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Third-generation CAR-CD30 T-cell therapy for CD30-positive lymphomas

Feb 25, 2020

The two currently approved chimeric antigen receptor (CAR) T-cell therapies for the treatment of relapsed/refractory (R/R) large B-cell lymphoma (LBCL) —  axicabtagene ciloleucel and  tisagenlecleucel— have demonstrated significant clinical benefit in patients with R/R disease. 1,2These CAR T-cell therapies are CD19-directed, but other types of lymphoma, such as classical Hodgkin lymphoma (cHL) and anaplastic large cell lymphoma (ALCL), lack CD19 expression and so alternative CAR T-cell-based therapies are required. 3

CD30 is a cell membrane protein and a member of the tumor necrosis factor receptor superfamily which has been identified as an attractive target for CAR T-cell therapy. This is due to its expression on most cHLs and ALCLs, as well as in a proportion of other lymphomas, such as cutaneous T-cell lymphomas (CTCLs) and diffuse LBCLs (DLBCLs). 4

CD30 is a target of a number of antibody-based therapies. Most notably, positive results have been achieved with brentuximab vedotin (BV), an antibody drug conjugate targeting CD30 +cells, which is approved by the EMA for the treatment of cHL, ALCL, and CTCL. 5 However, there are limitations associated with BV treatment, such as inadequate persistence and tumor penetration. Consequently, anti-CD30 CAR T cells are being investigated for the treatment of CD30 +lymphomas. 4

Thus far, two phase I/II clinical trials ( NCT01316146and  NCT02259556) have investigated the potential use of anti-CD30 CAR T-cell therapy in R/R cHL or ALCL. Despite small cohort sizes, the two trials reported tolerable CD30 CAR T-cell regimens, however clinical benefit was sub-optimal. 6–8Preclinical advances of a third-generation, anti-CD30 CAR T-cell therapy were presented by Biagio De Angelis, Bambino Gesù Pediatric Hospital , IRCCS, Rome, IT,at the 2 ndEuropean CAR T Cell Meeting. 3 The data are based on results previously shown at the  61 stASH Annual Meeting & Expositionin Orlando, US. 8

Study design 9

  • A  comparative preclinical study evaluating the stability and anti-tumor activity of two CD30 CAR-T constructs

CAR design 3

  • A third-generation CAR construct containing an anti-CD30 single-chain variable-fragment cassette linked to CD3ζ by the signaling domains of either:
    • CD28.4-1BB

or

    • CD28.OX40
  • The vector incorporates an inducible caspase-9 (iCasp9) suicide gene
  • A CD34 antibody recognition domain was incorporated in the hinge region of all vectors as a selection marker

Methods 9

  • In vitro anti-tumor efficacy was evaluated using non-Hodgkin lymphoma (Karpas299) and cHL (L428) cell lines, a short-term cytotoxic assay (51Cr release assay), and long-term co-cultures
    • Supernatant from co-culture experiments was analyzed by enzyme-linked immunosorbent assays (ELISA)
  • In vivoNSG mouse models were engrafted i.v.with lymphoma FF-luciferase cell lines, Karpas299 or L428
  • In vivotumor growth was monitored using bioluminescent imaging
  • Tumor re-challenging was initiated by i.v.infusion of 0.2x106 Karpas299 cells into mice that survived until Day +140 and were observed for an additional 110 days

Results 8,9

  • High transduction rates were obtained with all constructs (80–90%)
  • Insertion of a CD34 antigen allowed for convenient selection and monitoring of transduced cells
  • Introduction of the iCasp9 suicide gene allowed controlled apoptosis of CD30 CAR T cells, irrespective of their co-stimulatory domains
  • Incorporation of a CD28.OX40 vsCD28.4-1BB costimulatory domain into a CD30-CAR resulted in a more stable CAR expression 30 days after transduction (84.72 ± 5.30% vs63.98 ± 11.51; p = 0.002)
  • CAR CD30-CD28.OX40 elicited greater anti-lymphoma properties in comparison to CAR CD30-CD28.4-1BB in vitro
    • The level of IFNγ (pg/ml/1e6) T cells was significantly higher in CD30-CD28.OX40 vsCD30-CD28.4-1BB transfected T cells
    • In long-term co-culture experiments with various cell lines, CD28.OX40 CD30 CAR T cells showed significantly higher anti-lymphoma activity compared to CD28.41BB CAR T cells when challenged at high tumor/effector ratio
    • Secretion of Th1 cytokines (IFNγ, IL-2, and TNFα) after antigen stimulation was significantly higher in CD30-CD28.OX40 vsCD30-CD28.4-1BB transfected T cells (p = 0.040, p = 0.008, and p = 0.02, respectively)
  • At the experimental endpoint (Day +165), overall survival was significantly improved in CD30-CD28.OX40 vsCD30-CD28.4-1BB CAR T-cell-treated mice (60% vs0%), p = 0.0021
  • Persistence of CD28.OX40 CAR T-cell established long-term immunological memory as demonstrated by second tumor challenge experiments in mouse tumor models

Conclusions 3

  • The iCasp9 system stands as a promising approach to decrease toxicities across CAR T-cell therapies
  • The CD30-CD28.OX40 CAR construct could overcome the limited efficacy of previously investigated CD30 CAR T-cell therapies in CD30 +lymphomas if the preclinical data translates to clinical studies

How can third generation anti-CD30 CARs improve treatment of CD30+ lymphomas?

  1.   S. Food and Drug Administration. YESCARTA (axicabtagene ciloleucel). Available at: https://www.fda.gov/vaccines-blood-biologics/cellular-gene-therapy-products/yescarta-axicabtagene-ciloleucel. [Accessed Feb 7, 2020]
  2.   S. Food and Drug Administration. KIMRIAH (tisagenlecleucel) . Available at: https://www.fda.gov/vaccines-blood-biologics/cellular-gene-therapy-products/kymriah-tisagenlecleucel. [Accessed February 7, 2020]
  3.   De Angelis B. A new promising third generation CAR-CD30 T-cell therapy for CD30+ lymphoma. 2020 Jan 30: 2nd European CAR T cell Meeting, Sitges, ES
  4.   Grover N.S. et al.Challenges of driving CD30-directed CAR-T-cells to the clinic.  BMC Cancer. 2019 Mar 6; 19:203. DOI: https://doi.org/10.1186/s12885-019-5415-9
  5.   European Medicines Agency. Adcetris, brentuximab vedotin. Available at: https://www.ema.europa.eu/en/medicines/human/EPAR/adcetris. [Accessed February 19, 2020]
  6. Ramos C.A. et al. Clinical and immunological responses after CD30-specific chimeric antigen receptor-redirected lymphocytes. J Clin Invest.2017 Sep 1; 127(9):3462–3471. DOI: 1172/JCI94306
  7.   Wang C.M. Autologous T-cells expressing CD30 chimeric antigen receptors for relapsed or refractory Hodgkin lymphoma: An open-label phase I trial. Clin Cancer Res.2017 Mar 1; 23(5):1156–1166. DOI: 1158/1078-0432.CCR-16-1365
  8. De Angelis B. et al. A new promising third generation CAR-CD30 T-cell therapy for CD30+ lymphoma.  Blood .2019 Nov 13; 134(Supplement_1):2069. DOI: https://doi.org/10.1182/blood-2019-127969
  9.   De Angelis B. et al. A new promising third generation CAR-CD30 T-cell therapy for CD30+ lymphoma. Poster presented at: 61st American Society of Hematology (ASH) Meeting & Exposition; 2019 Dec; Orlando, US