Area Abbonati
LoginPassword
doi   10.1700/1283.14184
Tumori 2013;99(2):149-153




Sentinel lymph node biopsy in breast cancer:
review on various methodological approaches
Baha Zengel1, Ulkem Yararbas2, Ahmet Sirinocak1, Guliz Ozkok3,
Ali Galip Denecli
1, Hakan Postaci3, and Adam Uslu1
1Turkish Ministry of Health Izmir Bozyaka Research and Training Hospital, Department of General Surgery; 2Ege University, Medical Faculty, Department of Nuclear Medicine, Bornova; 3Turkish Ministry of Health Izmir Bozyaka Research and Training Hospital, Department of Pathology, Izmir, Turkey

Key words: breast cancer, isosulfan blue, radiocolloid, sentinel lymph node biopsy.

Correspondence to: Baha Zengel, MD, PhD, Turkish Ministry of Health Izmir Bozyaka Research and Training Hospital, Department of General Surgery, Saim Cikrikci Cad. No: 59, Bozyaka, Izmir, 35110 Turkey.
Tel +90-532-6148109;
fax +90-232-2614444;
email bahazengel@gmail.com;
bahazengel@yahoo.com

Received May 5, 2012;
accepted December 4, 2012

abstract

Aims and background. Sentinel lymph node biopsy has been accepted as a standard procedure for early stage breast cancer. In this retrospective analysis, the results obtained with different methodological approaches using radiocolloid with or without blue dye were examined. 
Methods. A total of 158 sentinel lymph node biopsies were performed in 152 patients. Group A (85 patients) underwent lymphatic mapping using a combination of periareolar intradermal radiocolloid and subareolar blue dye injections. Group B (73 patients) underwent only periareolar intradermal radiocolloid injection. One large tin colloid and two small radiocolloids (nanocolloid of serum albumin -NC- and colloidal rhenium sulphide -CS-) were used.
Results. Successful lymphatic mapping was attained in 157 of 158 procedures (99.4%). Radiocolloids localized sentinel lymph nodes in 99.4% and blue dye in 75.3% of the cases. The number of sentinel lymph nodes removed was greater in nanocolloid and colloidal rhenium sulphide groups (P ≤0.05). Among 60 metastatic sentinel lymph nodes, frozen section analysis using hematoxylin and eosin staining failed to detect 1 macro- and 10 micrometastasis. Radiocolloid uptake was higher in sentinel lymph nodes accumulating blue dye (1643 ± 3216 counts/10 sec vs 526 ± 1284 counts/10 sec, P <0.001). Higher count rates were obtained by using larger sized colloids (median and interquartile range: tin colloid, 2050 and 4548; nanocolloid, 835 and 1799; colloidal rhenium sulphide, 996 and 2079; P = 0.01). Only 2 extra-axillary sentinel lymph nodes were visualized using periareolar intradermal injection modality.
Conclusions. Radiocolloids were more successful than blue dye in sentinel lymph node detection. More sentinel lymph nodes were harvested with small colloids, but different sized radiocolloids were similarly successful. Sentinel lymph nodes having higher radiocolloid uptake tended to accumulate blue dye more frequently. Sentinel lymph nodes manifested higher count rates when a larger colloid was used. Frozen section was very successful in detecting macrometastatic disease in sentinel lymph nodes, but the technique failed in most of the micrometastates.
Introduction
Following the concept of breast conservation, sentinel lymph node biopsy (SLNB) has emerged as another major step in breast cancer surgery. Although it is accepted as a standard method in the evaluation of axillary status in T1-2N0M0 breast cancers, there is still no consent on a unique methodological approach1-5. The technique can be performed using radiocolloid, blue dye, or a combination of the two. Whereas some authors report higher SLN detection rates with radiocolloids, many others claim similar detection and accuracy rates with both blue dyes and radiocolloids6,7. Many radiocolloids have been commercially available for the detection of sentinel lymph nodes. There is general agreement that periaerolar intradermal injection route eventuates in a high detection rate, and a particle size ranging from 100 to 200 nm is ideal to achieve optimal retention in the sentinel lymph node (SNL)8.
In this retrospective analysis, with review of our experience in SLNB, we evaluated the success rates of blue dye and radiocolloid, factors that might influence blue dye accumulation, influence of different sized radiocolloids on harvested SLNs and efficacy of intra-operative frozen section analysis in the detection of metastatic foci in SLNs.
Patients and methods
The study group consisted of 152 breast cancer patients (150 females and 2 males) who underwent SLNB between June 2008 and December 2011. Six patients had bilateral lesions, thus a total of 158 SLNBs were performed. All lymphatic mapping procedures were performed by the first author. Thirty-four patients had prior excisional biopsy. The age range was 24-88 years (mean, 57 ± 5).
Lymphatic mapping was performed by the combined method (blue dye and radiocolloid) in 85 (group A) and by using only radiocolloid in the remaining 73 procedures (group B). Due to commercial availability, three different radiocolloids with respect colloid diameter and chemical composition had to be used. Thus, Tc99m tin colloid (TC) (Amerscan Hepatate II; Amersham International, Amersham, UK) was used in 16, Tc99m nanocolloid of serum albumin (NC) (Nanocoll; Nycomed Amersham Sorin srl, Saluggia, Italy) in 41, and Tc99m colloidal rhenium sulphide (CS) (Nanocis; CIS Bio International, Gif-sur-Yvette, France) in 101 procedures. Mean colloid diameter was 72-88 nm in TC, 8 nm in NC, and 22-29 nm in CS9. The radiocolloid was injected intradermally at four quadrants of the periareolar region on the day (18-24 hr) before surgery in all procedures. For each injection, 0.25 mCi of radiocolloid was prepared in 0.1 ml volume. Before the operation, lymphatic images were collected at anterior and lateral projections within an hour following injection. Isosulfan blue 1% (Lymphazurin®) was injected into the subareolar space in 5 ml volume following induction of anesthesia. A 5-minute efficient massage was performed in order to stimulate lymphatic drainage. In group A, all hot and/or blue nodes and in group B all hot nodes were accepted as SLNs and were harvested. Hot nodes were defined as nodes bearing radioactivity fourfold that of the background activity and were localized using a hand-held gamma probe (Navigator® GPS, Tyco Healthcare). ‘Successful lymphatic mapping’ was defined as localization of one or multiple SLNs by radiocolloid and/or blue dye. SLNs were evaluated with frozen section analysis intraoperatively. The remaining tissue of the SLNs was formalin-fixed, paraffin-embedded, and sectioned. Hematoxylin and eosin (H&E) staining was performed for histological evaluation.
Immunohistochemistry using cytokeratin antibody was used for lymph nodes which were negative with H&E staining. The detection of metastatic cells in one of these steps was defined as a ‘positive SLN’. Metastatic lymph nodes were classified according to the size of metastatic deposit as macrometastasis (>2 mm) or micrometastasis (0.2-2 mm). Cell clusters or isolated tumor cells <0.2 mm in diameter were defined as submicrometastasis. Patients having macro- or micrometastasis in SLNs underwent axillary dissection. No additional axillary clearance was performed in patients with submicroscopic deposits, which were considered to be nonmetastatic.
Statistical analysis was performed using SPSS 16.0 for Windows. Kruskal Wallis and Mann-Whitney U tests were used to compare the detection rate of SLNs by three different radiocolloids, to test the relationship between blue-dye avidity and the count rates obtained by gamma-probe imaging, and to explore the possible effect of tumor size on axillary nodal involvement. The relation between blue dye accumulation and existence of prior excisional biopsy was investigated using univariate analysis (chi-squared test). P ≤0.05 was accepted as significant.
Results
Successful lymphatic mapping was achieved in 157 of 158 procedures (99.4%). Radiocolloid uptake was observed in all 157 cases. The only patient with unsuccessful lymphatic mapping was prepared with a combined method and had a prior excisional biopsy incision at an upper outer quadrant. Among 85 cases in whom the combined method was used (group A), blue dye localized SLNs in 64 (75.3%) and radiocolloid in 84 (98.8%) patients. Among 73 procedures in whom only radiocolloid was used (group B), SLN localization was attained in all studies (100%) (Table 1).
Among all successful 157 SLNBs, the median number of SLNs harvested was 3 (interquartile range (IR), 2; range, 1-9). The median number of SLNs using different radiocolloids was as follows; 2.0 ± 2.0 in TC, 3.0 ± 2.0 in NC and 3.0 ± 2.0 in CS groups (Table 2). The differences between TC/NC and TC/CS groups were found to be statistically significant (P = 0.033). The median number of SLNs showing blue dye avidity was 2 (IR, 1, range, 1-7) (Table 2). Extra-axillary drainage was observed in 2 patients. In addition to axillary SLNs, a parasternal (internal mammary) and an intramammarian node were visualized, each in one case. The intramammarian node was harvested, but the parasternal node was left in situ. 
Median and IR of 10-second count values according to 3 subgroups were as follows; TC, 2050 and 4548; NC, 835 and 1799; CS, 996 and 2079, respectively. SLNs demonstrated higher counts when a larger colloid (TC) was used. The difference was statistically significant (P = 0.01).






SLNs were metastatic in 60 (38.2%) cases. Forty-eight of them were diagnosed during frozen section analysis and 12 during H&E staining or immunohistochemistry. Among 48 patients, 3 had micro- and 45 had macro­metastatis. Ten of 12 metastatic deposits that frozen section failed to demonstrate were micrometastases and 1 was a macrometastasis. In a single patient with 3 negative SLNs, a parasentinel node with a suspicious macroscopic appearance was harvested, and the evaluation of paraffin blocks revealed metastatic involvement.
Except for one patient with micrometastatic deposit, complementary axillary dissection was performed in all patients with positive SLNs and the particular patient with a positive parasentinel node.
Total axillary lymph node number (lymph nodes removed during SLNB and axillary dissection) ranged from 10-51 (median, 27.5; IR, 14.75). The distribution of patients with axillary involvement according to SLNB and axillary dissection procedures, the number of lymph nodes harvested and total metastatic lymph node results are shown in Table 3.
None of the 12 micrometastatic patients had additional metastatic lymph nodes in axillary dissection material. However, additional metastatic lymph nodes were detected (range, 1–24; median, 2; IR, 13) in 19 of 47 (40.4%) patients with macrometastasis in SLNs.
A metastatic SLN was found in 32 of 85 (37.6%) procedures in group A. Twenty-two patients (68.8%) had demonstrated both blue dye and radiocolloid uptake, and 10 patients (31.2%) had only radiocolloid accumulation in their SLNs. In group A, the effect of prior excisional biopsy on blue dye accumulation was evaluated. Isosulfan blue was frequently positive in patients without prior biopsy, but the difference was not statistically significant. Blue dye accumulation was observed in 11 of 18 (61.1%) patients with previous biopsy and in 53 of 67 (79.1%) cases without previous biopsy ( P = 0.116).
Correlation between isosulfan blue accumulation and count rates (counts/10 sec) of hot nodes was also evaluated. A total of 270 SLNs in 84 patients (group A) was examined. Mean counts obtained during 10 sec with the gamma probe were significantly higher in SLNs accumulating blue dye (127 nodes, median ± IR, 1643 ± 3216 counts/10 sec) than in SLNs that did not demonstrate blue dye accumulation (143 nodes, median ± IR, 526 ± 1284 counts/10 sec) (P <0.001).



Discussion
SLNB is currently accepted as the standard method in the evaluation of axillary status of stage I and II breast cancer3,10,11. Almost 75% of early stage breast cancer patients benefit from the technique in terms of axillary dissection-related morbidity12. Despite the cumulative experience on the subject, the ideal SLN localization technique has yet to be defined. A combined method using blue dye and radiocolloid has been proposed as the gold standard by some investigators11. However, many studies have reported no additional benefit with the synchronous use of blue dye or comparable results with the use of blue dye alone13-15.
SLN detection rate varies according to the type of colloid and injection modality. In a recent international multicenter study, overall SLN detection rate was reported to be 97%16. When different injection modalities were compared, periareolar intradermal injection was found superior for lymphatic mapping. In our previous two studies, we demonstrated 97.6% and 98% SLN identification rates using the periareolar intradermal injection technique6,8. The current study is also in concordance with our prior experience, and successful lymphatic mapping was obtained in 99.3% of the cases by injecting various radiocolloids intradermally at the periareolar area. However, our results failed to recommend the use of blue dye as a single agent in SLN identification, since successful SLN detection was observed only in 63 of 81 patients (77.7%). If we had used blue dye as a single agent, then 8 of 26 (30.7%) metastatic SLNs of group A would have been missed. The success rate of blue dye in SLN mapping varies in the literature 11,13-15. The time-dependant clearance of blue dye from the lymphatic basin may be a possible explanation for the different results. Depending on this hypothesis, extension of the interval between the injection of blue dye and SLN dissection will increase the clearance of blue dye. In surgical practice, the time consumed for frozen section analysis of the primary breast lesion is the major cause of such a delay. Our study group included both previously biopsied and non-biopsied patients. When the two groups were compared for blue dye avidity, although not statistically significant, blue dye positivity was more frequent in patients without prior biopsy (82.3% vs 66.7%).
Our results also showed a correlation between radiocolloid uptake and isosulfan blue accumulation. SLNs with isosulfan blue uptake had significantly higher count rates. Additionally, count rates obtained over SLNs were found to be influenced by the size of the colloid. Larger colloids yielded a significantly higher count rate probably due to slower clearance from the SLNs.
SLN number is closely related to the size of the radiocolloid17. In accordance with the relevant literature, the number of SLNs in our study was significantly higher in patients treated with small radiocolloids18,19. Injection modality and size of the colloid have been described as major factors that influence visualization of parasternal nodes20-22. By using deep injections and small colloids, the rate of parasternal node visualization increases. In our study groups, we used both small and large colloids. Parasternal drainage was observed in only one patient in whom a small colloid was used. Our results emphasize the significance of injection type in extra-axillary drainage and are in concordance with the literature and our previous works using superficial injections6,8,23.
We did not successfully detect micrometastatic deposits in SLNs by frozen section analysis. We therefore missed 75% of the micrometastasis during intraoperative frozen section. In spite of general agreement about the positive contribution of rapid immunohistochemistry on intraoperative SLN analysis, there are also some contrary reports24-26. Our technique using H&E staining during frozen section analysis yielded a sensitivity of 79.5% (particularly evident in micrometastatic disease), which was surprisingly comparable with prior reports based on the rapid-immunohistochemistry technique26. In this setting, the need of rapid-immunohistochemistry is not though be to be critical.
Removal of palpable lymph nodes with no radiocolloid or blue dye uptake is of great importance for reducing the false-negative rate in SLNB27. In our patient group, we harvested a non-sentinel and palpable lymph node which proved to be metastatic.
In conclusion, our study reviewing SLNB with a broad perspective revealed a very high SLN detection rate using periareolar intradermal radiocolloid injection. Large and small radiocolloids were comparabily effective in lymphatic mapping, but small colloid administration provided better marking of SLNs, thus more SLNs were harvested by the method. Although the use of blue dye facilitated the procedure by adding a visual component, isosulfan blue was inefficient as a single agent. SLNs possessing higher radiocolloid count rates tended to accumulate blue dye more frequently. SLNs were found to have higher count rates when a larger colloid was used. Frozen section analysis using H&E staining was very successful in the diagnosis of macrometastatic disease in SLNs, but the technique failed to detect most of the micrometastases.
References
1. Giuliano AE, Kirgan DM, Guenther JM, Morton DL: Lymphatic mapping and sentinel lymphadenectomy for breast cancer. Ann Surg, 220: 391-401, 1994.
 2. Haigh PI, Giuliano AE: Sentinel lymphadenectomy in node negative breast cancer. Cancer Treat Res, 103: 25-37, 2000.
 3. Veronesi U, Paganelli G, Viale G, Luini A, Zurrida S, Galimberti V, Intra M, Veronesi P, Maisonneuve P, Gatti G, Mazzarol G, De Cicco C, Manfredi G, Fernández JR: Sentinel lymph-node biopsy as a staging procedure in breast cancer: update of a randomised controlled study. Lancet Oncol, 7: 983-990, 2006.
 4. Motomura K, Egawa C, Komoike Y, Kataoka T, Nagumo S, Koyama H, Inaji H: Sentinel node biopsy for breast cancer: technical aspects and controversies. Breast Cancer, 14: 25-30, 2007.
 5. Benson JR, della Rovere GQ: Axilla Management Consensus Group. Management of the axilla in women with breast cancer. Lancet Oncol, 8: 331-348, 2007.
 6. Yararbas U, Argon AM, Yeniay L, Kapkac M: Problematic aspects of sentinel lymph node biopsy and its relation to previous excisional biopsy in breast cancer. Clin Nucl Med, 34: 854-858, 2009.
 7. Meyer-Rochow GY, Martin RC, Harman CR: Sentinel node biopsy in breast cancer: validation study and comparison of blue dye alone with triple modality localization. ANZ J Surg, 73: 815-818, 2003.
 8. Yararbas U, Argon AM, Yeniay L, Zengel B, Kapkaç M: The effect of radiocolloid preference on major parameters in sentinel lymph node biopsy practice in breast cancer. Nucl Med Biol, 37: 805-810, 2010.
 9. Jimenez IR, Roca M, Vega E, García ML, Benitez A, Bajén M, Martín-Comín J: Particle sizes of colloids to be used in sentinel lymph node radiolocalization. Nucl Med Commun, 29: 166-172, 2008.
10. Veronesi U, Paganelli G, Viale G, Luini A, Zurrida S, Galimberti V, Intra M, Veronesi P, Robertson C, Maisonneuve P, Renne G, De Cicco C, De Lucia F, Gennari R: A randomized comparison of sentinel-node biopsy with routine axillary dissection in breast cancer. N Engl J Med, 349: 546-553, 2003.
11. McMasters KM, Tuttle TM, Carlson DJ, Brown CM, Noyes RD, Glaser RL, Vennekotter DJ, Turk PS, Tate PS, Sardi A, Cerrito PB, Edwards MJ: Sentinel lymph node biopsy for breast cancer: a suitable alternative to routine axillary dissection in multi-institutional practice when optimal technique is used. J Clin Oncol, 18: 2560-2566, 2000.
12. Kell MR, Burke JP, Barry M, Morrow M: Outcome of axillary staging in early breast cancer: a meta-analysis. Breast Cancer Res Treat, 120: 441-447, 2010.
13. Krikanova M, Biggar M, Moss D, Poole G: Accuracy of sentinel node biopsy for breast cancer using blue dye alone. Breast J, 16: 384-388, 2010.
14. Cserni G, Rajtár M, Boross G, Sinkó M, Svébis M, Baltás B: Comparison of vital dye-guided lymphatic mapping and dye plus gamma probe-guided sentinel node biopsy in breast cancer. World J Surg, 26: 592-597, 2002.
15. Bines S, Kopkash K, Ali A, Fogg L, Wool N: The use of radioisotope combined with isosulfan blue dye is not superior to radioisotope alone for the identification of sentinel lymph nodes in patients with breast cancer. Surgery, 144: 606-610, 2008.
16. Straver ME, Meijnen P, van Tienhoven G, van de Velde CJ, Mansel RE, Bogaerts J, Duez N, Cataliotti L, Klinkenbijl JH, Westenberg HA, van der Mijle H, Snoj M, Hurkmans C, Rutgers EJ: Sentinel node identification rate and nodal involvement in the EORTC 10981-22023 AMAROS trial. Ann Surg Oncol, 17: 1854-1861, 2010.
17. Wilhelm AJ, Mijnhout GS, Franssen EJ: Radiopharmaceuticals in sentinel lymph-node detection - an overview. Eur J Nucl Med, 26 (4 Suppl): S36-42, 1999.
18. Leidenius MH, Leppänen EA, Krogerus LA, Smitten KA: The impact of radiopharmaceutical particle size on the visualization and identification of sentinel nodes in breast cancer. Nucl Med Commun, 25: 233-238, 2004.
19. Jastrzebski T, Lass P, Swierblewski M, Wydra D, Drucis K, Kruszewski W, Gulida G, Kopacz A: The influence of radioisotope vehicle on breast sentinel node detection. Eur J Surg Oncol, 32: 928-932, 2006.
20. Park C, Seid P, Morita E, Iwanaga K, Weinberg V, Quivey J, Hwang ES, Esserman LJ, Leong SP: Internal mammary sentinel lymph node mapping for invasive breast cancer: implications for staging and treatment. Breast J, 11: 29-33, 2005.
21. Koizumi M, Nomura E, Yamada Y, Takiguchi T, Makita M, Iwase T, Tada T, Tada K, Nishimura S, Takahashi K, Yoshimoto M, Kasumi F, Akiyama F, Sakamoto G: Radioguided sentinel node detection in breast cancer patients: comparison of 99mTc phytate and 99mTc rhenium colloid efficacy. Nucl Med Commun, 25: 1031-1037, 2004.
22. Mudun A, Sanli Y, Ozmen V, Turkmen C, Ozel S, Eroglu A, Igci A, Yavuz E, Tuzlali S, Muslumanoglu M, Cantez S: Comparison of different injection sites of radionuclide for sentinel lymph node detection in breast cancer: single institution experience. Clin Nucl Med, 33: 262-267, 2008.
23. Somasundaram SK, Chicken DW, Waddington WA, Bomanji J, Ell PJ, Keshtgar MR: Sentinel node imaging in breast cancer using superficial injections: technical details and observations. Eur J Surg Oncol, 35: 1250-1256, 2009.
24. Viale G, Bosari S, Mazzarol G, Galimberti V, Luini A, Veronesi P, Paganelli G, Bedoni M, Orvieto E: Intraoperative examination of axillary sentinel lymph nodes in breast carcinoma patients. Cancer, 85: 2433-2438, 1999.
25. Krishnamurthy S, Meric-Bernstam F, Lucci A, Hwang RF, Kuerer HM, Babiera G, Ames FC, Feig BW, Ross MI, Singletary E, Hunt KK, Bedrosian I: A prospective study comparing touch imprint cytology, frozen section analysis, and rapid cytokeratin immunostain for intraoperative evaluation of axillary sentinel lymph nodes in breast cancer. Cancer, 115: 1555-1562, 2009.
26. Choi YJ, Yun HR, Yoo KE, Kim JH, Nam SJ, Choi YL, Ko YH, Kim BT, Yang JH: Intraoperative examination of sentinel lymph nodes by ultrarapid immunohistochemistry in breast cancer. Jpn J Clin Oncol, 36: 489-493, 2006.
27. Carmon M, Olsha O, Rivkin L, Spira RM, Golomb E: Intraoperative palpation for clinically suspicious axillary sentinel lymph nodes reduces the false-negative rate of sentinel lymph node biopsy in breast cancer. Breast J, 12: 199-201, 2006.