Sentinel Lymph Node Biopsy for Squamous Cell Carcinoma

In the head and neck, squamous cell carcinoma (SCC) encompasses 2 distinct clinical entities: aerodigestive tract SCC (commonly referred to as head and neck SCC; HNSCC) and cutaneous SCC (cSCC). It is a malignant tumor of keratinizing epidermal cells. For both HNSCC and cSCC, the most common route of metastasis is lymphatic. However, both the investigation and the treatment of a patient with a clinically N0 nodal basin remain controversial. Particularly in oral cavity SCC, up to 30% of patients have subclinical (not found on examination or imaging) metastases in the neck, and knowledge of lymph node disease alters surgical and postoperative management.[1, 2, 3]

The sentinel lymph node

The sentinel lymph node (SLN) is the first lymph node to drain a metastatic tumor cell that drains via the lymphatic route. The concept of the SLN is based on the orderly progression of tumor cells within the lymphatic system. Mapping of the lymph flow from the tumor site to the regional lymphatic drainage area can be used to identify the primary draining lymph node (ie, SLN). If the SLN can be identified and examined for the presence of tumor metastases, an elective lymph node dissection for staging may not need to be performed.

The concept of the SLN originated in 1977 when Cabanas described mapping of the first lymph node–draining penile carcinoma. In 1977, Robinson et al described the use of cutaneous lymphoscintigraphy in the nodal basin for truncal melanomas using colloidal gold scanning. The development of lymphatic mapping at the end of the 1980s was a breakthrough in making the sentinel node concept applicable to various types of malignancies, particularly breast cancer and melanoma. In 1993, Alex et al introduced the use of technetium-99m sulfur colloid, a radioactive tracer, which is injected intradermally around a primary melanoma site, followed by an imaging study and subsequent intraoperative use of a gamma probe to localize the sentinel node.[4] This technique is now commonly practiced for early-stage melanoma without clinically evident nodal metastasis. Alex and Krag performed the first successful SLN biopsy (SNLB) of the head and neck on a patient with a supraglottic carcinoma.[5]

Relevance of SLNB in SCC

Approximately 3.3 million patients are diagnosed with 5.4 million cases of keratinocyte skin cancer (cSCC or basal cell carcinoma) in the United States each year. Approximately 80% of ultraviolet light–induced SCCs develop on the arms, head, or neck. The frequency of cSCC, as with all nonmelanoma skin cancers, is increasing. Recent projections suggest 300,000-400,000 new cases of cSCC per year.

cSCC is the second leading cause of skin cancer death after melanoma, and it is the second most common type of nonmelanoma skin cancer after basal cell carcinoma. Most cSCCs occur on the sun-exposed areas of the head and the neck. Nodal metastasis is relatively uncommon, only occurring approximately 3% of the time, and it is even less likely in early-stage disease (T1-T2).[6] While it is uncommon, patients with aggressive disease have a high risk of development of nodal metastasis that may be subclinical at initial presentation.[7]

By 2017 estimates (American Cancer Society Facts and Figures),[8] approximately 70,000 cases of HNSCC are diagnosed in the United States each year. SCC can occur in any mucosal area of the head and neck, including the nasal cavity and sinuses, nasopharynx, oral cavity (including lower lip), oropharynx, larynx, and hypopharynx. Smoking and alcohol use are historically the primary risk factors for HNSCC, but the incidence of human papillomavirus (HPV)–related cancers of the oropharynx is rising rapidly. The latter rarely present without nodal disease, so SLNB is unlikely to have a role in the management of HPV-related tumors of the oropharynx. SLNB has primarily been investigated in lesions of the oral cavity, given the historically high rates of nodal metastasis noted above and the accessibility of these tumors for radiotracer injection. It can have a significant impact on the need for lymph node dissection.

Positron-emission tomography (PET-CT), contrasted CT, and MRI are commonly used to classify tumors and nodal or distant metastases of SCC. Definitive staging of lymph node disease can only be achieved by performing a staging lymph node dissection. For mucosal disease in its early stages (T1-T3, N0), clinicians have to balance the benefit of performing an elective (prophylactic) lymph node dissection against the associated morbidity. For cSCC, elective neck dissection is uncommon.

For more information, see Head and Neck Cancer and Head and Neck Squamous Cell Carcinoma.

Etiology

Any of the following may cause SCC:

• Exposure to sunlight

• Chemical carcinogens, such as arsenic and hydrocarbons

• HPV

• Ionizing radiation

• Cigarette smoke

• Chronic irritation or ulceration

• Alcohol

In addition, immunocompromised patients have a much higher risk of developing SCC. Two genes, PATCHED and TP53, have been identified that usually prevent cancers but are inactivated in patients with SCC; TP53 is mutated in more than 90% of patients with SCC.

Clinical manifestations

cSCCs typically manifest on the head, neck, or arms. They usually have elevated and rolled edges with central ulceration. Well-differentiated SCCs are likely to manifest as firm erythematous nodules of varying sizes, sometimes with an area of central hyperkeratosis. The tumor is usually firm, although tumors in more advanced cases can be soft and friable. Erosion and ulceration are more common with SCCs. Poorly differentiated SCCs are more apt to manifest as faintly erythematous nodules or plaques that are not well defined; ulceration is also common.

See the image below.

Large, sun-induced squamous cell carcinoma (SCC) on the forehead/temple. Image courtesy of Glenn Goldman, MD.

HNSCC typically manifests as an irregular, at times ulcerated, friable mass of the oral, oropharyngeal, laryngeal, hypopharyngeal, nasopharyngeal, or sinonasal mucosa. Lesions appearing in the oral cavity are subclassified according to anatomic location; the lower lip can exhibit HNSCC on the mucosal surface or cSCC on the skin surface. Oral cavity HNSCC is most amenable to SLNB because of the generally easy accessibility of the primary lesions.

Successful SLNB

Defining a successful SLNB accurately is critical. Although some studies have examined the impact of an individual surgeon's experience on the SLN identification rate, SLN identification is clearly not an appropriate endpoint; many studies have documented excellent SLN identification rates with unacceptably high false-negative rates, typically defined as a negative SLNB when there is either a metastatic lymph node at the time of surgery or recurrence in the nodal basin where the SLN was identified.[9] The more important issue is the experience required to achieve an acceptably low false-negative rate. Such rates are typically identified through prospective studies in which the neck is dissected after an SLNB to provide a point of comparison. In cases in which the pathology results from the scintigram are unclear or are negative, a formal elective neck dissection should be considered for staging purposes.

This procedure is multidisciplinary. As such, surgeons must ensure that experienced nuclear medicine specialists, radiologists, and pathologists are involved. The learning curve for individual surgeons is undoubtedly associated with the experience levels of specialists within the multidisciplinary team. The implications of the SLN procedure must be effectively communicated to radiation oncologists and medical oncologists. The coordination of effort among the various specialists in each discipline is an essential component of the learning process.

The role of the SLNB in the management of SCC of the head and the neck will evolve as more centers actively perform the technique.[10, 11, 12]

Patient education

For patient education information, see the Cancer and Tumors Center, as well as Skin Cancer, Skin Biopsy, and Cancer of the Mouth and Throat.

SLNB in cSCC

SLNB has been used in larger cutaneous tumors (at least T2) with clinically negative lymph node evaluation. Data from a systematic review show a positive SLN rate of 13.5% in patients carefully selected for the procedure.[7]  A 5-year retrospective case series examining 93 SLNBs found a positive SLN rate of 5.4% (5 biopsies). The tumors associated with positive SLNBs were all staged as T2b.[13] Given that elective nodal basin dissection is rarely performed in cSCC, SLNB can be used to determine the need for therapeutic nodal dissection. While the survival benefit of such an approach has not yet been fully elucidated, it stands to reason that diagnosing subclinical nodal metastasis and providing complete treatment of locoregional disease is likely to improve both disease-specific and overall survival. However, a retrospective study of 720 patients with high-risk cSCC with a tumor thickness greater than 5 mm concluded that, based on the current data, SLNB does not provide any benefit as to future metastasis or tumor-specific survival.[14] As in melanoma, the challenge of performing SLNB in cSCC of the head and neck is that the predictability of SLN location is lower and the likelihood of multiple SLNs is higher than in other body locations.[9, 15] Identification of multiple SLNs can result in the need for multiple operations to evaluate disease status.

SLNB in HNSCC

SLNB has been studied primarily in HNSCC of the oral cavity. The primary role is in cases in which an elective neck dissection is considered, which often occurs when the primary tumor is staged as T1 or T2. In a large multicenter European trial, a negative SLNB in T1-2N0 oral cavity SCC was sufficient to rule out nodal metastasis in 86% of cases, producing a negative predictive value of 95%.[16] This approximates the negative predictive value of SLNB in melanoma. While historically floor-of-mouth tumors were believed to be inappropriate for SLNB because of “shine-through” (misleading detection of signal from the primary injection site), this association was not observed in the study. Yang et al found that SLNB had high sensitivity and high negative predictive value (NPV) for cT1/T2N0 SCC of the tongue.[17] Also, a prospective Japanese trial that included T2 and T3 tumors found benefit for directing neck dissection with SLNB.[1] Although shine-through is an acknowledged limitation of Tc-99m radiotracer use in SLNB, substituting SPECT CT with MR lymphography with a gadolinium-based contrast agent may overcome this limitation. A literature review found this MR lymphography approach had a sensitivity of 90.9% with a NPV of 92.8%.[18] Further trials are needed to verify that SLNB alone is sufficient to provide sufficient staging, and thus prognostic, information. Important to note, a 2015 multicenter trial in the United States demonstrated efficacy of SLNB in appropriately selected oral cavity cancers when using technetium (Tc) 99m tilmanocept.[19] Data from experienced centers continue to support further investigation and careful implementation of these techniques.[20]

Contraindications to SLN biopsy are clinical lymphadenopathy (based on physical examination or imaging), disruption of lymphatic drainage, prior extensive surgery (eg, dissection of the neck), previous radiation to the head and the neck, and adverse reaction to radiotracer compound.

SLNB can be a valuable staging technique for patients with cSCC or HNSCC whose lymphatic drainage has not been altered by previous surgery or radiotherapy. It provides reliable detection of micrometastasis, indicating the need for directing therapy (lymphadenectomy or radiotherapy) to the draining nodal basin. This technique also allows the surgeon to accurately plan a dissection of the neck, taking into consideration the pattern of lymphatic drainage of each patient, for example, understanding if the lymphatic drainage is unilateral or bilateral, or if multiple distinct nodal basins are involved. For example, a midface cSCC could drain to the parotid nodes, facial nodes, upper cervical nodes, or more than one of these sites. In the setting of elective surgical nodal treatment, all of these would need to be addressed, which would present substantial and often unnecessary morbidity to the patient. Sentinel lymph node mapping clarifies the drainage pathway(s) in this setting. In this way, unnecessary dissection and associated morphofunctional sequelae can be avoided in numerous patients.

Multiple studies have reviewed the outcome of patients undergoing sentinel lymph node biopsy of the head and neck. It has been shown that positive SLNB correlates with poorer disease-specific survival and overall survival in HNSCC, while not definitely affecting survival in cSCC.[1, 7, 16] Two major studies showed evidence of benefit from mapping of aberrant drainage pathways that directed dissection to levels not always included in an elective neck dissection (IIB, IV).[1, 16] At a minimum, data show that SLNB does not appear to adversely affect outcome. No studies exist that have directly compared morbidity associated with SLNB to that of elective nodal dissection, but it appears overall that SLNB provides a promising way to decrease morbidity while achieving the same or improved disease control for HNSCC and cSCC.

Complications of SLN biopsy

Allergic reactions to radiotracer, nerve injury, lymphedema, neuropathy, fat necrosis, seroma, and hematoma can result following SLNB.

Patients primarily are educated that sentinel lymph node (SLN) biopsy (SLNB) is most valuable for providing staging information to determine necessary treatment. It is important to educate patients that this is not a therapeutic technique, but rather a diagnostic procedure that allows the treatment team to plan more effective treatment. Risks of the surgery should be reviewed in the consent and should cover relevant risks of a neck dissection, including bleeding, infection, numbness, facial weakness, shoulder weakness, tongue weakness, injury to major vessels, chyle leak, and need for further surgery. The consent process needs to cover all potential procedures, particularly if a completion neck dissection may be performed within the same procedure. Patients also need to understand that additional surgery may be necessary based on the findings.

Patient education can be challenging regarding head and neck SLNB, owing to the "if, then" scenarios that are encountered depending on results. This can be improved by using preoperative single-photon emission computed tomography (SPECT) imaging, which better evaluates the position of the SLN(s) and allows surgeons to better predict the required procedure(s) compared with simple gamma imaging (see below). If reviewing SPECT images with patients, it must be made clear that although these images look similar to positron-emission tomography (PET-CT) imaging, the “hot spots” represent pathways to be evaluated and not sites of disease per se.

As noted in other sections, the technique of SLNB is dependent on successful coordination between the surgeon and the nuclear medicine team, including consistent injection of radiotracer, imaging, and review of results with the surgeon. Many surgeons argue for frozen section analysis with completion neck dissection if a positive node is found. However, only limited nodal assessment can be performed using intraoperative evaluation. As with melanoma and breast cancer, SLNs excised from patients with head and neck squamous cell carcinoma (HNSCC) or cutaneous squamous cell carcinoma (cSCC) should undergo step-sectioning (see below) to thoroughly evaluate for occult microscopic disease.

A radiotracer probe is necessary to identify the lymph node intraoperatively. Some surgeons prefer to have a separate probe available prior to surgery to verify the planned area of dissection before preparing and draping the patient. Typical instrumentation and equipment for a neck dissection are adequate for this procedure. Nerve monitoring is not routinely used, but it may be useful in some settings (eg, intraparotid nodes, level V cervical nodes).

If SLNB is performed without neck dissection, surgery is often performed in an outpatient setting, depending on the primary tumor resection. For SLNB performed with associated neck or other nodal basin dissection, patients are often admitted for 1-2 nights. Patients undergoing extensive dissection often have a drain placed postoperatively. Follow-up is typically performed 7-14 days postoperatively for wound check and to review results and discuss additional treatment.

Physical examination and imaging are performed to evaluate the expected draining nodal basin of the HNSCC or cSCC being assessed. CT, MRI, ultrasound, and PET-CT are all commonly used in this process based on local resources, skillsets, costs, and clinical preferences.

In the setting of SLNB, imaging studies are performed to detect macroscopic lymph node metastases. If the imaging studies fail to show lymph node metastases, an SLNB is indicated depending on the likelihood of occult nodal disease from the primary tumor. For example, cT2N0 oral cavity SCC has an estimated risk of greater than 20% of occult cervical nodal disease and elective neck dissection has been advocated for these patients. In this setting, SLNB has the potential to avoid neck dissection in nearly 80% of cases. Similarly, while all cSCC in the head and neck carries only a 3-5% rate of metastasis, for advanced cSCC having high-risk features (eg, >2 mm thickness, incomplete excision, perineural invasion), risk of occult nodal disease increases with each high-risk feature. In this setting, SLNB can direct the need for local-only versus locoregional therapy.

Lymphoscintigraphy

Patients undergo lymphoscintigraphy up to 1 day prior to surgery. A maximum of 40 MBq technetium-99m–labeled human serum albumin (radiotracer (ie, AlbuRES, Nanocoll, Lymphoseek) is injected throughout the normal skin or mucosa surrounding the tumor edge and the dermis or submucosa on the deep aspect of the tumor in a volume of approximately 0.5-1 mL. A syringe with a permanently secured needle is used for injection, thereby preventing inadvertent spillage of radiotracer. Colloid is injected at as many points as necessary in an attempt to completely surround the tumor. For oral cavity injections, a mouthwash is used immediately following injection to prevent pooling or swallowing of residual radioactive material by the patient.

Static lymphoscintigraphy is performed at 15 minutes, 30 minutes, and 1 hour postinjection in two planes or until the appearance of radioactive nodes. Hot spots are usually seen 15 minutes postinjection. If nodes are still absent 1 hour after injection, the lymph nodes are too close to the injection site or radiocolloid has leaked out of the injection site. SPECT has largely replaced static lymphoscintigraphy in which an additional CT is performed onto which the lymphoscintigraphy is overlain to improve the anatomic localization of SLNs.

Multiple colloids are commonly used for lymphoscintigraphy: AlbuRES, Nanocoll, and, more recently, Lymphoseek, have gained favor, particularly in oral cavity SCC. AlbuRES has a mean particle size of 500 nm and is a slower-moving particle that remains in first-echelon (sentinel) nodes but requires a high density of terminal lymphatic vessels at the injection site. For these reasons, AlbuRES is the colloid of choice on the tongue and on the floor of the mouth. Nanocoll has a mean particle size of 50 nm and is a faster-moving colloid, which finds lymphatic vessels despite injection into tissues with low densities of terminal lymphatics. Nanocoll also moves readily from sentinel nodes to subsequent-echelon nodes. For these reasons, Nanocoll is the colloid of choice for primary tumors that are not located on the floor of the mouth or on the tongue. Lymphoseek (technetium Tc 99m tilmanocept) is a diethylenetriaminepentaacetic acid and mannose compound that rapidly diffuses within 15 minutes and stays present in tissue up to 30 hours. For this reason, it is gaining popularity in use for sentinel node mapping.

While historically blue dye and radioactive mapping had been combined, the consistent success of SLNB by radioactive mapping (across multiple studies approaching 95%) has led to preferential use. Intraoperative use of blue dye has many associated problems that have resulted in a decline in its use, particularly as the standard radiotracer technique has improved.

Anatomic planning

Anatomic planning should follow similar principles to standard neck dissections. See Neck Dissection Classification.

At operation, 1-2 mL of Patent Blue V dye can be used, although this has decreased in popularity. To approximate the same injection sites as for radiocolloid, ensure that the same person performs all the injections. A suitable incision is made in the neck in such a position as to facilitate excision of the incision scar if a subsequent neck dissection is necessary.

The handheld gamma probe is used to identify radioactive sentinel nodes, including those marked preoperatively during lymphoscintigraphy. To reduce detection of radiation from the injection site, a series of malleable, sterilized lead plates may be used to mask the injection site, thus aiding in vivo identification of radioactive nodes. Radioactive nodes are excised, and radioactivity within the node is confirmed ex vivo.

If the blue dye is used, stained lymphatics, if seen, are followed to the first draining lymph node, which is then harvested. Sentinel nodes are labeled according to their color and radioactivity. The anatomic neck level of the sentinel nodes is noted. Although sentinel nodes are usually harvested prior to treatment of the primary tumor, the proximity of the sentinel node to the injection site may require a further search for sentinel nodes following excision of the primary tumor. If sentinel nodes are sought after excision of the injection site, the nodes are not likely to be stained blue.

Because of the relatively high radioactivity present in the injection sites and the proximity to the sentinel node, detection of scattered radiation must be avoided as far as possible. This is particularly true in the floor of mouth. In addition to the use of the lead plates described above, the gamma probe must have a well-collimated detector, which excludes gamma radiation except over a small angle in front of it. Set the pulse-height analysis window to only include the technetium-99m photopeak with a cutoff on the low-energy side at approximately 130 keV. Check the calibration at regular intervals of not more than 1 month (depending on the make and model of the instrument). Devise a quick check of calibration, and perform this quick check before each use. Calling on appropriate scientific or technical assistance may be necessary to ensure the gamma probe is at its optimal settings and to make an estimate of its sensitivity at these settings.

Sentinel nodes are fixed in 10% neutral, buffered formalin, and, after fixation, they are bisected through the hilum (if identifiable) or through the long axis of the node. If the halves are thicker than 2 mm, the slices are further trimmed to provide additional blocks of 2 mm. If sentinel nodes are found to be free from tumor after the initial histologic examination, step-serial sections are prepared at an additional 6 levels in the block at intervals of approximately 150 µm. One hematoxylin and eosin (H&E)–stained section is prepared at each level. If the nodes still appear negative after histologic examination, an adjacent section from each level is examined by immunocytochemistry using the multicytokeratin antibody AE1/AE3.

Step-sectioning sentinel lymph nodes

The current histopathologic routine is to step-section the sentinel lymph nodes (SLNs) at multiple levels and to perform immunohistochemical staining as appropriate to identify micrometastatic disease. If any question remains about abnormal cells after the first sections are taken, additional sections are obtained. Immunohistochemistry results identify an additional 10-20% of patients with positive SLNs, in whom micrometastases are not seen on routine sections stained with permanent H&E. At least some of the increased rate of detection of micrometastatic disease is attributable to step-sectioning at multiple levels.

The approach for sampling involves bivalving the lymph node, fixing the two halves face down, and subsequently sectioning each half into 10 sections, which are alternately used for H&E staining, immunohistochemistry testing, and molecular staging. The sensitivity of intraoperative frozen section examination of the SLN is disappointingly low (< 50%), although false-positive results are almost never reported. Because of concerns about tissue loss during the frozen section procedure, most centers eschew frozen sections and rely on permanent sections, except to confirm grossly suggestive metastatic disease.

Multiple histopathologic staging systems have been proposed.

In one, the interpretation of the histopathology and immunocytochemistry results for the SLN is categorized as stages 1 through 5, as follows:

• Stage 1 tumor: In stage 1, the sample is positive for tumor upon first examination using H&E stain.
• Stage 2 tumor: In stage 2, the sample initially appears negative for tumor, but it is noted to be positive for tumor upon examination of the H&E stain of step-serial sections.
• Stage 3 tumor: In stage 3, the sample is negative for tumor at stages 1 and 2 but positive for tumor based on immunohistochemistry results. To be categorized as positive for tumor, cells must be present that are positive based on immunocytochemistry results and are cytologically observed to be nucleated cells with the characteristics of viable epithelial cells in both the immunocytochemical preparation and the serial H&E-stained sections. Cytokeratin positivity lacking the cytological features of viable tumor cells is categorized as stage 4.
• Stage 4 tumor: In stage 4, cytokeratin positivity does not show the features of viable tumor cells. This positivity likely represents either dying tumor cells (possibly apoptotic cells), characterized by eosinophilic bodies lacking normal nuclei, or macrophages with phagocytosed tumor products. Usually, these cells are single and not in small, cohesive groups. The decision to allocate nodes to this category requires careful comparison of the serial H&E-stained section and the immunocytochemical preparation.
• Stage 5 tumor: The sample is negative for tumor at all stages.

Another histopathologic staging system divides findings into three categories,[21] which have shown progressively worsening disease-free and overall survival,[16] as follows:

• Isolated tumor cells: Isolated tumor cells or small clusters
• Micrometastasis: Lymph node infiltration by tumor less than 2 mm in diameter
• Macrometastasis: Lymph node infiltration by tumor greater than 2 mm in diameter

If any lymph node contains a viable tumor (ie, stages 1-3 according to the pathology code description) either based on routine histology studies or based on immunohistochemistry and multiple sectioning, the patient undergoes a completion dissection of the neck. Perform the dissection of the neck within 4 weeks of the SLN biopsy, and begin any adjuvant radiotherapy within 6 weeks of the dissection. Do not administer radiotherapy prior to a neck dissection.

For tumors in stage 4 or 5 according to the pathology code description, no further treatment to the neck is required and no prophylactic neck radiotherapy or additional surgery is necessary.

The current treatment for patients with squamous cell carcinoma (SCC) includes a proper diagnosis based on a high level of consideration and examination, with subsequent staging and development of a treatment paradigm. A treatment program of surgery, radiation, or chemotherapy is best developed by an oncology team that includes surgeons, radiation and medical oncologists, and rehabilitative specialists who all have significant experience in the care of patients with cancer involving the head and the neck. By using this approach, new protocols and surgical options can be appropriately offered to those patients with advanced cancers.

Patients are seen every 3 months for the first year, every 4 months for the following 2 years, and every 6 months until 5 years following SLN biopsy (SNLB). At any stage, if nodal disease is detected, patients are able to undergo further surgery.

SNLB for head and neck squamous cell carcinomas (HNSCCs) is assuming a growing role in the management of early-stage patients. Particularly, those with T1-2 and even early T3 tumors may benefit from the procedure since the rate of occult lymph node metastases in this group is much lower than in those with more advanced tumors. The accuracy of identification and false-negative rates of the procedure are good, but if there is concern by the surgeon performing the procedure that the node is difficult to locate (ie, lymphoscintigraphy findings are unclear or it is difficult to identify in the operating room), a formal neck dissection should be performed. SLNB will probably play an increasing role in the management of early-stage aerodigestive SCC and cutaneous HNSCC in the future as it gains general acceptance.