Over 99% of toxicity was grade 1 or 2. Median LVEF was unchanged pre- to post-treatment, with three patients (15%) experiencing asymptomatic decreases in LVEF; no patient experienced symptoms of left ventricular dysfunction. Experience in our Phase I trial evaluating the combination of CONFIDENTIAL Page 14 of 102 Protocol: HERCEPTIN® + E75/GM-CSF Version 3.1 ML25749 05 March 2017 trastuzumab and a cytotoxic T cell-eliciting HER2 peptide vaccine in the adjuvant setting has not shown any added cardiac toxicity with combination therapy during vaccine dose escalation. Finally, in preliminary Phase II trial results of the combination of Herceptin and either HER2 CD4+ helper or CD8+ T cell-eliciting vaccines conducted by our group, no additive cardiac toxicity has been found (Mittendorf et al. 2010). These early clinical trials have shown promise of efficacy in combination therapy as well. Disis and colleagues demonstrated prolonged increases in peptide-specific immunity after vaccination, as well as epitope-spreading within the HER2 protein and to other non-HER2 tumor antigens (Disis et al. 2004). Finally, we have investigated sequential therapy with Herceptin followed by HER2 vaccination in the adjuvant setting in our ongoing Phase II trials. Of 62 patients who received standard of care Herceptin, the 32 who received no vaccine have experienced a 12.5% breast cancer recurrence rate (4/32) – comparable with reported rates of similarly staged and treated patients. In contrast, of the 30 patients who were vaccinated with either E75+GM-CSF (n=12) or GP2+GM-CSF (n=18) after completing Herceptin therapy, the recurrence rate is 0% (0/30) (p=0.065) (Sears et al. 2011). The emerging evidence suggesting that patients with low-expressing HER2 breast cancer respond to Herceptin or HER2 vaccination combined with the encouraging results suggesting synergism in combination therapy are intriguing and are worthy of further investigation. Therefore, we believe that HER2 1+ and 2+ breast cancer patients, who have already shown a response to vaccination and possibly to Herceptin alone, could potentially show a dramatic response to the combination of Herceptin and E75 with the immunoadjuvant GM-CSF and that the combination can be delivered safely. 2. OBJECTIVES In this study, we intend to assess the ability of the combination of Herceptin and the HER2 vaccine E75 (administered with the immunoadjuvant GM-CSF) given in the adjuvant setting to prevent recurrences in NP (or NN if negative for both estrogen (ER) and progesterone (PR) receptors) breast cancer patients with tumors that express low (1+) or intermediate (2+) levels of HER2. Enrolled patients will be randomized to receive Herceptin and E75 + GM-CSF or Herceptin with GM-CSF alone (no E75). The safety of the combination therapy will be documented, specifically to ensure that no additive cardiac toxicity results from combination HER2-directed therapy. Efficacy will be documented by comparing the DFS and immunological responses between treatment groups. 2.1 PRIMARY OBJECTIVES The primary efficacy endpoint is to compare DFS at 24 months between treatment groups. The primary safety issue is to prove there is no additive cardiac toxicity with combination HER2-directed therapy. 2.2 SECONDARY OBJECTIVES A secondary endpoint of the trial is to compare DFS at 36 months. Immunologic responses to the vaccine will also be documented and correlated to clinical benefit. CONFIDENTIAL Page 15 of 102 Protocol: HERCEPTIN® + E75/GM-CSF Version 3.1 ML25749 05 March 2017 3. STUDY DESIGN 3.1 DESCRIPTION OF THE STUDY The study will be a multi-center, prospective, randomized, single-blinded, placebo-controlled Phase II trial of Herceptin + E75/GM-CSF versus Herceptin + GM-CSF alone. The vaccine to be used in this study, E75, is investigational and will be used in combination with Herceptin under the Investigational New Drug (IND) Application, BB-IND #14919. The Sponsor/Investigator of this IND is Dr.