A novel CD3xPSMA bispecific antibody for efficient T cell mediated killing of prostate tumor cells with minimal cytokine release.

2019 ◽  
Vol 37 (7_suppl) ◽  
pp. 324-324 ◽  
Author(s):  
Starlynn Clarke ◽  
Kevin Dang ◽  
Yuping Li ◽  
Preethi Sankaran ◽  
Duy Pham ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Prostate Tumor ◽  
Bispecific Antibodies ◽  
Therapeutic Window ◽  
Cytokine Release ◽  
Human T Cells ◽  
Prostate Tumor Cells

324 Background: Castration resistant prostate cancer (CRPC) remains an incurable disease and new therapeutics are urgently needed. Prostate specific membrane antigen (PSMA) is expressed on the surface of prostate cancer cells and expression increases with disease progression. Therapies directed against PSMA such as radiolabeled antibodies and T cell redirecting therapies including chimeric antigen receptor T cells (CAR-Ts) and T-cell engaging bispecific antibodies (T-BsAbs) have shown promising efficacy in clinical trials but also induce significant toxicity. In particular CAR-Ts and T-BsAbs potently kill tumor cells but induce cytokine release-related toxicities. Novel anti-CD3 engaging domains may be required to create T-BsAbs with a broader therapeutic window. We have developed fully human CD3xPSMA bispecific antibodies that efficiently eliminate prostate tumor cells while minimizing cytokine release. Methods: Antibodies targeting CD3 and PSMA were generated in transgenic rats that produce human antibodies (UniRat, OmniFlic) followed by repertoire deep sequencing of lymph nodes isolated from immunized animals and high-throughput gene assembly/expression. CD3xPSMA T-BsAbs were assembled and evaluated for T cell activation and ability to eliminate PSMA+ tumor cells in vitro. Results: Primary human T cells were activated only in the presence of both bispecific CD3xPSMA antibodies and PSMA (either plate-bound or on the surface of tumor cells). Potent and selective cytotoxicity against PSMA+ prostate tumor cells was observed in co-cultures of primary human T cells and tumor cells treated with CD3xPSMA bispecific antibodies. Strikingly, CD3xPSMA bispecifics containing a novel low affinity anti-CD3 domain produced similar levels of tumor cell cytotoxicity compared to CD3xPSMA bispecifics containing a traditional high affinity anti-CD3 domain, but with reduced cytokine production. Conclusions: We have created novel CD3xPSMA bispecific antibodies that mediate T-cell killing of PSMA+ tumor cells with minimal production of cytokines. Such T-BsAbs may improve safety, efficacy, and opportunities for combination therapy to treat CRPC.

Evaluation of monovalent versus biparatopic CD3xPSMA bispecific antibodies for t-cell mediated killing of prostate tumor cells with minimal cytokine release.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. e16519-e16519
Author(s):  
Ben Buelow ◽  
Starlynn Clarke ◽  
Kevin Dang ◽  
Jacky Li ◽  
Chiara Rancan ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Cell Activation ◽  
Bispecific Antibodies ◽  
Cytokine Release ◽  
Binding Domains ◽  
Prostate Tumor Cells

e16519 Background: Castration resistant prostate cancer (CRPC) remains an incurable disease and new treatments are needed. Therapies directed against Prostate specific membrane antigen (PSMA) -such as radiolabeled antibodies, chimeric antigen receptor T cells (CAR-Ts) and T-cell engaging bispecific antibodies (T-BsAbs)- have shown promising efficacy but also induce significant toxicity. In particular T-cell redirection leads to efficient killing of tumor cells but induces cytokine release-related toxicities. We have developed a panel of monovalent and biparatopic CD3xPSMA bispecific antibodies that eliminate prostate tumor cells while minimizing cytokine release. Methods: Antibodies targeting CD3 and PSMA were generated in transgenic rats (UniRat™, OmniFlic™) followed by deep sequencing of the antibody repertoire from draining lymph nodes in immunized animals, and high-throughput gene assembly/expression. PSMA x CD3 T-BsAbs were assembled and evaluated for stability, pharmacokinetics, and T cell activation and ability to eliminate PSMA+ tumor cells in vitro and in vivo. Results: Bispecific CD3xPSMA Abs. incorporating either monovalent or biparatopic anti-PSMA binding domains activated T-cells in the presence of PSMA (plate-bound or cell surface), while no T cell activation occurred in the absence of either PSMA antigen or bispecific antibody. Potent/selective cytotoxicity against PSMA+ cells was observed in co-cultures of primary human T cells and tumor cells treated with CD3xPSMA T-BsAbs. Similar results were observed in in vivo Xenograft models of prostate cancer. Strikingly, CD3xPSMA bispecifics containing a novel low affinity anti-CD3 domain produced similar levels of tumor cytotoxicity compared to those with a traditional high affinity anti-CD3 domain, but with reduced cytokine production. Conclusions: We have created novel CD3xPSMA bispecific antibodies incorporating both monovalent and biparatopic anti-PSMA binding domains that mediate T-cell killing of PSMA+ tumor cells with minimal production of cytokines. Such T-BsAbs may improve safety, efficacy, and opportunities for combination therapy to treat CRPC.

Effect of modulation of CD3 binding in a PSMAxCD3 T-cell engaging bispecific antibody on maintenance of efficient tumor cell kill and cytokine release.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e17583-e17583
Author(s):  
Ben Buelow ◽  
Kevin Dang ◽  
Pranjali Dalvi ◽  
Yuping Li ◽  
Alexander Cheung ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
T Cell ◽  
Tumor Cell ◽  
Tumor Cells ◽  
Bispecific Antibody ◽  
Cytokine Production ◽  
Bispecific Antibodies ◽  
Cytokine Release ◽  
Human T Cells

e17583 Background: Castration resistant prostate cancer (CRPC) is an incurable disease and represents a significant unmet need. Prostate specific membrane antigen (PSMA) is a protein highly expressed on the surface of prostate cancer cells; expression has been shown to increase with disease progression. Therapies targeting PSMA, such as anti-PSMA radioligand conjugates, have shown promise in clinical trials, validating this target for CRPC. T-cell recruiting bispecific antibodies (T-BsAbs) have demonstrated potent tumor killing activity against multiple tumor types, but immune-mediated toxicities have hampered T-cell redirecting therapies to date. Using Teneobio’s unique antibody discovery platform, we have developed a CD3xPSMA bispecific antibody (TNB-585) that retains the potent cytotoxicity of other T-BsAbs but with significantly reduced cytokine release. Methods: Antibodies targeting CD3 and PSMA were generated via immunization of our proprietary transgenic animals. Candidate antibodies were selected by repertoire deep sequencing of B-cells from draining lymph nodes, followed by high-throughput gene assembly and recombinant expression. Multiple bispecific antibodies targeting CD3 and PSMA were assembled and evaluated for their ability to selectively activate primary human T-cells and mediate killing of PSMA+ tumor cells in vitro, ex vivo, and in vivo. T-cell activation surface markers, cytokine production, and tumor cell cytotoxicity were measured. Results: In co-culture experiments, primary human T-cells were activated only in the presence of both the bispecifics and PSMA+ cells. These bispecifics mediated potent and selective cytotoxicity against PSMA-positive tumor cells, prostate tumor cell lines, or primary human prostate tumor cells isolated from patients. From among these we identified TNB-585, which showed attenuated binding to CD3. TNB-585 mediated comparable tumor cell cytotoxicity to CD3xPSMA T-BsAbs containing a high affinity anti-CD3 domain but with significantly reduced cytokine production. TNB-585 also showed tumor growth inhibition in xenograft models of prostate cancer in vivo. Conclusions: We have developed a novel CD3xPSMA T-BsAb that mediates T-cell killing of PSMA+ tumor cells with minimal production of cytokines. This molecule may improve safety, efficacy, and offer opportunities for combination therapy to treat CRPC. A Phase 1 clinical trial of this compound in CRPC is scheduled to begin in Q1 2021.

A CD3 x FRα T-cell engaging bispecific antibody for efficient killing of ovarian cancer cells with minimal cytokine release.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e18050-e18050
Author(s):  
Ben Buelow ◽  
Brian Avanzino ◽  
Aarti Balasubramani ◽  
Andrew Boudreau ◽  
Laura Davison ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cell ◽  
Tumor Cells ◽  
Cytokine Production ◽  
Cell Cytotoxicity ◽  
Cytokine Release ◽  
Limited Efficacy ◽  
Human T Cells

e18050 Background: Ovarian Cancer (OvCa) is the leading cause of gynecologic cancer mortality in women. Since the introduction of platinum-based chemotherapy there has been little change in the prognosis of OvCa patients, with < 30% overall survival in advanced disease, creating an urgent medical need for novel therapies. Few ovarian epithelium-specific surface proteins are suited for Ab targeting. However, studies have shown folate receptor α (FRα) to be highly over-expressed in OvCa; expression level and stage correlate, and FRα is absent or minimally expressed in normal tissues. However, naked Ab therapy has shown limited efficacy while CAR-T therapy has been plagued by toxicity and limited efficacy. ADCs have demonstrated some activity but present the risk of toxin-mediated side effects. Using Teneobio’s unique antibody discovery platform, we have developed a CD3 x FRα T-BsAb that retains the potent cytotoxicity of other T-cell redirecting therapies but with significantly reduced cytokine release. Methods: Antibodies targeting CD3 and FRα were generated via immunization of our proprietary transgenic animals. Candidate antibodies were selected by repertoire deep sequencing of B-cells from draining lymph nodes, high-throughput gene assembly, recombinant expression, and functional screening. Bispecific antibodies targeting CD3 and FRα were assembled and evaluated for their ability to selectively activate primary human T-cells and mediate killing of FRα+ tumor cells in vitro and in vivo. T-cell activation surface markers, cytokine production and tumor cell cytotoxicity were measured. Results: Primary human T-cells were activated only in the presence of both the CD3 x FRα T-BsAb and FRα (either recombinant or cell-surface protein). Potent and selective cytotoxicity against FRα+ tumor cells was observed in co-cultures of primary human T-cells and OvCa tumor cell lines. Strikingly, our T-BsAb mediated comparable tumor cell cytotoxicity to CD3 x FRα T-BsAbs containing a high affinity anti-CD3 domain but with significantly reduced cytokine production. Our Ab showed preliminary evidence of tumor growth inhibition in xenograft models of OvCa in vivo. Conclusions: We have created a novel CD3 x FRα T-BsAb that mediates T-cell killing of FRα+ tumor cells with minimal production of cytokines. This molecule may improve safety, efficacy, and offer opportunities for combination therapy to treat OvCa.

scholarly journals Attenuating CD3 affinity in a PSMAxCD3 bispecific antibody enables killing of prostate tumor cells with reduced cytokine release

2021 ◽  
Vol 9 (6) ◽  
pp. e002488
Author(s):  
Kevin Dang ◽  
Giulia Castello ◽  
Starlynn C Clarke ◽  
Yuping Li ◽  
Aarti Balasubramani ◽  
...  
Keyword(s):  
T Cell ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Cell Activation ◽  
Prostate Tumor ◽  
Cytokine Release ◽  
High Affinity ◽  
Prostate Tumor Cells ◽  
3D Spheroids

BackgroundTherapeutic options currently available for metastatic castration-resistant prostate cancer (mCRPC) do not extend median overall survival >6 months. Therefore, the development of novel and effective therapies for mCRPC represents an urgent medical need. T cell engagers (TCEs) have emerged as a promising approach for the treatment of mCRPC due to their targeted mechanism of action. However, challenges remain in the clinic due to the limited efficacy of TCEs observed thus far in solid tumors as well as the toxicities associated with cytokine release syndrome (CRS) due to the usage of high-affinity anti-CD3 moieties such as OKT3.MethodsUsing genetically engineered transgenic rats (UniRat and OmniFlic) that express fully human IgG antibodies together with an NGS-based antibody discovery pipeline, we developed TNB-585, an anti-CD3xPSMA TCE for the treatment of mCRPC. TNB-585 pairs a tumor-targeting anti-PSMA arm together with a unique, low-affinity anti-CD3 arm in bispecific format. We tested TNB-585 in T cell-redirected cytotoxicity assays against PSMA+ tumor cells in both two-dimensional (2D) cultures and three-dimensional (3D) spheroids as well as against patient-derived prostate tumor cells. Cytokines were measured in culture supernatants to assess the ability of TNB-585 to induce tumor killing with low cytokine release. TNB-585-mediated T cell activation, proliferation, and cytotoxic granule formation were measured to investigate the mechanism of action. Additionally, TNB-585 efficacy was evaluated in vivo against C4-2 tumor-bearing NCG mice.ResultsIn vitro, TNB-585 induced activation and proliferation of human T cells resulting in the killing of PSMA+ prostate tumor cells in both 2D cultures and 3D spheroids with minimal cytokine release and reduced regulatory T cell activation compared with a positive control antibody that contains the same anti-PSMA arm but a higher affinity anti-CD3 arm (comparable with OKT3). In addition, TNB-585 demonstrated potent efficacy against patient-derived prostate tumors ex vivo and induced immune cell infiltration and dose-dependent tumor regression in vivo.ConclusionsOur data suggest that TNB-585, with its low-affinity anti-CD3, may be efficacious while inducing a lower incidence and severity of CRS in patients with prostate cancer compared with TCEs that incorporate high-affinity anti-CD3 domains.

scholarly journals 121 ICAM-1-specific affinity tuned CAR T cells expressing SSTR2 for real-time imaging

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A130-A130
Author(s):  
Jingmei Hsu ◽  
Eric von Hofe ◽  
Michael Hsu ◽  
Koen Van Besien ◽  
Thomas Fahey ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Phase I ◽  
Tumor Cells ◽  
Somatostatin Receptor ◽  
Laboratory Animals ◽  
Therapeutic Window ◽  
Car T Cells ◽  
Car T

BackgroundThe use of CAR T cells for solid tumors has a number of challenges, such as lack of tumor-specific targets, CAR T cell exhaustion, and the immunosuppressive tumor microenvironment. To address these challenges, AffyImmune has developed technologies to affinity tune and track CAR T cells in patients. The targeting moiety is affinity tuned to preferentially bind to tumor cells overexpressing the target while leaving normal cells with low basal levels untouched, thereby increasing the therapeutic window and allowing for more physiological T cell killing. The CAR T cells are designed to express SSTR2 (somatostatin receptor 2), which allows for the tracking of CAR T cells in vivo via PET/CT scan using FDA-approved DOTATATE.MethodsAIC100 was generated by affinity tuning the I-domain of LFA-1, the physiological ligand to ICAM-1. Various mutants with 106-fold difference in affinity were evaluated for affinity. This allowed structure activity relationships to be conducted using CAR T cells expressing the various affinity mutants against targets with varying antigen densities. The variant with micromolar affinity was clearly the most effective in non-clinical animal models. AIC100 is currently being evaluated to assess safety, CAR T expansion, tumor localization, and preliminary activity in patients with advanced thyroid cancer in a phase I study (NCT04420754). Our study uses a modified toxicity probability interval design with three dosage groups of 10 x 106, 100 x 106, and 500 x 106 cells.ResultsPreclinical studies demonstrated greater in vivo anti-tumor activity and safety with lower affinity CAR T cells. A single dose of AIC100 resulted in tumor elimination and significantly improved survival of animals. AIC100 activity was confirmed in other high ICAM-1 tumor models including breast, gastric, and multiple myeloma. In a Phase I patient given 10-million CAR T cells, near synchronous imaging of FDG and DOTATATE revealed preliminary evidence of transient CAR T expansion and tumor reduction at multiple tumor lesions, with the peak of CAR T density coinciding with the spike in CAR T numbers in blood.ConclusionsWe have developed affinity tuned CAR T cells designed to selectively target ICAM-1 overexpressing tumor cells and to spatiotemporally image CAR T cells. Near-synchronous FDG and DOTATATE scans will enhance patient safety by early detection of off-tumor CAR T activity and validation of tumor response. We anticipate that our ‘tune and track’ technology will be widely applicable to developing potent yet safe CAR T cells against hard-to-treat solid cancers.Trial RegistrationNCT04420754Ethics ApprovalIRB number19-12021154IACUC (animal welfare): All animal experiments were performed in accordance with the National Institute of Health’s Guide for the Care and Use of Laboratory Animals. Animal handling protocols were approved by the Institutional Laboratory Animal Use and Care Committee of Weill Cornell Medicine (Permit Number: 2012–0063).

The secreted matrix protein mindin increases prostate tumor progression and tumor-bone crosstalk via ERK 1/2 regulation

2019 ◽  
Vol 40 (7) ◽  
pp. 828-839
Author(s):  
Juan A Ardura ◽  
Irene Gutiérrez-Rojas ◽  
Luis Álvarez-Carrión ◽  
M Rosario Rodríguez-Ramos ◽  
José M Pozuelo ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Tumor Progression ◽  
Tumor Cells ◽  
Matrix Protein ◽  
Bone Cells ◽  
Prostate Tumor ◽  
Dependent Manner ◽  
Prostate Tumors ◽  
Adenocarcinoma Cells ◽  
Prostate Tumor Cells

Abstract Advanced prostate cancer cells preferentially metastasize to bone by acquiring a bone phenotype that allows metastatic cells to thrive in the skeletal environment. Identification of factors that promote the expression of ectopic bone genes—process known as osteomimicry—leading to tumor progression is crucial to prevent and treat metastatic prostate cancer and prolong life expectancy for patients. Here, we identify the extracelular matrix protein mindin in the secretome of prostate adenocarcinoma cells and show that mindin overexpression in human and mouse TRAMP-C1-induced prostate tumors correlates with upregulated levels of bone-related genes in the tumorigenic prostate tissues. Moreover, mindin silencing decreased osteomimicry in adenocarcinoma cells and in the prostate tumor mice model, as well as reduced tumor cell proliferation, migration and adhesion to bone cells. Inhibition of the extracellular signal-regulated kinase 1/2 (ERK 1/2) phosphorylation decreased the proliferative, migratory and pro-adhesion actions of mindin on prostate tumor cells. In addition, conditioned media obtained by crosstalk stimulation of either osteocytes or osteoblasts with the secretome of TRAMP-C1 cells promoted osteomimicry in prostate tumor cells; an effect inhibited by mindin silencing of TRAMP-C1 cells. In vivo, tibiae of primary tumor-bearing mice overexpressed the pro-angiogenic and pro-metastattic factor vascular endothelial growth factor receptor 2 (VEGFR2) in a mindin-dependent manner. Our findings indicate that mindin is a novel regulator of osteomimicry in prostate tumors and potentially mediates tumor-bone cell crosstalk, suggesting its promising role as a target to inhibit bone metastases.

Preclinical evaluation of AMG 160, a next-generation bispecific T cell engager (BiTE) targeting the prostate-specific membrane antigen PSMA for metastatic castration-resistant prostate cancer (mCRPC).

2019 ◽  
Vol 37 (7_suppl) ◽  
pp. 301-301 ◽  
Author(s):  
Julie Bailis ◽  
Petra Deegen ◽  
Oliver Thomas ◽  
Pamela Bogner ◽  
Joachim Wahl ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Half Life ◽  
Cell Activation ◽  
Phase 1 ◽  
Xenograft Model ◽  
Cytokine Release ◽  
Single Chain

301 Background: mCRPC is a disease of high unmet medical need, especially for patients who fail novel hormonal therapies and chemotherapy. BiTE molecules provide an off the shelf therapy that activates a patient’s own immune system and redirects T cells to kill tumor cells. The BiTE mechanism of action is distinct from other immunotherapies and may unlock immune response in mCRPC. PSMA is a compelling BiTE target that is highly expressed on PCa compared to normal tissue and has increased expression in mCRPC. Methods: AMG 160 is a fully human, half-life extended (HLE) BiTE that targets PSMA on tumor cells and CD3 on T cells. AMG 160 comprises two tandem single chain variable fragments fused to an Fc domain. Results: AMG 160 binds human and non-human primate (NHP) PSMA and CD3, leading to T cell activation and proliferation and cytokine production. AMG 160 redirects T cells to kill PSMA-positive cancer cell lines in vitro, including those with low PSMA levels or androgen-independent signaling. Weekly dosing of AMG 160 induces significant antitumor activity in established PCa xenograft model. The pharmacokinetics (PK) and pharmacodynamics of AMG 160 were tested in NHP. AMG 160 treatment led to BiTE target engagement in vivo, including transient T cell activation and cytokine release in blood, and mixed cellular infiltrates in multiple organs known to express PSMA. AMG 160 treatment was well tolerated. Cytokine release associated with the first dose could be attenuated using a step dose regimen. The half-life of AMG 160 in NHP was about one week. Based on allometric scaling, the PK profile of AMG 160 may be projected to enable dosing every other week in humans. Conclusions: AMG 160 is a potent HLE BiTE with specificity for PSMA-positive tumor cells. A Phase 1 study is planned to evaluate the safety and efficacy of AMG 160 in patients with mCRPC.

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