Agonistic Antibody Research Articles

Abstract PR-10: Phase 1b study of maintenance soluble beta-glucan (Odetiglucan) in combination with a CD40 agonistic monoclonal antibody (CDX-1140) in patients with metastatic pancreatic adenocarcinoma that had not progressed on first-line chemotherapy

Abstract Background: Metastatic pancreatic ductal adenocarcinoma (mPDAC) has a dismal 5-year overall survival (OS) rate of 3%. Current standard of care provides only modest anti-cancer impact. Preclinical models show that combinations of myeloid agonists can induce strong anti-tumor activity in PDAC tumors resistant to immunotherapy, including immune checkpoint blockade (anti-CTLA4, anti-PD1). This study combines odetiglucan, a beta glucan pathogen-associated molecular pattern (PAMP), with CDX-1140, a fully human Ig2K agonistic monoclonal CD40 antibody, in the maintenance setting after cytotoxic chemotherapy induction. Preliminary findings are presented as the study was closed early for financial reasons. Methods: Patients with mPDAC who had not progressed after 4-8 months of 1L chemotherapy were enrolled and received weekly odetiglucan (4 mg/kg) plus every 3-week dosing of CDX-1140 (1.5 mg/kg). The primary endpoints were safety, MTD and RP2D, with secondary endpoints including DOR, PFS, ORR, and OS. Tissue and blood samples were collected at baseline and during treatment to evaluate immune pharmacodynamic responses. Results: A total of 5 patients received treatment on study including 4 patients who were efficacy evaluable and 1 patient who withdrew due to adverse event (arthralgia) after beginning treatment. Treatment-related emergent adverse events were seen in all patients with most common AEs including grade 1-2 arthralgia, chills, and fatigue. Grade 3 AEs included arthralgia (n=1), fatigue (n=1), and leukopenia (n=2). The ORR included 1 PR, 1 SD, and 2 PD. Two patients remained on treatment for >100 days. The pharmacodynamic profile was consistent with the mode of action of odetiglucan and CDX-1140, including a reduction in peripheral Dectin-1 expressing monocytes and transient decreases in peripheral B cells. Patients with disease control (PR or SD, n=2) compared to patients with progression (PD, n=2) showed expansion of peripheral Ki67+ CD8+ T cells following treatment. Analysis of a paired tumor biopsy from 1 patient with PD confirmed penetration of odetiglucan into a liver metastasis with expansion of CD11b+ macrophages and a reduction of cytokeratin+ tumor content, but without significant changes in T cell infiltration. Conclusions: The combination of odetiglucan and CDX-1140 was feasible and safe when administered as maintenance therapy in patients with mPDAC whose disease had not progressed on 1L chemotherapy. Treatment produced encouraging activity with changes in immunological correlates associated with disease control. These early results support further investigation combining odetiglucan with a CD40 agonist in mPDAC. Citation Format: Mark H O'Hara, Max Wattenberg, Ignacio Garrido-Laguna, Eileen M O'Reilly, Michael Yellin, Tibor Keler, Michele Gargano, Nick Niles, Jeremy Drees, Nandita Bose, Gregory L Beatty. Phase 1b study of maintenance soluble beta-glucan (Odetiglucan) in combination with a CD40 agonistic monoclonal antibody (CDX-1140) in patients with metastatic pancreatic adenocarcinoma that had not progressed on first-line chemotherapy [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr PR-10.

Abstract B061: Phase 1b study of maintenance soluble beta-glucan (Odetiglucan) in combination with a CD40 agonistic monoclonal antibody (CDX-1140) in patients with metastatic pancreatic adenocarcinoma that had not progressed on first-line chemotherapy

Abstract Background: Metastatic pancreatic ductal adenocarcinoma (mPDAC) has a dismal 5-year overall survival (OS) rate of 3%. Current standard of care provides only modest anti-cancer impact. Preclinical models show that combinations of myeloid agonists can induce strong anti- tumor activity in PDAC tumors resistant to immunotherapy, including immune checkpoint blockade (anti-CTLA4, anti-PD1). This study combines odetiglucan, a beta glucan pathogen- associated molecular pattern (PAMP), with CDX-1140, a fully human Ig2K agonistic monoclonal CD40 antibody, in the maintenance setting after cytotoxic chemotherapy induction. Preliminary findings are presented as the study was closed early for financial reasons. Methods: Patients with mPDAC who had not progressed after 4-8 months of 1L chemotherapy were enrolled and received weekly odetiglucan (4 mg/kg) plus every 3-week dosing of CDX-1140 (1.5 mg/kg). The primary endpoints were safety, MTD and RP2D, with secondary endpoints including DOR, PFS, ORR, and OS. Tissue and blood samples were collected at baseline and during treatment to evaluate immune pharmacodynamic responses. Results: A total of 5 patients received treatment on study including 4 patients who were efficacy evaluable and 1 patient who withdrew due to adverse event (arthralgia) after beginning treatment. Treatment-related emergent adverse events were seen in all patients with most common AEs including grade 1-2 arthralgia, chills, and fatigue. Grade 3 AEs included arthralgia (n=1), fatigue (n=1), and leukopenia (n=2). The ORR included 1 PR, 1 SD, and 2 PD. Two patients remained on treatment for >100 days. The pharmacodynamic profile was consistent with the mode of action of odetiglucan and CDX-1140, including a reduction in peripheral Dectin-1 expressing monocytes and transient decreases in peripheral B cells. Patients with disease control (PR or SD, n=2) compared to patients with progression (PD, n=2) showed expansion of peripheral Ki67+ CD8+ T cells following treatment. Analysis of a paired tumor biopsy from 1 patient with PD confirmed penetration of odetiglucan into a liver metastasis with expansion of CD11b+ macrophages and a reduction of cytokeratin+ tumor content, but without significant changes in T cell infiltration. Conclusions: The combination of odetiglucan and CDX-1140 was feasible and safe when administered as maintenance therapy in patients with mPDAC whose disease had not progressed on 1L chemotherapy. Treatment produced encouraging activity with changes in immunological correlates associated with disease control. These early results support further investigation combining odetiglucan with a CD40 agonist in mPDAC. Citation Format: Mark H O'Hara, Max Wattenberg, Ignacio Garrido-Laguna, Eileen M O'Reilly, Michael Yellin, Tibor Keler, Michele Gargano, Nick Niles, Jeremy Drees, Nandita Bose, Gregory L Beatty. Phase 1b study of maintenance soluble beta-glucan (Odetiglucan) in combination with a CD40 agonistic monoclonal antibody (CDX-1140) in patients with metastatic pancreatic adenocarcinoma that had not progressed on first-line chemotherapy [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B061.

Abstract B043: Synergy of Subasumstat and anti-CD40 improves survival by augmenting tumor macrophage infiltration

Abstract Background: Identification of effective immunomodulatory strategies remains a major unmet need in pancreatic ductal adenocarcinoma (PDAC). Agonistic anti-CD40 antibody (aCD40) exhibits direct cytotoxicity on tumor cells and augments antigen presentation. Subasumstat(SUB), a sumoylation inhibitor, augments innate and adaptive immune responses by increasing interferon signaling. We hypothesized that SUB+aCD40 combination would promote an effective antitumor immune response in an aggressive orthotopic model of PDAC. Methods: 2000 KPC 46 cells were implanted in the pancreas of four groups of F1 hybrid (initial survival, CD8+ T-cell depletion, and clodronate liposome depletion) and nude mice, enrolled when tumors had reached 3-5mm. The vehicle (veh) group received 80ul SUB vehicle (q48 hrs) and 100 ug blank IgG (qwk); SUB only group received 15mg/kg q48h; aCD40 only group received 100ug qwk; SUB+aCD40 group received 15mg/kg SUB q48h and 100 ug aCD40 qwk. Single cell RNA sequencing (scRNASeq) was conducted on n=2 tumors from each group. Immunohistochemistry (IHC) was performed on veh and SUB+aCD40 tumors for macrophage infiltration using F4/80. Flow cytometry (FC) was employed for the presence of lymphoid and myeloid markers in veh, SUB+aCD40, and clodronate depleted SUB+aCD40 (SUB+aCD40+clod) groups. Results: Mice receiving SUB+aCD40 had a significantly improved median survival over those receiving monotherapy or vehicle (24.5d vs. SUB:15.5d, aCD40:20d, veh: 19d). scRNASeq revealed increased tumor infiltrating CD8+ T-cell and myeloid populations when comparing veh and monotherapy to SUB+aCD40. F4/80 IHC confirmed increased macrophage infiltration in SUB+aCD40 vs veh (p = 0.01). The survival advantage conferred by SUB+aCD40 was preserved in both CD8+ T-cell depleted mice (p=0.025) and nude mice (p=0.037), while clodronate eliminated the treatment survival advantage (p=0.39). FC showed increased tumor macrophage/monocyte infiltration in SUB+aCD40 vs veh (veh:11.9%, SUB+aCD40:22.5%, p=0.03) and reduction in SUB+aCD40+clod (12.6%, p=0.01). CD8+ T-cell CD69 expression decreased in the SUB+aCD40+clod group (SUB+aCD40:19%, SUB+aCD40+clod:6%, p< 0.01). CD8 T-cell infiltration (SUB+aCD40:4.95%, SUB+aCD40+clod:3.85%, p=0.27) and CD4/CD8 T-cell ratio (veh: 1.26, SUB+aCD40:0.52, SUB+aCD40+clod:0.69, p=0.09) were not affected by clodronate. Conclusion: SUB+aCD40 therapy confers a survival advantage in the KPC46 model of PDAC in a T-cell independent manner by increasing macrophage/monocyte infiltration. SUB+aCD40 promotes CD8+ T-cell infiltration and activation, but they do not contribute to survival advantage, possibly due to exhaustion. Future work will include addition of therapeutics to mitigate/reverse T-cell exhaustion to harness both innate and adaptive immunity in treating PDAC. Citation Format: Asimina Courelli, Kevin Li, James Lee, Herve Tiriac, Yuan Chen, Andrew Lowy. Synergy of Subasumstat and anti-CD40 improves survival by augmenting tumor macrophage infiltration [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B043.

Agonist antibody to guanylate cyclase receptor NPR1 regulates vascular tone

Heart failure is a leading cause of morbidity and mortality1,2. Elevated intracardiac pressures and myocyte stretch in heart failure trigger the release of counter-regulatory natriuretic peptides, which act through their receptor (NPR1) to affect vasodilation, diuresis and natriuresis, lowering venous pressures and relieving venous congestion3–8. Recombinant natriuretic peptide infusions were developed to treat heart failure but have been limited by a short duration of effect9,10. Here we report that in a human genetic analysis of over 700,000 individuals, lifelong exposure to coding variants of the NPR1 gene is associated with changes in blood pressure and risk of heart failure. We describe the development of REGN5381, an investigational monoclonal agonist antibody that targets the membrane-bound guanylate cyclase receptor NPR1. REGN5381, an allosteric agonist of NPR1, induces an active-like receptor conformation that results in haemodynamic effects preferentially on venous vasculature, including reductions in systolic blood pressure and venous pressure in animal models. In healthy human volunteers, REGN5381 produced the expected haemodynamic effects, reflecting reductions in venous pressures, without obvious changes in diuresis and natriuresis. These data support the development of REGN5381 for long-lasting and selective lowering of venous pressures that drive symptomatology in patients with heart failure.

Photothermal therapy co-localized with CD137 agonism improves survival in an SM1 melanoma model without hepatotoxicity.

Aim: We investigate combining Prussian Blue nanoparticles (PBNPs), as photothermal therapy (PTT) agents, with agonistic CD137 antibodies (αCD137) on a single nanoparticle platform to deliver non-toxic, anti-tumor efficacy in SM1 murine melanoma.Methods: We electrostatically coated PBNPs with αCD137 (αCD137-PBNPs) and quantified their physicochemical characteristics, photothermal and co-stimulatory capabilities. Next, we tested the efficacy and hepatotoxicity of PTT using αCD137-PBNPs (αCD137-PBNP-PTT) in SM1 tumor-bearing mice.Results: The αCD137-PBNPs retained both the photothermal and agonistic properties of the PBNPs and αCD137, respectively. In vivo, SM1 tumor-bearing mice treated with αCD137-PBNP-PTT exhibited a significantly higher survival rate (50%) without hepatotoxicity, compared with control treatments.Conclusion: These data suggest the potential utility of co-localizing PBNP-PTT with αCD137-based agonism as a novel combination nanomedicine.

Pharmacologic HIF stabilization activates costimulatory receptor expression to increase antitumor efficacy of adoptive T cell therapy.

Adoptive cell transfer (ACT) is a therapeutic strategy to augment antitumor immunity. Here, we report that ex vivo treatment of mouse CD8+ T cells with dimethyloxalylglycine (DMOG), a stabilizer of hypoxia-inducible factors (HIFs), induced HIF binding to the genes encoding the costimulatory receptors CD81, GITR, OX40, and 4-1BB, leading to increased expression. DMOG treatment increased T cell killing of melanoma cells, which was further augmented by agonist antibodies targeting each costimulatory receptor. In tumor-bearing mice, ACT using T cells treated ex vivo with DMOG and agonist antibodies resulted in decreased tumor growth compared to ACT using control T cells and increased intratumoral markers of CD8+ T cells (CD7, CD8A, and CD8B1), natural killer cells (NCR1 and KLRK1), and cytolytic activity (perforin-1 and tumor necrosis factor-α). Costimulatory receptor gene expression was also induced when CD8+ T cells were treated with three highly selective HIF stabilizers that are currently in clinical use.

Human/mouse CD137 agonist, JNU-0921, effectively shrinks tumors through enhancing the cytotoxicity of CD8+ T cells in cis and in trans.

Agonistic antibodies against CD137 have been demonstrated to completely regress established tumors through activating T cell immunity. Unfortunately, current CD137 antibodies failed to benefit patients with cancer. Moreover, their antitumor mechanisms in vivo remain to be determined. Here, we report the development of a small molecular CD137 agonist, JNU-0921. JNU-0921 effectively activates both human and mouse CD137 through direct binding their extracellular domains to induce oligomerization and signaling and effectively shrinks tumors in vivo. Mechanistically, JNU-0921 enhances effector and memory function of cytotoxic CD8+ T cells (CTLs) and alleviates their exhaustion. JNU-0921 also skews polarization of helper T cells toward T helper 1 type and enhances their activity to boost CTL function. Meanwhile, JNU-0921 attenuates the inhibitory function of regulatory T cells on CTLs. Our current work shows that JNU-0921 shrinks tumors by enhancing the cytotoxicity of CTLs in cis and in trans and sheds light on strategy for developing CD137 small molecular agonists.

LTβR Agonism Promotes Anti-Tumor Immune Responses via Modulation of the Tumor Microenvironment.

The presence of high endothelial venules (HEV) and tertiary lymphoid structures (TLS) in solid tumors is correlated with favorable prognosis and better responses to immune-checkpoint blockade (ICB) in many cancer types. Elucidation of the molecular mechanisms underlying intratumoral HEV and TLS formation and their contribution to anti-tumor responses may facilitate development of improved treatment strategies. Lymphotoxin beta receptor (LTβR) signaling is a critical regulator of lymph node organogenesis and can cooperate with antiangiogenic and ICB treatment to augment tumor-associated HEV formation. Here, we demonstrated that LTβR signaling modulates the tumor microenvironment via multiple mechanisms to promote anti-tumor T cell responses. Systemic activation of the LTβR pathway via agonistic antibody treatment induced tumor-specific HEV formation, upregulated the expression of TLS-related chemokines, and enhanced dendritic cell (DC) and T cell infiltration and activation in syngeneic tumor models. In vitro studies confirmed direct effects of LTβR agonism on DC activation and maturation and associated DC-mediated T cell activation. Single agent LTβR agonist treatment inhibited syngeneic tumor growth in a CD8+ T cell- and HEV-dependent manner, and the LTβR agonist enhanced anti-tumor effects of anti-PD-1 and CAR T cell therapies. An in vivo tumor screen for TLS-inducing cytokines revealed that the combination of LTβR agonism and lymphotoxin alpha (LT⍺) expression promoted robust intratumoral TLS induction and enhanced tumor responses to anti-CTLA-4 treatment. Collectively, this study highlights crucial functions of LTβR signaling in modulating the tumor microenvironment and could inform future HEV/TLS-based strategies for cancer treatments.

Abstract Or120: CD40 is an immune checkpoint regulator that potentiates myocardial inflammation through activation and expansion of CCR2 + macrophages and CD8 T-cells

Novel immune checkpoint therapeutics including CD40 agonists have tremendous promise to elicit antitumor responses in patients resistant to current therapies. Conventional immune checkpoint inhibitors (PD-1/PD-L1, CTLA-4 antagonists) are associated with serious adverse cardiac events including life-threatening myocarditis. However, little is known regarding the potential for CD40 agonists to trigger myocardial inflammation or myocarditis. Here, we leveraged genetic mouse models, single cell sequencing, and cell depletion studies to demonstrate that an anti-CD40 agonist antibody reshapes the cardiac immune landscape through activation of CCR2 + macrophages and subsequent recruitment of effector memory CD8 T-cells. We identify a positive feedback loop between CCR2 + macrophages and CD8 T-cells driven by IL12b, TNF, and IFN-γ signaling that promotes myocardial inflammation and show that prior exposure to CD40 agonists sensitizes the heart to secondary insults and accelerates LV remodeling. Collectively, these findings highlight the potential for CD40 agonists to promote myocardial inflammation and potentiate heart failure pathogenesis.

Targeting TNFRSF25 by agonistic antibodies and multimeric TL1A proteins co-stimulated CD8+ T cells and inhibited tumor growth

BackgroundTumor necrosis factor receptor superfamily 25 (TNFRSF25) is a T-cell co-stimulatory receptor. Expression of its ligand, TNF-like cytokine 1A (TL1A), on mouse tumor cells has been shown to promote tumor...

Selenium nanoparticles enhance the anti-tumor immune responses of anti-4-1BB antibody and alleviate the adverse effects on mice

4-1BB agonists for cancer immunotherapy have shown good preliminary efficacy in clinical trials, but several of the first-generation 4-1BB agonistic antibodies entering the clinic have failed due to safety issues. Selenium nanoparticles (SeNPs) exhibit anti-inflammatory, anti-tumor, antioxidant, and immune-modulating properties. In addition, they have been shown to have detoxifying effects and prevent oxidative liver damage. In this study, we used an anti-4-1BB antibody in combination with SeNPs to evaluate the anti-lung cancer effects in in vitro and in vivo experiments and explore the underlying mechanisms by pathological analyses, quantitative PCR, and enzyme-linked immunoassay. We found that 5 μmol·L–1 anti-4-1BB antibody combined with 1 μmol·L–1 SeNPs increased the expression of IFN-γ and promoted the killing effects of peripheral blood mononuclear cells on Lewis lung carcinoma cells, with a lethality rate up to 56.88 %. Experiments in tumor-bearing mice showed that the tumor inhibition rate was 58.61 % after treatment with 3.5 mg/kg anti-4-1BB antibody combined with 0.25 mg/kg SeNPs, and the liver function index returned to normal. When the combined treatment was compared with the antibody treatment alone, detection of immune relevant factors demonstrated that the expression of FOXP3, IL-2, IL-12, and TNF-α in the spleen was downregulated, whereas the expression of IFN-γ in the spleen, serum, and tumor was upregulated, accompanied by increased Fas ligand expression in the tumor tissues. Based on these findings, we get the conclusion that anti-4-1BB antibody combined with SeNPs may alleviate the immunosuppression of regulatory T cells, promote the immune cell proliferation and metastasis to synergistically kill tumor cells. This combination also reduces the inflammatory damage to normal tissues and slows overstimulation of the splenic immune response.

CD137 expression and signal function drive pleiotropic γδ T-cell effector functions that inhibit intracellular M. tuberculosis growth

Co-activation signal that induces/sustains pleiotropic effector functions of antigen-specific γδ T cells remains unknown. Here, Mycobacteria tuberculosis (Mtb) tuberculin administration during tuberculosis (TB) skin test resulted in rapid expression of co-activation signal molecules CD137 and CD107a by fast-acting Vγ2Vδ2 T cells in TB-resistant subjects (Resisters), but not patients with active TB. And, anti-CD137 agonistic antibody treatment experiments showed that CD137 signaling enabled Vγ2Vδ2 T cells to produce more effector cytokines and inhibit intracellular Mtb growth in macrophages (Mɸ). Consistently, Mtb antigen (Ag) HMBPP stimulation induced sustainable high-level CD137 expression in fresh and activated Vγ2Vδ2 T cells from uninfected subjects, but not TB patients. CD137+Vγ2Vδ2 T-cell subtype predominantly displayed central memory phenotype and mounted better proliferative responses than CD137−Vγ2Vδ2 T-cells. In response to HMBPP, CD137+Vγ2Vδ2 T-cell subtype rapidly differentiated into greater numbers of pleiotropic effector cells producing anti-Mtb cytokines compared to CD137−Vγ2Vδ2 T subtype, with the non-canonical NF-κB pathway involved. CD137 expression in Vγ2Vδ2 T cells appeared to signal anti-Mtb effector functions leading to intracellular Mtb growth inhibition in Mɸ, and active TB disrupted such CD137-driven anti-Mtb effector functions. CD137+Vγ2Vδ2 T-cells subtype exhibited an epigenetic-driven high-level expression of GM-CSF and de novo production of GM-CSF critical for Vγ2Vδ2 T-cell controlling of Mtb growth in Mϕ. Concurrently, exosomes produced by CD137+Vγ2Vδ2 T cells potently inhibited intracellular mycobacterial growth. Furthermore, adoptive transfer of human CD137+Vγ2Vδ2 T cells to Mtb-infected SCID mice conferred protective immunity against Mtb infection. Thus, our data suggest that CD137 expression/signaling drives pleiotropic γδ T-cell effector functions that inhibit intracellular Mtb growth.

Impact of isotype on the mechanism of action of agonist anti-OX40 antibodies in cancer: implications for therapeutic combinations

BackgroundOX40 has been widely studied as a target for immunotherapy with agonist antibodies taken forward into clinical trials for cancer where they are yet to show substantial efficacy. Here, we...

PD-L1 promotes oncolytic virus infection via a metabolic shift that inhibits the type I IFN pathway.

While conventional wisdom initially postulated that PD-L1 serves as the inert ligand for PD-1, an emerging body of literature suggests that PD-L1 has cell-intrinsic functions in immune and cancer cells. In line with these studies, here we show that engagement of PD-L1 via cellular ligands or agonistic antibodies, including those used in the clinic, potently inhibits the type I interferon pathway in cancer cells. Hampered type I interferon responses in PD-L1-expressing cancer cells resulted in enhanced efficacy of oncolytic viruses in vitro and in vivo. Consistently, PD-L1 expression marked tumor explants from cancer patients that were best infected by oncolytic viruses. Mechanistically, PD-L1 promoted a metabolic shift characterized by enhanced glycolysis rate that resulted in increased lactate production. In turn, lactate inhibited type I IFN responses. In addition to adding mechanistic insight into PD-L1 intrinsic function, our results will also help guide the numerous ongoing efforts to combine PD-L1 antibodies with oncolytic virotherapy in clinical trials.

CD40 agonist engineered immunosomes modulated tumor microenvironment and showed pro-immunogenic response, reduced toxicity, and tumor free survival in mice bearing glioblastoma

CD40 agonist antibodies (αCD40) have shown promising anti-tumor response in both preclinical and early clinical studies. However, its systemic administration is associated with immune- and hepato-toxicities which hampers its clinical usage. In addition, αCD40 showed low tumor retention and induced PD-L1 expression which makes tumor microenvironment (TME) immunosuppressive. To overcome these issues, in this study, we have developed a multifunctional Immunosome where αCD40 is conjugated on the surface and RRX-001, a small molecule immunomodulator was encapsulated inside it. Immunosomes showed higher tumor accumulation till 96 h of administration and displayed sustained release of αCD40 in vivo. Immunosomes significantly delayed tumor growth and showed tumor free survival in mice bearing GL-261 glioblastoma by increasing the population of CD45+CD8+ T cells, CD45+CD20+ B cells, CD45+CD11c+ DCs and F4/80+CD86+ cells in TME. Immunosome significantly reduced the population of T-regulatory cells, M2 macrophage, and MDSCs and lowered the PD-L1 expression. Moreover, Immunosomes significantly enhanced the levels of Th1 cytokines (IFN-γ, IL-6, IL-2) over Th2 cytokines (IL-4 and IL-10) which supported anti-tumor response. Most interestingly, Immunosomes averted the in vivo toxicities associated with free αCD40 by lowering the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), IL-6, IL-1α and reduced the degree of liver damage. In addition, Immunosomes treated long-term surviving mice showed tumor specific immune memory response which prevented tumor growth upon rechallenge. Our results suggested that this novel formulation can be further explored in clinics to improve in vivo anti-tumor efficacy of αCD40 with long-lasting tumor specific immunity while reducing the associated toxicities.

A novel anti-CD47 protein antibody and toll-like receptor agonist complex detects tumor surface CD-47 changes in early stage lung cancer by in vivo imaging

CD47, a cell surface protein known for inhibiting phagocytosis, plays a critical role in the tumor microenvironment (TME) and is a potential biomarker for cancer. However, directly applying αCD47, a hydrophilic macromolecular antibody that targets CD47, in vivo for cancer detection can have adverse effects on normal cells, cause systemic toxicities, and lead to resistance against anti-cancer therapies. In this study, we developed a novel complex incorporating aluminum-based metal-organic frameworks (Al-MOF) loaded with indocyanine green (ICG), αCD47, and resiquimod (R848), a hydrophobic small molecule Toll-like receptor 7/8 (TLR7/8) agonist. Upon activation with an infrared 808 nm laser, the nanocomposites exhibited photothermal effects that triggered the release of the loaded reagents, induced ROS production, and induced changes in the TME. This led to the polarization of immune-suppressive M2 macrophages towards an immune-stimulatory M1 phenotype, promoted dendritic cell (DC) maturation, and enabled mature DCs to facilitate antigen presentation, T-cell activation, and critical roles in tumor immunity. Furthermore, in vivo imaging successfully detected the specific binding of αCD47 with CD47 on tumor cells. Overall, the complex composed of αCD47 antibody and toll-like receptor agonist showed promising efficacy in both tumor diagnosis and therapy, providing a potential strategy for detecting early lung cancer and modulating the tumor microenvironment for improved treatment outcomes.

TRLS-06. PHASE 1 STUDY OF THE CD40 AGONISTIC MONOCLONAL ANTIBODY APX005M (SOTIGALIMAB) IN PEDIATRIC, ADOLESCENT, AND YOUNG ADULT PATIENTS WITH RECURRENT OR PROGRESSIVE CNS TUMORS AND NEWLY DIAGNOSED BRAIN STEM GLIOMA: A PEDIATRIC BRAIN TUMOR CONSORTIUM STUDY (PBTC-051; NCT03389802)

Abstract BACKGROUND CD40 is a co-stimulatory receptor present on antigen-presenting cells. CD40 agonists are hypothesized to reverse cancer-induced immune suppression and induce tumor cell killing. The PBTC completed a first-in-pediatrics phase 1 study (PBTC-051) of the CD40 agonistic monoclonal antibody APX005M (sotigalimab). METHODS Patients 1-21.99 years old with recurrent/progressive/refractory malignant non-brainstem central nervous system (CNS) tumors were eligible for stratum 1, and those with post-radiation pre-progression diffuse intrinsic pontine glioma (DIPG) for stratum 2. Patients received APX005M (sotigalimab) via IV on day 1 of each 21 day cycle. A 3 + 3 statistical design was used for dose escalation through 3 planned dose levels (DL). Patients could receive APX005M (sotigalimab) for 36 cycles or until disease progression, unacceptable toxicity, or death. RESULTS 31 eligible patients enrolled (20 on stratum 1, 11 on stratum 2) between 2018-2023; 29 were evaluable for dose finding. The most common diagnosis in stratum 1 was ependymoma (35%). DL3 (0.6 mg/kg) was the recommended phase 2 dose (RP2D) for stratum 1 and DL2 (0.3 mg/kg) for stratum 2. Five patients had dose limiting toxicities—two stratum 1 patients at DL3 and three stratum 2 patients at DL3. The most common grade 3+ adverse events at least possibly attributable to APX005M (sotigalimab) were lymphopenia, neutropenia, leukopenia, and increased ALT. Median number of cycles was 2 (1-36) for stratum 1 and 3 (1-19) for stratum 2. No patients had an objective response to APX005M. 6 month progression-free survival (PFS) for stratum 2 patients (calculated from start of treatment) was 30.7±12.8%; median PFS was 3.7 months. CONCLUSIONS APX005M was well-tolerated in pediatric patients with both recurrent/refractory/progressive CNS tumors and post-radiation pre-progression DIPG. A RP2D was established for both populations. Future studies of this agent in pediatric neuro-oncology patients should prioritize combination therapy.

Intermittent MEK Inhibition with GITR Costimulation Rescues T-cell Function for Increased Efficacy with CTLA-4 Blockade in Solid Tumor Models.

MEK inhibitors (MEKi) have shown limited success as a treatment for MAPK/ERK pathway-dependent cancers due to various resistance mechanisms tumor cells can employ. CH5126766 (CKI27) is an inhibitor that binds to MEK and prevents release of RAF, reducing the relief of negative feedback commonly observed with other MEKis. We observed that CKI27 increased MHC expression in tumor cells and improved T cell-mediated killing. Yet, CKI27 also decreased T-cell proliferation, activation, and cytolytic activity by inhibiting the MAPK/ERK pathway that is activated downstream of T-cell receptor signaling. Therefore, we aimed to balance the positive and negative immunomodulatory effects of MEKis for optimal combination with immunotherapy. Intermittent administration of CKI27 allowed T cells to partially recover and costimulation via GITR and OX-40 agonist antibodies completely alleviated inhibition of function. In Kras mutant lung and colon tumor mouse models, intermittent CKI27 and anti-GITR significantly decreased tumor growth and prolonged survival when further combined with CTLA-4 immune checkpoint blockade. Moreover, this triple combination increased CD8+ and CD4+ T-cell proliferation, activation, and effector/memory subsets in the tumor-draining lymph nodes and tumors and led to intratumoral regulatory T-cell destabilization. These data, collectively, will allow for more informed decisions when optimizing combination regimens by overcoming resistance, reducing toxicity, and generating long-term immune responses.

PEG-lipid-modified agonistic antibody against tumor necrosis factor receptor family elicits superior apoptosis-inducing activity against human carcinoma

We have recently developed a novel PEG-lipid-modified antibody to enhance the induction of apoptosis by the agonistic antibody. The chemically modified TRA-8 antibody [anti-death receptor 5 (DR5) antibody] with PEG-lipid (DSPE-PEG) demonstrated significant cytotoxic activity in vitro without the need for crosslinking with a secondary antibody, which is typically required. We investigated the correlation between the PEG-lipid structure and the cytotoxic activity of the modified antibodies by varying the PEG length or lipid structure. However, when the DSPE-PEG-modified TRA-8 antibody was incubated with plasma, it lost its cytotoxic activity, likely due to degradation in the DSPE-PEG component. Nevertheless, by designing new PEG-lipids that are intended to be resistant to enzymatic degradation, we were able to prevent this degradation and restore the cytotoxic activity of the modified antibody. These findings provide valuable insights for the design of PEG-lipid-modified antibodies and suggest their potential effectiveness in enhancing cancer therapy.

Intratumoral Fc-optimized agonistic CD40 antibody induces tumor rejection and systemic antitumor immunity in patients with metastatic cancer.

While CD40 agonism is an attractive approach for activating antigen-presenting cells and initiating antitumor responses, previous attempts have encountered limited clinical efficacy coupled with toxicity. We previously demonstrated that interactions between the antibody Fc domain and the inhibitory receptor FcγRIIB are critical for enhanced antitumor activity. Here, we present the results of a phase 1 study on intratumoral administration of an anti-CD40 agonistic antibody (2141-V11) Fc-engineered to enhance FcγRIIB binding. Primary endpoints included safety, maximum tolerated dose (MTD), and recommended phase 2 dose. Secondary objectives included preliminary clinical activity and correlative studies from biospecimens. 2141-V11 was well-tolerated without dose-limiting toxicities and MTD was not reached. In ten evaluable patients with metastatic cancer, the overall response rate was 20%, with complete responses in two patients (melanoma and breast carcinoma) and stable disease in six patients. 2141-V11 induced tumor regression in injected and non-injected lesions, with increased leukocyte infiltration and tertiary lymphoid structures (TLS) formation in post-treatment biopsies. In a humanized mouse model for CD40 and FcγRs, 2141-V11 induced TLS formation in mice bearing orthotopic breast carcinoma, correlating with local and abscopal antitumor effects, systemic immune activation, and immune memory. These findings support the safety and efficacy of 2141-V11, warranting phase 2 studies and suggesting a unique mechanism of action for this Fc-enhanced immunotherapy (NCT04059588).