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01 juni 2020, Contains fulltext : 220786.pdf (Publisher’s version ) (Open Access), INTRODUCTION: Adoptive cellular immunotherapy could be an interesting new treatment option for ovarian carcinoma (OC), as research has demonstrated that OC is an immunogenic disease. In particular, natural killer (NK) cells have attracted attention due to their ability to kill tumor cells without prior sensitization. The therapeutic value of allogeneic NK cells has been first observed in hematological cancers and is increasingly being explored in solid tumors. METHODS: To substantiate the rationale for NK cell therapy in OC we performed a literature search in the Pubmed database and in the international trial register clinicaltrials.gov with attention for the effect of OC on NK cell function, the effect of current treatment on NK cell biology and the evidence on the therapeutic value of NK cell therapy against OC. RESULTS: In six clinical trials only 31 OC patients have been reported that received NK cell adoptive transfer. The majority of patients reached stable disease after NK cell therapy, with a mild pattern of side effects. In patients who received repeated infusions, more complete responses are described. All reported studies investigated the intravenous infusion of NK cells. Whereas the studies that are currently recruiting, investigate intraperitoneal infusion of allogeneic NK cells. CONCLUSION: In this review the pre-clinical evidence and current trials on NK cell immunotherapy in OC patients are summarized. Furthermore, challenges that have to be overcome for NK cell adoptive therapy to have a significant impact on disease outcome are discussed.

Contains fulltext : 202258.pdf (publisher's version ) (Closed access), Antibodies that block the interaction between programmed death ligand 1 (PD-L1) and PD-1 have shown impressive responses in subgroups of patients with cancer. PD-L1 expression in tumors seems to be a prerequisite for treatment response. However, PD-L1 is heterogeneously expressed within tumor lesions and may change upon disease progression and treatment. Imaging of PD-L1 could aid in patient selection. Previously, we showed the feasibility to image PD-L1(+) tumors in immunodeficient mice. However, PD-L1 is also expressed on immune cell subsets. Therefore, the aim of this study was to assess the potential of PD-L1 micro single-photon emission tomography/computed tomography (microSPECT/CT) using radiolabeled PD-L1 antibodies to (i) measure PD-L1 expression in two immunocompetent tumor models (syngeneic mice and humanized mice harboring PD-L1 expressing immune cells) and (ii) monitor therapy-induced changes in tumor PD-L1 expression. We showed that radiolabeled PD-L1 antibodies accumulated preferentially in PD-L1(+) tumors, despite considerable uptake in certain normal lymphoid tissues (spleen and lymph nodes) and nonlymphoid tissues (duodenum and brown fat). PD-L1 microSPECT/CT imaging could also distinguish between high and low PD-L1-expressing tumors. The presence of PD-L1(+) immune cells did not compromise tumor uptake of the human PD-L1 antibodies in humanized mice, and we demonstrated that radiotherapy-induced upregulation of PD-L1 expression in murine tumors could be monitored with microSPECT/CT imaging. Together, these data demonstrate that PD-L1 microSPECT/CT is a sensitive technique to detect variations in tumor PD-L1 expression, and in the future, this technique may enable patient selection for PD-1/PD-L1-targeted therapy.

Contains fulltext : 205181.pdf (publisher's version ) (Closed access), Ovarian cancer is the most lethal gynecological malignancy in the developed world. In spite of intensive research, the mortality has hardly decreased over the past twenty years. This necessitates the exploration of novel therapeutic modalities. Transient protein expression through delivery of mRNA is emerging as a highly promising option. In contrast to gene therapy there is no risk of integration into the genome. Here, we explore the expression of mRNA in models of ovarian cancer of increasing complexity. The cell-penetrating peptide (CPP) PepFect 14 (PF14) was used to formulate CPP-mRNA nanoparticles. Efficient expression of a reporter protein was achieved in two-dimensional tissue cultures and in three-dimensional cancer cell spheroids. PF14 nanoparticles greatly outperformed a lipid-based transfection agent in vivo, leading to expression in various cell types of tumor associated tissue. Protein expression was restricted to the peritoneal cavity. Messenger RNA expression across different cell types was confirmed in primary ovarian cancer explants. As ovarian cancer is confined to the peritoneal cavity in most cases, the results create the basis for applications in which the tumor microenvironment is transiently modified through protein expression.

Radboud University, 15 oktober 2019, Promotor : Jansen, J.H. Co-promotores : Dolstra, H., Waart, A.B. van der, Contains fulltext : 207786.pdf (publisher's version ) (Open Access)

Contains fulltext : 203697.pdf (publisher's version ) (Closed access)

Contains fulltext : 202816.pdf (publisher's version ) (Open Access), INTRODUCTION: Recurrent ovarian carcinoma has dismal prognosis, but control of disease and prolonged survival are possible in some patients. The estimated 5-year survival is 46% for all stages of ovarian cancer, and only 28% for metastasized disease. Notably, the majority of women with ovarian cancer are diagnosed with stage III or IV disease with a high recurrence rate. As most women with relapsed or metastatic cancer will die of progressive disease, there is an urgent need for novel therapeutic strategies. The primary aim of our study is to evaluate safety and toxicity of intraperitoneal infusion of ex vivo-expanded natural killer cells (NK), generated from CD34+ umbilical cord blood (UCB) progenitor cells, with and without a preceding non-myeloablative immunosuppressive conditioning regimen in patients suffering from recurrent ovarian cancer. The secondary objectives are to compare the in vivo lifespan, expansion, and biological activity of intraperitoneally infused NK cell products with or without preparative chemotherapy, as well as evaluate effects on disease load. METHODS: In this phase I safety trial, 12 patients who are suffering from recurrent ovarian cancer, detected by a significant rise in serum level of CA-125 on two successive time points, will be included. Prior to UCB-NK cell infusion, a laparoscopy is performed to place a catheter in the peritoneal cavity. The first cohort of three patients will receive a single intraperitoneal infusion of 1.5-3x10 UCB-NK cells, generated ex vivo from CD34+ hematopoietic progenitor cells obtained from an allogeneic UCB unit, without a preparative chemotherapy regimen. The second group of three patients will be treated with a similar dose of UCB-NK cells following a preparative four days non-myeloablative immunosuppressive conditioning regimen with cyclophosphamide and fludarabine (Cy/Flu). If no severe toxicity is seen in these 6 patients, an extension cohort of 6 patients will be included to answer the secondary o

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Contains fulltext : 208018.pdf (publisher's version ) (Open Access)

Contains fulltext : 202774.pdf (publisher's version ) (Closed access)

Contains fulltext : 200311.pdf (publisher's version ) (Open Access)

Radboud University, 18 mei 2018, Promotor : Jansen, J.H. Co-promotores : Hobo, W.A., Dolstra, H., Contains fulltext : 190672.pdf (publisher's version ) (Open Access)

Contains fulltext : 194897.pdf (publisher's version ) (Open Access)

Contains fulltext : 196608.pdf (publisher's version ) (Open Access)

Contains fulltext : 190712.pdf (publisher's version ) (Closed access), New immunotherapeutic interventions have revolutionized cancer treatment. The immune responsiveness of acute myeloid leukaemia (AML) was first demonstrated by allogeneic stem cell transplantation. In addition, milder immunotherapeutic approaches are exploited. However, the long-term efficacy of these therapies is hampered by various immune resistance and editing mechanisms. In this regard, co-inhibitory signalling pathways have been shown to play a crucial role. Via up-regulation of inhibitory checkpoints, tumour-reactive T cell and Natural Killer cell responses can be strongly impeded. Accordingly, the introduction of checkpoint inhibitors targeting CTLA-4 (CTLA4) and PD-1 (PDCD1, CD279)/PD-L1 (CD274, PDCD1LG1) accomplished a breakthrough in cancer treatment, with impressive clinical responses. Numerous new co-inhibitory players and novel combination therapies are currently investigated for their potential to boost anti-tumour immunity and improve survival of cancer patients. Although the challenge here remains to avoid severe systemic toxicity. This review addresses the involvement of co-inhibitory signalling in AML immune evasion and discusses the opportunities for checkpoint blockers in AML treatment.

Contains fulltext : 191170.pdf (publisher's version ) (Closed access)

Contains fulltext : 183885.pdf (publisher's version ) (Closed access)

Radboud University, 5 april 2018, Promotores : Jansen, J.H., Radstake, T.R.D.J. Co-promotores : Dolstra, H., Hobo, W.A., Contains fulltext : 187354.pdf (publisher's version ) (Open Access)

Contains fulltext : 182504.pdf (publisher's version ) (Closed access), Allogeneic stem cell transplantation (allo-SCT) with or without donor lymphocyte infusions (DLI) is the only curative option for several hematological malignancies. Unfortunately, allo-SCT is often associated with GvHD, and patients often relapse. We therefore aim to improve the graft-versus-tumor effect, without increasing the risk of GvHD, by targeting hematopoietic lineage-restricted and tumor-associated minor histocompatibility antigens using peptide-loaded dendritic cell (DC) vaccinations. In the present multicenter study, we report the feasibility, safety and efficacy of this concept. We treated nine multiple myeloma patients with persistent or relapsed disease after allo-SCT and a previous DLI, with donor monocyte-derived mHag-peptide-loaded DC vaccinations combined with a second DLI. Vaccinations were well tolerated and no occurrence of GvHD was observed. In five out of nine patients, we were able to show the induction of mHag-specific CD8+ T cells in peripheral blood. Five out of nine patients, of which four developed mHag-specific T cells, showed stable disease (SD) for 3.5-10 months. This study shows that mHag-based donor monocyte-derived DC vaccination combined with DLI is safe, feasible and capable of inducing objective mHag-specific T-cell responses. Future research should focus on further improvement of the vaccination strategy, toward translating the observed T-cell responses into robust clinical responses.

Contains fulltext : 177224.pdf (publisher's version ) (Closed access), Although allogeneic stem cell transplantation (allo-SCT) can elicit graft-versus-tumor (GVT) immunity, patients often relapse due to residual tumor cells. As essential orchestrators of the immune system, vaccination with dendritic cells (DC) is an appealing strategy to boost the GVT response. Nevertheless, durable clinical responses after DC vaccination are still limited, stressing the need to improve current DC vaccines. Aiming to empower DC potency, we engineered monocyte-derived DCs to deprive them of ligands for the immune checkpoint regulated by programmed death 1 (PD-1). We also equipped them with interleukin (IL)-15 "transpresentation" skills. Transfection with short interfering (si)RNA targeting the PD-1 ligands PD-L1 and PD-L2, in combination with IL15 and IL15Ralpha mRNA, preserved their mature DC profile and rendered the DCs superior in inducing T-cell proliferation and IFNgamma and TNFalpha production. Translated into an ex vivo hematological disease setting, DCs deprived of PD-1 ligands (PD-L), equipped with IL15/IL15Ralpha expression, or most effectively, both, induced superior expansion of minor histocompatibility antigen-specific CD8+ T cells from transplanted cancer patients. These data support the combinatorial approach of in situ suppression of the PD-L inhibitory checkpoints with DC-mediated IL15 transpresentation to promote antigen-specific T-cell responses and, ultimately, contribute to GVT immunity. Cancer Immunol Res; 5(8); 710-5. (c)2017 AACR.

Contains fulltext : 178035.pdf (publisher's version ) (Open Access)

Contains fulltext : 177260.pdf (publisher's version ) (Closed access), Purpose: Older acute myeloid leukemia (AML) patients have a poor prognosis; therefore, novel therapies are needed. Allogeneic natural killer (NK) cells have been adoptively transferred with promising clinical results. Here, we report the first-in-human study exploiting a unique scalable NK-cell product generated ex vivo from CD34+ hematopoietic stem and progenitor cells (HSPC) from partially HLA-matched umbilical cord blood units.Experimental Design: Ten older AML patients in morphologic complete remission received an escalating HSPC-NK cell dose (between 3 and 30 x 106/kg body weight) after lymphodepleting chemotherapy without cytokine boosting.Results: HSPC-NK cell products contained a median of 75% highly activated NK cells, with <1 x 104 T cells/kg and <3 x 105 B cells/kg body weight. HSPC-NK cells were well tolerated, and neither graft-versus-host disease nor toxicity was observed. Despite no cytokine boosting being given, transient HSPC-NK cell persistence was clearly found in peripheral blood up to 21% until day 8, which was accompanied by augmented IL15 plasma levels. Moreover, donor chimerism up to 3.5% was found in bone marrow. Interestingly, in vivo HSPC-NK cell maturation was observed, indicated by the rapid acquisition of CD16 and KIR expression, while expression of most activating receptors was sustained. Notably, 2 of 4 patients with minimal residual disease (MRD) in bone marrow before infusion became MRD negative (<0.1%), which lasted for 6 months.Conclusions: These findings indicate that HSPC-NK cell adoptive transfer is a promising, potential "off-the-shelf" translational immunotherapy approach in AML. Clin Cancer Res; 23(15); 4107-18. (c)2017 AACR.

Radboud University, 24 november 2017, Promotores : Blijlevens, N.M.A., Jansen, J.H. Co-promotores : Dolstra, H., Schaap, N.P.M., Contains fulltext : 178256.pdf (publisher's version ) (Open Access)

Contains fulltext : 174543.pdf (publisher's version ) (Open Access), Because of the potent graft-versus-tumor (GVT) effect, allogeneic stem cell transplantation (alloSCT) can be a curative therapy for hematological malignancies. However, relapse remains the most frequent cause of treatment failure, illustrating the necessity for development of adjuvant post-transplant therapies to boost GVT immunity. Dendritic cell (DC) vaccination is a promising strategy in this respect, in particular, where distinct biologic functions of naturally occurring DC subsets, i.e. myeloid DCs (mDCs) and plasmacytoid DCs (pDCs), are harnessed. However, it is challenging to obtain high enough numbers of primary DC subsets from blood for immunotherapy due to their low frequencies. Therefore, we present here an ex vivo GMP-compliant cell culture protocol for generating different DC subsets from CD34+ hematopoietic stem and progenitor cells (HSPCs) of alloSCT donor origin. High numbers of BDCA1+ mDCs and pDCs could be generated, sufficient for multiple vaccination cycles. These HSPC-derived DC subsets were highly potent in inducing antitumor immune responses in vitro. Notably, HSPC-derived BDCA1+ mDCs were superior in eliciting T cell responses. They efficiently primed naive T cells and robustly expanded patient-derived minor histocompatibility antigen (MiHA)-specific T cells. Though the HSPC-pDCs also efficiently induced T cell responses, they exhibited superior capacity in activating NK cells. pDC-primed NK cells highly upregulated TRAIL and possessed strong cytolytic capacity against tumor cells. Collectively, these findings indicate that HSPC-derived DC vaccines, comprising both mDCs and pDCs, may possess superior potential to boost antitumor immunity post alloSCT, due to their exceptional T cell and NK cell stimulatory capacity.

Contains fulltext : 177298.pdf (publisher's version ) (Closed access), OBJECTIVE: Plasmacytoid dendritic cells (PDCs) are a critical source of type I interferons (IFNs) that can contribute to the onset and maintenance of autoimmunity. Molecular mechanisms leading to PDC dysregulation and a persistent type I IFN signature are largely unexplored, especially in patients with systemic sclerosis (SSc), a disease in which PDCs infiltrate fibrotic skin lesions and produce higher levels of IFNalpha than those in healthy controls. This study was undertaken to investigate potential microRNA (miRNA)-mediated epigenetic mechanisms underlying PDC dysregulation and type I IFN production in SSc. METHODS: We performed miRNA expression profiling and validation in highly purified PDCs obtained from the peripheral blood of 3 independent cohorts of healthy controls and SSc patients. Possible functions of miRNA-618 (miR-618) on PDC biology were identified by overexpression in healthy PDCs. RESULTS: Expression of miR-618 was up-regulated in PDCs from SSc patients, including those with early disease who did not present with skin fibrosis. IFN regulatory factor 8, a crucial transcription factor for PDC development and activation, was identified as a target of miR-618. Overexpression of miR-618 reduced the development of PDCs from CD34+ cells in vitro and enhanced their ability to secrete IFNalpha, mimicking the PDC phenotype observed in SSc patients. CONCLUSION: Up-regulation of miR-618 suppresses the development of PDCs and increases their ability to secrete IFNalpha, potentially contributing to the type I IFN signature observed in SSc patients. Considering the importance of PDCs in the pathogenesis of SSc and other diseases characterized by a type I IFN signature, miR-618 potentially represents an important epigenetic target to regulate immune system homeostasis in these conditions.

Objective. Plasmacytoid dendritic cells (PDCs) are a critical source of type I interferons (IFNs) that can contribute to the onset and maintenance of autoimmunity. Molecular mechanisms leading to PDC dysregulation and a persistent type I IFN signature are largely unexplored, especially in patients with systemic sclerosis (SSc), a disease in which PDCs infiltrate fibrotic skin lesions and produce higher levels of IFN alpha than those in healthy controls. This study was undertaken to investigate potential microRNA (miRNA)-mediated epigenetic mechanisms underlying PDC dysregulation and type I IFN production in SSc. Methods. We performed miRNA expression profiling and validation in highly purified PDCs obtained from the peripheral blood of 3 independent cohorts of healthy controls and SSc patients. Possible functions of miRNA-618 (miR-618) on PDC biology were identified by overexpression in healthy PDCs. Results. Expression of miR-618 was up-regulated in PDCs from SSc patients, including those with early disease who did not present with skin fibrosis. IFN regulatory factor 8, a crucial transcription factor for PDC development and activation, was identified as a target of miR-618. Overexpression of miR-618 reduced the development of PDCs from CD34+ cells in vitro and enhanced their ability to secrete IFN alpha, mimicking the PDC phenotype observed in SSc patients. Conclusion. Up-regulation of miR-618 suppresses the development of PDCs and increases their ability to secrete IFN alpha, potentially contributing to the type I IFN signature observed in SSc patients. Considering the importance of PDCs in the pathogenesis of SSc and other diseases characterized by a type I IFN signature, miR-618 potentially represents an important epigenetic target to regulate immune system homeostasis in these conditions.