Research ArticleResearch Article

The Relationship between Serum VEGF Expression and PET/CT Images in TNM-staging of Lung Carcinoma

Zhengping Xiong, Yangde Zhang, Zhaoyu Liang, Aiming Xie, Qing He and Shuren Yang
Chinese Journal of Clinical Oncology October 2007, 4 (5) 313-317; DOI: https://doi.org/10.1007/s11805-007-0313-3

Abstract

OBJECTIVE To analyze the relationship between the characteristics of PET/CT images for lung carcinoma (LC) TNM staging and the expression of serum VEGF protein.

METHODS PET/CT examinations were performed before treatment of 53 patients with LC. The expression of serum VEGF protein was examined using a quantitative sandwich enzyme-linked immunosorbent assay (ELISA R and D system). The relationship was analyzed between PET/CT images for LC T-staging and metastasis (lymph nodes and distance) and serum VEGF expression.

RESULTS Based on PET/CT images for LC T-staging, 11 cases were staged as T1, 9 as T2, 18 as T3 and 15 as T4. Mediastinal nodal metastases were found in 22 patients, and distance metastasis in 9. The serum VEGF level in the LC patients was (378.02±180.79) ng/L, showing that there was a significant difference between the patients and healthy subjects (P<0.05). There was no significant difference in the level of serum VEGF between different low T-staged (T1, T2) patients. However, the level of serum VEGF was significantly different between the low T-staged (T1, T2) and high T-staged (T3, T4) patients (P<0.05). The level of the serum VEGF protein in the patients with mediastinal nodal metastasis was (561.50±104.55) ng/L, and indicating that there was a statistical significance (t=12.21, P<0.05) compared to those in the non-metastatic group. The level of serum VEGF in the patients with distance metastasis was (614.11±158.81) ng/L, demonstrating that there was a significant difference (t=5.30, P<0.05) compared to those in the nondistant metastatic group.

CONCLUSION ①There is a high level of serum VEGF expression in LC patients. ②There is a correlation between metastases (lymph nodes and distance) and the level of serum VEGF. ③With an upgrade of the TNM-stage, the level of serum VEGF protein is elevated.

KEYWORDS:

keywords

INTRODUCTION

Lung carcinoma (LC) has a high incidence in China and means to improve its diagnosis are of great interest. PET/CT technology is a functional molecular imaging system which integrates positron emission computerized tomography (PET) and X-ray computed tomography (CT), thus forming a synthetized fusion image with an integration of high resolution and complementation between structure and metabolic activity. Application of PET/CT improves the accuracy for LC diagnosis. Further studies on the relationship between PET/CT images (TNM-staging) and serum VEGF expression in LC can reflect energy metabolism of the tumor as well as display angiogenesis, the degree of metastatic spread for clinical staging and for radiotherapeutic treatment planning. Thus, this system can provide a theoretical basis for tumor prognosis.

MATERIALS AND METHODS

General data

Data were collected from 53 lung-cancer patients, diagnosed and treated in our hospital during the period from July 2005 to the beginning of 2007. These patients received no treatment before their first PET/CT examination in this hospital. Among the patients, 37 were male and 16 female, with ages ranging from 14 to 78, and a mean of 43.7. Pathological diagnosis showed that 42 of the total cases suffered from squamous cell carcinoma, while another 11 were diagnosed with LC of other pathologic classifications.

Methods of PET/CT examination

Equipment and parameters

18F-FDG with a radiochemical purity of over 95% was used as the PET developer. The GE Discovery-type PET/CT scanner was utilized in the diagnosis of all the cases. The patients were asked to fast over 5 h to stabilize the blood sugar level before scanning. An i.v. dose of 4 to 8 mCi of 18F-FDG, was then administered and the images taken, after lying in bed for one hour. Conventional transection scanning (thickness of 4 to 6 mm, interlamellar space 3 mm and pitch 1.0), was performed using a sagittal plane and coronal reconstruction imaging. Iohexol was used for enhancement scanning, (0.1 mmol/kg bolus injection via an ulnar vein) employing conditions like a plain scan.

T-staging standard

The 1997 TNM clinical staging of lung cancer was adopted, i.e. T1 meant that the maximum diameter (Max. Diam.) of the tumor was less than 3.0 cm. T2 meant that the Max Diam of the tumor was larger than 3.0 cm and that there was an encroachment on the main bronchus, with a space of 2.0 cm or more away from the eminence, encroachment of the visceral pleura, and the tumor was involved to the hilum of the lung, with a pulmonary atelectasis (not total lung). T3 meant that the tumor of any size was involved in any of the following sites, i.e. the chest wall, diaphragm and mediastinal pleura. T4 meant that the tumors of any size were involved in any of the following organs: heart, great vessels, trachea, esophagus and vertebral body.

Evaluation of PET/CT images for metastasis of the lung hilum and of the mediastinal lymph nodes

When the standard uptake value (SUV) was larger than or equivalent to 2.5, it usually was considered to be a lymph node metastasis.

M-staging standard

M0 meant there was no distant metastasis and M1 a distant metastasis.

Detection of serum VEGF expression

Drawing blood

The following procedures or conditions were used: the blood was taken via the median vein of the elbow in the morning, following overnight fasting and centrifuged at 2000 r/m for 5 min. The serum was then collected and stored at -40°C.

VEGF determination

Streptomycin antibiotin antibody-peroxidase enzymelinked immunosorbent assay sandwich method (ELISA) was used to determine the serum VEGF level.

Control group

The serum VEGF level of tumor-free 20 healthy subjects, was used as a control.

Experimental group

The serum VEGF level in 53 lung cancer cases was determined.

Statistical analysis

The SPSS for windows 10.0 was used for statistical analysis of the results. Statistical significance was accepted when the P value was less than 0.05.

RESULTS

Image development

①T-staging of 53 patients receiving a PET/CT examination: there were 11 cases at Stage-T1, 9 at T2, 18 at T3 and 15 at T4. ②Lymph node metastasis: mediastinal hilar lymph-node metastases from the lung cancer were detected in 22 patients. ③Distant metastases: bone metastases were found in 5 cases, liver metastasis in 1, adrenal metastasis in 1 and brain metastasis in 2 (Figs.1~3).

Fig.1.
Fig.1.

Male, 70, T4N2, left-lung cancer, mediastinal lymph node, pleural metastasis.

Fig.2.
Fig.2.

Male, 34, leftupper lung cancer, bladebone metastasis.

Fig.3.
Fig.3.

Male, 42, leftupper lung cancer, supraclavicular lymph node and liver metastasis.

Relationship between TNM staging and serum VEGF levels

The expression of serum VEGF: data from 20 LC patients were randomly drawn and compared to the healthy subjects in the control group. The comparison showed that there was an abnormal over-expression of VEGF in the LC patients, indicating that there was a significant difference between the LC patients and the controls (P<0.05, Table 1).

View this table:
Table 1.

Contrast in VEGF expression between the LC group patients and the healthy subjects.

Relationship between the T-staging and serum VEGF levels

It was found, using analysis of variance, that there was a statistical significance in the serum VEGF level among the groups with stage-T1, T2, T3 and T4 (F=38.84, P=0.00). Employing the q-test, it showed that there was an increase in the serum VEGF level in the stage-T3 and T4, compared to the stage-T1 and T2 groups. This difference between the two groups was significant (P<0.05), but there was no significant difference in the serum VEGF level comparing the satge-T1 with T2 patients (P>0.05, Table 2).

View this table:
Table 2.

Comparison between T-staging and serum VEGF expression.

Relationship between lymph node metastasis and VEGF serum levels

Using PET/CT examinations, mediastinal lymph node metastases were found in 22 cases. These cases were divided into two groups, i.e. one with a mediastinal lymph node SUV of larger than or equivalent to 2.5, and another with SUV of less than 2.5. There was a significant correlation between the lymph node metastases and the serum VEGF level (t=12.21, P<0.05, Table 3). Determination of the correlation between lymph node size and the SUV, suggests that part of the lymph node gradular enlargement might be caused by reactive hyperplasia and not by metastases (Table 4).

View this table:
Table 3.

Comparison between lymph node metastasis and the serum VEGF level (Cases).

View this table:
Table 4.

Comparison between size of lymph node and SUV (Cases).

Relationship between distant metastasis and serum VEGF levels

The total cases were divided into two groups, i.e., a group without distant metastases and those with distant metastases. There was apparent correlation between distant metastasis and serum VEGF expression, showing that there was a significant difference in the VEGF level between the two groups (t=5.30, P <0.05, Table 5).

View this table:
Table 5.

Relationship between distant metastases and serum VEGF levels.

DISCUSSION

Production of images with PET/CT scans

Correct diagnosis using plain X-ray films and CT scanning can be made for most pulmonary lesions, however, there is still some difficulty in determining the complete nature of a portion of these lesions. Application of PET-CT examinations improves the accuracy of diagnosis. PET/CT is a functional imaging system formed by an integration of positron emission tomography (PET) with X-ray computed tomography (CT). PET/CT is based on the specific functional PET image, and is combined with high-end CT technology for precise localization, forming a synthetized image with an integrative complementation between a fine resolution and the metabolic imaging.

Melloni et al.[1] reported that PET/CT imaging was quite important in the diagnosis and treatment of LC. Wang et al.[2] indicated that the accuracy for detection of a LC site and metastasis was 100% using PET/CT with their first 18F-FDG PET examination. PET/CT imaging was superior to simple PET and CT, with a sensitivity of 95.2%, 90.4% and 73.8%, respectively. Gu et al.[3] reported that sensitivity of PET/CT staging was 96.9%, while its accuracy was 90.6%. A study by Zhang et al.[4]showed that the diagnostic accordance rate of serum T-antigen detection in combination with CT scan and PET-CT images for LC patients was 67.44% and 90.70%. The positive rate of regional LC focus was 86.05% and 95.35%, and the positive rate of mediastinal lymph node detection was 65.12% and 83.72%, respectively.

Synthetized PET/CT image can concurrently demonstrate the metabolic status of pulmonary tumors and their anatomic localization, providing a sensitivity and specificity that is markedly higher compared to conventional CT and PET. Studies of Yi et al.[5] have shown that PET/CT enjoyed a better accuracy and specificity, compared to spiral CT, at 90% and 100%, respectively. Zhu et al.[6] suggested that there was close correlation between the degree of VEGF expression in non-small cell lung cancers (NSCLC), and that lymph node metastasis could be used as one of the biological parameters for predicting prognosis in the NSCLC patients. However, the PET/CT images indicated that there was no correlation between LC TNM staging and level of serum VEGF. PET/CT can distinguish direct invasion by a tumor and lymph-node metastasis, and can change TNM staging of the tumor. A PET/CT examination can provide information for more exact staging of LC patients, and thus eliminate non-cancerous tumor composition. This allows a better understanding of the tumor encroachment, and aids in defining the radiotherapeutic field. The most common metastatic site for LC is the mediastinal lymph node of the lung hilum. In routine examinations, the size of lymph nodes is usually regarded as the standard of lymph-node metastasis. At present, a diameter of over 1.0 cm is an indication of tumor metastasis. The authors suggest that there might be a false positive situation by accepting this standard. The SUV of part of the mediastinal lymph nodes of over 1.0 cm which was a hypometabolic region, being displayed in the PET/CT examination, was less than 2.5. So, part of the lymph node swelling was reactive hyperplasia.

Relationship between serum VEGF level and TNM staging

The level of serum VEGF in the LC patients: The level of serum VEGF usually is very low in the healthy people, and an over-expression can occur when there is a cancer metastasis in the human body. Most previous reports[5-7], suggested that VEGF was an important factor for promoting angiogenesis. The level of serum VEGF in cancer patients was an over expression, and was significantly higher compared to the healthy subjects. The level of serum VEGF in advanced LC patients was significantly higher compared to early LC patients. In addition it was higher in LC patients with lymph node metastasis and distant metastasis compared to those without metastases. There was no obvious correlation of the serum VEGF level with the sex, age and LC histopathological types. The data from our study showed that the serum VEGF level in the LC patients was higher, compared to the healthy control group, which was in accordance with most previous related literatures.

Relationship between the TNM staging of LC and serum VEGF expression

Zhu et al.[6] reported that the level of VEGF expression was higher in Stage-III patients compared to tose in Stage-I and II, and there was no obvious correlation between VEGF expression and pathological types. Studies by Liu et al.[7] showed that the level of serum VEGF expression (386.45±112.38) was markedly higher in LC patients compared to a control group (108.26±11.37, P<0.01). The serum VEGF level was respectively 621.41±76.71 and 636.48±160.20 in the Stage-IIIb and IV LC patients, which was apparently higher, compared to 264.12±172.08, 298.56 ±134.17 and 305.43±89.26 in the Stage-I, II and IIIa patients, respectively (P<0.01). The VEGF level in a group with a survival period of over 6 months was 276.32±56.47, which was significantly lower compared to 652.20±65.34 in a group with a survival period of less than 6 months. The serum VEGF level was 410.12±87.23 in cases with adenocarcinoma, 386.78±102.36 in squamous cancer patients, 365.64± 98.47 in small cell cancer patients and 373.91±138.65 in adenosquamous carcinoma cases. There was no significant difference in the VEGF content among various types. The higher the serum VEGF level, the higher the metastatic potency. The serum VEGF level appears to relate to the biological features of LC as supported by our study.

Relationship between LC lymph-node metastasis and the serum VEGF level

Studies of Yan et al.[8] demonstrated that the concentration of serum VEGF was obviously higher in LC patients, compared to a control group. There was no significant difference in the level of serum VEGF between different sexes, ages, histological types and degrees of differentiation, as well as T staging (primary tumors), but there was a significant difference in the serum VEGF content among different N stages (lymph node metastasis). In LC patients, there was a correlation between the serum VEGF level and lymph node metastasis. In our study, there was a correlation between mediastinal lymph-node metastasis and VEGF expression. In studies on the relationship between the size of lymph nodes and metastasis, reactive hyperplasia may play a role in large lymph nodes, which is not a result of metastasis, and thus of no statistical significance. This may be related to other genes that play a major role in the lymph-node metastasis.

Development of LC and serum VEGF

A significant increase in the serum VEGF level in patients with a distant liver or bone metastasis or non-distant metastasis, is an indication of enhanced vasoformation. The VEGF can further angiopoiesis, and stimulate new vascularization, which fosters cancer cell proliferation and metastasis. Over the past few years, many researchers[9-11] have investigated the role of VEGF expression on the prognosis and adjuvant chemotherapy of non-small cell lung cancer. In the group of patients with a distant metastasis, VEGF expression was markedly higher compared to that in non-distant LC metastatic cases, and high VEGF levels were rarely seen in patients with a survival rate of 5 years or over, compared to the cases with survival of less than 5 years. These researchers suggested that the serum level was a good prognostic index for LC lymph-node metastasis and distant metastasis. The study of Vokes et al.[12] aimed at a random three-phase test, using bevacizumab, an anti-VEGF monoclonal antibody, to inhibit lung cancer neovascularization.

Footnotes

  • This work was supported by a grant from the Scientic Fund of the Public Health Bureau of Hunan Province (No. B2005139).

  • Received June 6, 2007.
  • Accepted October 10, 2007.

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