Efficacy In Syngeneic Tumor Models Research Articles

Abstract 5090: TU2218, a dual inhibitor against ALK5/VEGFR2, increases anti-CTLA4 antitumor efficacy in syngeneic tumor models

Abstract The combination of pembrolizumab with low-dose ipilimumab shows substantial antitumor activity and manageable profile of toxicity in anti-PD-(L)1 antibody failure-setting (NCT02743819). This suggests a breakthrough for the absence of treatment option after relapsed/refractory anti-PD-(L)1 therapy. Accordingly, the anti-CTLA4 drug-based combination can be considered as a promising strategy for beneficial outcomes against resistance acquired from immunotherapy. Herein, we demonstrate that TU2218, a first-in-class, orally available inhibitor against ALK5 and VEGFR2, showed synergistic antitumor efficacy when combined with an anti-CTLA4 antibody in preclinical tumor models which is being accompanied by increasing ratio of CD8 T cell to regulatory T cell and enhanced immunological memory. In this work, antitumor efficacy of TU2218 combination with an anti-CTLA4 antibody was assessed with CT26-, 4T1-, B16F10- and WEHI-164-bearing mice. In the CT26 model, the combination of TU2218 with an anti-CTLA4 antibody significantly inhibited tumor growth up to 92% compared to vehicle, thus being superior to single treatments (e.g., tumor growth inhibition (TGI) 46% for TU2218, TGI 74% for anti-CTLA4). In this combination group, the complete regression (CR) rate was 75 % (i.e., six cases among eight mice), while single treatments showed lower CR rates (e.g., CR 10% (1/10) for TU2218, CR 30% (3/10) for anti-CTLA4). Meanwhile, the role of CD8+ T cell in antitumor activity was elucidated by in vivo depleting CD8+ T cell in mice treated with combination therapy. The depletion of CD8+ T cells reduced the antitumor response, which suggests the indispensable role of CD8+ T cells in the antitumor efficacy of TU2218 and anti-CTLA-4 antibody combination. In addition, the long-term immune-memory was evaluated by re-implanting tumor cells into both mice cured by combination therapy and age-matched tumor-naïve mice. In this case, 6 mice cured of original implantation with CT26 tumors showed complete resistance to the re-implantation of CT26 cells during an untreated period for 21 days, whereas all age-matched tumor-naïve mice have developed tumors after 10 days from cell-transplants. Importantly, we could confirm the positive correlation between the immunological memory-response of combination therapy and the increasing rate of effector memory CD4+ and CD8+ T cells in spleens compared to those of age-matched group. In 4T1-, B16F10- and WEHI-164-bearing mice, combination of TU2218 with an anti-CTLA4 antibody led to higher CR rate as well as enhanced inhibition of tumor growth. Overall, our findings showed that TU2218 plays multifaceted roles in inducing immune activation under combination with an anti-CTLA4 antibody, which may be attributed to the increased ratio of cytotoxic CD8 T cell to regulatory T cell and improvement of adaptive immunity with long-term immunological memory. Citation Format: Nam-Hoon Kim, Jihyun Lee, Hun-Taek Kim. TU2218, a dual inhibitor against ALK5/VEGFR2, increases anti-CTLA4 antitumor efficacy in syngeneic tumor models. [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 5090.

Abstract 3232: Combination therapy with onco-selective mRNA LNPs targets the complex immunosuppressive tumor microenvironment and is well tolerated at efficacious doses

Abstract Novel improved cancer immunotherapies are needed, since most cancer patients exhibit a treatment resistant tumor microenvironment to currently available immunotherapeutic modalities. Due to primary resistance or the development of acquired resistance to therapeutic approaches these patients unfortunately do not benefit from a long-lasting overall survival benefit. Combination treatments are a promising approach to induce a sustained anti-tumor immune response. However, dose limiting adverse events in healthy tissues and/or the complex expensive production requirements have limited the full therapeutic potential of combinatorial applications. mRNA-based combinatorial therapy options have evolved by recent advancements in the production, purification, and delivery of mRNA to cells. Today, mRNA therapies are a rapidly growing class of medications that can redefine how many diseases are treated. These therapies enable the production of biologics directly in the patient and mRNA LNPs are easy to manufacture at scale. Despite these key advantages, mRNA therapeutics are yet to show their true potential in oncology. Dose limiting toxicities of mRNA immunotherapies are driven, in part, by systemic payload (encoded protein) toxicity, which can be avoided by engineering the mRNA to improve its onco-selectivity and thereby reduce systemic target-mediated adverse events. Kernal Biologics develops novel onco-selective mRNA therapies directed to breach the immunosuppressive tumor microenvironment. Based on our proprietary machine learning-enabled computational pipeline, we designed our next generation mRNA therapeutics. These mRNAs have the potential to increase the depth and breadth of anti-PD1/PD-L1 treatment plus enable responses in patients that are currently non-responders or refractory to the clinically approved immune checkpoint blockade therapies. Here, we describe combination therapies of tumor-selective mRNA LNPs that achieve strong and lasting anti-tumor efficacy in syngeneic tumor models. We observed regression of established tumors, complete responses (CRs) and improved overall survival. At efficacious doses the mRNA LNPs were well tolerated while driving anti-tumor immune activation and modulation of the tumor microenvironment. Mouse blood hematology and chemistry analyses were within a normal range. Similarly, pathological immunohistochemistry analysis of liver, spleen and bone marrow revealed no findings. In summary, our data support the feasibility of onco-selective mRNA combination treatment of a variety of cancers with poor T cell infiltration and immunosuppressive TME, major obstacles in cancer immunotherapy. Citation Format: Manfred Kraus, Rudy Christmas, Tom A. Addison, Yulia Rybakova, Leona Lee, Jieni Xu, Mark Krimmer, Cafer Ozdemir, Burak Yilmaz, Yusuf Erkul. Combination therapy with onco-selective mRNA LNPs targets the complex immunosuppressive tumor microenvironment and is well tolerated at efficacious doses [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 3232.

Abstract 5542: Characterization of a long-acting IL-13 super-antagonist engineered to target tumor associated macrophages and myeloid cells

Abstract Introduction: Regulation of Th1/Th2 balance in the tumor microenvironment (TME) plays an important role in both tumor progression and response to immunotherapy. The IL-4/IL-13 pathway induces an anti-inflammatory (Th2) response by stimulating M2 skewing of tumor associated macrophage (TAM) and myeloid derived suppressor cells (MDSCs), thereby supporting tumor growth by suppressing Th1 response. Therefore, inhibition of the IL-4/IL-13 pathway has the potential to inhibit tumor growth by shifting the TME towards a pro-inflammatory response. To achieve this, we engineered a long-acting IL-13 super-antagonist (i.e., Fc-MDNA413) to target the IL-13Rα1 component of type II IL-4 receptor (IL-4Rα/IL-13α1) expressed on TAMs and MDSCs to inhibit their differentiation and expansion for immuno-oncological applications. Experimental Procedure: Studies conducted include binding analyses by Biacore/SPR, signaling analyses in IL-4/IL-13 cell reporters, in vitro cell proliferation, in vitro M1/M2 macrophage polarization and in vivo efficacy in syngeneic tumor models. Summary of Data: IL-13Rα1 and IL-4Rα constitute the type II IL-4R that drives a Th2 response upon activation by IL-4 and IL-13. Binding studies showed that in comparison to wild-type Fc-IL13, Fc-MDNA413 has higher affinity for IL-13Rα1 but lower affinity for the decoy IL-13Rα2 receptor, indicating a more favorable receptor selectivity. In a cell-based reporter assay, Fc-MDNA413 inhibits both IL-4 and IL-13 induced p-STAT6 signaling, a pathway that drives towards a Th2 response. The antagonism of Fc-MDNA413 was also observed in an IL-13 dependent TF-1 proliferation assay where dose-dependent inhibition of cell proliferation was demonstrated. In a macrophage polarization assay with human primary myeloid cells, Fc-MDNA413 inhibited both IL-4 and IL-13 induced M2 polarization in a dose-dependent manner, consistent with an inhibitory effect on IL-4/IL-13 signaling. Treatment of tumor bearing mice with Fc-MDNA413 inhibited tumor growth in multiple syngeneic tumor models (e.g., EMT6 breast cancer and KLN205 lung cancer) known to have higher percentage of TAMs and MDSCs in TME, hence demonstrating the therapeutic potential of this long-acting IL-4/IL-13 super-antagonist. Conclusion: Fc-MDNA413 is a long-acting IL-4/IL-13 super-antagonist that inhibits IL-4 and IL-13 induced signaling and M2 skewing of macrophages, resulting in tumor growth inhibition in vivo. Ongoing studies include testing in additional syngeneic tumor models, deciphering in vivo mechanism of action and investigating potential synergy with other therapies. Next generation multifunctional superkines that incorporate IL-4/IL-13 antagonism are under development to target immunologically challenging tumors. Citation Format: Aanchal Sharma, Manjunatha Ankathatti Munegowda, Fahar Merchant, Minh D. To. Characterization of a long-acting IL-13 super-antagonist engineered to target tumor associated macrophages and myeloid cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5542.

Abstract 5532: An ‘anti-PD1-IL2 beta-only super-agonist’ displays potent anti-tumor efficacy

Abstract Introduction: Combination studies with engineered IL-2 variants and immune-checkpoint blockade have demonstrated synergy in both pre-clinical models and the clinic. Co-administration of those 2 agents can potentially face challenges related to overlapping toxicities in obtaining optimal doses for synergistic effects. We propose to potentiate the synergy by enabling cis activation of IL-2 agonism and blockade of PD-1/PDL-2 binding with a single anti-PD1-IL2 to generate Bifunctional SuperKines for ImmunoTherapy (BiSKIT). Both human (h) and mouse (m) versions of anti-PD1 are fused with MDNA109FEAA, an IL-2 superkine with enhanced affinity for IL-2Rβ (CD122) and no binding to IL-2Rα (CD25). Experimental Procedure: Studies included binding analyses with Biacore/SPR, signaling analyses in an IL-2 cell reporter assay, cell proliferation assay, in vitro pSTAT5 signaling in human PBMCs, PD1/PDL-1 cell reporter assay and efficacy in syngeneic tumor models. Summary of Data: Anti-hPD1-MDNA109FEAA and surrogate anti-mPD1-MDNA109FEAA were tested in both in vitro and in vivo assays. Both constructs demonstrated selective binding to either hPD1 or mPD1 as expected, and the MDNA109FEAA motif retained its receptor selectivity (enhanced affinity to CD122 and no binding to CD25). In both HEK-Blue IL-2 reporter and CTLL-2 proliferation bioassays, anti-PD1-MDNA109FEAA exhibited similar agonistic effect as MDNA109FEAA-Fc, demonstrating that potency towards the IL-2 axis was not diminished. This was further confirmed with pSTAT5 signaling in primary human PBMCs where anti-PD1-MDNA109FEAA showed enhanced activation of CD8 T-cells and NK cells with reduced activity on Treg when compared to rhIL-2. Both constructs were also effective at inhibiting PD1/PDL-1 pathway in respective human and mouse specific cell-based reporter assays. In a CT26 syngeneic colon tumor model, anti-mPD1-MDNA109FEAA exhibited superior efficacy over monotherapy and combination with anti-mPD1 and MDNA109FEAA-Fc when administered at equal molar dosage and same schedule. Conclusion: We have engineered anti-PD1-MDNA109FEAA to enable cis engagement with IL-2 receptor and PD1 as an approach to activate immune cells while reducing their exhaustion. Anti-mPD1-MDNA109FEAA achieved potent efficacy in a CT26 colon tumor model and is currently being evaluated in additional syngeneic tumor models with varying responsiveness to immune checkpoint blockade. Citation Format: Minh D. To, Manjunatha Ankathatti Munegowda, Rosemina Merchant, Aanchal Sharma. An ‘anti-PD1-IL2 beta-only super-agonist’ displays potent anti-tumor efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5532.

ROR\u03b3t agonist enhances anti-PD-1 therapy by promoting monocyte-derived dendritic cells through CXCL10 in cancers

BackgroundThe overall response rate to checkpoint blockade remains unsatisfactory, partially due to the immune-suppressive tumor microenvironment. A retinoic acid-related orphan receptor γt (RORγt) agonist (LYC-55716) is currently used in clinical trials combined with anti-PD-1, but how the Th17 cell transcription factor RORγt enhances antitumor immunity of PD-1 in the tumor microenvironment remains elusive.MethodsThe expression of mRNA was analyzed using qPCR assays. Flow cytometry was used to sort and profile cells. Cell migration was analyzed using Transwell assays. Biacore was used to determine the binding affinity to the RORγt protein. The RORγt GAL4 cell-based reporter gene assay was used to measure activity in the RORγt driven luciferase reporter gene expression.ResultsWe designed a potent and selective small-molecule RORγt agonist (8-074) that shows robust antitumor efficacy in syngeneic tumor models and improves the efficacy of anti‑PD‑1 in a murine lung cancer model. RORγt agonist treatment increased intratumoral CD8+ T cells, which were correlated with CXCL10 and monocyte-derived dendritic cells (MoDCs). In addition, the RORγt agonist promoted Type 17 T cell migration by upregulating CCL20 and CCR6 expression, and Type 17 T cell tumor infiltration. CCL20 induces MoDCs migration, and CXCL10 derived from MoDCs promotes CD8+ T cell migration.ConclusionOur results revealed that the RORγt agonist improved the efficacy of anti-PD-1. The RORγt agonist increased the migration of MoDCs, which increased the local levels of CXCL10, thus promoting CD8+ T cell tumor infiltration. Our findings provide the mechanistic insights implicating the RORγt agonist in immunotherapy and offer a strategy for targeting the RORγt agonist to improve PD-1 antibody efficacy in cancers.

Abstract P198: MDNA11 is a long-acting ‘beta-only’ IL-2 agonist that demonstrates a safe and durable anti-tumor immune response

Abstract Background: MDNA11 is an engineered ‘beta-only’ IL-2 superkine with (1) enhanced affinity for IL-2Rb (CD122) to preferentially activate anti-cancer effector immune cells and (2) extended half-life by fusion to human albumin, avoiding the need for frequent administration. Methodology: MDNA11 was characterized in in vitro and in vivo studies including assessment of efficacy in syngeneic tumor models as single agent and in combination with immune checkpoint inhibitors (CPIs). A GLP study in non-human primate was conducted to evaluate the safety, pharmacodynamic and pharmacokinetic profiles of MDNA11. Results: MDNA11 does not bind IL-2Ra (CD25) but demonstrates a 30-fold higher affinity for IL-2Rb (CD122), resulting in enhanced in vitro STAT5 signaling in human NK and CD8+ T-cells with diminished activation in Tregs when compared to rhIL-2. In CT26 and MC38 syngeneic tumor models, MDNA11 demonstrated potent and durable anti-tumor activity as monotherapy (Q1W x 2) and synergized with anti-PD1 and anti-CTLA4 to achieve complete response and tumor clearance. These mice were protected against relapse and subsequent tumor re-challenges without any further treatment by inducing long-term antigen-specific CD8+ T-cells. MDNA11 binds to human and cynomolgus monkey IL-2 receptors with near-identical affinity. A GLP toxicology study involving intravenous administration on a Q2W x 3 schedule showed that MDNA11 was well tolerated up to the highest planned dose of 600 mcg/kg. The most common clinical observations were dehydration and diarrhea, which generally occurred following the first but not subsequent dose administration. There was a dose-proportional increase in Cmax and AUC. MDNA11 induced dose-dependent proliferation and expansion of peripheral CD4+ and CD8+ T-cells lasting beyond 7-days following each administration. Similar stimulatory effects on NK cells were observed but effects on Tregs were limited. Despite the robust immune response, there was no sign of cytokine release syndrome nor evidence of ADA. At the highest dose of MDNA11 (600 mcg/kg), an increase in eosinophil counts was observed following the second dose, but histopathological evaluation showed no evidence of pulmonary edema nor vascular leak syndrome in all organs examined. These data constitute a strong framework for the conduct of a phase 1/2 study in patients with advanced solid tumors that encompasses dose escalation, dose expansion, and combination with CPIs. Conclusions: MDNA11 is a long-acting ‘beta-only’ IL-2 superkine that exhibits robust anti-tumor activity in mouse tumor models as a single agent and in combination with anti-PD1 and anti-CTLA4. In a non-human primate GLP toxicology study, MDNA11 demonstrated durable and selective activation of immune effector cells (NK, CD4+ and CD8+ T-cells), and a favorable safety profile. Citation Format: Minh D. To, Fahar Merchant, Ma'an Muhsin, Carole Galligan, L. Bruce Pearce, Peter Lloyd, Rosemina Merchant. MDNA11 is a long-acting ‘beta-only’ IL-2 agonist that demonstrates a safe and durable anti-tumor immune response [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P198.

Abstract 1921: Armed oncolytic myxoma virus demonstrates efficacy in syngeneic tumor models alone and in combination with immune checkpoint inhibitors

Abstract Oncolytic viruses (OV) selectively replicate in and lyse tumor cells and provide stimulation to the immune system, representing a promising therapeutic option in development to treat cancers that do not respond well to treatment with immune checkpoint inhibitors. Myxoma virus (MYXV) is a member of the Pox family of double stranded DNA viruses. The natural host of MYXV is a subset of rabbits and hares, but MYXV can infect cancer cell lines of humans and other species. The genome of MYXV is relatively large and is amenable to engineering for expression of transgenes making it an excellent oncolytic virus for introduction of immunomodulatory proteins. Oncolytic activity and transgene production capability were examined in multiple mouse cancer cell lines. In vivo efficacy following intratumoral and intravenous administration of armed myxoma virus, both alone and with immune checkpoint inhibitors is demonstrated. Immunomodulatory mechanisms of actions are investigated. Multi-armed myxoma represents a novel, engineerable, oncolytic virus with preclinical characteristics that warrant further investigation as an immunomodulatory cancer therapeutic. Citation Format: Lina S. Franco, Joseph Mamola, Wazir Abdullahi, Ana de Matos, Mario Abrantes, Benjamin S. Walker, Zachary Tacner, Nicole Grigaitis, Cassandra Kien, Grant McFadden, Leslie L. Sharp. Armed oncolytic myxoma virus demonstrates efficacy in syngeneic tumor models alone and in combination with immune checkpoint inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1921.

Abstract 1711: Induction of anti-tumor immunity by intratumoral delivery of mRNA encoding cytokines and TNFRSF agonists

Abstract In the recent decade several immune checkpoint inhibitors, such as anti-PD1/PD-L1 antibodies, have been approved by regulatory authorities and are actively used in oncology clinical practice. However, these antibodies exhibit durable anti-tumor benefits in only a small fraction of cancer patients. A major unmet medical need in immune oncology (IO) is extending the benefits of these therapies to patients who never experience tumor regression (primary resistance) and those who derive initial benefit before experiencing tumor progression (acquired resistance). Novel IO combinations have the potential to both sensitize primary resistant tumors and combat the acquisition of resistance to checkpoint inhibition. Synthetic, in vitro transcribed messenger RNA (mRNA) is an emerging therapeutic modality that offers the ability to express a broad range of therapeutic combinations. In this project, we proposed to ignite anti-tumor immunity locally through intratumoral (i.t.) delivery of a mixture of mRNAs encoding cytokines and a TNF receptor superfamily (TNFRSF) agonist. Using in vitro and in vivo screening, we identified a combination therapy consisting of three cytokines IFNα, IL-7, and a fusion protein of IL-15 with its receptor alpha chain as well as one TNFRSF agonist, an Fc fusion hexameric OX40L for enhanced receptor clustering. The mRNA mixture enhanced human antigen-specific T cell responses in vitro and displayed potent anti-tumor efficacy in syngeneic tumor models when injected i.t., demonstrating the induction of protective immunity. Importantly, in a preclinical dual-flank syngeneic tumor model, we demonstrated that this protective immunity extended beyond tumors injected with the lead mRNA mixture to control growth of distant untreated lesions. Mechanistically, the lead candidate induced anti-tumor immunity by recruiting dendritic cells to the tumor, leading to activation and expansion of tumor specific CD8 T cells. In summary, this mRNA mixture, encoding IFNα, IL-7, IL-15 and OX40L, induced potent anti-tumor immunity in mice and will be further investigated as a combination partner to overcome resistance mechanisms to immune checkpoint inhibition. Citation Format: Julie-Ann Gavigan, Chaomei Shi, Michael Lampa, Natalia Malkova, Qunyan Yu, Hui Qu, Mikhail Levit, Timothy Wagenaar, Christian Hotz, Xiangming Li. Induction of anti-tumor immunity by intratumoral delivery of mRNA encoding cytokines and TNFRSF agonists [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1711.

Abstract PO091: Armed Myxoma virus demonstrates efficacy in syngeneic tumor models alone and in combination with immune checkpoint inhibitors

Abstract Oncolytic Viruses (OV) selectively replicate in and lyse tumor cells and provide stimulation to the immune system. This represents a promising therapeutic option for cancer patients that do not respond well to treatment with immune checkpoint inhibitors. Myxoma virus (MYXV) is a member of the Poxviridae family of double stranded DNA viruses. The natural host of MYXV is a subset of lagomorphs, but MYXV can infect cancer cell lines of humans and other species. The genome of MYXV is relatively large and is amenable to engineering for expression of transgenes making it an excellent oncolytic virus for introduction of immunomodulatory proteins. The current work describes the oncolytic activity, transgene production capability, in vivo activity and immunomodulatory mechanism of actions following intratumoral and intravenous administration of armed myxoma viruses in murine cancer models. Additional combination therapy with clinically relevant immune checkpoint inhibitors is demonstrated. Citation Format: Lina S. Franco, Lino E. Torres-Dominguez, Joseph Mamola, Ana L. de Matos, Mario Abrantes, Benjamin S. Walker, Zachary Tacner, Cassandra Kien, Natalie M. Elliott, Grant McFadden, Leslie L. Sharp. Armed Myxoma virus demonstrates efficacy in syngeneic tumor models alone and in combination with immune checkpoint inhibitors [abstract]. In: Abstracts: AACR Virtual Special Conference: Tumor Immunology and Immunotherapy; 2020 Oct 19-20. Philadelphia (PA): AACR; Cancer Immunol Res 2021;9(2 Suppl):Abstract nr PO091.

Abstract 1559: TLR7/8 immune-stimulating antibody conjugates elicit robust myeloid activation leading to enhanced effector function and anti-tumor immunity in pre-clinical models

Abstract Deficient anti-tumor immunity often results from an immunosuppressive tumor microenvironment (TME) that renders antigen presenting cells (APCs) unable to effectively stimulate T cells. Recent studies indicate that local delivery of immunostimulatory adjuvants can activate tumor resident APCs, driving uptake, processing and presentation of tumor neoantigens to T cells that mediate anti-tumor immunity. To overcome challenges associated with intratumoral delivery of such adjuvants, we developed a novel class of TLR immune-stimulating antibody conjugates (TAC) that comprise a TLR7/8 agonist conjugated to tumor-targeting monoclonal antibodies. In vitro co-cultures with human cancer cell lines and leukocytes revealed that TACs potently activate primary APCs, leading to increased co-stimulatory molecule expression (e.g. CD40, CD86) and secretion of pro-inflammatory cytokines (e.g. TNFα). The TACs also enhanced antibody-mediated effector functions such as ADCC and ADCP. Surprisingly, these constructs also induced dendritic cell (DC) differentiation from monocytes, as measured by changes in cellular morphology and DC surface markers (e.g. CD14 downregulation). CyTOF-based analysis of intracellular signaling in human leukocytes revealed a unique signaling signature of the conjugate compared to a mixture of its components, suggesting a novel biological mechanism by which the conjugate stimulates APCs. Finally, we demonstrated in vivo efficacy in syngeneic tumor models in which TAC treatment led to tumor clearance and development of immunologic memory. These results provide a strong rationale for this technology as a platform for cancer immunotherapy. Citation Format: Shelley E. Ackerman, Joseph C. Gonzalez, Josh D. Gregorio, Jason C. Paik, Felix J. Hartmann, Justin A. Kenkel, Arthur Lee, Angela Luo, Cecelia I. Pearson, Murray L. Nguyen, Benjamin Ackerman, Lauren Y. Sheu, Richard P. Laura, Steven J. Chapin, Brian S. Safina, Sean C. Bendall, David Dornan, Edgar G. Engleman, Michael N. Alonso. TLR7/8 immune-stimulating antibody conjugates elicit robust myeloid activation leading to enhanced effector function and anti-tumor immunity in pre-clinical models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1559.

Abstract B005: FPT155, a novel therapeutic CD80-Fc fusion protein with potent antitumor activity in preclinical models

Abstract Optimal activation of T cells requires both antigen-specific signaling and induction of costimulatory pathways. Costimulatory pathways are upregulated by foreign antigenic challenge but are not active in the normal immune state, thus maintaining peripheral tolerance to self-antigens. The absence of costimulatory signaling may likewise play a role in tolerance to cancer cells. Five Prime has developed a CD80 (B7.1) extracellular domain (ECD)-Fc fusion protein (FPT155) as a costimulatory molecule that can stimulate T-cell activation and break tumor immune tolerance. In vitro studies show that FPT155 induces T-cell activation and cytokine production via CD28. FPT155 activity requires T-cell receptor engagement, which indicates that FPT155 is not a CD28 “superagonist” that can drive T-cell activation independently of antigenic stimulation. To investigate the impact of CD80-Fc in preclinical models, we generated murine FPT155 (mFPT155), a surrogate fusion protein, composed of the mouse CD80 ECD fused with the murine Fc region of IgG2a. mFPT155 is well tolerated in mice and has potent efficacy in syngeneic tumor models, including the induction of complete tumor regression in the CT26 model. mFPT155 promotes the infiltration of T cells into the tumor core and increases the effector T:Treg ratio, thus inducing a favorable microenvironment for an effective antitumor immune response. Furthermore, combination of mFPT155 and anti-PD1 therapy leads to stronger antitumor efficacy compared to either therapy alone. Together these results indicate that mFPT155 is a potent T-cell costimulator with strong antitumor activity in mouse tumor models. We are currently performing IND-enabling studies and plan to initiate a clinical study with FPT155 in 2018. Citation Format: Susannah D. Barbee, Barbara Sennino, Jacqueline De La Torre, Monica Macal, Quinn Walker, Marc R. Jabon, Amanda Chen, David A. Busha, Margaret Best, Kristen L. Pierce, Luis Borges, Kevin P. Baker, Thomas Brennan. FPT155, a novel therapeutic CD80-Fc fusion protein with potent antitumor activity in preclinical models [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B005.