In Vivo Fate and Activity of Second- versus Third-Generation CD19-Specific CAR-T Cells in B Cell Non-Hodgkin's Lymphomas.

Second-generation (2G) chimeric antigen receptors (CARs) targeting CD19 are highly active against B cell malignancies, but it is unknown whether any of the costimulatory domains incorporated in the CAR have superior activity to others. Because CD28 and 4-1BB signaling activate different pathways, combining them in a single third-generation (3G) CAR may overcome the limitations of each individual costimulatory domain. We designed a clinical trial in which two autologous CD19-specific CAR-transduced T cell products (CD19.CARTs), 2G (with CD28 only) and 3G (CD28 and 4-1BB), were infused simultaneously in 16 patients with relapsed or refractory non-Hodgkin's lymphoma. 3G CD19.CARTs had superior expansion and longer persistence than 2G CD19.CARTs. This difference was most striking in the five patients with low disease burden and few circulating normal B cells, in whom 2G CD19.CARTs had limited expansion and persistence and correspondingly reduced area under the curve. Of the 11 patients with measurable disease, three achieved complete responses and three had partial responses. Cytokine release syndrome occurred in six patients but was mild, and no patient required anti-IL-6 therapy. Hence, 3G CD19.CARTs combining 4-1BB with CD28 produce superior CART expansion and may be of particular value when treating low disease burden in patients whose normal B cells are depleted by prior therapy.

Ramos CA ，Rouce R ，Robertson CS ，Reyna A ，Narala N ，Vyas G ，Mehta B ，Zhang H ，Dakhova O ，Carrum G ，Kamble RT ，Gee AP ，Mei Z ，Wu MF ，Liu H ，Grilley B ，Rooney CM ，Heslop HE ，Brenner MK ，Savoldo B ，Dotti G ... -  《-》

Treatment of patients with relapsed or refractory CD19+ lymphoid disease with T lymphocytes transduced by RV-SFG.CD19.CD28.4-1BBzeta retroviral vector: a unicentre phase I/II clinical trial protocol.

Chimeric antigen receptor (CAR) T cells spark hope for patients with CD19+ B cell neoplasia, including relapsed or refractory (r/r) acute lymphoblastic leukaemia (ALL) or r/r non-Hodgkin's lymphoma (NHL). Published studies have mostly used second-generation CARs with 4-1BB or CD28 as costimulatory domains. Preclinical results of third-generation CARs incorporating both elements have shown superiority concerning longevity and proliferation. The University Hospital of Heidelberg is the first institution to run an investigator-initiated trial (IIT) CAR T cell trial (Heidelberg Chimeric Antigen Receptor T cell Trial number 1 [HD-CAR-1]) in Germany with third-generation CD19-directed CAR T cells. Adult patients with r/r ALL (stratum I), r/r NHL including chronic lymphocytic leukaemia, diffuse large B-cell lymphoma, follicular lymphoma or mantle cell lymphoma (stratum II) as well as paediatric patients with r/r ALL (stratum III) will be treated with autologous T-lymphocytes transduced by third-generation RV-SFG.CD19.CD28.4-1BB zeta retroviral vector (CD19.CAR T cells). The main purpose of this study is to evaluate safety and feasibility of escalating CD19.CAR T cell doses (1-20×106 transduced cells/m2) after lymphodepletion with fludarabine (flu) and cyclophosphamide (cyc). Patients will be monitored for cytokine release syndrome (CRS), neurotoxicity, i.e. CAR-T-cell-related encephalopathy syndrome (CRES) and/or other toxicities (primary objectives). Secondary objectives include evaluation of in vivo function and survival of CD19.CAR T cells and assessment of CD19.CAR T cell antitumour efficacy.HD-CAR-1 as a prospective, monocentric trial aims to make CAR T cell therapy accessible to patients in Europe. Currently, HD-CAR-1 is the first and only CAR T cell IIT in Germany. A third-generation Good Manufacturing Practice (GMP) grade retroviral vector, a broad spectrum of NHL, treatment of paediatric and adult ALL patients and inclusion of patients even after allogeneic stem cell transplantation (alloSCT) make this trial unique. Ethical approval and approvals from the local and federal competent authorities were granted. Trial results will be reported via peer-reviewed journals and presented at conferences and scientific meetings. Eudra CT 2016-004808-60; NCT03676504; Pre-results.

Schubert ML ，Schmitt A ，Sellner L ，Neuber B ，Kunz J ，Wuchter P ，Kunz A ，Gern U ，Michels B ，Hofmann S ，Hückelhoven-Krauss A ，Kulozik A ，Ho AD ，Müller-Tidow C ，Dreger P ，Schmitt M ... -  《BMJ Open》

Phase 1 studies of central memory-derived CD19 CAR T-cell therapy following autologous HSCT in patients with B-cell NHL.

Myeloablative autologous hematopoietic stem cell transplantation (HSCT) is a mainstay of therapy for relapsed intermediate-grade B-cell non-Hodgkin lymphoma (NHL); however, relapse rates are high. In phase 1 studies designed to improve long-term remission rates, we administered adoptive T-cell immunotherapy after HSCT, using ex vivo-expanded autologous central memory-enriched T cells (TCM) transduced with lentivirus expressing CD19-specific chimeric antigen receptors (CARs). We present results from 2 safety/feasibility studies, NHL1 and NHL2, investigating different T-cell populations and CAR constructs. Engineered TCM-derived CD19 CAR T cells were infused 2 days after HSCT at doses of 25 to 200 × 10(6) in a single infusion. In NHL1, 8 patients safely received T-cell products engineered from enriched CD8(+) TCM subsets, expressing a first-generation CD19 CAR containing only the CD3ζ endodomain (CD19R:ζ). Four of 8 patients (50%; 95% confidence interval [CI]: 16-84%) were progression free at both 1 and 2 years. In NHL2, 8 patients safely received T-cell products engineered from enriched CD4(+) and CD8(+) TCM subsets and expressing a second-generation CD19 CAR containing the CD28 and CD3ζ endodomains (CD19R:28ζ). Six of 8 patients (75%; 95% CI: 35-97%) were progression free at 1 year. The CD4(+)/CD8(+) TCM-derived CD19 CAR T cells (NHL2) exhibited improvement in expansion; however, persistence was ≤28 days, similar to that seen by others using CD28 CARs. Neither cytokine release syndrome nor delayed hematopoietic engraftment was observed in either trial. These data demonstrate the safety and feasibility of CD19 CAR TCM therapy after HSCT. Trials were registered at www.clinicaltrials.gov as #NCT01318317 and #NCT01815749.

Wang X ，Popplewell LL ，Wagner JR ，Naranjo A ，Blanchard MS ，Mott MR ，Norris AP ，Wong CW ，Urak RZ ，Chang WC ，Khaled SK ，Siddiqi T ，Budde LE ，Xu J ，Chang B ，Gidwaney N ，Thomas SH ，Cooper LJ ，Riddell SR ，Brown CE ，Jensen MC ，Forman SJ ... -  《-》

In Vivo Expansion and Antitumor Activity of Coinfused CD28- and 4-1BB-Engineered CAR-T Cells in Patients with B Cell Leukemia.

Several recent clinical trials have successfully incorporated a costimulatory domain derived from either CD28 or 4-1BB with the original CD3ζ T cell activating domain to form second-generation chimeric antigen receptors (CARs) that can increase the responsiveness and survival of CAR-engineered T (CAR-T) cells. However, a rigorous assessment of the individual benefits of these costimulatory components relative to the in vivo performance of infused T cells in patients is still lacking. Therefore, we have designed a study that allows us to investigate and compare the impact of different costimulatory signal domains on CAR-T cells in vivo. Patients with B cell leukemia were infused with a mixture of two types of CD19-specific CAR-T cells, individually bearing CD28 (28ζ) and 4-1BB (BBζ) costimulatory signaling domains. We found that such a clinical procedure was feasible and safe. Complete remission (CR) was observed in five of seven enrolled patients, with two patients exhibiting durable CR lasting more than 15 months. The in vivo expansion pattern of 28ζ and BBζ CAR-T cells varied significantly among individual patients. These results confirm a feasible method of comparing different CAR designs within individual patients, potentially offering objective insights that may facilitate the development of optimal CAR-T cell-based immunotherapies.

Cheng Z ，Wei R ，Ma Q ，Shi L ，He F ，Shi Z ，Jin T ，Xie R ，Wei B ，Chen J ，Fang H ，Han X ，Rohrs JA ，Bryson P ，Liu Y ，Li QJ ，Zhu B ，Wang P ... -  《-》

Phase I Trial of Fourth-Generation Anti-CD19 Chimeric Antigen Receptor T Cells Against Relapsed or Refractory B Cell Non-Hodgkin Lymphomas.

The administration of second- or third-generation anti-CD19 chimeric antigen receptor (CAR) T cells has remarkably improved the survival of patients with relapsed or refractory B cell malignancies. However, there are limited clinical results from fourth-generation CAR-T cell therapy, and the factors affecting response rate and survival have not been fully determined. Lymphoma patients with progression or relapse after intensive treatments, including hematopoietic stem cell transplantation, and life expectancy >2 months were enrolled in the study. Peripheral lymphocytes were collected through apheresis, and magnetically selected T cells were lentivirally transduced with a 4th-generation CAR featuring an anti-CD19 CAR and the iCasp9 suicide switch (4SCAR19). The patients received 4SCAR19 T cell infusion after approximately seven days of expansion and a conditioning regimen comprising cyclophosphamide/fludarabine. The efficacy, safety, and risk factors were evaluated. A total of 21 patients with relapsed/refractory B cell non-Hodgkin lymphoma were enrolled and received 4SCAR19 T cell infusions at a median dose of 8.9×105 CAR-T cells/kg. The overall response rate was 67% [95% confidence interval (CI), 43 to 85], with 43% of patients achieving a complete response and 24% having a partial response. The overall and complete response rates were 58 and 33% in the diffuse large B-cell lymphoma (DLBCL) group and 78 and 56% in the non-DLBCL group, respectively. The median overall survival was 23.8 months (95% CI, not reached), with a median follow-up of 13.7 months. Factors affecting overall survival were International Prognostic Index (IPI), disease type, and remission status after CAR-T cell treatment. The most common adverse events of grade 3 or 4 during treatment were neutropenia (76%), leukopenia (71%), and thrombocytopenia (29%). The incidence of cytokine release syndrome (CRS) was 14%, and all cases were grade 1. One patient developed grade 3 neurotoxicity. No deaths were attributed to infusion of 4SCAR19 T cells, CRS, or neurotoxicity. In this study, patients with relapsed or refractory B cell non-Hodgkin's lymphoma who received 4SCAR19 T cell therapy had durable responses and few of adverse events. The IPI model is suitable for evaluating the prognosis of patients receiving CAR-T cell therapy. Chinese Clinical Trial Registry (http://www.chictr.org.cn): ChiCTR-OOC-16007779.

Zhou X ，Tu S ，Wang C ，Huang R ，Deng L ，Song C ，Yue C ，He Y ，Yang J ，Liang Z ，Wu A ，Li M ，Zhou W ，Du J ，Guo Z ，Li Y ，Jiao C ，Liu Y ，Chang LJ ，Li Y ... -  《Frontiers in Immunology》