Biochemical and biological activity of the anthracycline analog, 4-demethyl-6-0-methyl-doxorubicin

Summary The chromophore-modified derivative of doxorubicin, 4-demethyl-6-0-methyl-doxorubicin, has been tested for antitumor activity in a range of experimental murine tumor systems. In contrast to the inactive 6-0-methyl derivative of daunorubicin, 4-demethyl-6-0-methyl-doxorubicin provided antitumor effects comparable to that of the parent compound. In addition, detailed DNA-interaction studies showed that the doxorubicin derivative retains the ability to bind DNA by the intercalation mechanism. However, the binding affinity was appreciably reduced following structural modification in the anthraquinone chromophore. On the basis of the proposed models of intercalation, these results could be rationalized in terms of steric influence of the bulky methoxy group. The results of this study are in agreement with the correlation already observed between DNA binding and relative antitumor activity of anthracyclines.     

Characteristics of Acute Asthma Patients Attended Frequently Compared with Those Attended Only Occasionally in an Emergency Department

Asthma patients that depend on emergency department (ED) services are generally considered to have extremely poor disease control and prognosis. It is important to identify characteristics related to poor disease control and frequent visits to the ED to apply appropriate clinical management. This study comprised a cross-sectional survey of consecutive patients with asthma exacerbation (age ≥12 years) presenting at the adult ED of a large, tertiary care, university-affiliated hospital over a 2-month period. The frequent visitors (FV) were defined by ≥3 visits to the ED in the preceding year, and the occasional visitors (OV) by ≤2 visits. Eighty-six patients (61 females and 25 males) were included in the study (mean age 38 ± 18 years). Of these patients, 51.2% were FV and 48.8% were OV. Sixty-nine percent had annual income lower than A$3000 and 66.3% had ≤8 years of the formal education. Only 18.6% had used inhaled corticosteroids, 79.1% identified the asthma attack severity, 70.9% increased or initiated inhaled β-agonist, 20.9% increased or initiated steroid therapy, and 55.8% had an asthma action plan for attack. The number of hospital admissions in past year (OR 4.3, P = .02), use of home nebulizer (OR 3.6, P = .05) and the lack of a written asthma action plan (OR 3.3, P = .03) were independently associated with frequent visits to the ED. We conclude that a substantial proportion of the patients that visit the ED are FV. These patients are more likely to have hospital admission in the past year, to use a home nebulizer, and to lack a written asthma action plan. They should be considered the most important target for asthma education.     

Sequence dependence of the antitumor and toxic effects of 5-fluorouracil and cis-diamminedichloroplatinum combination on primary colon tumors in mice

Summary Primary colon tumors of different sizes and malignancy, chemically induced by methylazoxymethanol in outbred CF-1 mice, were used to investigate the antitumor effects of 5-Fluorouracil (5FU) and cis-diammine-dichloroplatinum (DDP), given weekly i.v. as single agents or in combination. When single-drug chemotherapy was tested, DDP showed higher efficacy than 5FU. In fact, in two separate experiments a significant reduction (P(0.05) of tumor number (TN) and tumor burden was obtained by treatment with the optimal dose of DDP (4.5 mg/kg per injection) and not by that of 5FU (52 mg/kg). When the two drugs were combined (24-h interval), studies carried out on healthy mice treated weekly i.v. showed a lower toxicity with the same doses given in the sequence 5FU-DDP than in the opposite sequence. The two drugs, delivered in the sequence 5FU followed by DDP, statistically reduced the TN and total tumor burden compared to control mice (P(0.05). On the other hand, the same doses in the sequence DDP followed by 5FU did not attain significant tumor reduction. The sequence dependence of the activity and toxicity of the 5FU and DDP combination observed in this experimental model should be taken into account in the design of clinical trials.Abbreviations used MAM methylazoxymethanol acetate - 5FU 5Fluorouracil - DDP cis-diamminedichloroplatinum - TN tumor number - TTB total tumor burden - Ara-C cytosine-arabinoside This work was partially supported by Grant N. 84.00746.44 of Finalized Project Oncology of CNR (Rome, Italy)     

Chromosomal assignment of human O6-methylguanine-DNA-methyltransferase gene by hamster-human somatic cell hybrids.

Using an in vitro assay to measure O6-methylguanine-DNA-methyltransferase (MT) activity in cell extracts from a panel of human-hamster cell hybrids, we were able to locate the human MT gene on chromosome 10. Chinese hamster cells have little or no MT activity and the presence of human chromosome 10 was a necessary condition for MT activity in cell hybrids. In some cell hybrids carrying chromosome 10, however, MT activity was not higher than that of hamster cells. As an explanation for this result, genetic determinants repressing MT expression and/or activity might be present in other human chromosomes carried by MT-negative cell hybrids. Partial hyperploidy of the hamster karyotype, variable activity of the parental human cell lines and changes during subculturing of the cell hybrids might also account for the lack of enzymatic activity in chromosome 10 containing hybrids.     

Electronic implementation of guidelines in the EsPeR system: a knowledge specification method

Despite initiatives to standardize methods for the development of clinical guidelines, several barriers hinder their integration in daily clinical practice: failure to fulfil quality criteria, poor effectiveness of their dissemination. Computerization of guidelines can favor their dissemination. The initial step of computerization is the knowledge specification from the text of the guideline. We describe the method of knowledge specification, which is used in EsPeR (Personalized Estimate of Risks), a web-based decision support system in preventive medicine, which allows, for a given person, to estimate risks and access recommendations, based on clinical profile. This method is based on a structured and systematic analysis of text allowing detailed specification of a decision tree. We use decision tables to validate the decision algorithm and decision trees to specify this algorithm, along with elementary messages of recommendation. Editing tools are used to facilitate the process of validation and the workflow between expert physicians and computer scientists. Applied to eleven different guidelines, the method allows a quick and valid computerization and integration in the EsPeR system. The method used for computerization could help to define a framework usable at the initial step of guideline development in order to produce guidelines ready for electronic implementation.     

Hydrogen peroxide: effects on DNA, chromosomes, cell cycle and apoptosis induction in Fanconi's anemia cell lines

Fanconi's anemia (FA) is an inherited autosomal recessive syndrome; cells from FA patients are very sensitive to crosslinking agents and to oxygen. Epstein-Barr virus (EBV)-transformed lymphoblasts belonging to different FA complementation groups and normal EBV-transformed lymphoblasts were studied for their response to treatment with the oxidizing agent hydrogen peroxide (H2O2). The analysis of 8-hydroxy-2'-deoxyguanosine (8-OHdG) content in the DNA of untreated cells showed an increased basal level of damage in cells from the complementation groups FA-C and FA-E. H2O2-induced 8-OHdG was higher in FA than in normal cell lines. The removal of 8-OHdG after H2O2 treatment was significantly reduced in the cells from complementation group E. However, all FA cell lines showed a normal ability in the resealing of DNA breaks, at least soon after treatment. All cell lines were also equally efficient in the removal of damaged pyrimidines. Compared with normal cells, FA cell lines showed an increase in the baseline level of micronuclei, but not in the number of micronuclei induced by H2O2. Micronuclei in FA cells originated prevalently from chromosomal fragmentation and, at a minor extent, from chromosome loss. After H2O2 treatment, FA cell lines accumulated in G(2) phase to a greater extent than normal lymphoblasts. However, reversion of mutation in FA-A and FA-C cells did not result in the correction of this phenotype. In cells evaluated for apoptosis no ladder formation was found in FA-C, FA-E and corrected FA-C cells. In conclusion, among the FA cell lines examined, only FA-E showed a defect in the repair of H2O2-induced damage. On the other hand, differences found in the cell cycle and apoptosis might be due to irreversible changes occurring in FA cell lines as a consequence of the primary defect.     

Economical, simple method for production of the gaseous environment required for cultivation of Campylobacter jejuni.

Campylobacter jejuni is an enteric pathogen recognized worldwide as a cause of diarrhea. Its isolation from stool samples requires a microaerophilic environment that heretofore has been expensive and cumbersome to create. An economical, portable, and simple method is described which involves the production of appropriate concentrations of oxygen and carbon dioxide. Inside a plastic bag are placed two cups, one containing fine steel wool (grade 0) previously soaked in a 2.5% aqueous solution of copper sulfate and the other containing an Alka-Seltzer tablet in tap water. As suggested by Jurgensen et al. (Rev. Bras. Pat. Clin. 18:58-63, 1982), we used the effervescent antacid to generate CO2. By plate counts, we found this method to be as reliable in the cultivation of 20 isolates of C. jejuni in pure and mixed fecal culture as the reference gas method (85% N2, 10% CO2, and 5% O2). Analyses of the gas mixture inside the bag after up to 24 h of incubation confirmed the creation of an atmosphere of reduced O2 and increased CO2 concentrations. This method is eminently suitable for field situations in which more costly supplies are not available.     

Requirement of HMGB1 and RAGE for the maturation of human plasmacytoid dendritic cells

Dendritic cells (DC) are key components of innate and adaptive immune responses. Plasmacytoid DC (PDC) are a specialized DC subset that produce high amounts of type I interferons in response to microbes. High mobility group box 1 protein (HMGB1) is an abundant nuclear protein, which acts as a potent pro-inflammatory factor when released extracellularly. We show that HMGB1 leaves the nucleus of maturing PDC following TLR9 activation, and that PDC express on the plasma membrane the best-characterized receptor for HMGB1, RAGE. Maturation and type I IFN secretion of PDC is hindered when the HMGB1/RAGE pathway is disrupted. These results reveal HMGB1 and RAGE as the first known autocrine loop modulating the maturation of PDC, and suggest that antagonists of HMGB1/RAGE might have therapeutic potential for the treatment of systemic human diseases.