AE37 Vaccine Research Articles

AE37 HER2-targeted vaccine in the prevention of breast cancer recurrence: A mini narrative review of current evidence

Breast cancer remains a significant global health challenge, necessitating innovative therapeutic strategies. This review synthesizes findings from multiple studies investigating the safety profile and efficacy of the AE37 human epidermal growth factor receptor 2 (HER2)-targeted vaccine, offering insights into its potential role in breast cancer immunotherapy. A systematic search of electronic databases, including PubMed, MEDLINE, Scopus, and Web of Science, was conducted to identify relevant articles published up to October 2023. The search strategy utilized a combination of keywords, including “AE37 HER2 vaccine,” “breast cancer recurrence prevention,” and related terms. Boolean operators (AND, OR) were employed to refine the search. The AE37 vaccine exhibited a favorable safety profile across all studies, with minimal adverse effects reported. Efficacy outcomes varied, with promising trends observed in specific breast cancer subgroups, such as advanced-stage, HER2 under-expressed, and triple-negative breast cancer patients. Subgroup analyses suggested potential benefits, emphasizing the need for precise patient stratification. While the AE37 HER2-targeted vaccine demonstrates a promising safety profile and potential efficacy in specific breast cancer subgroups, an understanding requires addressing identified limitations and advancing research in nuanced directions. This paper provides a foundation for navigating the complex landscape of breast cancer immunotherapy with the AE37 vaccine.

AE37: a HER2-targeted vaccine for the prevention of breast cancer recurrence

ABSTRACT Introduction HER2 is a prevalent growth factor in a variety of malignancies, most prominently breast cancer. Over-expression has been correlated with the poorest overall survival and has been the target of successful therapies such as trastuzumab. AE37 is a novel, HER2-directed vaccine based on the AE36 hybrid peptide (aa776-790), which is derived from the intracellular portion of the HER2 protein, and the core portion of the MHC Class II invariant chain (the Ii-Key peptide). This hybrid peptide is given with GM-CSF immunoadjuvant as the AE37 vaccine Areas covered This article describes in detail the preclinical science leading to the creation of the AE37 vaccine and examines use of this agent in multiple clinical trials for breast and prostate cancer. The safety profile of AE37 is discussed and opinions on the potential of the vaccine in breast and prostate cancer patient subsets along with other malignancies, are offered. Expert opinion Future trials utilizing the AE37 vaccine to treat other HER2-expressing malignancies are likely to see similar success, and this will be enhanced by combination immunotherapy. Ii-Key modification of other peptides of interest across oncology and virology could yield impressive results over the longer term.

Subgroup efficacy evaluation of the AE37 HER2 vaccine in breast cancer patients in the adjuvant setting.

3088 Background: AE37 is a Ii-Key hybrid of the HER2 peptide AE36 (HER2776-790), which stimulates peptide-specific T cells. We have completed the active phase of a prospective, randomized, multi-center, phase II trial of the AE37 vaccine in the adjuvant setting. The primary analysis, performed after a median follow up (f/u) of 25 months (mo), did not show a significant difference in disease free survival (DFS) between vaccinated and control patients (pts). However, demonstrating the efficacy of cancer vaccines may require more time than other therapies, especially in malignancies with relatively late recurrences like breast cancer. Here, we present updated efficacy data after extended f/u in subgroups of pts stratified by clinicopathologic characteristics. Methods: Clinically disease-free, node positive or high-risk node negative pts with any level of HER2 expression were randomized to receive AE37 + GM-CSF (VG) or GM-CSF alone (CG) following standard of care therapy. Pts received 6 monthly intradermal inoculations during the primary vaccine series (PVS) followed by 4 boosters administered every 6 mo. Kaplan Meier and log rank analyses were performed from the time of the first inoculation in pts who completed at least the PVS, according to stage, node status, tumor size, HER2 expression and ER/PR status. Results: There were no clinicopathologic differences between groups in the 298 enrolled pts (VG = 153, CG = 145). The vaccine is safe and well tolerated. After a median f/u of 55 mo, there was a trend toward improved DFS in the VG among stage IIB/III pts (VG, n = 73, DFS 82% vs CG, n = 61, 67%, HR = 0.48, p = 0.06) and those with low HER2 expression (HER2 LE, VG, n = 68, 89% vs CG, n = 66, 51%, HR = 0.47, p = 0.1). Improved DFS in the VG was documented in patients with both stage IIB/III disease and HER2 LE (VG, n = 39, 90% vs CG, n = 38, 32%, HR 0.3, p = 0.02) and triple negative (TNBC) pts (VG, n = 21, 89% vs CG, n = 21, 0%, HR 0.26, p = 0.05). Conclusions: The AE37 vaccine is safe and well tolerated and has statistically significant efficacy in stage IIB/III pts with HER2 LE and in TNBC pts. This justifies further evaluation in a phase III study enrolling stage IIb/III pts not eligible for trastuzumab treatment and the very high risk TNBC group. Clinical trial information: NCT00524277.

Unraveling the role of preexisting immunity in prostate cancer patients vaccinated with a HER-2/neu hybrid peptide

BackgroundCancer vaccines aim at eliciting not only an immune response against specific tumor antigens, but also at enhancing a preexisting immunity against the tumor. In this context, we recently reported...

Primary analysis of a prospective, randomized, single-blinded phase II trial evaluating the HER2 peptide AE37 vaccine in breast cancer patients to prevent recurrence

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. 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. 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). 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.

MHC class II tetramer analyses in AE37-vaccinated prostate cancer patients reveal vaccine-specific polyfunctional and long-lasting CD4+ T-cells

ABSTRACTRealizing the basis for generating long-lasting clinical responses in cancer patients after therapeutic vaccinations provides the means to further ameliorate clinical efficacy. Peptide cancer vaccines stimulating CD4+ T helper cells are often promising for inducing immunological memory and persistent CD8+ cytotoxic T cell responses. Recent reports from our clinical trial with the AE37 vaccine, which is a HER2 hybrid polypeptide, documented its efficacy to induce CD4+ T cell immunity, which was associated with clinical improvements preferentially among HLA-DRB1*11+ prostate cancer patients. Here, we performed in-depth investigation of the CD4+ T cell response against the AE37 vaccine. We used the DR11/AE37 tetramer in combination with multicolor flow cytometry to identify and characterize AE37-specific CD4+ T cells regarding memory and Tregs phenotype in HLA-DRB1*11+ vaccinated patients. To verify vaccine-specific immunological memory in vivo, we also assessed AE37-specific CD4+ T cells in defined CD4+ memory subsets by cell sorting. Finally, vaccine-induced AE37-specific CD4+ T cells were assessed regarding their functional profile. AE37-specific memory CD4+ T cells could be detected in peptide-stimulated cultures from prostate cancer patients following vaccination even 4 y post-vaccination. The vast majority of vaccine-induced AE37-specific CD4+ T cells exhibited a multifunctional, mostly Th1 cytokine signature, with the potential of granzyme B production. In contrast, we found relatively low frequencies of Tregs among AE37-specific CD4+ T cells. This is the first report on the identification of vaccine-induced HER2-specific multifunctional long-lasting CD4+ T cells in vaccinated prostate cancer patients.

Abstract 2899: Immunologic effects of a HER2 peptide (GP2) vaccine booster in previously vaccinated breast cancer patients

Abstract Background: We have conducted clinical trials of the HER2 peptide GP2 (MHC Class I restricted) vaccine in clinically disease-free breast cancer patients. Our Phase I/II trials showed the GP2+GM-CSF vaccine to be safe and effective in stimulating clonal expansion of GP2-specific cytotoxic CD8+ T-cells (GP2-CTLs) with anti-tumor activity. Here, we present data regarding the safety and immunologic response to boosters following completion of the primary vaccination series (PVS). Methods: Following completion of their PVS, patients enrolled on our phase II trial were administered booster inoculations every 6 months for 4 doses (500mcg GP2 peptide + 125mcg GM-CSF in the vaccine group (VG) or 125mcg GM-CSF alone in the control group (CG)). HLA A2- controls from the parallel study arm evaluating the HER2 peptide AE37 (MHC Class II restricted) vaccine were also eligible for evaluation as an extended CG (ECG). Patients were monitored for local and systemic toxicity, local reactions (LR) were measured, and GP2-CTLs were quantified using the HLA-A2:IgG dimer assay. Results: We have enrolled 294 patients (83 VG, 66 CG, and 145 ECG). Forty-six VG and 104 control (CG + ECG) have received booster doses. Boosters were well tolerated overall with toxicities comparable between VG and CG+ECG (Local: Gr 0: 2% vs 2%, Gr 1: 85% vs 93%, Gr 2: 13% vs 9%; Systemic: Gr 0: 52% vs 52%, Gr 1: 46% vs 45% , Gr 2: 0% vs 2%, Gr 3: 2% vs 1% ). Mean LR increased from booster 1 (B1) to booster 4 (B4) in the VG (B1: 63.8mm, B2: 66.0mm, B3: 85.2mm, B4: 72.6mm) and declined in the CG (B1: 47.8mm, B2: 43.8mm, B3: 40.2mm, B4: 37.8mm). Patients receiving B1 at 6 months after PVS in the VG group (n=46) were defined as early booster (EB, n=30), and those receiving B1 >6 months after PVS were defined as late booster (LB, n=16). GP2-CTLs prior to B1 were lower in LB patients (LB, 0.55±0.17 vs. EB, 1.00±0.16, p=0.07). Post-booster GP2-CTLs were compared between LB and EB at B1 (0.99±0.15 vs. 1.17±0.25, p=0.82), B2 (0.68±0.11 vs. 0.92±0.21, p=0.32), B3 (0.79±0.13 vs. 2.30±0.64, p=0.003), and B4 (1.08±0.43 vs. 3.37, p=na) respectively. EB post-booster GP2-CTLs levels were greater than LB at all time points although both groups sustained an increase in GP2-CTLs throughout the booster series. Conclusions: The GP2 peptide vaccine and boosters are safe with toxicities likely attributable to GM-CSF. The GP2 vaccine elicits a strong HER2-specific immune response that can be maintained with boosters. Based on these data, an optimal booster regimen is being devised. Citation Format: Erika Schneble, John S. Berry, Alfred F. Trappey, Timothy J. Vreeland, Diane F. Hale, Alan K. Sears, Guy T. Clifton, Sathibalan Ponniah, Elizabeth A. Mittendorf, George Peoples. Immunologic effects of a HER2 peptide (GP2) vaccine booster in previously vaccinated breast cancer patients. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2899. doi:10.1158/1538-7445.AM2014-2899

Primary analysis of the prospective, randomized, single-blinded phase II trial of AE37 vaccine versus GM-CSF alone administered in the adjuvant setting to high-risk breast cancer patients.

638 Background: AE37 is the Ii-Key hybrid of the HER2 peptide AE36 (HER2 aa:776-790). It is a MHC class II epitope capable of stimulating CD4+ helper T-cells. We have completed accrual to a prospec...

Effect of immunization with Ii-key modified HER2 (776-790) peptide vaccine (AE37) on immunologic responses in prostate cancer patients.

e16037 Background: We have shown that the AE37 vaccine (Ii-Key modified HER2(776-790) peptide) is safe and induces HER2/neu–specific cellular immune responses in patients with prostate cancer (Perez SA et al Clin. Cancer Res. 2010, 16:3495). We now present data from 4-year immunological assessments of prostate cancer patients who received AE37. Methods: Seventeen patients in a phase I study were given 6 doses of AE37 at monthly intervals and one additional dose a year after initiating treatment. Immunological testing to assess active versus suppressive immunity was conducted one month (intermediate-term immunomonitoring [ITI]) and 3 years (long-term immunomonitoring [LTI]) after the final dose of AE37. ELISPOT and proliferation assays were conducted to assess cytokine secretion and mitogenic response to antigen. DTH reactions were measured to assess in vivo immune response to antigen. All assays were conducted using native HER2(776-790) peptide (AE36). The percent Treg cells and ng/ml TGFβ were determined as markers for immune suppression. Results: Neither ELISPOT nor proliferation assays were statistically different at LTI compared to ITI. While clearly above pre-vaccine responses, the drop in DTH was statistically significant (p < 0.05). Similarly, the increase in Treg cells and circulating TGFβ was also statistically significant. An increase of >200 % in PSA-doubling time at any point during the study was observed in 6/17 patients, with 3 retaining this effect to 5 years. Conclusions: AE37 generates immunological memory associated with possible clinical efficacy in spite of Tregs and TGF-β levels returning at 4 years after being decreased for up to 6 months after initial AE37 vaccination. These results support further randomized testing of the AE37 vaccine. Clinical trial information: 2006-003299-37. [Table: see text]

Booster inoculations of the AE37 peptide vaccine enhance immunological responses in a phase II study.

3095 Background: We are conducting a multicenter randomized phase II trial of AE37, the Ii-Key hybrid peptide of HER2 776-790 (AE36). The purpose of the study is to determine if the AE37 vaccine can prevent recurrence in disease-free conventionally treated node-positive (NP) and high-risk node-negative (NN) breast cancer patients at significant risk for recurrence. Since clinical efficacy is anticipated to occur as the result of long lasting memory immune responses induced by vaccination, repeated booster inoculations were scheduled as part of the trial. Here we present data on immune responses in patients who received boosters up to 24 months after completion of the primary vaccination series (PVS). Methods: The trial is enrolling NP or high-risk NN patients with any degree of HER2 expression (IHC 1-3+ or FISH > 1.2) rendered disease-free following standard of care therapy. The vaccine group (VG) received AE37+GM-CSF and control group (CG) GM-CSF alone in 6 monthly i.d. inoculations followed by boosters administered every 6 months x 4. Immunologic responses were assessed in vivo by dermal reactions at the inoculation site, and in vitro, against the AE36 peptide, with proliferation and IFN-γ ELISPOT assays. Results: 25 patients in the VG and 23 in the CG have completed their boosters. After the last booster (BRC24), 100%, 54% and 54% in the VG (vs. 9%, 18% and 27% in the CG) responded by dermal reaction, proliferation and IFN-γ ELISPOT, respectively. Mean dermal reactions (orthogonal mean in mm) in vaccinated patients was 25.9±3.13 at completion of the PVS (R6) and increased to 35.47±4.35 at BRC24 (p=0.01). VG patients increased their proliferation response (stimulation index, SI) to AE36 from 0.97±0.046 at baseline (R0) before vaccination to 2.27±0.57 at R6 (p=0.0003) which was maintained until BRC24 (SI 2.21±0,33, p<0.0001). The number of IFN-γ specific spots/106 PBMC increased from 26.88±12.36 at R0 to 40.35±17.02 (p=0.07) at R6, up to 62±16.82 (p=0.0076) at BRC24. Conclusions: Our data demonstrate that AE37 vaccine boosters enhance the immune responses against HER elicited during the PVS, thus sustaining long lasting immunity, a prerequisite for possible clinical efficacy which is currently being evaluated. Clinical trial information: NCT00524277.

Early efficacy analysis of the AE37 vaccine in patients with HER2 low-expressing and triple-negative breast cancer.

109 Background: Peptide vaccines comprised of HLA class II epitopes, which elicit CD4+ T cell responses, play a critical role in potentiating immune responses. We are conducting a randomized phase II trial of AE37, a hybrid peptide created by the addition of the Ii-Key moiety (LRMK) to the HER2 helper epitope, AE36 (HER2 aa776-790). Here, we present efficacy data focusing on outcomes in patients with low HER2 (IHC 1+ or 2+) expression and triple negative breast cancer (TNBC). Methods: The trial is enrolling node positive or high risk node negative breast cancer patients with any degree of HER2 expression (IHC 1+, 2+ or 3+ or FISH > 1.2) rendered disease-free following standard of care therapy. Patients are randomized to receive either AE37+GM-CSF or GM-CSF alone in 6 monthly intradermal inoculations followed by booster inoculations administered every 6 months. Results: The trial has enrolled 254 patients; 105 in the vaccine group (VG) and 149 in the control group (CG). After a median follow-up of 22.3 months, the disease-free survival (DFS) rate in the VG is 90.3% vs 81.1% in the CG (p=.46), a 49% risk reduction. Evaluating patients with low HER2 expression (IHC 1+ or 2+), there are 53 VG patients and 77 CG patients. The groups are well-matched with respect to the percentage of patients with high grade tumors, tumors > 2cm, the rate of node positivity and ER/PR status (all p>.5). The DFS rate in the VG of low HER2 expressers is 89.8% vs 68.2% in the CG (p=.12), a 68% risk reduction. When limiting analyses to patients with TNBC (ER/PR negative, HER2 1+ or 2+), there are 13 VG patients and 23 CG patients. The groups are again well-matched with the exception of control patients having a larger percentage of tumors > 2 cm (70% vs 31%; p=.02). The DFS rate in the VG of TNBC patients is 83.3% vs 47.6% in the CG (p=.23), a 68% risk reduction. Conclusions: Early analyses suggest clinical benefit to vaccination with AE37, particularly in patients with low HER2-expressing tumors. Importantly, the benefit appears to persist in TNBC patients. Patients will continue to be followed per protocol for 5 years; however, these data suggest that a subsequent phase III trial should evaluate the vaccine in patients with low HER2-expressing disease to include TNBC.

An assessment of disease features and immune response in breast cancer patients that did not recur after receiving HER2 peptide, AE37 vaccine in a randomized phase II trial.

625 Background: In a phase I trial of AE37, the Ii-Key hybrid of HER2 derived peptide AE36 (776-790), administered with immunoadjuvant GMCSF demonstrated the vaccine to be safe and capable of stimulating CD4+helper T cells with HER2 specific anti-tumor activity. Here we present analysis of immune markers and patient feature that may impact recurrence from an ongoing prospective, randomized, single-blinded Phase IIb trial of AE37+GMCSF v GMCSF alone in adjuvant high risk breast cancer (BC) patients. Methods: After completion of standard therapy; disease-free, node positive or high risk node negative BC patients (pts) were randomized to receive either AE37+GMCSF or GMCSF in 6 monthly intradermal inoculations. Immunologic responses were measured using [3H]-thymidine incorporation assay (in vitro), delayed-type hypersensitivity (DTH) reactions (in vivo) and T regulatory cells (Tregs). Among those vaccinated recurrent pts (VR) are compared to non recurrent (VNR) pts. Results: We have vaccinated 109 pts with 8.3% recurrence rate at 2 year median follow up. VR v VNR were younger (44 v 50 yo p=0.11), had higher grade (67% v 44% p=0.32), more ER/PR- (44% v 38% p=0.75), larger tumors (89% v 50% p=0.06), and node positive (89% v 70% p=0.37). No difference for HER2 status (IHC 3+, 44% v 49% p=0.64). Both VR and VNR responded to vaccine though the mean DTH and proliferative stimulation index was approximately 10% less in VR pts (18 v 20 p=0.73; 1.96 v 2.2 p=0.77 respectively). The most predictive measure was change in Tregs with VR pts less likely to decrease their Tregs levels (50% v 76% p=0.17) after vacination and more likely experience increased Tregs (17% v 8% p=0.48). A decrease in Tregs had an inverse trend towards recurrence (p=0.17). Conclusions: Preliminarily, it appears most pts immunologically respond to vaccine though slightly less for VR in most assays. The changes of Tregs appear to correlate best with disease recurrence. Whether this reflects an association with disease status or a failure of the vaccine is yet to be seen. These levels may become important in predicting risk for clinical recurrence in future vaccine trials.

Effect of immunization with the ii-key modified HER2/neu(776-790) (AE37) peptide vaccine on pre-existent immunity to PSA in HLA-A24+ prostate cancer patients: Association with increased AE37-specific immunologic responses.

e15125 Background: In solid tumors such as prostate cancer, novel biomarkers are needed for assessing immunological responses and clinical efficacy. By using dextramers specific for PSA we have retrospectively analyzed CD8+ T cell frequencies in patients with HER-2/neu+ prostate cancer who had received the AE37 vaccine in a phase I study (Perez SA et al Clin. Cancer Res. 2010, 16:3495). AE37 binds to numerous MHC alleles (Perez SA et al Clin. Cancer Res. 2010, 16:3495) (Salazar LG, Clin Cancer Res. 2003, 9:5559) encouraging its use as a therapeutic vaccine for general HER-2/neu+ cancer patients. Methods: CD8+ Dextramer+ T cells were measured, retrospectively, in the blood (which was stored in N2 liquid) of HLA-A24+ (n= 12) and HLA-A2+ (n=12) patients. Blood lymphocytes collected before, during and 6-months post vaccination were all analyzed with the same method involving direct labeling with HLA A24/PSA(153-161)-FITC and PerCP-conjugated anti-CD8 monoclonal antibody. Results: 8 of 12 HLA-A24+ patients had preexistent immunity to PSA as evidenced by the increased percentages of CD8+ HLA-A24/PSA(153-161)+ T cells in pre-vaccination samples. All of these patients had shown increased in vitro (IFNγ-based ELISPOT assay) and in vivo (DTH) immunological responses to the AE37 vaccine in the phase I study (Perez SA et al Clin. Cancer Res. 2010, 16:3495). Importantly, this preexistent immunity was significantly boosted in 5 patients during vaccinations with AE37, was slightly increased in 2 others, whereas in one patient no changes were observed during vaccinations. Conclusions: These results suggest that HLA-A24 expression along with preexistent immunity to PSA(153-164) may represent biomarkers based on which HER-2/neu+ patients can be selected most likely to benefit from vaccination with AE37. A phase II trial is warranted for validating these results. Given the promiscuity of AE37, it would be intriguing to examine if preexistent immunity to other prostate antigens restricted by various MHC alleles exist and to demonstrate an increase in preexistent immunity in association with a positive immunologic response to vaccination with AE37.

P1-13-01: An Update of a Phase II Trial of the HER2 Peptide AE37 Vaccine in Breast Cancer Patients To Prevent Recurrence.

Abstract Introduction AE37 is the Ii-Key hybrid of the HER2−derived peptide AE36 (HER2:776–790). A phase I trial administering AE37 with the immunoadjuvant GM-CSF demonstrated the vaccine to be safe and capable of stimulating CD4+helper T-cells with HER2−specific anti-tumor activity. Here we present an update of our prospective, randomized, single-blinded, phase II trial of the AE37 vaccine for the prevention of breast cancer recurrence in disease-free, high risk patients. Methods After completion of standard therapy, disease-free, node positive or high risk node negative breast cancer patients were randomized to receive either AE37+GM-CSF (vaccine) or GM-CSF alone (control) in six monthly intradermal inoculations. Patients were enrolled with any level of HER2 expression, (IHC 1+ 2+ or 3+). Specific immunologic responses to both AE36 and AE37 were evaluated in all patients at pre-determined intervals: before (RO), mid-series (R3), upon completion (R6), and at six (RC6) and 12 (RC12) months after completion of the vaccine series. In vitro responses were measured using the [3H]-thymidine incorporation assay and in vivo responses using delayed-type hypersensitivity (DTH) reactions. The trial's primary endpoint is disease recurrence. Results To date, 215 patients have enrolled (vaccine=92, control=123). 99% of local and systemic toxicities were ≤grade 2 or less. There were no grade 4–5 local or systemic toxicities and no difference between toxicity profiles of vaccine and control groups. Vaccine patients exhibited a statistically significant increase from baseline in AE36 and AE37 proliferative responses at each time point, including maintenance of this response up to 12 months post-vaccination (AE36 (cpm): R0=0, R3=1335, R6=1242, RC6=1586, RC12=1360; AE37: R0=0, R3=2859, R6=2300, RC6=3235, RC12=3279, p<0.001) while there have been no proliferative changes for control patients (AE36: R0=91, R3=95, R6=97, RC6=126, RC12=48; AE37: R0=291, R3=399, R6=319, RC6=103, RC12=0). Vaccine patients also had statistically significant increases in DTH reactions to both AE36 and AE37 (AE36 (mm): R0=0, R6=15, RC6=15, RC12=15; AE37: R0=0, R6=24, RC6=17, RC12=20 p=<0.001) while controls had no response (AE36: R0, R6, RC6, RC12=0; AE37: R0, R6, RC6, RC12=0). With a median follow up of 17 months, breast cancer recurrences were reduced by 42% in vaccine patients compared to control patients (7.6% vs. 13.2%, p=0.15). In an analysis of patients with low HER2 expression (IHC 1 or 2+), vaccine patients experienced a 49% reduction in recurrence compared to controls (9.5% vs. 18.6%, p=0.16) with no reduction seen in HER2 over-expressing patients (6.0% vs. 7.9%, p=0.49). Conclusions The AE37 vaccine is safe and well tolerated with only mild toxicity, which is attributable to the GM-CSF immunoadjuvant. The AE37 vaccine elicits strong peptide specific in-vivo and ex-vivo immune responses, which are maintained for 12 months after completion of the vaccine series. While the number of recurrences are still low, the recurrence rate appears to decrease in vaccinated patients. Administration of the AE37 vaccine may reduce the risk of breast cancer recurrence with the greatest benefit in patients with low levels of HER2 expression. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-13-01.

AE37: a novel T-cell-eliciting vaccine for breast cancer

Introduction:Immunotherapy, including vaccines targeting the human EGFR2 (HER-2/neu) protein, is an active area of investigation in combatting breast cancer. Several vaccines are currently undergoing clinical trials, most of which are CD8+ T-cell-eliciting vaccines. AE37 is a promising primarily CD4+ T-cell-eliciting HER-2/neu breast cancer vaccine currently in clinical trials.Areas covered:This article reviews preclinical investigations as well as findings from completed and ongoing Phase I and Phase II clinical trials of the AE37 vaccine.Expert opinion:Clinical trials have shown the AE37 vaccine to be safe and capable of generating peptide-specific, durable immune responses. This has been shown in patients with any level of HER-2/neu expression. Early clinical findings suggest there may be benefit to AE37 vaccination in preventing breast cancer recurrence.

Abstract LB-413: Circulating regulatory T cells (CD4+CD25high CD127low) decrease in breast cancer patients after vaccination with a modified HER-2/neu HLA class II peptide (AE37) vaccine

Abstract BACKGROUND: Regulatory T cells (TReg) are elevated in breast cancer patients with the number of TReg cells being inversely associated with clinical outcome. We are conducting a prospective, randomized phase II trial of the HER-2/neu peptide vaccines GP2 and AE37. GP2 is an HLA-A2+ -restricted peptide (HER-2/neu aa: 654–662) capable of eliciting a CD8+ cytotoxic T cell response directed against HER-2/neu expressing tumors. AE37 is the Ii-Key hybrid of the HER-2/neu peptide AE36 (HER-2/neu aa:776–790) and is capable of stimulating CD4+ helper T-cells with anti-tumor activity. Given the ability of AE37 to activate CD4+ helper T cells, concern exists that unwanted stimulation of TReg cells may occur with vaccination; although, this was not seen in our phase I trial. We have sought to confirm this finding by evaluating peripheral TReg cell levels in patients from our larger phase II breast cancer vaccine trial. METHODS: Node positive or high-risk node negative breast cancer patients with any level of HER-2/neu expression rendered disease-free by standard treatments were enrolled. HLA-A2+ patients were placed in the GP2 arm and randomized to either GP2 + GM-CSF (immunoadjuvant) or GM-CSF alone. HLA-A2− patients were placed in the AE37 arm and randomized to either AE37 + GM-CSF or GM-CSF alone. Vaccinations were given as six monthly intradermal inoculations. CD4+CD25high CD127low (CD127low) TReg cell levels were measured before (R0), mid-series (R3), upon completion (R6), and at six (RC6) and twelve (RC12) months after completion of the vaccine series using antibody staining and flow cytometry analysis. p values were calculated using the Student's t-test. RESULTS: TReg cells were measured in 65 patients in the AE37 arm (34 vaccine, 31 adjuvant) and 33 patients in the GP2 arm (17 vaccine, 16 adjuvant). Patients receiving the AE37 vaccine exhibited a decrease in CD127low TReg cells from baseline at all measured timepoints (R0=4.2, R3=3.7, p<0.01; R6=3.7, p=0.01; RC6=3.5, p<0.01; RC12=3.7, p<0.00001) while the randomized adjuvant patients demonstrated no decrease at any time point except from R0 to R3 (R0=4.3, R3=3.6, p=0.02; R6=4.1, p=0.58; RC6=3.9, p=0.06; RC12=3.9, p=0.21). In the GP2 arm, CD127low TReg cell levels did not change for either GP2 vaccinated patients (R0=3.6, R3=3.7, p=0.47; R6=4.1, p=0.67; RC6=4.3, p=0.56; RC12=4.3, p=0.16) or in randomized adjuvant patients (R0=4.4, R3=4.0, p=0.58; R6=4.4, p=0.95; RC6=3.8, p=0.25; RC12, p=0.59). CONCLUSIONS: Administration of a CD4 T cell-eliciting HER-2/neu peptide (AE37) vaccine to breast cancer patients induced a sustained decrease in circulating CD127low TReg cells. No such decrease was seen in randomized adjuvant-treated patients. Use of the CD8-eliciting, GP2 vaccine did not change circulating TReg cell levels. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-413. doi:10.1158/1538-7445.AM2011-LB-413

Results from a Phase I Clinical Study of the Novel Ii-Key/HER-2/neu(776–790) Hybrid Peptide Vaccine in Patients with Prostate Cancer

Active immunotherapy is emerging as a potential therapeutic approach for prostate cancer. We conducted the first phase I trial of an Ii-Key/HER-2/neu(776-790) hybrid peptide vaccine (AE37) with recombinant granulocyte macrophage colony-stimulating factor as adjuvant in patients with HER-2/neu(+) prostate cancer. The primary end points of the study were to evaluate toxicity and monitor patients' immune responses to the vaccine. Thirty-two HER-2/neu(+), castrate-sensitive, and castrate-resistant prostate cancer patients were enrolled. Of these, 29 patients completed all six vaccination cycles with AE37. Immunologic responses in the total patient population were monitored by delayed-type hypersensitivity and IFN-gamma ELISPOT and intracellular staining. Regulatory T-cell (Treg) frequency and plasma HER-2/neu and transforming growth factor-beta levels were also determined. Immunologic responses were also analyzed among groups of patients with different clinical characteristics. Local/systemic toxicities were monitored throughout the study. Toxicities beyond grade 2 were not observed. Seventy-five percent of patients developed augmented immunity to the AE37 vaccine and 65% to the unmodified AE36 peptide as detected in the IFN-gamma-based ELISPOT assay. Intracellular IFN-gamma analyses revealed that AE37 elicited both CD4(+) and CD8(+) T-cell responses. Eighty percent of the patients developed a positive delayed-type hypersensitivity reaction to AE36. Additionally, significant decreases could be detected in circulating Treg frequencies, plasma HER-2/neu, and serum transforming growth factor-beta levels. Patients with less extensive disease developed better immunologic responses on vaccination. AE37 vaccine is safe and can induce HER-2/neu-specific cellular immune responses in patients with castrate-sensitive and castrate-resistant prostate cancer, thus emphasizing the potential of AE37 to target HER-2/neu for the immunotherapy of prostate cancer.

S86 Results from the first phase I clinical study of the novel Ii-Key/HER-2(776–790) hybrid peptide vaccine in patients with prostate cancer

Purpose: Active immunotherapy is emerging as a potential therapeutic approach for prostate cancer. We conducted the first phase I trial of an Ii-Key/HER-2/neu(776–790) hybrid peptide vaccine (AE37) with recombinant granulocyte macrophage colony-stimulating factor as adjuvant in patients with HER-2/neu + prostate cancer. The primary end points of the study were to evaluate toxicity and monitor patients' immune responses to the vaccine. Experimental Design: Thirty-two HER-2/neu + , castrate-sensitive, and castrate-resistant prostate cancer patients were enrolled. Of these, 29 patients completed all six vaccination cycles with AE37. Immunologic responses in the total patient population were monitored by delayed-type hypersensitivity and IFN-γ ELISPOT and intracellular staining. Regulatory T-cell (Treg) frequency and plasma HER-2/neu and transforming growth factor-β levels were also determined. Immunologic responses were also analyzed among groups of patients with different clinical characteristics. Local/systemic toxicities were monitored throughout the study. Results: Toxicities beyond grade 2 were not observed. Seventy-five percent of patients developed augmented immunity to the AE37 vaccine and 65% to the unmodified AE36 peptide as detected in the IFN-γ–based ELISPOT assay. Intracellular IFN-γ analyses revealed that AE37 elicited both CD4 + and CD8 + T-cell responses. Eighty percent of the patients developed a positive delayed-type hypersensitivity reaction to AE36. Additionally, significant decreases could be detected in circulating Treg frequencies, plasma HER-2/neu, and serum transforming growth factor-β levels. Patients with less extensive disease developed better immunologic responses on vaccination. Conclusion: AE37 vaccine is safe and can induce HER-2/neu–specific cellular immune responses in patients with castrate-sensitive and castrate-resistant prostate cancer, thus emphasizing the potential of AE37 to target HER-2/neu for the immunotherapy of prostate cancer. Clin Cancer Res; 16(13); 3495–506. ©2010 AACR.

Results from the first phase I clinical study of the novel Ii-Key/HER2/neu(776–790) hybrid peptide vaccine in patients with prostate cancer

3011 Background: HER-2/neu(776–790) represents an immunogenic epitope from the HER-2/neu oncoprotein whose immunogenicity is highly potentiated upon linkage with the Ii-Key moiety (LRMK) from the MHC class II invariant chain. Herein, we present the results of the first phase I clinical trial of the Ii-Key/HER-2/neu(776–790) (AE37) vaccine in patients (pts) with prostate cancer. Methods: Androgen-dependent (AD) and androgen-independent (AI) pts with HER-2/neu+ (IHC: 1–3) prostate adenocarcinomas were eligible. Concomitant medication with bicatulamide and LHRH or docitaxel was allowed. All pts received 6 monthly vaccinations with the AE37 vaccine (500 μg of AE37 plus 125 μg of GM-CSF) administered in two doses intradermally 5cm apart in the same extremity for each vaccination cycle. Immunologic responses were measured monthly in vitro by the IFNγ-based ELISPOT assay using pts’ PBMC and in vivo at the beginning and end of immunizations using DTH. Local dermal reactions were also measured after each vaccination. Results: Eligible pts [AD (n=18), AI (n=10)] were at stages T1–3N0M0, GS: 3–7 (n=18); T1–3N+M0, GS: 6–7 (n=2); T1–3N0M+, GS: 6–9 (n=4) and T1–3N+M+, GS: 6–9 (n=4). All pts had standard treatment prior to vaccinations, including surgery (S) (n=9); hormonal treatment (HT) (n=4); S+HT (n=6); S+HT+radiotherapy (RT); (n=2); S+chemotherapy (CH) (n=2); HT+RT (n=2); CH (n=3) and S+HT+CH (n=2). During vaccinations, 11 pts were free of any treatment, while 5 pts who had progressive disease received additional chemotherapy; the remainder received HT alone or combined with RT. 25 pts completed all vaccinations. Toxicity and side effects beyond grade-2 were not observed. GM-CSF was reduced by 50% for subsequent vaccinations when a local reaction of 100mm or greater was observed. DTH reactions to the parent HER-2/neu(776–790) peptide were increased (compared to pre-vaccination) for all pts, while approximately half the pts responded with significantly increased IFNγ responses to AE37 and/or parent HER-2/neu(776–790) peptide in at least 2 sequential vaccination cycles. Conclusions: The AE37 vaccine is safe and well tolerated. AE37 is also capable of eliciting potent and specific immunologic responses in prostate cancer pts. [Table: see text]

Circulating regulatory (CD4+CD25+FOXP3+) T cells decrease in breast cancer patients after vaccination with an Ii-Key-modified class II HER2/neu peptide (AE37).

Abstract Abstract #3134 Background: CD4+CD25+FOXP3+ regulatory T cells (Tregs) have been implicated in the suppression of immune responses against various tumors. To monitor the potential induction of Tregs in breast cancer (BrCa) patients receiving a modified HLA Class II HER2/neu peptide (AE37) vaccine in a clinical trial, we have analyzed peripheral blood lymphocytes (PBL) from vaccinated patients for the presence of Tregs and correlated our findings with ex vivo immune assays and in vivo delayed type hypersensitivity (DTH) responses to the vaccine. Methods: Fifteen BrCa patients have completed 6 monthly injections of the AE37+GM-CSF vaccine in a dose escalation safety study. The AE37 peptide consists of a HER2/neu peptide (776-790) linked to the Ii-Key moiety of the HLA Class II-associated invariant chain, which enhances epitope interaction with the Class II molecule. PBL obtained pre- and post-vaccination were stained with anti-CD4/CD25 (n=15) and FOXP3 (n=9) antibodies (PCH101 and 236A/E7) and analyzed by flow cytometry. Cells were also stimulated ex vivo with AE37 peptide to measure IFN-γ ELISPOT, proliferation (3H-thymidine-cpm) and cytokine secretion (TGF-β). DTH responses to the AE37 peptide pre- and post-vaccination were also recorded. Results: The mean CD4+ and CD4+CD25+ T cell populations for all patients (n=15) did not change from pre- to post-vaccination (CD4+ = 52.3+3.3% vs. 50.5+3.9%, p=0.6; CD4+CD25+ = 1.9+0.2% vs. 2.4+0.5%, p=0.2). Tregs (CD4+CD25+FOXP3+) were reduced in all 9 patients tested pre- to post-vaccination for both FOXP3 antibodies (Ab) (FOXP3 Ab1 = 2.1+0.2% vs. 1.1+0.1%, p=0.002; FOXP3 Ab2 = 2.0+0.2% vs. 1.0+0.2%, p=0.0009). There was no difference in pre- to post-vaccination levels of TGF-β (2720+582 pg/ml vs. 3387+848 pg/ml; p=0.9). AE37-specific proliferative responses increased from pre- to post-vaccination (34+23cpm vs. 6427+1431 cpm; p<0.001). ELISPOT demonstrated an increased response from pre- to long term (6-12mo.) post-vaccination (Median – 2 vs. 34 spots/106 cells; p=0.003). DTH responses increased in all patients from pre- to post-vaccination (3.6+1.4 mm vs. 56.0+9.4 mm; p<0.0001), and there appeared to be an inverse relationship between the degree of Treg reduction and the size of DTH response to AE37 (R2=0.83).
 Discussion: The novel AE37 HER2/neu peptide vaccine does not result in increased levels of Tregs. Furthermore, the reduced levels of Tregs in vaccinated patients appear to be associated with more robust responses in ex vivo immune assays and in vivo DTH reactions suggesting that the AE37 vaccine may be clinically useful. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 3134.