Sentinel node tumour burden quantified based on cytokeratin 19 mRNA copy number predicts non-sentinel node metastases in breast cancer: Molecular whole-node analysis of all removed nodes
Introduction
Sentinel lymph-node (SN) biopsy is the standard axillary staging procedure for patients with clinically node-negative breast cancer.1 Until recently, guidelines had recommended complete axillary lymph-node dissection (CALND) for patients with any SN metastases.1 Recently, the American College of Surgeons Oncology Group Z-0011 randomised trial, which was designed to compare survival in SN-positive patients who did or did not undergo CALND, found no difference in locoregional recurrence or survival rates between the study arms.2, 3 However, it is controversial whether the results of the Z-0011 trial change daily practice.4 At this time, the results of this trial may only be applicable for clinically node-negative patients who have one or two positive SNs and who are receiving adjuvant systemic chemotherapy and breast-conserving surgery with tangential irradiation.5, 6
To select the SN-positive patients who can safely be excluded from CALND treatment, several mathematical models to estimate the likelihood of non-sentinel node (non-SN) metastases have been produced.7 In almost all models, the SN tumour burden, which may be defined as the number of positive SNs or the size of the SN metastasis, is a predictor of non-SN metastasis.7, 8 However, these models have a critical limitation in terms of the evaluation of lymph-node metastasis. Routine histopathological examinations are non-standardised and limited in their ability to accurately detect metastases, particularly micrometastases, because they only partially evaluate each node. This may lead to the underestimation of the SN and non-SN status and poorly reproducible measurements of the micrometastases. In fact, most of these models were not reliably predictive for the subgroup of patients with micrometastasis in the SN.7 Furthermore, the non-SN tumour burden (macro- or micrometastasis) is not considered in these models.
The one-step nucleic acid amplification (OSNA) assay (Sysmex, Kobe, Japan) was developed to overcome these limitations of the histopathological examination of lymph nodes. This assay is approved and commercialised for clinical use throughout Europe and Japan. This assay can assess the whole lymph node and yields quantitative results in the form of cytokeratin 19 (CK19) mRNA copy number.9 Calibration and validation studies9, 10 have provided reasonable evidence that the CK19 mRNA copy numbers detected by the OSNA assay can be good estimates of macro-, micrometastasis and isolated tumour cells defined by the American Joint Committee on Cancer Staging Manual.11 We have shown that the OSNA whole-node assay detects more cases of SN micrometastases than frozen-section histology using a 2-mm-sectioned node.12, 13 Furthermore, the OSNA assay of all removed non-SNs detected significantly more cases of metastases, particularly micrometastases, which can be missed by single-section histology.14
Therefore, the OSNA whole-node assay of all SNs and non-SNs allows for the accurate measurement of tumour burden in either situation. In the present single-institute retrospective study, we elucidated the relationship between SN and non-SN tumour burdens and determined the predictors of non-SN metastases; this was performed to reveal the usefulness of the OSNA assay regarding the prediction of non-SN metastasis in breast cancer.
Section snippets
Patients and tumours
The study population included patients with clinically and ultrasonographically node-negative breast cancer who had undergone CALND after a metastatic SN biopsy and whose SNs and non-SNs were examined using the OSNA assay between September 2009 and April 2011 at the Cancer Institute Hospital, Tokyo, Japan. The exclusion criteria were as follows: (1) SN mapping without the use of a radioisotope tracer, (2) previous excision of a primary tumour, (3) heterochronous ipsilateral breast cancer
Patient characteristics and non-SN status
Between September 2009 and April 2011, 1251 breast cancer patients underwent SN biopsy with the OSNA assay; 1122 of them did not meet the exclusion criteria. Among these 1122 patients, 211 (18.8%) were diagnosed as SN-positive; all the patients consequently underwent CALND. Among these 211 patients, there were 185 patients whose all non-SNs were assessed using the OSNA assay.
The demographic characteristics are presented in Table 1. All of the patients were Japanese women. Of these 185 patients,
Discussion
In the present study, all removed nodes were evaluated using molecular whole lymph node analysis using the OSNA assay and without using any histopathological examination. With this approach, the node tumour burden could be accurately quantified based on the CK19 mRNA copy number. All patients underwent SN mapping and identification with a radioisotope tracer to rigorously classify the axillary nodes as SN or non-SN.17 Thus, the present study further characterises the relationship between SN and
Conflict of interest statement
None declared.
Acknowledgements
This work was supported in part by a Grant-in-Aid for Young Scientists (B) (No. 21791264) from the Japanese Ministry of Education, Culture, Sports, Science, and Technology and a Research Grant from the Foundation for the Promotion of Cancer Research in Japan. The Sysmex Corporation contributed funding for laboratory consumables used for the OSNA assay. We thank all staff members working in the Breast Oncology Center of the Cancer Institute Hospital, including Masujiro Makita, Seiichiro
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The use of onestep nucleic acid amplification (OSNA) and tumour related factors in the treatment of axillary breast cancer: A predictive model
2016, European Journal of Surgical OncologyCitation Excerpt :In the last 2 years, the relationship between CK19 mRNA copy numbers as a molecular measure of tumour load in the sentinel lymph node and prediction of non-sentinel nodal involvement in the axilla has been under investigation. Several studies have shown that increasing tumour load in the sentinel node appears to predict the likelihood of non-sentinel node involvement based on CK19 copy number threshold values13–21 (Supplementary Table A). Several of the studies agree that mRNA copy number thresholds higher than those attributed to a positive result by the manufacturer need to be considered as a threshold for ANC.18,20,21,23
Keratins in health and disease
2015, Current Opinion in Cell BiologyBreast cancer metastasis burden in sentinel nodes analysed using one-step nucleic acid amplification predicts axillary nodal status
2015, BreastCitation Excerpt :However, this study does not suggest a similar significant effect, and reaffirms that further axillary metastasis risk is more accurately determined, according to the established micrometastasis/macrometastasis dichotomy. In concordance with the literature, patients undergoing mastectomy, patients with larger primary tumours and patients with more positive lymph nodes on SLNB were all found to be at greater risk of having further axillary disease [24–29]. However, these factors are of limited clinical utility for risk stratification.
Tumoral load quantification of positive sentinel lymph nodes in breast cancer to predict more than two involved nodes
2014, BreastCitation Excerpt :In our study, with a higher sample size (n = 726) focused on cases with more than two non-SLN detected after lymphadenectomy, the multivariate analysis did show an association with multifocality, TTL and lymphovascular infiltration. In the study by Ohi et al. [31], 130 patients with positive SLN that underwent complete ALND were investigated, and the CK19 mRNA copy number ≥ 5.0 × 103 in the SLN was the most significant predictor of non-SLN metastases (p = 0.003), while the CK19 mRNA copy number ≥ 1.0 × 105 in the SLN was the only independent predictor of ≥4 metastatic non-SLN nodes (p = 0.014) [32]. Recently, a multivariate model to predict non-SLN MI based on log TTL, tumour size, number of affected SLN, presence of lymphovascular infiltration and Her2 status has been proposed [33,34].
One-step nucleic acid amplification assay for intraoperative prediction of non-sentinel lymph node metastasis in breast cancer patients with sentinel lymph node metastasis
2014, BreastCitation Excerpt :Moreover, our finding that the number of SLNs identified as positive by the OSNA assay was significantly associated with non-SLN metastasis was also consistent with that of previous reports on the findings of post-operative histological examinations [8,9]. We adopted the cut-off value of 5000 copies for the differentiation between macrometastasis and micrometastasis since this cut-off value has been widely used [13–21,27,28]. But a recent study has suggested a possibility that this cut-off value might lead to overdiagnosis of micrometastasis as macrometastasis [32].