Axillary Lymph Node Nanometastases Are Prognostic Factors for Disease-Free Survival andMetastatic Relapse in Breast Cancer Patients

Purpose: Early breast cancer presents with a remarkable heterogeneity of outcomes. Undetected, microscopic lymphnode tumor deposits may account for a significant fractionof this prognostic diversity. Thus, we systematically evaluated the presence of lymph node tumor cell deposits V0.2 mm in diameter [pN0(i+), nanometastases] and analyzed their prognostic effect. Experimental Design: Single-institution, consecutive patients with 8 years of median follow-up (n = 702) were studied.Tomaximize chances of detectingmicrometastases andnanometastases, whole-axilla dissections were analyzed. pN0 cases (n = 377) were systematically reevaluatedby lymphnode (n =6676) step-sectioning and anticytokeratin immunohistochemical analysis. The risk of first adverse events and of distant relapse of bona fide pN0 patients was compared with that of pN0(i+), pN1mi, and pN1cases. Results:Minimal lymphnode deposits were revealed in13% of pN0 patients.The hazard ratio for all adverse events of pN0(i+) versus pN0(i ) was 2.51 (P =0.00019). Hazards of pN1mi andpN0(i+) cases were not significantly different. A multivariate Cox model showed a hazard ratio of 2.16 for grouped pN0(i+)/pN1mi versus pN0(i ) (P = 0.0005). Crude cumulative incidence curves for metastatic relapse were also significantly different (Gray’s test m = 5.54, P = 0.019). Conclusion:Nanometastases are a strong risk factor for disease-free survival and for metastatic relapse. These findings support the inclusion of procedures for nanometastasis detection in tumor-node-metastasis staging. The tumor-node-metastasis (TNM) staging system for breast cancer (1) has proven invaluable in categorizing the extent of neoplastic disease, and as a basis to estimate prognosis and to direct treatment (2). However, this has not lead to the definition of tightly homogeneous prognostic classes, as considerable heterogeneity of outcomes can be observed among disease cases currently categorized as similar. This is particularly evident in the case of small breast tumors (2). We argued that a diverse extent of lymph node dissemination at early stages of disease may account for diverse disease recurrence dynamics. The principle that the macroscopic burden of metastatic cells (e.g., number of invaded lymph nodes) dictates different risks of disease recurrence has been recognized (1, 3). This principle might be equally important at the low end of the spectrum, i.e., in the case of microscopic tumor cell deposits (1, 4). Serial sectioning coupled to immunohistochemical analysis has considerably improved the detection of small tumor cell clusters in lymph nodes (5–10). Occult metastases can indeed be identified in up to 30% of cases previously classified as pN0 (7–9), in 14% to 20% of the cases by single lymph node sections (9, 10). Studies based on these procedures have shown that axillary lymph node microinvasion is a prognostic factor for breast cancer patients, and is associated with poorer diseasefree and overall survival (7, 8, 11–13). As a consequence, micrometastases (0.21-2 mm in diameter) have been identified as a relevant risk factor and their detection has been introduced in TNM staging procedures (2003 TNM edition; ref. 1). Following these guidelines, isolated tumor cells or cell deposits smaller than 0.2 mm are currently required to be classified as pN0(i+) (1). This is because the associated risk is believed to be small or altogether nil. However, little clinical data are available to support this point of view, and the question of the lower size limit of lymph node metastases that may bear clinical significance remains open. Imaging, Diagnosis, Prognosis Authors’Affiliations: Department of Experimental and Diagnostic Medicine, Section of Anatomic Pathology, University of Ferrara, Ferrara, Italy ; Unit of Medical Statistics and Biometry, National Cancer Institute of Milano; Institute of Medical Statistics and Biometry, University of Milano, Milan, Italy ; and Department of Oncology and Neurosciences and Unit of Cancer Pathology, CeSI, ‘‘G. d’Annunzio’’Chieti University Foundation, Chieti, Italy Received 3/9/06; revised 7/21/06; accepted 8/28/06. Grant support: Fondazione of the Cassa di Risparmio della Provincia di Chieti, Italian Association for Cancer Research, MIUR-FIRB Postgenomica grant RBNE0157EH, and EUNoEBiopattern (FP6-2002-IST-1no. 508803). The costs of publication of this article were defrayed in part by the payment of page charges.This article must therefore be hereby marked advertisement in accordance with18 U.S.C. Section1734 solely to indicate this fact. Note: Supplementary data for this article are available at Clinical Cancer Research Online (http://clincancerres.aacrjournals.org/). Requests for reprints: Saverio Alberti, Unit of Cancer Pathology, Center of Excellence in Research on Aging, University ‘‘G. d’Annunzio,’’ Via Colle dell’Ara, 66013 Chieti Scalo (Chieti), Italy. Phone: 39-0871-541-551; Fax: 39-0871-541551; E-mail: s.alberti@unich.it. F2006 American Association for Cancer Research. doi:10.1158/1078-0432.CCR-06-0569 www.aacrjournals. org Clin Cancer Res 2006;12(22) November15, 2006 6696 Research. on May 2, 2017. © 2006 American Association for Cancer clincancerres.aacrjournals. org Downloaded from To tackle this issue, we systematically investigated the presence and prognostic effect of tumor cell deposits V0.2 mm in diameter in the axillary lymph nodes of breast cancer patients conventionally classified as pN0. As these tumor deposits are up to six orders of magnitude smaller in volume than micrometastases, we propose to indicate them as ‘‘nanometastases. ’’