Diagnosis Of Lymph Node Micrometastasis Research Articles

Specific diagnosis of lymph node micrometastasis in breast cancer by targeting activatable near-infrared fluorescence imaging

Axillary lymph node metastasis has always been defined as the most important prognostic factor in the treatment of early breast cancer. Ultrasound and MRI can detect only 10% of lymph node micrometastases in early breast cancer. Therefore, it is crucial to detect early breast cancer with lymph node metastasis, however, there is no current examination method for accurate diagnosis. When breast cancer presents a malignant tendency, colony stimulating factor-1 and chemokine CCL-2 absorb mononuclear cells from the surrounding environment and differentiate into M2 Tumor associated macrophages (TAM), which increase the invasion of tumor cells and further promote the development of tumors. Mannose, as a simple natural ligand, can selectively bind to TAM surface CD206 (macrophage mannose receptor, MMR). In this study, mannose was connected with near infrared dye (NIR) IR780 via disulfide bond to obtain Mannose-IR780 conjugate (MR780), which was further self-assembled into near infrared nanoprobe (MR780 NPs) with quenched fluorescence. When selectively targeting CD206 highly expressed on the surface of TAM, disulfide bond was cleaved by the glutathione enriched in the microenvironment, resulting in fluorescence recovery, thus achieving NIR fluorescence molecular imaging of TAM and diagnosis of tumor lymph node metastasis in mouse models. Our findings suggest that targeted imaging of TAM enable noninvasive and sensitive detection of metastatic lymph nodes in vivo, which is instructive for tumor therapy.

Current status of lymph node micrometastasis in gastric cancer.

Lymph node metastasis is one of the most important prognostic factors in patients with gastric cancer. An inadequate number of dissected lymph nodes is an independent risk factor affecting recurrence, even in patients who are node negative. Oddly, certain early-stage patients still experience recurrence or metastasis within a short time, even if they have undergone standard radical mastectomy. Many researchers have attributed these adverse events to lymph node micrometastasis (LNM), which is defined as a microscopic deposit of malignant cells of less than 2 mm in diameter. With the development of diagnostic tools such as immunohistochemistry and reverse transcription-polymerase chain reaction, the rate of detection of LNM has been constantly increasing. Although there is no clear consensus about risk factors for or the definitive clinical significance of LNM, the clinical impact of LNM is remarkable in gastric cancer. For minimally invasive treatment in particular, such as endoscopic submucosal dissection and laparoscopic surgery, accurate diagnosis of LNM is regarded as the potential key to maintaining the balance between curability and safety. This review provides an overview of the definition, detection and significance of LNM in gastric cancer. We also summarize several attention-drawing controversies regarding the treatment of patients who may have LNM.

Diagnosis of lymph node micrometastasis at the pN₀ stage of lung adenocarcinoma using a combination of markers.

This study aimed to detect micrometastatic tumor cells in the lymph nodes of patients with pN0 lung adenocarcinoma using a combination of thyroid transcription factor-1 (TTF-1) expression and cytokeratin 7 (CK7) expression and to investigate the association of lymph node micrometastasis with the clinicopathological characteristics of patients with lung adenocarcinoma. A total of 54 patients with pN0 lung adenocarcinoma and whose primary tumors were positive for both TTF-1 and CK7 expression were included in this study. In total, 893 lymph nodes were obtained from these 54 patients and were analyzed for micrometastasis by immunohistochemical staining with anti-CK7 and anti-TTF-1 antibodies. CK7- and TTF-1-positive cells were found in the lymph nodes of 9 (16.7%) of 54 patients, and 21 (2.4%) of 893 lymph nodes exhibited positivity for these factors. No cells positive for both CK7 and TTF-1 were detected in the 5 lymph nodes obtained from patients with benign lung tumors. Lymph node micrometastasis was found to be associated with the differentiation grade and primary tumor position (P < 0.05). The detection of lymph node micrometastasis by a combination of CK7 and TTF-1 immunohistochemical staining provides a more accurate assessment of tumor staging for pN0 lung adenocarcinoma.

Advantages of the rapid double-staining method for intraoperative detection of micrometastasis in sentinel lymph nodes

For rapid intraoperative diagnosis of lymph node micrometastasis, we refined the rapid immunohistochemistry method by combining anti-cytokeratin antibody-labeled nanocrystal beads with rapid hematoxylin and eosin (H&E) staining on the same section, referred to as the rapid double staining (RDS) technique. Two frozen-section slices each were obtained from 372 lymph nodes of 100 breast cancer patients. We performed RDS for 1 slide and rapid H&E staining for the other. The results were compared with the corresponding final pathological data obtained from the permanent specimens. For specimens from patients with pN1(mi) as determined by final pathological examination, the false‑negative rate was 33.3% for rapid H&E staining and 16.7% for RDS. For specimens from patients with pN0(i+) as determined by final pathological examination, the false‑negative rate was 80% for rapid H&E staining and 0% for RDS. These results indicate that RDS is superior to conventional rapid H&E staining for intraoperative diagnosis of nodal micrometastasis and isolated tumor cells.

Lymph node micrometastasis in gastrointestinal tract cancer—a clinical aspect

Lymph node micrometastasis (LNM) can now be detected thanks to the development of various biological methods such as immunohistochemistry (IHC) and reverse transcription-polymerase chain reaction (RT-PCR). Although several reports have examined LNM in various carcinomas, including gastrointestinal (GI) cancer, the clinical significance of LNM remains controversial. Clinically, the presence of LNM is particularly important in patients without nodal metastasis on routine histological examination (pN0), because patients with pN0 but with LNM already in fact have metastatic potential. However, at present, several technical obstacles are impeding the detection of LNM using methods such as IHC or RT-PCR. Accurate evaluation should be carried out using the same antibody or primer and the same technique in a large number of patients. The clinical importance of the difference between LNM and isolated tumor cells (≤0.2 mm in diameter) will also be gradually clarified. It is important that the results of basic studies on LNM are prospectively introduced into the clinical field. Rapid diagnosis of LNM using IHC and RT-PCR during surgery would be clinically useful. Currently, minimally invasive treatments such as endoscopic submucosal dissection and laparoscopic surgery with individualized lymphadenectomy are increasingly being performed. Accurate diagnosis of LNM would clarify issues of curability and safety when performing such treatments. In the near future, individualized lymphadenectomy will develop based on the establishment of rapid, accurate diagnosis of LNM.

胃癌センチネルリンパ節生検における術中リンパ節微小転移診断

胃癌センチネルリンパ節生検における術中リンパ節微小転移診断

The value of step-sectioning in the diagnosis of lymph node micrometastasis in breast cancer

Objectives: Routine practice is to examine one histological section for evaluating secondary tumor deposits in the axillary lymph nodes in patients with breast cancer. The aim of the present study is to evaluate whether multiple levels of histological section detect significantly more metastatic deposits in axillary lymph nodes from breast cancer resection specimen than the standard practice of examining one section. Methods: A retrospective study of 30 patients with node negative breast cancer was performed whose specimens were received at AL-Jamhuri Teaching Hospital and private laboratories in Mosul city between the years 2008 and 2010. The original slides and the paraffin wax blocks containing the lymph nodes were retrieved from the archive, and 4 extra levels (separated by 30 µm) were cut from each block and were stained with H & E stain. Results: Of the 30 cases with node negative breast carcinoma, 4 (13.3%) were found to contain extra tumor deposits at deeper levels that were not detected at the original sections. Conclusion: Multiple levels of histological sections separated by relatively small intervals detect more tumor deposits in the axillary lymph nodes than the current practice of examining a single section.

The efficiency of micrometastasis by sentinel node navigation surgery using indocyanine green and infrared ray laparoscopy system for gastric cancer

The clinical application of sentinel node navigation surgery (SNNS) for patients with gastric cancer requires accurate intraoperative diagnosis of lymph node metastasis. However, the clinical significance of the diagnosis of lymph node micrometastasis for gastric cancer has not been established. In this study, we evaluated lymph nodes dissected during SNNS by immunohistochemistry with anti-cytokeratin antibody (IHC) staining for gastric cancer to investigate the usefulness of SNNS. The subjects were 130 patients with gastric cancer (3,381 lymph nodes) who underwent SNNS with infrared ray observation and lymph node dissection of D1+α or more. The dissected lymph nodes were stained with IHC (CAM 5.2), and the results were compared with intra- and postoperative diagnoses by hematoxylin and eosin (H&E) staining. In addition, the association of metastatic lymph nodes and ICG-positive lymph nodes was examined. The number of patients (lymph nodes) with lymph node metastasis by HE and IHC staining was 16 (52 nodes) and 31 (91 nodes), respectively. Fifteen patients (27 nodes) diagnosed with pN0 by HE staining were diagnosed to be metastatic by IHC staining. The tumor depth of these patients was pT1 in ten patients (m, 3; sm, 7) and pT2 in five (mp, 4; ss, 1). Regarding the histological type, three patients were classified as well-differentiated type, while six patients each had moderately and poorly differentiated types. The grade of lymphatic invasion was ly0 in 5, ly1 in 6, and ly2 in 4, respectively. Histological assessment of 27 IHC-positive and HE-negative cells indicated 5 single cells, 16 clusters, and 6 micrometastases. These lymph nodes were all included in the sentinel nodes (SN) identified during surgery. All but one patient (0.8%) were recurrence-free at 2-8 years after surgery (median 74.7 months). The one patient developed anastomotic recurrence 4.5 years after the first operation and died. Since all 27 lymph nodes in 15 patients with metastasis by IHC staining but not by HE staining were micrometastasis or less and included in the SN, ICG-positive lymphatic basin dissection by SNNS with infrared ray observation seems to be an adequate method of lymph node dissection for gastric cancer.

Clinical study on lymph node metastatic recurrence in patients with N0 esophageal squamous cell cancer

Esophageal squamous cell cancer (ESCC) is one of the most common malignancies treated by thoracic surgeons. It is aggressive and generally associated with a poor prognosis. One of the most important prognostic factors is the presence of the lymph node metastasis (LNM). The purpose of the present study was to investigate the risk factor with lymph node metastatic recurrence in patients with N0 esophageal squamous cancer after Ivor Lewis esophagectomy based on the detection of mucin1 (MUC1) mRNA. The subjects were 82 patients with pN0 ESCC who underwent Ivor Lewis esophagectomy with two-field lymph node dissection from January 2001 to January 2005. All lymph nodes (501 stations) obtained from these patients were reevaluated by reverse transcriptase-polymerase chain reaction (RT-PCR) to detect MUC1mRNA. The diagnosis of lymph node micrometastasis (LNMM) was based on the detection of MUC1 mRNA. The Kaplan-Meier method was used to calculate the survival rate and lymph nodal metastatic rate. Log-rank test was performed to compare the recurrence rate, and Cox regression multivariate analysis was performed to determine independent prognostic factors. The overall 3-year survival rates of 82 patients were 78.0%, and the first recurrence exhibiting lymph nodal metastasis was recognized in 37 patients (45.1%) in the first 3 years after operation. Lymph node metastatic rate in patients in the first 3 years after operation was significantly associated with the T status (P < 0.05). MUC1 mRNA was identified in at least one lymph node station from 23 (28.1%) patients. Also, lymph node metastatic rate of the patients with LNMM was significantly higher than that of the patients without LNMM (P < 0.01).The results of multivariate analysis confirmed that LNMM and T status in patients with N0 ESCC were independent risk factors for 3-year lymph node metastatic recurrence after Ivor Lewis esophagectomy. Adjunctive therapy might be beneficial in controlling the locoregional recurrence and elevated healing rates for certain patients.

Mucin 1 and Vascular Endothelial Growth Factor C Expression Correlates with Lymph Node Metastatic Recurrence in Patients with N0 Esophageal Cancer After Ivor‐Lewis Esophagectomy

The purpose of the present study was to investigate the risk factors associated with lymph node metastatic recurrence in patients with N0 esophageal cancer after Ivor-Lewis esophagectomy based on the detection of Mucin 1 mRNA and vascular endothelial growth factor (VEGF) C mRNA. The subjects were 82 patients with pN0 esophageal cancer who underwent Ivor-Lewis esophagectomy with two-field lymph node dissection from January 2001 to January 2005. A total of 501 lymph nodes obtained from these patients were re-evaluated by reverse transcriptase-polymerase chain reaction (RT-PCR) to detect mucin l (MUC1) mRNA; VEGF-C mRNA was also detected in esophageal cancer issues by RT-PCR. The diagnosis of lymph node micrometastasis (LNMM) was based on the detection of MUC1 mRNA. The Kaplan-Meier method was used to calculate the survival rate and lymph nodal metastatic rate, the log-rank test was performed to compare the recurrence rate, and Cox regression multivariate analysis was performed to determine independent prognostic factors. MUC1 mRNA was detected in 29 lymph nodes from 23 patients, which accounted for 5.79% of all the 501 lymph nodes and 28.05% of all 82 patients, respectively. Vascular endothelial growth factor C mRNA was identified in esophageal cancer issues from 42 (51.22%) patients. The overall 3- and 5-year survival rates of 82 patients were 78.0 and 51.2%, respectively. First recurrence exhibiting lymph nodal metastasis was recognized in 37 patients (45.1%) in the first 3 years after operation. The lymph node metastatic rate in patients in the first 3 years after operation was significantly associated with T status (p < 0.05) and the lymph node metastatic rate of the patients with LNMM was significantly higher than that of the patients without LNMM (p < 0.01). The lymph node metastatic rate of the patients with VEGF-C mRNA expression in esophageal cancer tissues was significantly higher than that of the patients without VEGF-C mRNA expression (p < 0.01).The results of multivariate analysis confirmed that VEGF-C mRNA expression in esophageal cancer tissues, LNMM, and T status in patients with N0 esophageal cancer were independent relevant factors for 3-year lymph node metastatic recurrence after Ivor-Lewis esophagectomy. Vascular endothelial growth factor C mRNA expression in esophageal cancer tissues, LNMM, and T status in patients with N0 esophageal cancer were independent risk factors for 3-year lymph node metastatic recurrence after Ivor-Lewis esophagectomy. Adjunctive therapy might be beneficial in controlling the locoregional recurrence and elevated healing rates for certain patients.

Gene Diagnosis and Prognostic Significance of Lymph Node Micrometastasis after Complete Resection of Histologically Node‐negative Non‐small Cell Lung Cancer

The purpose of the present study was to investigate the prevalence of lymph node micrometastasis (LNMM) based on the detection of MUC1 mRNA, and assess the impact of these micrometastases on prognosis after resection of pathologic N0 (pN0) non-small cell lung cancer (NSCLC). The subjects were 89 patients who underwent complete resection of pN0 NSCLC at our department between January 2000 and January 2002. All lymph nodes (402 stations) obtained from these patients were re-evaluated by reverse transcriptase-polymerase chain reaction (RT-PCR) to detect MUC1 mRNA. The diagnosis of LNMM was based on the detection of MUC1 mRNA. The Kaplan-Meier method was used to calculate the survival rate, and Cox regression multivariate analysis was performed to determine independent prognostic factors. Micrometastases were detected in 36 lymph node stations (9.0%) from 21 patients (23.6%). The TNM staging of these 21 patients was upregulated from stage IA-IIB to stage IIIA. The 5-year survival rate of patients with LNMM was significantly lower than that of patients without LNMM (23.8% versus 44.1%; p < 0.05). The results of multivariate analysis confirmed that T status, histology, and LNMM were independent prognostic factors. The prevalence of LNMM in patients with pN0 NSCLC was 23.6% (21/89). T status, histology, and LNMM were independent prognostic factors.

Lymph Node Micrometastasis: A Predictor of Early Tumor Relapse After Complete Resection of Histologically Node-Negative Esophageal Cancer

To investigate the prevalence of lymph node micrometastasis (LNMM) on the basis of the detection of MUC1 mRNA, and assess the impact of these micrometastases on disease-free interval after resection of pathologic N0 (pN0) esophageal squamous cell cancer (ESCC). The subjects were 93 patients who underwent complete resection of pN0 ESCC at our department between January 1999 and January 2001. All lymph nodes (426 stations) obtained from these patients were reevaluated by reverse transcription-polymerase chain reaction to detect MUC1 mRNA. The diagnosis of LNMM was based on the detection of MUC1 mRNA. A log-rank test was performed to compare the disease-free interval, and Cox regression multivariate analysis was performed to determine the independent prognostic factors. Micrometastasis was detected in 40 lymph node stations (9.4%) from 32 patients (34.4%). Disease-free interval was significantly associated with LNMM (P = 0.0138). The 5-year survival rate of patients with LNMM was significantly lower than that of those without LNMM (P = 0.004). The results of multivariate analysis confirmed that T status and LNMM were independent prognostic factors. The prevalence of LNMM in patients with pN0 ESCC was 34.4% (32/93). Thus, LNMM was significantly associated with the disease-free interval. T status and LNMM were both independent prognostic factors.

Rapid immunohistochemical detection of tumor cells in gastric carcinoma

The detection of single tumor cells or tumor cell clusters represents an important issue in intraoperative frozen section analysis. For example, surgical margins may be evaluated in order to minimize the number of additional operations. Furthermore, intraoperative diagnosis of lymph node micrometastasis (LNM) may help to define the area of appropriate lymph node dissection. In addition to haematoxylin and eosin (H&E)-stained sections, immunohistochemical detection of single tumor cells or cell clusters may be helpful in this context. The aim of this study was to evaluate the clinical significance, reliability and sensitivity of intraoperative rapid immunostaining of frozen sections. Therefore, we compared the results of rapid immunohistochemical staining of frozen sections and paraffin sections applying the EnVision and Histofine(R) detection systems. In a prospective immunohistochemical study, paraffin and frozen sections of 20 gastric cancer specimens were analyzed. Paraffin as well as frozen sections were stained immunohistochemically applying the EnVision and Histofine detection systems. As primary antibodies, AE1/AE3 (anti-cytokeratin), EMA (anti-MUC1) and B lymphocyte marker anti-CD20 were applied. The rapid immunostaining procedure was able to be completed within 10-13 min. Rapid immunohistochemical staining of frozen and paraffin sections of the same tumors resulted in comparable immunoreactivity. The rapid EnVision and Histofine procedures allowed immunostaining of frozen sections in less than 13 min. These methods can represent useful additional tools in routine surgical pathology and research, enabling a more accurate frozen section diagnosis compared to staining with H&E alone. Intraoperative rapid immunostaining can be a simple and useful technique to detect LNM.

Detection of mRNA expression of CK-19 and MUC1 gene for diagnosis of lymph node micrometastasis in NSCLC patients by reverse transcriptase-polymerase chain reaction

To investigate gene diagnosis of micrometastasis in lymph nodes in patients with non-small cell lung cancer (NSCLC) and the feasibility of mucin 1 (MUC1) mRNA and cytokeratin 19 (CK19) mRNA as molecular marker to detect micrometastasis of lung cancer. Expression of MUC1 mRNA and CK19 mRNA was detected in 119 lymph nodes taken from 31 patients with NSCLC, 35 lymph nodes from 10 patients with pulmonary benign diseases as controls by nested reverse transcriptase-polymerase chain reaction (RT-PCR). In the 119 lymph nodes from lung cancer patients, CK19 mRNA expression was detected in 66 lymph nodes (55.5%) and MUC1 mRNA expression was detected in 65 lymph nodes (54.5%) by RT-PCR. Neither CK19 mRNA nor MUC1 mRNA expression was observed in all the 35 lymph nodes in the benign pulmonary lesion group. The results suggest that the detection of both MUC1 and CK19 mRNA might be helpful to diagnose NSCLC micrometastasis in lymph nodes. The establishment of this method may lead to an earlier diagnosis of metastasis for lung cancer.

Rapid immunohistochemical detection of lymph node micrometastasis during operation for upper gastrointestinal carcinoma.

The intraoperative diagnosis of lymph node micrometastasis (LNM) may help guide the area of appropriate lymph node dissection. This study aimed to evaluate the rapid immunohistochemical detection of LNMs using frozen sections during operation for gastro-oesophageal cancer. Rapid immunostaining with anticytokeratin (AE1/AE3) antibody was compared with conventional immunostaining. A total of 210 lymph nodes obtained from 47 patients with oesophageal squamous cell carcinoma and from 32 with gastric adenocarcinoma were examined during operation. Lymph nodes were frozen, sectioned, and examined by histological and immunohistochemical methods. It took 30 min to complete the rapid immunostaining procedure; the expression of cytokeratin by rapid immunostaining was similar to that by conventional immunostaining. The incidence of lymph node metastasis detected by histological and immunohistochemical examination was 17 and 23 per cent respectively. LNM was solely detected in 12 lymph nodes by immunostaining: three micrometastases and nine with tumour cell microinvolvement. : Intraoperative rapid immunostaining is a simple and useful technique for detecting LNMs. Further study should investigate the role of rapid immunostaining during cancer surgery to select appropriate areas for lymphadenectomy.

Real-time rapid reverse transcriptase—polymerase chain reaction for intraoperative diagnosis of lymph node micrometastasis: Clinical application for cervical lymph node dissection in esophageal cancers

Background. New molecular techniques have been designed to detect cancer micrometastases that are otherwise missed by conventional histologic examination. The aim of this study was to establish a sensitive and rapid genetic assay to detect lymph node micrometastasis and to assess its usefulness clinically for cervical lymphadenectomy in esophageal cancer. We have recently shown that metastasis in the lymph node chain along the recurrent laryngeal nerves (rec LNs) is a predictor of cervical node metastasis in esophageal cancer. In our retrospective study, the positive rate of cervical lymph node metastasis with rec LNs metastasis was 51.6%, and the rate without rec LNs metastasis was 11.6%. There was a significant difference in both positive rates (P =.0002). Methods. Rec LNs obtained from 50 patients with esophageal cancer were assessed prospectively by intraoperative histopathologic examination (HE) and genetic analysis. The latter involved a real-time quantitative reverse transcriptase—polymerase chain reaction (RT-PCR) system with multiple markers, carcinoembryonic antigen, squamous cell carcinoma, and melanoma antigen-3, whose messenger RNAs are highly and frequently expressed in esophageal cancers. Cervical lymphadenectomy was subsequently performed in a subset of these patients. Results. Ten of 50 patients (20%) were scored as node positive by HE, and 24 patients (48%) were scored positive by genetic diagnosis, including 9 HE-positive cases. Genetic diagnosis of rec LNs accurately predicted all 9 cases with cervical lymph node metastasis and 2 cases with cervical lymph node recurrence, whereas HE missed 2 cases with cervical lymph node metastasis and 2 cases with cervical lymph node recurrence. Conclusions. Our real-time rapid RT-PCR assay can improve the sensitivity of HE for detection of lymph node metastasis and might be potentially useful for intraoperative genetic diagnosis for subsequent cervical lymphadenectomy in esophageal cancer surgery. (Surgery 2002;132:34-40.)