Keith B. Gorden

Oligodeoxynucleotides Differentially Modulate Activation of TLR7 and TLR8 by Imidazoquinolines

Among the 11 human TLRs, a subfamily TLR7, TLR8, and TLR9 display similarities in structure and endosomal localization. Natural agonists consisting of nucleic acids, such as ssRNA or DNA with CpG motifs, activate the innate immune cells through these TLRs. Immune response modifiers (IRMs) of imidazoquinoline class compounds 3M-001, 3M-002, and 3M-003 have been shown to activate the innate immune system via TLR7, TLR8, and TLR7/8, respectively. In looking at the effect of the agonists of the TLR7, TLR8, and TLR9 on the activation of NF-κB of transfected HEK cells, we discovered that some oligodeoxynucleotides (ODNs) could modulate imidazoquinoline effects in a negative or positive manner. In this study we demonstrate that poly(T) ODNs can inhibit TLR7 and enhance TLR8 signaling events involving NF-κB activation in HEK cells and cytokine production (IFN-α, TNF, and IL-12) by human primary PBMC. In contrast, TLR3 agonist poly(I:C) does not affect imidazoquinoline-induced responses. The modulation of TLR7 and TLR8 responses is independent of CpG motifs or the nature of the ODN backbone structure. Furthermore, we show that to be an effective modulator, the ODNs need to be in the cell at the same time with either of the TLR7 or TLR8 agonist. We have also demonstrated that there is a physical interaction between IRMs and ODNs. The cross-talk between ODNs, IRMs, and TLR7 and TLR8 uncovered by this study may have practical implications in the field of microbial infections, vaccination, and tumor therapy.

Binding of Soluble Yeast β-Glucan to Human Neutrophils and Monocytes is Complement-Dependent.

The immunomodulatory properties of yeast β-1,3/1,6 glucans are mediated through their ability to be recognized by human innate immune cells. While several studies have investigated binding of opsonized and unopsonized particulate β-glucans to human immune cells mainly via complement receptor 3 (CR3) or Dectin-1, few have focused on understanding the binding characteristics of soluble β-glucans. Using a well-characterized, pharmaceutical-grade, soluble yeast β-glucan, this study evaluated and characterized the binding of soluble β-glucan to human neutrophils and monocytes. The results demonstrated that soluble β-glucan bound to both human neutrophils and monocytes in a concentration-dependent and receptor-specific manner. Antibodies blocking the CD11b and CD18 chains of CR3 significantly inhibited binding to both cell types, establishing CR3 as the key receptor recognizing the soluble β-glucan in these cells. Binding of soluble β-glucan to human neutrophils and monocytes required serum and was also dependent on incubation time and temperature, strongly suggesting that binding was complement-mediated. Indeed, binding was reduced in heat-inactivated serum, or in serum treated with methylamine or in serum reacted with the C3-specific inhibitor compstatin. Opsonization of soluble β-glucan was demonstrated by detection of iC3b, the complement opsonin on β-glucan-bound cells, as well as by the direct binding of iC3b to β-glucan in the absence of cells. Binding of β-glucan to cells was partially inhibited by blockade of the alternative pathway of complement, suggesting that the C3 activation amplification step mediated by this pathway also contributed to binding.

Imprime PGG-Mediated Anti-Cancer Immune Activation Requires Immune Complex Formation

Imprime PGG (Imprime), an intravenously-administered, soluble β-glucan, has shown compelling efficacy in multiple phase 2 clinical trials with tumor targeting or anti-angiogenic antibodies. Mechanistically, Imprime acts as pathogen-associated molecular pattern (PAMP) directly activating innate immune effector cells, triggering a coordinated anti-cancer immune response. Herein, using whole blood from healthy human subjects, we show that Imprime-induced anti-cancer functionality is dependent on immune complex formation with naturally-occurring, anti-β glucan antibodies (ABA). The formation of Imprime-ABA complexes activates complement, primarily via the classical complement pathway, and is opsonized by iC3b. Immune complex binding depends upon Complement Receptor 3 and Fcg Receptor IIa, eliciting phenotypic activation of, and enhanced chemokine production by, neutrophils and monocytes, enabling these effector cells to kill antibody-opsonized tumor cells via the generation of reactive oxygen species and antibody-dependent cellular phagocytosis. Importantly, these innate immune cell changes were not evident in subjects with low ABA levels but could be rescued with exogenous ABA supplementation. Together, these data indicate that pre-existing ABA are essential for Imprime-mediated anti-cancer immune activation and suggest that pre-treatment ABA levels may provide a plausible patient selection biomarker to delineate patients most likely to benefit from Imprime-based therapy.

Abstract 5034: Imprime PGG decreases regulatory T cell suppression and enhances T cell proliferation and differentiation revealing additional mechanisms for its anti-tumor activity

Imprime PGG is a soluble yeast-derived β-glucan immunomodulator being developed for treatment of cancer in combination with anti-tumor antibodies (Abs). Imprime PGG binds to complement receptor (CR)3 on innate immune cells (neutrophils and monocytes), and primes them to exert anti-tumor activity against Ab-targeted, complement opsonized tumor cells. Numerous studies in syngeneic and human xenogeneic tumor models in mice have demonstrated that administration of Imprime PGG in combination with anti-tumor Ab treatment reduces tumor growth and prolongs survival beyond that observed with either agent alone. In clinical trials, Imprime PGG in combination with tumor-targeting Abs has also been associated with improvements in objective tumor response and survival. Imprime PGG9s mechanism of action has been extensively studied in the context of the innate immune system. Whether Imprime PGG may also play a role in the modulation of adaptive immune responses and the cross-talk that exists between the innate immune system and T-cell responses was unclear. Recent work has revealed that Imprime PGG is capable of activating not only the innate immune system but also has the potential of inhibiting the polarization of macrophages into the immunosuppressive M2 phenotype. Since considerable cross-talk exists between the innate and adaptive immune cells in the immunosuppressive tumor microenvironment, we investigated whether Imprime PGG may also play a role in direct or indirect modulation of one of the adaptive immunosuppressive cell types, regulatory T cells (Tregs). We demonstrated that plasma from Imprime PGG treated whole blood (WB) was able to decrease the suppressive capability of Tregs on conventional CD4+ T cells. We also determined that plasma from Imprime PGG treated WB was able to enhance the proliferation of both CD4+ and CD8+ T cells in response to CD3/28 beads and to peptide stimulation in vitro. This enhanced proliferation was accompanied by upregulation of the transcription factor Tbet and increased IFN-γ production suggesting Imprime PGG may drive polarization of T cells to a Th1, anti-tumor phenotype. Interestingly, these effects were greatest when Imprime PGG was added to WB and not to isolated plasma, indicating that the effects of Imprime PGG on the T cell compartment are indirect and may involve cross-talk from the innate immune system. These data are the first to show that Imprime PGG may exert anti-tumor activity not only by re-directing the innate immune system against opsonized tumor cells, but also by re-orienting the adaptive immune compartment toward a Th1, anti-tumor phenotype. Citation Format: Steven M. Leonardo, Keith Gorden, Ross Fulton, Lindsay Wurst. Imprime PGG decreases regulatory T cell suppression and enhances T cell proliferation and differentiation revealing additional mechanisms for its anti-tumor activity. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5034. doi:10.1158/1538-7445.AM2015-5034

Abstract B019: Imprime PGG, a soluble β-glucan, binds to and activates dendritic cells resulting in enhanced T cell priming, expansion, and cytokine production

Imprime PGG (Imprime), a soluble yeast 1,3/1,6 β-glucan, is being developed as a novel cancer immunotherapy in conjunction with anti-tumor antibodies in several cancers. Randomized Phase 2 clinical trials of Imprime in the 1st-line treatment of stage IV non-small cell lung cancer have shown promising efficacy in both objective tumor response and survival. Mechanistic work has revealed that Imprime acts to stimulate a coordinated, anti-cancer immune response in conjunction with antibody therapy. Imprime represents a pathogen associated molecular pattern (PAMP) and, as such, can be efficiently and effectively recognized by cells of the innate immune system, triggering macrophage repolarization, neutrophil activation, monocyte-derived dendritic cell maturation, NK cell activation and, via cross-talk with the adaptive immune system, expansion of CD4 and CD8 T cells. As PAMPs have been shown to stimulate dendritic cells (DCs), which are critical for generating robust and durable anti-tumor responses, we sought to better understand the effects of Imprime on DCs. Here we demonstrate that Imprime binds to various DC subsets, stimulating the critical antigen presenting functional activity of these DC subsets. Following i.v. administration in mice, Imprime bound both classical splenic DCs, including CD8α+ cross-presenting DCs, and migratory DC subsets within peripheral lymph nodes. In vitro treatment of human whole blood showed binding in the inflammatory DC subset (Lin-HLA-DR+CD11c+CD16+) as well as the classical (Lin-HLA-DR+CD11c+CD1c+) DC subset. Binding of Imprime to both mouse and human DCs resulted in the upregulation of MHC class II and the co-stimulatory molecules CD80/86 that are critical for antigen presentation and T cell activation. Furthermore, in vivo treatment of mice with Imprime in combination with H-2Kb-restricted OVA257-264 peptide resulted in enhanced expansion of adoptively transferred OVA-specific OT-I CD8 T cells. In contrast to mice immunized with OVA peptide alone, which did not generate a functional CD8 T cell response, Imprime co-administration resulted in functional OT-I capable of ex vivo degranulation and production of the cytokines IFN-γ and IL-2. These data suggest that, in addition to previous studies showing Imprime primes monocytes, macrophages, and neutrophils, Imprime also enhances T cell activation and expansion by directly stimulating dendritic cell maturation and efficient antigen presentation. These data demonstrate that Imprime PGG treatment may enhance the adaptive immune response necessary for durable tumor control and, together with previously published data, indicate that Imprime PGG treatment triggers an orchestrated anti-cancer immune response involving both the innate and adaptive immune systems. Citation Format: Ross B. Fulton, Steven M. Leonardo, Kyle S. Michel, Michael E. Danielson, Keith B. Gorden, Jeremy R. Graff. Imprime PGG, a soluble β-glucan, binds to and activates dendritic cells resulting in enhanced T cell priming, expansion, and cytokine production. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B019.

Abstract A160: Imprime PGG binds to neutrophils through complement, Fc, and dectin-1 receptors, priming these cells for enhanced ROS production and tumor cell cytotoxicity

Imprime PGG (Imprime), a soluble yeast 1,3/1,6 β-glucan, is being developed as a novel cancer immunotherapy in conjunction with anti-tumor antibodies in several cancers. In clinical studies, including randomized Phase 2 clinical trials in the 1st-line treatment of stage IV non-small cell lung cancer with bevacizumab, Imprime treatment has shown promising efficacy in both objective tumor response rates and survival. In numerous syngeneic and xenogeneic mouse tumor models, Imprime treatment in combination with an anti-tumor antibody reduced tumor growth and prolonged survival beyond that observed with antibody alone. Mechanistic studies have now demonstrated that, with endogenous anti-β-glucan antibodies (ABA), Imprime forms an immune complex. This Imprime-ABA complex then activates the classical complement pathway and is subsequently opsonized by iC3b. This immune complex then binds to and primes innate immune cells, including macrophages, monocytes and neutrophils, triggering a coordinated immune attack against antibody-targeted cancer cells. In this study, we sought to explore more fully the functional consequences of Imprime-ABA immune complex binding to, and priming of, neutrophils. Neutrophils are the first line of defense against fungal infections. Neutrophils detect fungal infections through recognition of β-glucans on the surface of yeast and fungi. These β-glucans serve as fungal pathogen associated molecular patterns (PAMPs) that are efficiently and effectively recognized by receptors on innate immune cells, initiating a coordinated immune response to infection. Here we demonstrate that Imprime, in individuals with high ABA levels, is recognized as a PAMP, directly interacting with neutrophils via multiple receptors, priming these cells and ultimately leading to enhanced effector responses including reactive oxygen species (ROS) generation and tumor cell killing. Our data now show that Imprime elicits enhanced neutrophil survival as measured by cell viability dye exclusion using flow cytometry. Imprime treatment also enhances the responsiveness of neutrophils to Fc receptor activation. As measured by luminol-based read-outs, ROS production in response to anti-Fc receptor antibody-coated beads is profoundly enhanced by Imprime treatment. Moreover, Imprime treated neutrophils show a remarkable surge in ROS production compared to vehicle–treated neutrophils when exposed to B cell lymphoma cells decorated with anti-CD20 monoclonal antibodies (i.e. rituximab). These Imprime-treated neutrophils also exhibit a robust increase in cytotoxicity against these B cell lymphoma cells coated with rituximab using in vitro Calcein AM dye release assays. We next sought to delineate which receptors might be most critical for Imprime binding and priming of neutrophils. As demonstrated by antibody blocking studies, the Imprime-ABA complex can bind to both complement receptors (notably CR3) and Fc receptors. Imprime can also block binding of a dectin-1 agonizing antibody to dectin-1 and can bind directly to HL-60 and HEK cells lines engineered to overexpress dectin-1 by retroviral transduction. Collectively, these data show for the first time that Imprime directly primes neutrophils to recognize and kill antibody-decorated tumor cells and interacts with the Fc, complement and dectin-1 receptors on the surface of neutrophils. Citation Format: Steven M. Leonardo, Ross B. Fulton, Lindsay R. Wurst, Keith B. Gorden, Adria Bykowski Jonas, Xiaohong Qui, Anissa SH Chan, Jeremy R. Graff. Imprime PGG binds to neutrophils through complement, Fc, and dectin-1 receptors, priming these cells for enhanced ROS production and tumor cell cytotoxicity. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr A160.

Abstract LB-080: Imprime PGG, a β-glucan PAMP (pathogen-associated molecular pattern) activates the direct killing functions of innate immune cells in concert with tumor targeting antibodies

Imprime PGG (Imprime) is a soluble yeast 1,3/1,6 β-glucan PAMP (pathogen-associated molecular pattern). As a PAMP, Imprime triggers innate immune function, activating the direct killing functions of innate immune cells, facilitating MDSC differentiation and macrophage repolarization as well as enabling dendritic cell maturation and antigen presentation, driving T cell expansion and activation. In the clinic, Imprime is administered intravenously and is well-tolerated. In multiple clinical trials (> 400 subjects), including randomized phase 2 studies in NSCLC, Imprime has consistently shown promising increases in both objective tumor response and patient survival. To date, the clinical experience with Imprime has centered on combinations with tumor targeting monoclonal antibodies (Mabs). For instance, Imprime combined with rituximab and alemtuzumab in CLL patients yielded a 65% complete response rate (vs 37% historical CR rate for alemtuzumab plus rituximab). We sought to better characterize the effect of Imprime in concert with tumor-targeting mAbs. We show that Imprime enhances the effector functions of multiple innate immune cell lineages. We first evaluated the generation of Reactive Oxygen Species (ROS) in neutrophils isolated from human healthy volunteer whole blood. These neutrophils, but not those from vehicle treated whole blood, specifically recognized B cell lymphomas (Raji) only after opsonization with anti-CD20 Mabs (rituximab, ofatumumab, obinatuzumab), generating a substantial ROS burst that coincided with enhanced tumor cell cytotoxicity. Similarly, increased antibody dependent cellular phagocytosis (ADCP) mediated by monocyte-derived macrophages was evident against antibody-opsonized lymphomas (Z138 B cell lymphomas with obinutuzumab) and solid tumor cells (SKBr3 breast cancer cells with trastuzumab) from Imprime-treated whole blood. Likewise, increased Natural Killer (NK) cell-mediated antibody dependent cellular cytotoxicity (ADCC) was evident only after Imprime treatment against antibody-opsonized cancer cells (SKBR3 with trastuzumab, K562 erythroleukemia cells with anti-glycophorin-A). In vivo, we now show that Imprime administered intravenously significantly enhances the anti-tumor efficacy of trastuzumab in a patient derived xenograft model of breast cancer, reducing mean tumor volume to ∼ 50% of that achieved by trastuzumab alone. In the B16 lung experimental metastasis model, the addition of Imprime to the anti-TRP1 tumor targeting antibody TA-99 significantly reduces both the number and size of B16 lung metastases. Together, these data show that Imprime stimulates the innate immune system, augmenting the anti-tumor efficacy of a diverse array of tumor targeting antibodies in multiple tumor types. Citation Format: Steven M. Leonardo, Ross B. Fulton, Keith B. Gorden, Katy Fraser, Ben Harrison, Takashi Kangas, Adria Jonas, Yumi Yokoyama, Nadine Ottoson, Nandita Bose, Jeremy R. Graff. Imprime PGG, a β-glucan PAMP (pathogen-associated molecular pattern) activates the direct killing functions of innate immune cells in concert with tumor targeting antibodies. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-080.

Abstract 2834: Identification of a critical level of anti-beta glucan IgG antibody necessary for response to soluble beta-glucan therapy and its application as a biomarker for analysis in clinical trials

Soluble beta-glucan is a pathogen associated molecular pattern that can activate the innate immune system and has been shown to enhance anti-tumor antibody function in animal models and human clinical trials. Imprime-PGG is yeast derived soluble beta-glucan that is being developed as an immunotherapuetic drug for administration with approved monoclonal anti-tumor antibodies. We have previously demonstrated that in vitro binding and functional response to Imprime-PGG in whole blood correlates with the level of anti-beta-glucan antibodies (ABA) as determined by ELISA. In donors with high ABA levels, stimulation with Imprime-PGG corresponds with higher levels of complement cascade activation, neutrophil binding, modulation of surface markers and IL-8 production. Additionally we have shown that when intravenous immunoglobulin preparations (IVIG) containing high levels of ABA is added to a whole blood sample, it increases the binding and functional response to Imprime-PGG in vitro. Based on these data, we hypothesized that the level of ABA in patient serum could serve as a potential biomarker predictive of positive clinical outcome. The objective of this study was to determine a range of IgG anti-β glucan at which an individual would be considered to be biomarker positive with regards to being responsive to Imprime-PGG treatment. A panel of 8 donors was selected with low IgG ABA and the levels of ABA were increased by the addition of IVIG to whole blood samples at 2, 5 and 10 mg/mL final concentrations. Significant differences were observed in Imprime-PGG activation with increasing IgG levels in complement cascade activation as measured by C5b-9, neutrophil binding of Imprime-PGG, surface marker modulation and IL-8 expression. Addition of IVIG to 2.5mg/mL showed a difference in complement cascade activation where a level of 5mg/mL was necessary to see a significant difference in the other endpoints measured. Analyzing previous data in healthy donors, this ABA range predicted individuals responsive to Imprime-PGG stimulation. Analysis of clinical trial patient data, segregating individuals into a biomarker positive group based on this range of IgG ABA showed a significant survival benefit over the biomarker negative population. These results confirm that the level of ABA is critical to Imprime-PGG response and provides evidence for an IgG ABA range where response to stimulation occurs that can be used a clinical biomarker. They also provide evidence that in a clinical setting, addition of exogenous ABA could provide a benefit to patients with low ABA levels receiving Imprime-PGG treatment. Citation Format: Keith Gorden. Identification of a critical level of anti-beta glucan IgG antibody necessary for response to soluble beta-glucan therapy and its application as a biomarker for analysis in clinical trials. [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 2834. doi:10.1158/1538-7445.AM2014-2834

Abstract 3767: Imprime PGG, a soluble yeast b-glucan PAMP, enhancement of anti-tumor responses in combination with tumor targeting antibody is highly dependent on NK cell killing

Cancer therapy has been reshaped by checkpoint inhibitors (CPIs), making it possible for durable responses against cancers with traditionally low cure rates. Current efforts are focused on combination therapies in the hopes of evading resistance to CPIs and improving overall response. One escape mechanism attributed to acquired resistance to CPIs includes defective antigen presentation, namely a loss in MHC class I expression. This leads to loss of CD8 T cell-mediated tumor killing and disease relapse. This recent revelation has stimulated a need for therapies that activate other cytotoxic effector cells such as NK cells to kill tumors. Imprime PGG (Imprime) is a soluble, systemically delivered yeast 1,3/1,6 β-glucan PAMP (pathogen-associated molecular pattern) capable of triggering innate immune cell function leading to a cascade of immune activation and enhanced tumor killing. Imprime activates the innate immune system via dectin-1, eliciting production of a variety of chemokines and cytokines, including type I IFN, leading to the mobilization and stimulation of innate cell types including dendritic cells and monocytes. Unlike other PAMPs which systemic administration often leads to toxic side effects, Imprime has been administered safely by intravenous infusion to >400 human subjects. Currently, Imprime PGG is being evaluated in combination with αPD1 therapy in multiple clinical trials. Previously we have shown that combination therapy of anti-Trp1 antibody and Imprime leads to a significant reduction in both number and size of lung metastases in the B16F10 metastatic melanoma tumor model over anti-Trp1 antibody alone. This reduction of metastases is highly dependent on NK cells but not CD8 T cells. To explore the impact of Imprime on NK-mediated cytotoxicity, we further evaluated in vivo killing of MHC class I deficient TapKO cells after intravenous administration of Imprime. In these experiments Imprime was able to enhance the NK cytotoxic killing of the targets. All NK cell killing observed was dependent on type I IFN, phagocytic cells and dectin 1. Imprime treatment increased cytokines that drive enhanced NK activation and effector phenotype. Significant increases were observed in the cytokines IL15/IL15rα, IL18, IL12p70 in lymph node lysates as well as increases in the effector molecules CD69, GrB, and CD107a on splenic NK cells. The upregulation of all of these molecules during Imprime treatment was dependent on dectin 1. Additionally, IL15/IL15rα production was also dependent on type I IFN, and phagocytic cells. Interestingly, Ly6c hi monocytes, which are increased after Imprime treatment, also show enhancement of IL15rα expression. Collectively, these data demonstrate that Imprime contributes to enhanced NK functionality and killing which may provide a unique immunotherapeutic approach to complement existing therapies. Citation Format: Kathryn A. Fraser, Takashi Kangas, Ross B. Fulton, Steven M. Leonardo, Ben Harrison, Yumi Yokoyama, Nandita Bose, Jeremy R. Graff, Mark Uhlik, Keith B. Gorden. Imprime PGG, a soluble yeast b-glucan PAMP, enhancement of anti-tumor responses in combination with tumor targeting antibody is highly dependent on NK cell killing [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 3767.

Abstract 1703: A novel tumor vaccine platform: direct conjugation of antigens to the β glucan PAMP Imprime PGG enhances antigen presentation and T cell priming

Significant clinical and pre-clinical research has shown that PAMPs- Pathogen Associated Molecular Patterns- can trigger an integrated anti-cancer immune response involving both innate and adaptive immunity. Imprime PGG is a soluble yeast β-1,3/1,6 glucan currently in multiple phase 2 clinical studies in combination with the immune checkpoint inhibitor (CPI), pembrolizumab. Preclinical mechanistic research has shown that Imprime PGG can re-polarize immunosuppressive myeloid cells in the tumor microenvironment and activate antigen presenting cells to prime antigen-specific CD8 T cells thereby boosting effector T cell function and expansion. Based on this ability to activate dendritic cells and induce type I interferon, we sought to explore the use of Imprime as an immune activating/antigen-directing scaffold onto which we could attach tumor antigen to drive an antigen-specific T cell based immune response. Imprime PGG has been safely administered intravenously to more than 400 human subjects and thus represents a safe starting point for the development of a cancer vaccine. Imprime’s 1,3/1,6 β-glucan polymeric structure allows for the straightforward attachment of peptides and proteins via three different conjugation routes. To explore this function we covalently linked the chicken ovalbumin (OVA) protein to Imprime to generate a β-glucan-protein conjugate (Imprime-OVA). Using T cell receptor transgenic OT-I CD8 and OT-II CD4 T cells to track responses to OVA, we treated mice with Imprime-OVA intravenously and examined the expansion and functional quality of the T cell response 7 days later at the peak of expansion. Following Imprime-OVA treatment, both OVA-specific CD8 and CD4 T cells underwent vigorous expansion. OT-I CD8 T cells upregulated the transcription factor Tbet, which is central to developing effector functions, and were highly multifunctional in their ability to produce IFN-γ, TNF-α and IL-2. OT-I T cells responded similarly to vaccination with the minimal H-2Kb/OVA257-264 peptide covalently attached to Imprime. By comparison, vaccination with unconjugated OVA (protein or peptide) and Imprime was much less effective in driving T cell expansion and differentiation. The CD8 T cell response required Batf3-dependent cross-presenting DCs whereas the CD4 T cell response did not, and both CD4 and CD8 responses required the C-type lectin receptor Dectin-1. These data show that an Imprime PGG-protein conjugate can effectively elicit the expansion and functional activation of cytotoxic T cells and may have utility as a potential cancer vaccine platform. These data provide evidence that Imprime PGG not only serves as a combination therapy for CPIs, anti-angiogenics, and tumor targeting antibodies but may also be a robust platform for multiple tumor vaccine strategies. Citation Format: Kyle S. Michel, Ross B. Fulton, Steven M. Leonardo, Keith B. Gorden, Jeremy R. Graff, Michael E. Danielson. A novel tumor vaccine platform: direct conjugation of antigens to the β glucan PAMP Imprime PGG enhances antigen presentation and T cell priming [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 1703. doi:10.1158/1538-7445.AM2017-1703