Preclinical In Vivo Modeling of Cytokine Release Syndrome Induced by ErbB-Retargeted Human T Cells: Identifying a Window of Therapeutic Opportunity?

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.

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Faculty Opinions recommendation of In vivo generation of human CD19-CAR T cells results in B-cell depletion and signs of cytokine release syndrome.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.734036173.793581845 ◽

2021 ◽

Author(s):

David Teachey

Keyword(s):

T Cells ◽

B Cell ◽

Cell Depletion ◽

B Cell Depletion ◽

Cytokine Release ◽

Cytokine Release Syndrome ◽

Car T Cells ◽

Car T

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Refractory Cytokine Release Syndrome in Recipients of Chimeric Antigen Receptor (CAR) T Cells

Blood ◽

10.1182/blood.v124.21.2296.2296 ◽

2014 ◽

Vol 124 (21) ◽

pp. 2296-2296 ◽

Cited By ~ 27

Author(s):

Noelle V. Frey ◽

Bruce L. Levine ◽

Simon F. Lacey ◽

Stephan A. Grupp ◽

Shannon L Maude ◽

...

Keyword(s):

T Cells ◽

Research Funding ◽

Disease Burden ◽

High Dose ◽

Cytokine Release ◽

Cytokine Release Syndrome ◽

Remission Rates ◽

Significant Disease ◽

Durable Remission

Abstract CTLO19 cells are CAR-modified T cells which recognize CD19 and produce high durable remission rates for pts with relapsed or refractory acute lymphoblastic leukemia (ALL). Cytokine Release Syndrome (CRS) has emerged as the major treatment related effect from CTL019, with symptoms that include high fevers and malaise but can progress to capillary leak, hypoxia and hypotension. CRS occurs hours to days after CTL019 infusion and correlates with rapid in vivo CTL019 expansion and marked elevation of serum IL6. In most cases, CRS is self-limited or rapidly reversed with anti-cytokine directed therapies. Here we report 3 cases of refractory CRS in adult pts with ALL. Our experience offers insight into clinical and investigational parameters describing this syndrome; highlights the variance of CRS across disease types and illustrates complexities of CRS management during concurrent infectious illness. As of 7/1/14, 97 pts (30 pediatric ALL, 12 adult ALL, 41 CLL, 14 NHL) have been treated with CTLO19. To capture clinical manifestations of CRS across protocols, we developed a novel CRS grading scale which will be described. Severe CRS (Gr 3-5) occurred in 27 (64%) of ALL pts and only 16 (29%) of CLL/NHL pts (p=0.001). 12 adults with ALL received CTL019; 8/9 evaluable pts achieved CR (MRD negative) at 1 month and 1 pt with extramedullary disease had marked reduction of PET avid disease which is maintained at 1 yr. Severe CRS occurred in 11 of 12 adult ALL pts. CRS was self-limited in 2 pts, rapidly reversed with anti-IL6 directed therapy in 6 pts and was refractory to therapy, contributing to death in 3 pts who were not evaluable for disease response. No baseline attributes differentiate these 3 pts from the 9 adult ALL pts with manageable Gr1-4 CRS. We have shown however that ALL disease burden correlates with CRS severity (in press) and all 3 pts had significant disease burden at baseline. All received lymphodepleting chemotherapy with cyclophosphamide 300 mg/m2 q12h x 6 followed by infusion of CTLO19 cells. These 3 pts each received 6.50E+06, 6.70E+06 and 8.45E+06 CTLO19 cells/kg compared to median CTL019 dose of 3.62E+06 in the 9 adult ALL pts with manageable CRS. Pt 21413-03 developed CRS 12 hrs after infusion and tested positive for influenza B on D3. Despite broad spectrum antimicrobials (including oseltamivir) and anticytokine directed therapy with tocilizumab (4mg/kg x 2) and steroids, he died with refractory hypotension on D5. Pt 21413-06 had extensive disease after 2 prior allogeneic SCTs and developed CRS within 12 hrs of infusion. In addition to broad spectrum antibiotics, she received tocilizumab 8mg/kg (D 3, 6 and 12); intermittent high dose steroids (D 4-15) and etanercept (D14). She died D15 with hypotension, hypoxic respiratory failure and concurrent MDR pseudomonas sepsis and pneumonia. Pt 21413-11 developed CRS within 24 hrs of infusion. He received tocilizumab 8mg/kg (D3&4); siltuximab (D5&15) and intermittent high dose steroids (D 4-15). After an initial response, he developed recurrent fever, pulmonary infiltrates and blood cultures positive for stenotrophomonas. He died D15 with refractory hypoxia and hypotension. All 3 pts’ clinical CRS correlated with marked in vivo CTL019 expansion and progressive serum cytokine elevations (data to be shown). CONCLUSIONS: CRS is the major toxicity of CTL019 therapy and its clinical course varies depending on disease type (more frequent and severe in ALL) and disease burden (in ALL). The 3 refractory CRS cases described here (of 97 total pts treated) have all occurred in adult ALL pts with significant disease burden who received relatively high doses of CTL019 cells. In addition, all 3 had significant infectious complications which potentially fueled underlying CRS and/or were made more virulent due to impairment of immunity with administration of anti-cytokine directed therapies. Future protocol modifications will be made goal of limiting severity of CRS while maintaining high durable remission rates. Further exploration is planned to better correlate timing and choice of anticytokine directed therapy in relation to clinical and investigation parameters of CRS. Disclosures Frey: Novartis: Research Funding. Off Label Use: USe of CART19 cells to treat CLL. Levine:Novartis: Patents & Royalties, Research Funding. Lacey:Novartis: Research Funding. Grupp:Novartis: Consultancy, Research Funding. Schuster:Novartis: Research Funding. Hwang:NVS: Research Funding. Leung:Novartis: Employment. Shen:Novartis: Employment. Ericson:Novartis: Employment. Melenhorst:Novartis: Research Funding. June:Novartis: Patents & Royalties, Research Funding. Porter:Novartis: Patents & Royalties, Research Funding.

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Chimeric Antigen Receptor T Cells Directed Against CD19 Induce Durable Responses and Transient Cytokine Release Syndrome in Relapsed, Refractory CLL and ALL

Blood ◽

10.1182/blood.v120.21.717.717 ◽

2012 ◽

Vol 120 (21) ◽

pp. 717-717 ◽

Cited By ~ 2

Author(s):

David L Porter ◽

Stephan A. Grupp ◽

Michael Kalos ◽

Alison W. Loren ◽

Lester Lledo ◽

...

Keyword(s):

T Cells ◽

Research Funding ◽

University Of Pennsylvania ◽

Cytokine Release ◽

Cytokine Release Syndrome ◽

Variable Effect ◽

To Receive ◽

Signaling Domains

Abstract Abstract 717 Background: Chimeric antigen receptors (CARs) combine the antigen recognition domain of an antibody with intracellular signaling domains into a single chimeric protein. CD19 is an ideal target for CARs since expression is restricted to normal and malignant B cells. Inclusion of the CD137 (4-1BB) signaling domain results in potent antitumor activity and in vivo persistence of anti-CD19 CARs in mice. We reported anti-tumor activity of CAR-modified autologous T cells targeted to CD19 (CART19 cells) in 3 patients (pts) with CLL with relatively short follow up (Porter, et al NEJM 2011; Kalos et al Sci Trans Med 2011). We now report on outcomes and longer follow up from 10 pts treated with CART19 cells. Methods: Autologous T cells collected by leukapheresis were transduced with a lentivirus encoding anti-CD19 scFv linked to 4-1BB and CD3-z signaling domains. Gene-modified T cells were expanded and activated ex-vivo by exposure to anti-CD3/CD28 beads. Pts had CLL or ALL with persistent disease after at least 2 previous treatments. Results: 10 pts have received CART19 cells; 9 adults median age 65 yrs (range 51–78) were treated for relapsed, refractory CLL and one 7 yr old was treated for relapsed refractory ALL. CLL pts had received a median of 5 prior regimens (range 2–10) and all had active disease at the time of infusion. 3/9 CLL patients had deletion of the p53 gene. The ALL pt had chemorefractory relapse, having received chemotherapy 6 weeks prior to infusion. All CLL pts received lymphodepleting chemotherapy 4–6 days before infusions (FC, PC or bendamustine, while the ALL pt had an ALC <10 after prior chemotherapy and did not require further lymphodepletion). A median of 7.5 × 108 total cells (range 1.7–50) corresponding to 1.45 × 108 (range 0.14–5.9) genetically modified cells were infused on day 0. Median follow-up as of 8/12/2012 was 5.6 mo (range 1–24 mo). 9 pts are evaluable for response (<30d follow up in 1 pt). No pt has died. There were no infusional toxicities >grade 2. CART19 homed to the marrow in the CLL pts and marrow and CSF for the ALL patient with detectable CART19 cells in the CSF (21 lymphs/uL, 78% CAR+) day 23 after infusion. 4/9 evaluable pts achieved CR. (3 CLL, 1 ALL). 2 CLL pts had a PR lasting 3 and 5 months, and 3 pts did not respond. In the 4 pts who achieved CR, maximal expanded cells in the blood were detected at an average of 27 fold higher than the infused dose (range 21–40-fold) with maximal in-vivo expansion between day 10 and 31 post infusion. No patient with CR has relapsed. All pts who responded developed a cytokine release syndrome (CRS) manifested by fever, and variable degrees of nausea, anorexia, and transient hypotension and hypoxia. In responding CLL pts the maximal fold elevation from baseline for IFN-γ was 89–298x, IL-6 6–40x, and IL2R 5– 25x, while no significant elevation in systemic levels of TNFα or IL2 were observed. For the ALL pt, maximal elevations from baseline were: IFNγ: 6040x; IL-6: 988x; IL2R: 56x, while significant elevations in TNFα (17x) and IL2 (163x) were also observed. The timing for maximum cytokine elevation differed but in all cases correlated with peak T cell expansion in the PBMC. 5 pts with CRS required treatment; patient 03 was treated with high dose steroids with resolution of symptoms but only achieved a PR. While steroid treatment had a variable effect on the CRS, we noted that these symptoms were temporally associated with significant elevations in serum IL-6. Accordingly, 4 of these pts were treated with the IL6-receptor antagonist tocilizumab on day 3–10 with prompt resolution of fevers, hypotension and hypoxia. 3 of these patients are evaluable for response and 2 achieved a CR. For the pts in CR, CART19 expression in the blood was documented by flow cytometry at the most recent follow up for each patient: 24 mo (pt 01), 22 mo (pt 02), 3 mo (pt 100), and 2 mo (pt 09). Conclusions: Autologous T cells genetically engineered to express an anti-CD19 scFv coupled to 4-1BB/CD3-z signaling domains can undergo robust in-vivo expansion, persist for at least up to 2 yrs, and can be associated with a significant CRS that responds to anti-cytokine therapy. CART19 cells can induce potent and sustained responses (6/9 responses, 4 CR) for patients with advanced, refractory and high risk CLL and relapsed refractory ALL. Disclosures: Porter: Novatis: Patents & Royalties; Celgene: Honoraria; Genentech: Employment; Pfizer: Research Funding. Off Label Use: The use of CART19 cells to treat CD19+ malignancy and the use of tocilizumab to treat cytokine activation syndrome related to CART19 cells. Kalos:University of Pennsylvania: Employment, Patents & Royalties. Levine:TxCell: Consultancy, Membership on an entity's Board of Directors or advisory committees; University of Pennsylvania: financial interest due to intellectual property and patents in the field of cell and gene therapy. Conflict of interest is managed in accordance with University of Pennsylvania policy and oversight Patents & Royalties. June:Novartis: Research Funding, entitled to receive royalties from patents licensed to Novartis, entitled to receive royalties from patents licensed to Novartis Patents & Royalties.

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Development of CAR-T Cell Constructs with Broad Anti-Tumor Tropism

Blood ◽

10.1182/blood-2020-136704 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 24-24

Author(s):

Ameet K. Mishra ◽

Iris Kemler ◽

David Dingli

Keyword(s):

Multiple Myeloma ◽

T Cells ◽

T Cell ◽

Cytokine Release ◽

Cytokine Release Syndrome ◽

Car T Cells ◽

Car T Cell ◽

Nsg Mice ◽

Human T Cells ◽

Car T

Chimeric antigen receptor T (CAR-T) cell therapy is a transformative approach to cancer eradication. CAR-T is expensive in part due to the restricted use of each CAR construct for a specific set of tumors such as B cell lymphoma targeted with CD19 and multiple myeloma targeted with BCMA. A CAR construct with broad anti-tumor activity can be advantageous due to wide applicability and scalability of production. We show that CD126, the IL-6 receptor alpha, is an antigen that is expressed by many hematologic and solid malignancies including multiple myeloma, non-Hodgkin lymphoma, acute myeloid leukemia, pancreatic and prostate adenocarcinoma, non-small cell lung cancer and malignant melanoma amongst others. High CD126 expression is a negative prognostic marker in many malignancies. The two CD126 targeting CAR-T cell constructs contain the CD28 anchoring domain followed by 4-1BB and CD3 zeta signaling domain. Lentiviral vectors were generated with triple plasmid (CAR, psPAX2 and pMD2.G) transfection of 293T cells and the vector concentrated by ultracentrifugation and used to transduce human T cells. T cells were isolated from leuko-reduction cones using negative selection with magnetic beads. The transduction efficiency was around 60%. The T cells were activated with anti-CD3/CD28 beads and expanded for two weeks before using for downstream experiments. CD126 CAR-T cells are able to kill many tumor cells in an antigen specific manner and with an efficiency that is directly proportional to the cell surface expression of CD126 expression (rho = 0.6, p = 0.0019). The presence of soluble CD126 in the culture media did not interfere with CAR-T cell killing. The CAR-T constructs bind murine CD126. However, injection of CD126 targeting CAR-T cells in NSG mice did not lead to any evidence of hepatotoxicity and weight loss despite possible expression of this antigen on hepatocytes. In vivo studies in NSG mice with multiple myeloma (RPMI-8226) and prostate adenocarcinoma (DU-145) xenograft models (n=10 tumors per group) showed that the intravenously injected CD126 targeted CAR-T cells (107) infiltrated the tumors, expanded, produced human interferon gamma and killed the tumor cells (p<0.001). Bioluminescence imaging showed control of tumor growth in the actively treated tumors compared to the controls (p<0.05). At post mortem, mice injected with CD126 targeted CAR-T cells had smaller residual tumors compared to controls injected with non-engineered human T cells from the same donor. Binding of sIL-6R by CAR-T cells could mitigate cytokine release syndrome. In support of this, murine SAA-3 levels (the equivalent of human CRP) were lower in mice injected with CD126 CAR-T compared to controls (p<0.05), suggesting that binding of sIL-6R by CAR-T cells could mitigate cytokine release syndrome. CD126 provides a novel therapeutic for CAR-T cells in a broad variety of tumors with low risk of toxicity. Disclosures Dingli: Apellis: Consultancy; Millenium: Consultancy; Janssen: Consultancy; Bristol Myers Squibb: Research Funding; Sanofi-Genzyme: Consultancy; Alexion: Consultancy; Rigel: Consultancy; Karyopharm Therapeutics: Research Funding.

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Upregulation of FcγRIIb on monocytes is necessary to promote the superagonist activity of TGN1412

Blood ◽

10.1182/blood-2014-08-593061 ◽

2015 ◽

Vol 125 (1) ◽

pp. 102-110 ◽

Cited By ~ 27

Author(s):

Khiyam Hussain ◽

Chantal E. Hargreaves ◽

Ali Roghanian ◽

Robert J. Oldham ◽

H. T. Claude Chan ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Activation ◽

Cell Activation ◽

Close Association ◽

Lymphoid Tissues ◽

Cytokine Release ◽

Cytokine Release Syndrome ◽

Key Points

Key Points TGN1412-induced T-cell activation following high-density preculture of PBMCs is a consequence of FcγRIIb upregulation on monocytes. In vivo, cytokine release syndrome may be due to the close association of FcγRIIb-bearing cells with T cells in lymphoid tissues.

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Monocytes Are Required for Both Optimal Anti-Leukemic Efficacy and the Cytokine Release Syndrome By CAR-T Cells: Lessons from an Innovative Xenotolerant Mouse Model

Blood ◽

10.1182/blood.v128.22.997.997 ◽

2016 ◽

Vol 128 (22) ◽

pp. 997-997

Author(s):

Margherita Norelli ◽

Monica Casucci ◽

Barbara Camisa ◽

Laura Falcone ◽

Catia Traversari ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Cytokine Release ◽

Cytokine Release Syndrome ◽

Car T Cells ◽

Car T Cell ◽

Car T ◽

Secondary Transfer

Abstract Background: Chimeric antigen-receptor (CAR)-engineered T cells promise to cure chronic and acute leukemias refractory to standard treatments. Before this promise is fulfilled, however, two crucial issues need to be solved: i) how to circumvent the emergence of secondary resistance (e.g. due totarget-antigen loss; leukemic lineage switch); ii) how to manage associated toxicities (e.g. the cytokine release syndrome, CRS; lineage aplasias). Unfortunately, all these issues cannot be addressed pre-clinically in currently available NSG mouse models, because they lack human hematopoiesis and, furthermore, ultimately develop xenograft-versus-host disease (X-GVHD), preventing the evaluation of long-term effects. Methods: We have developed an innovative xenotolerant model by transplanting human hematopoietic stem cells (HSCs) intraliver in newborn NSG mice triple transgenic for human SCF, GM-SCF and IL-3 (SGM3). Differently from "classical" NSG, SGM3 mice reconstituted high levels of human T cells (>1000 cells per microL at week 8), which, once transferred in secondary recipients, persisted up to 200d without causing X-GVHD, even after irradiation. Robust and specific xenotolerance was confirmed by in vitrohyporesponsiveness to NSG, bot not to C57/Bl6 antigens (irradiated splenocytes) or human HLAs (PBMCs). Secondary transfer experiments in leukemic and/or HSC-humanized SGM-3 mice have been then designed for studying the determinants of CAR-T cell efficacy and associated toxicities in the absence of confounding xenoreactivity. Results: SGM3-derived T cells were transduced ex vivo with either a CD19 or a CD44v6 CAR (both having a CD28 2G design) after activation with CD3/CD28-beads and IL-7/IL-15, resulting in a preferential and functional CD45RA+/CD62L+/CD95+ stem memory T cell (TSCM) phenotype. Once transferred in secondary recipients previously engrafted with a CD19+/CD44v6 leukemic cell line, CD19 or CD44v6 CAR-T cells equally mediated rapid tumor clearance both in low and high tumor-burden settings, in the absence of malaise or elevated human IL-6 levels in vivo. At later time points (after 100d), however, approximately 50% of responding mice relapsed despite significant CAR-T cell persistence in vivo (>50 cells per microL). A significant fraction of leukemia relapses were characterized by post-transcriptional down-regulation of CD44v6 expression or CD19 loss, respectively. Conversely, secondary transfer of SGM3-derived CAR-T cells in leukemic SGM3 mice that had been previously humanized with HSCs resulted in the development of a clinical syndrome similar to the CRS observed in clinical trials (high fevers, elevated IL-6, TNF-alpha and serum amyloid A levels - mouse analog of C-reactive protein in humans), resulting in 30% lethality. This CRS was anticipated and shortened for CD44v6 compared with CD19 CAR-T cells and worse in the case of 4-1BB compared with the original CD28 2G CAR designs. Strikingly, mice recovering from the CRS benefited from durable leukemic remissions, yet experienced long-lasting CD19+ B-cell or CD44v6+ monocyte aplasias. Deepness of remission was confirmed in "tertiary" recipients, which did not develop leukemia after the infusion of bone-marrow cells from mice in remission 150d since CAR-T cell infusion. Interestingly, in this model, tocilizumab administration at the time of either CD19 or CD44v6 CAR-T cell infusion efficiently prevented the CRS, but did not interfere with their comparable and long-term anti-leukemic effects. Conversely, depleting monocytes/macrophages before therapeutic CAR-T cell infusion by either lyposomal clodronate or by the prophylactic CD44v6 CAR-T cells inhibited CRS development, but also resulted in significantly worse leukemia-free survival (at 250d, 0% vs 80%, P<0.0001). Conclusions: A number of lessons can be learned from this innovative xenotolerant mouse model of CAR-T cell immunotherapy: monocytes are required for both i) optimal anti-leukemic efficacy, and ii) the occurrence of CRS; iii) tocilizumab prevents the CRS without interfering with efficacy; iv) monocyte aplasia induced by CD44v6 CAR-T cells does not impact on their efficacy, at least in the theraeputic setting, and may ameliorate CRS toxicity. As for CD44v6 CAR-T cells, this model could be used for effectively predicting the efficacy and associated toxicities of new CAR-T cell therapies, speeding up their clinical development. Disclosures Traversari: MolMed SpA: Employment. Bordignon:MolMed SpA: Employment. Ciceri:MolMed SpA: Consultancy. Bonini:TxCell: Membership on an entity's Board of Directors or advisory committees; Molmed SpA: Consultancy. Bondanza:Formula Pharmaceuticals: Honoraria; TxCell: Research Funding; MolMed SpA: Research Funding.

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627 The DLL3-targeted half-life extended bispecific T cell engager (HLE BiTE®) immune-oncology therapy AMG 757 has potent antitumor activity in neuroendocrine cancer

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-sitc2020.0627 ◽

2020 ◽

Vol 8 (Suppl 3) ◽

pp. A663-A663

Author(s):

Keegan Cooke ◽

Juan Estrada ◽

Jinghui Zhan ◽

Jonathan Werner ◽

Fei Lee ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Tumor Growth ◽

Mouse Models ◽

T Cell Activation ◽

Phase 1 ◽

Phase 1 Study ◽

Human T Cells

BackgroundNeuroendocrine tumors (NET), including small cell lung cancer (SCLC), have poor prognosis and limited therapeutic options. AMG 757 is an HLE BiTE® immune therapy designed to redirect T cell cytotoxicity to NET cells by binding to Delta-like ligand 3 (DLL3) expressed on the tumor cell surface and CD3 on T cells.MethodsWe evaluated activity of AMG 757 in NET cells in vitro and in mouse models of neuroendocrine cancer in vivo. In vitro, co-cultures of NET cells and human T cells were treated with AMG 757 in a concentration range and T cell activation, cytokine production, and tumor cell killing were assessed. In vivo, AMG 757 antitumor efficacy was evaluated in xenograft NET and in orthotopic models designed to mimic primary and metastatic SCLC lesions. NSG mice bearing established NET were administered human T cells and then treated once weekly with AMG 757 or control HLE BiTE molecule; tumor growth inhibition was assessed. Pharmacodynamic effects of AMG 757 in tumors were also evaluated in SCLC models following a single administration of human T cells and AMG 757 or control HLE BiTE molecule.ResultsAMG 757 induced T cell activation, cytokine production, and potent T cell redirected killing of DLL3-expressing SCLC, neuroendocrine prostate cancer, and other DLL3-expressing NET cell lines in vitro. AMG 757-mediated redirected lysis was specific for DLL3-expressing cells. In patient-derived xenograft and orthotopic models of SCLC, single-dose AMG 757 effectively engaged human T cells administered systemically, leading to a significant increase in the number of human CD4+ and CD8+ T cells in primary and metastatic tumor lesions. Weekly administration of AMG 757 induced significant tumor growth inhibition of SCLC (figure 1) and other NET, including complete regression of established tumors and clearance of metastatic lesions. These findings warranted evaluation of AMG 757 (NCT03319940); the phase 1 study includes dose exploration (monotherapy and in combination with pembrolizumab) and dose expansion (monotherapy) in patients with SCLC (figure 2). A study of AMG 757 in patients with neuroendocrine prostate cancer is under development based on emerging data from the ongoing phase 1 study.Abstract 627 Figure 1AMG 757 Significantly reduced tumor growth in orthotopic SCLC mouse modelsAbstract 627 Figure 2AMG 757 Phase 1 study designConclusionsAMG 757 engages and activates T cells to kill DLL3-expressing SCLC and other NET cells in vitro and induces significant antitumor activity against established xenograft tumors in mouse models. These preclinical data support evaluation of AMG 757 in clinical studies of patients with NET.Ethics ApprovalAll in vivo work was conducted under IACUC-approved protocol #2009-00046.

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