Verena Pfirrmann
Goethe University Frankfurt
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Publication
Featured researches published by Verena Pfirrmann.
International Journal of Cancer | 2016
Sarah Oelsner; Juliane Wagner; Miriam E. Friede; Verena Pfirrmann; Sabrina Genßler; Eva Rettinger; Christian J. Buchholz; Heike Pfeifer; Ralf Schubert; Oliver G. Ottmann; Evelyn Ullrich; Peter Bader; Winfried S. Wels
Pre‐emptive cancer immunotherapy by donor lymphocyte infusion (DLI) using cytokine‐induced killer (CIK) cells may be beneficial to prevent relapse with a reduced risk of causing graft‐versus‐host‐disease. CIK cells are a heterogeneous effector cell population including T cells (CD3+ CD56−), natural killer (NK) cells (CD3−CD56+) and natural killer T (T‐NK) cells (CD3+ CD56+) that exhibit non‐major histocompatibility complex (MHC)‐restricted cytotoxicity and are generated by ex vivo expansion of peripheral blood mononuclear cells in the presence of interferon (IFN)‐γ, anti‐CD3 antibody, interleukin‐2 (IL‐2) and interleukin‐15 (IL‐15). To facilitate selective target‐cell recognition and enhance specific cytotoxicity against B‐cell acute lymphoblastic leukemia (B‐ALL), we transduced CIK cells with a lentiviral vector encoding a chimeric antigen receptor (CAR) that carries a composite CD28‐CD3ζ domain for signaling and a CD19‐specific scFv antibody fragment for cell binding (CAR 63.28.z). In vitro analysis revealed high and specific cell killing activity of CD19‐targeted CIK/63.28.z cells against otherwise CIK‐resistant cancer cell lines and primary B‐ALL blasts, which was dependent on CD19 expression and CAR signaling. In a xenograft model in immunodeficient mice, treatment with CIK/63.28.z cells in contrast to therapy with unmodified CIK cells resulted in complete and durable molecular remissions of established primary pre‐B‐ALL. Our results demonstrate potent antileukemic activity of CAR‐engineered CIK cells in vitro and in vivo, and suggest this strategy as a promising approach for adoptive immunotherapy of refractory pre‐B‐ALL.
Cytotherapy | 2015
Verena Pfirrmann; Sarah Oelsner; Eva Rettinger; Sabine Huenecke; Halvard Bonig; Michael Merker; Winfried S. Wels; Jindrich Cinatl; Ralf Schubert; Thomas Klingebiel; Peter Bader
BACKGROUND AIMS Human cytomegalovirus (CMV) infection and reactivation is a leading complication of allogeneic hematopoietic stem cell transplantation (HSCT). In addition to drug treatment, the adoptive transfer of virus-specific T cells to restore cellular immunity has become a standard therapy after allogeneic HSCT. We recently demonstrated potent anti-leukemic activity of interleukin (IL)-15-activated cytokine-induced killer (CIK) cells. With the use of the same expansion protocol, we asked whether concurrent CMV antigen-pulsing might generate CIK cells with anti-leukemic and anti-CMV activity. METHODS CIK cells expanded in the presence of interferon-γ, IL-2, IL-15 and anti-CD3 antibody were pulsed once with CMV(pp65) peptide pool. CMV-specific CIK (CIK(pp65)) and conventional CIK cells were phenotypically and functionally characterized according to their cytokine secretion pattern, degranulation capacity and T-cell receptor (TCR)-mediated and NKG2D-mediated cytotoxicity. RESULTS We demonstrated that among CIK cells generated from CMV-seropositive donors, a single stimulation with CMV(pp65) protein co-expanded cytotoxic CMV-specific cells without sacrificing anti-tumor reactivity. Cells generated in this fashion lysed CMV(pp65)-loaded target cells and CMV-infected fibroblasts but also leukemic cells. Meanwhile, the alloreactive potential of CIK(pp65) cells remained low. Interestingly, CMV reactivity was TCR-mediated and CMV-specific cells could be found in CD3(+)CD8(+)CD56(+/-) cytotoxic T-cell subpopulations. CONCLUSIONS We provide an efficient method to generate CIK(pp65) cells that may represent a useful cell therapy approach for preemptive immunotherapy in patients who have both an apparent risk of CMV and impending leukemic relapse after allogeneic stem cell transplantation.
Frontiers in Immunology | 2017
Sabine Huenecke; Claudia Cappel; Ruth Esser; Verena Pfirrmann; Emilia Salzmann-Manrique; Sibille Betz; Eileen Keitl; Julia Banisharif-Dehkordi; Shahrzad Bakhtiar; Christoph Königs; Andrea Jarisch; Jan Soerensen; Evelyn Ullrich; Thomas Klingebiel; Peter Bader; Melanie Bremm
Natural killer (NK) cells play an important role following allogeneic hematopoietic stem cell transplantation (HSCT) exerting graft-versus-leukemia/tumor effect and mediating pathogen-specific immunity. Although NK cells are the first donor-derived lymphocytes reconstituting post-HSCT, their distribution of CD56++CD16− (CD56bright), CD56++CD16+ (CD56intermediate=int), and CD56+CD16++ (CD56dim) NK cells is explicitly divergent from healthy adults, but to some extent comparable to the NK cell development in early childhood. The proportion of CD56bright/CD56int/CD56dim changed from 15/8/78% in early childhood to 6/4/90% in adults, respectively. Within this study, we first compared the NK cell reconstitution post-HSCT to reference values of NK cell subpopulations of healthy children. Afterward, we investigated the reconstitution of NK cell subpopulations post-HSCT in correlation to acute graft versus host disease (aGvHD) and chronic graft versus host disease (cGvHD) as well as to viral infections. Interestingly, after a HSCT follow-up phase of 12 months, the distribution of NK cell subpopulations largely matched the 50th percentile of the reference range for healthy individuals. Patients suffering from aGvHD and cGvHD showed a delayed reconstitution of NK cells. Remarkably, within the first 2 months post-HSCT, patients suffering from aGvHD had significantly lower levels of CD56bright NK cells compared to patients without viral infection or without graft versus host disease (GvHD). Therefore, the amount of CD56bright NK cells might serve as an early prognostic factor for GvHD development. Furthermore, a prolonged and elevated peak in CD56int NK cells seemed to be characteristic for the chronification of GvHD. In context of viral infection, a slightly lower CD56 and CD16 receptor expression followed by a considerable reduction in the absolute CD56dim NK cell numbers combined with reoccurrence of CD56int NK cells was observed. Our results suggest that a precise analysis of the reconstitution of NK cell subpopulations post-HSCT might indicate the occurrence of undesired events post-HSCT such as severe aGvHD.
Frontiers in Pediatrics | 2014
Eva Rettinger; Andreas Glatthaar; Behnaz Ahangarian Abhari; Sarah Oelsner; Verena Pfirrmann; Sabine Huenecke; Selim Kuçi; Hermann Kreyenberg; Andre Willasch; Thomas Klingebiel; Simone Fulda; Peter Bader
Allogeneic hematopoietic stem cell transplantation (HSCT) is an established treatment option for high-risk hematological malignancies, and may also be offered to patients with solid malignancies refractory to conventional therapies. In case of patients’ relapse, refractory tumor cells may then be targeted by cellular therapy-based combination strategies. Here, we investigated the potential of small molecule IAP (SMAC mimetic) BV6 in increasing cytokine-induced killer (CIK) cell-mediated cytotoxicity against different tumor targets. Four-hour pre-incubation with 2.5 μMol BV6 moderately enhanced CIK cell-mediated lysis of hematological (H9, THP-1, and Tanoue) and solid malignancies (RH1, RH30, and TE671). However, BV6 also increased apoptosis of non-malignant cells like peripheral blood mononuclear cells and most notably had an inhibitory effect on immune cells potentially limiting their cytotoxic potential. Hence, cytotoxicity increased in a dose-dependent manner when BV6 was removed before CIK cells were added to tumor targets. However, cytotoxic potential was not further increasable by extending BV6 pre-incubation period of target cells from 4 to 12 h. Molecular studies revealed that BV6 sensitization of target cells involved activation of caspases. Here, we provide evidence that SMAC mimetic may sensitize targets cells for CIK cell-induced cell death. However, BV6 also increased apoptosis of non-malignant cells like CIK cells and peripheral mononuclear cells. These findings may therefore be important for cell- and small molecule IAP-based combination therapies of resistant cancers after allogeneic HSCT.
Bone Marrow Transplantation | 2017
Andre Willasch; Emilia Salzmann-Manrique; Thomas Krenn; M Duerken; Joerg Faber; J Opper; Hermann Kreyenberg; R Bager; Sabine Huenecke; Claudia Cappel; Melanie Bremm; Verena Pfirrmann; Michael Merker; Evelyn Ullrich; Shahrzad Bakhtiar; Eva Rettinger; Andrea Jarisch; Jan Soerensen; Thomas Klingebiel; Peter Bader
Therapy for post-transplant relapse of paediatric ALL is limited. Standardised curative approaches are not available. We hereby describe our local procedure in this life-threatening situation. A total of 101 ALL patients received their first allogeneic stem cell transplantation (SCT) in our institution. After relapse, our primary therapeutic goal was to cure the patient with high-dose chemotherapy or specific immunotherapy (HDCHT/SIT) followed by a second SCT from a haploidentical donor (transplant approach). If this was not feasible, low-dose chemotherapy and donor lymphocyte infusions (LDCHT+DLI) were offered (non-transplant approach). A total of 23 patients suffered a post-transplant relapse. Eight patients received HDCHT/SIT, followed by haploidentical SCT in 7/8. Ten received LDCHT+DLI. The eight patients treated with a second transplant and the ten treated with the non-transplant approach had a 4-year overall survival of 56% and 40%, respectively (P=0.232). Prerequisites for successful treatment of post-transplant relapse by either a second transplant or experimental non-transplant approaches are good clinical condition and the capacity to achieve haematological remission by the induction treatment element.
Pediatric Blood & Cancer | 2016
Claudia Cappel; Sabine Huenecke; Anica Suemmerer; Stephanie Erben; Eva Rettinger; Verena Pfirrmann; Annekathrin Heinze; Olga Zimmermann; Thomas Klingebiel; Evelyn Ullrich; Peter Bader; Melanie Bremm
Neuroblastoma (NB) is the most common solid extracranial tumor in childhood. Despite advances in therapy, the prognosis is poor and optimized therapies are urgently needed. Therefore, we investigated the antitumor potential of interleukin‐15 (IL‐15)‐activated cytokine‐induced killer (CIK) cells against different NB cell lines.
Oncotarget | 2017
Michael Merker; Verena Pfirrmann; Sarah Oelsner; Simone Fulda; Thomas Klingebiel; Winfried S. Wels; Peter Bader; Eva Rettinger
Pediatric patients with recurrent, refractory or advanced soft tissue sarcoma (STS) who are simultaneously showing signs of cumulative treatment toxicity are in need of novel therapies. In this preclinical analysis, we identified ErbB2 as a targetable antigen on STS cells and used cytokine-induced killer (CIK) cells transduced with the lentiviral 2nd-generation chimeric antigen receptor (CAR) vector pS-5.28.z-IEW to target ErbB2-positive tumors. Solely CIK cell subsets with the CD3+ T cell phenotype showed up to 85% cell surface expression of the respective CAR. A comparison of wildtype (WT), mock-vector and ErbB2-CAR-CIK cells showed, that engineered cells exhibited diminished in vitro expansion, retained WT CIK cell phenotype with higher percentages of differentiated effector memory/effector cells. Activating natural killer (NK) cell receptor NKG2D-restricted target cell recognition and killing of WT and ErbB2-CAR-CIK cells was maintained against ErbB2-negative tumors, while ErbB2-CAR-CIK cells demonstrated significantly increased cytotoxicity against ErbB2-positive targets, including primary tumors. ErbB2-CAR- but not WT CIK cells proliferated, infiltrated and efficiently lysed tumor cell monolayers as well as 3D tumor spheroids. Here, we demonstrate a potential cell therapeutic approach using ErbB2-CAR-CIK cells for the recognition and elimination of tumor cells expressing ErbB2, which we identified as a targetable antigen on high-risk STS cells.
Cytotherapy | 2018
Lisa-Marie Pfeffermann; Verena Pfirrmann; Sabine Huenecke; Melanie Bremm; Halvard Bonig; Hans-Michael Kvasnicka; Thomas Klingebiel; Peter Bader; Eva Rettinger
BACKGROUND Prolonged immunosuppression or delayed T-cell recovery may favor Epstein-Barr virus (EBV) infection or reactivation after allogeneic hematopoietic stem cell transplantation (HSCT), which can lead to post-transplant lymphoproliferative disease (PTLD) and high-grade malignant B-cell lymphoma. Cytokine-induced killer (CIK) cells with dual specific anti-tumor and virus-specific cellular immunity may be applied in this context. METHODS CIK cells with EBV-specificity were generated from peripheral blood mononuclear cells (PBMCs), expanded in the presence of interferon-γ, anti-CD3, interleukin (IL)-2 and IL-15 and were pulsed twice with EBV consensus peptide pool. CIK cells with EBV-specificity and conventional CIK cells were phenotypically and functionally analyzed. Additionally, CIK cells with EBV-specificity were applied to a patient with EBV-related PTLD rapidly progressing to highly aggressive B-cell lymphoma on a compassionate use basis after approval and agreement by the regulatory authorities. RESULTS Pre-clinical analysis showed that generation of CIK cells with EBV-specificity was feasible. In vitro cytotoxicity analyses showed increased lysis of EBV-positive target cells, enhanced proliferative capacity and increased secretion of cytolytic and proinflammatory cytokines in the presence of EBV peptide-displaying target cells. In addition, 1 week after infusion of CIK cells with EBV-specificity, the patients highly aggressive B-cell lymphoma persistently disappeared. CIK cells with EBV-specificity remained detectable for up to 32 days after infusion and infusion did not result in acute toxicity. DISCUSSION The transfer of both anti-cancer potential and T-cell memory against EBV infection provided by EBV peptide-induced CIK cells might be considered a therapy for EBV-related PTLD.
Bone Marrow Transplantation | 2018
Shahrzad Bakhtiar; Emilia Salzmann-Manrique; Martin Hutter; Thomas Krenn; Matthias Duerken; Joerg Faber; Harald Reinhard; Hermann Kreyenberg; Sabine Huenecke; Claudia Cappel; Melanie Bremm; Verena Pfirrmann; Michael Merker; Anke Barnbrock; Stefan Schöning; Andre Willasch; Eva Rettinger; Jan Soerensen; Thomas Klingebiel; Andrea Jarisch; Peter Bader
Allogeneic hematopoietic stem cell transplantation (alloHSCT) has become a well-established treatment option for many patients suffering from malignant and non-malignant diseases. In the past decade, high-resolution HLA-typing, remission surveillance, pre-emptive immune intervention, and standardisation in supportive care measures have substantially improved transplant outcomes. This retrospective study evaluated transplant procedures in 162 paediatric patients with acute lymphoblastic leukaemia (n = 124) or acute myeloid leukaemia (n = 38) who received their first alloHSCT in our institution over an 11-year period. We observed a significant reduction in risk of non-relapse mortality (NRM) over time (HR = 0.34, 95% CI 0.12–0.98; P = 0.05), the 4-year NRM estimate decreased from 20% in 2005–2008 to 7% in 2012–2016 (P = 0.02) and an increase in survival after relapse. There was no significant difference in patients who received a graft from a sibling, haplo, or an unrelated donor with regard to their overall survival (P = 0.45), event-free survival (P = 0.61), and non-relapse mortality (P = 0.19). Our data suggest that a specific transplant infrastructure with a highly experienced team in an accredited transplant centre likely contributes to better transplant outcomes for acute leukaemia patients in complete remission regardless of donor type.
Cancer immunology research | 2016
Sarah Oelsner; Juliane Wagner; Miriam E. Friede; Verena Pfirrmann; Eva Rettinger; Ralf Schubert; Heike Pfeifer; Evelyn Ullrich; Peter Bader; Winfried S. Wels
Pre-emptive immunotherapy after HSC transplantation based on minimal residual disease (MRD) status with donor lymphocyte infusions (DLI) using cytokine-induced killer (CIK) cells may be beneficial to prevent relapse with a reduced risk of causing graft-versus-host-disease (GvHD) compared to conventional T cell infusion. CIK cells are a heterogeneous effector cell population including T cells (CD3+CD56-), natural killer (NK) cells (CD3-CD56+) and natural killer T (T-NK) cells (CD3+CD56+), that exhibit non-MHC-restricted cytotoxic activity and are generated by ex vivo expansion of peripheral blood mononuclear cells (PBMC) through the addition of interferon (IFN)-γ, anti-CD3 antibody, IL-2 and IL-15. While CIK cells have shown potent in vivo activity against various cancer types such as lymphomas or colorectal cancer, their cytotoxicity against B-ALL, characterized by the expression of CD19, has been limited. Hence, retargeting of CIK cells using chimeric antigen receptors (CARs) to facilitate selective target cell recognition and enhance specific cytotoxicity represents a promising approach. CAR comprise an extracellular scFv antibody fragment as an antigen-binding domain, linked via a flexible hinge region and a transmembrane domain to an intracellular signaling moiety such as CD3 zeta chain (first generation CAR), or zeta chain fused to a co-stimulatory protein domain such as CD28 (second generation CAR). We established an optimized protocol for transduction of CIK cells with CD19-specific lentiviral CAR constructs, and characterized cells for expression of an EGFP marker gene and CAR surface expression. Effects of exposure to lentiviral vector particles on the development of CIK cell subpopulations were monitored. In in vitro cytotoxicity assays we could demonstrate potent and selective cytotoxicity of retargeted CIK cells towards established cancer cell lines expressing CD19 and primary pre-B-ALL blasts. In addition, we observed significantly enhanced degranulation of CAR-CIK cells upon target cell contact and showed increased secretion of pro-inflammatory cytokines, while no secretion of immunosuppressive IL-10 could be detected. Cytotoxity towards non-malignant allo-PBMCs remained low. In addition, we investigated anti-leukemic activity of retargeted CIK cells in vivo in NOD/SCID common gamma chain knockout (NSG) mouse models using bioluminescence and fluorescence imaging. We observed potent reduction of tumor load after only two CIK cell applications in mice with an engrafted primary pre-B-ALL yielding long-term survivors with a negative MRD status (4/6). In a separate model, we demonstrated migration of CD19-specific CIK cells into local, subcutaneous tumor sites. In conclusion, CAR-CIK cells represent a promising alternative to CAR-T cell therapy, as relatively high numbers of conventional CIK cells have already been infused in clinical trials, which demonstrated a low risk of causing severe side effects. Citation Format: Sarah Oelsner, Juliane Wagner, Miriam E. Friede, Verena Pfirrmann, Eva Rettinger, Ralf Schubert, Heike Pfeifer, Evelyn Ullrich, Peter Bader, Winfried S. Wels. Genetically modified cytokine-induced killer (CIK) cells for targeted cancer therapy. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr A164.