Synthesis of antitumor-active conjugates of adriamycin or daunomycin with the copolymer of divinyl ether and maleic anhydride

Polymeric conjugates of adriamycin (ADR) ( 2 ) or daunomycin (DM) ( 3 ) were synthesized by reaction of the drugs with the copolymer of divinyl ether and maleic anyhdride (DIVEMA) ( 1 ). The content of ADR moieties in the DIVEMA conjugate ( 4 ) could be varied depending on the reaction conditions up to 35,8 wt.-%. Considering the low toxicity and the high possibility of renal excretion, DIVEMA with M?_w of 7000 and M?_w/M?_n = 1,6 was used for the conjugation. The rate of drug release from the conjugate was determined under physiological conditions by reversed phase HPLC. Within 14 days only 15% of the attached ADR was released from conjugate 4 . The antitumor activity of the conjugates was tested in vitro and in vivo against mouse P388 leukemia. Conjugate 4 proved to be 28 times less active than ADR in vitro, which could be explained from the slow drug-release. On the contrary 50% of the leukemic mice treated by 4 survived more than 60 days, whereas no mice given ADR alone or the admixture of ADR and DIVEMA survived 30 days. An antitumor activity of the polymeric conjugate better than that of the free drug was also observed in vivo with DM. Such a polymeric effect can be attributed either to the change in body distribution, the difference in pharmacokinetics, or the slow drugrelease.     

Acute hepatic failure in IFN-gamma-deficient BALB/c mice after murine coronavirus infection

We previously showed that an intraperitoneal infection with mouse hepatitis virus (MHV) persists in interferon-gamma (IFN-gamma)-deficient C57BL/6 (B6-GKO) mice and results in subacute fatal peritonitis, which bears a resemblance to feline infectious peritonitis. To examine the role of other host factors in MHV infection in mice, IFN-gamma-deficient mice with a BALB/c background (BALB-GKO) were infected intraperitoneally with MHV and compared with B6-GKO mice. In contrast to B6-GKO mice, BALB-GKO mice died within 1 week due to acute hepatic failure. The viral titer of the liver in BALB-GKO mice was significantly higher than that in B6-GKO mice. All hepatocytes in BALB-GKO mice were necrotic at 5 days post-infection, which was clearly distinct from large but limited lesion in the liver from infected B6-GKO mice. The serum alanine aminotransferase activity of infected BALB-GKO mice were higher than that of B6-GKO mice and was paralleled with the severity of the pathological changes and viral titers in infected mice. Administration of exogenous IFN-gamma to BALB-GKO partially inhibited the acute death. These results indicate that BALB-GKO and B6-GKO mice clearly show different diseases following MHV infection, although wild type counterparts of both mice apparently showed the same clinical course after MHV infection.     

Isolation, characterization, and disruption of dnr1, the areA/nit-2-like nitrogen regulatory gene of the zoophilic dermatophyte, Microsporum canis.

A homolog of the major nitrogen regulatory genes areA from Aspergillus nidulans and nit-2 from Neurospora crassa was isolated from the zoophilic dermatophyte, Microsporum canis. This gene, dnr1, encodes a polypeptide of 761 amino acid residues containing a single zinc-finger DNA-binding domain, which is almost identical in amino acid sequence to the zinc-finger domains of AREA and NIT-2. The functional equivalence of dnr1 to areA was demonstrated by complementation of an areA loss-of-function mutant of A. nidulans with dnr1 cDNA. To further characterize this gene, dnr1 was disrupted by gene replacement based on homologous recombination. Of 100 transformants analyzed, two showed the results expected for replacement of dnr1. The growth properties of the two dnr1(-) mutant strains on various nitrogen sources were examined. Unlike the A. nidulansareA(-) mutant, these dnr1(-) mutants showed significantly reduced growth on ammonia, a preferred nitrogen source for fungi. These mutant strains were also able to utilize various amino acids for growth. In comparison with wild-type M. canis, the two dnr1(-) mutants showed reduced growth on medium containing keratin as the sole nitrogen source. This is the first report describing successful production of targeted gene-disrupted mutants by homologous recombination and their phenotypic analysis in dermatophytes.     

N-Glycosylation contributes to the limited cross-reactivity between hemagglutinin neuraminidase proteins of human parainfluenza virus type 4A and 4B

cDNAs encoding human parainfluenza virus type 4B (hPIV-4B) hemagglutinin neuraminidase (HN) protein were cloned and the nucleotide sequences were determined. A high degree of identity (81.4%) was observed between the nucleotide sequences of hPIV-4A and -4B HN proteins, and an 87.3% identity was found between the deduced amino acid sequences. This degree of identity is considered to be greater than immunological similarity between hPIV-4A and -4B HN proteins determined using monoclonal antibodies. To elucidate the causes of the antigenic difference between HN proteins of hPIV-4A and -4B, we constructed three cDNAs of hPIV-4B HN whose potential N-glycosylation sites were partially or completely the same as in hPIV-4A HN cDNA. We compared the antigenicity of the expressed wild-type and mutant proteins, and found that the antigenicities of the mutant hPIV-4B HN proteins were more similar to the hPIV-4A HN protein than to the non-mutant hPIV-4B HN protein. This study indicated that the antigenic diversity between hPIV-4A and -4B was partly caused by deletion or creation of glycosylation sites, showing that the point mutations resulting in deletion or creation of glycosylation sites is one of the initial steps leading to the division of virus into subtypes. Received: 21 January 2000     

Turtle-chicken chimera: an experimental approach to understanding evolutionary innovation in the turtle.

Turtles have a body plan unique among vertebrates in that their ribs have shifted topographically to a superficial layer of the body and the trunk muscles are greatly reduced. Identifying the developmental factors that cause this pattern would further our understanding of the evolutionary origin of the turtles. As the first step in addressing this question, we replaced newly developed epithelial somites of the chicken at the thoracic level with those of the Chinese soft-shelled turtle Pelodiscus sinensis (P. sinensis somites into a chicken host) and observed the developmental patterning of the grafted somites in the chimera. The P. sinensis somites differentiated normally in the chicken embryonic environment into sclerotomes and dermomyotomes, and the myotomes differentiated further into the epaxial and hypaxial muscles with histological morphology similar to that of normal P. sinensis embryos and not to that of the chicken. Epaxial dermis also arose from the graft. Skeletal components, however, did not differentiate from the P. sinensis sclerotome, except for small fragments of cartilage associated with the host centrum and neural arches. We conclude that chicken and P. sinensis share the developmental programs necessary for the early differentiation of somites and that turtle-specific traits in muscle patterning arise mainly through a cell-autonomous developmental process in the somites per se. However, the mechanism for turtle-specific cartilage patterning, including that of the ribs, is not supported by the chicken embryonic environment.     

Tacrolimus reduces urinary excretion of leukotriene E(4) and inhibits aspirin-induced asthma to threshold dose of aspirin

BACKGROUND: The exact mechanism of aspirin-induced asthma is not clear. It has been postulated that precipitation of asthma attacks by aspirin is linked to inhibition of COX activity and massive release of cysteinyl leukotriene into the airway. Tacrolimus, a macrolide-derived immunosuppressant, is used for immunosuppression in organ transplantation and also for allergic diseases such as atopic dermatitis. OBJECTIVE: We evaluated the effects of tacrolimus in aspirin-induced asthma by using a double-blind, crossover study design. METHODS: Twelve patients with aspirin-induced asthma (male:female, 3:9; mean age +/- SD, 36.7 +/- 7.2 years) received either tacrolimus (0.1 mg/kg) or placebo 2 hours before the threshold dose of oral aspirin. RESULTS: In the placebo arm, oral aspirin significantly decreased FEV 1 concomitant with significant increases in sputum eosinophilic cationic protein and urinary leukotriene E(4) levels. Tacrolimus significantly inhibited bronchoconstriction and abrogated aspirin-induced increase in both sputum eosinophilic cationic protein and urinary leukotriene E(4) levels. CONCLUSION: The current study suggested that tacrolimus inhibited bronchoconstriction to a threshold dose of aspirin by inhibition of cysteinyl leukotriene excretion.     

In vitro and in vivo antifungal activities of FX0685, a novel triazole antifungal agent with potent activity against fluconazole-resistant Candida albicans.

To evaluate the therapeutic potential of FX0685, a new triazole antifungal agent, for the treatment of opportunistic fungal infections, particularly systemic candidiasis and aspergillosis, in vitro and in vivo studies were performed using fluconazole (FLC), itraconazole (ITC) and/or amphotericin B (AMB) as reference drugs. A preliminary in vitro study showed that the antifungal activity of FX0685 against FLC-susceptible Candida albicans, several non-C. albicans Candida species and Cryptococcus neoformans was superior to that of FLC and comparable or superior to those of ITC and AMB, while the anti-Aspergillus fumigatus activity of FX0685 was to varying degrees lower than that of ITC. FX0685 appeared to be comparable to FLC and ITC in the treatment of murine systemic C. albicans and pulmonary A. fumigatus infection, respectively. The biological property of FX0685 was characterized by its potent in vitro and in vivo activity against FLC-resistant C. albicans. Part of this unique property was explained by the finding that it retained potent inhibitory activity against those CYP51 molecules in which amino acid substitutions confer a phenotype of resistance to FLC and some other azole derivatives. All of these results lead to the possibility that FX0685 may be a potential antifungal drug candidate for the treatment of various clinical forms of systemic candidiasis, including those caused by FLC-resistant C. albicans, as well as for the treatment of pulmonary aspergillosis.     

Fermented grain products, production, properties and benefits to health

Fermented foods such as Japanese traditional food “miso (fermented soy bean paste)” have been shown to be rich source of micronutrients with the potential to prevent various human diseases. We have introduced effects of a new dietary supplement of fermented grain foods mixture containing extracts from wheat germ, soybeans, rice bran, tear grass, sesame, wheat, citrus lemon, green tea, green leaf extract and malted rice under the trade name of antioxidant biofactor (AOB). Chemical analysis of AOB shows the presence of various phenolic compounds (catechins, rutin, genistin, daidzin, etc.). AOB has strong antioxidant properties and additional biological effects, which might be of importance in context with the prevention of degenerative diseases. This paper focuses on the effect of supplementing AOB in various animal models and humans.