Detection of the sentinel lymph node in breast cancer

in breast cancer Santosh K. Somasundaram, Dennis W. Chicken, and Mohammed R. S. Keshtgar * Department of Surgery, Royal Free and University College Medical School, University College London, London, UK Introduction *Correspondence to: Mohammed R. S. Keshtgar, Department of Surgery, Royal Free Hospital NHS Trust, Pond Street, London NW3 2QG, UK. E-mail: m.keshtgar@ucl.ac.uk Introduction: Introduction: Axillary lymph node status for lymphatic staging in breast cancer is the best prognostic indicator and guides systemic treatment. Sentinel lymph node (SLN) biopsy is a novel, minimally invasive technique for lymphatic staging proven to improve quality of life. The accurate detection of the SLN is paramount for the success of the procedure. Methods: Relevant literature was reviewed with regards to the different dyes and techniques used for the detection of SLN in breast cancer. Results: Highest identification rates and lowest false negative rates are achieved by using the combined blue dye and radiocolloid technique with pre-operative imaging using a gamma camera. There is a well-recognized learning curve to successfully perform SLN biopsy. Conclusions: The concept of SLN has been well validated and is the standard of care in early breast cancer. A multidisciplinary approach and structured training is the key to the successful introduction of the technique. Keywords: sentinel lymph node/breast cancer/sentinel node imaging/new START The primary aims of modern breast cancer surgery are to obtain local and regional control of the cancer and gather sufficient information to make an accurate prediction of the risk of distant metastases in order to guide systemic therapy. In breast cancer, this has traditionally been achieved by resection of the primary tumour (either by mastectomy or by wide local excision) and axillary lymph node dissection (ALND). ALND, however, has significant short- and long-term morbidity, the most significant being lymphoedema. With the trend towards earlier detection and presentation of breast cancer, most patients do not have lymphatic metastases at diagnosis. In these patients, ALND is purely British Medical Bulletin 2007; 84: 117 131 DOI:10.1093/bmb/ldm032 & The Author 2007. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oxfordjournals.org

S. K. Somasundaram et al. a diagnostic procedure, with no therapeutic benefit. Herein lies an irony: with widespread use of breast conserving surgery, the staging procedure carries greater morbidity than the therapeutic procedure of the primary cancer. Sentinel lymph node biopsy (SLNB) is a minimally invasive technique to stage the axilla in breast cancer, without compromising the prognostic information obtained from ALND. The sentinel node concept The sentinel node was defined by Morton et al. as any lymph node(s) receiving direct lymphatic drainage from the primary tumour, and therefore is the first node to become involved when a tumour metastasises. The concept behind SLNB is that lymphatic metastases occur in an orderly manner and that the sentinel node status predicts the histological status of the regional lymph nodes. If the sentinel node does not contain metastases, the draining nodal basin is highly unlikely to harbour metastases and complete nodal dissection is not required. The term sentinel node was suggested by Cabanas 1 30 years ago in his work on penile cancer. He reported an anatomical description of the sentinel node, although subsequent studies by others showed this anatomical approach to have insufficient predictive value for clinical use. The concept was revived by Morton et al., 2 who used vital dyes to simulate the spread of malignant melanoma. This functional approach to identification of the sentinel node proved highly accurate, and was soon adapted to breast cancer by Giuliano et al. 3 Over the last 10 years, the concept of a sentinel node has been extensively validated. Turner et al. 4 performed a histopathological validation of the SLNB and showed that if the SLN is free of tumour, the probability of the non-sentinel node involvement is,0.1%. Numerous clinical studies have compared SLNB with ALND in breast cancer. These studies have been combined into large metanalyses which have validated the technique, demonstrating high sentinel node identification rates and more importantly consistently low false negative rates (Table 1). The results of three randomized controlled trials comparing SLNB with ALND in 2352 patients have been published to date. 9 11 All three have demonstrated that SLNB is an accurate staging technique which results in a significant reduction in morbidity with less postoperative pain, paraesthesia, shoulder stiffness and lymphoedema. Drain usage, operative time, length of hospital stay and time to 118 British Medical Bulletin 2007;84

Table 1 Published metanalyses of clinical validation studies of sentinel lymph node biopsy compared with axillary lymph node dissection for lymphatic staging in breast carcinoma Author Year No. of patients SLN identification rate (%) Sensitivity (%) False negative rate (%) McMasters et al. 5 1998 1385 86 94 6.2 Miltenburg et al. 6 1999 912 84 95 5.1 Gemignani and Borgen 7 2001 3800 88 93 7.6 Kim et al. 8 2006 8059 96 93 7.3 resumption of normal duties after surgery were significantly less in the SLNB patients. The rate of axillary recurrence following a negative sentinel node biopsy is reassuringly low in two large published studies. The Memorial Sloane Kettering Cancer Centre reported a series of 2340 patients with a negative sentinel node biopsy and not subjected to ALND. At a median follow-up of 31 months, three patients (0.12%) had developed an axillary recurrence. 12 The axillary recurrence rate reported by Veronesi et al. for 953 similar patients followed for a median of 38 months was 0.31% (n ¼ 3). The 5 year survival of sentinel node negative patients was 98%. 13 A further benefit of SLNB is the potential for targeted intensive histopathological examination (ultrastaging), by examination of multiple sections of the node and use of immunohistochemical (IHC) stains. This increases the sensitivity of detection of small metastatic deposits, which could go undetected if using traditional pathological examination. Methods for identification of the sentinel node Injection methods The original injection techniques aimed to mimic tumour metastases; hence, the agents were injected adjacent to the tumour in a peritumoural fashion. Better understanding of the lymphatic drainage of the breast allowed the development of superficial injection techniques. Lymph drains from the deep breast parenchyma superficially to the rich subdermal lymphatics. Subdermal lymphatics drain to the axilla via the subareolar plexus through one or more final common lymphatic channels. Rapid tracer migration and superior identification rates are achieved by injection of the blue dye and colloid either intradermally overlying the tumour or superficially at the periareolar margin. 14,15 Most units have now adopted these superficial injection techniques, which are easy, British Medical Bulletin 2007;84 119

S. K. Somasundaram et al. highly reproducible, and often show the lymphatic tracts, which may be useful for differentiating the sentinel node from second echelon nodes. The main drawback of the superficial technique is that it rarely demonstrates internal mammary nodes. If an internal mammary drainage is suspected, a deeper intra or peri-tumoural injection technique is recommended. Radiocolloids The use of radiocolloids for sentinel node identification offer several advantages. The colloids are efficiently trapped in the sentinel node (whereas blue dyes typically pass into second echelon nodes). Radiocolloids enable pre-operative sentinel node imaging, and facilitate rapid and easy intraoperative identification by the surgeon using a gamma probe (described later under surgical technique ). Several studies have shown better sentinel node identification rates, when compared with blue dye alone. 16 There is variation in the colloids used worldwide. The original studies conducted within the USA used sulphur colloids, and these are still used to date. In Europe, albumin colloids are used most widely, and are sold as a commercially available kit. Antimony trisulphide is the colloid of choice in Australia. The particle size of the colloid used is important: particles of 40 80 nm are small enough to migrate rapidly, yet large enough to be efficiently trapped within the sentinel node. The universal radioactive tracer used is Technetium-99 m. Technetium has several advantages. (i) It is a pure gamma radiation emitter, hence offers excellent tissue penetration. (ii) Comparatively safe form of radiation when compared with alpha and beta particles. (iii) Short half-life of 6 h, hence decays rapidly. Surgery therefore needs to be performed within 24 h of radiocolloid injection. (iv) Cheap and readily available in every nuclear medicine department. Although the benefits of using radiocolloids are apparent, there are several disadvantages, particularly complicated and restrictive legislation governing the administration of radiopharmaceuticals to patients and the handling of radioactive materials. The use of radiocolloids results in radioactive contamination of all swabs and drapes used for an operation, and careful protocols need to be designed to appropriately deal with this waste material. Surgeons and theatre staff need to be appropriately trained to manage waste. A further disadvantage is 120 British Medical Bulletin 2007;84

Sentinel node imaging the need for an additional pre-operative investigation, the timing of which needs to be coordinated with operating lists. Radiocolloid injection allows pre-operative sentinel node imaging using a gamma camera. Sentinel node imaging is different from lymphoscintigraphy, although the terms are erroneously used interchangeably. Classical lymphoscintigraphy aims to demonstrate the entire lymphatic tree, to identify abnormalities in lymphatic drainage. This is quite distinct from sentinel node imaging, where the aim is to identify the first draining lymph node. Figure 1 shows an example of sentinel node imaging. The imaging criterion for a sentinel node is a radioactive ( hot ) node ideally with a tract draining from the injection site. Pre-operative imaging identifies the draining lymphatic basin and number of sentinel nodes. This is helpful when sentinel nodes lie in unusual locations (such as intramammary, internal mammary or supraclavicular nodes) (Fig. 2). Frequently more than one sentinel node is shown on sentinel node imaging. Identifying a sentinel node on pre-operative imaging is highly predictive of the success of the subsequent surgical procedure. It provides the surgeon with a road map, although the precise anatomical location of Fig. 1 Sentinel node imaging. Left anterior oblique views above and left lateral views below. Radiocolloid has been injected in the periareolar region. Two radioactive ( hot ) nodes can be seen in the axilla. A single direct tract leads from the injection site to the lower node, which is therefore the sentinel node on imaging criteria. British Medical Bulletin 2007;84 121

S. K. Somasundaram et al. Blue dye Fig. 2 Sentinel node imaging showing two sentinel nodes: Direct lymphatic tracts from the injection site lead to a lymph node in the left axilla and another to an intramammary node (arrowed). the node can only be determined using the intra-operative gamma probe. There is continuing debate about the precise value of pre-operative sentinel node imaging. Gamma cameras are expensive capital items. Some authors have argued that sentinel node identification rate is neither enhanced nor management altered by this expensive step in sentinel node identification. 17 Injection of blue dye enables the surgeon to identify blue-stained lymphatic tracts draining from the tumour. Following these tracts allows identification of the first draining lymph node. Several dyes which have a common characteristic of weak binding to albumin are used for this purpose. In Europe, Patent blue dye is usually used. Patent blue dye is a purple/blue food colourant (E131), which has been banned from foods in the USA, Norway and Australia due to the risk of allergy. Isosulphan blue is a widely used alternative. Both these dyes carry a risk of allergy in 1 2% of patients, which may range from trivial skin rashes to life-threatening anaphylaxis. 18 Methylene blue has been suggested as an alternative, and has been successfully used for sentinel 122 British Medical Bulletin 2007;84

Combination technique node identification. The risk of allergy appears lower, although it does induce an intense tissue reaction, which may result in skin necrosis if injected superficially. 19 Injection of blue dye causes skin staining, which typically lasts several months, but may rarely cause permanent tattooing. All blue dyes enter the circulation, which may make patients appear cyanotic and interfere with pulse oximetry. Patients should be warned that the dye is excreted in urine post-operatively. Sentinel node identification using blue dye alone is a difficult technique to learn and requires a wider exposure of the surgical wound to trace the afferent lymphatics to the tail of the breast. Metanalysis shows that the sentinel node identification rate is lower and the false negative rate higher than using radiocolloid in isolation or a combination of techniques. 20 Employing two complimentary techniques for sentinel node identification will logically improve the sentinel node identification rate and reduce false negative biopsies. 21 This has been clearly shown in numerous studies. In a metanalysis reported by Cody et al., 20 the combined technique gave the best identification rate of 91% and the lowest false negative rate of 5%. Using two modalities is easier, and there is evidence that the wellrecognized learning curve to perform sentinel node biopsy is shorter when using the combination technique; if one technique fails, the other may succeed. The combination technique enables identification of sentinel nodes in unusual locations. There is therefore consensus that the combined blue dye and radiocolloid is the preferred technique. 22 Within the UK, this is the technique advocated in the national training programme (NEW START) for sentinel node biopsy in breast cancer. Surgical technique Patent blue dye is injected in the periareolar region after anaesthetizing the patient. The injection site is gently massaged to aid migration of the dye through the lymphatics. The patient is positioned with the ipsilateral arm abducted. The axilla is systematically scanned with a gamma probe (Fig. 3). Prior to this, a background count of the radioactivity is recorded for comparison with any residual activity following the procedure. The site of highest radioactivity is marked and a small British Medical Bulletin 2007;84 123

S. K. Somasundaram et al. Fig. 3 Systematic scanning of the axilla with a gamma probe enables the site of highest radioactivity to be determined. incision made. The probe provides audio feedback, which enables the surgeon to localize the SLN using the principle of line of sight. It provides three-dimensional orientation about the precise location of the SLN in the axilla. The surgical criteria for an SLN are a hot and blue node, a blue node with blue afferent lymphatic tracts or a hot node with 5 10 times the background radioactivity or more than 10% of the activity of the hottest node (Fig. 4). After harvesting the SLN, the probe is used to identify any other hot nodes by checking the residual counts in the axilla. After biopsy of the hot and blue nodes, the axilla is carefully palpated to identify any grossly involved node(s) which may be neither hot nor blue. A surgical drain is rarely required after an SLNB. In many centers, sentinel lymph node biopsy is done prior to the breast surgery to allow time for an intra-operative examination of the SLN. Intra-operative diagnosis of SLN metastasis enables the surgeon to proceed with ALND. This avoids a second operative procedure 124 British Medical Bulletin 2007;84

Fig. 4 A blue afferent lymphatic vessel draining into a blue-stained node which is radioactive ( hot ) is the best operative definition of a sentinel node. and anaesthetic for these patients. Intra-operative diagnosis, however, introduces uncertainty for patients, as on induction of anaesthetic, they do not know the extent of the operation they are to undergo. Waking to find a surgical drain implies a more extensive surgical procedure as well as worse prognosis node-positive disease. Despite this, given the choice, most patients opt for intra-operative diagnosis, although the psychological impact of this is yet to be determined. 23 Operation lists have to be scheduled accordingly to allow extra time for ALND in patients with SLN metastases detected intra-operatively. Histo-pathological examination of the sentinel lymph node The commonly used techniques for intra-operative detection of sentinel lymph node metastases are touch imprint cytology (TIC) and frozen section histology. The main drawback is that they require expert, welltrained cyto-pathologists to report results accurately in a short time. A further novel technology that is being used is the reverse transcriptase polymerase chain reaction (rt PCR), a rapid nucleic acid amplification method that appears to accurately reflect the metastatic status of the sentinel nodes intra-operatively. Clinical trials of the technique are underway. The biopsied sentinel lymph nodes are serially sectioned at a minimum of three levels depending on the size of the SLN so as British Medical Bulletin 2007;84 125

S. K. Somasundaram et al. not to miss any metastases less than 2 mm. They undergo routine Haematoxylin and Eosin (H&E) staining and nodes with no evidence of metastases on H&E staining undergo further IHC staining for cytokeratin antibodies. The SLNs are thus ultrastaged as H&E positive, IHC positive or negative. Pre-operative imaging of axilla One of the exclusion criteria for SLNB is clinical, histological or radiological evidence of regional nodal metastases. There are clear advantages in identifying patients with nodal metastases preoperatively, as node positive patients do not benefit from SLNB and can therefore proceed directly to ALND. Clinical examination of the axilla, even by experienced surgeons, is notoriously inaccurate. The axilla can be more accurately assessed by an ultrasound scan, which is simple, inexpensive and readily available. The ultrasound criteria for a suspicious node include size.5 mm and morphological characteristics (round, hypoechoic, eccentric cortical hypertrophy with loss of central hilum). Ultrasound in isolation has a low specificity which can be enhanced by proceeding to guided fine needle aspiration cytology or trucut biopsy. 24 Currently, no imaging technique is capable of identifying microscopic tumor spread and hence SLNB remains the diagnostic procedure of choice for axillary staging. Limitations/problems with current standard techniques A well-recognized pitfall of sentinel node identification using the combination technique is a grossly metastatic sentinel node causing a blockage to the flow of lymphatic fluid through the afferent lymphatics. This leads to opening of alternative lymphatic channels and consequently both blue dye and radiocolloid may be diverted away from the true sentinel node. To overcome this, careful palpation of the axilla through the operative wound is recommended, to identify grossly metastatic nodes. The handling, disposal and administration of radioactive materials to patients are governed by a series of complex regulations. These regulations can appear daunting to the uninitiated, although it should be recognized that every nuclear medicine department functions within these laws every day! Some have argued for identification of the sentinel node by using blue dye only, to avoid the challenges introduced by the use of radiocolloid. The authors view is that denying patients the 126 British Medical Bulletin 2007;84

benefit of using the combined technique to avoid legislative restrictions is not acceptable practice. Research continues into alternative tracers to identify the sentinel node. Fluorescent dyes (such as Indocyanine Green) and more recently fluorescent quantum dots may enable combination technique sentinel node biopsy, without radiocolloid nor the need for an expensive gamma camera. 25,26 An alternative investigational approach is the use of magnetic particles, which may be detected by an instrument sensitive to magnetism. 27 Training in sentinel node biopsy There is a well-documented learning curve to sentinel node biopsy. 28 Successful identification of the SLN is directly related to the surgeon s experience. The key to success is a multi-disciplinary approach with the surgeon, nuclear medicine physician, histopathologist, nursing and theatre staff acquiring the knowledge and skills to enable successful introduction of the technique. The aim of structured training is to avoid a false negative sentinel node biopsy. A false negative sentinel node biopsy would result in understaging, with a consequent false sense of security and the possibility of systemic undertreatment for those patients likely to benefit most. In the UK, a structured training programme called NEW START has been developed to teach a standardized technique. This programme consists of theoretical teaching, on-site proctored training of the surgeon for five cases, followed by performing an audit series of sentinel node biopsy and immediate ALND in a further 25 patients. The aim of the audit series is to verify that the surgeon with the assistance of the multidisciplinary team is able to identify the sentinel node with a high identification rate and more importantly a low false negative rate. A localization rate of.90% and false negative rate of,10% is required for certification. A simulator has been developed, which is able to simulate all key new skills required to successfully perform the procedure. 29 This simulator enables surgical skills laboratory training which acts as an interface between theoretical training and performing the procedure on live patients. Controversies In the wake of SLNB, new controversies have arisen. Ultrastaging of the axilla by enhanced pathological examination of the SLN enables British Medical Bulletin 2007;84 127

S. K. Somasundaram et al. the identification of micrometastases (,2 mm), submicrometastases (,0.2 mm) and isolated tumour cells (detectable by IHC staining only). The significance of these tiny metastases for predicting prognosis (hence influencing systemic therapy) or predicting further axillary metastases (hence influencing the decision to proceed to ALND) is, however, unclear. The American College of Surgeons Oncology Group (ACOSOG) is examining this issue in a large trial (Z10) which is yet to be reported. Biopsy of internal mammary chain sentinel nodes remains an area for research. A small percentage of patients are upstaged by internal mammary node biopsy, but changes in systemic therapy are made in an even smaller proportion. There is the potential for additional morbidity such as haemothorax or pneumothorax as a result of internal mammary chain biopsy. The value of radiotherapy to the internal mammary nodes is the subject of ongoing trials in Europe and Canada. 30 The role of SLNB in ductal carcinoma in situ (DCIS) is controversial. Pure DCIS has by definition no capacity for metastasis. Microinvasive DCIS has a very low rate of nodal positivity. Ten per cent of patients pre-operatively diagnosed with DCIS are found to have a co-existing invasive carcinoma, and these patients benefit from nodal staging. SLNB is warranted for extensive DCIS requiring treatment by mastectomy and microinvasive DCIS. 31 Opinions differ about SLNB in patients undergoing neoadjuvant chemotherapy. These patients typically have large tumours, and hence are more likely to be node positive. Although it appears that sentinel node biopsy can still be accurately performed after neoadjuvant chemotherapy, nodal metastases may respond well to chemotherapy and therefore regress completely. Regression of nodal metastases is a positive prognostic sign. Sentinel node biopsy as a separate procedure prior to neoadjuvant therapy enables accurate lymphatic staging, although of course does not alter decisions about chemotherapy! The optimal management of patients with a positive sentinel node is yet to be established. Most surgeons currently proceed to completion ALND. Unless intra-operative diagnosis of sentinel nodes is utilized, this may require a second surgical procedure. Axillary irradiation may offer equivalent regional control with lower morbidity and without the need for a second operation. This question is being addressed by the AMAROS trial, conducted by the European Organisation for the Research and Treatment of Cancer (EORTC). An alternative approach is close observation of the sentinel node positive axilla, reserving axillary dissection for those patients who develop overt axillary metastases. The rationale for this is that ALND confers no clear survival benefit, and finding additional metastatic 128 British Medical Bulletin 2007;84

nodes beyond the sentinel node is unlikely to change decisions about systemic therapy. This question was addressed by the American College of Surgeons Oncology Group (ACOSOG) multicentre trial Z11. Results are awaited, but target patient recruitment was not achieved and the study may therefore be underpowered to provide an answer. A further approach to patients with a positive sentinel node is to attempt to predict those patients with non-sentinel node axillary metastases, and to reserve ALND for those patients at high risk. A predictive nomogram has been developed by Van Zee et al. 32 and calculations can be performed using a web-based tool. External validation of this predictive tool has shown conflicting results, suggesting that it may require further refinement. 33 Until the results of the trials underway are known, most surgeons proceed to axillary dissection when the sentinel node shows metastases. Key points for clinical practice Surgical lymphatic staging remains integral to modern cancer management. Sentinel node biopsy is the standard of care for lymphatic staging of breast cancer. The sentinel node concept in breast cancer has been extensively validated. Current techniques for sentinel node identification have been optimized. Sentinel node biopsy enables accurate, minimally invasive lymphatic staging while avoiding the morbidity of routine lymph node dissection for node negative breast cancer patients. The improved quality of life after breast cancer surgery has been confirmed in several large randomized controlled trials. Intensive pathological examination of the sentinel node enables the detection of low volume metastases (ultrastaging). The clinical significance of isolated tumour cells and micrometastases less than 0.2 mm is yet to be determined in beast cancer. The current standard of care for sentinel node positive breast cancer patients is ALND, although this may change with the result of major trials which are underway. The major disadvantage of sentinel node biopsy is understaging as a result of a false negative sentinel node biopsy. Adequate multidisciplinary training is the key to avoid this. Sentinel node biopsy research continues, and may yield new tracers and result in changes in the management of patients with a metastatic sentinel node. British Medical Bulletin 2007;84 129

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