Ultrastaging and Low-Volume Metastatic Disease in Early-Stage Cervical Cancer: State of the Art

Abstract

The sentinel lymph node technique in early-stage cervical cancer—when to perform it, its process, as well as the surgical specimen—continues to be a challenge for gynecologists, oncologists, and pathologists in order to plan the therapeutic strategy. The objective of this paper is to describe the state of the art and provide a critical point of view about these topics.

1. Introduction

The sentinel lymph node technique in early-stage cervical cancer is a safe method that has practically become standard in most of the centers in the United States and Europe.

According to the latest 2023 NCCN guideline update, the sentinel lymph node technique may be used in tumors of up to 4 cm [1]. However, the best detection rates and mapping results are obtained when tumors are smaller than 2 cm [2]. This technique includes direct cervical injection of patent blue dye, technetium-99 colloid (99Tc), or indocyanine green. The injection may be applied at two sites (3 o’clock and 9 o’clock) at four cardinal points (12 o’clock, 3 o’clock, 6 o’clock, and 9 o’clock) or in four quadrants. After the injection, the sentinel node is usually medial to the external iliac vessels, ventral to the hypogastric vessels, or in the upper region of the obturator fossa.

The advantages of the sentinel lymph node technique are [3]:

• Determining the histopathological status of the first lymph drainage site from the primary tumor.
• Identifying the lymph node in aberrant sites (which might not have been included in the systemic lymphadenectomy).
• Lowering morbidity.
• Enabling the routine use of ultrastaging.

Intraoperative assessment of the sentinel nodes is a reliable procedure; however, micrometastases and isolated tumor cells (ITCs) may go undetected. As mentioned above, detection is no longer an issue, but the point is when is the best time to do so. As for detection, some groups in Argentina are studying the sentinel lymph node technique. To date, Anchezar et al. reported the largest case series in 2023 [4].

At present, the 8th edition of the 2017 AJCC, which describes three types of node involvement related to tumor cells, is applicable:

• Macromeastases measuring > 2 mm
• Micrometastases measuring > 0.2 mm but not greater than 2 mm and/or including >200 cells.
• Isolated tumor cells (ITCs) measuring ≤ 0.2 mm or including ≤ 200 cells are generally detected on immunohistochemistry (IHC).

Intraoperative assessment should include both macroscopically healthy sentinel lymph nodes and macroscopically suspicious nodes. Upon confirmation of node involvement, radical surgery will be canceled or the treatment approach will be reevaluated at some centers. The sentinel lymph node should be sent to the pathologist in a container without fixative for intraoperative assessment.

The pathologist will then perform a macroscopic dissection of the perinodal fat tissue and identify and select the lymph node. From a surgical standpoint, good practice states that some perinodal tissue should be spared for the precise diagnosis of extranodal spread.

Only one section is enough when one lymph node presents macroscopic involvement. The study may be combined with cytologic evaluation.

However, non-suspicious lymph nodes should either be divided in two (if small) or sectioned at (approximately) every 2 mm of width. Each tissue section should be cut and stained by H&E. After freezing, the section should be fixed (preferably in 4% buffered formalin), processed, and embedded in paraffin.

If no metastasis is found in the first section, the sentinel node should then be ultrastaged. [5]. It is important to consider that the intraoperative frozen biopsy has an estimated 20% sensitivity and a negative predictive value of about 93%. For this reason, these new techniques with immunohistochemistry (IHC) (together with the development of biomolecular markers) should be performed for an accurate study [6].

Ultrastaging includes a thorough assessment of a lymph node in order to detect metastases not usually diagnosed with a standard technique.

The study includes the following [5]:

• Serial sectioning of the node at different levels (at least five serial sections at 200 μm).
• Staining with H&E (hematoxylin and eosin).
• Use of IHC staining techniques: at least a wide-spectrum cytokeratin antibody (e.g., AE1/AE3).

Ultrastaging has the advantage of detecting low-volume metastases (micrometastases and isolated tumor cells, ITCs).

In 2022, The European Society of Gynecological Oncology (ESGO) published the SENTIX trial: an international prospective cohort study of the sentinel lymph node biopsy in cervical cancer.

Six hundred forty-seven (647) patients with tumors <4 cm (or <2 cm with sparing fertility) with no suspicion of lymph node disease on images, and only those with bilateral lymph node detection were included.

Conventional studies of the sentinel lymph node (staining with H&E) and ultrastaging were performed in all cases. Eighty-two (82) patients had a positive node: 46 (56%) were diagnosed with the standard study, whereas 36 more cases (44%) were detected with ultrastaging [7].

2. What Is the Impact of Low-Volume Metastatic Disease?

The medical literature today reports favorable rates of false negatives, as confirmed by the SENTICOL-1 trial [8,9].

In this study, using the double staining technique (Tc99-labeled radiotracer and patent blue), the sentinel node was detected in 136 of 139 patients (97.8%).

Two (2) of the 23 patients with negative sentinel lymph nodes were positive in one different node in the lymphadenectomy (true false negatives). For this reason, this study reported 92% sensitivity. However, in cases of bilateral sentinel lymph node detection (104 out of 134 patients), the rate of false negatives was 0% (100% sensitivity).

In 2011, Cibula et al. published one of the most important papers on the prognostic impact of low-volume lymph node disease. The study included 645 retrospective patients from eight centers.

Macrometastases and micrometastases were identified in 47 and 46 patients, respectively. After a mean follow-up of 40 months, the presence of macrometastases and micrometastases was associated with a significant decrease in overall survival [10].

In 2018, FIGO (International Federation of Gynecology and Obstetrics) introduced a modification in the staging of cervical cancer. The Stage IIIc was added for cases of positive adenopathy.

In this sense, a difference was included based on the location of metastases. Cases of positive nodes in the pelvis should be reported as Stage IIIc1, and cases of lumbar aortic adenopathy should be reported as Stage IIIc2.

Importantly, updated states in which cases of node metastases are confirmed by pathology should be named with the letter “p” as a prefix (for example, pIIIc1 or pIIIc2). However, when the diagnosis is confirmed by images, the letter “r” should be added (for example: rIIIc1 or rIIIc2) [11].

3. What About Micrometastases and Isolated Tumor Cells (ITCs)?

The 2018 FIGO report on cervical cancer does not mention these entities. Therefore, in 2019, FIGO published a correction [12] of their own report stating that the presence of micrometastases in the lymphadenectomy specimen (or sentinel node) should be staged as Stage IIIc.

The publication stated that the presence of isolated tumor cells (ITCs) should be reported; however, staging is not modified since no evidence is available today in terms of decreased survival or disease-free survival.

It is important to remember that when performing the sentinel lymph node technique, the node should be processed with ultrastaging for adequate treatment planning, regardless of whether either surgical or non-surgical management is selected.

4. Something to Think About and Take into Consideration

For many years now, the detection of the sentinel lymph node has not been an issue; however, the important point is determining how to study the node and what the impact is in both decision-making and treatment planning.

In 2007, Schneider stated that detection is better in tumors smaller than 2 cm. Moreover, Schneider suggested studying the node with IHCs and developing molecular markers related to the finding of viral HPV ARN [2].

In 2010, Bats et al. published similar concepts showing that the sensitivity of intraoperative frozen biopsy was only 20% [13].

That same year, Díaz et al. performed ultrastaging of the sentinel node and radical trachelectomy specimens and published a metastasis and micrometastases rate of 8%, which was not diagnosed by classical H&E [14].

At present, we know that persistent infection with high-oncogenic-risk serotypes of HPV is necessary for the occurrence of cervical cancer. HPV carcinogenesis is based on the capacity of the E6 and E7 proteins to interfere with cell growth control.

In 2015, Basiletti-Soderini et al. [15] performed a prospective multicenter study including 50 patients with a diagnosis of cervical cancer with a squamous etiology and adenocarcinoma, stages Ia2 and IIb, undergoing surgical management.

The hysterectomy specimens and lymph nodes of all the patients were assessed with molecular techniques and PCR in order to determine the expression of E6-E7 in high-grade HPV in the lymph nodes through the detection of mRNA and its predictive value for node metastasis.

In patients whose pelvic nodes presented metastasis, as diagnosed by histology, viral mRNA was detected corresponding to the same viral type found in the primary tumor, as well as in patients with histologically negative nodes.

A high level of agreement was seen between the histological result and the mRNA measurement in E6 and E7. The presence of mRNA indicates viral genome transcription, a process that, in a lymph node, would occur only in cervical carcinoma metastatic cells, for these viruses are active in epithelial cells only.

Therefore, the finding of mRNA in normal pelvic lymph nodes might indicate very early metastasis, detectable at the molecular level [15,16]. It should be underlined that the virus is in a proliferative stage, and for this, the virus requires the presence of a squamous cell.

Therefore, the presence of isolated cells in lymph nodes with the expression of these oncoproteins might be considered an early molecular marker of metastasis (and even a potential treatment target). Considering this evidence, we should think about the following questions:

• Are we not treating a disease that might be then left to its natural course?
• Could there be more lymph nodes involved apart from the sentinel node?
• Should the lymphadenectomy be wider based on this information?

However, the ConCerv [17] trial proposed cervical conization for early stages with tumors smaller than 2 cm as a treatment option, with a 3.5% recurrence rate.

Following a less radical treatment approach, the SHAPE trial [18] proposes extrafascial hysterectomy as a management option, with a recurrence rate of 2.56%; however, the difference is not significant compared to radical hysterectomy.

However, ultrastaging was not used to process the surgical specimen. Therefore, regarding the recurrences mentioned, are they really recurrences or persistent disease?

In conclusion, we are in a position to say that ultrastaging and molecular studies are here to stay and help us with more selective and patient-customized treatment planning.