scholarly journals P06.03 Bispecific antibody-driven synthetic agonistic receptor engineered T cells lead to specific and conditional therapy in melanoma cancer models

2021 ◽  
Vol 9 (Suppl 1) ◽  
pp. A20.1-A20
Author(s):  
M Benmebarek ◽  
F Märkl ◽  
J Keyl ◽  
B Loureiro Cadilha ◽  
M Geiger ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumour Cell ◽  
Bispecific Antibody ◽  
Cell Lysis ◽  
Adoptive T Cell Therapy ◽  
Melanoma Cancer ◽  
Cancer Models ◽  
Overlap Extension

BackgroundImmunotherapeutic approaches, including immune checkpoint blockade and adoptive T cell therapy (ACT) in the form of tumor-infiltrating lymphocytes (TIL), have had marked success in the treatment of melanoma. Despite these successes, many patients are refractory to treatment or relapse with therapy-resistant disease. To overcome said limitations, we propose a controlled ACT approach, where T cells are armed with synthetic agonistic receptors (SAR) that are conditionally activated only in the presence of a target melanoma-associated antigen, and a cross-linking bispecific antibody (BiAb) specific for both SAR T cell and tumour cell.Materials and MethodsA SAR composed of an extracellular EGFRvIII, trans- membrane CD28, and intracellular CD28 and CD3z domains was fused via overlap- extension PCR cloning. T cells were retrovirally transduced to stably express our SAR construct. We validated our approach in two murine as well as two human cancer models expressing our melanoma-associated target antigens TYRP (murine) and MCSP (human). We confirmed conditional and specific stimulation and proliferation of our T cells, as well as their tumour-antigen-directed cytotoxicity, in vitro and in vivo.ResultsCrosslinking TYRP-EGFRvIII (murine) and MCSP-EGFRvIII (human) BiAb, monovalently selective for our SAR, induced conditional antigen-dependent activation, proliferation of SAR-T cells and directed tumour cell lysis with specificity towards two TYRP-expressing murine melanoma and two MCSP-expressing human melanoma cancer models. In vivo, anti-tumoural activity was mediated by the co-administration of SAR-T cells and BiAb, in A375 and MV3 melanoma xenograft models. Further, we could show that SAR T cells exhibited resistance to MDSC-induced suppression of activation and proliferation.ConclusionsHere we apply the SAR x BiAb approach in efforts to deliver specific and conditional activation of SAR transduced T cells, and targeted tumour cell lysis. The modularity of our platform is key for a targeting approach in a tumor entity with a high mutational load such as melanoma and is fundamental in our drive towards personalised immunotherapies. Further, the SAR approach has demonstrated resistance to MDSC-induced suppression, an interesting axis that requires further investigation.Disclosure InformationM. Benmebarek: None. F. Märkl: None. J. Keyl: None. B. Loureiro Cadilha: None. M. Geiger: None. C. Karches: None. S. Endres: None. C. Klein: None. S. Kobold: None. A. Klüver: None. M. Schwerdtfeger: None.

scholarly journals P06.01 Bispecific antibody-driven synthetic agonistic receptor – transduced T cells mediate specific and conditional therapy in melanoma cancer models

2020 ◽  
Vol 8 (Suppl 2) ◽  
pp. A41.2-A42
Author(s):  
M Benmebarek ◽  
J Keyl ◽  
F Märkl ◽  
M Geiger ◽  
C Karches ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumour Cell ◽  
Bispecific Antibody ◽  
Cell Lysis ◽  
Melanoma Cancer ◽  
Cancer Models ◽  
Overlap Extension ◽  
Part Time

BackgroundImmunotherapeutic approaches, including immune checkpoint blockade and adoptive T cell therapy (ACT) in the form of tumor-infiltrating lymphocytes (TILs), have had marked success in the treatment of melanoma. Despite these successes, many patients are refractory to treatment or relapse with therapy-resistant disease. To overcome these limitations, we propose a controlled ACT approach, where T cells are armed with synthetic agonistic receptors (SARs) that are conditionally activated only in the presence of a target melanoma-associated antigen, and a cross-linking bispecific antibody (BiAb) specific for both (SAR) T cell and tumour cell.Materials and MethodsA SAR composed of an extracellular EGFRvIII, trans- membrane CD28, and intracellular CD28 and CD3z domains was fused via overlap- extension PCR cloning. T cells were retrovirally transduced to stably express our SAR construct. We validated our approach in two murine as well as two human cancer models expressing our melanoma-associated target antigens TYRP (murine) and MCSP (human). We confirmed conditional and specific stimulation and proliferation of our T cells, as well as their tumour-antigen-directed cytotoxicity, in vitro and in vivo.ResultsCrosslinking TYRP-EGFRvIII (murine) and MCSP-EGFRvIII (human) BiAb, monovalently selective for our SAR, induced conditional antigen-dependent activation, proliferation of SAR-T cells and directed tumour cell lysis with specificity towards two TYRP-expressing murine melanoma and two MCSP-expressing human melanoma cancer models. In vivo, anti-tumoural activity was mediated by the co-administration of SAR-T cells and BiAb, in an A375 melanoma xenograft model. Further, overexpression of IDO (a key immunosuppressive enzyme implicated in the suppression of T cell function in the tumor microenvironment) in a melanoma model did not influence the killing kinetics of SAR T cells.ConclusionsHere we apply the SAR x BiAb approach in efforts to deliver specific and conditional activation of synthetic agonistic receptor transduced T cells, and targeted tumour cell lysis. The modularity of our platform is key for a targeting approach in a tumor entity with a high mutational load such as melanoma and is fundamental in our drive towards personalised immunotherapies. Further, the SAR approach has demonstrated resistance to IDO-mediated inhibition in the context of melanoma, an interesting axis that requires further investigation.Disclosure InformationM. Benmebarek: None. J. Keyl: None. F. Märkl: None. M. Geiger: A. Employment (full or part-time); Significant; Roche. C. Karches: None. S. Rausch: None. A. Gottschlich: None. A. Öner: None. M. Feinendegen: None. J. Dörr: None. B. Cadilha: None. S. Endres: None. C. Klein: A. Employment (full or part-time); Significant; Roche. S. Kobold: None.

scholarly journals P07.01 A modular and controllable T cell therapy platform for AML

2021 ◽  
Vol 9 (Suppl 1) ◽  
pp. A22.2-A23
Author(s):  
M Benmebarek ◽  
B Loureiro Cadilha ◽  
M Herrmann ◽  
S Schmitt ◽  
S Lesch ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Therapy ◽  
T Cell Activation ◽  
Tumour Cell ◽  
Cell Activation ◽  
Cell Lysis ◽  
Target Cells ◽  
Single Chain

BackgroundTargeted immunotherapies have shown limited success in the context of acute myeloid leukemia (AML). The mutational landscape, heterogeneity attributed to this malignancy and toxicities associated with the targeting of myeloid lineage antigens, it has become apparent that a modular and controllable cell therapy approach with the potential to target multiple antigens is required. We propose a controlled ACT approach, where T cells are equipped with synthetic agonistic receptors (SARs) that are selectively activated only in the presence of a target AML-associated antigen, and a cross-linking tandem single chain variable fragment (taFv) specific for both (SAR) T cell and tumour cell.Materials and MethodsA SAR composed of an extracellular EGFRvIII, trans- membrane CD28, and intracellular CD28 and CD3z domains was fused via overlap- extension PCR cloning. T cells were retrovirally transduced to stably express our SAR construct. SAR-specific taFvs that target AML-associated antigens were designed and expressed in Expi293FTM cells and purified by nickel affinity and size exclusion chromatography (SEC). We validated our approach in three human cancer models and patient-derived AML blasts expressing our AML-associated target antigens CD33 and CD123.ResultsAnti-CD33-EGFRvIII and anti-CD123 EGFRvIII taFv, monovalently selective for our SAR, induced conditional antigen-dependent activation, proliferation and differentiation of SAR-T cells. Further, SAR T cells bridged to their target cells by taFv could form functional immunological synapses, resulting in efficient tumor cell lysis with specificity towards CD33-expressing AML cells. SAR-taFv combination could also mediate specific cytotoxicity against patient-derived AML blasts and leukemic stem cells whilst driving SAR T cell activation. In vivo, treatment with SAR-taFv combination could efficiently eradicate leukemia and enhance survival in an AML xenograft models. Furthermore, we could show selective activation of SAR T cells, as well as a controllable reversibility and modularity of said activation upon depletion of the T cell engaging molecule, both in vitro and in vivo.ConclusionsHere we apply the SAR-taFv platform in efforts to deliver specific and conditional activation of SAR-transduced T cells, and targeted tumour cell lysis. The modularity of our platform will allow for a multi-targeting ACT approach with the potential to translate the ACT successes of B cell malignancies to AML. With a lack of truly specific AML antigens, it is invaluable that this approach possesses an intrinsic safety switch via its taFv facet. Moreover, we are able to circumvent pan-T cell activation due to the specific targeting and activation of SAR T cells.Disclosure InformationM. Benmebarek: None. B. Loureiro Cadilha: None. M. Herrmann: None. S. Schmitt: None. S. Lesch: None. S. Stoiber: None. A. Darwich: None. C. Augsberger: None. B. Brauchle: None. M. Schwerdtfeger: None. A. Gottschlich: None. A. Gottschlich Rataj: None. N.C. Fenn: None. C. Klein: None. M. Subklewe: None. S. Endres: None. K. Hopfner: None. S. Kobold: None.

scholarly journals IMMU-31. QUANTITATIVE EVALUATION OF ROUTES OF ADMINISTRATION OF IMMUNOTHERAPEUTIC T CELLS TO ORTHOTOPIC GLIOMA

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii111-ii111
Author(s):  
Lan Hoang-Minh ◽  
Angelie Rivera-Rodriguez ◽  
Fernanda Pohl-Guimarães ◽  
Seth Currlin ◽  
Christina Von Roemeling ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Growth ◽  
Ex Vivo ◽  
Adoptive T Cell Therapy ◽  
Whole Body ◽  
T Cell Therapy ◽  
Clinical Responses

Abstract SIGNIFICANCE Adoptive T cell therapy (ACT) has emerged as the most effective treatment against advanced malignant melanoma, eliciting remarkable objective clinical responses in up to 75% of patients with refractory metastatic disease, including within the central nervous system. Immunologic surrogate endpoints correlating with treatment outcome have been identified in these patients, with clinical responses being dependent on the migration of transferred T cells to sites of tumor growth. OBJECTIVE We investigated the biodistribution of intravenously or intraventricularly administered T cells in a murine model of glioblastoma at whole body, organ, and cellular levels. METHODS gp100-specific T cells were isolated from the spleens of pmel DsRed transgenic C57BL/6 mice and injected intravenously or intraventricularly, after in vitro expansion and activation, in murine KR158B-Luc-gp100 glioma-bearing mice. To determine transferred T cell spatial distribution, the brain, lymph nodes, heart, lungs, spleen, liver, and kidneys of mice were processed for 3D imaging using light-sheet and multiphoton imaging. ACT T cell quantification in various organs was performed ex vivo using flow cytometry, 2D optical imaging (IVIS), and magnetic particle imaging (MPI) after ferucarbotran nanoparticle transfection of T cells. T cell biodistribution was also assessed in vivo using MPI. RESULTS Following T cell intravenous injection, the spleen, liver, and lungs accounted for more than 90% of transferred T cells; the proportion of DsRed T cells in the brains was found to be very low, hovering below 1%. In contrast, most ACT T cells persisted in the tumor-bearing brains following intraventricular injections. ACT T cells mostly concentrated at the periphery of tumor masses and in proximity to blood vessels. CONCLUSIONS The success of ACT immunotherapy for brain tumors requires optimization of delivery route, dosing regimen, and enhancement of tumor-specific lymphocyte trafficking and effector functions to achieve maximal penetration and persistence at sites of invasive tumor growth.

scholarly journals PD-L1 Checkpoint Blockade Augments Anti-Tumor Immune Response of GD2 or HER2-Bsab Ex Vivo Armed T-Cells (EVAT) Therapy in Osteosarcoma

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1959-1959
Author(s):  
Jeong A Park ◽  
Hong fen Guo ◽  
Hong Xu ◽  
Nai-Kong V. Cheung
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Immune Checkpoint ◽  
Bispecific Antibody ◽  
Ex Vivo ◽  
Activated T Cells ◽  
The Impact ◽  
Anti Tumor Activity

Background Ex Vivo Armed T-cells (EVAT) carrying zeptomoles (10-21M) of T-cell engaging GD2-bispecific antibody (GD2-EVAT) or HER2-bispecific antibodies (HER2-EVAT) have potent anti-tumor activity against GD2(+) and/or HER2(+) solid tumors. Strategies to further optimize this approach are highly relevant. PD-1 is a key immune checkpoint receptor expressed mainly by activated T-cells and mediates immune suppression by binding to its ligands PD-L1 or PD-L2. Upregulation of PD-L1 has been found in many cancers including osteosarcoma and associated with aggressive disease and poor outcome. While the use of immune checkpoint inhibitors (ICIs) seems logical, the ideal timing when combined with T-cell engaging bispecific antibody (T-BsAb) or EVAT has yet to be defined. Here, we described the effects of anti-PD-1 or anti-PD-L1 antibodies on GD2-EVAT or HER2-EVAT therapy and explored the impact of its timing in the treatment of osteosarcoma which is GD2(+), HER2(+) and PD-L1(+). Methods GD2-BsAb and HER-BsAb were built using the IgG(L)-scFv format (Can Immunol Res, 3:266, 2015, Oncoimmunology, PMID:28405494). T-cells from healthy volunteer donors were isolated, and cultured ex vivo in the presence of CD3/CD28 beads plus 30 IU/mL of interleukin 2 (IL-2). Between day 7 and day 14, activated T-cells (ATCs) were harvested and armed for 20 minutes at room temperature with GD2-BsAb or HER2-BsAb. In vivo anti-tumor activity against GD2(+), HER2(+), and PD-L1(+) osteosarcoma cell line xenografts was tested in BALB-Rag2-/-IL-2R-γc-KO mice. Anti-human PD-1 antibody (pembrolizumab, anti-PD-1) or anti-human PD-L1 antibody (atezolizumab, anti-PD-L1) were tested for synergy with GD2-EVAT or HER2-EVAT therapy. Results The PD-1 expression increased among T-cells that circulated in the blood, that infiltrated the spleen or the tumor after EVAT therapy. While anti-PD-L1 combination therapy with GD2-EVAT or HER2-EVAT improved anti-tumor response against osteosarcoma (P=0.0123 and P=0.0004), anti-PD-1 did not (all P>0.05). The addition of anti-PD-L1 significantly increased T-cell survival in blood and T-cell infiltration of tumor when compared to GD2-EVAT or HER2-EVAT alone (all P<0.0001). Treatment of GD2-EVAT or anti-PD-L1 plus GD2-EVAT downregulated GD2 expression on tumors, but anti-PD-1 plus GD2-EVAT did not. For the next step we tested the impact of different combination schedules of ICIs on GD2-EVAT therapy. Concurrent anti-PD-1 (6 doses along with GD2-EVAT therapy) interfered with GD2-EVAT, while sequential anti-PD-1 (6 doses after GD2-EVAT) did not make a significant effect (P>0.05). On the other hand, while the concurrent use of anti-PD-L1 did not show benefit on GD2-EVAT, sequentially administered anti-PD-L1 produced a significant improvement in tumor control when compared to anti-PD-L1 or GD2-EVAT alone (P=0.002 and P=0.018). When anti-PD-L1 treatment was extended (12 doses after GD2-EVAT), the anti-tumor effect was most pronounced compared to GD2-EVAT alone (P <0.0001), which translated into improved survival (P=0.0057). These in vivo anti-tumor responses were associated with increased CD8(+) tumor infiltrating lymphocytes (TILs) of tumor. Conclusion In the arming platform, large numbers of target-specific T-cells can be generated, and this EVAT therapy is a highly effective cellular treatment with high potency in preclinical models. In addition, the advantage of ex vivo cytokine release following T-cell arming and activation could reduce or avoid life threatening cytokine storm if such activation was to proceed in vivo. Adoptive T-cell therapy induced immune response upregulates the inhibitory immune checkpoint PD-1/PD-L1 pathway, and combination treatment with anti-PD-L1 antibody, especially when combined as sequential therapy and continuously treated, significantly improved anti-tumor effect of EVAT, partly through increase in CD8(+) TILs infiltration. Disclosures Xu: MSK: Other: co-inventors in patents on GD2 bispecific antibody and HER2 bispecific antibody. Cheung:Ymabs: Patents & Royalties, Research Funding.

scholarly journals L4 Synthetic agonistic receptor-activating BiTEs – a modular platform for the efficient targeting of acute myeloid leukemia

2020 ◽  
Vol 8 (Suppl 2) ◽  
pp. A2.2-A3
Author(s):  
M Benmebarek ◽  
BL Cadilha ◽  
M Hermann ◽  
S Lesch ◽  
C Augsburger ◽  
...  
Keyword(s):  
T Cells ◽  
Acute Myeloid Leukemia ◽  
T Cell ◽  
T Cell Activation ◽  
Tumour Cell ◽  
Myeloid Leukemia ◽  
Cell Activation ◽  
Cell Lysis ◽  
Target Cells ◽  
Acute Myeloid

BackgroundTargeted immunotherapies have shown limited success in the context of acute myeloid leukemia (AML). Due to the mutational landscape and heterogeneity attributed to this malignancy and toxicities associated with the targeting of myeloid lineage antigens, it has become apparent that a modular and controllable cell therapy approach with the potential to target multiple antigens is required. We propose a controlled ACT approach, where T cells are armed with synthetic agonistic receptors (SARs) that are conditionally activated only in the presence of a target AML-associated antigen, and a cross-linking bispecific T cell engager (BiTE) specific for both (SAR) T cell and tumour cell.Materials and MethodsA SAR composed of an extracellular EGFRvIII, trans-membrane CD28, and intracellular CD28 and CD3z domains was fused via overlap-extension PCR cloning. T cells were retrovirally transduced to stably express our SAR construct. SAR-specific bispecific T cell engagers (BiTE) that target AML-associated antigens were designed and expressed in Expi293FTMcells and purified by nickel affinity and size exclusion chromatography (SEC). We validated our approach in three human cancer models and patient-derived AML blasts expressing our AML-associated target antigen CD33.ResultsCD33-EGFRvIII BiTE, monovalently selective for our SAR, induced conditional antigen-dependent activation, proliferation and differentiation of SAR-T cells. Further, SAR T cells bridged to their target cells by BiTE could form functional immunological synapses, resulting in efficient tumor cell lysis with specificity towards CD33-expressing AML cells. SAR.BiTE combination could also mediate specific cytotoxicity against patient-derived AML blasts whilst driving SAR T cell activation. In vivo, treatment with SAR.BiTE combination could efficiently eradicate leukemia and enhance survival in an AML xenograft model. Furthermore, we could show selective activation of SAR T cells, as well as a controllable reversibility of said activation upon depletion of the T cell engaging molecule.ConclusionsHere we apply the SAR x BiAb approach in efforts to deliver specific and conditional activation of agonistic receptor-transduced T cells, and targeted tumour cell lysis. The modularity of our platform will allow for a multi-targeting ACT approach with the potential to translate the ACT successes of B cell malignancies to AML. With a lack of truly specific AML antigens, it is invaluable that this approach possesses an intrinsic safety switch via its BiTE facet. Moreover, we are able to circumvent pan-T cell activation due to the specific targeting and activation of SAR T cells.Disclosure InformationM. Benmebarek: None. B.L. Cadilha: None. M. Hermann: None. S. Lesch: None. C. Augsburger: None. B. Brauchle: None. S. Stoiber: None. A. Darwich: None. F. Rataj: None. C. Klein: A. Employment (full or part-time); Significant; Roche. K. Hopfner: None. M. Subklewe: None. S. Endres: None. S. Kobold: None.

AMG 701 Potently Induces Anti-Multiple Myeloma (MM) Functions of T Cells and IMiDs Further Enhance Its Efficacy to Prevent MM Relapse In Vivo

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 135-135 ◽  
Author(s):  
Shih-Feng Cho ◽  
Liang Lin ◽  
Lijie Xing ◽  
Kenneth Wen ◽  
Tengteng Yu ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Growth ◽  
Cd4 T Cells ◽  
Xenograft Model ◽  
Cell Lysis ◽  
Cell Surface Expression ◽  
Effector Memory ◽  
T Effector Cells

AMG 701 is a half-life extended BiTE® (bispecific T-cell engager) targeting the B cell maturation antigen (BCMA). Here, we confirmed AMG 701-mediated T cell-redirected lysis of MM cells, defined immunomodulatory effects of AMG 701, and investigated combination potential of AMG 701 with immunomodulatory drugs (IMiDs) in human MM. Firstly, AMG 701 induced specific and efficacious T cell-dependent cytotoxicity (TDCC) against all MM cell lines tested, regardless of sensitivity to current anti-MM agents and expression levels of BCMA. AMG 701-induced TDCC was minimally affected in the presence of myeloma-supporting cells and cytokines in the bone marrow (BM) microenvironment, including osteoclasts (OCs), BM stromal cells (BMSCs), and a proliferation-inducing ligand (100 ng/ml). Importantly, AMG 701 induced lysis of autologous patient cells from the relapse and refractory stage of MM (RRMM). AMG 701 rapidly upregulated cell surface expression of CD107a and the production of IFNγ and TNFα, more so in CD8 than CD4 T subsets. It stimulated the proliferation and activation of T cells, to a greater extent in CD8 vs CD4 T cells, leading to significantly increased ratios of CD8/CD4 T cells. Significantly, AMG 701 induced differentiation of naive T cells (CD4 and CD8) to T cells with memory phenotype. This includes central memory (CM), effector memory (EM) T cells, and stem cell like memory cells. Time-course immunophenotyping studies showed that AMG 701 transiently upregulated the expression of key immune checkpoint and costimulatory markers on both CD4 and CD8 T cells. The induced T cells purified from ex vivo co-cultures still effectively lysed MM cells with lower BCMA levels. This may suggest an increased T cell clonality. Furthermore, IMiDs (len or pom) enhanced AMG 701-mediated TDCC against MM cells at earlier time points, lower E/T ratios, lower concentrations, or in the presence of immunosuppressive OCs or BMSCs. The combination AMG 701 and IMiDs maximized MM cell lysis accompanied with a decreased EC50 value. Combined treatments induce a more pronounced immunomodulation than AMG 701 alone in the presence of OCs, as evidenced by higher percentage of CM+EM and CD8/CD4 ratio at d8. AMG 701 with IMiDs combination significantly enhance AMG 701-mediated autologous patient MM cell lysis in a synergistic manner (combination index &lt; 1). In the human NCI-H929 xenograft model reconstituted with human T effector cells, AMG 701 effectively blocked tumor growth 5d after the first injection, regardless of doses (0.02-2 mg/kg). Tumors were completely eradicated following 3 separate injections in the host without weight loss. Next, sub-optimal doses and treatment schedules for AMG 701 and len were then used to investigate in vivo anti-MM effects by the combination vs monotherapy. Mice receiving MM cells were treated, from d15 until the end of the study, with len once daily, AMG 701 once weekly, or combination of AMG 701 and len. Two days after the first drug administration, all three treatments significantly inhibited MM tumor growth in mice (p&lt;0.001). Most importantly, while AMG 701 or len group showed tumor progress eventually, the combination of AMG 701 with len continuously suppressed tumor growth (p&lt;0.05 after d26; p&lt;0.001 after d40 for combination vs either agent alone). Combination of AMG 701 and len significantly induced superior MM cell regression, compared to either monotherapy, resulting in enhanced tumor regression and prevention of disease relapse. Taken together, these results strongly support AMG 701-based clinical studies, both as monotherapy (NCT03287908) and in combination with IMiDs to enhance elimination of residual diseases and prolong long-term durable responses in MM. Disclosures Munshi: Oncopep: Consultancy; Janssen: Consultancy; Abbvie: Consultancy; Takeda: Consultancy; Adaptive: Consultancy; Amgen: Consultancy; Celgene: Consultancy. Wahl:Amgen Research GmbH: Employment. Matthes:Amgen Research GmbH: Employment. Anderson:Sanofi-Aventis: Other: Advisory Board; Bristol-Myers Squibb: Other: Scientific Founder; Oncopep: Other: Scientific Founder; Amgen: Consultancy, Speakers Bureau; Janssen: Consultancy, Speakers Bureau; Takeda: Consultancy, Speakers Bureau; Celgene: Consultancy, Speakers Bureau. Chapman-Arvedson:Amgen Research: Employment.

A Novel Bioluminescent Mouse Model for Optimization of Adoptive T Cell Therapy

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2162-2162
Author(s):  
Martin Szyska ◽  
Stefanie Herda ◽  
Stefanie Althoff ◽  
Andreas Heimann ◽  
Tra My Dang ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Therapy ◽  
Tumor Model ◽  
Effector Function ◽  
Tumor Rejection ◽  
Adoptive T Cell Therapy ◽  
T Cell Therapy ◽  
The Impact

Abstract Adoptive T cell therapy (ATT) is a promising option for the treatment of solid cancers. However, various defense mechanisms acquired by the tumor during evolution prevent transferred T cells (TC) to unfold their full potential. A combination of ATT with accessory therapeutic approaches including checkpoint inhibition and targeted therapy could lift TC inhibition and efficiently shift the immune balance towards tumor rejection. An in-vivo analysis of the impact of combination strategies on the outcome of ATT would greatly enhance the search for an optimal accessory to ATT therapy. We generated the transgenic mouse line BLITC (bioluminescence imaging of T cells) expressing an NFAT (nuclear factor of activated T cell)-dependent Click-beetle luciferase (Na et. al, 2010) and a constitutive Renilla Luciferase, allowing us to monitor migration and activation of transferred TCs in vivo. In order to analyze crucial ATT parameters in a clinically relevant tumor model, BLITC mice were crossed to the two HY-TCR transgenic mice Marilyn (CD4: H-2Ab-Dby) and MataHari (CD8: H-2Db-Uty) to generate TCs that could be monitored for in-vivo infiltration, local activation and rejection of established (> 0,5 cm x 0,5 cm / ≥10 days growth) H-Y expressing MB49 tumors. In order to better reflect the clinical situation, we lymphodepleted tumor-bearing immunocompetent albino B6 mice with fludarabine (FLu) and/or cyclophosphamide (CTX) prior to ATT. Transferred TCs were FACSorted and injected after an optional culture expansion phase. As shown before for freshly injected tumor cells (Perez-Diez, 2007), we observed a superior response of tumor-antigen specific CD4+ TCs compared to CD8+ TCs against established tumors. Whereas 5*106 CD8+ T cells hardly attenuated tumor growth, even as few as 5000 H-Y TCR-transgenic CD4+ T cells rejected tumors in most mice, depending on the lymphodepleting treatment (Figure A - remission rates in parentheses). Tumor infiltration and activation of adoptively transferred TCs was monitored in-vivo by the respective bioluminescent reporters. Around day 4 and 6, CD4+ TCs migrated from tumor-draining lymph nodes into the tumor environment and persisted until rejection. Interestingly, activation of CD4+ TCs was only transient (between days 4 and 7) in all mice, independent of therapy outcome (in Figure B shown for refractory tumor). Whereas loss of activation signal during remission was correlated with tumor clearance and decline of effector function, in refractory tumors it suggests a rapid inactivation of infiltrating TCs by the tumor microenvironment. Our data indicate that the failure of tumor rejection is not caused by impaired peripheral expansion or tumor homing but rather by inhibition of TC effector function. Responsible mechanisms and counter-acting therapeutic interventions are the focus of ongoing studies. In summary, the BLITC reporter system facilitates analysis of therapeutic parameters for ATT in a well-established solid tumor model. Using BLITC mice for transduction with TCR or CAR expression cassettes could allow rapid monitoring of on-target as well as undesired off-target effects in virtually any tumor setting. Future experiments will focus on the beneficial effects of combination treatments on the activation of adoptively transferred TCs. Figure. Figure. Disclosures No relevant conflicts of interest to declare.

scholarly journals Engraftment of human central memory-derived effector CD8+ T cells in immunodeficient mice

Blood ◽  
2011 ◽  
Vol 117 (6) ◽  
pp. 1888-1898 ◽  
Author(s):  
Xiuli Wang ◽  
Carolina Berger ◽  
ChingLam W. Wong ◽  
Stephen J. Forman ◽  
Stanley R. Riddell ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Effector T Cells ◽  
Central Memory ◽  
Model Systems ◽  
Adoptive T Cell Therapy ◽  
Effector Memory ◽  
T Cell Therapy ◽  
Effector Cells

Abstract In clinical trials of adoptive T-cell therapy, the persistence of transferred cells correlates with therapeutic efficacy. However, properties of human T cells that enable their persistence in vivo are poorly understood, and model systems that enable investigation of the fate of human effector T cells (TE) have not been described. Here, we analyzed the engraftment of adoptively transferred human cytomegalovirus pp65-specific CD8+ TE cells derived from purified CD45RO+CD62L+ central memory (TCM) or CD45RO+CD62L− effector memory (TEM) precursors in an immunodeficient mouse model. The engraftment of TCM-derived effector cells (TCM/E) was dependent on human interleukin-15, and superior in magnitude and duration to TEM-derived effector cells (TEM/E). T-cell receptor Vβ analysis of persisting cells demonstrated that CD8+ TCM/E engraftment was polyclonal, suggesting that the ability to engraft is a general feature of TCM/E. CD8+ TEM/E proliferated extensively after transfer but underwent rapid apoptosis. In contrast, TCM/E were less prone to apoptosis and established a persistent reservoir of functional T cells in vivo characterized by higher CD28 expression. These studies predict that human CD8+ effector T cells derived from TCM precursors may be preferred for adoptive therapy based on superior engraftment fitness.

scholarly journals Melanoma reactive TCR-modified T cells generated without activation retain a less differentiated phenotype and mediate a superior in vivo response

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Siao-Yi Wang ◽  
Tamson V. Moore ◽  
Annika V. Dalheim ◽  
Gina M. Scurti ◽  
Michael I. Nishimura
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Viral Vectors ◽  
Cell Activation ◽  
Adoptive T Cell Therapy ◽  
In Vitro Assays ◽  
T Cell Therapy

AbstractAdoptive T cell therapy with T cell receptor (TCR)-modified T cells has shown promise in treating metastatic melanoma and other malignancies. However, studies are needed to improve the efficacy and durability of responses of TCR-modified T cells. Standard protocols for generating TCR-modified T cells involve activating T cells through CD3 stimulation to allow for the efficient transfer of tumor-reactive receptors with viral vectors. T cell activation results in terminal differentiation and shortening of telomeres, which are likely suboptimal for therapy. In these studies, we demonstrate efficient T cell transduction with the melanoma-reactive TIL1383I TCR through culturing with interleukin 7 (IL-7) in the absence of CD3 activation. The TIL1383I TCR-modified T cells generated following IL-7 culture were enriched with naïve (TN) and memory stem cell populations (TSCM) while maintaining longer telomere lengths. Furthermore, we demonstrated melanoma-reactivity of TIL1383I TCR-modified cells generated following IL-7 culture using in vitro assays and a superior response in an in vivo melanoma model. These results suggest that utilizing IL-7 to generate TCR-modified T cells in the absence of activation is a feasible strategy to improve adoptive T cell therapies for melanoma and other malignancies.

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