Molecular Remission and B Cell Aplasia Induced in a First Cohort of Adults with Relapsed B-ALL Treated with 19–28z CAR-Targeted T Cells

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3566-3566
Author(s):  
Marco L Davila ◽  
Isabelle Riviere ◽  
Xiuyan Wang ◽  
Jae H. Park ◽  
Jolanta Stefanski ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Induction Chemotherapy ◽  
High Dose ◽  
Molecular Remission ◽  
Hematopoietic Stem ◽  
Allogeneic Hsct ◽  
Initial Cohort

Abstract Abstract 3566 B cell acute lymphoblastic leukemia (B-ALL) in adults has a dismal prognosis. Intensified, combinatorial chemotherapy yields remarkable results in pediatric ALL but less so in adults. As allogeneic hematopoietic stem cell transplantation (HSCT) is curative in only a minority of patients, there remains a dire need for effective therapies in this patient population. Building on our results obtained with genetically targeted T cells in xenogeneic models of ALL and our experience in patients with chronic lymphocytic leukemia (Brentjens, Rivière, Blood, 2011), we have recently initiated a clinical trial (NCT01044069) treating adults with relapsed or refractory B-ALL with autologous T cells expressing the 19–28z chimeric antigen receptor (CAR), a dual CD3zeta/CD28 signaling receptor specific for the B cell antigen CD19. Human peripheral blood T cells retrovirally transduced to express the 19–28z receptor specifically lyse normal and malignant B cells in vitro and eradicate established tumors in vivo in pre-clinical animal models. We report the results from the initial cohort of patients treated on this protocol. The first two patients had relapsed disease that achieved morphologic remission following re-induction chemotherapy. Following adoptive therapy with CAR-modified T cells, one patient achieved a B cell aplasia and molecular remission documented by deep sequencing, which demonstrated the disappearance of IgH rearrangements associated with the malignant clone. The other patient had achieved a molecular remission after re-induction chemotherapy but further developed a complete B cell aplasia following modified T cell infusion. While in molecular remission, both patients successfully underwent allogeneic HSCT and were therefore removed from the study. In contrast to these two patients, the third patient failed to achieve a morphologic remission after re-induction chemotherapy and had >60% blasts in the bone marrow (BM) at the time of T cell infusion. Within 12 hours of completing T cell infusion, the patient developed high-grade fevers, hypotension, and rigors. Serum analyses demonstrated a sharp rise in cytokines (IFNg, TNFa, IL6, IL2, and IL8), reflecting rapid and robust onset of T cell activation. High-dose corticosteroids initiated 5 days after T cell infusion controlled these symptoms. Post-T cell BM aspirate on day 8 demonstrated undetectable blasts. Flow cytometry could not detect blasts or B lineage cells, but readily identified 19–28z CAR+ T cells. Significantly, the patient's B-ALL tumor cells harbored a unique 9p21 cytogenetic deletion that was detected by FISH in 28% of nuclei immediately prior to T cell infusion and was reduced to 5% 8 days after T cell infusion. The next BM aspirate, performed on day 24 after T cell infusion, demonstrated a persistent morphologic remission, a recovering BM, and complete absence of any detectable 9p21 deletions. Furthermore, PCR amplification for IgH rearrangements in the BM 24 days after T cell infusion confirmed a molecular remission and a B cell aplasia. Collectively, the clinical outcomes from this initial cohort of 3 patients demonstrate for the first time the in vivo efficacy of CD19-targeted T cells to induce clinical and molecular remissions as well as B cell aplasia in adults with relapsed or refractory B-ALL. These early results strongly support further investigation of 19–28z targeted T cells to treat leukemias and suggest this is a potential salvage therapy for patients with relapsed/refractory B-ALL who are failing chemotherapy or are ineligible for allogeneic HSCT. Disclosures: No relevant conflicts of interest to declare.

Gene Targeting of RhoA Reveals Its Essential Role In Lymphopoiesis

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 282-282
Author(s):  
Shuangmin Zhang ◽  
Yi Zheng ◽  
Richard Lang ◽  
Fukun Guo
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Cell Development ◽  
B Cell Development ◽  
Brdu Incorporation ◽  
Hematopoietic Stem ◽  
Cell Functions

Abstract Abstract 282 RhoA GTPase is an intracellular signal transducer capable of regulating a wide range of cell functions including cytoskeleton dynamics, proliferation, and survival. In lymphocytes, studies by using dominant negative mutant or C3 transferase expressing transgenic mice suggest that RhoA is involved in TCR and BCR signaling and related T cell functions such as polarization, migration, survival, and proliferation. To date, the physiological role of RhoA in lymphocyte development remains unclear. In this study, we have achieved T cell, B cell, and hematopoietic stem cell-specific deletion of RhoA by conditional gene targeting with CD2, CD19 and Mx1 promoter-driven Cre expression, respectively, in the RhoAloxP/loxP mice. First, we found that RhoA gene disruption in early T cells caused a drastic decrease in thymocyte cellularity, with the numbers of CD4−CD8− double negative (DN), CD4+CD8+ double positive (DP), CD4+CD8− single positive (SP), and CD4−CD8+ SP T cells decreased by 88.8% ± 6.0%, 99.4% ± 1.0%, 99.3% ± 1.2%, and 98.6% ± 2.0%, respectively. Among DN subpopulations, CD44+CD25− (DN1), CD44+CD25+ (DN2), CD44−CD25+ (DN3), and CD44−CD25− (DN4) cells were reduced by 91.7% ± 6.0%, 54.9% ± 27.7%, 50.9% ± 33.3%, and 96.7% ± 3.4%, respectively. Further, RhoA knockout led to a significant loss of DP thymocytes at the initial stage (CD69highTCRint) of positive selection, suggesting that RhoA is required for positive selection. The decreased thymocyte cellularity in mutant mice is associated with increased apoptosis of all thymic T lineages. RhoA deficiency also resulted in a perturbation in thymocyte cell cycle progression as manifested by increased BrdU incorporation in DN1 and DN2 cells and decreased BrdU incorporation in DN4 and DP cells. Concomitantly, RhoA-deficient thymocytes showed a 59.8% ± 26.3% reduction in proliferative potential in response to TCR crosslinking. Western blot analysis revealed that the activities of ZAP70, LAT, Akt, Erk, and p38 were impaired in RhoA-/- thymocytes. In periphery, spleens of the RhoA null mice contained 7.4% ± 8.0% of CD4+ T cells and 3.7% ± 2.7% of CD8+ T cells compared with that of wild type (WT) mice. Loss of peripheral mature T cells in mutant mice is reflected by a marked reduction of naive T cells, whereas effector and memory phenotype cells were marginally affected by RhoA deficiency. RhoA-deficient naïve T cells were more susceptible to apoptosis, suggesting that homeostatic defect of naïve T cells in RhoA-/- mice is attributed to impaired cell survival. Abrogation of RhoA caused an increased in vivo BrdU incorporation in naïve T cell compartments. Thus, RhoA deficiency induces naïve T cell homeostatic proliferation, possibly due to a compensatory effect of lymphopenia. In contrast to that in thymocytes, Erk was constitutively activated in RhoA-deficient splenic T cells. These observations implicate RhoA in the multiple stages of T cell development and the proper assembly of early TCR signaling complex. Second, deletion of RhoA in pre-proB cells had no effect on early B cell development in bone marrow but significantly inhibited late B cell development in spleen, resulting in 78.2% ± 13.6%, 78.6% ± 16.9%, and 93.2% ± 3.4% reduction in transitional, follicular, and marginal zone B cells, respectively. Plasma cells in spleen were decreased by 50.9 % ± 25.9% in RhoA null mice. However, we did not detect any changes in survival of in vivo RhoA-/- B cells or RhoA-/- B cells cultured in vitro with survival factor BAFF. Distinct from previously characterized Cdc42 knockout mice, BAFF-R expression was not altered in RhoA-/- B cells. Moreover, RhoA-/- B cells appeared to be normal in proliferation and Akt and Erk activation in response to BCR crosslinking. These data suggest that RhoA is important for late B cell development through regulation of differentiation but not cell survival or proliferation. Finally, deletion of RhoA from hematopoietic stem cells did not affect common lymphoid progenitor production, indicating that RhoA is not required for early lymphoid progenitor commitment. Taken together, these lineage-specific mouse genetic studies demonstrate that RhoA critically regulates T and B cell development by distinct cellular mechanisms at multiple stages of lymphopoiesis. Disclosures: No relevant conflicts of interest to declare.

Hematopoietic Multipotent Progenitor Cells Mobilized by G-CSF Can Inhibit Gvhd

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2969-2969
Author(s):  
Maud D'Aveni ◽  
Julien Rossignol ◽  
Ruddy Montandon ◽  
Marie Bouillie ◽  
Flora Zavala ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Progenitor Cells ◽  
Mechanisms Of Action ◽  
Activated T Cells ◽  
Hematopoietic Stem ◽  
Allogeneic Hsct ◽  
Ifn Γ

Abstract Abstract 2969 Backgound. Acute graft-versus-host-disease (aGVHD) is a frequent life threatening complication of allogeneic hematopoietic stem cell transplantation (HSCT). Despite the infusion of higher doses of T cells with the use of G-CSF-mobilized HSC grafts, the incidence of aGVHD is not increased. The mechanisms by which G-CSF-mobilized HSC can control GVHD are imperfectly elucidated. We previously described the mobilization of murin hematopoïetic progenitor cells (HPCs) by G-CSF and FLT3 ligand capable of inducing tolerance against autoimmune diabetes in the nude mice (Kared, Immunity 2006). We now show that G-CSF can mobilize murin HPCs with immunoregulatory functions in the allogeneic immune response and describe their mechanisms of action. Methods. Mobilization of HPCs is performed by subcutaneous administration of human recombinant G-CSF at 200μg/kg per day, for 4 consecutive days in the C57BL6 (H-2b) mouse. HPCs are collected in the spleen by FACS sorting according to their phenotype: Lin- Sca1high cKithigh FLT3low CD34+ CD106+ CD127−. In vitro, functions and mechanisms of action were analyzed by co-cultures with i) T cells (from C57BL6) activated by anti-CD28 and -CD3 mAbs or activated by BALB/c (H-2d) allogeneic splenic LPS matured dendritic cells, ii) C57BL6 splenic selected CD4+CD25high T regulatory T cells activated by anti-CD28 and -CD3 mAbs iii) activated antitumor specific CD8 T cells (C57BL6 ovalbumin specific TCR transgenic T cells). These different cultures were performed in the presence or absence of inhibitors of selective cytokines or other regulatory molecules. In vivo, we assessed the effect of donor HPCs on GVHD development by injecting C57BL6 derived HPCs (0.5×106/mouse), splenic T cells (1×106/mouse) and T depleted bone marrow cells (5×106/mouse) into lethally irradiated (8 Gy) Balb/c recipients. Results. In vitro, as compared to controls without HPCs, after 3 days of culture, HPCs: 1) promote the proliferation of natural T regs activated by anti-CD3 and anti-CD28 (>80% at 3 days of culture compared to control <50%), 2) inhibit the proliferation of activated T cells (>80% T cells blocked before 4 divisions as compared to control-T cells alone >80% after 4 divisions- p<0, 001) and 3) induce the apoptosis of activated T cells (30% increased, p=0, 01). The proliferation of T regs was cell contact dependant and required the presence of TGF-b. The inhibition of T cell activation required IFN γ produced by activated T-cells and some contact-dependent stimuli. In such pro-inflammatory conditions, HPCs differentiate after 4 days in myeloid derived suppressor cells (MDSC). These cells could then produce NO in response to IFN γ and suppress the proliferation of activated T cell. However, T cell suppression was not dependant on L-arginine depletion. Induction of apoptosis of T cells was Fas/Fas-L dependant. Although in the presence of HPCs the proliferation of CD8+ T TCR transgenic against the dominant ovalbumin epitope SIINFEKL was reduced, the cytotoxic response against the SIINFEKL-pulsed EL4 cell line was enhanced (cytotoxicity >90% with HPCs versus <90% w/o HPCs, p<0, 001). In addition, HPCs express CCR7 and CD62L, which should allow their migration to the sites of allopriming. In vivo, none of the mice that had received allogeneic HSCT with HPCs developed clinical or histological GVHD signs as compared to 50% of the control allografted mice without HPCs. Conclusion. Hematopoietic progenitor cells acquire an immunosuppressive potential after G-CSF mobilization. These cells can be isolated from mobilized peripheral stem cells and suppress GVHD while possibly preserving the GVL effect. Work is underway in humans to identify and amplify this population ex vivo for potential therapeutic application in allogeneic HSCT. Disclosures: No relevant conflicts of interest to declare.

B Cell Aplasia In a Patient with Relapsed B Cell Acute Lymphoblastic Leukemia Following Re-Induction and Consolidation with Autologous T Cells Genetically Targeted to the CD19 Antigen

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3268-3268
Author(s):  
Marco L Davila ◽  
Clare Taylor ◽  
Xiuyan Wang ◽  
Jolanta Stefanski ◽  
Malgorzata Olszewska ◽  
...  
Keyword(s):  
Clinical Trial ◽  
T Cells ◽  
Stem Cell ◽  
Acute Lymphoblastic Leukemia ◽  
T Cell ◽  
B Cell ◽  
Induction Chemotherapy ◽  
Lymphoblastic Leukemia ◽  
High Dose ◽  
Cell Acute Lymphoblastic Leukemia

Abstract Abstract 3268 Despite high initial remission rates following induction chemotherapy, most adults with B cell acute lymphoblastic leukemia (B-ALL) ultimately relapse and the overall prognosis is poor. In light of the overall poor outcomes seen with currently available chemotherapy regimens as well as allogeneic stem cell transplantation, novel and effective treatment approaches are needed for these patients. To this end, we have developed a program utilizing a patient's own T cells genetically modified ex vivo to express a chimeric antigen receptor (CAR), termed 19–28z, specific to the CD19 antigen expressed on normal B cells as well as most B-ALL tumors. In preclinical studies, human T cells modified to express the 19–28z CAR effectively eradicated systemic human B-ALL NALM-6 tumors in SCID-Beige mice. Based on these findings we have recently opened a phase I clinical trial (IRB #09-114) wherein patients with relapsed B-ALL are initially treated with re-induction chemotherapy followed by consolidation with high dose cyclophosphamide (3gm/m2) and a subsequent infusion of autologous T cells genetically modified to express the 19–28z CAR. Herein, we report the initial findings of the first patient treated on this clinical trial. This patient, a 67-year-old male, with B-ALL (normal cytogenetics), achieved a complete remission following induction chemotherapy with mitoxantrone and high-dose cytarabine. The patient remained in remission following treatment with vincristine (consolidation B) and cyclophosphamide (consolidation C). However, he was noted to have relapsed disease following consolidation cycle D with cytarabine and etoposide. At the time of relapse the patient was leukapheresed to obtain autologous T cells, and subsequently achieved a second remission following re-induction with a modified PEG-asparaginase, vincristine, and prednisone regimen. Upon recovery, the patient, as stipulated by the clinical trial, received lymphodepleting consolidation with high dose cyclophosphamide followed, 2 and 3 days later, by a split dose infusion of 3 × 106/kg autologous 19–28z+ T cells, the lowest planned T cell dose on this trial. Over the next 2 weeks, FACS and Q-PCR detected gene-modified T cells in the peripheral blood. Significantly, over the next 5 weeks, despite recovery of neutrophils and T cells, the patient exhibited a persistent B cell aplasia consistent with CD19-targeted cytotoxic activity of the infused autologous 19–28z+ T cells. The patient subsequently received an allogeneic stem cell transplant from a HLA-identical sibling effectively abrogating further analysis of modified T cell function. Despite this limitation, we conclude that following lymphodepleting chemotherapy, modified CD19-targeted T cells exhibit effective anti-CD19 cytotoxic activity, as demonstrated by the persistent B cell aplasia, in the clinical setting. These findings support the promise of this novel adoptive T cell therapy in patients with relapsed B-ALL. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Overcoming CD19 Antigen Loss in B-Cell Malignancies with CAR T Cells Targeting BAFF-R

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3871-3871
Author(s):  
Hong Qin ◽  
Zhenyuan Dong ◽  
Xiuli Wang ◽  
Wesley Cheng ◽  
Diane Lynne Smith ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
High Dose ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T ◽  
Equity Ownership ◽  
Antigen Loss

Background: The current success in treating hematological malignancies with CD19 targeted chimeric antigen receptor (CAR) T cells is diminished by tumor relapse from target antigen loss. The B-cell activating factor-receptor (BAFF-R), a tumor necrosis factor receptor superfamily protein (TNFRSF13C), is a particularly interesting alternative target in CD19 relapsed disease and in B cell malignancies in general. BAFF-R null mice exhibit greatly reduced normal B-cell numbers and mouse strains expressing a mutant BAFF-R exhibited decreased B-cell life spans and a dramatically reduced peripheral B-cell compartment. Thus, BAFF-R signaling is a driver of B-cell survival, which may limit the capacity of clonal B-cell tumors to escape therapy by down-regulation of antigen expression. We have exploited the potential of the BAFF/BAFF-R axis and developed a proprietary BAFF-R targeting CAR T cell using a single-chain variable fragment (scFv) of a humanized anti-human BAFF-R antibody engineered into a second generation CAR construct containing 4-1BB costimulatory and CD3ζ intracellular signaling domains (Qin et al. Sci. Transl. Med., In Press). BAFF-R CAR T cells were cytotoxic against a panel of human leukemia and lymphoma lines. Adoptively transferred BAFF-R-CAR T cells eradicated 10-day pre-established tumor xenografts after a single treatment and retained efficacy against xenografts deficient in CD19 expression, including CD19-negative variants within a background of CD19-positive lymphoma cells. BAFF-R-, but not CD19-CAR T cells, also demonstrated antitumor effects against an additional CD19 antigen loss primary patient-derived xenograft (PDX) in vivo. Methods and Results: A first-in-human clinical trial of this BAFF-R CAR T cell in B-ALL is planned. To this end, we cloned the BAFF-R CAR construct into a previously FDA approved clinical lentiviral vector (Figure, part A), and produced BAFF-R CAR T cells using both the research prototype and clinical vectors for head-to-head analysis. We observed comparable cytotoxic T lymphocyte activity against previously established leukemia and lymphoma models in vitro. In vivo, NSG mice challenged with Nalm-6 human B-ALL tumors were randomized (n=5/group) for treatment with either BAFF-R CAR produced from the research prototype or clinical vectors. The two experimental groups were indistinguishable in their antitumor efficacy (data not shown). Further, to advance IND enabling studies we also compared clinical CAR T cell manufacturing strategies. We used an early stage TN/MEM subset for CAR T cell production, as used in previous CAR T cell trials at our institution, and using TN/MEM isolated from healthy donor PBMCs achieved 36% transduction efficiency, comparable to clinical experience at City of Hope. Finally, we assessed the therapeutic effects of BAFF-R CAR TN/MEM cells in vivo. We challenged NSG mice on day 0 with 1 x 105 Nalm-6-CD19 deficient (Nalm-6-CD19KO) human ALL tumor cells and randomized them (n=5/group) to receive a single infusion of either low dose (1 x 106) or high dose (2 x 106) BAFF-R-CAR TN/MEM cells on day 10. Because transduction efficiency was 36%, 2.8 or 5.6 x 106 total T cells were infused to yield 1 or 2 x106 BAFF-R-CAR TN/MEM, respectively. Non-transduced T cells from the same donor were used as allogeneic controls (non-CAR). Tumors were monitored by bioluminescent imaging on the days indicated. Both the low and the high dose BAFF-R CAR T cell groups demonstrated complete Nalm-6 CD19KO tumor regressions, compared with controls (Figure, part B). Conclusions: Our studies support the clinical development of BAFF-R CAR T cells and a pending clinical trial of BAFF-R CAR T cell therapy in relapsed/refractory B-ALL patients who have failed prior CD19-targeted immunotherapy. Targeting BAFF-R may thereby add to existing alternative strategies to overcome relapse from CD19 antigen loss. While BAFF-R antigen loss by tumor cells is unlikely, because its expression is critically required for normal B-cell survival, this hypothesis can only be established by clinical testing. Disclosures Qin: InnoLifes: Consultancy, Equity Ownership; Pepromene Bio: Consultancy, Equity Ownership. Aldoss:AUTO1: Consultancy; Helocyte: Consultancy, Honoraria, Other: travel/accommodation/expenses; Jazz Pharmaceuticals: Honoraria, Other: travel/accommodation/expenses, Speakers Bureau; Agios: Consultancy, Honoraria. Kwak:Pepromene Bio: Consultancy, Equity Ownership, Research Funding; InnoLifes: Consultancy, Equity Ownership; Xeme BioPharma, Inc: Consultancy, Equity Ownership; Enzychem LifeSciences: Consultancy; Celltrion, Inc.: Consultancy; Celltrion Healthcare: Consultancy.

scholarly journals Switchable control over in vivo CAR T expansion, B cell depletion, and induction of memory

2018 ◽  
Vol 115 (46) ◽  
pp. E10898-E10906 ◽  
Author(s):  
Sophie Viaud ◽  
Jennifer S. Y. Ma ◽  
Ian R. Hardy ◽  
Eric N. Hampton ◽  
Brent Benish ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Rest Period ◽  
Cell System ◽  
Cellular Activity ◽  
Dosing Regimen ◽  
Car T Cells ◽  
Car T

Chimeric antigen receptor (CAR) T cells with a long-lived memory phenotype are correlated with durable, complete remissions in patients with leukemia. However, not all CAR T cell products form robust memory populations, and those that do can induce chronic B cell aplasia in patients. To address these challenges, we previously developed a switchable CAR (sCAR) T cell system that allows fully tunable, on/off control over engineered cellular activity. To further evaluate the platform, we generated and assessed different murine sCAR constructs to determine the factors that afford efficacy, persistence, and expansion of sCAR T cells in a competent immune system. We find that sCAR T cells undergo significant in vivo expansion, which is correlated with potent antitumor efficacy. Most importantly, we show that the switch dosing regimen not only allows control over B cell populations through iterative depletion and repopulation, but that the “rest” period between dosing cycles is the key for induction of memory and expansion of sCAR T cells. These findings introduce rest as a paradigm in enhancing memory and improving the efficacy and persistence of engineered T cell products.

Pax5 determines B- versus T-cell fate and does not block early myeloid-lineage development

Blood ◽  
2003 ◽  
Vol 101 (11) ◽  
pp. 4342-4346 ◽  
Author(s):  
Claudiu V. Cotta ◽  
Zheng Zhang ◽  
Hyung-Gyoon Kim ◽  
Christopher A. Klug
Keyword(s):  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Cell Fate ◽  
Nk Cell ◽  
Cell Lineage ◽  
Blood Analysis ◽  
Hematopoietic Stem ◽  
Peripheral Blood Analysis

Abstract Progenitor B cells deficient in Pax5 are developmentally multipotent, suggesting that Pax5 is necessary to maintain commitment to the B-cell lineage. Commitment may be mediated, in part, by Pax5 repression of myeloid-specific genes. To determine whether Pax5 expression in multipotential cells is sufficient to restrict development to the B-cell lineage in vivo, we enforced expression of Pax5 in hematopoietic stem cells using a retroviral vector. Peripheral blood analysis of all animals reconstituted with Pax5-expressing cells indicated that more than 90% of Pax5-expressing cells were B220+ mature B cells that were not malignant. Further analysis showed that Pax5 completely blocked T-lineage development in the thymus but did not inhibit myelopoiesis or natural killer (NK) cell development in bone marrow. These results implicate Pax5 as a critical regulator of B- versus T-cell developmental fate and suggest that Pax5 may promote commitment to the B-cell lineage by mechanisms that are independent of myeloid gene repression.

scholarly journals Early Reconstitution of Antibody Secreting Cells after Allogeneic Stem Cell Transplantation

2022 ◽  
Vol 11 (1) ◽  
pp. 270
Author(s):  
Martina Hinterleitner ◽  
Clemens Hinterleitner ◽  
Elke Malenke ◽  
Birgit Federmann ◽  
Ursula Holzer ◽  
...  
Keyword(s):  
Innate Immunity ◽  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
B Cells ◽  
Stem Cell Transplantation ◽  
B Cell ◽  
Cell Transplantation ◽  
Antibody Secreting Cells

Immune cell reconstitution after stem cell transplantation is allocated over several stages. Whereas cells mediating innate immunity recover rapidly, adaptive immune cells, including T and B cells, recover slowly over several months. In this study we investigated kinetics and reconstitution of de novo B cell formation in patients receiving CD3 and CD19 depleted haploidentical stem cell transplantation with additional in vivo T cell depletion with monoclonal anti-CD3 antibody. This model enables a detailed in vivo evaluation of hierarchy and attribution of defined lymphocyte populations without skewing by mTOR- or NFAT-inhibitors. As expected CD3+ T cells and their subsets had delayed reconstitution (<100 cells/μL at day +90). Well defined CD19+ B lymphocytes of naïve and memory phenotype were detected at day +60. Remarkably, we observed a very early reconstitution of antibody-secreting cells (ASC) at day +14. These ASC carried the HLA-haplotype of the donor and secreted the isotypes IgM and IgA more prevalent than IgG. They correlated with a population of CD19− CD27− CD38low/+ CD138− cells. Of note, reconstitution of this ASC occurred without detectable circulating T cells and before increase of BAFF or other B cell stimulating factors. In summary, we describe a rapid reconstitution of peripheral blood ASC after CD3 and CD19 depleted haploidentical stem cell transplantation, far preceding detection of naïve and memory type B cells. Incidence before T cell reconstitution and spontaneous secretion of immunoglobulins allocate these early ASC to innate immunity, eventually maintaining natural antibody levels.

scholarly journals Corticosteroids Do Not Influence the Efficacy and Kinetics of CAR-T Cells for B-Cell Acute Lymphoblastic Leukemia

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 228-228 ◽  
Author(s):  
Shuangyou Liu ◽  
Biping Deng ◽  
Jing PAN ◽  
Zhichao Yin ◽  
Yuehui Lin ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Lymphoblastic Leukemia ◽  
High Dose ◽  
Car T Cells ◽  
Steroid Group ◽  
Car T ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Kinetics Of

Cytokine release syndrome (CRS) is the most prominent and potentially life-threatening toxicity caused by chimeric antigen receptor (CAR) T cell therapy, therefore, effectively controlling severe CRS is critical to ensure patient safety. Tocilizumab, an interleukin-6 receptor antagonist, has been widely used to treat CRS, whereas it is not clear if corticosteroids could be as another optimal choice for managing CRS. We applied corticosteroids instead of tocilizumab as the first-line agent to control CRS in patients with relapsed/refractory B-cell acute lymphoblastic leukemia during CAR-T therapy. The impacts of steroids on treatment efficiency and kinetics of CAR-T cells were assessed by comparing two groups of patients who did (42 cases) or did not (26 cases) receive steroids. Patients followed up less than one month (went to other hospitals for transplantation or died within one month) were excluded. Treatment effects were evaluated on day 30 after T-cell infusion and then monthly in follow-up patients. Minimal residual disease (MRD) was detected by multiparameter flow cytometry (FCM) and quantitative PCR for fusion genes. The dynamic monitoring of CAR-T cells was performed through flow cytometric quantitation of FITC+CD3+ T cells. B-cell aplasia (BCA) was assayed by FCM. Dexamethasone or methylprednisolone or both (alternately) were administrated. Dexamethasone was used in most cases especially for patients with neurologic symptoms; methylprednisolone was preferred for patients with pulmonary or liver dysfunction, and patients accepting high dose steroids. Steroids started with low dose and could be increased if symptoms were not resolved, for severe CRS, steroids would be escalated up to dexamethasone 20mg/m2/d or more higher up to methylprednisolone 10mg/kg/d. Once CRS was improved, steroids were rapidly reduced and stopped. A total of 68 patients (28 adults and 40 children younger than 18 years) were included, 22 (32.4%) presented with extramedullary diseases (EMD), bone marrow blasts in patients without EMD varied between 5%-96.5%, 31 (45.6%) patients had an allogeneic transplantation, 54 (79.4%) cases received CD19-specific and 14 (20.6%) received CD22-specific CAR-T therapy. Forty-two (61.8%) cases, including all (10) of grade III CRS, 68.2% (30/44) of grade II CRS and 2 patients with no CRS but with GVHD (1 case) or neurotoxicity (1 case), were administered steroids, among them, 23/42 (54.8%) received high dose steroids (&gt;10mg/m2/d dexamethasone or equivalent), the duration of steroid use was 1-16 days (78.6% &lt;= 7 days); whereas 26 (38.2%) patients were not given any steroids but the supportive care. We found that there was no difference either in complete remission (CR) rate (95.2% vs 92.3%, p=.344) or in MRD negative CR rate (80.0% vs 79.2%, p=.249) between steroid and non-steroid group, verified that corticosteroids even high dose steroids did not influence the treatment response. Furthermore, we investigated the dynamics of CAR-T cells. Firstly, the expansion of CAR-T cells in peripheral blood (PB) was evaluated, the average CAR-T cell counts in steroid group were significantly higher than those in non-steroid group on D11 (p=.0302), D15 (p=.0053), D20 (p=.0045) and D30 (p=.0028), except for D7 when CAR-T cells began to expand (p=.9815), this demonstrated that steroids did not suppress the proliferation of CAR-T cells in PB. Secondly, the percentages of patients with detectable CAR-T cells in bone marrow (BM) and cerebrospinal fluid (CSF) were compared between steroid and non-steroid group, there were no differences both in BM (85.2% vs 78.6%, p=.923) and in CSF (68.6% vs 57.9%, p=.433), which implied steroids did not influence the trafficking of T-cells to BM and CSF. Thirdly, we monitored B-cell aplasia (BCA) in part of patients followed-up more than 2 months without further treatments, the percentages of patients with BCA in steroid group had no significant differences compared to non-steroid group at 2-month (p=.086) and 3-month (p=.146). Later, although limited cases left, in the steroid group, 100% of patients (4-month, 7/7; 5-month, 7/7; 6-month, 5/5) still maintained BCA and CR, indicating that corticosteroids did not impact the duration of functional CAR-T cells. In conclusion, corticosteroids do not compromise the treatment efficacy and kinetics of CAR-T cells, could be as a feasible and effective approach to manage CAR-T associated CRS. Disclosures No relevant conflicts of interest to declare.

scholarly journals High levels of interleukin 10 production in vivo are associated with tolerance in SCID patients transplanted with HLA mismatched hematopoietic stem cells.

1994 ◽  
Vol 179 (2) ◽  
pp. 493-502 ◽  
Author(s):  
R Bacchetta ◽  
M Bigler ◽  
J L Touraine ◽  
R Parkman ◽  
P A Tovo ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
T Cell ◽  
Mrna Expression ◽  
Fetal Liver ◽  
Hla Antigens ◽  
Interleukin 10 ◽  
Hematopoietic Stem

Transplantation of HLA mismatched hematopoietic stem cells in patients with severe combined immunodeficiency (SCID) can result in a selective engraftment of T cells of donor origin with complete immunologic reconstitution and in vivo tolerance. The latter may occur in the absence of clonal deletion of donor T lymphocytes able to recognize the host HLA antigens. The activity of these host-reactive T cells is suppressed in vivo, since no graft-vs. -host disease is observed in these human chimeras. Here it is shown that the CD4+ host-reactive T cell clones isolated from a SCID patient transplanted with fetal liver stem cells produce unusually high quantities of interleukin 10 (IL-10) and very low amounts of IL-2 after antigen-specific stimulation in vitro. The specific proliferative responses of the host-reactive T cell clones were considerably enhanced in the presence of neutralizing concentrations of an anti-IL-10 monoclonal antibody, suggesting that high levels of endogenous IL-10 suppress the activity of these cells. These in vitro data correlate with observations made in vivo. Semi-quantitative polymerase chain reaction analysis carried out on freshly isolated peripheral blood mononuclear cells (PBMC) of the patient indicated that the levels of IL-10 messenger RNA (mRNA) expression were strongly enhanced, whereas IL-2 mRNA expression was much lower than that in PBMC of healthy donors. In vivo IL-10 mRNA expression was not only high in the T cells, but also in the non-T cell fraction, indicating that host cells also contributed to the high levels of IL-10 in vivo. Patient-derived monocytes were found to be major IL-10 producers. Although no circulating IL-10 could be detected, freshly isolated monocytes of the patient showed a reduced expression of class II HLA antigens. However, their capacity to stimulate T cells of normal donors in primary mixed lymphocyte cultures was within the normal range. Interestingly, similar high in vivo IL-10 mRNA expressions in the T and non-T cell compartment were also observed in three SCID patients transplanted with fetal liver stem cells and in four SCID patients transplanted with T cell-depleted haploidentical bone marrow stem cells. Taken together, these data indicate that high endogenous IL-10 production is a general phenomenon in SCID patients in whom allogenic stem cell transplantation results in immunologic reconstitution and induction of tolerance. Both donor T cells and host accessory cells contribute to these high levels of IL-10, which would suppress the activity of host-reactive T cell in vivo.

scholarly journals Transcription factor Etv6 regulates functional differentiation of cross-presenting classical dendritic cells

2018 ◽  
Vol 215 (9) ◽  
pp. 2265-2278 ◽  
Author(s):  
Colleen M. Lau ◽  
Ioanna Tiniakou ◽  
Oriana A. Perez ◽  
Margaret E. Kirkling ◽  
George S. Yap ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
Transcription Factor ◽  
T Cell ◽  
Cd8 T Cells ◽  
Functional Differentiation ◽  
Specific Gene ◽  
Hematopoietic Stem

An IRF8-dependent subset of conventional dendritic cells (cDCs), termed cDC1, effectively cross-primes CD8+ T cells and facilitates tumor-specific T cell responses. Etv6 is an ETS family transcription factor that controls hematopoietic stem and progenitor cell (HSPC) function and thrombopoiesis. We report that like HSPCs, cDCs express Etv6, but not its antagonist, ETS1, whereas interferon-producing plasmacytoid dendritic cells (pDCs) express both factors. Deletion of Etv6 in the bone marrow impaired the generation of cDC1-like cells in vitro and abolished the expression of signature marker CD8α on cDC1 in vivo. Moreover, Etv6-deficient primary cDC1 showed a partial reduction of cDC-specific and cDC1-specific gene expression and chromatin signatures and an aberrant up-regulation of pDC-specific signatures. Accordingly, DC-specific Etv6 deletion impaired CD8+ T cell cross-priming and the generation of tumor antigen–specific CD8+ T cells. Thus, Etv6 optimizes the resolution of cDC1 and pDC expression programs and the functional fitness of cDC1, thereby facilitating T cell cross-priming and tumor-specific responses.

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