26P - Gene Expression of ESR1, PGR and HER2 in Lymph Node Homogenates and Concordance with IHC in Primary Breast Tumours

Abstract

Aim: To determine whether extracts from the molecular assessment of sentinel lymph node (LN) status may be used (i) to assess ER, PR or HER2 status and (ii) as a resource for assessing other molecular markers in axillary LNs. Background: One-step nucleic acid amplification (OSNA) is used for rapid detection of sentinel LN metastases. Based on the RNA level of cytokeratin 19 (KRT19) an epithelial marker, in LN extracts, the LN is classified as negative, ‘+’ positive or ‘+ +’ positive. Measurement of molecular markers in these extracts could be clinically useful when insufficient invasive tissue is present in the primary to characterize these markers and may have research applications. Patients and methods: 800 consecutive LN homogenates remaining after OSNA tests yielded 39 OSNA ++ cases. Additional HER2 + (IHC 3+) and ER -ve cases were identified from 400 further LN homogenates to give a final set of 48 OSNA ‘+ +’ cases (7 HER2 +, 7 ER -ve and 9 PR -ve). 23 OSNA -ve cases were used as controls. RNA was extracted from the homogenates and gene expression of KRT19, ESR1, HER2 and PGR was assessed by QPCR. IHC data was available from routine analysis of the primary tumours. Results: As expected, KRT19 expression levels in OSNA ++ samples were much higher (mean 1840-fold) than in OSNA -ve samples. Expression levels of ESR1, HER2 and PGR were 30-40-fold higher in OSNA ++ samples compared to OSNA -ve samples. 6 of the 7 IHC HER2 + cases had the highest HER2 gene expression levels. When HER2 expression levels were normalized to KRT19 level (to account for the amount of tumour present in each LN homogenate) there was complete concordance between LN RNA and primary tumour IHC. The 7 ER -ve cases were among the 9 lowest ESR1 gene expression levels; normalization to KRT19 did not improve the association. The 9 PR -ve cases were among the 13 lowest PGR gene expression levels; normalization to KRT19 lost any concordance for PR. Conclusion: These novel data indicate that LN extracts remaining after OSNA may have clinical utility for the measurement of HER2 and possibly ER and PR status. The data also suggest that RNA extracts from LNs are a resource for understanding the molecular characteristics of those cells with the greatest potential for metastatic spread. Disclosure: All authors have declared no conflicts of interest.