Endoscopic Ultrasonography-Guided Fine-Needle Aspiration for Duodenal Subepithelial Lesions Showing a Hypoechoic Mass on Endoscopic Ultrasound Imaging

Background/Aims: For duodenal subepithelial lesions showing a hypoechoic mass on endoscopic ultrasound imaging, the utility of endoscopic ultrasound-guided fine-needle aspiration and the frequency of histological types have not been the focus of previous literature. This study aimed to clarify this. Materials and Methods: This prospective observational study enrolled 22 consecutive patients who underwent endoscopic ultrasound-guided fine-needle aspiration for duodenal subepithelial lesions with hypoechoic mass on endoscopic ultrasound. Immunohistochemical analysis was performed for all endoscopic ultrasound-guided fine-needle aspiration and surgically resected specimens. The main outcome measures were the technical results of endoscopic ultrasound-guided fine-needle aspiration and the frequency of histological types of duodenal subepithelial lesions with hypoechoic mass. Results: Thirteen fine-needle aspiration specimens were obtained from the duodenal bulb and eight from the descending duodenal region. The puncture was not performed because of intervening vessels in one patient. The diagnostic rate was 81% (95% confidence interval: 58.1-94.6, 17/21 patients). In 12 patients receiving surgical resection (excluding one cancellation of endoscopic ultrasound-guided fine-needle aspiration), the diagnostic accuracy of endoscopic ultrasound-guided fine-needle aspiration was 75% (95% confidence interval: 42.8-94.5, 9/12 patients). No complications were observed. The histopathological diagnoses included 11 cases of gastrointestinal stromal tumor (50%), 2 cases of leiomyoma (9%), 2 cases of metastatic cancer (9%), 2 cases of benign inconclusive, and 1 case each of carcinoid, malignant lymphoma, leiomyosarcoma, gauzeoma, and aberrant pancreas (4.5% each). The frequency of malignant tumors in the duodenal subepithelial lesions with hypoechoic mass group was 73% (16/22 patients). Conclusions: Endoscopic ultrasound-guided fine-needle aspiration for duodenal subepithelial lesions with hypoechoic mass was safe and accurate. As duodenal subepithelial lesion with hypoechoic mass has a reasonably high possibility of containing malignant tumors, it is desirable to perform endoscopic ultrasound-guided fine-needle aspiration.


INTRODUCTION
Duodenal subepithelial lesions (DSELs) are rare, 1 although their detection rate is increasing with recent advances in endoscopy and observation technology.2][3][4] Duodenal subepithelial lesions have a broad spectrum of histologic types, and corresponding management according to these types is needed.However, endoscopic and histologic diagnoses using conventional endoscopic biopsy are difficult because of the overlying normal mucosa. 1 Endoscopic ultrasound (EUS) is the most important imaging modality for the differential diagnosis of gastrointestinal subepithelial lesions (SELs). 2,5,6The EUS can determine the origin of the gastrointestinal wall layer (i.e., within the submucosal layer, in continuity with the muscularis propria, or outside the wall), its content (i.e., liquid, fat, solid tumor, or blood vessel), and size of the gastrointestinal SEL. 7Therefore, EUS can provide a conclusive diagnosis of some lesions using echo findings only, including lipoma (high echoic mass), cystic lesion (anechoic mass), and varices.However, hypoechoic masses (HM) are also observed in malignant tumors, such as GIST, malignant lymphoma, metastatic cancer, neuroendocrine tumors, and SEL-like cancer, and benign conditions, such as leiomyoma, schwannoma, and aberrant pancreas. 2,7It is difficult to distinguish between these lesions using EUS findings alone 8,9 and tissue acquisition is needed.Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) is a reliable procedure for the conclusive immunohistochemical diagnosis of HMs in gastrointestinal SELs. 2,4,8Although histological features of HMs in gastric SELs using EUS-FNA have been reported, 10,11 no studies have focused on duodenal subepithelial lesions with hypoechoic masses (DSELHMs).Herein, we prospectively evaluated 22 patients who underwent EUS-FNA with the detection of DSELHM by EUS at Aso Iizuka Hospital.

MATERIALS AND METHODS Patients
Patients with DSELs were managed according to our institutional diagnostic and therapeutic algorithm for gastrointestinal SELs (Figure 1). 12Surgical resection was recommended for patients with histologically confirmed GIST (guided by immunohistochemical analysis of EUS-FNA specimen) according to the Japanese GIST guidelines 13 after discussion with each patient.In our algorithm, EUS-FNA was performed for all DSELHMs >10 mm in diameter.This prospective study enrolled 22 consecutive patients (male:female, 15:7; mean age, 61.6 years) diagnosed with a DSELHM >10 mm in diameter by EUS who underwent EUS-FNA for differentiation of DSELHMs at our institution from October 2004 to June 2020.

Immunohistochemical Analysis
The EUS-FNA and surgical resection specimens were fixed in 10% formaldehyde, and tissue blocks were embedded in paraffin.The sections were stained with hematoxylin and eosin.Immunoperoxidase staining was performed on the cell blocks and representative histological sections of the tumor using commercially available antibodies.The details of the antibodies used have been previously described. 10,11A tumor with a positive reaction to c-kit, CD34, or DOG1 was diagnosed as a GIST.A tumor with a negative reaction to c-kit, CD34, DOG1, and S-100, and a positive reaction to muscle actin was diagnosed as a myogenic tumor (leiomyoma).A tumor with a negative reaction to c-kit, CD34, DOG1, and muscle actin, and a positive reaction to S-100 was diagnosed as a neurogenic tumor (schwannoma).

Assesment of Clinical Outcome
The histological diagnostic rate, complications of EUS-FNA, and the frequency of the histological types of DSELHMs were evaluated in all 22 cases.The accuracy of the differential diagnosis of DSELHMs was calculated in 12 surgically resected patients with a diagnosis based on preoperative EUS-FNA (Figure 3).

Statistical Analysis
95% confidence intervals (CIs) were appropriately calculated by statistical analysis using Stata version 15.0 (Stata Corp LLC, Tex, USA).

Ethical Approval
This study was performed at our institution and approved by the Ethics Committee of Aso Iizuka Hospital (registration no.17129).This study is registered with the University Hospital Medical Information Network (UMIN) Clinical Trials Registry, number UMIN 000009972.Written informed consent was obtained from all the patients and the study performed in accordance with the Declaration of Helsinki.

RESULTS
The characteristics of the 22 patients who underwent EUS-FNA for DSELHMs are shown in Table 1.Lesions were present in the duodenal bulb in 13 patients and the descending part of the duodenum in 9 patients.The mean tumor size measured using EUS was 29.7 mm (range, 11-100 mm).
The 22G FNA needles were solely used in 16 cases, 25G needles were solely used in 3 cases, and both were used in 2 cases.The mean number of passes in EUS-FNA was 2.57 (range, 1-5), excluding 1 discontinuation.The histological diagnosis rate of EUS-FNA was 81% (95% CI: 58.1-94.6,17/21 patients), excluding 1 patient in whom  the puncture was terminated (a safe puncture route could not be obtained because of blood vessels in the puncture route).No EUS-FNA procedure-related complications were noted.One patient who could not undergo EUS-FNA (due to an intervening blood vessel in the puncture route) underwent surgical resection because of clinical suspicion of GIST and the diagnosis was confirmed as GIST in the resected tissue.Two patients who underwent EUS-FNA without a conclusive histological diagnosis were surgically resected because of suspicion of a GIST (1 patient had GIST, and the other had a gauzeoma).Two patients diagnosed with malignant tumors by EUS-FNA did not undergo resection.One patient with a carcinoid tumor was followed up at his request.The remaining patients had metastatic cancer and had received chemotherapy.Preoperative histological diagnosis was correctly obtained by EUS-FNA using immunohistochemical analysis in 9 of the 12 surgically resected cases (diagnostic accuracy: 75%, 95%; CI: 42.8-94.5),excluding the case with puncture cancellation (Table 1).The mean time interval between EUS-FNA and surgery was 8.3 weeks (range, 3-15 weeks).
The histological types of DSELHMs assessed using EUS-FNA or surgically resected specimens are shown in Table 2.

DISCUSSION
Duodenal subepithelial lesions are rare; hence, the frequency of malignant disease in DSEL remains unknown.
Currently, there are no guidelines or policies for the treatment of DSEL.Large DSELs and those associated with symptoms (e.g., bleeding and passage obstruction) are candidates for resection. 3,14By contrast, asymptomatic submucosal tumors ≤20 mm in diameter are treated on a case-by-case basis. 5,14In most cases, surgical resection is the treatment of choice when malignancy is suspected, but there are reports of less invasive endoscopic resection (ER) when the tumor size is small. 15,16Pre-treatment tissue diagnosis is important for selecting an appropriate treatment for DSEL.However, since DSELs are usually covered with normal epithelium, making a histological diagnosis using conventional endoscopic biopsy is difficult.These are often evaluated using EUS-FNA or other biopsy methods. 5The EUS-FNA is the standard tissue sampling method for SELs.This study reports our experience with EUS-FNA for DSELHM in 22 patients.
The diagnostic yields of EUS-FNA using various needle types to evaluate gastrointestinal SELs range from 52% to 87%, 11,[17][18][19][20][21] whereas the diagnostic accuracy of EUS-FNA in surgically resected patients ranges from 91% to 100%. 10,11,21,22De Moura et al 23 also reported diagnostic accuracy of 73% for EUS-FNA in surgically resected patients with DSEL (n = 18).Comparatively, the diagnostic yield and accuracy of EUS-FNA in patients with DSELHM in this study were 81% and 75%, respectively.This study used conventional type 22-or 25-gauge needles to obtain histological samples.Recently available new needles, such as Franseen or Fork-tip type needles (fine-needle biopsy needles), could further improve the diagnostic rate (85%-89%) of all GI tract SELs. 24,25The incidence of EUS-FNA-related adverse events using 22-25-gauge needles for SELs was reported to be close to 0%, 10,26,27 and no adverse events were observed in this study.Thus, EUS-FNA is an accurate and safe histological test for the definitive diagnosis of DSELHMs.
The frequency of histological types of duodenal submucosal tumors has not been sufficiently investigated.
Li et al 16  of malignant disease was 13.9% (neuroendocrine neoplasm, 12.9%; GIST, 1%).El Chafic et al 28 reported the frequency of histologic types using surgical pathology or EUS-FNA of 5 endosonographically suspected duodenal GI stromal tumors (HM on EUS).All of them were malignant tumors, including 3 neuroendocrine tumors (60%) and 1 (20%) each of GIST and metastatic tumors (20%).Differences in histological types in the earlier 2 studies were derived from the inclusion criteria (endoscopically resectable lesions vs. GIST-suspected lesions by EUS).Miettinen et al 29 reported the frequency of immunohistologic types of 190 duodenal mesenchymal tumors coded as leiomyomas, leiomyosarcomas, smooth muscle tumors, schwannomas, neurofibromas, nerve sheaths, or stromal tumors retrieved from the files of the Armed Forces Institute of Pathology and Haartman Institute of the University of Helsinki from 1970 to 1996.It showed malignant lesion as 94.2% (GIST 82.1%; leiomyosarcoma 2.6%; miscellaneous malignant tumors 9.5%) and benign lesions as 4.2% (leiomyoma 3.2%; schwannoma 1.1%).In this study, 73% of patients with DSELHM had malignant tumors.There were no cases of Brunner's gland hyperplasia or lipomas in this study because Brunner's gland hyperplasia may have been diagnosed by typical endoscopic and EUS findings (multiple cystic mass), 30 and lipomas were excluded by typical EUS findings (hyperechoic mass).A previous study reported the histological typing of 90 gastric SELs <20 mm in diameter and HMs on EUS, with 47 cases (52%) of malignant SEL (44 GISTs, 1 glomus tumor, 1 SEL-like cancer, and 1 malignant lymphoma), 19 cases (21%) of benign SEL (14 leiomyomas, 4 ectopic pancreases, and 1 neurinoma), and 24 cases (27%) of indeterminate SEL. 10 In the present study, although there were only a small number of DSELHM cases, there was a high proportion of malignant tumors in the DSELHMs (73%; Funding: This study was conducted with the assistance of a research grant from Aso Iizuka Hospital.

Figure 2 .
Figure 2. The EUS-FNA of a small duodenal GIST (surgically resected case) in a 48-year-old woman.(A) EGD showing a small SEL in the bulbus of the duodenum.(B) EUS showing an 11 mm diameter subepithelial hypoechoic solid tumor with continuity to the proper muscle layer.(C) Puncture of the small GIST under EUS guidance.(D) Immunohistochemical findings of the EUS-FNA specimen showing diffusely stained c-kit positive spindle and epithelioid tumor cells.EGD, esoph agoga strod uoden oscop y; EUS, endoscopic ultrasound; EUS-FNA, endoscopic ultrasound-guided fine-needle aspiration; GIST, gastrointestinal stromal tumor; SEL, subepithelial lesion.

Table 1 .
Characteristics of 22 Patients who Underwent EUS-FNA for DSELHM performed ER in combination with ligation in 101 patients and reported the frequency of histological diagnosis using resected specimens.The frequency of benign disease was 86.1% (Brunner's gland hyperplasia, 50.5%; lipoma, 18.8%; ectopic pancreas, 16.8%) and that