In vitro cytotoxic dose-relation of cisplatin and sodium thiosulphate in human tongue and oesophageal squamous carcinoma cell lines.

BACKGROUND: Intraarterial chemotherapy of oral and oropharyngeal cancer with cisplatin (cis-diamminedichloroplatinum [II]) has experienced a revival in the last decade. Side-effects of the therapy were very low with concomitant systemic infusion of the neutralizing agent sodium thiosulphate. The requisite dose of the chemotherapeutic agent which safely leads to apoptosis of oral cancer cells has not yet been assessed in vitro, nor has the combination of cisplatin and sodium thiosulphate been examined for the potential reduction of cytotoxicity in oral cancer cells. STUDY DESIGN: In a panel of two tongue squamous cancer cell lines and an oesophageal cancer cell line as control and comparison, cisplatin (0.2-10 microgram/ml) was combined with sodium thiosulphate (0-0.5 mg/ml). RESULTS: 10 microgram/ml of cisplatin proved to be 100% antiproliferative, while any additional concentration of sodium thiosulphate decreased this effect. At the maximum dose of cisplatin, a sodium thiosulphate/cisplatin concentration relation of less than 6:1 still effected cytotoxic activity of >80%. An increase of cisplatin concentration led to higher cytotoxicity irrespective of sodium thiosulphate concentration. The oesophageal cell line was more sensitive to cisplatin and to sodium thiosulphate than the tongue cell lines. CONCLUSIONS: In this study, it was found that high concentrations of cisplatin are necessary in oral cancer to reach cytotoxic levels which support high-dose intraarterial chemotherapy by which these levels might be reached. A sodium thiosulphate/cisplatin concentration ratio within the tumour of less than 6:1 may be allowed without compromising the cytotoxic activity of cisplatin.     

Simple and efficient method for the detection of diethylenetriaminepentaacetic acid

Diethylenetriaminepentaacetic acid (DTPA) is a commonly used chelating agent. Its antiviral, antibacterial and immunomodulatory effects are well documented. DTPA forms a highly stable complex with lead (II) with an increased absorption coefficient and a bathochromic shift of the absorption maximum compared to pure DTPA. Based on this complex a high-performance liquid chromatographic method for the quantitative detection of DTPA in biological fluids was developed. A calibration curve was prepared and linearity was shown in the concentration range between 10 mg l(-1) and 1000 mg l(-1) DTPA. The recovery in water and in human plasma showed the method to be suitable for routine use.     

Co-expression of different types of Fc receptors on murine peritoneal macrophages

Simultaneous expression of particular immunoglobulin Fc receptors (FcR) was studied on the plasma membranes of murine peritoneal macrophages. This was facilitated by the use of sheep red blood cells (SRBC) and/or synthetic microspheres coated with monoclonal antibodies of different isotypes. It was concluded that a majority of macrophages bear more than one type of FcR; macrophages bearing at least three types of FcR were present in the peritoneal cavity; macrophages bearing Fc mu R did not bind IgE, IgA or IgG; all macrophages bearing Fc alpha R also expressed Fc gamma 2bR, Fc gamma 3R and Fc epsilon R; all macrophages bearing Fc epsilon R also expressed Fc gamma 2bR and Fc alpha R. Except for Fc alpha R, essentially equivalent numbers of FcR-bearing macrophages were detected when antibody-coated SRBC or polymeric microspheres were used. Simultaneous applications of these reagents permitted the most detailed and direct investigations yet performed of multiple FcR expression on individual cells.     

Differential effects of intravenous anesthetics on hepatosplanchnic microcirculation in rats: sidestream dark-field imaging study

This study aimed to investigate the effects of intravenous anesthetics on hepatosplanchnic microcirculation in laparotomized mechanically ventilated rats using Sidestream Dark-field (SDF) imaging. Thirty male Wistar rats were divided into 5 groups (n = 6 each). All rats were initially anesthetized with 60 mg/kg pentobarbital (i.p.) for instrumentation. This was followed by either ketamine, propofol, thiopental, midazolam or saline+fentanyl (iv bolus over 5 min and then maintenance over 90 min). SDF imaging of the liver and distal ileum microcirculation was performed at the baseline and at t = 5, 35, 65 and 95 min. In propofol group there was increase of functional sinusoidal density (FSD) following induction (+25%, P < 0.05) and maintenance at t = 95 min (+10.3%, P < 0.05), in ketamine and midazolam group decrease of FSD was observed after induction (-20.4%, P < 0.05; -10.1%, P < 0.05) and during maintenance at t = 65 min (-11.6%, P < 0.05; -11.4%, P < 0.05) when compared to baseline. Following induction with propofol functional capillary density (FCD) of ileal longitudinal muscle layer increased (+10.6%, P < 0.05) and returned to baseline values during maintenance. Ketamine and midazolam decreased FCD of longitudinal layer after induction (-24.6%, P < 0.05; -21.1%, P < 0.05) and remained decreased during maintenance at t = 95 min (-10.8%, P < 0.05; -15.5%, P < 0.05). In thiopental and control group, changes in microcirculatory parameters were not significant throughout the study. In conclusion, intravenous anesthetics affect the hepatosplanchnic microcirculation differentially, propofol has shown protective effect on the liver and intestinal microcirculation.     

Exosomal vaccines containing the S protein of the SARS coronavirus induce high levels of neutralizing antibodies

Infection with the SARS-associated coronavirus (SARS-CoV) induces an atypical pulmonary disease with a high lethality rate. Although the initial SARS epidemic was contained, sporadic outbreaks of the disease still occur, suggesting a continuous need for a vaccine against this virus. We therefore explored exosome-based vaccines containing the spike S proteins of SARS-CoV. S-containing exosomes were obtained by replacing the transmembrane and cytoplasmic domains of the S protein by those of VSV-G. The immunogenicity and efficacy of the S-containing exosomes were tested in mice and compared to an adenoviral vector vaccine expressing the S protein. Both, S-containing exosomes and the adenoviral vector vaccine induced neutralizing antibody titers. After priming with the SARS-S exosomal vaccine and boosting with the adenoviral vector the neutralizing antibody titers exceeded those observed in the convalescent serum of a SARS patient. Both approaches were effective in a SARS-S-expressing tumor challenge model and thus warrant further investigation.     

Cisplatin-resistant neuroblastoma cells express enhanced levels of epidermal growth factor receptor (EGFR) and are sensitive to treatment with EGFR-specific toxins

PURPOSE: Neuroblastomas frequently show expression of the epidermal growth factor receptor (EGFR) and may therefore be susceptible to EGFR-targeted therapies. Here, EGFR expression and functionality was investigated in parental chemosensitive neuroblastoma cell lines (UKF-NB-3, IMR-32, NLF, SH-SY5Y) and their cisplatin-resistant sublines (UKF-NB-3(r)CDDP(1000), IMR-32(r)CDDP(1000), NLF(r)CDDP(1000), and SH-SY5Y(r)CDDP(500)). Moreover, the EGFR antibody cetuximab, the EGFR tyrosine kinase inhibitor Tyrphostin B46, and recombinant EGFR-targeted toxins were investigated for their influence on the viability and growth of neuroblastoma cells. EXPERIMENTAL DESIGN: EGFR expression and function was measured by flow cytometry or Western blot. Cell viability was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Apoptosis was examined by immunostaining for active caspase-3 or cleaved poly(ADP-ribose) polymerase. Cellular binding of FITC-labeled immunotoxins was studied by flow cytometry, and cellular uptake was studied by confocal laser scanning microscopy. RESULTS: The EGFR-targeted antibody and growth factor toxins scFv(14E1)- Pseudomonas exotoxin A (ETA) and TGF-alpha-ETA exerted anti-cancer effects in neuroblastoma cell lines that were insensitive to cetuximab or EGFR tyrosine kinase inhibitors. Furthermore, adaptation of chemosensitive neuroblastoma cells to cisplatin increased EGFR expression and sensitivity to both recombinant toxins. Treatment of chemosensitive neuroblastoma cells with cisplatin reversibly increased EGFR expression, whereas cisplatin-resistant cells showed enhanced EGFR expression independent of the presence of cisplatin. Combination treatment with scFv(14E1)-ETA or TGF-alpha-ETA and cisplatin exerted significantly improved anticancer effects compared with either single treatment in parental neuroblastoma cells, cisplatin-resistant sublines, and primary cultures. CONCLUSIONS: EGFR-targeted cytotoxic reagents such as scFv(14E1)-ETA and TGF-alpha-ETA represent promising candidates for further development as antineuroblastoma agents, especially in combination with cisplatin.     

Prospects for cationic polymers in gene and oligonucleotide therapy against cancer

Gene and antisense/ribozyme therapy possesses tremendous potential for the successful treatment of genetically based diseases, such as cancer. Several cancer gene therapy strategies have already been realized in vitro, as well as in vivo. A few have even reached the stage of clinical trials, most of them phase I, while some antisense strategies have advanced to phase II and III studies. Despite this progress, a major problem in exploiting the full potential of cancer gene therapy is the lack of a safe and efficient delivery system for nucleic acids. As viral vectors possess toxicity and immunogenicity, non-viral strategies are becoming more and more attractive. They demonstrate adequate safety profiles, but their rather low transfection efficiency remains a major drawback. This review will introduce the most important cationic polymers used as non-viral vectors for gene and oligonucleotide delivery and will summarize strategies for the targeting of these agents to cancer tissues. Since the low efficiency of this group of vectors can be attributed to specific systemic and subcellular obstacles, these hurdles, as well as strategies to circumvent them, will be discussed. Local delivery approaches of vector/DNA complexes will be summarized and an overview of the principles of anticancer gene and antisense/ribozyme therapy as well as an outline of ongoing clinical trials will be presented.