Abstract P3-07-08: Quantitative HER family proteins assessment as prognostic and predictive biomarkers in the EGF30008 clinical trial
Authors: Nuciforo P , Thyparambil S , Galván P , Vilaro M , Jimenez J , Liao W-L , Cecchi F , Blackler A , Press MF , Gagnon R , Ellis C , Hembrough T , Johnston S , Prat A 
Published: May 2016

Abstract
Background
Combined targeted strategy with letrozole (Le) and lapatinib (La) improves progression-free survival (PFS) in patients with metastatic breast cancer (MBC) co-expressing hormone receptor-positive (HR+) and HER2+ but not in HR+/HER2-negative (HER-) disease (Johnston J Clin Oncol 2009). However, among HER2+ tumors, quantitative levels of HER2 are heterogeneous with a broad dynamic range corresponding to approximately 163.7 to 17446.7 amol/µg as previously reported (Nuciforo SABCS 2014). In addition, within HER2- tumors, quantitative measurement of HER family proteins may identify those patients most likely to benefit from the addition of La to Le. In this retrospective study, we tested the prognostic and predictive ability of HER proteins quantification in clinically HER2+ tumor samples from the EGF30008 study.
Methods
Formalin-fixed paraffin-embedded primary tumor tissues sections from HER2+ MBC population were used. After laser microdissection, tissue lysates were prepared for selected reaction monitoring mass spectrometry (SRM-MS) analysis. Absolute quantitation was accomplished through simultaneous detection of endogenous target and synthetic labeled heavy peptide identical to analytical targets (EGFR, HER2, HER3). HER2 protein levels were correlated with PAM50 molecular subtypes, ERBB2 and ESR1 genes by nCounter. PFS and overall survival (OS) were analyzed by Kaplan–Meier and log-rank test. To determine whether HER2 protein levels were predictive of La benefit, we tested the interaction term of HER2 protein as a continuous variable by treatment arm in a Cox model.
Results
Within the HER2+ study cohort (n=219), 107 had an available tumor block; 84 cases had sufficient material for HER expression measurement by SRM-MS. Average HER2 levels were
2321.1 amol/ug (median, 817.6). HER2 levels were lower in Le+La (n=43; mean, 1761 amol/ug) compared to Le (n=41; mean, 2908 amol/ug) arms, although the difference was non-significant (p=0.108). No expression of EGFR and HER3 was observed. HER2 protein levels were significantly different among PAM50 subtypes with HER2-enriched (HER2E) tumors showing the highest expression followed by Basal-like, Luminal A, Luminal B, and Normal-like (p&lt;0.001). A correlation between HER2 protein, ERBB2 (r=0.5, p&lt;0.001) and ESR1 (r=-0.5, p=0.001) gene expression was found. In patients with disease that expresses HER2 protein levels above the median a trend towards worse PFS (2.9 vs 7.7 months, p=0.092) and OS (21 vs 39 months, p=0.071) were observed. A statistically significant interaction was observed between HER2 protein levels and La treatment for both PFS (p=0.049) and OS (p&lt;0.001). HER2+ tumors with lower expression of HER2 benefited more from La than those with higher expression.
Conclusions
Levels of HER2 protein in HER2+ MBC are extremely heterogeneous. An association between HER2 protein and gene expression by nCounter was observed. HER2E tumors by PAM50 showed the highest levels of HER2 protein. Within the group of HER2+ MBC by standard IHC/FISH, tumors with high HER2 protein had a statistically non-significant worse outcome and do not seem to benefit from La. Further validation of these findings is warranted.
Citation Format: Nuciforo P, Thyparambil S, Galván P, Vilaro M, Jimenez J, Liao W-L, Cecchi F, Blackler A, Press MF, Gagnon R, Ellis C, Hembrough T, Johnston S, Prat A. Quantitative HER family proteins assessment as prognostic and predictive biomarkers in the EGF30008 clinical trial. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P3-07-08.