|Year : 2015 | Volume
| Issue : 8 | Page : 234-238
Contribution of ultrasound-guided fine-needle aspiration cell blocks of metastatic supraclavicular lymph nodes to the diagnosis of lung cancer
Hai-Ying Tian1, Dong Xu2, Jun-Ping Liu2, Wei-Min Mao3, Li-Yu Chen2, Chen Yang2, Li-Ping Wang2, Kai-Yuan Shi2
1 The Second Clinical Medical College of Imaging and Nuclear Medicine, Zhejiang Chinese Medical University, Hangzhou, Zhejiang Province, 310053, China
2 Department of Ultrasonography, Zhejiang Cancer Hospital, Hangzhou, 310022, China
3 Department of Thoracic Oncology Surgery, Zhejiang Cancer Hospital, Hangzhou, 310022, China
|Date of Web Publication||26-Nov-2015|
Department of Ultrasonography, Zhejiang Cancer Hospital, Hangzhou, Zhejiang Province, 310022
Department of Thoracic Oncology Surgery, Zhejiang Cancer Hospital, Hangzhou, 310022
Source of Support: None, Conflict of Interest: None
Purposes: Routine smears of fine-needle aspiration (FNA) specimens of supraclavicular lymph nodes with ultrasound (US) real-time guidance have proven useful in lung cancer staging, but the clinical value of additional information from cell-block of FNA samples has been little researched. This study mainly focused on the contribution of cell block analysis to the diagnosis and staging in lung cancer.
Materials and Methods: Clinical data about 211 lung cancer patients with supraclavicular lymph node enlargement admitted to ultrasonography in the Zhejiang Cancer Hospital and recommended a needle biopsy under US-guided, the adequacy of the specimens for preparing cell blocks was acquireded, and the additional immunohistochemistry or genetic information provided from cell block analysis was examined.
Results: In 211 lung cancer patients referred for US-guided FNA (median age 61.8 ± 10.0 years, range 30-88) 279 aspirations were performed. Conventional smears could be obtained from 185 aspirates (66.3%) and contained 176 (95.1%) diagnostic smears. Cell blocks could be obtained from 94 aspirates (33.7%) and contained diagnostic material in 88 (93.6%) aspirates. Above all, cell blocks also made epithelial growth factor receptor gene mutation analysis in 17 patients with FNA samples, and the positive rate was 70.6%. Overall, cell blocks provided clinically significant information for 51 of the 211 patients participating in the study (24.2%).
Conclusion: Cell-block samples from US-guided FNA is a promising, relatively noninvasive technique to provide additional information in lung cancer diagnosis. Analysis of cell blocks allows for genetic analysis of the patients with supraclavicular lymph nodes metastasis.
Keywords: Cell block, fine-needle aspiration, lung cancer, supraclavicular lymph node ultrasound-guided
|How to cite this article:|
Tian HY, Xu D, Liu JP, Mao WM, Chen LY, Yang C, Wang LP, Shi KY. Contribution of ultrasound-guided fine-needle aspiration cell blocks of metastatic supraclavicular lymph nodes to the diagnosis of lung cancer. J Can Res Ther 2015;11, Suppl S4:234-8
|How to cite this URL:|
Tian HY, Xu D, Liu JP, Mao WM, Chen LY, Yang C, Wang LP, Shi KY. Contribution of ultrasound-guided fine-needle aspiration cell blocks of metastatic supraclavicular lymph nodes to the diagnosis of lung cancer. J Can Res Ther [serial online] 2015 [cited 2020 Feb 17];11:234-8. Available from: http://www.cancerjournal.net/text.asp?2015/11/8/234/170544
Hai-Ying Tian and Jun-Ping Liu contributed equally to this work.
| > Introduction|| |
Despite all efforts, lung cancer is remains the most common fatal malignancy worldwide and approximately 80-90% of cases involve nonsmall cell lung cancer (NSCLC).  Operative resection is the most effective method for early stage of lung cancer, but the majority of patients have advanced disease at the time of diagnosis. According to the latest tumor, node, and metastasis classification of International Association for the Study of Lung Cancer, supraclavicular lymph node metastasis is considered N3 disease, and the long-term effect of surgery treatment was not satisfactory for this stage.  The initial treatment for advanced lung cancer is systemic chemotherapy.  With the introduction of neoteric targeted therapies for NSCLC, pathologists have had to deal with a corresponding promotion in the need for accurate diagnosis and appropriate classification of subtypes. The analysis of gene mutations in cancer cells, such as epithelial growth factor receptor (EGFR) or anaplastic lymphoma kinase gene,  has become crucial for the choice of treatment. Studies have shown that (the) combination (of) chemotherapy with EGFR tyrosine kinase inhibitors (TKI) could be improved the living state for patients with EGFR mutations. ,
Ultrasound-guided fine-needle aspiration (US-guided FNA) is currently the first-line ancillary diagnostic procedure performed in patients with cervival and supraclavicular nodes and masses or surgery may not be the first treatment option, and can often provide prognostic and therapeutic information as well.  Material obtained during US-guided FNA can be processed additionally as a cell block. The usefulness of cell blocks has been acknowledged in FNA procedures, and several medical societies have recently recommended its routinely use for lung cancer diagnosis. , However, this processing technique is not yet widely used on US-guided FNA of lymph nodes and there is little research about its contribution to the additional diagnostic information. The aim of our study was to evaluate that contribution in a retrospectively recruited series of patients with supraclavicular lymph node enlargement undergoing US-guided FNA for the diagnosis or staging of lung cancer.
| > Materials and methods|| |
Over a 18-month period from January 1, 2014, a total of 211 patients (159 men) of median age 61.8 ± 10.0 years (range 30-88) were included in the study. All patients who had a suspicion of lung cancer or has been diagnosed with lung cancer, with nonpalpable supraclavicular nodes seen on the US scan were enrolled in the study. US-guided FNA was used as a diagnostic procedure in these patients and additionally used for staging in all lung cancer patients who did not show distant metastasis at the first examination. Patients with hemorrhagic diseases or coagulation disorders were excluded from staging by FNA.
Ultrasound evaluation and fine-needle aspiration technique
US evaluation of the supraclavicular region was performed using a 7-12 MHz linear probe with an Color Doppler Ultrasonic Diagnostic Apparatus (model:LOGIQ E9,GE Health Care). In addition, the cervical region was examined on both sides up to the submental region if enlarged supraclavicular nodes US were detected. According to Vassallo et al., criteria for suspicious lymph nodes were a short-axis diameter of more than 5 mm, a rounded shape (longitudinal-axis to short-axis <2) and a missing echogenic hilum, the lymph node was called pathological and FNA was performed. , A cut off size of 5 mm is reasonable as it is often technically impossible to sample smaller nodes. Node shape, blood flow pattern and the size was recorded using the software of the US unit. If enlarged supraclavicular lymph nodes were detected by US, biopsy was performed under US guidance if the result could influence further patient management. Additional informed consent was obtained for all patients prior to biopsy. Ethical approval was obtained from the Ethical Committees of Zhejiang Cancer Hospital. The patients were placed in a supine position with their necks hyperextended. Enlarged nodes were aspirated with fine 22-gauge cytology needles. The cytological specimen was aspirated with the free-hand technique by 2 US interventional physicians. Once the needle tip was inside the target, negative pressure was maintained with a syringe at the proximal end of the catheter while the needle was pushed forth and back, releasing the suction before the needle was removed from the target structure.
The aspiration of material in the needle was obtained and all specimens were placed on slides and fixed with 95% alcohol. The cytologist classified nodes as "normal tissue negative for malignancy" when the sample included 40 lymphocytes per high-power field in cellular areas of the smear and/or clusters of pigmented macrophages, and no cancerous cells,  or as "metastatic" when identifiable groups of cancer cells were present. Nodes containing only isolated anisotropic cells were regarded as "suspicious" but nondiagnostic. Nodes containing only blood cells were also classified a nondiagnostic. Stations showing only nodes with a short-axis diameter <5 mm during US-guided FNA were not sampled and marked as normal, in agreement with previous research. 
All cell blocks were essentially dried up and agglomerated the specimens on filter paper and then placed them into 10% formalin just after for subsequent processing in the laboratory.  Cell blocks were embedded in paraffin and obtained for sections of 5 μm thickness. Conventional hematoxylin-eosin staining was used on cell-block sections and, when necessary, immunohistochemical stainings were applied for the identification or phenotyping of malignant cells. For some cases of NSCLC, genomic DNA were extracted from the tumor cells and EGFR mutations in exons 18(G719), 19(deletion), 20(T790), and 21(L858 and L861) , were detected by ARMS methods. All EGFR mutation analysis was performed in the pathology laboratory.
Data were analyzed using SPSS statistical software (Version 19.0; SPSS, Chicago, IL). Results were expressed as absolute and relative frequencies for categorical variables, and as medians and standard deviations for continuous variables. First, availability of cell blocks containing adequate tissue samples from nodes or masses sampled by means of US-guided FNA was assessed. Second, the provision of new pathologic information from these cell blocks was analyzed. Finally, the impact of the additional information provided by the analysis of cell blocks over patient staging was assessed. The recovery of a cell block suitable for performance of genetic analysis of EGFR mutations in patients with metastatic NSCLC was considered as additional genetic information.
| > Results|| |
US-guided FNA of supraclavicular lymph nodes was performed on 211 patients with a final diagnosis of lung cancer; the patient's median age was 61.8 ± 10.0 years (range 30-88) and the male-to-female ratio was 3.1:1 [Table 1]. US-guided FNA diagnosed metastasis in 89 out of 117 patients with evidence of enlargement in supraclavicular lymph nodes on the color Doppler US (76.1%). Of 279 FNA procedures performed, the average short-axis dimension of the 129 malignant supraclavicular lymph nodes was 7.5 ± 2.9 cm (range 3-21 mm), 135 (48.4%) showed lymphocytes and were negative for malignancy, 6 gave isolated atypical cells (2.2%) and 9 (3.2%) gave only nonrepresentative material [Table 2].
|Table 2: Diagnoses in routine smears of supraclavicular lymph nodes with US - guided FNA (n=279, %)|
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Cell blocks could be prepared from 94 aspirates (33.7%) and adequate material for diagnosis was recovered from 88 (93.6%) of them [Figure 1]. The median short-axis diameter of nodes from which material for cell block processing was 8.2 cm ± 3.2 cm (range 4-19 mm). Malignancy was diagnosed at the examination of 72 of the obtained cell blocks, being the block sample diagnostic and negative for lung cancer in 22 of the performed aspirations. In 3 cases the cell block showed only isolated atypical cells and was considered nondiagnostic, 3 cases showed only nonrepresentative. There were 51 cases of cell blocks achieved the subtype and also provided additional pathologic and clinically significant information. Additionally, cell blocks provided material suitable for EGFR gene mutation analysis in 17 of the 64 patients with metastatic NSCLC in the sampled nodes (26.6%), and the positive rate was 70.6%. Based on the identification of such mutations, these patients received subsequent therapy with a TKI in clinic, and all patients remitted in different degrees. US evaluation following biopsy did not reveal any sign of hematoma or other possible complications. None of the patients reported any form of discomfort, pain or swelling.
|Figure 1: Diagnoses from routine smear and cell block acquired with ultrasound-guided fine-needle aspiration of metastatic supraclavicular lymph nodes (EGFR: epithelial growth factor receptor)|
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| > Discussion|| |
US has proven to be a valuable diagnostic technique in the evaluation of patients with cervival and supraclavicular masses ,, and have a higher detection rate than palpation or computed tomography.  Using a state-of-the-art US technique, a wide range of abnormality prevalence in supraclavicular lymph nodes has been reported in lung cancer patients ranging from 20% to 59%. ,,, In addition, metastases were proven in up to 45.5%. 
It was not technically possible to perform US-guided core-needle biopsy or surgical biopsy in patients with suspicious lymph node enlargement on account of the proximity of their lymph nodes to vascular structures or short-axis is too short, cytological material may be the only specimen available for diagnostic workup. A number of recent reports demonstrated that US evaluation of the supraclavicular region and US-guided aspiration are simple and safe procedures for detecting and proving a nonresectable tumor stage. ,, This technique provides conventional smears for cytology that have a good correlation with histological diagnoses. Thus, more invasive and unnecessary diagnostic procedures can be avoided, especially in patients with advanced disease.
Accurate diagnosis and staging of the patient with suspected lung cancer is the basis for individual, optimal therapy. Some experts pointed out that the presence of lymphatic micrometastases and/or isolated tumor cells is associated with distant recurrence in patients with early stage NSCLC. They recommend the routine use of serial sectioning and immunohistochemistry in lymph nodes assessment to improve the accuracy of staging.  For those with unresectable or micrometastases lymph nodes, US-guided FNA is the best choice, multi-point and multi-directional puncture can effectively found the pathological cells. Due to the complex background, poor reproducibility, and overlapping cells structure that results in poor detection of conventional smears. However, cell block can clear some blood cells and inflammatory cells, to a certain extent, provide clinically significant information and improve the positive rate of cytological diagnosis. Cell blocks can be acquired by means of US-guided FNA, and, compared with conventional smears, allow the performance of sections suitable for larger immunohistochemical staining batteries , to determine the tumor types, and, in patients with NSCLC, EGFR genetic analysis in cell block samples to provide the basis for clinical targeted therapy. We found that over a 93.6% of the recovered cell blocks contained diagnostic cellular material, a percentage similar to those in other series where cell blocks from needle core biopsies have been processed. ,
Cell-block processing allowed for the performance of EGFR mutational analysis with a diagnosis of metastatic NSCLC and in 12 of the 17 patients confirmed the presence of an EGFR mutation, which confer sensitivity to the TKIs.  Targeted therapies are under active development as a means to improve treatment efficacy in selected patient populations. The strong association of EGFR gene mutation with NSCLC has made it a candidate for TKI therapy, whereas patients without EGFR mutations respond poor to chemotherapy.  Small-molecule TKIs that target the EGFR including the reversible inhibitors gefitinib and erlotinib, were the first targeted drugs to enter clinical use for the treatment of unselected patients with NSCLC.
| > Conclusion|| |
The present date confirms that cervical and supraclavicular US and US-guided FNA are safe, easy to perform and potentially valuable tools in the diagnosis and staging. Cell-block preparation is a simple method that provides important additional information after US-guided FNA in lung cancer. Overall, cell-block processing provided clinically significant information and allowed for the performance of genetic analyses of EGFR mutations, confirming the advantages of this processing method for the diagnosis and staging of lung cancer, and may allow clinicians to optimize targeted therapy of lung cancer.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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