CD19-specific Chimeric Antigen Receptor Research Articles

Treatment of Testicular Relapse of B-cell Acute Lymphoblastic Leukemia With CD19-specific Chimeric Antigen Receptor T Cells

BackgroundIrradiation has been a standard treatment for testicular relapse but is associated with severe hypogonadism. Because CD19-specific chimeric antigen receptor T (CAR-T) cells can eradicate leukemic blasts in cerebrospinal fluid, a pharmacologic sanctuary site, we tested the efficacy of this therapy in 7 boys with isolated testicular relapse of B-cell acute lymphoblastic leukemia. Patients and MethodsCD19-specific CAR-T cells were generated with the use of autologous T cells transduced with a lentiviral vector to express a CAR molecule containing anti-CD19 scFv derived from the HI19α murine monoclonal antibody, human CD8α hinge, and human 4-1BB (CD137) and CD3ζ costimulatory signaling transmembrane domains. After the conditioning regimen, which consisted of intravenous fludarabine and intravenous cyclophosphamide, 7 patients with a median age of 9 years (range, 2-10 years) with isolated testicular relapse received a single infusion of CD19 CAR-T cells at a total dose of 5 × 106 all T cells per kilogram. ResultsAll 7 patients achieved complete remission with normal testes. Six patients remained in second remission for 5 to 23 months (median, 14 months), and 1 patient subsequently relapsed in the bone marrow. The probability of event-free survival for all patients at 12 months of follow-up was 83.3% ± 15.2% (standard error). The treatment was well-tolerated, with grade 1 cytokine-release syndrome developing in 5 patients. ConclusionThese results suggest that CAR-T cell therapy is a treatment option for patients with testicular relapse.

Establishment of a simple and efficient platform for car-t cell generation and expansion: from lentiviral production to in vivo studies

IntroductionAdoptive transfer of T cells expressing a CD19-specific chimeric antigen receptor (CAR) has shown impressive response rates for the treatment of CD19 + B-cell malignancies in numerous clinical trials. The CAR molecule, which recognizes cell-surface tumor-associated antigen independently of human leukocyte antigen (HLA), is composed by one or more signaling molecules to activate genetically modified T cells for killing, proliferation, and cytokine production. ObjectivesIn order to make this treatment available for a larger number of patients, we developed a simple and efficient platform to generate and expand CAR-T cells. MethodsOur approach is based on a lentiviral vector composed by a second-generation CAR that signals through a 41BB and CD3-ζ endodomain. ConclusionsIn this work, we show a high-level production of the lentiviral vector, which was successfully used to generate CAR-T cells. The CAR-T cells produced were highly cytotoxic and specific against CD19+ cells in vitro and in vivo, being able to fully control disease progression in a xenograft B-cell lymphoma mouse model. Our work demonstrates the feasibility of producing CAR-T cells in an academic context and can serve as a paradigm for similar institutions. Nevertheless, the results presented may contribute favoring the translation of the research to the clinical practice.

The CNS can be a safe space for CARs

In this issue of Blood, Frigault et al report that tisagenlecleucel, a CD19-specific chimeric antigen receptor (CAR) T-cell product, has a safety profile in a small series of patients with aggressive B-cell non-Hodgkin lymphomas (B-NHLs) and secondary central nervous system (CNS) involvement similar to that seen in patients without active CNS disease. 1 The Frigault et al study suggests that this product may benefit this previously excluded group of patients without an increased risk of neurotoxicity.

CD19-Directed CAR T Cells Treatment in a Patient With Refractory DLBCL and CNS Involvement

ABSTRACT Introduction: Diffuse large B-cell lymphoma (DLBCL) is the most common neoplasm of the lymphatic system. Treatment and clinical management are difficult in the relapsed/refractory (R/R) setting. Chimeric antigen receptor (CAR) T cells are genetically engineered using autologous patient lymphocytes and have shown very promising results in the treatment of relapsed and refractory cases of DLBCL. Methods: A 64-year-old male patient with refractory DLBCL and central nervous system (CNS) involvement after 9 lines of therapy was treated with CD19-specific CAR T cell therapy at the Department of medical oncology and hematology at the University Hospital of Zurich and followed-up for 10 weeks. Results: Autologous lymphocytes were successfully harvested and transfected/expanded for CAR T cell production. Conditioning chemotherapy and CAR T infusion was well tolerated. Post-infusion side effects were mild (cytokine release syndrome [CRS] grade 1−2), with limited signs of neurotoxicity. Ten weeks after CAR T cell therapy, an excellent response could be documented via PET-CT. The CNS lesion disappeared as assessed via cranial MRI. Conclusion: CD19-targeted CAR T cell therapy is a revolutionary treatment option for heavily pre-treated R/R DLBCL even in the setting of CNS involvement.

CD19 CAR T cells following autologous transplantation in poor-risk relapsed and refractory B-cell non-Hodgkin lymphoma

AbstractHigh-dose chemotherapy and autologous stem cell transplantation (HDT-ASCT) is the standard of care for relapsed or primary refractory (rel/ref) chemorefractory diffuse large B-cell lymphoma. Only 50% of patients are cured with this approach. We investigated safety and efficacy of CD19-specific chimeric antigen receptor (CAR) T cells administered following HDT-ASCT. Eligibility for this study includes poor-risk rel/ref aggressive B-cell non-Hodgkin lymphoma chemosensitive to salvage therapy with: (1) positron emission tomography–positive disease or (2) bone marrow involvement. Patients underwent standard HDT-ASCT followed by 19-28z CAR T cells on days +2 and +3. Of 15 subjects treated on study, dose-limiting toxicity was observed at both dose levels (5 × 106 and 1 × 107 19-28z CAR T per kilogram). Ten of 15 subjects experienced CAR T-cell–induced neurotoxicity and/or cytokine release syndrome (CRS), which were associated with greater CAR T-cell persistence (P = .05) but not peak CAR T-cell expansion. Serum interferon-γ elevation (P < .001) and possibly interleukin-10 (P = .07) were associated with toxicity. The 2-year progression-free survival (PFS) is 30% (95% confidence interval, 20% to 70%). Subjects given decreased naive-like (CD45RA+CCR7+) CD4+ and CD8+ CAR T cells experienced superior PFS (P = .02 and .04, respectively). There was no association between CAR T-cell peak expansion, persistence, or cytokine changes and PFS. 19-28z CAR T cells following HDT-ASCT were associated with a high incidence of reversible neurotoxicity and CRS. Following HDT-ASCT, effector CD4+ and CD8+ immunophenotypes may improve disease control. This trial was registered at www.clinicaltrials.gov as #NCT01840566.

Abstract 2320: Avadomide (CC-122) increases effector function and reverses exhaustion in chronically stimulated lisocabtagene maraleucel anti-CD19 CAR T drug product

Abstract Lisocabtagene maraleucel (liso-cel) is an investigational drug product composed of autologous CD8+ and CD4+ T cells expressing a CD19-specific chimeric antigen receptor (CAR), being evaluated in clinical trials for treatment of B cell malignancies. The CELMoD CC-122 is being evaluated in non-Hodgkin lymphoma (NHL) and can augment T cell function through Cereblon-mediated degradation of Ikaros and Aiolos. Combination approaches aimed at enhancing CAR T cell function may increase the rate, depth, and durability of clinical responses. Here we report pre-clinical results supporting the potential combination of CC-122 an liso-cel. To determine potentiating effects of CC-122 following acute activation (anti-idiotypic Ab or CD19+ targets), T cells expressing the CAR of liso-cel were cultured with CC-122 and assessed for intracellular signaling, cytolytic activity, cytokine production, and activation marker expression. CC-122 significantly increased NFκB, Nur77, and STAT5 as well as effector cytokine production and activation marker expression. Activation-induced exhaustion of CAR T may decrease durable responses in patients. We developed a chronic stimulation assay to render CAR T cells hypofunctional (reduced cytolysis and IL-2 secretion) and examined whether enhanced activation mediated by CC-122 would exhaust the CD19-directed CAR T cells more rapidly. CC-122 was added in one of two regimens: during chronic stimulation (“concurrent”) or during re-challenge (“rescue”). Effects on exhaustion in “concurrent” or “rescue” cultures were evaluated, including by RNAseq and measuring of effector function in re-challenge assays with CD19+ targets. Surprisingly, “concurrent” CC-122 partially reversed a gene signature associated with CAR T cell hypofunctionality and preserved more effector function relative to controls. In “rescue” assays, CC-122 dose-dependently increased cytokine production and restored cytolytic function of exhausted CAR T cells upon re-challenge with CD19+ tumor spheroids. In a disseminated tumor xenograft model, concurrent or delayed dosing of CC-122, in combination with two sub-curative doses of liso-cel, significantly increased median survival by 30 to 40 days. In some cases, survival endpoints were not reached with the combination. Taken together, CC-122 treatment of CD19-directed CAR T cells was shown to enhance CAR T activation in acute stimulation assays; limit CAR T exhaustion onset during chronic stimulation as assessed by gene expression; and reverse an exhausted, hypofunctional CAR T state induced by chronic stimulation. The data suggest that liso-cel pharmacologic activity may both be enhanced and extended by this combinatorial approach. These findings support an ongoing clinical investigation evaluating the addition of CC-122 to liso-cel in patients with R/R, aggressive NHL (NCT03310619). Citation Format: Heidi K. Jessup, Evan P. Thomas, Jim S. Qin, Yue Jiang, Oleksandr Baturevych, Neha Soni, Aye T. Chen, Ruth A. Salmon, Melissa G. Works, Michael O. Ports. Avadomide (CC-122) increases effector function and reverses exhaustion in chronically stimulated lisocabtagene maraleucel anti-CD19 CAR T drug product [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2320.

CSPG4-Specific CAR T Cells for High-Risk Childhood B Cell Precursor Leukemia.

The advent of CD19-specific chimeric antigen receptor (CAR) T cells has proven to be a powerful asset in the arsenal of cancer immunotherapy of acute lymphoblastic leukemia and certain B cell lymphomas. However, a sizable portion of patients treated with CD19-CAR T cells relapse with CD19-negative cancer cells, necessitating the quest for back-up antigens. Chondroitin sulfate proteoglycan 4 (CSPG4) expression has been reported on leukemic blasts bearing the ill-fated MLL 11q23 rearrangement. We aimed at exploring the use of CSPG4-specific CAR T cells against mixed-lineage leukemia (MLL)-rearranged leukemic blasts, using the precursor B cell leukemia cell line KOPN8 (MLL–MLLT1 translocation) as a model. First, we confirmed CSPG4 expression on KOPN8 cells. Bulk T cells electroporated with mRNA encoding a CSPG4-specific CAR upregulated activation markers and secreted the Th1 cytokines TNF and IFNγ in an antigen-specific manner upon co-culture with KOPN8 cells. More importantly, CSPG4-specific CAR T cells evinced specific degranulation towards KOPN8 cells and specifically lysed KOPN8 target cells in chromium lysis experiments. CSPG4 is a well-established CAR target in cutaneous melanoma. Here, we provide proof-of-principle data for the use of CSPG4-specific CAR T cells against MLL-translocated leukemias.

S1634 A PHASE I FIRST‐IN‐HUMAN CLINICAL TRIAL OF CD19‐TARGETED 19–28Z/4–1BBL “ARMORED” CAR T CELLS IN PATIENTS WITH RELAPSED OR REFRACTORY NHL AND CLL INCLUDING RICHTER TRANSFORMATION

Background:Second generation, CD19‐specific chimeric antigen receptor (CAR) T cell therapy containing either the CD28 or 41BB co‐stimulatory domain have demonstrated long‐term complete remission (CR) rates of 40–50% in relapsed or refractory (R/R) NHL and 20–30% in CLL. The suboptimal activity of the 2nd generation CAR T cells in NHL and in particular CLL may be due, in part, to poor proliferation and cytolytic activity of T cells in the inhibitory tumor microenvironment. We have previously demonstrated that binding of 4–1BBL to its cognate receptor enhances T cell proliferation, IL‐2 secretion, survival and cytolytic activity of the T cells compared to 19–28z, 19–41BBz and 1928BBz (Zhao Z et al. Cancer Cell 2015;28:415–428). Herein, we report updated results ona study of adult patients with CLL and NHL treated with escalating doses of autologous 19–28z/4–1BBL+ CAR T cells (NCT03085173).Aims:The primary objective of this study was to determine the toxicity and MTD of 1928z‐41BBL CAR T cells in patients with relapsed or refractory CD19+ hematologic malignancies. Secondary objectives was to assess the anti‐tumor efficacy and in vivo persistence of the CAR T cells. Exploratory objectives were to assess the level of MRD when applicable and to assess the impact of infused CAR T cells on endogenous anti‐tumor immune response and changes in cellular and cytokine tumor microenvironment.Methods:Pts with R/R NHL (DLBCL, follicular lymphoma (FL), transformed FL (tFL), Waldenström's macroglobulinemia (WM)), CLL including Richter's transformation and B‐ALL were eligible for the trial. Pts received conditioning chemotherapy of cyclophosphamide (Cy) alone or combined with fludarabine (Flu) followed by escalating doses of CAR T cells. CAR T cells were administered at dose level (DL) 1 (1x105 CAR T cells/kg), DL2 (3x105 CAR T cells/kg), DL 3 (1x106 CAR T cells/kg), and DL4 (3x106 CAR T cells/kg).Results:28 pts were enrolled with R/R CLL (n = 9), de novo DLBCL (n = 9), tFL (n = 3), FL and WM (n = 4), Richter's transformation (n = 3) and a single pt with B‐ALL. Median age was 70 (range, 53–81), and median number of prior treatments was 5 (range, 2–17). 3 pts received DL1 CAR T cells, 3 pts DL2 CAR T cells, 3 pts DL3 CAR T cells and remaining 19 pts received DL4 CAR T cells. No dose‐limiting toxicity (DLT) was observed. All 28 pts are at least 2 weeks from T cell infusion and evaluable for toxicity. Overall, 11 pts experienced CRS (39.3%), which were mostly grade 1 (n = 6) and grade 2 (n = 4) and 11 pts (39.3%) experienced neurotoxicity (NTX), including 8 pts with grade 1–2 and 3 pts with grade 3 NTX. 16 of 28 pts (57%) achieved a CR, including 7 of 9 pts (78%) with large cell lymphoma, 2 of 3 pts with FL (67%), 3 of 10 pts (30%) with CLL, 2 pts with Richter's transformation (67%) and the one pt with B‐ALL. The pt with WM achieved a VGPR. With a median follow‐up of 169 days (24–534 days), 8 pts (29%) remain in CR. Peak CAR T cell expansion occurred at a median of 9 days after CAR T cell infusion (range, 2–82). CAR T cell detection beyond 160 days has been noted.Summary/Conclusion:Treatment with 19–28z/41BBL armored CAR T cells is safe. No severe CRS was observed and severe NTX occurred in 8% of the pts. The overall CR rate of 57% is encouragingCR rates were higher in pts with large cell lymphoma (78%) compared to CLL (20%), though small number of pts limits any firm conclusions. Pts with CLL may require higher doses of CAR T cells or incorporation of the CAR therapy in earlier lines of treatments. Detailed cytokine and CAR T cell expansion analysis as well as updated data will be presented.image

ZUMA-8: A phase 1/2 multicenter study evaluating KTE-X19 in patients (pts) with relapsed/refractory (R/R) chronic lymphocytic leukemia (CLL).

TPS7566 Background: Despite treatment advances, CLL is largely incurable. First-line targeted therapy with ibrutinib mostly produces durable remissions, but high-risk disease or many prior therapies increases relapse risk (Ghia P, et al. Haematologica. 2014). Relapse after ibrutinib is associated with a poor outcome (Maddocks KJ, et al. JAMA Oncol. 2015). Allogeneic stem cell transplantation potentially offers long-term remissions but has a high risk of morbidity/mortality (Shustik C, et al. Ann Hematol. 2017). Autologous T cells expressing a CD19-specific chimeric antigen receptor (CAR) with a CD28 costimulatory domain may be efficacious against CLL (Kochenderfer JN, et al. Blood. 2012). KTE-X19 is an autologous anti-CD19 CAR T cell therapy under investigation for R/R hematologic malignancies and may offer longer durable remissions with manageable safety in pts with R/R CLL. ZUMA-8 is a Phase 1/2 multicenter study for pts with R/R CLL. Methods: Adult pts must have R/R CLL with ≥ 2 prior treatment regimens, disease progression on ibrutinib, ECOG 0-1, and adequate organ function. Phase 1 will enroll 12-18 pts to assess dose-limiting toxicities (DLTs) with a 6 + 3 dose escalation/de-escalation design; 30 more pts may be enrolled to further assess safety. Phase 2 will enroll ≈60 pts to evaluate efficacy and safety. Pts will undergo leukapheresis followed by optional bridging therapy. Cyclophosphamide (500 mg/m2) and fludarabine (30 mg/m2) conditioning chemotherapy will be given on Days -5, -4 and -3. KTE-X19 will be given on Day 0 at 0.5, 1 or 2 × 106 KTE-X19 cells/kg. The primary endpoint is incidence of DLTs for Phase 1 and independent review committee-assessed objective response rate (ORR) per iwCLL 2018 criteria for Phase 2. Secondary endpoints include complete remission (CR) rate, investigator-assessed ORR, minimal residual disease (MRD) negativity rate, MRD-negative CR rate, duration of response, progression-free survival, overall survival, safety, and patient-reported outcomes (Phase 2). Serum cytokine and blood KTE-X19 cell levels over time and level of anti-KTE-X19 antibodies are exploratory endpoints. Accrual is ongoing. Clinical trial information: NCT03624036.

Imaging of CAR T-Cells in Cancer Patients: Paving the Way to Treatment Monitoring and Outcome Prediction.

In 2013, the journal Science named cancer immunotherapy as the “breakthrough of the year” based on targeted approaches using chimeric antigen receptor (CAR) T-cells. CD19-specific CAR T-cell therapy has revolutionized the treatment landscape for patients with relapsed B-cell acute lymphocytic

Piggybat transposase for the generation of CD19 specific chimeric antigen receptor T cells

Background & Aim Introduction DNA transposon systems offer an alternative to viral vectors for genetic modification of eukaryotic cells. The Piggybac system is being used in CAR T-cell clinical trials presently, but its widespread use may be limited by licensing restrictions. Transposon mining of the little brown bat (Myotis lucifugus) genome has revealed the presence of an active DNA transposase named Piggybat. Aims To optimise manufacturing conditions and evaluate the genomic integration profile of the Piggybat transposase in the generation of CD19 specific CAR T-cells (CAR19-T) compared to Piggybac transposase. Methods, Results & Conclusion Methods CAR19-T were generated from healthy donors (n=3) by co-electroporation of two plasmids encoding the transposase and a CAR19 transposon. T-cells were co-cultured with irradiated feeder cells and cytokines. CAR19-T expansion, CAR expression, memory phenotype, CD19 specific activation and cytotoxicity were assessed after a 2-week culture period. CAR gene integration copy number was determined by droplet digital polymerase chain reaction and integration site analysis was conducted by next generation sequencing of PCR amplicons spanning genomic DNA/CAR gene intersection sites. Results Over 2 weeks, Piggybat CAR19-T expanded a mean of 26 fold with 52% CAR expression, and Piggybac CAR19-T expanded a mean of 11 fold with 67% CAR expression. Piggybat and Piggybac CAR19-T had a mean 59% and 48% CD8+ T-cells respectively, with a similar CAR19-T memory phenotype. Using 50ug/ml concentration of each transposon and transposase plasmids, Piggybat had fewer genomic integrants than Piggybac (mean 3.4 and 10.1 copies per cell respectively). Integrant copy number could be titrated by altering transposon plasmid concentration. Peak CAR19-T expansion, expression and MFI were achieved using 1ug Piggybat transposase plasmid and 5ug transposon plasmid. Piggybat and Piggybac CAR19-T demonstrated similar CD19 specificity and cytotoxicity. Piggybat CAR19-T showed 1920 unique genomic integration sites, 11.13% within 2.5kb of transcription start sites (TSS) and 43.72% being intragenic, compared to Piggybac with 2697 unique integration sites, 20.33% TSS localised and 47.77% intragenic. Conclusions Compared to Piggybac, Piggybat transposase can generate CD19 specific CAR T-cells with similar CAR expression, function and insertion site profile, but fewer integrants per cell. Thus, Piggybat is a new DNA transposon that may allow cost-effective CAR T-cell production for clinical use.

Factors associated with durable EFS in adult B-cell ALL patients achieving MRD-negative CR after CD19 CAR T-cell therapy

AbstractAutologous T cells engineered to express a CD19-specific chimeric antigen receptor (CAR) have produced impressive minimal residual disease–negative (MRD-negative) complete remission (CR) rates in patients with relapsed/refractory B-cell acute lymphoblastic leukemia (B-ALL). However, the factors associated with durable remissions after CAR T-cell therapy have not been fully elucidated. We studied patients with relapsed/refractory B-ALL enrolled in a phase 1/2 clinical trial evaluating lymphodepletion chemotherapy followed by CD19 CAR T-cell therapy at our institution. Forty-five (85%) of 53 patients who received CD19 CAR T-cell therapy and were evaluable for response achieved MRD-negative CR by high-resolution flow cytometry. With a median follow-up of 30.9 months, event-free survival (EFS) and overall survival (OS) were significantly better in the patients who achieved MRD-negative CR compared with those who did not (median EFS, 7.6 vs 0.8 months; P < .0001; median OS, 20.0 vs 5.0 months; P = .014). In patients who achieved MRD-negative CR by flow cytometry, absence of the index malignant clone by IGH deep sequencing was associated with better EFS (P = .034). Stepwise multivariable modeling in patients achieving MRD-negative CR showed that lower prelymphodepletion lactate dehydrogenase concentration (hazard ratio [HR], 1.38 per 100 U/L increment increase), higher prelymphodepletion platelet count (HR, 0.74 per 50 000/μL increment increase), incorporation of fludarabine into the lymphodepletion regimen (HR, 0.25), and allogeneic hematopoietic cell transplantation (HCT) after CAR T-cell therapy (HR, 0.39) were associated with better EFS. These data allow identification of patients at higher risk of relapse after CAR T-cell immunotherapy who might benefit from consolidation strategies such as allogeneic HCT. This trial was registered at www.clinicaltrials.gov as #NCT01865617.

19. Unexpected karyotype findings and lineage switching during anti-CD19 CAR-T immunotherapy for lymphoid malignancies

Background: CD19-specific chimeric antigen receptor (CAR)-modified T cells infusion is a promising therapy for patients with relapsed/refractory CD19+ B cell malignancies. Early in our institution's CAR-T program, 2 anecdotal cases of B-ALL patients developed AML post CAR-T with karyotypic aberrations noted prior to infusion.

Evolution of chimeric antigen receptor (CAR) T cell therapy: current status and future perspectives.

Engineering T cells with a chimeric antigen receptor (CAR) that reprograms their antigen selectivity and signaling has recently emerged as one of the most promising therapeutic approaches for treating cancers. For example, two CD19-specific CAR T cell (CAR-T) therapies have shown remarkable responses in patients with relapsed/refractory B-cell cancers, and were approved by the US Food and Drug Administration in 2017. This initial clinical success has spurred an explosion of interests in this novel therapy from both academia and industry, and results from basic and clinical research have enabled the rapid evolution of the CAR-T field. In this review, we describe the basic structure of the CAR and discuss how each of its domains affect the efficacy and safety of CAR-T therapies. In addition, we discuss some of the novel concepts and other considerations that are essential for ensuring the future success of CAR-T therapy.

Orphan designation: Autologous CD4+ and CD8+ T cells expressing a CD19-specific chimeric antigen receptor, Treatment of diffuse large B-cell lymphoma

Orphan designation: Autologous CD4+ and CD8+ T cells expressing a CD19-specific chimeric antigen receptor, Treatment of diffuse large B-cell lymphoma

Orphan designation: Autologous CD4+ and CD8+ T cells expressing a CD19-specific chimeric antigen receptor, Treatment of diffuse large B-cell lymphoma

Orphan designation: Autologous CD4+ and CD8+ T cells expressing a CD19-specific chimeric antigen receptor, Treatment of diffuse large B-cell lymphoma

Orphan designation: Autologous CD4+ and CD8+ T cells expressing a CD19-specific chimeric antigen receptor, Treatment of follicular lymphoma

Orphan designation: Autologous CD4+ and CD8+ T cells expressing a CD19-specific chimeric antigen receptor, Treatment of follicular lymphoma

Orphan designation: Autologous CD4+ and CD8+ T cells expressing a CD19-specific chimeric antigen receptor, Treatment of follicular lymphoma

Orphan designation: Autologous CD4+ and CD8+ T cells expressing a CD19-specific chimeric antigen receptor, Treatment of follicular lymphoma

Genetic Modification Strategies to Enhance CAR T Cell Persistence for Patients With Solid Tumors.

Immunotherapy with chimeric antigen receptor (CAR) T cells offers a promising method to improve cure rates and decrease morbidities for patients with cancer. In this regard, CD19-specific CAR T cell therapies have achieved dramatic objective responses for a high percent of patients with CD19-positive leukemia or lymphoma. Most patients with solid tumors however, have experienced transient or no benefit from CAR T cell therapies. Novel strategies are therefore needed to improve CAR T cell function for patients with solid tumors. One obstacle for the field is limited CAR T cell persistence after infusion into patients. In this review we highlight genetic engineering strategies to improve CAR T cell persistence for enhancing antitumor activity for patients with solid tumors.

Abstract A025: Eradication of neuroblastoma by T-cells redirected with an optimized GD2-specific chimeric antigen receptor and IL-15

Abstract The treatment with CD19-specific chimeric antigen receptor T-cells (CAR-Ts) has shown remarkable antitumor activity in patients with B-cell malignancies. However, the clinical benefits of CAR-Ts in solid tumors remain unclear. A compelling concern is that in patients with solid tumors CAR-Ts do not immediately encounter their cognate antigen in the circulation and thus lack the appropriate costimulatory signals necessary for full activation. We here sought to explore if expressing IL-15 in CAR-Ts would provide them with sufficient sustained survival until they engage the cognate antigen. Using the GD2-specific CAR and neuroblastoma (NB) as a tumor model, we explored the benefits of incorporating the IL-15 cytokine (and the iCaspase9 suicide gene for safety) within the CAR molecule. CAR-Ts from 12 healthy donors were transduced with an optimized GD2.CAR (encoding the CD28 endodomain) without (GD2-Ts) or with the IL15 (GD2.15.iC9-Ts) and expanded ex vivo with IL-7/IL-15 for 14 days. CAR transduction (82% ± 8% and 82% ± 12%, respectively) and ex vivo antitumor activity in 4 days co-culture at different E:T ratios were comparable. However, upon repetitive stimulation with GD2+ NB tumors (CHLA-255 and LAN-1), only GD2.15.iC9-Ts showed significantly superior expansion and antitumor activity (p&amp;lt;0.05). In vivo in a NB metastatic model in NSG mice engrafted with FireflyLuciferase+ CHLA-255 or LAN-1, IL-15 producing CAR-Ts showed superior antitumor activity at low T-cell doses. Tumor rechallenge experiments clearly highlighted the superior survival of mice receiving GD2.15.iC9-Ts, which was paralleled by higher detection of IL-15 in the circulation and frequency of CAR-Ts in vivo. Remarkably, although CAR-Ts were expanded in the presence of the same amount of exogenous IL-7/IL-15, we observed a significantly higher frequency of “stem-like cells” in GD2.15.iC9-Ts (25% ± 11%, CD45RA+CCR7+) compared to GD2-Ts (6% ± 2%, CD45RA+CCR7+, p&amp;lt;0.001), suggesting that transgenic IL-15 expression favors the preservation of T-cell “stem-ness.” Importantly, CAR-Ts were controllable as the dimerizing drug AP20187 induced cell death of GD2.15.iC9-Ts both in vitro and in vivo. Overall, our data suggest that transgenic IL-15 within the CAR construct can increase the persistence of CAR-Ts targeting NB tumors and preserve more stem-ness. Our results will not only guide therapeutic options for patient with NB, but also CAR-T developmental research for a broad range of solid tumors. Citation Format: Yuhui Chen, Chuang Sun, Leonid Metelitsa, Gianpietro Dotti, Barbara Savoldo. Eradication of neuroblastoma by T-cells redirected with an optimized GD2-specific chimeric antigen receptor and IL-15 [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A025.