Efficacy Of Therapeutic Cancer Vaccines Research Articles

Inhibition of DNMTs increases neoantigen-reactive T-cell toxicity against microsatellite-stable colorectal cancer in combination with radiotherapy

The therapeutic efficacy of immunotherapy is limited in the majority of colorectal cancer patients due to the low mutational and neoantigen burdens in this immunogenically “cold” microsatellite stability-colorectal cancer (MSS-CRC) cohort. Here, we showed that DNA methyltransferase (DNMT) inhibition upregulated neoantigen-bearing gene expression in MSS-CRC, resulting in increased neoantigen presentation by MHC class I in tumor cells and leading to increased neoantigen-specific T-cell activation in combination with radiotherapy. The cytotoxicity of neoantigen-reactive T cells (NRTs) to DNMTi-treated cancer cells was highly cytotoxic, and these cells secreted high IFNγ levels targeting MSS-CRC cells after ex vivo expansion of NRTs with DNMTi-treated tumor antigens. Moreover, the therapeutic efficacy of NRTs further increased when NRTs were combined with radiotherapy in vivo. Administration of DNMTi-augmented NRTs and radiotherapy achieved an ∼50 % complete response and extended survival time in an immunocompetent MSS-CRC animal model. Moreover, remarkably, splenocytes from these mice exhibited neoantigen-specific T-cell responses, indicating that radiotherapy in combination with DNMTi-augmented NRTs prolonged and increased neoantigen-specific T-cell toxicity in MSS-CRC patients. In addition, these DNMTi-augmented NRTs markedly increase the therapeutic efficacy of cancer vaccines and immune checkpoint inhibitors (ICIs). These data suggest that a combination of radiotherapy and epi-immunotherapeutic agents improves the function of ex vivo-expanded neoantigen-reactive T cells and increases the tumor-specific cytotoxic effector population to enhance therapeutic efficacy in MSS-CRC.

Engineered polymer‐based scaffold as an implantable therapeutic vaccine against melanoma

AbstractAlthough tumor‐antigen‐based therapeutic cancer vaccines are a potential cancer immunotherapy strategy, recent clinical trials show low efficacy for multiple reasons. One method that has been recently investigated to improve the efficacy of therapeutic cancer vaccines is the development of implantable vaccines for sustained delivery of antigens and CD8 T cell activation. Here, we optimized the composition for an implantable vaccine scaffold composed of alginate, polyvinyl alcohol, and poly(methyl vinyl ether‐ alt‐maleic anhydride) loaded with tumor antigens. Considering the adjuvant property of aluminum compounds, aluminum ion was used to crosslink alginate in the scaffold. The scaffold showed an effective antigen incorporation efficiency of 90.34 ± 0.55% using ovalbumin as the model antigen and 89.67 ± 2.8% using B16‐F10 cell lysate. SEM analysis of the scaffold showed pore size ranging from 5 to 10 μm. Cell viability analysis using mouse RAW 264.7 macrophages proved the cytocompatibility of the scaffold. In vitro antigen release studies using ovalbumin showed 8.42% release for a period of 14 days. In vivo antitumor analysis carried out in subcutaneous mouse B16‐F10 melanoma model demonstrated that the scaffold vaccine reduced the rate of tumor growth and improved survival in tested animals. The median survival time increased from 29 days in untreated animals to 58 days in scaffold vaccine‐implanted animals.

Extracellular vesicles hybrid plasmid-loaded lipid nanovesicles for synergistic cancer immunotherapy

Combination immunotherapy of cancer vaccines with immune checkpoint inhibitors (ICIs) represents a promising therapeutic strategy for immunosuppressed and cold tumors. However, this strategy still faces challenges, including the limited therapeutic efficacy of cancer vaccines and immune-related adverse events associated with systematic delivery of ICIs. Herein, we demonstrate the antitumor immune response induced by outer membrane vesicle from Akkermansia muciniphila (Akk-OMV), which exhibites a favorable safety profile, highlighting the potential application as a natural and biocompatible self-adjuvanting vesicle. Utilizing tumor cell-derived exosome as an antigen source and Akk-OMV as a natural adjuvant, we construct a cancer vaccine formulation of extracellular vesicles hybrid lipid nanovesicles (Lipo@HEV) for enhanced prophylactic and therapeutic vaccination by promoting dendritic cell (DC) maturation in lymph node and activating cytotoxic T cell (CTL) response. The Lipo@HEV is further loaded with plasmid to enable gene therapy-mediated PD-L1 blockade upon peritumoral injection. Meanwhile, it penetrates into lymph node to initiate DC maturation and CTL activation, synergistically inhibiting the established tumor. The fabrication of extracellular vesicles hybrid plasmid-loaded lipid nanovesicles reveals a promising gene therapy-guided and vesicle-based hybrid system for therapeutic cancer vaccination and synergistic immunotherapy strategy.

Skin-Grafting and Dendritic Cell "Boosted" Humanized Mouse Models Allow the Pre-Clinical Evaluation of Therapeutic Cancer Vaccines.

Vaccines have been hailed as one of the most remarkable medical advancements in human history, and their potential for treating cancer by generating or expanding anti-tumor T cells has garnered significant interest in recent years. However, the limited efficacy of therapeutic cancer vaccines in clinical trials can be partially attributed to the inadequacy of current preclinical mouse models in recapitulating the complexities of the human immune system. In this study, we developed two innovative humanized mouse models to assess the immunogenicity and therapeutic effectiveness of vaccines targeting human papillomavirus (HPV16) antigens and delivering tumor antigens to human CD141+ dendritic cells (DCs). Both models were based on the transference of human peripheral blood mononuclear cells (PBMCs) into immunocompromised HLA-A*02-NSG mice (NSG-A2), where the use of fresh PBMCs boosted the engraftment of human cells up to 80%. The dynamics of immune cells in the PBMC-hu-NSG-A2 mice demonstrated that T cells constituted the vast majority of engrafted cells, which progressively expanded over time and retained their responsiveness to ex vivo stimulation. Using the PBMC-hu-NSG-A2 system, we generated a hyperplastic skin graft model expressing the HPV16-E7 oncogene. Remarkably, human cells populated the skin grafts, and upon vaccination with a DNA vaccine encoding an HPV16-E6/E7 protein, rapid rejection targeted to the E7-expressing skin was detected, underscoring the capacity of the model to mount a vaccine-specific response. To overcome the decline in DC numbers observed over time in PBMC-hu-NSG-A2 animals, we augmented the abundance of CD141+ DCs, the specific targets of our tailored nanoemulsions (TNEs), by transferring additional autologous PBMCs pre-treated in vitro with the growth factor Flt3-L. The Flt3-L treatment bolstered CD141+ DC numbers, leading to potent antigen-specific CD4+ and CD8+ T cell responses in vivo, which caused the regression of pre-established triple-negative breast cancer and melanoma tumors following CD141+ DC-targeting TNE vaccination. Notably, using HLA-A*02-matching PBMCs for humanizing NSG-A2 mice resulted in a delayed onset of graft-versus-host disease and enhanced the efficacy of the TNE vaccination compared with the parental NSG strain. In conclusion, we successfully established two humanized mouse models that exhibited strong antigen-specific responses and demonstrated tumor regression following vaccination. These models serve as valuable platforms for assessing the efficacy of therapeutic cancer vaccines targeting HPV16-dysplastic skin and diverse tumor antigens specifically delivered to CD141+ DCs.

Multiparametric flow cytometry to characterize vaccine-induced polyfunctional T cell responses and T cell/NK cell exhaustion and memory phenotypes in mouse immuno-oncology models.

Suitable methods to assess in vivo immunogenicity and therapeutic efficacy of cancer vaccines in preclinical cancer models are critical to overcome current limitations of cancer vaccines and enhance the clinical applicability of this promising immunotherapeutic strategy. In particular, availability of methods allowing the characterization of T cell responses to endogenous tumor antigens is required to assess vaccine potency and improve the antigen formulation. Moreover, multiparametric assays to deeply characterize tumor-induced and therapy-induced immune modulation are relevant to design mechanism-based combination immunotherapies. Here we describe a versatile multiparametric flow cytometry method to assess the polyfunctionality of tumor antigen-specific CD4+ and CD8+ T cell responses based on their production of multiple cytokines after short-term ex vivo restimulation with relevant tumor epitopes of the most common mouse strains. We also report the development and application of two 21-color flow cytometry panels allowing a comprehensive characterization of T cell and natural killer cell exhaustion and memory phenotypes in mice with a particular focus on preclinical cancer models.

Abstract 6799: A porous silicon particle based particulate anti-cancer vaccine for immunotherapy

Abstract Background: Anti-cancer vaccine is a widely-studied strategy that activates innate and adaptive host immunity via its adjuvant components and tumor antigens against cancer. Unfortunately, its response rate in clinical trials was not promising. On the vaccine part, sufficient stimulation of strong and long-lasting local and systemic immunity and efficient promotion of intratumoral infiltration of cytotoxic T lymphocytes are among the critical issues determining the therapeutic efficacy of cancer vaccines in addition to the innate features of tumor. Our project aims to develop a porous silicon particle based particulate vaccine against cancer, and investigate the therapeutic mechanism and combinational effects with current first-line strategies. Method: We first screened the combinations of ligands of Toll-like receptors (TLRs) expressing on antigen presenting cells (APCs) and selected the most powerful combo for immune stimulation. We then loaded the identified adjuvants in addition to antigen peptides to produce vaccine particles. Stimulation and licensing of APCs were tested in vitro, and its therapeutic efficacies were evaluated on multiple orthotopic and systemic metastasis cancer murine models of breast and colon cancers. Furthermore, we investigated the vaccine’s synergistic effects and impacts on tumor microenviroment (TME) with an immunogenic chemotherapeutic oxaliplatin on a CT26 colon cancer model. Result: According to the adjuvant screening assays, we identified a capable combination of CpG1826 (CpG) and 2’3’-cGAMP (cGAMP), which extensively stimulated the activation, maturation and type-I interferon secretion of APCs. A µGCVax formulation composed by these two adjuvants and antigen peptides was then applied on primary and metastatic HER2+ breast cancer TUBO bearing mice. µGCVax potently promoted the activation and lymph node migration of CD8+ and CD103+ dendritic cells in vivo. Distal metastatic tumor nodules in all different tissues vanished after two vaccinations. Functional cure of cancer and extension of animal survival were achieved. Moreover, oxaliplatin treatment on colon cancer caused immunogenic cell death and generated an immune favorable TME in a subcutaneous CT26 colon cancer model. The combination of µGCVax with the oxaliplatin effectively blocked the progression of large tumor mass. We finally revealed that the reprogramming of TME and the promotion of efficacy were mostly achieved from diminishing myeloid-derived suppressor cells (MDSCs) in tumor. Conclusion: µGCVax is an effective and board-spectrum formulation for cancer immunotherapy. It effectively induces APC activation and licensing, and stimulates the innate and adaptive immunity against multiple cancers. The combination with oxaliplatin further promotes the therapeutic efficacy of µGCVax by decreasing tumor MDSCs. Citation Format: Junhua Mai, Yongbin Liu, Dongfang Yu, Haifa Shen. A porous silicon particle based particulate anti-cancer vaccine for immunotherapy. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6799.

Cell membrane-camouflaged liposomes and neopeptide-loaded liposomes with TLR agonist R848 provides a prime and boost strategy for efficient personalized cancer vaccine therapy

Recent advances in bioinformatics and nanotechnology offer great opportunities for personalized cancer vaccine development. However, the timely identification of neoantigens and unsatisfactory efficacy of therapeutic cancer vaccines remain two obstacles for clinical transformation. We propose a “prime and boost” strategy to facilitate neoantigen-based immunotherapy. To prime the immune system, we first constructed personalized liposomes with cancer cell membranes and adjuvant R848 to provide immunostimulatory efficacy and time for identifying tumor antigens. Liposomes loaded with personalized neopeptides and adjuvants were used to boost the immune response. In vitro experiments verified potent immune responses, including macrophage polarization, dendritic cell maturation, and T lymphocyte activation. In vivo B16F10 and TC-1 cancer model were used to investigate efficient tumor growth suppression. Liposomal vaccines with neopeptides could stimulate human dendritic cells and T lymphocytes in vitro. These results demonstrate that the “prime and boost” strategy provides simple, quick, and efficient personalized vaccines for cancer therapy.

In situ phase transitional polymeric vaccines for improved immunotherapy.

Cancer vaccines have exhibited immense potential in cancer treatment. Through activating the host's immune system, vaccines stimulate extensive functional T cells to eliminate cancer. However, the therapeutic efficacy of cancer vaccines is limited by their inferior lymph node delivery and inadequate uptake of dendritic cells. Herein, we propose an in situ phase transitional strategy on vaccine manufacturing to maximally enhance lymph node drainage while ensuring adequate dendritic cell uptake. The phase transitional vaccines, with dynamic size modulation property, retain a small size (24.4 ± 3.1 nm) during lymph node draining then transform into larger particles (483.0 ± 41.6 nm) on-site by external signal input. Results show that this strategy induced rapid and robust immune response in a mouse melanoma tumor model. Furthermore, a stronger humoral immune response was observed in mice when immunized with MHC-II restricted antigen, which demonstrated that lymph node-targeted cancer vaccine delivery could be effectively manipulated through dynamic size modulation.

Selecting Target Antigens for Cancer Vaccine Development.

One of the principal goals of cancer immunotherapy is the development of efficient therapeutic cancer vaccines that are able to elicit an effector as well as memory T cell response specific to tumor antigens. In recent years, the attention has been focused on the personalization of cancer vaccines. However, the efficacy of therapeutic cancer vaccines is still disappointing despite the large number of vaccine strategies targeting different tumors that have been evaluated in recent years. While the preclinical data have frequently shown encouraging results, clinical trials have not provided satisfactory data to date. The main reason for such failures is the complexity of identifying specific target tumor antigens that should be unique or overexpressed only by the tumor cells compared to normal cells. Most of the tumor antigens included in cancer vaccines are non-mutated overexpressed self-antigens, eliciting mainly T cells with low-affinity T cell receptors (TCR) unable to mediate an effective anti-tumor response. In this review, the target tumor antigens employed in recent years in the development of therapeutic cancer vaccine strategies are described, along with potential new classes of tumor antigens such as the human endogenous retroviral elements (HERVs), unconventional antigens, and/or heteroclitic peptides.

Intratumoral immune activation with TLR4 agonist synergizes with effector T cells to eradicate established murine tumors

Effective T cell-based immunotherapy of solid malignancies requires intratumoral activity of cytotoxic T cells and induction of protective immune memory. A major obstacle to intratumoral trafficking and activation of vaccine-primed or adoptively transferred tumor-specific T cells is the immunosuppressive tumor microenvironment (TME), which currently limits the efficacy of both anti-tumor vaccines and adoptive cell therapy (ACT). Combination treatments to overcome TME-mediated immunosuppression are therefore urgently needed. We combined intratumoral administration of the synthetic toll-like receptor 4 agonist glucopyranosyl lipid A (oil-in-water formulation, G100) with either active vaccination or adoptive transfer of tumor-specific CD8 T cells to mice bearing established melanomas or orthotopically inoculated glioblastomas. In combination with cancer vaccines or ACT, G100 significantly increased expression of innate immune genes, infiltration and expansion of activated effector T cells, antigen spreading, and durable immune responses. Complete tumor regression of both injected and non-injected tumors was observed only in mice receiving combination immunotherapy. TLR4-based intratumoral immune activation may be a viable approach to enhance the efficacy of therapeutic cancer vaccines and ACT in patients.

Nanoparticle mediated cancer immunotherapy.

The versatility and nanoscale size have helped nanoparticles (NPs) improve the efficacy of conventional cancer immunotherapy and opened up exciting approaches to combat cancer. This review first outlines the tumor immune evasion and the defensive tumor microenvironment (TME) that hinders the activity of host immune system against tumor. Then, a detailed description on how the NP based strategies have helped improve the efficacy of conventional cancer vaccines and overcome the obstacles led by TME. Sustained and controlled drug delivery, enhanced cross presentation by immune cells, co-encapsulation of adjuvants, inhibition of immune checkpoints and intrinsic adjuvant like properties have aided NPs to improve the therapeutic efficacy of cancer vaccines. Also, NPs have been efficient modulators of TME. In this context, NPs facilitate better penetration of the chemotherapeutic drug by dissolution of the inhibitory meshwork formed by tumor associated cells, blood vessels, soluble mediators and extra cellular matrix in TME. NPs achieve this by suppression, modulation, or reprogramming of the immune cells and other mediators localised in TME. This review further summarizes the applications of NPs used to enhance the efficacy of cancer vaccines and modulate the TME to improve cancer immunotherapy. Finally, the hurdles faced in commercialization and translation to clinic have been discussed and intriguingly, NPs owe great potential to emerge as clinical formulations for cancer immunotherapy in near future.

Abstract A97: The inhibitory receptor NKG2A acts as a checkpoint on CD8 T cells in the context of cancer vaccines

Abstract We previously demonstrated that high HLA-E expression in ovarian carcinoma and NSCLC may neutralize the survival benefit of T-cell infiltration, suggesting NKG2A could represent an immune checkpoint. Here, we demonstrate higher frequencies of NKG2A+CD8+ T cells in TIL samples obtained from human HNSCC biopsies in patients with measurable immune reactivity to HPV16-viral antigens. CyTOF analysis suggested that this T-cell subset harbored an CD103+ effector phenotype. In mouse tumor models, therapeutic cancer vaccines increased the frequency of NKG2A+-positive CD8+ TIL up to 80%. Expression of Qa-1, the mouse homologue of HLA-E, on tumor cells also increased after cancer vaccinations. In vitro studies revealed that upregulation of Qa-1 was mediated by T cell-derived IFNγ. Blockade of this inhibiting NKG2A-Qa-1 axis by antibody improved the T-cell mediated rejection of tumors and, in addition, genetic knockdown of Qa-1 in three different tumor models recapitulated this finding. Blocking of NKG2A as a stand-alone treatment was not effective, indicating that NKG2A constitutes an adaptive resistance mechanism in response to CD8 T-cell activation. In conclusion, NKG2A is enriched on CD8+ TIL and functions as an immune checkpoint that restrains therapeutic efficacy of cancer vaccines. Citation Format: Thorbald van Hall, Nadine van Montfoort, Linda Borst, Michael J. Korrer, Young Kim, Pascale André, Nicolai Wagtmann, Sytse Piersma, Sjoerd H. van der Burg. The inhibitory receptor NKG2A acts as a checkpoint on CD8 T cells in the context of cancer vaccines [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2018 Nov 27-30; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(4 Suppl):Abstract nr A97.

Safety and Efficacy of Therapeutic Cancer Vaccines Alone or in Combination With Immune Checkpoint Inhibitors in Cancer Treatment.

Therapeutic cancer vaccines have proven to seldom induce dramatic clinical response when used alone, and therefore, they are being studied in combination with additional treatment modalities to achieve optimal treatment activities. Growing preclinical data show that combining vaccines and immune checkpoint inhibitors (ICIs) can prime intensified immunogenicity and modulate immunosuppressive tumor microenvironment. Herein, we focus on the safety and efficacy of approved and promising cancer vaccines alone or combined with ICIs in the treatment of several malignancies. Generally, the majority of clinical trials support the concept of synergy that combination therapy of vaccines and ICIs holds maximized potential to improve clinical outcomes. Importantly, the combination has acceptable safety and minimal additional toxicity compared with single-agent vaccines or ICIs. Additionally, the potential strategies of combining personalized tumor vaccines with ICIs will become priority option and future direction of vaccine development and application and the urgent need to develop effective biomarkers to screen appropriate patient populations and predict response to combination therapy.

Co-delivery of immunomodulators in biodegradable nanoparticles improves therapeutic efficacy of cancer vaccines

To improve the efficacy of cancer vaccines we aimed to modulate the suppressive tumor microenvironment. In this study, the potential of intratumoral immune modulation with poly (I:C), Resiquimod (R848) and CCL20 (MIP3α) was explored. Biodegradable polymeric nanoparticles were used as delivery vehicles for slow and sustained release of these drugs in the tumor area and were combined with specific immunotherapy based on therapeutic peptide vaccination in two aggressive murine carcinoma and lymphoma tumor models. Whereas nanoparticle delivery of poly (I:C) or R848 improved therapeutic efficacy, the combination with MIP3α remarkably potentiated the cancer vaccine antitumor effects. The long-term survival increased to 75–100% and the progression free survival nearly doubled on mice with established large carcinoma tumors. The potent adjuvant effects were associated with lymphoid and myeloid population alterations in the tumor and tumor-draining lymph node. In addition to a significant influx of macrophages into the tumor, the phenotype of the suppressor tumor-associated macrophages shifted towards an acute inflammatory phenotype in the tumor-draining lymph node. Overall, these data show that therapeutic cancer vaccines can be potentiated by the combined nanoparticle mediated co-delivery of poly (I:C), R848 and MIP3α, which indicates that a more favorable milieu for cancer fighting immune cells is created for T cells induced by therapeutic cancer vaccines.

Combination of Sunitinib and PD-L1 Blockade Enhances Anticancer Efficacy of TLR7/8 Agonist-Based Nanovaccine.

Cancer vaccines composed of tumor-associated antigens (TAAs) and toll-like receptor (TLR) agonists have shown promising antitumor efficacy in preclinical studies by generating antigen-specific CD8 T cells, but translation of cancer vaccines to the clinic has been limited due to variables responses and development of resistance. The tumor microenvironment deploys various immune escape mechanisms that neutralize CD8 T cell-mediated tumor rejection. Therefore, we hypothesized that modulation of the tumor microenvironment can augment CD8 T cell activation and enhance therapeutic efficacy of cancer vaccines. To accomplish this, we aimed to eliminate immune suppressive cells and block their inhibitory signaling. Combination of the tyrosine kinase inhibitor (TKI) sunitinib with a nanoparticle-based cancer vaccine (nanovaccine) resulted in the reduction of immune-suppressive myeloid-derived suppressive cells (MDSCs) and regulatory T cells (Tregs). Blockade of programmed death-ligand 1 (PD-L1) using anti-PD-L1 antibody was used to reduce CD8 T cell exhaustion. Combination of nanovaccine+sunitinib+PD-L1 antibody treatment reduced PD-L1high M2 macrophages and MDSCs and upregulated activation of CD8 T cells in the tumor. Nanovaccine+sunitinib+PD-L1 antibody treatment also stimulated antigen-specific CD8 T cell response, which led to improved therapeutic efficacy in MB49 and B16F10 murine tumor models. These results suggest that modulation of tumor microenvironment using sunitinib and PD-L1 blockade can significantly enhance the antitumor efficacy of cancer nanovaccine.

Abstract 4936: Combination CD40 agonist and PD-1 antagonist antibody therapy enhances vaccine induced T cell responses in non-immunogenic cancers

Abstract A hallmark of many non-immunogenic cancers is the lack of tumor infiltrating lymphocytes (TIL) and/or failure to mount a robust anti-tumor T cell response via multiple mechanisms. The presence of T regulatory cells and myeloid derived suppressor cells (MDSCs) serve to dampen the immune response, and furthermore, tumor antigen-specific T cell tolerance limits the efficacy of therapeutic cancer vaccines. CD40 signaling is critical to the decision of whether cytotoxic T lymphocytes become primed or tolerized. Administration of monoclonal CD40 agonistic antibody (Ab) has been shown to promote CD8 activation in vivo, and likely alters the myeloid component of the tumor microenvironment. Our study asks the question of whether combining a T cell inducing vaccine and PD1 inhibition with CD40 agonistic Ab can induce T cell priming and TIL activation in non-immunogenic solid malignancies. We utilized mouse models of pancreatic ductal adenocarcinoma (PDAC) and breast cancer to assess the effects of drug combinations on intratumoral immune responses. Tumor-bearing mice were treated with a GM-CSF secreting vaccine (GVAX) + anti-PD1 Ab alone or in combination with CD40 Ab, or isotype control Ab, and monitored for survival. A separate cohort of mice were analyzed by immunohistochemistry and multi-color flow cytometry to assess T cell infiltration/activation and myeloid maturation. In a hemisplenectomy model of PDAC in which tumor cells were surgically implanted into wild-type recipients, mice treated with isotype control Abs succumbed to disease with extensive liver and peritoneal metastases at 35-70 days. GVAX + anti-PD1 Ab treatment displayed some efficacy, although 70% of mice eventually developed fatal liver metastases. In contrast, CD40 Ab was highly active, with 90% long-term survival afforded by a single administration of Ab, and mice treated with GVAX + anti-PD1 Ab + CD40 Ab had 100% survival. Similar trends in treatment efficacy were observed following subcutaneous tumor implantation of PDAC tumor cells in the lower limb. In an orthotopic model in which HER2/neu-expressing breast tumor cells were implanted into the mammary fat pad of syngeneic neu-N mice, we demonstrated delayed tumor progression and increased median survival in mice treated with GVAX + anti-PD1 Ab + CD40 Ab relative to either therapy alone. Further characterization of immune populations was carried out by high dimensional flow cytometric analysis utilizing PhenoGraph clustering and visualized by t-SNE. Changes were observed in monocytic and dendritic cell infiltration and maturation in the tumors of combination-treated mice. A significant decrease in granulocytic MDSCs was associated with response, as well as an increase in mature antigen presenting cells. In conclusion, GVAX, anti-PD1 and CD40 agonist Ab have potential synergy in modulating anti-tumor immunity in non-immunogenic cancers. Citation Format: Hayley S. Ma, Evanthia Roussos Torres, Bibhav Poudel, Tara Robinson, Brian Christmas, Kayla Cruz, Skylar Woolman, Christine Rafie, Blake Scott, Valerie Wall, Todd Armstrong, Elizabeth Jaffee. Combination CD40 agonist and PD-1 antagonist antibody therapy enhances vaccine induced T cell responses in non-immunogenic cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4936.

Abstract 2613: Combination agonist and antagonist antibody therapy enhances vaccine induced T cell responses in non-immunogenic cancers

Abstract Immune tolerance by multiple mechanisms is a major obstacle in the implementation of successful immunotherapy. T regulatory cells (Tregs) and myeloid derived suppressor cells (MDSCs) suppress the immune response, and the presence of extensive stromal networks within the tumor microenvironment (TME) can prevent T cell infiltration in many tumor types. Furthermore, tumor antigen-specific T cell tolerance limits the efficacy of therapeutic cancer vaccines. CD40 is expressed on antigen presenting cells, and CD40 signaling is critical to the decision of whether cytotoxic T lymphocytes become primed or tolerized. Administration of monoclonal CD40 agonistic antibody (Ab) has been shown to promote CD8 activation in vivo, and likely alters the myeloid component of the TME. Our study asks the question of whether combining a T cell inducing vaccine and PD-1 inhibition with CD40 agonistic Ab can induce T cell priming and tumor infiltrating lymphocyte (TIL) activation in non-immunogenic solid malignancies. We utilized an orthotopic transplant model in which HER2/neu-expressing breast tumor cells were implanted into the mammary fat pad of syngeneic neu-N mice to assess the effects of drug combinations on intratumoral immune responses. Tumor-bearing mice were treated with a granulocyte macrophage colony-stimulation factor secreting vaccine (GVAX) + anti-PD-1 Ab alone or in combination with CD40 agonistic Ab, and monitored for tumor clearance and survival. A separate cohort of mice were analyzed by IHC and multi-color flow cytometry to assess T cell infiltration and activation, myeloid maturation, and MDSCs after one week of treatment. We demonstrated delayed tumor progression and increased median survival in mice treated with GVAX + anti-PD-1 Ab + CD40 Ab relative to CD40 Ab alone, although endogenous T cell infiltration remained low across treatment groups. However, adoptive transfer of neu-specific TCR transgenic cells revealed 100% tumor clearance in the combination-treated mice, along with significant T cell infiltration and activation as measured by IFNγ, Granzyme B and TNFα-secreting CD8+ T cells. In contrast, only 20% of mice treated with GVAX + anti-PD-1 Ab were able to clear tumor, and none of the mice receiving vehicle or CD40 Ab alone had long-term survival. Trends were also observed in monocytic and dendritic cell infiltration and maturation in the tumors of combination-treated mice, and warrant further investigation. In conclusion, GVAX, anti-PD-1 Ab and CD40 agonist Ab have potential synergy in modulating anti-tumor immunity in breast cancer. These results support further studies more broadly in non-immunogenic solid tumors. Citation Format: Hayley S. Ma, Evanthia Roussos Torres, Brian Christmas, Blake Scott, Tara Robinson, Todd Armstrong, Elizabeth Jaffee. Combination agonist and antagonist antibody therapy enhances vaccine induced T cell responses in non-immunogenic cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2613. doi:10.1158/1538-7445.AM2017-2613

Antitumor effect of a whole tumor cell vaccine expressing human fibroblast activation protein in murine tumor models.

e14542 Background: Immunosuppressive factors in the tumor microenvironment reduce the therapeutic efficacy of cancer vaccines; therefore, dampening the tumor immunosuppressive environment while activating antitumor immunity should be a useful approach for cancer therapy. Cancer-associated fibroblasts (CAFs) are among the most important cellular components of the tumor microenvironment and play an important role in the development and progression of tumors. Fibroblast activation protein α (FAPα) is a type II transmembrane protein specifically expressed by CAFs in most epithelial cancers. Gene sequence homology between human and murine FAPα is 90%. Heterogeneity of genes in the evolution of the formation of such nuances can be utilized to break immune tolerance, enhance immunogenicity, induce tumour cell autoimmune responses and thus inhibit tumor growth. Methods: Tumor cells were transfected with human FAPα plasmids employing the cationic lipid DOTAP. Its antitumor effects were investigated in three established tumor models. The potential immune mechanisms were investigated through adoptive immunotherapy and 51Cr release assay. The distributions of the immune cells in the tumor microenvironment were detected by immunohistochemical staining and flow cytometry. Results: Our results shown that whole tumor cell vaccine expressing human FAPα significantly inhibit tumor growth and prolong the survival of tumor bearing mice. This antitumor immune response was involved both of cellular and humoral immune responses. FAPα specific neutralizing antibodies were found in the serum of vaccinated mice and CAFs were significantly reduced within the tumors. Furthermore, this vaccine enhanced the infiltration of CD4+ T cells and CD8+ T cells, and suppressed the accumulation of immunosuppressive cells in the tumor microenvironment. Conclusions: These findings suggest that whole tumor cell vaccine expressing human FAPα inhibit tumor growth by producing FAPα specific neutralizing antibodies and CTLs, and targeting tumor cells and CAFs. These observations provide a new strategy for the clinical use of genetically modified tumor cells as cancer vaccines.

A Bayesian adaptive phase I-II trial design for optimizing the schedule of therapeutic cancer vaccines.

Phase I-II clinical trials refer to the class of designs that evaluate both the safety and efficacy of a novel therapeutic agent in a single trial. Typically, Phase I-II oncology trials take the form of dose-escalation studies, where initial subjects are treated at the lowest dose level and subsequent subjects are treated at progressively higher doses until the optimal dose is identified. While dose-escalation designs are well-motivated in the case of traditional chemotherapeutic agents, an alternate approach may be considered for therapeutic cancer vaccines, where an investigator's main objective is to evaluate the safety and efficacy of a set of dosing schedules or adjuvant combinations rather than to compare the safety and efficacy of progressively higher dose levels. We present a two-stage, Bayesian adaptive Phase I-II trial design to evaluate the safety and efficacy of therapeutic cancer vaccines. In the first stage, we determine whether a vaccination schedule achieves a minimum level of performance by comparing the toxicity and immune response rates to historical benchmarks. Vaccination schedules that achieve a minimum level of performance are compared using their magnitudes of immune response. If the superiority of a single schedule cannot be established after the first stage, Bayesian posterior predictive probabilities are used to determine the additional sample size required to identify the optimal vaccination schedule in a second stage. Copyright © 2016 John Wiley & Sons, Ltd.

B cells promote tumor progression in a mouse model of HPV-mediated cervical cancer.

Enhancing anti-tumor immunity and preventing tumor escape are efficient strategies to increase the efficacy of therapeutic cancer vaccines. However, the treatment of advanced tumors remains difficult, mainly due to the immunosuppressive tumor microenvironment. Regulatory T cells and myeloid-derived suppressor cells have been extensively studied, and their role in suppressing tumor immunity is now well established. In contrast, the role of B lymphocytes in tumor immunity remains unclear because B cells can promote tumor immunity or display regulatory functions to control excessive inflammation, mainly through IL-10 secretion. Here, in a mouse model of HPV-related cancer, we demonstrate that B cells accumulated in the draining lymph node of tumor-bearing mice, due to a prolonged survival, and showed a decreased expression of MHC class II and CD86 molecules and an increased expression of Ly6A/E, PD-L1 and CD39, suggesting potential immunoregulatory properties. However, B cells from tumor-bearing mice did not show an increased ability to secrete IL-10 and a deficiency in IL-10 production did not impair tumor growth. In contrast, in B cell-deficient μMT mice, tumor rejection occurred due to a strong T cell-dependent anti-tumor response. Genetic analysis based on single nucleotide polymorphisms identified genetic variants associated with tumor rejection in μMT mice, which could potentially affect reactive oxygen species production and NK cell activity. Our results demonstrate that B cells play a detrimental role in anti-tumor immunity and suggest that targeting B cells could enhance the anti-tumor response and improve the efficacy of therapeutic cancer vaccines.