The Diagnostic Efficiency of 99mTc-EDDA/HYNIC-Octreotate SPECT-CT in Comparison with 111In-Pentetrotide in the Detection of Neuroendocrine Tumours

Objective: The aim of this study was to assess the diagnostic efficiency of 99mTc-EDDA/HYNIC-Octreotate in comparison with 111Inpentetrotide scintigraphy in the detection of neuroendocrine tumors. This study also evaluates the impact of SPECT-CT hybrid imaging on somatostatin receptor scintigraphy (SRS) interpretation and clinical management of these tumors. Methods: Fourteen patients were included in the study. All patients underwent a whole body and SPECT-CT imaging with both 99mTc- EDDA/HYNIC-octreotate and 111In-pentetrotide. Images were evaluated both visually and semiquantitatively. Results: On patient basis, the diagnostic results of both studies were similar. The number of lesions detected by 99mTc- EDDA/HYNICOctreotate were higher than the number of lesions detected by 111In-pentetrotide however the difference was not significant (40/43( 93%), 36/43 (83%) p=0.109). Semiquantitative analysis showed higher tumor/organ count ratios for both whole-body and SPECT 99mTc- EDDA/HYNIC-Octreotate scans. Conclusion: The results of this study suggested that, 99mTc- EDDA/HYNIC-Octreotate may be a better alternative to 111In- pentetrotide due to high image quality and lower radiation dose. SPECT/CT is a valuable tool for the assessment of neuroendocrine tumors by providing the precise anatomic localization of scintigraphic findings thus improving lesion detectability and characterization. Conflict of interest:None declared.


Introduction
Neuroendocrine tumors (NET) are a wide group of tumors which can involve different organs and systems. In the diagnosis, treatment and follow up of these tumors different common features such as expression of unique receptors, possession of special metabolic pathways, synthesis and secretion of different materials in peptide and amine structure are utilized. Imaging of NET can be done by various methods of nuclear medicine. The most commonly used target is somatostatin receptors.
The first somatostatin analog designed as a radiopharmaceutical was 111 In-DTPA-octreotide ( 111 Inpentetreotide, Octreoscan®) (1). However, high radiation exposure, production cost of In-111, and the difficulties in supplying are its limitations. Beside these, waiting period up to 48 hours for imaging after the injection hampers patient compliance.
Somatostatin analogs labeled with Technetium-99m( 99m Tc) provide a new perspective in somatostatin receptor imaging. First results showed that 99m Tc labeled somatostatin analogs has a sensitivity and specificity equivalent to 111 In-pentetreotide. Moreover, in some studies it was found to be even better than 111 In-pentetreotide in terms of success in lesion detection. Among 99m Tc labeled somatostatin analogs 99m Tc-depreotide (P-829) was the first radiopharmaceutical which was proven to be valuable for the evaluation of solitary pulmonary nodules (2,3). The other radiopharmaceutical was 99m Tc-HYNIC-Tyr3 octreotide ( 99m Tc-HYNIC TOC) which was granted marketing authorization and made available for the wide use. 99m Tc-HYNIC-Tyr3-octreotate ( 99m Tc-HYNIC-TATE) is another analogue which differs from 99m Tc-HYNIC TOC with terminal amino acid in the peptide sequence and showed higher receptor affinity and better internalization (4). Moreover, it was reported that 99m Tc-HYNIC-TATE has higher affinity to somatostatin receptor subtype-2 than octreotide (5).
Tc-99m has lower energy than In-111 which enables higher dose administration, better image quality and suitable radiation dosimetry. Higher doses of 99m Tc also increase the image quality and lesion detection capability of SPECT imaging. Availability as an instant cold kit and labeling with 99m Tc obtained from an onsite generator enables imaging of the patients on the same day of admission to the Nuclear Medicine departments. Radioguided surgery of neuroendocrine tumors with 99m Tc labeled somatostatin analogs is also more successful compared to radioguided surgery with In-111 because of higher injected dose of the tracer and better physical properties of 99m Tc for the detection with a gamma probe.
Ga-68 labeled PET tracers -in particular 68Ga-DOTATOC, 68 Ga-DOTATATE, and 68 Ga-DOTANOC-have been shown to have higher detection rate compared with conventional SRS and diagnostic CT (6). However, both 68 Ge/ 68 Ga generators and PET scanners are still lower in number than 99 Mo/ 99 Tc generators and SPECT cameras.
In this study we evaluated the accuracy of Tc-99m labelled EDDA/HYNIC-Octreotate using SPECT-CT and compared the results with 111 In-pentetreotide scintigraphy in the same patient population.

Materials and Method Patients
This study has been approved by the Local Ethic Committee and was supported in part by Turkish Scientific and Research Council (TUBITAK) project no: 109S498 and 109S101. Fourteen patients (7 men, and 7 women) with diagnosis of NET who were referred to Nuclear Medicine Department for somatostatin receptor scintigraphy (SRS) were included in the study. All patients gave their informed consent and underwent SRS with 99m Tc-EDDA/HYNIC-Octreotate and 111 In-pentetreotide. Radiopharmaceuticals
Imaging with 111 In-pentetreotide After intravenous administration of 222-250 MBq of 111 In-pentetreotide, whole body, planar and SPECT/CT imaging at 4 th and 24 th hours were carried out using doubleheaded hybrid gamma camera system integrated with x-ray CT (Infinia VC and Hawkeye, GE Medical Systems, U.S.A). Scintigraphic images were acquired with medium energy, parallel hole collimator, set to 172 and 245 keV energy peak, with 20% energy window and CT imaging was acquired in 4.5-5 minutes by taking simultaneous multiple slices in 512x512 matrix in helical mode on, by using 2,5 mA current at 140 keV, provided that the beam scope is 2 cm in each gantry rotation. After CT acquisition, SPECT images were recorded in a protocol of 128x128 matrix, 15-20 seconds/frame rotation with an angle of 360 degree.
Imaging with 99m Tc-EDDA/HYNIC Octreotate was performed within 5-15 days after 111 In pentetreotide imaging. After intravenous administration of 370-400 MBq of 99m Tc-EDDA/ HYNIC octreotate, whole body, planar and SPECT-CT imaging at fourth hour were carried out using double headed hybrid gamma camera system integrated with X-Ray CT (Infinia VC and Hawkeye, GE Medical Systems, U.S.A). Scintigraphic images were acquired with low energy high resolution parallel hole collimator peaked at 140 keV with a 10% energy window. CT images were acquired in 4.5-5 minutes by taking simultaneous multiple slices in 512x512 matrix in helical mode on, by using 2,5 mA current at 140 keV, provided that the beam scope is 2 cm in each gantry rotation. After CT acquisition, SPECT images were recorded in a protocol of 128x128 matrix, 15-20 seconds/frame rotation with an angle of 360 degree.
The image reconstruction was performed using iterative reconstruction (OSEM iteration numbers 2 and 10) on a Xeleris® work station (GE Medical Systems).

Scintigraphic Evaluation
The planar and SPECT-CT images of 99m Tc-EDDA/HYNIC Octreotate and 111 In pentetreotide studies were separately evaluated and compared to each other.
In the visual evaluation, the areas that showed higher uptake than the background were accepted as pathological uptake. CT component of hybrid imaging was used correlatively to verify the localization and characterization of the pathological uptake in scintigraphic studies.
Regions of interests (ROIs) were drawn over the areas which showed pathological uptake in scintigraphic studies for semiquantitative analysis. Identical ROIs were also placed over liver, spleen and right thigh in order to calculate background counts. For semiquantitative analysis of SPECT images, transaxial slices were used, where identical ROIs were placed over areas which showed pathological uptake as well as over liver and kidney and over the area with the least counts in that particular transaxial slice to represent the background. From these calculations tumor/ liver, tumor/kidney, tumor/spleen, tumor/thigh, tumor/ background (for transaxial images of SPECT) activity ratios were obtained. The ratios which were obtained from planar and SPECT images in both imaging studies were compared to each other.
The contribution of planar, SPECT and SPECT-CT images to the diagnosis were separately assessed. The uptake patterns were scored according to whether it can be exactly localized or not. Also another scoring scale was used to assess the intensity of uptake as (0: no suspicious uptake in the imaging area, 1: it is enough to differ between suspicious uptake/physiologic uptake, 2: it is not enough to differ between suspicious uptake/ physiologic uptake).

Statistical Analysis
For the measurable variables (tumor uptake counts, scintigraphic uptake ratios), the correlation between the values was evaluated by using correlation test between independent variables (Spearman test), the assessment of the difference between values were evaluated by the significance of difference between dependent variables (Wilcoxon test).
The assessment of the difference between different imaging methods was performed by using the significance of the difference between dependent variables test (McNemar test).
For all statistical tests that were used, p<0.05 value was accepted as statistically significant.

Results
Indication for SRS imaging was for the staging of five patients who had histopathological NET diagnosis and restaging residual disease/relapse evaluation of 9 patients who had been previously diagnosed and treated. The primary tumor was located in lungs in two patients, in gastrointestinal system in 10 patients, in neck in one patient and in kidney in the rest one patient (Table 1).
All patients were examined by one or more of the other radiological imaging methods which were given in Table 2.
The histopathological diagnosis were bronchial carcinoid, well differentiated neuroendocrine neoplasm, paraganglioma, poorly-differentiated neuroendocrine carcinoma and carcinoid tumor. The serum biochemical analyses were given in Table 2.

Comparison of the Radiopharmaceuticals in Terms of Tumor Uptake
The patient-based visual comparison of the tumor uptake of radiopharmaceuticals revealed that all patients who showed pathological uptake with 111 In-pentetreotide imaging also showed pathologic uptake in the same region with 99m Tc-EDDA/HYNIC-Octreotate imaging (Figure1 A,B). One patient (patient no: 9) who had no pathologic uptake with, 111 In-pentetreotide displayed pathological uptake on 99m Tc-EDDA/HYNIC-Octreotate planar and SPECT images (Figure 2 a,b). 99m Tc-EDDA/HYNIC-Octreotate imaging results were highly concordant with 111 In-pentetreotide imaging. When the concordance between the images with two radiopharmaceuticals were examined in terms of capability of detecting pathological uptake, Spearman correlation factor was found to be 62% (good), which was statistically significant (p<0.05).
The lesion-based comparative analysis revealed that scintigrapy detected 40 out of 43 lesions which were detected with anatomical imaging (contrast enhanced diagnostic CT and MRI together).
Thirty six of 40 lesions (83%) were shown with both 99m Tc-EDDA/HYNIC-Octreotate, and 111 In-pentetreotide. However, 4 lesions were only visualised with 99m Tc-EDDA/ HYNIC-Octreotate. Although the number of tumoral uptake sites were more for 99m Tc-EDDA/HYNIC-Octreotate compared to 111 In-pentetreotide the difference was not statistically significant (p=0.109). Three lesions were only visualized with anatomical imaging (contrast enhanced diagnostic CT and MRI together). Two of these lesions were cervical lymph nodes which were reported as suspected lymph nodes containing calcifications. Another lesion was in the liver but MRI could not distinguish whether the lesion was a metastatic focus or fibrotic area. SRS results were negative for these lesions. But these foci were not confirmed by histopathological examination.
The tumor uptake ratios on both 99m Tc-EDDA/HYNIC-Octreotate and 111 In pentetreotide studies were compared with each other using semiquantitative analysis. In planar images, the tumor/liver and tumor/kidney uptake ratios were significantly higher with 99m Tc-EDDA/HYNIC-Octreotate compared to 111 In-pentetreotide (p=0.015 and p=0.003, respectively Wilcoxon test). On the other hand, the difference between the tumor/spleen and tumor/thigh uptake ratios with 99m Tc-EDDA/HYNIC-

The contribution of 99m Tc-EDDA/HYNIC-Octreotate to Patient Management
The contribution of 99m Tc-EDDA/HYNIC-Octreotate to diagnosis and treatment were as follows: On a patient with the diagnosis of ectopic Cushing's syndrome, (Patient no: 1) the left hilar lymph node involvement was seen on whole-body planar images with 111 In pentetreotide and 99m Tc-EDDA/HYNIC-Octreotate.
Conventional imaging   methods also detected hilar lymph node. On SPECT-CT study, the primary tumor focus was detected on the lower lobe of the left lung with both agents. But conventional imaging methods did not detect the primary focus, most probably due to the localization and anatomical appearance of the lesion. Because the lesion was 10 mm long and not enhanced with contrast which was located at the basal lobe of the lung. After 185 MBq 99m Tc-EDDA/HYNIC Octreotate administration, the patient underwent gamma probe-guided surgery. The primary tumor focus and the hilar lymph node were successfully removed. As a result of orientation to the lesion promptly, surgery and anesthesia time was relatively reduced. Compared to pre-planned area, a significant contribution was achieved to patient morbidity and mortality by excising much smaller area (Figure 3). Following the 99m Tc-EDDA/HYNIC-Octreotate imaging studies, 177 Lu-Octreotate treatment was suggested for four patients who were diagnosed with extensive tumor involvement of the whole body, and 177 Lu-Octreotate treatment was applied for two patients who (gave consent) accepted ( Figure 4).

Discussion
Neuroendocrine tumors are heterogenous family of neoplasms that present with a wide range of morphologic, functional and behavioural characteristics (8). Because NETs have ability to express specific receptors on their cell membrane, such as somatostatin receptors, functional imaging plays an important role for both diagnosis and follow up of these tumors (9). Researchers are working to develop new somatostatin analogues labeled with different radionuclides to get higher diagnostic yield. 99m Tclabeled somatostatin analogs give a new perspective on somatostatin receptor imaging. In our study, a patient based comparison revealed that 99m Tc-EDDA/HYNIC-Octreotate has a similar biodistribution with that of 111 In-pentetreotide, which is consistent with the results of other studies (10). Patients who had tumoral uptake with 111 In-pentetreotide imaging had also 99m Tc-EDDA/HYNIC-Octreotate uptake within the same location.
A lesion based comparison of two agents revealed that 99m Tc-EDDA/HYNIC-Octreotate showed higher number of tumoral involvement especially in the liver (Patient 9).
Similarly other studies in the literature also reported that more metastatic lesions are detected in the liver using 99m Tc labeled somatostatin analogs as compared to 111 Inpentetreotide (10,11). However, the results of our study are not statistically significant probably due to small patient group.
Having ideal imaging characteristics, 99m Tc presents an advantage for SRS imaging compared to 111 In which gives better results with hybrid imaging system SPECT-CT for correct characterization and correct localization of pathological lesions. The correct anatomic location will eliminate false positive results, and improves the diagnostic accuracy. On our study, SPECT-CT detection efficiency of the anatomical localization was significantly higher than SPECT study.
SPECT-CT studies yielded more accurate information in particular the involvement of the abdominal region (12) ( Figure 5).
Semiquantiative analysis for the evaluation of tumoral involvement revealed that tumor/organ uptake ratios with 99m Tc-EDDA/HYNIC-Octreotate on both planar and SPECT studies were higher than 111 In-pentetreotide. It was reported that Octreotate has a higher receptor affinity and internalization capacity due to its terminal threoninol (13). In vitro studies that were done with SS receptor-positive cell culture showed that octreotate chelated with DTPA or DOTA shows 14-17 times more affinity to SStr2 than octreotide, and 8-10 times greater affinity than TOC (5). Due to these features, TATE is also preferred for treatment studies (14). The higher tumor/non-tumor uptake ratios    99m Tc-EDDA/HYNIC-Octreotate SRS . Wide spread metastatic lesions were detected by both agents. The foci at the right lower extremity were thought to be due to contamination. also provide an advantage for imaging one the same day (15). In limited studies, 99m Tc-EDDA/HYNIC-Octreotate was shown to be a promising agent for both imaging and gamma-probe applications (16,17).
In this study we showed that 99m Tc-EDDA/HYNIC-Octreotate is a good alternative for SRS with the advantages of better pharmacokinetic properties, lower radiation dose and higher diagnostic accuracy especially when hybrid imaging methods are used. Figure 5. 34-year-old woman with metastases after resection of primary focus which was in the ileum. a) Whole body planar images revealed pathological tracer uptake in the lower part of the abdomen (arrows). b) The same area was seen with SPECT images but distinguishing physiological uptake from tumoral involvement was not possible. c) SPECT-CT fusion images revealed that the involvement was due to physiological bowel activity