P3-07-23: Intraoperative Molecular Analysis of Sentinel Lymph Nodes in Breast Cancer Using One Step Nucleic Acid Amplification (OSNA).
Authors: Chaudhry A , Massey E , Jenkins M , Calder C , Winters ZE , Rayter Z 
Published: June 2012

Abstract
Introduction: The OSNA method for the intra-operative analysis of sentinel lymph nodes (SLNs) in breast cancer has been introduced in 3 UK centres since 2007. The methodology uses a polymerase chain reaction to quantitate CK19, a cytokeratin specific to breast duct epithelial cells. OSNA provides “real-time” results on SLNs analysed as negative (−) or positive with either micro (+) or macrometastases (++).
Method: This is a single-centre prospective pilot study of all patients undergoing breast cancer surgery including sentinel node biopsy from February 2010 to May 2011. SLN identification was performed using a dual localization technique with peri-areolar Patente V blue dye and Tc99 radio-active isotope. SLNs were cut into 4 slices labeled as A, B, C and D, respectively after the removal of all perinodal fat. In all SLNs, slices A and C were processed in OSNA and slices B and D underwent histological assessment by H&E staining. Slices A and C were prepared for OSNA analysis with the required reagent (Lynorhag), followed by centrifugation and homogenization. Micro-pipetted samples were processed against control specimens, to produce normalized and quantitative curve results. The primary outcome measure was to correlate the results of OSNA with histopathology for each SLN. A negative SLN resulted in no further axillary surgery compared to a level I axillary sampling following the presence of micrometastases and a level II dissection following the detection of macrometastases. The total duration of OSNA for all SLNs was recorded in relation to delays in the completion of the intended breast surgery in each patient.
Results: 251 SLNs were analysed in 112 patients (mean age of 55 years). Comparisons between OSNA and histopathology were made in 116 nodes (34 SLNs analysed by OSNA only). SLN positivity was evident in 30 nodes (26%) comprising either macrometastases (n=13) or micrometastases (n=17). The OSNA sensitivity and specificity of was 93% and 89%, respectively. Accounting for a tissue allocation bias in the presence of micrometastases only, the specificity rose to 94% if these cases were excluded in the analysis. There was no correlation between SLN positivity and tumour grade, size or receptor status. The time to OSNA results were analysed in 75 patients undergoing 45 wide local excisions (WLE), of which 18 were wire localized; 23 mastectomies and 22 SLNB alone. The mean time for OSNA was 40.5, 51.8, 54 and 61.5 minutes for 1,2,3, and 4 sentinel lymph nodes respectively. Operation time was prolonged by a median of 20 minutes (range −48 to +65 minutes) WLEs were delayed by the greatest time Exclusions: Nodes that were not available for histological comparison i) nodes weighing &lt;0.05g (n=34) were processed whole. ii) Departmental agreement from mid March 2011 to process nodes whole via OSNA (n=51); 6 had micro or macrometastases.
Conclusion: OSNA prevented staged axillary surgery in 24 (21%) of patients. A median time of 20 minutes for the OSNA procedure is comparable with acceptable operating times. Current experience supports the use of OSNA for each individual whole SLN analysis. The pilot data has resulted in a prescribed change in policy to analyse the whole SLN using this technique.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-07-23.