe better combinations in improving diagnostic accuracy of GC