Martin Cremer

A fusogenic dengue virus-derived peptide enhances antitumor efficacy of an antibody–ribonuclease fusion protein targeting the EGF receptor

Due to its frequent overexpression in a variety of solid tumors the epidermal growth factor receptor (EGFR) is a well-established target for therapeutic interventions in epithelial cancers. In order to target EGFR in head and neck cancer, we have generated a ribonuclease (RNase) fusion protein comprising a humanized anti-EGFR antibody single-chain Fv fragment (scFv) and Ranpirnase, an RNase from Rana pipiens. Fusion of Ranpirnase to the N-terminus of the scFv via a flexible glycine-serine linker (G4S)3 resulted in very poor cytotoxicity of the fusion protein. As endosomal accumulation and lysosomal degradation have been reported to diminish the antitumor efficacy of ribonuclease or toxin-based immunoagents, we explored a fusion peptide from dengue virus that has been reported to be involved in the endosomal escape of the virus. This peptide was introduced as a linker between Ranpirnase and the scFv moiety. The modified immunoRNase exhibited exceptionally high cytotoxicity toward EGFR-expressing head and neck cell lines without affecting specificity. These results indicate that endosomal entrapment needs to be considered for Ranpirnase-based immunoagents and might be overcome by the use of tailored transduction domains from viral proteins.

An EGF receptor targeting Ranpirnase-diabody fusion protein mediates potent antitumour activity in vitro and in vivo.

Cytotoxic ribonucleases such as the leopard frog derivative Ranpirnase (Onconase(®)) have emerged as a valuable new class of cancer therapeutics. Clinical trials employing single agent Ranpirnase in cancer patients have demonstrated significant clinical activity and surprisingly low immunogenicity. However, dose-limiting toxicity due to unspecific uptake of the RNase into non-cancerous cells is reached at relatively low concentrations of > 1 mg/m(2). We have in the present study generated a dimeric anti-EGFR Ranpirnase-diabody fusion protein capable to deliver two Ranpirnase moieties per molecule to EGFR-positive tumour cells. We show that this compound mediated far superior efficacy for killing EGFR-positive tumour cells than a monomeric counterpart. Most importantly, cell killing was restricted to EGFR-positive target cells and no dose-limiting toxicity of Ranpirnase-diabody was observed in mice. These data indicate that by targeted delivery of Ranpirnase non-selective toxicity can be abolished and suggests Ranpirnase-diabody as a promising new drug for therapeutic interventions in EGFR-positive cancers.

Successful collection of peripheral blood stem cells upon VIDE chemomobilization in sarcoma patients

In patients with Ewing sarcoma and some distinct subgroups of soft tissue sarcoma (STS), a quantitatively sufficient autologous peripheral blood stem cell (PBSC) collection for stem cell support might facilitate treatment continuation, dose‐intensification, and high‐dose chemotherapy. Here, we provide a detailed evaluation of PBSC collection upon vincristine, ifosfamide, doxorubicin, and etoposide (VIDE) chemomobilization.

Giant Cell Arteritis Related to Granulocyte-Colony-Stimulating FactorAdministration

Objective: Granulocyte-colony-stimulating factor (G-CSF) is routinely used to mobilize stem cells for peripheral blood stem cell (PBSC) collection by leukapheresis. Although generally considered safe and effective, G-CSF has been reported to cause severe side-effects in rare cases. Methods and Results: We report a case of a 65-year-old woman with diffuse large B-cell lymphoma, who received G-CSF for PBSC mobilization for ten days and developed fever of unknown origin. She was diagnosed with giant cell arteritis (GCA) related to G-CSF with aortic involvement based on typical findings obtained by contrastenhanced computed tomography and treated with high-dose prednisone. Conclusion: GCA might have to be considered as a rare but severe side effect of G-CSF administration. Imaging studies may help to identify large vessel vasculitis in cases that cannot be confirmed by tissue biopsy.

In MALDI–Mass Spectrometry Imaging on Formalin‐Fixed Paraffin‐Embedded Tissue Specimen Section Thickness Significantly Influences m/z Peak Intensity

In matrix‐assisted laser desorption/ionization–mass spectrometry imaging (MALDI–MSI) standardized sample preparation is important to obtain reliable results. Herein, the impact of section thickness in formalin‐fixed paraffin embedded (FFPE) tissue microarrays (TMA) on spectral intensities is investigated.

NKT cells - New players in CAR cell immunotherapy?

Low levels of peripheral blood natural killer T (NKT) cells in cancer patients and a favorable outcome associated with a high number of tumor‐infiltrating NKT cells demonstrated in several studies indicated the important role of these immune cells in the antitumor response. With effective antitumor immunity via direct tumor lysis, cytokine modulation of effector cells and regulation of immunosuppressive cells, type I NKT cells display interesting features/properties for the rapidly developing chimeric antigen receptor (CAR) technology. Due to their restriction to the monomorphic HLA‐like molecule CD1d, but not to the polymorphic human leukocyte antigen (HLA), NKT CAR cells show potential for enabling autologous and allogeneic/off‐the‐shelf cancer immunotherapy. Promising results were obtained in preclinical NKT CAR cell studies, but clinical trials have not yet been conducted. In this review, we summarize the biological features of NKT cells, their role in antitumor immunity and recent advances in the development of NKT CAR cells.

Selective contrast-enhanced computed tomography is appropriate in diffuse large B-cell lymphoma therapy response assessment

Although not the gold standard, contrast‐enhanced CT of neck, thorax, and abdomen/pelvis is routinely performed in diagnosis and response assessment of DLBCL. PD during first‐line treatment is a relatively rare event. The question arises if the imaging of initially involved regions only might be sufficient for response evaluation.

HCV load as a possible prognostic factor in patients with HCV-related DLBCL

Dear Editor, Several studies have suggested a potential causal link between HCV infection and diffuse large B cell lymphoma (DLBCL) [1, 2]. Currently, the optimal management of HCV-related DLBCL and the role of antiviral therapy (AVT) are still under debate [3], and factors that might predict the outcome of HCV-related DLBCL have only been described in few published studies [4–7]. The aims of our study were to analyze the clinical characteristics and outcome of patients who were diagnosed with both HCV and DLBCL and to identify possible factors associated with an adverse outcome, in particular focusing on HCV load. This is a retrospective single-center analysis of patients with DLBCL co-diagnosed with a HCV infection. We identified and retrieved the data of those patients who were treated or followed as inor outpatient from January 2000 to May 2014 at the University Hospital Heidelberg. The following data were collected: patients’ demographic characteristics, DLBCL-related clinical data (date of diagnosis, stage [8, 9], ECOG performance status [10], first-line therapy, response to therapy [11], time point of relapse), HCV infection-related data (date of diagnosis, genotype, viral load at different time points), and date of death or last follow-up at our hospital. Data were carefully evaluated in particular with regard to the chronology of HCV and DLBCL diagnoses, HCV load at different time points of DLBCL therapy, and overall survival (OS) in dependency of the viral load. Descriptive statistics was performed with Microsoft Excel 2013. OS was calculated and plotted using the Kaplan-Meier survival method (GraphPad Prism 5.0). To compare the survival rate between the groups, univariate log-rank tests were used. A P value less than 0.05 was considered to be statistically significant. All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the Helsinki Declaration of 1975, as revised in 2008 [5]. Fifteen patients with a diagnosis of DLBCL and HCV infection were identified. Three patients (20%) had secondary DLBCL and FL was the primary lymphoma diagnosis. The median age at DLBCL diagnosis was 47 (range 24–78) years. The majority of patients (n = 10, 67%) had advanced stage disease. Thirteen patients (87%) received systemic chemotherapy, and a CHOP-like regimen was administered in ten patients (67%). One patient was diagnosed with DLBCL post-transplant lymphoproliferative disorder (PTLD) in a transplant kidney (no. 9, Table 1). This patient received local therapy. Another patient died before the administration of chemotherapy due to progression of DLBCL (no. 10). No relapse or refractory disease was observed in 11 patients (74%) after first-line therapy (Table 2). Two thirds of patients (n = 10, 67%) were diagnosed with HCV infection prior to and one patient (no. 15) post DLBCL diagnosis. In three cases, HCV infection and DLBCL were diagnosed simultaneously (Table 2). In * Xiang Zhou zhou8285@126.com

In Burkitt lymphoma patients who relapse after induction with a short-intensive chemoimmunotherapy protocol, aggressive salvage chemotherapy therapy is ineffective: a single-center retrospective study

Dear Editor, Burkitt lymphoma (BL) is a highly aggressive non-hodgkin lymphoma (NHL) representing about 1% of mature NHLs [1]. The German Multicenter Study Group for Adult ALL conducted the multicenter GMALL-B-ALL/NHL2002 trial, which used a short-intensive protocol based on rituximab, high-dose methotrexate, and triple intrathecal therapy, with a reduced regimen for patients >55 years old. This trial reported an 88% complete response (CR) rate, and 5-year overall survival (OS) and progression-free survival (PFS) rates of 80% and 75%, respectively [2]. Thus, treatment according to this protocol, which has become the standard of care for adult BL patients at our institution, is curative in the majority of patients. However, almost identical rates for OS and PFS reported from recent trials [2–5] indicate that there is a lack of effective salvage therapies, and there is an extreme paucity of data regarding treatment options for patients who relapse with BL after induction with modern, short-intensive induction regimens. As a result, it seems questionable whether it is advisable to employ aggressive salvage chemotherapy for relapsed/ refractory BL. Therefore, we retrospectively analyzed the electronic records of all patients (n = 38) who were newly diagnosed with Burkitt lymphoma, Burkitt-like lymphoma, B-cell lymphoma unclassifiable with features intermediate between DLBCL, and Burkitt lymphoma (BCLU) or Burkitt leukemia between 2003 and 2013 in our institution and were treated with curative intent. Patients were enrolled in the GMALL-B-ALL/NHL2002 trial if eligible until 2011; afterwards, we continued to treat patients according to the trial protocol. We then looked in detail at patients with relapsed/ refractory disease (n = 9, 24%). There was (and is to date) no standard treatment for relapsed/refractory BL at our institution, so that a variety of curative or palliative approaches were used, based on individual treatment decisions (Fig. 1). Five patients received intensive salvage chemotherapy (with a curative intent), only two of which achieved a second remission: one CR and one PR, the latter of which lasted only a few weeks. The (only) patient who achieved a second CR proceeded to receive an allogeneic bone marrow transplantation from an HLA-compatible unrelated donor after conditioning therapy with busulfan/fludarabine/cyclophosphamide/ATG in 2007. Post-transplant, she suffered from gastrointestinal graft-versus-host disease and unfortunately died from infectious complications (pneumonia) 7 months after receiving the allogeneic bone marrow graft (recorded as death in CR). In general, the role of hematopoietic stem cell or bone marrow transplant, whether autologous or allogeneic, is difficult to assess in BL since most studies published to date present retrospective data * Martin Cremer martin.cremer@med.uni-heidelberg.de