anti-CD27 Monoclonal Antibody Research Articles

Abstract 4261: CD27 a new immuno-oncology target shaping innate and adaptive anti-tumor immune responses

Abstract CD27 is a member of the TNF-receptor superfamily, highly expressed on CD4+ and CD8+ T cells as well as on NK and NKT cells. It plays a key role on T cell proliferation and differentiation after stimulation with its ligand CD70. The co-stimulatory signal of CD27 on T cell is mediated via the NFκB pathway but also via the phosphatidylinositol 3 kinase and the protein kinase B. CD27/CD70 co-stimulation has the potential to boost immunity by T-cell activation, clonal expansion and enhanced differentiation into antigen specific cytotoxic and memory T cells. CD27/CD70 also influences the innate immune response via a direct activation of the NK cells and a subsequent secretion of interferon-gamma (IFN-γ). Therefore, CD27 signaling promotes cytotoxic T cell based anti-tumor immunity. With its central role in an immunological response, CD27 is a promising target for antitumor therapy. Previous works have demonstrated the efficacy of an agonistic CD27 antibody in controlling tumor growth and metastasis in different mice models including melanoma, renal cell carcinoma, breast cancer and lymphomas. This anti-tumor effect is mediated in part by an effective recruitment of IFN-γ producing CD8+ T cells within the tumor. Moreover, CD27 stimulation of Tumor Infiltrating Lymphocytes (TILs) can lower their threshold of activation and provide a broader repertoire of Ag-reactive T cells within the tumor. We have selected a lead therapeutic antibody from our library of 147 fully human anti-CD27 monoclonal antibodies generated in the Trianni mice. After confirming its binding potency and selectivity as well as its cross-reactivity with Non-Human Primate (NHP)-CD27 but not with the mouse-CD27, this lead candidate demonstrated strong agonistic proprieties. This was shown by its ability to induce a strong NFκB signal as well as to induce T cell proliferation and activation with secretion of pro-inflammatory cytokines. This antibody demonstrated agonistic proprieties without cross-linking confirming its potency. T cell activation observed after treatment with our anti-CD27 antibody only occurs in the presence of TCR engagement, preventing the risk of spontaneous activation of naïve T cells in vivo. The ability of our CD27 monoclonal antibody to increase an immune response was confirmed in a Mixed Lymphocyte Reaction assay with multiple donors. The role played by the NK cells and their activation via CD27 antibody was also demonstrated. To evaluate the anti-tumor functions of our lead antibody as a single agent or in combination with other immuno-therapies in vivo we have used human CD27 transgenic mice. We have demonstrated in the MB49 bladder tumor model as well as the EG7 thymoma model that our lead antibody induces a strong single agent anti-tumor activity and these tumors were totally controlled in combination with an anti-PD1 antibody. We are now analyzing the pharmacokinetic and pharmacodynamic of our antibody as well as its safety in a NHP model Citation Format: Thierry Guillaudeux, Yulia Ovechkina, Shaarwari Sridhar, David Peckham, Jessica Cross, Nathan Eyde, Emily Frazier, Neda Kabi, Remington Lance, Kurt Lustig, Mei Xu, Tarcha Eric, Shawn Iadonato. CD27 a new immuno-oncology target shaping innate and adaptive anti-tumor immune responses [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 4261.

Abstract P035: A promising cancer immunotherapy target: Novel fully human agonist antibodies against the human T-cell costimulatory receptor CD27

Abstract CD27 is a member of the TNF receptor superfamily and plays a critical role in T-cell activation by providing a costimulatory signal. CD27 signaling enhances T-cell proliferation, activation and differentiation of effector and memory T cells and therefore promotes cytotoxic T cell (CTL)-based anti-tumor immunity. Agonistic stimulation of CD27 is a promising cancer immunotherapy approach to boost specific T cell driven anti-tumor responses. In this study, we generated a series of 147 fully human monoclonal anti-CD27 antibodies and tested their agonist properties to stimulate T cell activation. Using a NF-κB reporter Jurkat cell line, we evaluated in vitro the ability of anti-CD27 antibodies to induce CD27 receptor activation. With this assay, five antibodies have been selected for their agonist properties. When combined with suboptimal T cell receptor (TCR) stimulation, agonist antibodies induced CD27 receptor activation with an EC50 of 1-5 ug/mL. We also used human peripheral blood T cells to characterize the CD27-mediated costimulatory effects of agonist antibodies in combination with TCR stimulation. Our anti-CD27 monoclonal antibodies boosted T cell proliferation and induced IL-2 and TNFalpha secretion only in a presence of TCR engagement. Moreover, CD27 agonists induce strong T cell proliferation in a Mixed Lymphocyte Reaction. CD27 antibodies were shown to bind human and cynomolgus monkey CD27 with a KD value of 5-20 nM as determined by BioLayer Interferometry, but do not bind to mouse CD27. In vivo experiments are currently ongoing to demonstrate the efficient anti-tumor activity of the selected CD27 agonist antibodies in different mice tumor models. In conclusion, we have developed and successfully selected efficient fully human immunostimulatory agonist CD27 mAbs as a promising cancer immunotherapy. Citation Format: Yulia Ovechkina, Shaarwari Sridhar, David Jurchen, David Peckham, Eric Tarcha, Shawn Iadonato, Thierry Guillaudeux. A promising cancer immunotherapy target: Novel fully human agonist antibodies against the human T-cell costimulatory receptor CD27 [abstract]. In: Abstracts: AACR Virtual Special Conference: Tumor Immunology and Immunotherapy; 2021 Oct 5-6. Philadelphia (PA): AACR; Cancer Immunol Res 2022;10(1 Suppl):Abstract nr P035.

133P Novel fully human agonist antibodies against the T-cell costimulatory receptor CD27 shape adaptive anti-tumor immunity

133P Novel fully human agonist antibodies against the T-cell costimulatory receptor CD27 shape adaptive anti-tumor immunity

PD-1 Blockade and CD27 Stimulation Activate Distinct Transcriptional Programs That Synergize for CD8+ T-Cell-Driven Antitumor Immunity.

Purpose: PD-1 checkpoint blockade has revolutionized the field of cancer immunotherapy, yet the frequency of responding patients is limited by inadequate T-cell priming secondary to a paucity of activatory dendritic cells (DC). DC signals can be bypassed by CD27 agonists, and we therefore investigated if the effectiveness of anti-PD-1/L1 could be improved by combining with agonist anti-CD27 monoclonal antibodies (mAb).Experimental Design: The efficacy of PD-1/L1 blockade or agonist anti-CD27 mAb was compared with a dual-therapy approach in multiple tumor models. Global transcriptional profiling and flow cytometry analysis were used to delineate mechanisms underpinning the observed synergy.Results: PD-1/PD-L1 blockade and agonist anti-CD27 mAb synergize for increased CD8+ T-cell expansion and effector function, exemplified by enhanced IFNγ, TNFα, granzyme B, and T-bet. Transcriptome analysis of CD8+ T cells revealed that combination therapy triggered a convergent program largely driven by IL2 and Myc. However, division of labor was also apparent such that anti-PD-1/L1 activates a cytotoxicity-gene expression program whereas anti-CD27 preferentially augments proliferation. In tumor models, either dependent on endogenous CD8+ T cells or adoptive transfer of transgenic T cells, anti-CD27 mAb synergized with PD-1/L1 blockade for antitumor immunity. Finally, we show that a clinically relevant anti-human CD27 mAb, varlilumab, similarly synergizes with PD-L1 blockade for protection against lymphoma in human-CD27 transgenic mice.Conclusions: Our findings suggest that suboptimal T-cell invigoration in cancer patients undergoing treatment with PD-1 checkpoint blockers will be improved by dual PD-1 blockade and CD27 agonism and provide mechanistic insight into how these approaches cooperate for CD8+ T-cell activation. Clin Cancer Res; 24(10); 2383-94. ©2018 AACR.

A phase II study of glembatumumab vedotin (GV), an antibody-drug conjugate (ADC) targeting gpNMB, in advanced melanoma.

109 Background: gpNMB is an internalizable transmembrane glycoprotein expressed in melanoma and multiple other tumor types. The ADC GV (CDX-011) delivers the potent cytotoxin MMAE to gpNMB+ cells. GV has shown promising activity in advanced melanoma and breast cancer. Methods: This Phase II study (CDX011-05) assessed the efficacy and safety of GV monotherapy (1.9 mg/kg q3w) for patients (pts) with advanced melanoma progressive after ≤1 chemotherapy, ≥1 checkpoint inhibitor (CPI) and if BRAFV600mutated, ≥1 BRAF/MEK inhibitor. Central IHC determined gpNMB expression in archival and/or pre-treatment tumor. Primary endpoint was objective response rate (ORR) (RECIST 1.1) with ≥6 responders out of 52 evaluable pts as threshold for antitumor activity. Additional endpoints: progression free survival (PFS), overall survival (OS), duration of response (DOR), safety, PK/PD and correlation of tumor gpNMB expression with efficacy. Results: 62 pts enrolled (all evaluable) had median age of 67 years; 55% male; 21% BRAFV600mutated; 63% with ≥3 lines prior therapy; 100% had prior CPI; 100% Stage IV; 89% M1c. One confirmed complete response (CR) and 6 confirmed partial responses (PR, including 1 unconfirmed CR) were seen (confirmed ORR = 11%, p = 0.035 comparing to reference ORR 5%); 33 pts had stable disease including 3 unconfirmed PR. Median DOR = 6.0 (range: 4.1, 14+) months (mos), median PFS = 4.3 mos and median OS = 9.8 mos; 26 pts continue to be followed for survival. All pts with available tissue (60/60) had gpNMB+ tumors; 47/60 had 100% gpNMB+ epithelial cells; no clear correlation with outcome was seen in this population with consistent high expression. Toxicities included alopecia, neuropathy, rash, fatigue and neutropenia. Treatment-related rash in cycle 1 was associated with improved ORR (rash = 22%; no rash = 7%), PFS (p = 0.007) and OS (p = 0.035). Conclusions: GV has promising activity (primary endpoint of ORR was met) with a manageable safety profile in heavily pre-treated melanoma pts. Additional cohorts evaluating GV with either varlilumab, an activating anti-CD27 monoclonal antibody, or PD-1 inhibitors are open to accrual to provide further insights into the synergy of ADC and immunotherapy. Clinical trial information: NCT02302339.

Crystal structure of CD27 in complex with a neutralizing noncompeting antibody.

CD27 is a T-cell and B-cell co-stimulatory glycoprotein of the tumor necrosis factor (TNF) receptor superfamily that is dependent on the availability of the TNF-like ligand CD70. Therapeutic approaches to treating autoimmune diseases and cancers with antagonistic and agonistic anti-CD27 monoclonal antibodies (mAbs), respectively, have recently been developed. Mouse anti-human CD27 mAb 2177 shows potency in neutralizing CD70-induced signaling; however, it does not block the binding of soluble CD70. To provide insight into the mechanism of action of the mAb, the crystal structure of the CD27 extracellular domain in complex with the Fab fragment of mAb 2177 was determined at 1.8 Å resolution. CD27 exhibits the assembly of cysteine-rich domains characteristic of the TNF receptor superfamily. The structure reveals a unique binding site of mAb 2177 at the edge of the receptor molecule, which allows the mAb to sterically block the cell-bound form of CD70 from reaching CD27 while leaving the ligand epitope clear. This mode of action suggests a potential dual use of mAb 2177 either as an antagonist or as an agonist.

Anti-CD27 Antibody Potentiates Antitumor Effect of Dendritic Cell-Based Vaccine in Prostate Cancer-Bearing Mice.

In the current study, we investigated whether anti-CD27 monoclonal antibody can enhance the antitumor efficacy of a dendritic cell–based vaccine in prostate cancer–bearing mice. The overall therapeutic effect of a dendritic cell–based vaccine for prostate cancer remains moderate. A prostate cancer model was established by subcutaneous injection of RM-1 tumor cells into male C57BL/6 mice on day 0. After 4 days, tumor-bearing mice were treated with RM-1 tumor lysate–pulsed dendritic cells (i.e., dendritic cell–based vaccine), anti-CD27 monoclonal antibody, or a combination of RM-1 tumor lysate–pulsed dendritic cells with anti-CD27 monoclonal antibody. Mice were killed at 21 days after tumor cell implantation. Tumor size was measured for assessment of antitumor effect. Spleens were collected for analysis of antitumor immune responses. The antitumor immune responses were evaluated by measuring the proliferation and activity of T cells, which have the ability to kill tumor cells. The combination therapy with RM-1 tumor lysate–pulsed dendritic cells and anti-CD27 antibody significantly enhanced T-cell proliferation and activity, and significantly reduced tumor growth, compared with monotherapy with RM-1 tumor lysate–pulsed dendritic cells or anti-CD27 antibody. Our results suggest that combined treatment can strengthen antitumor efficacy by improving T-cell proliferation and activity.

Therapeutic efficacy of a fully human anti-CD27 monoclonal antibody in a murine colon carcinoma model (162.23)

Abstract The costimulatory molecule CD27 is constitutively expressed on the majority of mature T cells, memory B cells, and a portion of NK cells. The interaction of CD27 with its ligand CD70 plays an important role in effector capacity and memory in T cells; in clonal B cell expansion and germinal center formation; and in NK cell cytolytic activity. Agonistic anti-mouse CD27 mAbs have been shown to have potent anti-tumor activity. A panel of fully human antibodies recognizing human CD27 was generated and we have previously shown that our lead clone (1F5) can mediate anti-tumor effects using the BCL1 B-lymphoma line in human CD27-transgenic (hCD27-Tg) mice. In order to further investigate the mechanism of this anti-hCD27 monoclonal and its activity in a therapeutic setting we assessed anti-tumor activity using the CT26 murine colon carcinoma line. CT26 cells were delivered s.c. to hCD27-Tg animals and five doses of 1F5 were delivered to mice i.p. beginning on days 3, 5, or 7. In multiple experiments, tumor growth in mice treated with 1F5 was substantially delayed or prevented compared to that of control-treated mice. When 1F5-treated surviving mice were rechallenged with CT26 cells, without additional anti-CD27 antibody treatment, tumors did not grow in most animals. These data support the continued clinical development of this anti-CD27 mAb to enhance immune responses in cancer therapy. Investigations of immune cell subset depletions and combination therapies are ongoing.

Abstract 4560: Anti-tumor activity of a fully human anti-CD27 monoclonal antibody in a transgenic mouse model

Abstract The costimulatory molecule CD27 is a member of the TNF receptor superfamily, and is constitutively expressed on the majority of mature T cells, memory B cells, and a portion of NK cells. The interaction of CD27 with its ligand CD70 plays an important role in the activation, proliferation, survival, and maturation of effector capacity and memory in T cells; in clonal B cell expansion and germinal center formation; and in NK cell cytolytic activity. Previous published work demonstrated that agonistic anti-mouse CD27 mAbs, given without a DC maturation signal, have potent anti-tumor activity through boosting of T cell immunity. To explore the therapeutic potential of this target, a panel of fully human antibodies recognizing human CD27 was generated using human Ig transgenic mice immunized with recombinant CD27. These anti-CD27 mAbs showed specific and high affinity binding to recombinant CD27 and CD27-expressing lymphoma cells. Human CD27-transgenic (hCD27-Tg) mice were generated to evaluate the anti-CD27 mAbs in vivo. The CD27-Tg mouse line was back-crossed onto a Balb/c background in order to assess anti-tumor activity using the BCL1 B-lymphoma line. Approximately 107 BCL1 cells were delivered i.v. to backcrossed animals on day 0. Doses of 150 to 600 ug of our lead anti-CD27 mAb (clone 1F5) were delivered to mice i.p. on days 3, 5, 7, 9 and 11. In multiple experiments, tumor growth in mice treated with the anti-CD27 mAb 1F5 was substantially delayed compared to that of control-treated mice. Further, many anti-CD27 treated animals did not show evidence of tumor growth several weeks past the point where all control animals had been euthanized. These data confirm that anti-human CD27 mAbs can mediate similar anti-tumor effects as the anti-mouse CD27 agonist antibodies. The 1F5 antibody was also demonstrated to cross-react with CD27 from cynomolgus monkeys, and in a small pilot study was well tolerated and without evidence of significant lymphocyte activation. Taken together the data support the therapeutic potential of this anti-CD27 mAb to enhance immune responses in cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4560. doi:10.1158/1538-7445.AM2011-4560

Anti-tumor effects of depleting and non-depleting anti-CD27 monoclonal antibodies in immune-competent mice

CD27 plays an important role in T-cell co-stimulation and is also expressed on lymphomas. In the present study, we generated novel depleting and non-depleting monoclonal antibodies (mAbs) against mouse CD27 and characterized their co-stimulatory activity in vitro and anti-tumor effects in immune-competent mice bearing syngeneic T-cell lymphoma (EG7) expressing or lacking CD27. A profound anti-tumor effect was observed with a non-depleting mAb (RM27-3E5), but not with a depleting mAb (RM27-3C1), against either EG7/CD27(+) or EG7/CD27(−) tumors, which was associated with the induction of EG7-specific cytotoxic T lymphocytes (CTLs). Consistently, the anti-tumor effect of RM27-3E5 was abolished in T cell-deficient nude mice. These results indicate that a non-depleting agonistic mAb against CD27 is promising for cancer therapy by co-stimulating tumor-specific CTL induction.

CD27 synergizes with CD40 to induce IgM, IgG, and IgA antibody responses of peripheral blood B cells in the presence of IL-2 and IL-10

CD40, a member of the tumor necrosis factor receptor (TNFR) family, promotes IgM, IgG, and IgA antibody (Ab) synthesis in combination with a variety of cytokines. Another TNFR family member, CD27, causes B cells to differentiate into antibody-forming cells, with marginal effects on proliferation. In the present study, we examined whether anti-CD27 monoclonal antibody (mAb) modulates the antibody production induced by anti-CD40 mAb immobilized on L cells expressing FcγRII (CDw32) in the presence of IL-2 and/or IL-10. The anti-CD40 mAb substantially enhanced IgM, IgG, and IgA production in combination with IL-2 and IL-10, whereas anti-CD27 mAb augmented it only marginally, as assessed by enzyme-linked immunosorbent assay. The addition of anti-CD27 mAb enhanced the anti-CD40-mediated IgM, IgG, and IgA antibody production only when both IL-2 and IL-10 were present in the culture. The CD27-positive B cell compartment generated synergistic antibody responses in response to four different stimulants, anti-CD27/anti-CD40 mAb and cytokines IL-2/IL-10, whereas the CD27-negative B cell compartment failed to do so. Kinetic analysis showed that anti-CD40 might function in the early phase of B cell activation, while anti-CD27 mAb functioned in the late stage. The addition of CD27 − to CD27 + B cells in various ratios did not have any effect on the antibody production, suggesting that CD27 + to CD27 − B cell interaction does not occur in this system. Our findings suggest that a member of the TNFR family, CD27, cooperates with CD40 to induce efficient antibody production in combination with cytokines IL-2 and IL-10.

Alternative molecular form of human T cell‐specific antigen CD27 expressed upon T cell activation

The CD27 membrane antigen is exclusively present on a large subset of human peripheral blood T lymphocytes and on mature thymocytes. Several observations point towards a specific role of this molecule on activated T cells. Upon T cell activation induced via the T cell receptor complex, CD27 expression greatly increases, while addition of anti-CD27 monoclonal antibodies amplifies the proliferative response. Within the CD4+ subset, only CD27+ T cells provide helper activity for B cell differentiation. Interestingly, CD27 expression differs not only quantitatively, but also qualitatively between resting and activated T cells. On resting cells, CD27 is a disulfide-linked homodimer with subunits of 55 kDa molecular mass. Upon activation, also a 55-kDa monomer seems to occur, while in addition a 32-kDa component is found. The relationship between these two proteins has been investigated. The 55-kDa and 32-kDa molecules do not seem to be physically associated. The two CD27 components are structurally highly homologous as determined by two-dimensional mapping of tryptic peptides. They both express the epitopes recognized by anti-CD27 antibodies, indicating that the 32-kDa molecule is also T cell specific. Both 55-kDa and 32-kDa molecules carry N-linked carbohydrate groups. However, they differ in other, not yet specified, post-translational modifications, and do not arise from a common precursor simply by alternative N-glycosylation. The 32-kDa form may be derived by proteolytic processing from the 55-kDa protein, but most likely not from a larger (common) precursor. Alternatively, both molecules may arise from a common gene by alternative mRNA splicing, or be derived from highly homologous genes. The dramatic change in molecular composition of CD27 suggests a newly acquired function for CD27 on activated T cells.

Anti‐CD27 monoclonal antibodies identify two functionally distinct subpopulations within the CD4+ T cell subset

Anti-CD27 monoclonal antibodies react with a cell surface molecule expressed on medullary thymocytes and a large subpopulation (75%) of peripheral blood T lymphocytes. This study was undertaken to analyze the functional capacities of CD27+ and CD27- subpopulations within the CD4+ subset. In addition, we investigated whether CD27 subpopulations belong to two mutually exclusive T cell sublineages. Proliferation upon lectin stimulation by either phytohemagglutinin or pokeweed mitogen (PWM) was found to be consistently higher in CD4+CD27+ cells compared to CD4+CD27- cells. In contrast, CD27+ and CD27- cells did not differ in anti-CD3, soluble antigen or interleukin (IL)2-induced proliferation. In PWM-driven B cell differentiation, CD4+CD27+ cells provided helper activity on IgM production, while CD4+CD27- cells did not. The differences observed between CD4+CD27+ and CD4+CD27- cells in both proliferation and T helper activity on IgM production can, at least in part, be explained by the inadequate production of IL2 by CD27- cells upon lectin stimulation. In contrast, CD27+ and CD27- subpopulations did not differ in the production of interferon-gamma. CD27- cells could be induced to express the CD27 antigen through stimulation of these cells with immobilized anti-CD3 antibodies. After 3 days of culture, approximately 50% of the cells had CD27 membrane expression, whereas after 6 days 80% of the cells express the antigen. We conclude that anti-CD27 antibodies identify two functionally distinct T cell populations within the CD4+ subset. These subpopulations apparently do not belong to two separate T cell sublineages, but may reflect differences in the activation state of CD27+ vs. CD27- peripheral blood T lymphocytes.