2006). E75, GP2, and AE37 peptides have all been utilized as the basis of separate anti-cancer vaccines combined with the immunoadjuvant, granulocyte macrophage colony-stimulating factor (GM-CSF), and tested by our group in Phase I/II trials. E75 has been shown to reduce clinical recurrences in the adjuvant setting similar in magnitude to the benefit proven with Herceptin (Peoples et al. 2008) While Herceptin has previously been reserved for HER2 3+ expressing tumors, emerging evidence suggests a potential benefit may exist for HER2 2+ expressing tumors as well CONFIDENTIAL Page 6 of 102 Protocol: HERCEPTIN® + E75/GM-CSF Version 3.1 ML25749 05 March 2017 (Paik et al. 2008). In our vaccine trials, we have shown that HER2-derived, CD8-eliciting, peptide-based vaccines used in the adjuvant setting may decrease the recurrence rate in breast cancer patients with tumors expressing any level of HER2, especially HER2 1+ and 2+ tumors (Benavides et al. 2009). Finally, in ongoing Phase II trials of our HER2 vaccines, 30 patients with HER2 3+ tumors treated with standard of care Herceptin followed by HER2 vaccination, have had no breast cancer recurrences in 48 months of follow up (Sears et al. 2011). Given this potential of patients with HER2 1+ and 2+ expressing tumors to respond to both Herceptin and HER2-derived peptide vaccines, we believe that the combination of Herceptin and a HER2-derived, CD8-eliciting peptide vaccine could potentially show a dramatic improvement in preventing breast cancer recurrence. 1.2 HER2 AND BREAST CANCER Growth factors and their receptors play critical roles in development, cell growth, differentiation, and apoptosis (Cross et al. 1991). Such receptors span the cell membrane, with the extracellular domain binding specific growth factors and the intracellular domain transmitting growth signals. Interaction of the extracellular domain with its cognate ligand often results in intracellular activation of tyrosine kinase activity. Overexpression of human epidermal growth factor receptor 2 (HER2, also known as erbB2, neu, and p185HER2) is observed in approximately 25-30% of human breast cancers (Slamon et al. 1987). HER2 overexpression has been reported to only rarely occur in the absence of gene amplification (Kallioniemi et al. 1992; Pauletti et al.