Shared target antigens on cancer cells and tissue stem cells: go or no-go for CAR T cells?

2016 ◽  
Vol 13 (2) ◽  
pp. 151-155 ◽  
Author(s):  
Andreas A. Hombach ◽  
Hinrich Abken
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
Cancer Cells ◽  
Car T Cells ◽  
Car T

105 A third-generation human GUCY2C-targeted CAR-T cell for colorectal cancer immunotherapy

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A116-A116
Author(s):  
Trevor Baybutt ◽  
Adam Snook ◽  
Scott Waldman ◽  
Jonathan Stem ◽  
Ellen Caparosa ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
T Cells ◽  
Cancer Cells ◽  
Inflammatory Cytokines ◽  
Department Of Defense ◽  
Thomas Jefferson ◽  
Third Generation ◽  
Car T Cells ◽  
Car T

BackgroundColorectal cancer (CRC) presents a significant public health burden, responsible for the second most cancer-related deaths in the United States, with an increasing incidence in young adults observed globally.1,2 While the blockade of immune checkpoints received FDA approval as a CRC therapeutic, only patients with microsatellite instability, accounting for 15% of sporadic cases, demonstrate partial or complete responses.3 We present a third-generation chimeric antigen receptor (CAR)-T cell directed towards the extracellular domain of the mucosal antigen guanylyl cyclase C (GUCY2C), which is over-expressed in 80% of CRC cases, as a therapeutic alternative for late stage disease. Here, we demonstrate that human GUCY2C CAR-T cells can selectively kill GUCY2C-expressing colorectal cancer cells in vitro and produce inflammatory cytokines in response to antigenic stimulation.MethodsPeripheral blood mononuclear (PBMCs) cells were isolated from leukoreduction filters obtained from the Thomas Jefferson University Hospital Blood Donor Center (IRB #18D.495). Magnetic Activated Cell Sorting (MACS) technology was used to negatively select pan-T cells (Miltenyi Biotec), followed by activation and expansion using anti-CD3, anti-CD28, and anti-CD2 coated microbeads (Miltenyi Biotec) and supplemented with IL-7 and IL-15 (Biological Resources Branch Preclinical Biologics Repository – NCI). T-cells were transduced with a lentiviral vector encoding the anti-GUCY2C CAR. Our CAR utilizes a single chain variable fragment of human origin directed towards the extracellular domain of GUCY2C, the CD28 hinge, transmembrane, and intracellular signaling domain (ICD), 4-1BB (CD137) ICD, and CD3ζ ICD. CAR-T cells were used for experiments between 10 to 14 days after activation in vitro using the xCELLigence real time cytotoxicity assay and intracellular cytokine staining.ResultsGUCY2C-directed CAR-T cells specifically lysed the GUCY2C-expressing metastatic CRC cell line T84, while the control CAR did not. GUCY2C-negative CRC cells were not killed by either. In addition to cell killing, GUCY2C-directed CAR-T cells of both the CD8+ and CD4+ co-receptor lineage produced the inflammatory cytokines IFN-γ and TNFα in response to GUCY2C antigen.ConclusionsWe demonstrate that human GUCY2C-directed CAR-T cells can selectively target GUCY2C-expressing cancer cells. We hypothesize that GUCY2C-directed CAR-T cells present a viable therapeutic option for metastatic CRC. In vivo animal models to examine this potential are currently on-going.AcknowledgementsThis work was supported by the Department of Defense Congressionally Directed Medical Research Programs (W81XWH-17-1-0299, W81XWH-191-0263, and W81XWH-19-1-0067) to AES and Targeted Diagnostic & Therapeutics to SAW. AES is also supported by a DeGregorio Family Foundation Award. SAW is supported by the National Institutes of Health (NIH) (R01 CA204881, R01 CA206026, and P30 CA56036), and the Department of Defense Congressionally Directed Medical Research Program W81XWH-17-PRCRP-TTSA. SAW and AES were also supported by a grant from The Courtney Ann Diacont Memorial Foundation. SAW is the Samuel M.V. Hamilton Professor of Thomas Jefferson University. JS, EC, and AZ were supported by an NIH institutional award T32 GM008562 for Postdoctoral Training in Clinical Pharmacology.Ethics ApprovalThis study was approved by the Thomas Jefferson University Institutional Review Board (IRB Control #18D.495) and the Institutional Animal Care and Use Committee (Protocol #01529).ReferencesSiegel RL, Miller KD, Jemal A. Cancer statistics, 2020. CA Cancer J Clin2020;70: 7–30. doi:10.3322/caac.21590Araghi M, Soerjomataram I, Bardot A, Ferlay J, Cabasag CJ, Morrison DS, et al. Changes in colorectal cancer incidence in seven high-income countries: a population-based study. Lancet Gastroenterol Hepatol 2019;4: 511–518. doi:10.1016/S2468-1253(19)30147-5Overman MJ, McDermott R, Leach JL, Lonardi S, Lenz H-J, Morse MA, et al. Nivolumab in patients with metastatic DNA mismatch repair-deficient or microsatellite instability-high colorectal cancer (CheckMate 142): an open-label, multicentre, phase 2 study. Lancet Oncol 2017;18: 1182–1191. doi:10.1016/S1470-2045(17)30422-9

scholarly journals CRISPR Screening of CAR T Cells and Cancer Stem Cells Reveals Critical Dependencies for Cell-Based Therapies

2020 ◽  
pp. CD-20-1243
Author(s):  
Dongrui Wang ◽  
Briana C Prager ◽  
Ryan C Gimple ◽  
Brenda Aguilar ◽  
Darya Alizadeh ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
Cancer Stem Cells ◽  
Car T Cells ◽  
Car T

Targeting glycosylated antigens on cancer cells using siglec‐7/9‐based CAR T‐cells

2020 ◽  
Vol 59 (7) ◽  
pp. 713-723 ◽  
Author(s):  
Sara Meril ◽  
Ortal Harush ◽  
Yishai Reboh ◽  
Tatyana Matikhina ◽  
Tilda Barliya ◽  
...  
Keyword(s):  
T Cells ◽  
Cancer Cells ◽  
Car T Cells ◽  
Car T

scholarly journals Visualization of human T lymphocyte-mediated eradication of cancer cells in vivo

2020 ◽  
Vol 117 (37) ◽  
pp. 22910-22919
Author(s):  
Xingkang He ◽  
Xin Yin ◽  
Jing Wu ◽  
Stina L. Wickström ◽  
Yanhong Duo ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cancer Cells ◽  
Cancer Patients ◽  
Metastatic Cancer ◽  
Car T Cells ◽  
Human T Cell ◽  
Car T ◽  
Metastatic Cancer Cells

Lymphocyte-based immunotherapy has emerged as a breakthrough in cancer therapy for both hematologic and solid malignancies. In a subpopulation of cancer patients, this powerful therapeutic modality converts malignancy to clinically manageable disease. However, the T cell- and chimeric antigen receptor T (CAR-T) cell-mediated antimetastatic activity, especially their impacts on microscopic metastatic lesions, has not yet been investigated. Here we report a living zebrafish model that allows us to visualize the metastatic cancer cell killing effect by tumor- infiltrating lymphocytes (TILs) and CAR-T cells in vivo at the single-cell level. In a freshly isolated primary human melanoma, specific TILs effectively eliminated metastatic cancer cells in the living body. This potent metastasis-eradicating effect was validated using a human lymphoma model with CAR-T cells. Furthermore, cancer-associated fibroblasts protected metastatic cancer cells from T cell-mediated killing. Our data provide an in vivo platform to validate antimetastatic effects by human T cell-mediated immunotherapy. This unique technology may serve as a precision medicine platform for assessing anticancer effects of cellular immunotherapy in vivo before administration to human cancer patients.

scholarly journals Patient-derived glioblastoma stem cells are killed by CD133-specific CAR T cells but induce the T cell aging marker CD57

Oncotarget ◽  
2014 ◽  
Vol 6 (1) ◽  
pp. 171-184 ◽  
Author(s):  
Xuekai Zhu ◽  
Shruthi Prasad ◽  
Simone Gaedicke ◽  
Michael Hettich ◽  
Elke Firat ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
T Cell ◽  
Cell Aging ◽  
Glioblastoma Stem Cells ◽  
Car T Cells ◽  
Car T

Choice of Better T-cells for Adoptive Immunotherapy

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. SCI-22-SCI-22 ◽  
Author(s):  
Dirk Hans Busch
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
B Cell ◽  
Adoptive Immunotherapy ◽  
Car T Cells ◽  
Car T ◽  
Selective Depletion

Abstract Adoptive transfer of primary (unmodified) or genetically engineered antigen-specific T cells has demonstrated astonishing clinical results in the treatment of infections and some malignancies. The definition of optimal targets and antigen receptors as well as the differentiation status of transferred T cells are emerging as crucial parameters for generating cell products with predictable efficacy and safety profiles. Our laboratory has demonstrated that defined subsets within the memory CD8+ T cell compartment fulfill all key characteristics of adult tissue stem cells and are essential for robust and long-term maintained responses upon adoptive transfer. We have developed clinical multi-parameter enrichment technologies to purify these memory stem cells for clinical applications. In my presentation I will report on the status of ongoing clinical trials using such purified cell products either as a primary T cell population for the treatment of infections upon allogeneic stem cell transplantation or after genetic modification with a CD19 CAR for the treatment of malignancies (collaboration with Stan Riddell, FHCC/Seattle). Infusing small numbers of T cells within a memory stem cell product can be highly effective therapeutically, but bears some risk of toxicity. Therefore, safeguards that allow selective depletion of transferred cells in the case of un-tolerable side effects may be needed to further improve adoptive immunotherapy. I will present results exploring the capacity of a truncated version of EGFR (EGFRt) co-expressed with T cells expressing a CD19-CAR. In pre-clinical mouse models we demonstrate that application of Cetuximab, which binds to EGFRt, confers selective depletion of adoptively transferred CAR-T cells in vivo. Long-term B cell aplasia, which is a main side effect of CD19-CAR T cell therapy, can be completely reverted with this strategy. Vaccination studies upon B cell recovery demonstrate full functionality of antigen-specific antibody formation. EGFRt co-expressing CD19-CAR T cells have been successfully transferred into first human patients, providing the option to test for the first time in a clinical setting whether treatment of B cell aplasia after long-term leukemia remission can be achieved by selective depletion. Disclosures Busch: STAGE cell therapeutics: Other: I was share holder of STAGE cell therapeutics, a company that was recently bought by Juno therapeutics.. Off Label Use: CD19 CAR T cells.

Interleukin-12 armored chimeric antigen receptor (CAR) T cells for heterogeneous antigen-expressing ovarian cancer.

2018 ◽  
Vol 36 (5_suppl) ◽  
pp. 12-12 ◽  
Author(s):  
Oladapo O. Yeku ◽  
Terence Purdon ◽  
David R. Spriggs ◽  
Renier J. Brentjens
Keyword(s):  
Ovarian Cancer ◽  
T Cells ◽  
T Cell ◽  
Cancer Cells ◽  
Chimeric Antigen Receptor ◽  
Second Generation ◽  
Antigen Expression ◽  
Interleukin 12 ◽  
Car T Cells ◽  
Car T

12 Background: Immune escape via downregulation of tumor associated antigens (TAAs) is an important mechanism of resistance to Chimeric Antigen Receptor (CAR) T cell therapy. Particularly in solid tumor malignancies where antigen expression could be heterogeneous, the risk of antigen-low or antigen-negative relapse is significantly high. One strategy to overcome this limitation is to reengineer CAR T cells to engage other arms of the immune system such as endogenous cytotoxic T cells and dendritic cells (DC) to broaden the antitumor response beyond the TAA targeted by CAR T cells. This could be achieved by co-modifying CAR T cells with Interleukin-12 (IL-12). IL-12 is a proinflammatory cytokine produced by DCs, and macrophages, and has been shown to promote maturation of DCs and increase T-cell proliferation. We hypothesized that CAR T cells genetically engineered to constitutively secrete IL-12 will be efficacious against Muc16ecto low (MLo) and Muc16ecto high (MHi) heterogeneous tumors in a syngeneic mouse model of ovarian peritoneal carcinomatosis. Methods: ID8 mouse ovarian cancer cells with either low endogenous Muc16ecto or transduced to express high levels of Muc16ecto were generated. Mouse T cells were transduced with plasmids encoding second generation Muc16 or Muc16/IL-12-directed CARs. C57BL/6 mice were inoculated i.p with tumor cells and subsequently treated with CAR T cells. Results: Second generation and IL-12 armored CAR T cells (4H1128?-IL12) were cytotoxic against both MLo and MHi cells in vitro. However, 4H1128?-IL12 were significantly more efficacious at killing both MLo and MHi cancer cells. In vivo, treatment with 4H1128?-IL12 led to significantly improved survival in mice inoculated with a 50:50 mix of MLo and MHi cells. Peritoneal washes performed on mice that succumbed to disease showed equivalent eradication of MLo and MHi. Treatment with 4H1128?-IL12 resulted in increased mature peritoneal DC’s (CD11b+ MHCII+). Finally, surviving mice from 4H1128?-IL12 cohorts were found to have increased T-cell receptor (TCR-β) productive clonality. Conclusions: IL-12-secreting CAR T cells are efficacious against tumors with low and heterogeneous antigen expression.

Preclinical evaluation of anti-ROR1 CAR T cells employing a ROR1 binding SCFV derived from the clinical stage mab cirmtuzumab.

2020 ◽  
Vol 38 (5_suppl) ◽  
pp. 41-41
Author(s):  
Charles E. Prussak ◽  
Christopher Oh ◽  
Juliana Velez Lujan ◽  
Sharon Lam ◽  
Jieyu Zhang ◽  
...  
Keyword(s):  
T Cells ◽  
Cancer Cells ◽  
In Vivo Studies ◽  
Tumor Escape ◽  
Cancer Stemness ◽  
2Nd Generation ◽  
Car T Cells ◽  
Car T

41 Background: Chimeric antigen receptor (CAR)-modified T cells (CAR-T) were generated targeting cells expressing ROR1, which is present on many malignant cancers and has been associated with cancer stemness and chemo-resistance. The ROR1 CAR utilizes the humanized single-chain fragment variable (scFv) binding domain of UC-961 (cirmtuzumab), which exhibits high affinity and specificity for human ROR1 and has demonstrated an excellent safety profile in Phase 1 studies. Methods: CAR constructs with varying spacer regions and intracellular co-stimulatory domains, using the scFV of cirmtuzumab, were constructed and used to generate CAR-T cells from healthy donors. These ROR1 CAR-T cells were tested for cytotoxicity against lymphoid cancer cells in vitro and in vivo studies that employed immune-deficient mice engrafted with labeled human leukemia cells MEC1 or MEC1-ROR1, which had been transfected to stably express ROR1. Results: The 2nd generation and 3rd generation CAR-T-cells with analogous spacer regions were comparably potent and selectively cytotoxic for cells bearing the ROR1 target antigen. But the 2nd generation CARs demonstrated greater potency in vitro even at low effector to target ratios. For the in vivo studies, mice received a single injection of ROR1 CAR-T cells or activated T cells from the same donor as a control. The ROR1 CAR-T cells rapidly cleared the leukemic cells from the animals, whereas animals receiving control T cells or no therapy quickly succumbed to progressive disease within 3 weeks. The administered CAR-T products remained highly active following administration and could be detected for ≥ 3 months without evidence for T cell exhaustion. Conclusions: The generated CAR-T cells utilizing constructs with the Fv of cirmtuzumab, a humanized mAb highly specific for ROR1, onco-embryonic surface antigen, effectively and selectively killed neoplastic cells bearing ROR1 both in vitro and in vivo. As ROR1 expression and signaling has been associated with cancer stemness and chemo-resistance utilizing ROR1 CAR-T therapy to target cancer cells might mitigate tumor escape. These data strongly support the rationale for continued development of our ROR1 CAR-T.

scholarly journals IKZF3 deficiency potentiates chimeric antigen receptor T cells targeting solid tumors

2021 ◽  
Author(s):  
tian chi ◽  
yan zou
Keyword(s):  
T Cells ◽  
Cancer Cells ◽  
Solid Tumors ◽  
Chimeric Antigen Receptor ◽  
Antigen Receptor ◽  
Xenograft Model ◽  
Cytokine Signaling ◽  
General Strategy ◽  
Car T Cells ◽  
Car T

Chimeric antigen receptor (CAR) T cell therapy has been successful in treating hematological malignancy, but solid tumors remain refractory. Here, we demonstrated that knocking out transcription factor IKZF3 in HER2-specific CAR T cells targeting breast cancer cells did not affect proliferation or differentiation of the CAR T cells in the absence of tumors, but markedly enhanced killing of the cancer cells in vitro and in a xenograft model. Furthermore, IKZF3 KO had similar effects on the CD133-specific CAR T cells targeting glioblastoma cells. AlphaLISA and RNA-seq analyses indicate that IKZF3 KO increased the expression of genes involved in cytokine signaling, chemotaxis and cytotoxicity. Our results suggest a general strategy for enhancing CAR T efficacy on solid tumors.

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