Background: LIV-1 is an estrogen-inducible gene that has been implicated in epidermal-to-mesenchymal transition (EMT) in preclinical models of progression and metastasis. Its expression is associated with node-positivity in breast cancer; and has been detected in a variety of cancer types, including estrogen receptor positive breast cancers. SGN-LIV1A is a novel antibody drug conjugate targeting LIV-1 that is currently being evaluated in the I-SPY 2 TRIAL. In this pilot study, we evaluated LIV-1 levels by IHC within HR/HER2/MammaPrint (MP) defined subtypes among patients screening for the I-SPY 2 TRIAL and its correlation to microarray assessed LIV-1 expression levels.

Method: In a pilot study, LIV-1IHC staining was performed by Quest Diagnostics on the pre-treatment samples of38 patients screening for the I-SPY 2 TRIAL. Pre-treatment expression data generated on a custom Agilent 44K platform was also available. We summarized the LIV-1 H-Scores and percent (%)-positivity across the population and within HR/HER2/MP subtypes; and we assessed the Pearson correlation between LIV-1 H-Score and LIV-1 gene expression levels. In addition, we compared the pre-treatment LIV-1 expression levels withinHR/HER2/MP subtypes across I-SPY 2 TRIAL patients from completed arms and their relevant controls (n=989) using ANOVA and post-hoc Tukey tests. Our statistics are descriptive rather than inferential; and does not take into account multiplicities of other biomarkers outside of this study.

Results: Of the 38 patients evaluated,37 have LIV-1 %-positivity > 0; and 18 (47%) have 100% LIV1 positivity. The medianLIV-1 H-Score is 200; and 89% of patients (34/38) have moderate/high LIV-1 staining(with H-Score≥100). Of the 34 patients who proceeded onto the trial (and have knownHR/HER2/MP status), 9 are triple negative, 19 are HR+HER2-, and 6 are HER2+. Due to our small sample size, we did not further subset the triple negative and HER2+cases; but within the HR+HER2- patients, 10 are MP1 compared to 9 who are MP2 class.LIV1 H-Score appears highest within the HR+HER2-MP1 cases (median: 290), followed by the HER2+ (median: 216), then the HR+HER2-/MP2 (median: 155), and the TN (median:120) subtype. LIV1 H-score is significantly correlated with LIV-1 mRNA expression levels (Rp=0.79, p<0.0001). Consistent with these observations, LIV-1 pre-treatment expression levels are significantly higher in the HR+HER2-MP1 group relative toall other HR/HER2/MP defined subtypes (Tukey HSD p < 0.0001) across the I-SPY2 TRIAL population. The HR+HER2+MP1 group also have high LIV-1 expression levels.

Conclusion: Our result suggest that although LIV-1 expression differs by subtype, it is expressed at a moderate/high level in the majority of patients. The good correlation between IHC and array-basedLIV-1 expression levels enables us to leverage the entire existing I-SPY 2 data set and confirm the high rates of LIV-1 expression across the I-SPY 2 population. Expression is highest in the subset of patients with the lowest pCR rates in the trial to date.Further studies to evaluate LIV-1 expression as a biomarker of response to LIV-1targeting therapies for the neoadjuvant treatment of breast cancer are warranted and ongoing in I-SPY 2.

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