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Background Melanoma therapy has changed dramatically over the last decade with improvements in immunotherapy, yet many patients do not respond to current therapies. This novel vaccine strategy may prime a patient’s immune system against their tumor and work synergistically with immunotherapy against advanced-stage melanoma. Methods This was a prospective, randomized, double-blind, placebo-controlled, phase IIb trial of the tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine administered to prevent recurrence in patients with resected stage III/IV melanoma. Patients were enrolled and randomized 2:1 to the TLPLDC vaccine or placebo (empty yeast cell wall particles and autologous dendritic cells). Both intention-to-treat (ITT) and per treatment (PT) analyses were predefined, with PT analysis including patients who remained disease-free through the primary vaccine/placebo series (6 months). Results A total of 144 patients were randomized (103 vaccine, 41 control). Therapy was well-tolerated with similar toxicity between treatment arms; one patient in each group experienced related serious adverse events. While disease-free survival (DFS) was not different between groups in ITT analysis, in PT analysis the vaccine group showed improved 24-month DFS (62.9% vs. 34.8%, p = 0.041). Conclusions This phase IIb trial of TLPLDC vaccine administered to patients with resected stage III/IV melanoma shows TLPLDC is well-tolerated and improves DFS in patients who complete the primary vaccine series. This suggests patients who do not recur early benefit from TLPLDC in preventing future recurrence from melanoma. A phase III trial of TLPLDC + checkpoint inhibitor versus checkpoint inhibitor alone in patients with advanced, surgically resected melanoma is under development. Trial Registration NCT02301611. Supplementary information The online version contains supplementary material available at (10.1245/s10434-021-09709-1).

Background Colorectal cancer (CRC) tumor microenvironment (TME) characteristics, such as tumor infiltrating lymphocyte (TIL) densities and PD-L1 status, are predictive of recurrence, disease-free survival, and overall survival. In many malignancies, TME characteristics are also predictive of response to immunotherapy. As window of opportunity studies using neoadjuvant immunotherapy become more common and treatment guidelines incorporate TME features, accurate assessment of the pre-treatment TME using the biopsy specimen is critical. However, no study has thoroughly evaluated the correlation between the TMEs of the biopsy and resection specimens. Methods We conducted a retrospective analysis of patients with stage I–III CRC with matched biopsy and resection specimens. CD3+, CD4+, CD8+, and FoxP3+ lymphocyte populations at the center of tumor (CT) and invasive margin (IM) and tumor PD-L1 status in the biopsy and resection specimens were evaluated. TIL populations were compared using Mann–Whitney U tests or Student’s t tests and correlated using Pearson r. Results CD3+ and CD4+ densities were significantly higher in the CT of the biopsy relative to the resection specimen Comparing biopsy and resection specimens, no TIL population at either the CT or IM had a correlation coefficient > 0.5. Determining PD-L1 status based on biopsy tissue resulted in a sensitivity of 37.1%, specificity of 81.4%, and accuracy of 61.5%. Conclusions These findings demonstrate significant discordance between the TME of the biopsy and resection specimens. Caution should be used when basing treatment decisions on pre-treatment endoscopic biopsy findings and when interpreting changes in the TME between pre-treatment biopsy and resection specimens after neoadjuvant therapy. Electronic supplementary material The online version of this article (10.1007/s00262-020-02784-5) contains supplementary material, which is available to authorized users.

Purpose: Preclinical data provide evidence for synergism between HER2-targeted peptide vaccines and trastuzumab. The efficacy of this combination was evaluated in patients with HER2 low-expressing breast cancer in the adjuvant setting. Patients and Methods: A phase IIb, multicenter, randomized, single-blinded, controlled trial enrolled disease-free patients after standard therapy completion (NCT01570036). Eligible patients were HLA-A2, A3, A24, and/or A26+, and had HER2 IHC 1+/2+, FISH nonamplified breast cancer, that was node positive and/or hormone receptor–negative [triple-negative breast cancer (TNBC)]. Patients received trastuzumab for 1 year and were randomized to placebo (GM-CSF, control) or nelipepimut-S (NPS) with GM-CSF. Primary outcome was 24-month disease-free survival (DFS). Secondary outcomes were 36-month DFS, safety, and immunologic response. Results: Overall, 275 patients were randomized; 136 received NPS with GM-CSF, and 139 received placebo with GM-CSF. There were no clinicopathologic differences between groups. Concurrent trastuzumab and NPS with GM-CSF was safe with no additional overall or cardiac toxicity compared with control. At median follow-up of 25.7 (interquartile range, 18.4–32.7) months, estimated DFS did not significantly differ between NPS and control [HR, 0.62; 95% confidence interval (CI), 0.31–1.25; P = 0.18]. In a planned exploratory analysis of patients with TNBC, DFS was improved for NPS versus control (HR, 0.26; 95% CI, 0.08–0.81, P = 0.01). Conclusions: The combination of NPS with trastuzumab is safe. In HER2 low-expressing breast cancer, no significant difference in DFS was seen in the intention-to-treat analysis; however, significant clinical benefit was seen in patients with TNBC. These findings warrant further investigation in a phase III randomized trial.

Purpose AE37 and GP2 are HER2 derived peptide vaccines. AE37 primarily elicits a CD4+ response while GP2 elicits a CD8+ response against the HER2 antigen. These peptides were tested in a large randomized trial to assess their ability to prevent recurrence in HER2 expressing breast cancer patients. The primary analyses found no difference in 5-year overall disease-free survival (DFS) but possible benefit in subgroups. Here, we present the final landmark analysis. Methods In this 4-arm, prospective, randomized, single-blinded, multi-center phase II trial, disease-free node positive and high-risk node negative breast cancer patients enrolled after standard of care therapy. Six monthly inoculations of vaccine (VG) vs. control (CG) were given as the primary vaccine series with 4 boosters at 6-month intervals. Demographic, safety, immunologic, and DFS data were evaluated. Results 456 patients were enrolled; 154 patients in the VG and 147 in CG for AE37, 89 patients in the VG and 91 in CG for GP2. The AE37 arm had no difference in DFS as compared to CG, but pre-specified exploratory subgroup analyses showed a trend towards benefit in advanced stage (p = 0.132, HR 0.573 CI 0.275–1.193), HER2 under-expression (p = 0.181, HR 0.756 CI 0.499–1.145), and triple-negative breast cancer (p = 0.266, HR 0.443 CI 0.114–1.717). In patients with both HER2 under-expression and advanced stage, there was significant benefit in the VG (p = 0.039, HR 0.375 CI 0.142–0.988) as compared to CG. The GP2 arm had no significant difference in DFS as compared to CG, but on subgroup analysis, HER2 positive patients had no recurrences with a trend toward improved DFS (p = 0.052) in VG as compared to CG. Conclusions This phase II trial reveals that AE37 and GP2 are safe and possibly associated with improved clinical outcomes of DFS in certain subgroups of breast cancer patients. With these findings, further evaluations are warranted of AE37 and GP2 vaccines given in combination and/or separately for specific subsets of breast cancer patients based on their disease biology.

A randomized, double-blind, placebo-controlled phase 2b trial of the tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine was conducted in patients with resected stage III/IV melanoma. Dendritic cells (DCs) were harvested with and without granulocyte-colony stimulating factor (G-CSF). This analysis investigates differences in clinical outcomes and RNA gene expression between DC harvest methods.The TLPLDC vaccine is created by loading autologous tumor lysate into yeast cell wall particles (YCWPs) and exposing them to phagocytosis by DCs. For DC harvest, patients had a direct blood draw or were pretreated with G-CSF before blood draw. Patients were randomized 2:1 to receive TLPLDC or placebo. Differences in disease-free survival (DFS) and overall survival (OS) were evaluated. RNA-seq analysis was performed on the total RNA of TLPLDC + G and TLPLDC vaccines to compare gene expression between groups.144 patients were randomized: 103 TLPLDC (47 TLPLDC/56 TLPLDC + G) and 41 placebo (19 placebo/22 placebo + G). Median follow-up was 27.0 months. Both 36-month DFS (55.8% vs. 24.4% vs. 30.0%, p = 0.010) and OS (94.2% vs. 69.8% vs. 70.9%, p = 0.024) were improved in TLPLDC compared to TLPLDC + G or placebo, respectively. When compared to TLPLDC + G vaccine, RNA-seq from TLPLDC vaccine showed upregulation of genes associated with DC maturation and downregulation of genes associated with DC suppression or immaturity.Patients receiving TLPLDC vaccine without G-CSF had improved OS and DFS. Outcomes remained similar between patients receiving TLPLDC + G and placebo. Direct DC harvest without G-CSF had higher expression of genes linked to DC maturation, likely improving clinical efficacy.

The coronavirus disease 2019 pandemic has profoundly impacted surgical education. We assessed resident perceptions of our virtual academic program, which consists of daily lectures or case conferences held via a videoconferencing platform.A survey evaluating attitudes and practices for virtual academics was administered to general surgery residents. A focus group was conducted to identify benefits, barriers to engagement, and opportunities for improvement for virtual education. A total of 19 residents completed the education survey, and seven residents participated in the focus group.While expressing preference toward in-person academics (84.2%), residents felt the virtual academics were of good quality (median rating 4/5) and preferred virtual academics to no academic sessions (94.7%). Of respondents, 57.9% believe that the coronavirus pandemic negatively impacted their surgical education. They believe their American Board of Surgery In-Training Examination preparation was not impacted. Residents preferred using a computer over a phone for academics (79% versus 16%). The focus group identified the benefits of virtual academics, including the ability to participate while away and having recordings available. Areas for improvement included reinforcement of protected time for academics, requiring cameras be on, increasing in-lecture polls, and creation of an online repository of recordings for review. Residents hoped a virtual component of academics and recordings would continue past the pandemic.Although virtual academics are not the preferred mode of learning in our residency, there are multiple unintended benefits. We recommend a hybrid academic model with in-person didactics and recorded video for later review.

BackgroundThe tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine is an autologous tumor vaccine that decreased recurrence in stage III/IV melanoma when granulocyte-colony stimulating factor (G-CSF) was not used to harvest the dendritic cells in a randomized phase 2B adjuvant trial.1The tumor lysate (TL) particle only (TLPO) vaccine utilizes a similar mechanism, but with autologous TL-loaded yeast cell wall particles; this eliminates the need for dendritic cell (DC) collection and ex-vivo loading and reduces production costs and time. The TLPO vaccine was compared to TLPLDC in an embedded bridging portion of the trial. Here, we examine 36-month outcomes of the ongoing randomized, double-blind phase 2 trial in patients (pts) with resected stage III/IV melanoma.MethodsPts were randomized 2:1 to receive TLPO or TLPLDC as a continuation of a previously established clinical trial comparing TLPLDC versus placebo. The TLPLDC group was analyzed separately based on use (or not) of G-CSF for collection of DC. Safety was measured by the Common Terminology Criteria for Adverse Events (CTCAE). Kaplan-Meier and log-rank analysis was used to compare 36-month disease-free survival (DFS) and overall survival (OS) in the intention-to-treat (ITT) main arms as well as pre-specified subgroups.ResultsA total of 187 pts were randomized with 41, 47, 56, and 43 pts enrolled in the placebo, TLPLDC without G-CSF (TLPLDC), TLPLDC with G-CSF (TLPLDC+G), and TLPO arm, respectively. Pts randomized to the TLPO arm were more likely to have stage IV melanoma (22.0% for placebo, 20.4% for TLPLDC and TLPLDC+G, and 44.2% for TLPO; p = 0.002) and to receive prior immunotherapy (36.6% for placebo, 39.8% for both TLPLDC and TLPLDC+G, and 83.7% for TLPO; p < 0.001). Grade 3+ adverse events were not significantly different between arms. In the ITT analysis, 36-month DFS was 30.0% for placebo, 55.8% for TLPLDC, 24.4% for TLPLDC+G, and 64.0% for TLPO (p < 0.001). OS at 36 months was 70.9% for placebo, 94.2% for TLPLDC, 69.8%% for TLPLDC+G, and 94.8% for TLPO (p = 0.011) (figure 1).Abstract 542 Figure 1Kaplan-Meier curves demonstrating DFS (A) and OS (B) between Placebo (n=41), TLPLDC (n=47), TLPLDC+G (n=56), and TLPO (n=43)ConclusionsThe TLPO and TLPLDC (without G-CSF) vaccines improved 36-month DFS and OS in this randomized phase 2 trial. The efficacy of the TLPO and TLPLDC vaccines will be confirmed in a phase III trial in resected Stage III/IV melanoma pts.Trial RegistrationNIH, clinicaltrials.gov, NCT02301611ReferencesO’Shea AE, Chick RC, Clifton GT, et al. The effect of pretreatment with G-CSF prior to dendritic cell collection during the phase IIb trial of an autologous DC-based vaccine for advanced, resectable melanoma. Presented at: Society for Immunotherapy of Cancer 35th Anniversary Annual Meeting & Preconference Programs (SITC 2020); November 11–14, 2020. Abstract 310. J Immunother Cancer. 2020;8(Suppl 3):A656–A959.Ethics ApprovalThe clinical trial protocol was approved by the Western Institutional Review Board (2014–1932). All participants provided informed consent prior to enrollment in the trial.

Immunotherapy has revolutionized the treatment of melanoma, yet survival remains poor for patients with metastatic disease. The autologous tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine has been shown to be safe adjuvant therapy for patients with resected stage III/IV melanoma who complete the primary vaccine series. Here, we describe an open-label trial of patients with metastatic melanoma treated with TLPLDC vaccine in addition to standard of care (SoC) therapies. The TLPLDC vaccine is created by loading autologous tumor lysate into yeast cell wall particles, which are phagocytosed by autologous dendritic cells ex vivo. Patients who recurred while enrolled in a phase IIb trial of adjuvant TLPLDC vaccine (crossover cohort) and patients with measurable metastatic melanoma cohort were offered TLPLDC vaccine along with SoC therapies. Tumor response was measured by RECIST 1.1 criteria. Overall survival (OS) and progression-free survival (PFS) were estimated by intention-to-treat analysis. Fifty-four patients were enrolled (28 in crossover cohort; 26 in metastatic melanoma cohort). The vaccine was well-tolerated with no grade ≥3 adverse events when given with SoC therapies to include checkpoint inhibitors, BRAF/MEK inhibitors, tyrosine kinase inhibitors, intralesional therapy and/or radiation. In the crossover arm, OS was 76.5% and PFS was 57.1% (median follow-up of 13.9 months). In the metastatic melanoma arm, OS was 85.7% and PFS was 52.2% (median follow-up 8.5 months). The TLPLDC vaccine is well-tolerated and safe in combination with SoC therapies. Future trials will determine the efficacy of TLPLDC in combination with SoC therapies in metastatic melanoma.

Background E39, an HLA‐A2‐restricted, immunogenic peptide derived from the folate‐binding protein (FBP), is overexpressed in multiple malignancies. We conducted a phase I/IIa trial of the E39 + GM‐CSF vaccine with booster inoculations of either E39 or E39′ (an attenuated version of E39) to prevent recurrences in disease‐free endometrial and ovarian cancer patients(pts). Here, we present the final 24‐month landmark analysis. Patients and methods HLA‐A2 + patients receiving E39 + GM‐CSF were included in the vaccine group (VG), and HLA‐A2‐ pts (or HLA‐A2 + patients refusing vaccine) were followed as the control group (CG). VG group received 6 monthly inoculations as the primary vaccine series (PVS) and were randomized to receive either E39 or E39′ booster inoculations. Demographic, safety, immunologic, and disease‐free survival (DFS) data were collected and evaluated. Results Fifty‐one patients were enrolled; 29 in the VG and 22 in the CG. Fourteen patients received

Background:HER2 low-expressing (LE) (IHC 1-2+, FISH non-amplified) breast cancer (BC) patients (pts) have not benefited from HER2-directed therapy despite HER2 antigen availability. Triple negative BC (TNBC), in particular, is immunogenic and in need of additional therapeutic options. We have previously shown the HER2-derived nelipeptimut-S (E75) + GM-CSF (NeuVax) to be synergistic with trastuzumab (Tz) in pre-clinical and pilot clinical studies. In a planned interim analysis of a multi-center, prospective, randomized, single-blinded, placebo-controlled phase 2b trial of Tz + NeuVax vs Tz to reduce recurrence in HER2 LE, node-positive (NP) and/or triple negative BC (TNBC) pts, we previously reported that the NeuVax + Tz was safe without added cardiac toxicity and demonstrated a significant reduction of recurrences in TNBC pts. This analysis examines additional subsets in this trial. Methods:HER2 LE, NP and/or TNBC pts who were clinically disease-free after standard therapy were randomized to receive Tz+NeuVax (vaccine group; VG) or Tz+GM-CSF (control group; CG). All pts received 1 yr of Tz per label. NeuVax or GM-CSF was given every 3 weeks x 6 starting with the 3rdTz dose, and then boosted every 6 months x 4. This pre-specified interim analysis was triggered 6 months after last enrollment. The primary endpoint is intention-to-treat 24 month disease-free survival (DFS) evaluated by log rank. Results: Of 275 pts randomized in the study (VG n=136, CG n=139), 98 had TNBC (VG=53, CG=45). In the interim analysis, estimated disease-free survival (DFS) was assessed with a median follow up of 18.8 months. No significant clinicopathologic differences were seen between treatment groups. In the TNBC group, estimated DFS was higher overall in VG vs CG (91.9% v 69.9%, p=0.023; hazard ratio [HR] 0.29, 95% confidence interval [CI] 0.09-0.90). On TNBC subgroup analysis, estimated DFS was higher in VG vs CG among pts who received neoadjuvant chemotherapy (VG n=35, CG n=31; HR 0.26, CI 0.07-0.93; p=0.03), HER2 IHC 1+ BC (VG n=34, CG n=28; HR 0.20, CI 0.04-0.96; p=0.03), pts who were AJCC 7thedition stage I/II (VG n=37, CG n=27; HR incalculable, no recurrences in the VG, p=0.008), and pts 351yr of age (VG n=32 & CG n = 26; HR 0.26 CI 0.07,0.94; p=0.009). HRs did not appreciably vary based on the histologic grade or presence of lymphovascular invasion. Conclusion:Examining the subgroups from the pre-specified interim analysis demonstrates a highly significant clinical benefit in TNBC pts overall. Within the TNBC cohort, specific benefit was seen in pts who received chemotherapy neoadjuvantly, expressed lower HER2, were earlier stage, and were older in age. These factors may help enrich the TNBC population targeted in a definitive Phase 3 study in TNBC patients with residual disease after neoadjuvant chemotherapy. Citation Format: Clifton GT, Kemp Bohan PM, Hale DF, Myers JW, Brown TA, Holmes JP, Vreeland TJ, Litton JK, Murthy RK, Mittendorf EA, Peoples GE. Subgroups analysis of a multicenter, prospective, randomized, blinded phase 2b trial of trastuzumab + nelipeptimut-S (NeuVax) vs trastuzumab for prevention of recurrence in breast cancer patients [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P2-09-01.

Background Checkpoint inhibitors (CPI) in combination with cell‐based vaccines may produce synergistic antitumor immunity. The primary analysis of the randomized and blinded phase IIb trial in resected stage III/IV melanoma demonstrated TLPLDC is safe and improved 24‐month disease‐free survival (DFS) in the per treatment (PT) analysis. Here, we examine efficacy within pre‐specified and exploratory subgroups. Methods Stage III/IV patients rendered disease‐free by surgery were randomized 2:1 to TLPLDC vaccine versus placebo. The pre‐specified PT analysis included only patients completing the primary vaccine/placebo series at 6 months. Kaplan–Meier analysis was used to compare 24‐month DFS among subgroups. Results There were no clinicopathologic differences between subgroups except stage IV patients were more likely to receive CPI. In stage IV patients, 24‐month DFS was 43% for vaccine versus 0% for placebo (p = 0.098) in the ITT analysis and 73% versus 0% (p = 0.002) in the PT analysis. There was no significant difference in 24‐month DFS when stratified by use of immunotherapy or CPI. For patients with resected recurrent disease, 24‐month DFS was 88.9% versus 33.3% (p = 0.013) in the PT analysis. All benefit from vaccination was in the PT analysis; no benefit was found in patients receiving up to three doses. Conclusion The TLPLDC vaccine improved DFS in patients completing the primary vaccine series, particularly in the resected stage IV patients. The efficacy of the TLPLDC vaccine will be confirmed in a phase III study evaluating adjuvant TLPLDC + CPI versus Placebo + CPI in resected stage IV melanoma patients., TLPLDC is an autologous tumor lysate vaccine made with autologous dendritic cells to prevent recurrence in patients with high‐risk resected melanoma. In this analysis, the vaccine was found to have the most benefit in stage IV resected patients and may have a synergistic effect in combination with checkpoint inhibitors. ​

Background We have completed a prospective, randomized, multi-center, double-blind, placebo-controlled phase IIb trial of the tumor lysate, particle loaded, dendritic cell (TLPLDC) vaccine given to prevent recurrences in patients with resected stage III/IV melanoma. During the trial, granulocyte colony stimulating factor (G-CSF) was administered to some patients to mobilize dendritic cells (DCs) precursors prior to harvest, allowing for similar DC yield with reduced blood draws. This study examines the impact of DC collection methods on vaccine effectiveness. Methods TLPLDC is produced by loading tumor lysate into pre-prepared yeast cell wall particles (YCWPs) and exposing them to autologous DCs. DC precursors were isolated either by collection of 50–70 mL of blood following pre-administration of 300µg of G-CSF 24–48 hrs prior, or collection of 120 mL of peripheral blood without G-CSF pretreatment based on patient and provider preference. Patients were randomized 2:1 to receive TLPLDC or placebo (DCs exposed to empty YCWPs). 1–1.5 × 106 cells/dose were injected intradermally at 0, 1, 2, 6, 12, and 18 months. Differences in disease free survival (DFS) and overall survival (OS) were analyzed by log rank. Results Of 144 patients randomized, 103 received TLPLDC and 41 received placebo. Within the TLPLDC group, 57 received pretreatment with G-CSF (TLPLDC+G-CSF) and 46 did not (TLPLDC–G-CSF). There were no significant clinicopathologic or treatment differences between the three treatment arms. 36-month DFS was significantly better in TLPLDC–G-CSF vs. TLPLDC+G-CSF or placebo (51.8% vs. 23.4% and 27.1% respectively, p=0.027) (figure 1). TLPLDC–G-CSF had correspondingly improved OS (92.9% vs. 62.8% and 72.3% respectively, p=0.022) (figure 2). Subgroup analysis revealed TLPLDC–G-CSF had increased DFS over TLPLDC+G-CSF or placebo in Stage IV (68.6% vs. 18.8% and 0.0% respectively, p=0.058). Similarly, the DFS survival benefit of TLPLDC–G-CSF was enhanced in patients who received prior immunotherapy (IO) (61.9% vs. 11.5% and 35.7% respectively, p=0.007) or checkpoint inhibitors (CPI) (48.5% vs. 10.6% and 37.5% respectively, p=0.039). Conclusions TLPLDC vaccine created without G-CSF pretreatment significantly improved 36-month DFS and OS compared to TLPLDC+G-CSF or placebo in stage III/IV (resected) melanoma patients. On further subgroup analysis, the increases in OS and DFS were more profound in patients who received additional immune therapies to include CPI. Ongoing evaluation will determine if G-CSF mobilization leads to collection of phenotypically different DCs. Based on these results, we are planning a phase III trial of TLPLDC–G-CSF + CPI vs. placebo + CPI in advanced melanoma post-resection. Trial Registration ClinicalTrials. gov Identifier: NCT02301611 Ethics Approval This study was reviewed and approved by the IRB or Independent Ethics Committee (IEC) of each participating center prior to study initiation.

Background The autologous tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine is safe and effective in improving 24 and 36-month disease-free survival (DFS) in patients (pts) with resected stage III/IV melanoma who completed the primary vaccine series. The tumor lysate, particle only (TLPO) vaccine has been developed to accelerate production by omitting DC isolation and ex vivo loading in favor of in vivo phagocytosis of the TL-loaded particles. We are currently conducting a randomized and double-blind trial of the TLPO vs TLPLDC to improve DFS and overall survival (OS) in patients with resected late stage melanoma. Methods Patients with stage III/IV melanoma who were clinically disease-free after standard of care therapies were randomized to receive TLPO vs TLPLDC (2:1) as a continuation of the phase IIb trial comparing TLPLDC vs placebo (2:1). For the TLPLDC vaccine, autologous TL was loaded into yeast cell wall particles (YCWP) which were then phagocytized by isolated autologous DC ex vivo. For the placebo DC were loaded with empty YCWP. For TLPO, the autologous TL-loaded YCWP were coated with a chemoattractant and injected intradermally for in vivo phagocytosis. Some patients in the TLPLDC arm received G-CSF prior to DC harvest to minimize blood draw (60 mL instead of 120 mL without G-CSF). For all arms, six vaccine/placebo doses were administered intradermally at 0, 1, 2, 6, 12, and 18 mos. Data was analyzed by an intention-to-treat (ITT) analysis for DFS and OS by the Kaplan-Meier method and compared by log-rank test. Results 63 pts were randomized to TLPO (n=43) vs TLPLDC (n=20). The TLPO cohort contained more females and received less chemotherapy (0% vs 10%), but otherwise were comparable. There were no differences in DFS (p=0.948) or OS (p=0.779) between the two vaccines (figures 1&2). Comparing the TLPO pts to all other pts in the phase IIb trial [TLPLDC+G-CSF (n=57), TLPLDC-G-CSF (n=46), and placebo (n=41)] the TLPO arm had improved DFS compared to placebo (p=0.019) and TLPLDC+G-CSF (p=0.001), but roughly equivalent to the TLPLDC-G-CSF arm (p=0.276) (figure 3). A similar trend was seen in OS analysis, though differences were not statistically significant (figure 4). Conclusions TLPO and TLPLDC vaccines (without the use of G-CSF) improve DFS in patients with resected stage III/IV melanoma compared to placebo. The TLPO vaccine may offer advantages via reduced cost and vaccine production time. TLPO should be closely considered for further clinical trials. Trial Registration NCT02301611: Phase IIB TL + YWCP + DC in MelanomaTLPLDC IND#16101TLPO IND#17274 Ethics Approval This study was approved by WIRB; protocol #20141932

Background The tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine is created ex vivo by loading autologous dendritic cells (DC) with yeast cell wall particles (YCWP) containing autologous tumor lysate, thus delivering tumor antigens to the DC cytoplasm via phagocytosis. TLPLDC then activates a robust T cell response against the unique antigens for each patient. The primary analysis of the prospective, randomized, multi-center, double-blind, placebo-controlled phase IIb trial in patients with resected stage III/IV melanoma showed TLPLDC improved 24-month disease-free survival (DFS) in the per-treatment (PT) analysis (patients completing the 6-month primary vaccine series). Here, we examine the secondary endpoint of 36-month DFS and overall survival (OS). Methods Patients with resected stage III/IV melanoma were randomized 2:1 to TLPLDC vaccine or placebo (autologous DC loaded with empty YCWP). Treatments were given at 0, 1, 2, 6, 12 and 18 months. The protocol was amended to include patients receiving concurrent checkpoint inhibitors (CPIs) to follow changes in standard of care. The co-primary endpoints were 24-month DFS by intention-to-treat (IT) analysis and per-treatment (PT) analysis, with secondary endpoints including 36-month DFS and OS by ITT and PT analysis, pre-specified analysis by stage, and safety as measured by CTCAE v4.03. Results Overall, 103 patients received TLPLDC and 41 placebo. In PT analysis, 65 patients received TLPLDC and 32 placebo. Total adverse events (AEs), grade 3+ AEs, and serious AEs (SAEs) were similar in placebo vs TLPLDC groups, with one related SAE per treatment arm. By ITT analysis, 36-month OS was 76.2% for TLPLDC vs 70.3% for placebo (HR 0.72, p=0.437) and 36-month DFS was 35.6% vs 27.1% (HR 0.95, p=0.841). By PT analysis, 36-month DFS was improved with TLPLDC (57.5% vs 35.0%; HR 0.50, p=0.025, figure 1). This effect was even more dramatic in resected stage IV patients (36-month DFS: 60.9% vs 0%; HR 0.12, p=0.001, figure 2). Conclusions This phase IIb trial again demonstrates the safety of the TLPLDC vaccine, and an improved 36-month DFS in patients with resected stage III/IV melanoma who complete the primary vaccine series, particularly in the stage IV subgroup. Next, a phase III trial will evaluate the efficacy of TLPLDC vaccine as adjuvant treatment for resected stage IV melanoma, with patients randomized to receive standard of care PD-1 inhibitors + TLPLDC versus PD-1 inhibitors + placebo. Trial Registration This is a phase IIb clinical trial registered under NCT02301611 Ethics Approval This study was approved by Western IRB, protocol 20141932.

HER2-targeted therapy has not benefited patients with low levels of HER2 expression; however, combination therapy may be effective. Primary analysis of a phase IIb trial investigating the HER2-derived vaccine nelipepimut-S (NPS) did not benefit the intention-to-treat population, but subset analysis showed a benefit in triple-negative breast cancer (TNBC) patients. The subset analysis of this multicenter, randomized, single-blind, phase IIb trial identified significant improvement in 36-month disease-free survival (DFS) between NPS (n = 55) and placebo (n = 44) in TNBC (HR 0.25, p = 0.01) and those who express HLA-A24 (HR 0.41, p = 0.05). The TNBC cohort demonstrated improved 36-month DFS in those with HER2 1+ expression (HR 0.17, p = 0.01), HLA-A24 positivity (HR 0.08, p 0.01), or in those who received neoadjuvant chemotherapy (HR 0.21, p 0.01). NPS vaccination with trastuzumab was associated with improved 36-month DFS among patients with TNBC. The observed benefit to this high-risk subgroup warrants confirmation in a phase III trial.

No approved medical therapies prevent progression of low-grade prostate cancer. Rapamycin inhibits cell proliferation and augments immune responses, producing an antitumor effect. Encapsulated rapamycin (eRapa) incorporates rapamycin into a pH-sensitive polymer, ensuring consistent dosing. Here, we present results from a phase I trial evaluating the safety and tolerability of eRapa in patients with prostate cancer. Patients with Gleason ≤7 (3+4) disease (low and intermediate risk) under active surveillance were enrolled in a 3+3 study with three eRapa dosing cohorts (cohort 1, 0.5 mg/week; cohort 2, 1 mg/week; and cohort 3, 0.5 mg/day). Patients were treated for 3 months and followed for an additional 3 months to assess safety, pharmacokinetics, quality of life (QoL), immune response, and disease progression. Fourteen patients (cohort 1, n = 3; cohort 2, n = 3; and cohort 3, n = 8) were enrolled. In cohort 3, one dose-limiting toxicity (DLT; neutropenia) and two non-DLT grade 1–2 adverse events (AE) occurred that resulted in patient withdrawal. All AEs in cohorts 1 and 2 were grade 1. Peak serum rapamycin concentration was 7.1 ng/mL after a 1 mg dose. Stable trough levels (∼2 ng/mL) developed after 48–72 hours. Daily dosing mildly worsened QoL, although QoL recovered after treatment cessation in all categories, except fatigue. Weekly dosing increased naïve T-cell populations. Daily dosing increased central memory cell populations and exhaustion markers. No disease progression was observed. In conclusion, treatment with eRapa was safe and well-tolerated. Daily dosing produced higher frequencies of lower grade toxicities and transient worsening of QoL, while weekly dosing impacted immune response. Future studies will verify clinical benefit and long-term tolerability. Prevention Relevance: There is an unmet medical need for a well-tolerated treatment capable of delaying progression of newly diagnosed low-grade prostate cancer. This treatment would potentially obviate the need for future surgical intervention and improve the perception of active surveillance as a more acceptable option among this patient population.

Background The COVID-19 pandemic presents a unique challenge to surgical residency programs. Due to the restrictions recommended by the Centers for Disease Control and Prevention and other organizations, the educational landscape for surgical residents is rapidly changing. In addition, the time course of these changes is undefined. Methods We attempt to define the scope of the problem of maintaining surgical resident education while maintaining the safety of residents, educators, and patients. Within the basic framework of limiting in-person gatherings, postponing or canceling elective operations in hospitals, and limiting rotations between sites, we propose innovative solutions to maintain rigorous education. Results We propose several innovative solutions including the flipped classroom model, online practice questions, teleconferencing in place of in-person lectures, involving residents in telemedicine clinics, procedural simulation, and the facilitated use of surgical videos. Although there is no substitute for hands-on learning through operative experience and direct patient care, these may be ways to mitigate the loss of learning exposure during this time. Conclusions These innovative solutions utilizing technology may help to bridge the educational gap for surgical residents during this unprecedented circumstance. The support of national organizations may be beneficial in maintaining rigorous surgical education.

Variability exists in opioid prescribing practices among surgeons, frequently resulting in the prescription of excessive opioids. This study evaluated the ability of a single educational intervention targeted toward general surgery residents to reduce the quantity of postoperative opioids prescribed.This retrospective cohort study evaluated opioid prescribing practices 12 mo prior to and 6 mo following a 30-min lecture for general surgery residents that discussed prescribing guidelines and multimodal analgesia. Opioid volumes (normalized to oral morphine equivalents, OME), opioid type, nonopioid pain medications, and refills requested were analyzed for opioid-naïve adult patients undergoing excisional breast biopsy (EB), mastectomy (M), laparoscopic appendectomy (LA), laparoscopic cholecystectomy (LC), open umbilical hernia repair (OUHR), open inguinal hernia repair (OIHR), or laparoscopic inguinal hernia repair (LIHR).695 and 376 patients preintervention and postintervention were included, respectively. Median OME prescribed decreased for EB (150 mg to 75 mg, P 0.001), M (225 mg to 150 mg, P = 0.85), LA (150 mg to 94 mg, P 0.001), LC (150 mg to 82 mg, P 0.001), OUHR (150 mg to 103 mg, P 0.001), OIHR (175 mg to 100 mg, P = 0.001), and LIHR (200 mg to 113 mg, P 0.001). Fewer patients received opioids alone and more patients received an opioid with two nonopioid adjuncts (P 0.001). More patients received oxycodone as fewer received acetaminophen-containing opioid combinations (P 0.001). Patients requiring refills decreased (11.9% to 7.2%) (P = 0.014).Following this targeted intervention, patients were discharged with fewer OME and more nonopioid analgesics, even as refill requests decreased. Educating residents on opioid prescription guidelines and multimodal therapy is effective and should be part of the annual didactic curriculum.

Introduction Tumor vaccines use various strategies to generate immune responses, commonly targeting generic tumor-associated antigens. The tumor lysate, particle-loaded, dendritic cell (TLPLDC) vaccine is produced from DC loaded with autologous tumor antigens, creating a patient-specific vaccine. Here, we describe initial phase I/IIa trial results. Methods This trial includes patients with any stage solid tumors, ECOG ≤1, and >4 months life-expectancy. A personalized vaccine is created using 1 mg of tumor and 120 ml blood (to isolate DC). Primary vaccination series (PVS) is four monthly inoculations. Patients are followed per standard of care (SOC). Endpoints include safety and tumor response (RECIST v1.1). Results 44 patients were enrolled and vaccinated consisting of 31 late stage patients with residual/measurable disease, and 13 disease-free patients after SOC therapies. While 4 patients progressed before completing the PVS, 12/31 (39%) demonstrated clinical benefit (2 complete responses, 4 partial responses, 6 stable disease). In the adjuvant setting, 46% of late stage patients remain disease free at a median of 22.5 months. Conclusions The TLPLDC vaccine is scalable, generates a personalized DC vaccine, and requires little autologous tumor tissue and few DC. The vaccine is safe, with primarily grade 0–2 toxicities, and nearly 40% clinical benefit rate in varied tumors, warranting further study. Trial Registration: ISRCTN81339386, Registered 2/17/2016.

Aim: We developed a novel approach to efficiently deliver autologous tumor antigens to the cytoplasm of dendritic cells (DC) using yeast cell wall particles (YCWP). Materials and Methods: Loading of YCWP, leakage of protein from loaded YCWP and cytoplasmic delivery of YCWP content was assessed using fluorescent-tagged experiments. Spectrophotometric analysis compared the epitope-specific T-cell responses following antigen presentation via YCWP versus exogenous loading. The in vivo effectiveness of tumor lysate (TL) particle loaded DC (TLPLDC) vaccine was assessed using murine melanoma models. Results: In fluorescence-tagged experiments, YCWP efficiently delivered antigen to the cytoplasm of DC. TLPLDC loading was more effective than conventional exogenous loading of DC. Finally, in murine melanoma models, TLPLDC outperformed an analogous dendritoma vaccine. Conclusion: The TLPLDC vaccine is commercially scalable and holds the potential of producing personalized vaccines.

BACKGROUND FBP is overexpressed in 20-50% of breast(B) cancers(Ca) and roughly 90% of endometrial(E) and ovarian (Ov) Ca. E39 (FBP191-199, EIWTHSYKV)+GM-CSF is an HLA-A2 restricted FBP peptide vaccine, which has been shown to generate significant in vivo immunologic response(IR) in a phase I/IIa trial in E Ca and Ov Ca patients (pts). There is a risk of inducing immunologic tolerance after multiple inoculations with a highly immunogenic vaccine. Thus, we are investigating a novel vaccination series using combinations of E39 and E39' (EIWTFSTKV, an attenuated version of E39) in a phase Ib, randomized, single-center trial. We are assessing short and long-term IR. Here, we present the initial IR analysis to the primary vaccination series (PVS) within B Ca pts. METHODS HLA-A2 positive B or Ov Ca pts were enrolled after completion of standard of care therapy and randomized into three arms: EE (6 inoculations of E39); EE'(3 inoculations of E39, then 3 of E39'); or E'E(3 of E39', then 3 of E39). Theoretically, due to lower FBP expression and less aggressive chemotherapy regimens, B Ca pts are more antigen naïve and have a less suppressed immune system. Thus, only B Ca pts were included in this analysis. The PVS includes 6 inoculations total (R1-R6), one every 3-4 weeks, and containing 250mcg GM-CSF+500mcg peptide in the first 5 pts per arm and 1000mcg of peptide in second 5 pts. To assess the in vivo IR, local reaction(LR) was measured 48 hours after each inoculation (R1-R6), and delayed type hypersensitivity(DTH) was measured pre-PVS (R0), 1, and 6-months post-PVS (RC1, RC6). Ex vivo IR was measured via dextramer assay for E39-specific CD8+ T-cells at R0, RC1, and RC6. Statistical analyses were completed using appropriate tests. RESULTS Thirty-five B Ca pts were enrolled, with 27 completing the PVS (EE n=10, EE' n=8, E'E n=9). No clinicopathologic differences between groups or significant toxicities > grade 2 were appreciated. LR increased from R1 to R6 in all groups (ΔEE= 24.80mm, p=0.14; ΔEE'=38.13mm, p=0.07; ΔE'E=8.05mm, p=0.38), the greatest increase approaching statistical significance in the EE' arm. The only arm with a statistically significant increase for in vivo DTH from R0-RC1-RC6 was in the EE' arm (ΔEE=-6.17mm, p=0.27; ΔEE'= 44.58mm, p CONCLUSION In this phase Ib trial analyzing the IR of B Ca pts receiving a different vaccination strategy, both peptides were noted to be safe and immunogenic. While no difference was seen in E39-specific CD8+ T cells between groups, the in vivo response was enhanced with the use of E39' after E39; this may indicate expansion of more effective clonal populations of CD8+ T cells with this strategy. These results may be specific to B Ca pts who are relatively antigen-naïve with relatively intact immune systems. Further analysis of these pts as this trial continues will determine the optimal vaccination strategy capable of stimulating and maintaining an IR to prevent B Ca recurrence. Citation Format: Jackson DO, Qiao N, Peace KM, Hale DF, Vreeland TJ, Greene JM, Berry JS, Trappey AF, Clifton GT, Ibrahim N, Toms A, Peoples GE, Mittendorf EA. Determining the optimal vaccination strategy using a combination of the folate binding protein (FBP) peptide vaccine (E39+GM-CSF) and an attenuated version (E39') to maximize the immunologic response in breast cancer patients [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P6-10-04.

Introduction: Pre-operative MRI is being used with increasing frequency to evaluate breast cancer patients, but the debate surrounding risks and benefits of this use continues. At our institution, we instituted a standardized protocol for pre-operative MRI. Here, we compare patients seen prior to routine use of MRI to those seen after and examine effects on surgical choices, timing and outcomes. Methods: This is a retrospective review of a prospectively collected database of all new invasive breast cancers seen from January 2007 to December 2012. The control group (CG) did not receive MRI, while the MRI group (MRG) underwent MRI according to our pretreatment protocol. Groups were compared with regards to basic demographics, initial surgical choices, need for re-excision, and surgical timing. The electronic medical records of patients in the MRG who underwent mastectomy as their initial surgery were examined closely to determine the main factors leading to their choice of surgery. Finally, correlation between findings on MRI and final surgical pathology was analyzed. Results: Of 282 patients included, 38 were in the CG and 244 in the MRG; the groups were well matched. The MRG had a significantly higher percentage of patients choosing initial mastectomy (MRG: 47.1% vs CG 21.1%, p=0.003). Patients seen in the first 2 years of the study were less likely to choose mastectomy than those enrolled in the latter years (29.2%vs 48.6%, p=0.004). The MRG had a lower chance of return to the operating room for re-excision (15.2% vs 28.9%, p=0.035). The average time from initial imaging to initial surgery was approximately the same between groups (MRG: 39.7 days vs CG 42.1 days, p=0.45) and the MRG actually had shorter time to definitive (margin-negative) surgical management (MRG: 43.5 days vs CG: 50.3 days, p=0.079). One hundred-fifteen patients in the MRG underwent mastectomy as initial surgery. Of these, 64 (55.7%) had no additional findings on MRI and chose mastectomy based on patient preference; 30 patients (26.1%) (29 unilateral, 1 bilateral) had mastectomy because of MRI findings. Of the 31 breasts removed (29 unilateral and 1 bilateral mastectomies) because of MRI findings, 26 (83.9%) had histologic findings that correlated with the MRI findings, while 5 (16.1%) did not. Conclusion: Patients receiving routine pre-treatment MRI had an increased mastectomy rate, but had a lower re-excision rate. We found no delay to initial surgical therapy and, perhaps more importantly, a slight decrease in time to margin-negative surgical therapy in the MRI group. Women choosing mastectomy after MRI did so because of personal preference over half of the time, while MRI findings influenced this choice in 26% of these women. When MRI findings did lead to mastectomy, these findings were confirmed by pathology results in the vast majority of cases.

9548 Background: A randomized, double-blind, placebo-controlled phase IIb trial of the tumor lysate, particle loaded, dendritic cell (TLPLDC) vaccine was conducted to prevent recurrence in patients (pts) with resected stage III/IV melanoma. Two methods for dendritic cell (DC) harvest were used for vaccine production, including no pretreatment or pretreatment with granulocyte-colony stimulating factor (G-CSF) in an attempt to reduce blood draw volumes. This analysis investigates differences in clinical outcomes and RNA gene expression between these DC harvest methods for TLPLDC vaccine creation. Methods: The TLPLDC vaccine is created by loading autologous tumor lysate into yeast cell wall particles (YCWPs) and exposing them to phagocytosis by DCs. By investigator/pt choice, pts had 120mL of blood drawn for DC harvest, or pts received 300μg of G-CSF for pre-DC mobilization and a 50-70 mL blood draw 24-48 hours later. Total vaccine production time was 72 hrs. Pts were randomized 2:1 to receive TLPLDC or placebo (DCs exposed to empty YCWPs). 1-1.5 x10^6 cells/dose were injected intradermally at 0, 1, 2, 6, 12, and 18 months. Differences in disease free survival (DFS) and overall survival (OS) were evaluated by Kaplan Meier analysis between pts who did not receive pretreatment (TLPLDC), pts who did receive pretreatment with G-CSF (TLPLDC+G), and pts receiving placebo. RNA-seq analysis was performed on the total RNA of pts’ prepared TLPLDC vaccines to assess gene expression. Relative RNA expression (RRE) was compared between TLPLDC and TLPLDC+G. Results: As previously reported, 144 pts were randomized: 103 received TLPLDC (46 TLPLDC, 57 TLPLDC+G) and 41 received placebo. There were no significant clinicopathologic or treatment differences between the three treatment arms. Survival was significantly improved in TLPLDC compared to TLPLDC+G or placebo, including 36-month OS (92.9% vs 62.8% vs 72.3% respectively, p = 0.022) and DFS (51.8% vs 23.4% vs 27.1%, p = 0.027). When compared to TLPLDC+G (n = 3) vaccine, RNA-seq from TLPLDC vaccine (n = 3) showed upregulation of genes associated with DC maturation, including HLA-DMB (RRE: 3.60), IFIT1 (3.38), CD27 (3.26), IFI44L (3.24), MX1 (2.96), HLA-DQA1 (2.67), HLA-DRA (2.40), CD49D (2.34) and CD74 (2.09), while downregulated genes were associated with DC suppression or immaturity including SERPINA1 (RRE:7.8), TLR2 (6.65), CCR1 (5.11), IL10 (4.19), CD93 (3.84) and CD14 (3.25). Conclusions: Pts receiving TLPLDC vaccine had significantly improved OS and DFS, while outcomes remained similar between those who received TLPLDC+G vs placebo. Pts who did not receive G-CSF had higher expression of genes linked to DC maturation and antigen processing and presentation, likely explaining the improvement in clinical efficacy. A phase III trial to further assess the TLPLDC vaccine to prevent recurrence is planned. Clinical trial information: NCT02301611.

// Elizabeth A. Mittendorf 1,* , Alexandros Ardavanis 2 , Jennifer K. Litton 3 , Nathan M. Shumway 4 , Diane F. Hale 5 , James L. Murray 3 , Sonia A. Perez 2 , Sathibalan Ponniah 6 , Constantin N. Baxevanis 2 , Michael Papamichail 2 and George E. Peoples 7,* 1 Department of Breast Surgical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA 2 Cancer Immunology and Immunotherapy Center, St. Savas Cancer Hospital, Athens, Greece 3 Department of Breast Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA 4 Department of Hematology/Oncology, Brooke Army Medical Center, Ft. Sam Houston, TX, USA 5 Department of Surgery, Brooke Army Medical Center, Ft. Sam Houston, TX, USA 6 Cancer Vaccine Development Laboratory, Department of Surgery, Uniformed Services University of the Health Sciences, Bethesda, MD, USA 7 Cancer Vaccine Development Program, San Antonio, TX, USA * These authors have contributed equally to this work Correspondence to: Elizabeth A. Mittendorf, email: // George E. Peoples, email: // Keywords : vaccine, cytotoxic T lymphocytes, breast cancer, HER2, trastuzumab Received : February 07, 2016 Accepted : July 09, 2016 Published : August 31, 2016 Abstract GP2 is a HER2-derived, HLA-A2+ restricted peptide. Phase I studies showed GP2 administered with GM-CSF to be safe and immunogenic. Here we report the primary analysis of a prospective, randomized, multicenter phase II adjuvant trial conducted to determine the vaccine’s efficacy. The trial enrolled HLA-A2+, clinically disease-free, node-positive and high-risk node-negative breast cancer patients with tumors expressing HER2 (immunohistochemistry[IHC] 1+-3+). Patients were randomized to GP2+GM-CSF versus GM-CSF alone. Disease-free survival (DFS) was analyzed in intention-to-treat (ITT) and per-treatment cohorts; pre-specified subgroup analyses were performed for patients with IHC 3+ or FISH+ disease. The trial enrolled 180 patients; 89 received GP2+GM-CSF and 91 received GM-CSF alone. The groups were well-matched for clinicopathologic characteristics. Toxicities have been minimal. The Kaplan-Meier estimated 5-year DFS rate in the ITT analyses was 88% (95% CI:78-94%) in vaccinated vs . 81% (95% CI:69-89%) ( P = 0.43) in control patients after a 34 month median follow-up. In the per-treatment analysis, the estimated 5-year DFS rates were 94% (95% CI:83-98%) and 85% (73-92%) ( P = 0.17). In IHC 3+/FISH+ patients, the estimated 5-year DFS rate was 94% (82-98%) in vaccinated patients ( n = 51) vs . 89% (71-96%) in control patients ( n = 50), ( P = 0.86) in the ITT analyses and 100% vs . 89% (71-96%) in vaccinated vs . control patients in the per-treatment analyses ( P = 0.08). While the overall ITT analysis did not demonstrate benefit to vaccination, this trial confirmed that the GP2 vaccine is safe and suggests that vaccination may have clinical activity, particularly in patients with HER2 overexpression who received the full vaccine series (ie per-treatment group).

Introduction: Great advances have been made in screening for and treatment of colorectal cancer (CRC), but recurrence rates remain high and additional therapies are needed. There is great excitement around the field of immunotherapy and many attempts have been made to bring immunotherapy to CRC through a cancer vaccine.Areas covered: This is a detailed review of the last decade’s significant CRC vaccine trials.Expert commentary: Monotherapy with a CRC vaccine is likely best suited for adjuvant therapy in disease free patients. Vaccine therapy elicits crucial tumor infiltrating lymphocytes, which are lacking in microsatellite-stable tumors, and therefore may be better suited for these patients. The combination of CRC vaccines with checkpoint inhibitors may unlock the potential of immunotherapy for a much broader range of patients. Future studies should focus on vaccine monotherapy in correctly selected patients and combination therapy in more advanced disease.

Background AE37 is the Ii-Key hybrid of the MHC class II peptide, AE36 (HER2 aa:776–790). Phase I studies showed AE37 administered with granulocyte macrophage colony-stimulating factor (GM-CSF) to be safe and highly immunogenic. A prospective, randomized, multicenter phase II adjuvant trial was conducted to evaluate the vaccine's efficacy. Methods Clinically disease-free node-positive and high-risk node-negative breast cancer patients with tumors expressing any degree of HER2 [immunohistochemistry (IHC) 1–3+] were enrolled. Patients were randomized to AE37 + GM-CSF versus GM-CSF alone. Toxicity was monitored. Clinical recurrences were documented and disease-free survival (DFS) analyzed. Results The trial enrolled 298 patients; 153 received AE37 + GM-CSF and 145 received GM-CSF alone. The groups were well matched for clinicopathologic characteristics. Toxicities have been minimal. At the time of the primary analysis, the recurrence rate in the vaccinated group was 12.4% versus 13.8% in the control group [relative risk reduction 12%, HR 0.885, 95% confidence interval (CI) 0.472–1.659, P = 0.70]. The Kaplan–Meier estimated 5-year DFS rate was 80.8% in vaccinated versus 79.5% in control patients. In planned subset analyses of patients with IHC 1+/2+ HER2-expressing tumors, 5-year DFS was 77.2% in vaccinated patients (n = 76) versus 65.7% in control patients (n = 78) (P = 0.21). In patients with triple-negative breast cancer (HER2 IHC 1+/2+ and hormone receptor negative) DFS was 77.7% in vaccinated patients (n = 25) versus 49.0% in control patients (n = 25) (P = 0.12). Conclusion The overall intention-to-treat analysis demonstrates no benefit to vaccination. However, the results confirm that the vaccine is safe and suggest that vaccination may have clinical benefit in patients with low HER2-expressing tumors, specifically TNBC. Further evaluation in a randomized trial enrolling TNBC patients is warranted.

Cancer vaccines have the potential to provide a nontoxic treatment for the prevention of cancer recurrence in the adjuvant setting. Many cancer vaccines have been tested in multiple phase III trials with minimal success. However, through these failed clinical trials, we have learned that the ideal setting for vaccine therapy is the adjuvant setting. Also, we have learned important lessons about patient selection to maximize the probability of success. This article will highlight some of the successes, our trial results in the adjuvant setting, and future directions.

Stage IV melanoma has high mortality, largely unaffected by traditional therapies. Immunotherapy including cytokine therapies and checkpoint inhibitors improves outcomes, but has significant toxicities. In this phase I/IIa trial, we investigated safety and efficacy of a dendritoma vaccine, an active, specific immunotherapy, in stage IV melanoma patients. Autologous tumor lysate and dendritic cells were fused creating dendritoma vaccines for each patient. Phase I patients were vaccinated every 3 months with IL-2 given for 5 days after initial inoculation. Phase IIa patients were vaccinated every 6 weeks with IL-2 given on days 1, 3 and 5 after initial inoculation. Toxicity and clinical outcomes were assessed. Twenty-five patients were enrolled and inoculated. All dendritoma and IL-2 toxicities were

Introduction: We are conducting a phase II clinical trial of the GP2+GM-CSF vaccine for the prevention of breast cancer recurrence in disease-free, node-positive or high-risk node negative patients (pts). GP2 is an HLA-A2+-restricted immunogenic peptide from the HER2 protein (aa: 654-662) capable of stimulating CD8+ cytotoxic T-lymphocytes (CTLs) to recognize and destroy HER2-expressing tumor cells. Here, we present the final pre-specified analysis of our study, disease free survival (DFS) at one year from last enrolled pt. Methods: The trial is a prospective, randomized, multi-center, placebo-controlled, single-blinded, phase I/II trial designed to evaluate the safety and clinical efficacy of GP2 in breast cancer patients. Pts with any level of HER2 (immunohistochemistry [IHC] 1-3+) expression were enrolled after standard of care therapy. Blinded HLA-A2+ pts were randomized 1:1 to receive GP2 + GM-CSF or GM-CSF alone. Pts in both treatment arms received 6 monthly intradermal inoculations during the primary vaccine series (PVS) followed by 4 booster inoculations administered every 6 months. Data was collected for demographic, safety, immunologic, and clinical recurrence. Per the statistical plan, DFS was analyzed for both intention-to-treat population (ITT; all randomized pts) and per-treatment population (PT; excluding recurrences during the PVS and second malignancies) using Kaplan-Meir methods. Continuous variables are compared using analysis of variance techniques and proportions compared with Fisher's exact test. Results: This trial enrolled 180 pts (vaccine group n=89 and control group n=91). There were no differences in age, grade, hormone receptor status, tumor size or nodal status between groups (all p>0.2). With median follow up of 36.2 months, the five-year DFS estimates did not demonstrate a difference between vaccinated (VG) and control groups (CG) with ITT (82.6% v 80.4%, p=0.96, respectively) or PT (88.9% v 84.3%, p=0.52, respectively). On further subset analysis, in HER2 3+ by IHC (or positive via FISH) pts, ITT analysis showed a DFS of 93.8% v 87.2% (VG, n=50 v CG, n=51; p=0.67, respectively) while the PT analysis revealed a DFS of 100% v 87.2% (VG, n=48 v CG, n=50; p=0.052, respectively). In ER/PR+ pts, ITT analysis showed a DFS of 88.1% v 80.6% (VG, n=54 v CG, n=60; p=0.41, respectively) while PT analysis demonstrated a DFS of 91.5% v 85.2% (VG, n=52 v CG, n=57; p=0.42, respectively). Conclusions: The final pre-specified analysis of this randomized phase II trial of the GP2+GM-CSF adjuvant breast cancer vaccine shows no statistical differences between treatment arms. However, it does identify a particular patient population where the vaccine may have efficacy; HER2 3+ patients appear to derive the greatest clinical benefit from GP2 vaccination after trastuzumab treatment with the PT analysis nearing statistical significance. Further studies are required to confirm these findings and investigate the potential link between hormone receptor status and the induction of clinically beneficial anti-tumor immunity with this vaccine. Citation Format: Greene JM, Schneble EJ, Perez S, Murray JL, Berry JS, Trappey AF, Hale DF, Vreeland TJ, Clifton GT, Ardavanis A, Litton JK, Shumway NM, Papamichail M, Peoples GE, Mittendorf EA. Final pre-specified analysis of the phase II trial of the GP2+GM-CSF peptide vaccine in high risk breast cancer patients to prevent recurrence. [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 P2-11-01.

Background: The treatment of melanoma has changed drastically with the advent of immunotherapy, particularly checkpoint inhibition (CPI). Unfortunately, only 50-60% of tumors respond to current immunotherapy. The autologous tumor lysate, particle loaded, dendritic cell (TLPLDC) vaccine utilizes the whole antigenic panel of the patient's tumor to stimulate T-cells and may work synergistically with other immunotherapies. We have previously demonstrated both safety and efficacy of TLPLDC vaccine as adjuvant therapy for patients (pts) with resected stage III or IV melanoma who complete the primary vaccine series. Here, we describe results in pts with metastatic melanoma treated with the TLPLDC vaccine together with other approved therapies. Methods: The TLPLDC vaccine is created by loading autologous tumor lysate into yeast cell wall particles which are then phagocytosed by autologous dendritic cells ex-vivo. Pts who recurred while enrolled in a randomized phase IIb trial of adjuvant TLPLDC vaccine (crossover arm) and pts with metastatic melanoma with measurable disease (MM arm) were offered TLPLDC vaccine in an open-label fashion in addition to other approved therapies as determined by their treating physician. The vaccine is given via intradermal injection monthly x4 followed by boosters at six and nine months. Tumor response is measured by RECIST 1.1 criteria. Results: To date, 53 pts have been enrolled (28 in the crossover arm and 25 in the MM arm). Within the crossover arm, 18 pts had received TLPLDC as part of the phase IIb trial, 8 were from the placebo group, and 2 patients withdrew prior to vaccination. Within the MM arm, 2 were excluded for no evidence of disease at time of vaccination and 1 withdrew. The vaccine was well tolerated, with 24% of patients experiencing a treatment-related adverse event (AE), with no grade ≥3 AE. Concurrent therapies were varied in the crossover arm, but 36% received CPI, compared with 68% in the MM arm. Of those receiving CPI, 59% had >1 immune-related AE, none of which were grade ≥3. In the crossover arm, 12-month overall survival (OS) was 83.3% in the vaccine-naïve cohort at median follow-up of 6.0 months vs. 94.7% in the re-vaccinated cohort at median follow-up of 9.0 months (p=0.961). In the MM arm, 12-month OS was 61.2% at a median follow-up of 7.7 months. In a pooled analysis of 33 evaluable pts, vaccine-naïve pts (n=21) had a 42.9% non-progression rate compared to a 66.7% non-progression rate in re-vaccinated pts (n=12). Conclusion: Vaccination with the TLPLDC vaccine in combination with approved systemic therapies in MM pts is well tolerated, with acceptable rates of immune-related AE when combined with CPI. Additionally, vaccination, and specifically re-vaccination, may provide clinical benefit particularly if used in combination with other approved therapies. Citation Format: Robert Connor Chick, Annelies T. Hickerson, Guy Travis Clifton, Phillip M. Kemp Bohan, Tommy A. Brown, Jessica L. Cindass, John W. Myers, Timothy J. Vreeland, Doreen O. Jackson, Diane F. Hale, Kaitlin M. Peace, Garth S. Herbert, Xianzhong Yu, Thomas Wagner, George E. Peoples. Safety and efficacy of autologous tumor lysate particle loaded dendritic cell (TLPLDC) vaccination in combination with systemic therapies in patients with metastatic melanoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6536.

Background: Immunotherapy has become a mainstay of adjuvant treatment of advanced melanoma, but recurrence rates remain high. TLPLDC is a personalized vaccine that was shown in a multi-center, double-blind, placebo-controlled phase IIb trial to be safe, well tolerated, and effective for preventing recurrence in patients with resectable stage III or IV melanoma. The trial enrolled patients who had either a new diagnosis of melanoma or recurrent disease. It is hypothesized that the length of disease-free interval (DFI) may be a marker of more indolent disease and therefore a good prognostic factor for vaccine efficacy. In this study, we examined the impact of disease-free interval on response to the vaccination. Methods: Vaccine production and dosing has been described elsewhere. Patients were characterized as having either primary or recurrent disease at the time of enrollment. For recurrent patients, DFI was defined as the length of time between their initial biopsy establishing a diagnosis of melanoma and complete surgical resection. Patients were grouped by DFI into a primary and recurrent group, where the recurrent group had DFI >3 months and the primary group had no prior disease or recurrence with DFI Results: Patients were randomized 2:1 (103 vaccine group, 41 control group) and divided into a recurrent group (n=48) and a primary group (n=96). There was no difference between groups in stage, age, margin status, BRAF mutation, or use of concurrent immunotherapy including checkpoint inhibitors. Patients in the recurrent group had numerically improved 24-month DFS with vaccine vs. placebo (52.6% vs. 23.5%, p=0.214) whereas the primary group had no difference (32.9% vs. 31.8%, p=0.451). Patients in the recurrent group had significantly improved overall survival with vaccine vs. placebo (94.4% vs. 50.5%, p=0.011) whereas the primary group had no difference (83.4% vs. 90.2%, p=0.779). Conclusions: The TLPLDC vaccine improved overall survival in patients with DFI >3 months prior to recurrence. These patients have more indolent disease biology and a less immunosuppressive tumor microenvironment, where monotherapy with a cancer vaccine is thought to be more effective. Further study is required to identify patients at initial presentation who are more likely to benefit from the TLPLDC vaccine. Meanwhile, combination therapy with vaccine and checkpoint inhibitor may be required for those with more aggressive disease biology. Citation Format: Robert Connor Chick, Phillip M. Kemp Bohan, Timothy J. Vreeland, Annelies T. Hickerson, Jessica L. Cindass, Diane F. Hale, Guy Travis Clifton, George E. Peoples. Impact of disease-free interval on recurrence in high-risk melanoma patients in a phase IIb trial of the tumor lysate particle loaded dendritic cell (TLPLDC) vaccine [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6537.

34 Background: At low doses, rapamycin inhibits cell proliferation and angiogenesis while augmenting CD8 T-cell responses, cumulatively producing an antitumor effect. Oral rapamycin is limited by variable bioavailability. Encapsulated rapamycin (eRapa) incorporates submicron rapamycin particles into a pH-sensitive polymer, improving bioavailability and allowing for consistent and lower dosing. Here, we present results of a phase Ib trial evaluating safety and treatment effects of eRapa in patients with low-grade prostate cancer (PCa). Methods: PCa patients with Gleason ≤7 (3+4) under active surveillance were enrolled in a 3+3 study with 3 dosing cohorts (0.5mg weekly, 1mg weekly, and 0.5mg daily) to determine the optimal dosing. Patients were treated for 3 months (m) and followed for 6m. Safety, labs (including PSA), pharmacokinetics, immune response, and quality of life (QOL) were assessed for each cohort. Results: 14 patients were enrolled; 3 in cohort 1, 3 in cohort 2, and 8 in cohort 3. 2 patients withdrew for non-dose limiting toxicity (DLT) (grade 1-2) adverse events (AEs) in cohort 3, leaving 6 evaluable. A single grade 3 DLT (neutropenia) occurred in cohort 3. No AEs > Grade 1 occurred in cohorts 1/2. Peak serum rapamycin concentration ([Rapa]) was 7 ng/mL after a 1 mg dose (2h after administration). Stable trough levels (2 ng/mL) were established after 48 hrs and persisted to 13wks. Central memory CD8 T cells and CD3+/CD56+ NK cells were more prevalent in cohort 3 than other cohorts at 1m (p = 0.027 and p = 0.041) and 3m (p = 0.023 and p = 0.049). There was no significant change in PSA level; no patients clinically progressed on therapy. In cohort 3, there were no differences between baseline and 3m QOL assessments but there was a suggestion of withdrawal effects at 6m. Conclusions: Treatment with low dose eRapa is safe and well-tolerated. The dose of 0.5mg daily produced stable serum [Rapa] through the duration of treatment and resulted in a positive immune impact. Further investigation with low dose and/or intermittent dosing of eRapa as a preventive agent in PCa and other indications will be required to establish clinical benefit. Clinical trial information: NCT03618355.

63 Background: A novel vaccine strategy may prevent recurrence in high-risk melanoma patients (pts). The TLPLDC vaccine uses yeast cell wall particles (YCWP) to load tumor lysate into autologous dendritic cells (DC). In this phase IIb trial of TLPLDC vs. placebo in resected stage III/IV pts, TLPLDC increased 24 month (mo) disease free survival (DFS) in the per treatment (PT) population. Here, we present a 24mo DFS subgroup analysis and estimated overall 36mo DFS. Methods: Disease-free pts were randomized 2:1 to the TLPLDC vaccine vs. unloaded YCWP+DC at 0, 1, 2, 6, 12, and 18mo. The protocol was amended to allow concurrent adjuvant checkpoint inhibitor (CPI) therapy once approved. The pre-specified PT population included only pts completing the primary vaccine/placebo series (PVS) at 6 mo. Kaplan-Meier estimates of DFS were used to compare treatment arms by stage (III or IV) and CPI therapy (yes/no) in the ITT and PT populations. Results: 144 pts were randomized (103 TLPLDC, 41 placebo); 98 pts (66 TLPLDC, 32 placebo) completed the PVS. There were no clinicopathologic differences between treatment groups. There was no difference in 24mo DFS in stage III pts (n = 112), but in stage IV pts (n = 32), the 24mo DFS was 44% vs 0% (TLPLDC vs placebo) (p = 0.41) in ITT and 73.3% vs. 0% (HR 0.14, p = 0.002) in PT. Stage IV pts were more likely to receive CPI than stage III pts (50% vs. 30%, p = 0.003). There was no difference in 24mo DFS for pts who did not receive CPI (n = 102), but in pts who received CPI (n = 42), the 24mo DFS was 49.3% vs. 31.3% (p = 0.71) in ITT and 68.8% vs. 41.7% (HR 0.46, p = 0.28) in PT, showing a trend toward improved DFS in pts who completed the PVS and received CPI (n = 31). Overall, the 36mo estimated DFS was 34.2% vs. 21.6% (p = 0.89) for ITT and 56.9% vs. 27.9% (p = 0.021) for PT. Conclusions: The TLPLDC vaccine improved DFS in patients completing the PVS at 24 and 36 mos, particularly in the resected stage IV subset. The apparent synergistic effect with TLPLDC + CPI will be confirmed in a phase III study evaluating adjuvant TLPLDC + CPI vs. CPI alone in resected stage IV melanoma pts. Clinical trial information: NCT02301611.

The development of HER2-targeted therapy has decreased recurrence rates and improved survival, transforming the natural history of HER2-positive breast cancer. However only a minority of breast cancer patients benefit as these agents are not used in patients with tumors expressing low levels of HER2. Preclinical data suggests a synergistic action of HER2-targeted vaccination with trastuzumab. We report the initial safety interim analysis of a phase II trial that enrolled patients with HER2 low-expressing (IHC 1+/2+) breast cancer who were clinically disease-free after standard therapy. Patients were randomized to receive the HER2-peptide vaccine nelipepimut-S + GM-CSF with trastuzumab (vaccine arm) or trastuzumab + GM-CSF (control arm) and were followed for recurrence. A planned analysis that occurred after enrollment of 150 patients showed no significant differences in toxicity between the two arms, including cardiac toxicity. The clinical efficacy of this combination will be reported 6 months after the final patient was enrolled.

In this randomized phase Ib trial, we tested combining the E39 peptide vaccine with a vaccine created from E39’, an attenuated version of E39. Patients with breast or ovarian cancer, who were disease-free after standard of care therapy, were enrolled and randomized to one of three arms. Arm EE received six E39 inoculations; arm EE’ received three E39 inoculations followed by three E39’; and arm E’E received three E39’ inoculations, followed by three E39. Within each arm, the first five patients received 500 μg of peptide and the remainder received 1000 μg. Patients were followed for toxicity, and immune responses were measured. This initial analysis after completion of the primary vaccination series has confirmed the safety of both vaccines. Immune analyses suggest incorporating the attenuated version of the peptide improves immune responses and that sequencing of E39 followed by E39’ might produce the optimal immune response. Trial Registration: NCT02019524

Immunotherapy, using peptide-based cancer vaccines is being studied to assess its potential in breast cancer. Trials of HLA-restricted peptide vaccines have been difficult to enroll given HLA subtype restrictions. It is necessary to determine the prognostic significance of HLA-status in breast cancer if patients who are ineligible to receive a vaccine due to their HLA-status are used as controls. The impact of targeted tumor associated antigen expression, when it effects eligibility is also important. We examined control patients from two randomized phase II trials that tested HER2-peptide vaccines to determine the effect of HLA-A2 status and HER2 expression on disease-free survival. The analysis showed that HLA-A2-status does not affect disease-free survival, regardless of HER2 expression suggesting that HLA-A2 negative patients can be used as control patients. Additionally, HER2 over-expression was associated with a better disease-free survival in this population, underscoring the need for additional therapies in HER2 low-expressing breast cancer. ClinicalTrials.gov Identifier: NCT00524277.

Background: Melanoma is an immunogenic cancer, yet only 50-60% of tumors respond to current immunotherapy. The autologous tumor lysate, particle loaded, dendritic cell (TLPLDC) vaccine may potentiate an immune response by stimulating T-cells. We are conducting a phase IIb double-blinded randomized trial of TLPLDC to prevent recurrence after resection. Patients who recurred were offered enrollment in an open-label TLPLDC vaccine trial along with standard therapy. Here, we describe characteristics and clinical outcomes of recurred patients that continued on open label TLPLDC vaccination. Methods: Patients with resected no evidence of disease (NED) stage III/IV melanoma were randomized to TLPLDC vs. empty yeast cell wall particle (YCWP) loaded DC in a 2:1 fashion. TLPDC is created by loading autologous tumor lysate into YCWP. This is then introduced ex vivo to the patient’s DC for phagocytosis. 1-1.5x106TLPLDCs are given via intradermal injection monthly x4 followed by boosters at 6 and 9 months. Patients who recurred after receiving TLPLDC (vaccine group, VG) or after empty YCWP (control group, CG) on the trial (primary endpoint) were offered open label TLPLDC vaccination with same dosing schedule along with standard therapy as determined by the patient’s treatment team. Disease status is measured by RECIST criteria. Results: To date, 23 (14 VG and 11 CG) patients have enrolled and been vaccinated, with median follow up (fu) of 7.86 months (mo). The patients received the following additional therapies: surgery, gamma-knife, check point inhibitor, radiation, TVEC, BRAF/MEK, and imantinib. No related toxicities > grade 2 were observed. Of 14 VG patients, 5 were NED when re-enrolled, 9 had measurable disease (MD). Of these, 11 have followed up data, 9 (82%) show disease control (5/5 NED patients remain NED, 4/6 with MD show SD), while 2 (18%) have progressed. Of 9 CG patients, 4 were NED and 5 had measurable disease at time of re-enrollment. Of these, 8 have followed up, 5 (62%) show disease control (2/4 NED remain NED, 1/4 with MD shows regression, 2/4 SD), while 3 (38%) have progressed (2/4 NED, 1/3 with MD). Conclusion: Open label administration of TLPLPDC vaccine after recurrence is demonstrated as a safe therapy and shows potential clinical benefit when combined SOC therapy. Further study is warranted to determine if prior exposure to TLPLDC may provide clinical benefit after the first episode of disease recurrence. Citation Format: Annelies T. Hickerson, Guy T. Clifton, Tommy A. Brown, Jessica L. Cindass, John W. Myers, Timothy J. Vreeland, Diane F. Hale, Kaitlin M. Peace, Doreen O. Jackson, Garth Herbert, Xianzhong Yu, Thomas E. Wagner, George E. Peoples. Prior vaccination with the autologous Tumor Lysate Particle Loaded Dendritic Cell (TLPLDC) Vaccine may impact clinical outcomes in melanoma patients treated with systemic therapies and re-vaccination [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT208.

e21025 Background: The treatment of melanoma has changed drastically with the advent of immunotherapy. The autologous tumor lysate, particle loaded, dendritic cell (TLPLDC) vaccine stimulates T-cells and may work synergistically with other immunotherapies. Here, we describe results in patients (pts) with metastatic melanoma (MM) treated with the TLPLDC vaccine together with other approved therapies. Methods: The TLPLDC vaccine is created using autologous tumor lysate loaded yeast cell wall particles to prime autologous dendritic cells ex-vivo. 1-1.5x106 TLPLDCs are given via intradermal injection monthly x 4 followed by boosters at six and nine months (mo). Pts who recurred while enrolled in our adjuvant phase IIb trial of the TLPLDC vaccine and pts with MM with measurable disease enrolled in a separate phase I/IIa trial were offered vaccination of TLPLDC vaccine in an open-label fashion in addition to other approved therapies as determined by their treating physician. Tumor response is measured by RECIST 1.1 criteria. Results: To date, 50 pts have been enrolled in the two trials (25 pts in each). Of the 42 pts with measureable disease, 30 pts received at least one dose of the vaccine, 11 progressed prior to vaccine administration, and 1 is pending. 2 pts withdrew at 2 and 7 mo. Of the remaining 28 evaluable pts, 13 pts had progressive disease with a median follow-up (f/u) of 3 (range 0-12) mo, 12 pts had stable disease with a median f/u of 7.5 (range 1-23) mo, 2 pts had a partial response with f/u of 7 and 13 mo, and one pt had a complete response with 18 mo of f/u. Overall, in pts with measureable disease, the disease control rate was 54% (15/28) and objective response rate was 11% (3/28). 8 pts were without measurable disease at enrollment, 3 recurred at a median f/u of 8 mo and 5 remain disease-free at a median of 26 mo f/u. No grade ≥ 3 toxicities were observed with combination TLPLDC vaccination and approved systemic therapies. Conclusions: Vaccination with the TLPLDC vaccine in combination with systemic approved therapies in MM pts is well tolerated and may provide clinical benefit in patients with and without measurable disease. Clinical trial information: NCT02678741.

556 Background: Breast cancer (BC) patients (pts) expressing low levels of HER2 by (immunohistochemistry (IHC) 1-2+) are not eligible for trastuzumab (Tz). However, in a randomized phase 2b trial, triple negative BC (TNBC) pts demonstrated a significantly better DFS with nelipepimut-S (NPS) + Tz vs Tz alone. Here, we assess the ex vivo and in vivo immune responses (IR) in both arms. Methods: Disease-free pts (n = 275) with HER2 IHC 1-2+, non-amplified BC who were node positive and/or had TNBC were randomized 1:1 to granulocyte-macrophage-colony stimulating factor (GM-CSF) or NPS+GM-CSF. ±NPS was given every 3 weeks x 6 followed by 4 boosters every 6 months (mo). All pts received Tz concurrently for 1 year per label regimen and were followed for recurrence. IR were evaluated ex vivo by clonal expansion of NPS-specific cytotoxic T lymphocytes (CTL) by dextramer-staining/flow cytometry at time points over 3 years. In vivo IR were assessed by delayed type hypersensitivity (DTH) reactions periodically. Results: The trial enrolled 97 TNBC pts; 60 had 4 timepoints available for analysis (37 NPS + Tz pts; 23 Tz pts). The NPS+Tz group exhibited increases in CTL frequencies vs baseline: 208%, 303%, 379% at 18, 24 and 30 mo, respectively. NPS+Tz pts’ mean CTL frequencies increased from 0.029 ±0.001% at baseline to 0.112±0.026% at 30 mo (p = 0.01) compared to Tz pts who were 0.027 ±0.001% at baseline and 0.057 ±0.016% at 30 mo (p = 0.71). Only 4 NPS+Tz pts recurred as compared to 13 in the Tz arm. While limited by low numbers, recurrent NPS + Tz pt did not mount an IR by ex vivo assessment (range: 0.0 - 0.026%) or by DTH (all measurements: 0 mm), while non-recurrent pts mounted both clonal CTL expansion (range: 0.000- 0.33%) and enhanced DTH (range: 0.0- 80.5mm). Conclusions: NPS+Tz combination is more efficacious in generating time-dependent antigen (NPS)-specific CTL by both ex vivo and in vivo measures vs Tz. Based on these preliminary data, it appears that both ex vivo and in vivo IR to NPS were attenuated in pts with TNBC recurrence. Further analysis of read-outs from these assays to validate the relationship of IRs to clinical effect seen with NPS+Tz in TNBC pts is underway. Clinical trial information: NCT02297698.

1 Background: Preclinical data shows synergism between trastuzumab (Tz) and HER2-targeted vaccines. We evaluated adjvuant nelipepimut-S (NPS) + GM-CSF with Tz compared to Tz with GM-CSF alone in HER2 low-expressing (LE) breast cancer (BC) patients (pts) to prevent recurrences. After a planned interim analysis showed benefit in triple negative BC (TNBC) pts, the decision was made to close the trial with guidance from the independent DSMB. Here, we report the final analysis of the trial with 7 months (mo) of added follow-up (f/u). Methods: The phase 2b trial enrolled clinically disease-free BC pts after standard therapy. Pts were HLA-A2, A3, A24, and/or A26+, had HER2-LE (IHC 1-2+, FISH non-amplified) BC and were node positive and/or TNBC. Pts were randomized to placebo with GM-CSF(control group, CG) or NPS with GM-CSF (vaccine group, VG), while all received Tz Q3wk for 1 year. GM-CSF or NPS + GM-CSF were given q3wks x 6 starting with the 3rd Tz dose, and boosters every 6 months x 4. Safety was assessed and pts were followed clinically for recurrences. The primary outcome was DFS at 24 mo. Results: 589 pts were screened at 26 sites. 275 pts were enrolled and randomized (VG:136, CG:139). There were no clinicopathologic differences between groups. Concurrent Tz and NPS was safe with no added overall or cardiac toxicity compared to CG and no grade 4/5 toxicities. In the ITT analysis (median f/u 25.7 mo), the estimated DFS was favorable but did not reach significance in the VG compared with the CG (HR 0.62, 95% CI: 0.31-1.25, p = 0.18). In the TNBC pts, the VG had statistically improved DFS compared to the CG (HR 0.26, 95% CI: 0.08-0.81, p = 0.013). Conclusions: The combination of NPS with Tz is safe with no added toxicity compared to Tz alone, even after prolonged exposure (25.7 mo). In this final analysis, there was a trend towards benefit in the ITT population that improved since the interim analysis with added f/u. The significant benefit seen at interim in the TNBC pts continued to strengthen in the VG group. These findings could position the NPS + Tz combination as an adjuvant therapy for early-stage TNBC and warrant further study. Clinical trial information: NCT01570036.

Background: HER2 over-expression is associated with more aggressive malignant disease. The introduction of trastuzumab and other HER2-directed therapies, however, has led to improved prognosis for patients (pts) with HER2 over-expressing (OE) tumors. Currently, no HER2-targeted therapies are available for patients with HER2 low-expressing (LE) (1+, 2+ by IHC) tumors. We are conducting a randomized, controlled Phase II trial of multiple peptide vaccines enrolling patients with any level of HER2 expression (1+, 2+ and 3+). Here, we report survival data based on levels of HER2 expression in our unvaccinated, control pts. Methods: After standard of care therapy, disease-free, high-risk BCa pts were randomized to receive either peptide+GM-CSF (Vaccine Group, VG) or GM-CSF alone (Control Group, CG) in six, monthly doses followed by four boosters every six months. Pts were prospectively followed for recurrence. Demographic information was available for all pts and was compared between groups using chi square or fisher exact tests. Disease-Free Survival (DFS) was compared using log rank. Results: To date, we have enrolled 196 pts in the CG. 96 pts had HER2 OE tumors, 100 had LE tumors. The only significant demographic difference between the CG OE and LE groups was more ER/PR positive patients in LE (LE 72% vs OE 51%, p = 0.008). 83% of CG OE pts received trastuzumab, 3% of CG LE pts received trastuzumab. At a median f/u of 30 mo, DFS was significantly higher for CG OE vs CG LE (92.5% v 65.5%, p = 0.001). Conclusions: In the cohort of control pts from our ongoing vaccine trial, conducted in an era when Tz has been standard of care therapy for patients with HER2 OE tumors, we have shown that HER2 LE pts are at higher risk of recurrence than OE pts, despite having more ER/PR positive. This calls for increased efforts to develop novel therapies for patients with HER2 LE disease. We have previously shown a trend towards increased DFS with the HER2 vaccines, AE37 (p = 0.13, median f/u 22 mo) and E75 (p = 0.16, median f/u 60mo) in HER2 LE pts, suggesting that these vaccines may represent one such novel therapeutic approach. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P2-14-01.

BACKGROUND: HER2 is a commonly expressed tumor-associated antigen in breast (BrCa) and ovarian cancer (OvCa) and, therefore, an attractive target for immunotherapy. We have investigated HER2-derived peptides as vaccines mixed with GM-CSF to include GP2 (a HLA-A2 and HLA-A3 restricted, CD8+ eliciting epitope) and AE37 (a HLA unrestricted, MHC class II, CD4+ eliciting epitope). Both peptide vaccines (PV) have shown clinical promise individually. There is clear rationale for combining GP2 and AE37 to elicit a more robust immune response (IR) of both CD4+ and CD8+ T cells. Here, we summarize initial toxicity (tox) and in vivo IR data from a phase 1 trial of the combined PV. METHODS: The trial is being performed as a five cohort, 3+3 dose-escalation, safety trial. Clinically disease-free, HLA-A2+ and A3+, BrCa and OvCa patients with tumors expressing any level of HER2 (IHC 1-3+) and who have completed standard-of-care therapy are accrued. In the first cohort, three patients received six, monthly intradermal inoculations (R1-R6) of 100mcg of AE37, 100mcg of GP2, and 125mcg of GM-CSF or 100:100:125. The second cohort received 250mcg of AE37, 100mcg of GP2, and 125mcg of GM-CSF or 250:100:125. Three additional cohorts were vaccinated: 250:250:125, 500:250:125, and 500:500:125. Toxicity was graded 48-72 hours post vaccination using NCI Toxicity Criteria v4.0. After each inoculation, local reactions (LR) are measured via the sensitive ballpoint pen method and reported as the orthogonal mean (OM). IR is assessed in vivo by delayed type hypersensitivity (DTH) reactions with separate intradermal inoculations of AE37, AE36, and GP2 antigens, measured both pre-vaccination (R0) and after the vaccine series (R6) via the sensitive ballpoint pen method, and reported as the OM. Means were compared using paired t-tests. RESULTS: 28 patients enrolled; 8 withdrew consent, 1 recurred prior to completing R6, 3 had an intercurrent illness, 14 patients completed R1-R6, and the vaccine series is ongoing in 2 patients. Six patients did not receive any inoculations and, therefore, are not included in this safety analysis. In 22 patients, the vaccine was well tolerated (max local tox: 23% Grade (Gr) 1, 73% Gr 2, 4% Gr 3; max systemic tox: 14% Gr 0, 50% Gr 1, 36% Gr 2). No dose-limiting toxicity was observed. For the 14 patients who completed the VS, the median age was 51(35-83). Breast tumor size was 3.3±1.1cm and ovarian tumor size was 10.0±2.3cm. Compared to GP2 LR at R1 (15.5±4.1mm), LR increased at R2 (31.7±5.9mm), R3 (42.9±7.4mm), R4 (35.3±7.3mm), R5 (45.0±9.9mm), and R6 (25.9±6.7mm, p = 0.17). Compared to the AE37 LR at R1 (18.5±3.8mm), LR increased at R2 (37.3±6.7mm), R3 (36.4±4.6mm), R4 (42.2±5.9mm), R5 (46.0±8.9mm), and R6 (36.2±6.6mm). Unless stated, all LR p-values < 0.05. After the VS, AE37 DTH increased from 0.0±0.0mm to 19.6±6.7mm (p CONCLUSIONS: Initial results from a phase I trial of a vaccine combining GP2 and AE37 peptides show that dual administration of the peptides is well tolerated at all tested dosing levels. Additionally, the combination is capable of stimulating strong peptide-specific in vivo immune responses. Continued testing of this vaccination strategy is underway. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P4-13-02.

O1 IL-15 primes an mTOR-regulated gene-expression program to prolong anti-tumor capacity of human natural killer cells #### Andreas Lundqvist1, Vincent van Hoef1, Xiaonan Zhang1, Erik Wennerberg2, Julie Lorent1, Kristina Witt1, Laia Masvidal Sanz1, Shuo Liang1, Shannon Murray3, Ola Larsson1

// Doreen O. Jackson 1 , Kevin Byrd 2,3 , Timothy J. Vreeland 4 , Diane F. Hale 1 , Garth S. Herbert 1 , Julia M. Greene 1 , Erika J. Schneble 1 , John S. Berry 1 , Alfred F. Trappey 1 , Guy Travis Clifton 5 , Mark O. Hardin 6 , Jonathan Martin 7 , John C. Elkas 8,9 , Thomas P. Conrads 2,3,8,10 , Kathleen M. Darcy 2,3 , Chad A. Hamilton 2,3 , George L. Maxwell 2,3,8,10 and George E. Peoples 7 1 Department of Surgery, San Antonio Military Medical Center, San Antonio, TX, USA 2 National Capital Consortium Fellowship in Gynecologic Oncology, Walter Reed National Military Medical Center Bethesda, MD, USA 3 Gynecologic Cancer Center of Excellence, Annandale, VA, USA 4 Department of Surgery, Womack Army Medical Center, Fayetteville, NC, USA 5 Department of Surgical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA 6 Department of Surgery, Madigan Army Medical Center, Tacoma, WA, USA 7 Cancer Vaccine Development Program, San Antonio, TX, USA 8 Department of Obstetrics and Gynecology, Inova Fairfax Hospital Annandale, VA, USA 9 Mid-Atlantic Gynecologic Oncology and Pelvic Surgical Associates, Annandale, VA, USA 10 Inova Schar Cancer Institute, Inova Health System, Annandale, VA, USA Correspondence to: Doreen O. Jackson, email: // Keywords : cancer, ovarian, endometrial, folate binding protein, immunotherapy Received : July 01, 2016 Accepted : October 19, 2016 Published : November 11, 2016 Abstract BACKGROUND: Folate binding protein(FBP) is an immunogenic protein over-expressed in endometrial(EC) and ovarian cancer(OC). We are conducting a phase I/IIa trial of E39 (GALE 301)+GM-CSF, an HLA-A2-restricted, FBP-derived peptide vaccine to prevent recurrences in disease-free EC and OC patients. This interim analysis summarizes toxicity, immunologic responses, and clinical outcomes to date. METHODS: HLA-A2+ patients were vaccinated(VG), and HLA-A2- or -A2+ patients were followed as controls(CG). Six monthly intradermal inoculations of E39+250mcg GM-CSF were administered to VG. Demographic, safety, immunologic, and recurrence rate(RR) data were collected and evaluated. RESULTS: This trial enrolled 51 patients; 29 in the VG and 22 in the CG. Fifteen patients received 1000mcg E39, and 14 received