DNA-Directed alkylating agents. 7. Synthesis, DNA interaction, and antitumor activity of bis(hydroxymethyl)- and bis(carbamate)-substituted pyrrolizines and imidazoles

A series of bis(hydroxymethyl)-substituted imidazoles, thioimidazoles, and pyrrolizines and related bis(carbamates), linked to either 9-anilinoacridine (intercalating) or 4-(4-quinolinylamino)benzamide (minor groove binding) carriers, were synthesized and evaluated for sequence-specific DNA alkylation and cytotoxicity. The imidazole and thioimidazole analogues were prepared by initial synthesis of [(4-aminophenyl)alkyl]imidazole-, thioimidazole-, or pyrrolizine dicarboxylates, coupling of these with the desired carrier, and reduction to give the required bis(hydroxymethyl) alkylating moiety. The pyrrolizines were the most reactive alkylators, followed by the thioimidazoles, while the imidazoles were unreactive. The pyrrolizines and some of the thioimidazoles cross-linked DNA, as measured by agarose gel electrophoresis. Strand cleavage assays showed that none of the compounds reacted at purine N7 or N3 sites in the gpt region of the plasmid gpt2Eco, but the polymerase stop assay showed patterns of G-alkylation in C-rich regions. The corresponding thioimidazole bis(carbamates) were more selective than the bis(hydroxymethyl) pyrrolizines, with high-intensity bands at 5'-NCCN, 5'-NGCN and 5'-NCGN sequences in the PCR stopping assay ( indicates block sites). The data suggest that these targeted compounds, like the known thioimidazole bis(carbamate) carmethizole, alkylate exclusively at guanine residues via the 2-amino group, with little or no alkylation at N3 and N7 guanine or adenine sites. The cytotoxicities of the compounds correlated broadly with their reactivities, with the bis(hydroxymethyl)imidazoles being the least cytotoxic (IC50s >1 microM; P388 leukemia) and with the intercalator-linked analogues being more cytotoxic than the corresponding minor-groove-targeted ones. This was true also for the more reactive thioimidazole bis(carbamates) (IC50s 0.8 and 11 microM, respectively), but both were more active than the analogous "untargeted" carmethizole (IC50 20 microM). The bis(hydroxymethyl)pyrrolizine analogues were the most cytotoxic, with IC50s as low as 0.03 microM.     

'98 Escherichia coli SWISS-2DPAGE database update

The combination of two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), computer image analysis and several protein identification techniques allowed the Escherichia coli SWISS-2DPAGE database to be established. This is part of the ExPASy molecular biology server accessible through the WWW at the URL address http://www.expasy.ch/ch2d/ch2d-top.html . Here we report recent progress in the development of the E. coli SWISS-2DPAGE database. Proteins were separated with immobilized pH gradients in the first dimension and sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the second dimension. To increase the resolution of the separation and thus the number of identified proteins, a variety of wide and narrow range immobilized pH gradients were used in the first dimension. Micropreparative gels were electroblotted onto polyvinylidene difluoride membranes and spots were visualized by amido black staining. Protein identification techniques such as amino acid composition analysis, gel comparison and microsequencing were used, as well as a recently described Edman "sequence tag" approach. Some of the above identification techniques were coupled with database searching tools. Currently 231 polypeptides are identified on the E. coli SWISS-2DPAGE map: 64 have been identified by N-terminal microsequencing, 39 by amino acid composition, and 82 by sequence tag. Of 153 proteins putatively identified by gel comparison, 65 have been confirmed. Many proteins have been identified using more than one technique. Faster progress in the E. coli proteome project will now be possible with advances in biochemical methodology and with the completion of the entire E. coli genome.     

Comparison of the frequency and enjoyability of pleasant events in cocaine abusers vs. non-abusers using a standardized behavioral inventory

AIMS: To examine whether cocaine abusers differ from non-abusers in their frequency and enjoyability of engaging in various "pleasant events", in order to approximate the density of positive reinforcement experienced in their natural environment. DESIGN: Comparisons of cocaine abusers to normative data and matched controls. SETTING: An outpatient substance abuse treatment center in Burlington, Vermont, USA. PARTICIPANTS: Subjects included 100 individuals enrolled in outpatient treatment for cocaine abuse or dependence and 50 community volunteers without histories of drug abuse or other major psychiatric illness and matched to cocaine-dependent patients on age, sex and SES. MEASUREMENTS: Diagnostic assessments were based upon clinical interviews using the DSM-III-R checklist. The primary focus of this study was the Pleasant Events Schedule (PES), a self-rated behavioral inventory of the frequency and enjoyability of engaging in "pleasant" activities. Cocaine use history, treatment outcome and other relevant variables were also assessed. FINDINGS: Cocaine abusers reliably reported lower frequency of non-social, introverted, passive outdoor and mood-related activities than controls. These differences remained after controlling for demographic and life-style differences between groups, with the exception of mood-related activities. Perceived enjoyability of the activities did not differ across groups. Intravenous cocaine use and prior treatment for cocaine abuse predicted particularly low frequency of pleasant activities. Greater frequency of non-social activities predicted better treatment outcome. CONCLUSIONS: Drug abuse is associated with low density of certain types of non-drug reinforcement. Systematic increases in these activities may improve treatment outcome.     

Operative management of Haglund's deformity in the nonathlete: a retrospective study

Haglund's deformity, or "pump bump," is a common cause of posterior heel pain. Management of the condition usually consists of nonoperative therapy. This study presents a retrospective study of 65 cases (53 patients), with symptomatic Haglund's deformity in nonathletes (13 male and 40 female), who presented during a 4-year period (1989-1994). Sixty-five percent (39 heels) of these patients failed to respond to nonoperative therapy for an average of 62 weeks, (range, 4-260 weeks). This group of patients went on to operative treatment. Surgical management consisted of excision of the posterior calcaneal tuberosity through a medial longitudinal incision with debridement, reattachment of the Achilles tendon using bone anchors, and 4 weeks of postoperative immobilization. Thirty-nine patients (74%) were contacted for follow-up. The average follow-up period for these patients was 155 weeks, (range, 92-335 weeks). There were 50% excellent results, 47% good results, 3% fair results (1 patient), and no poor results. The Maryland Foot Score for operated heels was an average of 67/100 preoperative and an average of 92/100 postoperative. On unoperated heels the score was an average of 81/100 at first evaluation and an average of 86/100 at final evaluation. Complications included one recurrence of painful prominence, one wound infection, and one incisional neuroma. The outcome of these cases demonstrated that in those patients who fail nonoperative treatment, surgical treatment of Haglund's deformity produces a predictably good surgical result when performed using the technique described.     

Desired concentration-dependent ion exchange for micro-optic lenses

The required concentration-dependent diffusion coefficients for both ideal one-dimensional and ideal radial gradient-index profiles are determined. The modified quasi-chemical diffusion model is used to relate the diffusion coefficient to optimum glass composition. Adding aluminum to sodium silicate glasses facilitates the approach to the desired concentration dependence of the diffusion coefficient for silver-sodium ion exchange. A parabolic one-dimensional index profile is fabricated in one of the glasses. It deviates from ideal values by less than 2%.     

Eucapnic voluntary hyperventilation as a bronchoprovocation technique: development of a standarized dosing schedule in asthmatics

A variety of dosing schedules have been reported for the hyperventilation method of broncho-provocation testing. To evaluate the effect of challenge technique on the bronchoconstrictive response, we had 16 subjects perform eucapnic voluntary hyperventilation (EVH) with dry, room temperature gas using four different dosing schedules. The hyperventilation challenge dosages included the following: (1) a target minute ventilation (VE) of 20 x FEV1 for 6 min; (2) a target VE of 15 x FEV1 for 12 min; (3) an interrupted challenge with a target VE of 30 x FEV1 for 2 min repeated 3 times; and (4) a target VE of 30 x FEV1 for 6 min. Challenges 2, 3, and 4 gave identical absolute ventilatory challenges (identical factor FEV1 x minutes) but at different VE dosages or time. Challenges 1 and 4 were of identical length, but different target VE. The mean postchallenge fall in FEV1 was 16.6 +/- 10.9%, 11.0 +/- 8.1%, 19.6 +/- 9.9%, and 26.7 +/- 11.3% for challenges 1, 2, 3, and 4, respectively. The response to an identical EVH challenge (FEV1 x 30 for 6 min) was reproducible when performed on separate days. We conclude that the challenge technique used for hyperventilation testing will have a significant impact on the bronchoconstrictive response and must be taken into account when interpreting study results. Tests may be quantitatively comparable over a narrow range of challenge time and VE. We recommend that a 6-min uninterrupted EVH challenge using dry, room temperature gas at a target VE of 30 x FEV1 be adopted as the "standard" challenge.     

Analysis of factors contributing to success in a program of micromanipulation-assisted fertilization

OBJECTIVE: To determine factors important to clinical success in micromanipulation-assisted in vitro fertilization (IVF). DESIGN: Procedures invoked in two separate series of micromanipulation-assisted IVF cycles, one unsuccessful (series I) and the other successful (series II), were compared in an effort to identify changes that led to clinical success. SETTING: University-based IVF clinic. PATIENTS: In both IVF series involving micromanipulation, patients consisted of infertile couples who fit any of five categories of male-factor related infertility. The female patients underwent controlled hyperstimulation for oocyte retrieval and the oocytes were inseminated normally or were subjected either to partial zona dissection or subzonal sperm insertion to assist fertilization. Results in all groups were compared between the two patient series. RESULTS: In the successful series II, a noticeable improvement in fertilization rate and embryo quality was observed compared with series I. A significant increase in the percentage of patients reaching embryo transfer, the pregnancy rate per transfer, and the pregnancy rate per retrieval were noted in series II; a 25% ongoing pregnancy rate per retrieval was observed overall in this successful group, with "ongoing" defined as manifestation of at least a fetal sac on ultrasound with no detectable problems. Patients with a mixed transfer of embryos derived from manipulated and normally inseminated oocytes had a 75% rate of pregnancy per transfer in series II. Differences between the two series could not be attributed to patient selection or biases in selection of oocytes relegated to micromanipulation. However, oocyte handling, micromanipulation, and culture protocols differed significantly between the two series in that temperature and pH of oocytes was better controlled, and micromanipulation time was minimized in series II. CONCLUSION: Success in micromanipulation depends on maintenance of the oocyte in a stable and supportive environment throughout the micromanipulation procedure. It is also important to minimize trauma to the eggs by performing micromanipulation rapidly and with minimal distortion of the egg. Patients with a poor fertilization rate in standard IVF may experience a substantial increase in the likelihood of pregnancy when micromanipulation-assisted fertilization is performed on some eggs.     

Educated insane: a nineteenth-century psychiatric paradigm

In the 1830s the time-honored notion that excess study could lead to madness underwent a significant change in America. Under the influence of Enlightenment pedagogy and phrenology, influential superintendents like Amariah Brigham and Isaac Ray feared that the "unnatural" overstimulation of children in schools would ruin their development. In the second half of the nineteenth century, as belief in environmental determinism waned and assumptions about what is "natural" changed, this psychiatric etiology was debated; then, overthrown. By the turn of the century, education was thought to aid, not harm, the mentally ill.