Human cytokine-induced killer cells have enhanced in vitro cytolytic activity via non-viral interleukin-2 gene transfer

Modulation of the immune system by genetically modified immunological effector cells is of potential therapeutic value in the treatment of malignancies. Interleukin-2 (IL-2) is a crucial cytokine which induces potent antitumor response. Cytokine-induced killer cells (CIK) have been described as highly efficient cytotoxic effector cells capable of lysing tumor cell targets and are capable of recognizing these cells in a non-MHC restricted fashion. Dendritic cells (DC) are the major antigen presenting cells. This study evaluated the antitumor effect of CIK cells which were non-virally transfected with IL-2 and co-cultured with pulsed and unpulsed DC. Human CIK cells generated from peripheral blood were transfected in vitro with plasmid encoding for the human IL-2. Transfection involved a combination of electrical parameters and a specific solution to deliver plasmid directly to the cell nucleus by using the Nucleofector(® )electroporation system. Nucleofection resulted in the production of IL-2 with a mean of 478.5 pg/10(6 )cells (range of 107.6–1079.3 pg /10(6 )cells/24 h) compared to mock transfected CIK cells (31 pg/10(6 )cells) (P = 0.05). After co-culturing with DC their functional ability was assessed in vitro by a cytotoxicity assay. On comparison with non-transfected CIK cells co-cultured with DCs (36.5 ± 5.3 %), transfected CIK cells co-cultured with DC had a significantly higher lytic activity of 58.5 ± 3.2% (P = 0.03) against Dan G cells, a human pancreatic carcinoma cell line

Effects of recombinant adenovirus-mediated expression of IL-2 and IL-12 in human B lymphoma cells on co-cultured PBMC

BACKGROUND: Modulation of the immune system by genetically modified lymphoma cell vaccines is of potential therapeutic value in the treatment of B cell lymphoma. However, the anti-tumor effect of any single immunogene transfer has so far been limited. Combination treatment of recombinant IL-2 and IL-12 has been reported to be synergistic for inducing anti-tumor responses in solid tumors but the potential of IL-2/IL-12 gene modified B cell lymphoma cells has not been explored yet. METHODS: Using three different human B cell lymphoma cell lines and primary samples from patients with B cell neoplasms, expression levels of the coxsackie B-adenovirus receptor (CAR) and alpha (v) integrins were analyzed by fluorescence-activated cell sorter (FACS). Adenoviral transduction efficiencies were determined by GFP expression analysis and IL-2 and IL-12 cytokine production was quantified by enzyme-linked immunosorbent (ELISA) assays. Proliferative activities of peripheral blood mononuclear cells (PBMC) stimulated with either cytokine derived from supernatants of transduced lymphoma cells were measured by cell proliferation (MTT) assays. An EuTDA cytotoxicity assay was used to compare cytotoxic activities of IL-2 and/or IL-12 stimulated PBMC against unmodified lymphoma cells. RESULTS: We found that B cell lymphoma cell lines could be transduced with much higher efficiency than primary tumor samples, which appeared to correlate with the expression of CAR. Adenoviral-expressed IL-2 and IL-12 similarly led to dose-dependent increases in proliferation rates of PBMC obtained from healthy donors. IL-2 and/or IL-12 transduced lymphoma cells were co-cultured with PBMC, which were assayed for their cytolytic activity against unmodified lymphoma cells. We found that IL-2 stimulated PBMC elicited a significant anti-tumor effect but not the combined effect of IL-2/IL-12 or IL-12 alone. CONCLUSION: This study demonstrates that the generation of recombinant adenovirus modified lymphoma cell vaccines based on lymphoma cell lines expressing IL-2 and IL-12 cytokine genes is technically feasible, induces increases in proliferation rates and cytotoxic activity of co-cultured PBMC, and warrants further development for the treatment of lymphoma patients in the future

High-dose chemotherapy with autologous peripheral blood stem cell support for recurrent primary AFP-producing intracranial germinoma

We report of a 34-year old man with second intracranial relapse of a suprasellar germinoma. Despite of extensive pretreatment with radiation and conventional chemotherapy relapse occurred and was treated with sequential high-dose chemotherapy followed by transfusion of autologous peripheral stem cells. The high-dose chemotherapy course was complicated by refractory derailment of pineal gland insufficiency. The patient achieved a complete remission after high dose chemotherapy which lasted for 13 months. Subsequently, he developed a third relapse and died

Invasive fungal infections in neutropenic enterocolitis: A systematic analysis of pathogens, incidence, treatment and mortality in adult patients

BACKGROUND: Neutropenic enterocolitis is a life-threatening complication most frequently occurring after intensive chemotherapy in acute leukaemias. Gramnegative bacteria constitute the most important group of causative pathogens. Fungi have also been reported, but their practical relevance remains unclear. The guidelines do not address concrete treatment recommendations for fungal neutropenic enterocolitis. METHODS: Here, we conducted a metaanalysis to answer the questions: What are frequency and mortality of fungal neutropenic enterocolitis? Do frequencies and microbiological distribution of causative fungi support empirical antimycotic therapy? Do reported results of antimycotic therapy in documented fungal neutropenic enterocolitis help with the selection of appropriate drugs? Following a systematic search, we extracted and summarised all detail data from the complete literature. RESULTS: Among 186 articles describing patients with neutropenic enterocolitis, we found 29 reports describing 53 patients with causative fungal pathogens. We found no randomised controlled trial, no good quality cohort study and no good quality case control study on the role of antifungal treatment. The pooled frequency of fungal neutropenic enterocolitis was 6.2% calculated from all 860 reported patients and 3.4% calculated from selected representative studies only. In 94% of the patients, Candida spp. were involved. The pooled mortality rate was 81.8%. Most authors did not report or perform antifungal therapy. CONCLUSION: In patients with neutropenic enterocolitis, fungal pathogens play a relevant, but secondary role compared to bacteria. Evidence concerning therapy is very poor, but epidemiological data from this study may provide helpful clues to select empiric antifungal therapy in neutropenic enterocolitis

Primary small cell carcinoma of the esophagus: patient data metaanalysis and review of the literature [Primäres kleinzelliges Ösophaguskarzinom: Patientendaten-Metaanalyse und Review der Literatur ]

[english] We analysed the typical features of primary small cell carcinoma of the esophagus (SCCE) with emphasis on occurrence, behaviour, outcome and treatment options. This metaanalysis was aimed at collecting and analyzing information from international studies about handling this disease. This seems necessary due to the rarity of this disease. Studies were acquired from electronic databases and reference lists. We finally analysed 313 patient cases from the literature with oesophageal SCC. A data extraction was accomplished referring to 13 evaluable features that are described in the “methods”, whereof 7 were analyzed with univariate and multivariate tests. Three hundred thirteen cases were analyzed, 109 patients (35%) had limited stage (LS), whereas 167 (54%) had extensive stage (ES). There is no information about the remaining 35 patients concerning the stage. Univariate and multivariate analysis showed only age (<50 years vs. >50 years, HR 1.024; 95% CI 1.000–1.041, <0.0001) and disease stage (LS vs. ES, HR 4.884; 95% CI 2.572–9.27, <0.0001) as significant prognostic factors. There also was a statistically significant difference in survival between those patients who received therapy compared to those who only received best supportive care (11.6 months vs. 0.8 months, HR 0.093, CI 95% 0.053–0.16, <0.001). In this first multivariate analysis for SCCE we show that SCCE is an aggressive type of tumour with a shorter survival rate compared to its counterpart from the lung. It is demonstrated that only disease stage (limited vs. extensive stage), age (<50 years vs. >50 years) and therapy are independent significant predictors of prognosis.<br>[german] Gegenstand unserer Untersuchungen war die Erhebung typischer Eigenschaften des kleinzelligen Ösophaguskarzinoms mit Berücksichtigung des Auftretens, des Verlaufs, der Auswirkung und der Behandlungsoptionen. Ziel unserer Metaanalyse bestand darin, Informationen aus internationalen Studien, die sich mit dieser Krankheit befassten, zu sammeln und auszuwerten. Dies scheint notwendig aufgrund der Seltenheit der Krankheit. Die verwendeten Studien wurden von elektronischen Datenbanken und Referenzlisten ermittelt. Insgesamt wurden 313 Patientenfälle mit einem kleinzelligen Ösophaguskarzinom aus der Literatur ausgewertet. Die erhobenen Daten beziehen sich auf 13 auswertbare Kriterien, die in den „Methoden“ beschrieben sind, wovon sieben Kriterien mit univariaten und multivariaten Tests untersucht worden sind. Von 313 analysierten Patientenfällen waren 109 (35%) Patienten im limitierten Stadium der Erkrankung und 167 (54%) im ausgedehnten Stadium. Über die übrigen 35 Patienten sind keine Informationen bezüglich des Stadiums bekannt. Univariate und multivariate Analysen zeigten nur das Alter (<50 Jahre vs. >50 Jahre, HR 1.024; 95% CI 1.000–1.041, <0.0001) und das Krankheitsstadium (LS vs. ES, HR 4.884; 95% CI 2.572–9.27, <0.0001) als signifikante prognostische Faktoren. Ebenfalls gab es einen statistisch signifikanten Unterschied in der Überlebenschance zwischen jenen Patienten, die therapiert worden sind, und jenen, die außer bestmöglicher unterstützender Fürsorge nicht therapiert worden sind (11,6 Monate vs. 0,8 Monate, HR 0.093, CI 95% 0.053–0.16, <0.001). In dieser ersten multivariaten Analyse für das kleinzellige Ösophaguskarzinom zeigen wir, dass diese Form des Krebses eine sehr aggressive Form ist, die mit einer kürzeren Überlebensrate einhergeht verglichen mit seinem Pendant, dem kleinzelligen Bronchialkarzinom. Es wurde gezeigt, dass nur das Krankheitsstadium (limitiert vs. ausgedehnt), das Alter (<50 Jahre vs. >50 Jahre) und die Therapie als Kriterien eigenständige, signifikante Anzeichen für eine Prognose sind