Preparing healthcare professional students for rural, regional and remote practice: demonstrating the effectiveness of an interprofessional simulation learning experience.

Undertaking an authentic interprofessional simulation experience may be a useful and consistent strategy for healthcare professional students to build competencies required for a rural healthcare context. An observational comparative study design was adopted to evaluate a clinical simulation experience created to develop the interprofessional competencies of a sample of healthcare professional students at a regional university situated on multiple campuses in New South Wales (NSW), Australia. Over 200 students across three campuses of the university were involved in a simulation experience that included four interprofessional activities. Of these students, 189 (89%) agreed to participate in the study. The healthcare professional students who participated in the study were from second year occupational therapy, physiotherapy, and podiatry, and third year speech pathology programs. Retrospective pre and post self-assessed interprofessional collaborative competencies were compared for all students using the revised Interprofessional Collaborative Attainment Survey (ICCAS). Results demonstrated a statistically significant improvement in self-perceived scores using the validated revised ICCAS survey. The findings of this study suggest that carefully designed and authentic interprofessional simulation experiences can facilitate the development of competencies required for effective interprofessional practice, which are necessary for successful rural practice.

Impact of the Pharmacists' Patient Care Process on Medication Adherence in Older Adults With Multimorbidity.

OBJECTIVE: To determine whether the pharmacistled Geriatric Education and Medication Management (GEMM) clinic utilizing the Pharmacists' Patient Care Process (PPCP) improves therapeutic and safety outcomes in ambulatory older adults. DESIGN/PATIENTS: This is a retrospective, case series of veterans newly enrolled into the GEMM clinic from September 2013 to September 2015. SETTING: This study was conducted in an ambulatory pharmacist-led clinic at the Baltimore Veteran Affairs Medical Center in Maryland. INTERVENTION: Demographic, clinical, laboratory, and pharmacy fill data were collected from the computerized patient record system. Medication use patterns were collected using the clinic's supplemental medication reconciliation flowsheet. Descriptive statistics were used to analyze data. MAIN OUTCOME MEASURE(S): Change in adherence rate, number of potentially inappropriate medications, and number of care transitions within the 12-month study period. RESULTS: The primary outcome of medication adherence was high throughout the 12-month study period; beginning at 91.1% at baseline and increasing to 99.0% by the fourth quarter. There was a 36.4% decrease in number of potentially inappropriate medications (PIMs) from 22 at baseline to 14 at 12 months. Medication-related, short-term care transitions decreased from 10 to 4 in the pre- to post-clinic enrollment time frame. Only one subject experienced a long-term care transition during the study period. CONCLUSIONS: Implementing the PPCP in the pharmacist-led GEMM clinic improved medication adherence and persistence, decreased number of PIMs, and assisted in preventing care transitions in ambulatory older adults with multimorbidity and polypharmacy.

Hello darkness my old friend: preferences for darkness vary by neuroticism and co-occur with negative affect.

Metaphors frequently link negative affect with darkness and associations of this type have been established in several experimental paradigms. Given the ubiquity and strength of these associations, people who prefer dark to light may be more prone to negative emotional experiences and symptoms. A five study investigation (total N = 605) couches these ideas in a new theoretical framework and then examines them. Across studies, 1 in 4 people preferred the perceptual concept of dark over the perceptual concept of light. These dark-preferring people scored higher in neuroticism (Studies 1 and 2) and experienced greater depressive feelings in daily life (Study 3). Moreover, dark preferences shared a robust relationship with depressive symptoms (Study 4) as well as generalised anxiety symptoms (Study 5). The results provide novel insights into negative affectivity and extend conceptual metaphor theory in a way that is capable of making individual difference predictions.

Cost-Effectiveness of Screening and Treatment for Cervical Cancer in Tanzania: Implications for other Sub-Saharan African Countries.

OBJECTIVES: To compare the institutional cost per person of screening and treatment between two groups of patients-those screened and those not screened before treatment for cervical cancer at Ocean Road Cancer Institute (ORCI) in Dar es Salaam, Tanzania-and to perform a cost-effectiveness analysis of the ORCI cervical cancer screening program. METHODS: The study included 721 screened and 333 unscreened patients treated at ORCI for cervical cancer from 2002 to 2011. We compared the cost of cervical cancer treatment per patient with life-years gained for patients screened at ORCI versus not screened. RESULTS: Patients with cancer were diagnosed at an earlier stage after participating in screening compared with nonparticipants. For example, 14.0% of stage I cancer patients had received screening by ORCI compared with 7.8% of unscreened cases. For stage IV cancer, these percentages were 1.4% and 6.9%, respectively. Average screening and treatment cost for patients receiving cancer screening ($2526) was higher than that for unscreened patients ($2482). However, we calculated an incremental cost-effectiveness ratio of $219 per life-year gained from receiving cervical cancer screening compared with not being screened, and thus the ORCI screening program was highly cost-effective. Furthermore, the screening program was associated with averting 1.3 deaths from cervical cancer each year resulting from earlier diagnoses of cancer cases, with the incremental cost-effectiveness ratio of $4597 per life saved. CONCLUSIONS: Although Sub-Saharan Africa faces substantial challenges in population health management, our study highlights the potential benefits from expanding access to regular cervical cancer screening for women in this region.

Inactivation of a GAL4-like transcription factor improves cell fitness and product yield in glycoengineered Pichia pastoris strains.

With a completely reengineered and humanized glycosylation pathway, glycoengineered Pichia pastoris has emerged as a promising production host for the manufacture of therapeutic glycoproteins. However, the extensive genetic modifications have also negatively affected the overall fitness levels of the glycoengineered host cells. To make glycoengineered Pichia strains more compatible with a scalable industrial fermentation process, we sought to identify genetic solutions to broadly improve cell robustness during fermentation. In this study, we report that mutations within the Pichia pastoris ATT1 (PpATT1) gene (a homolog of the Saccharomyces cerevisiae GAL4 [ScGAL4] transcriptional activator) dramatically increased the cellular fitness levels of glycoengineered Pichia strains. We demonstrate that deletion of the PpATT1 gene enabled glycoengineered Pichia strains to improve their thermal tolerance levels, reduce their cell lysis defects, and greatly improve fermentation robustness. The extension of the duration of fermentation enabled the PpATT1-modified glycoengineered Pichia strains to increase their product yields significantly without any sacrifice in product quality. Because the ATT1 gene could be deleted from any Pichia strains, including empty hosts and protein-expressing production strains alike, we suggest that the findings described in this study are broadly applicable to any Pichia strains used for the production of therapeutic proteins, including monoclonal antibodies, Fc fusions, peptides, hormones, and growth factors.

A phenylalanine to serine substitution within an O-protein mannosyltransferase led to strong resistance to PMT-inhibitors in Pichia pastoris.

Protein O-mannosyltransferases (PMTs) catalyze the initial reaction of protein O-mannosylation by transferring the first mannose unit onto serine and threonine residues of a nascent polypeptide being synthesized in the endoplasmic reticulum (ER). The PMTs are well conserved in eukaryotic organisms, and in vivo defects of these enzymes result in cell death in yeast and congenital diseases in humans. A group of rhodanine-3-acetic acid derivatives (PMTi) specifically inhibits PMT activity both in vitro and in vivo. As such, these chemical compounds have been effectively used to minimize the extent of O-mannosylation on heterologously produced proteins from different yeast expression hosts. However, very little is known about how these PMT-inhibitors interact with the PMT enzyme, or what structural features of the PMTs are required for inhibitor-protein interactions. To better understand the inhibitor-enzyme interactions, and to gain potential insights for developing more effective PMT-inhibitors, we isolated PMTi-resistant mutants in Pichia pastoris. In this study, we report the identification and characterization of a point mutation within the PpPMT2 gene. We demonstrate that this F664S point mutation resulted in a near complete loss of PMTi sensitivity, both in terms of growth-inhibition and reduction in O-mannosylglycan site occupancy. Our results provide genetic evidence demonstrating that the F664 residue plays a critical role in mediating the inhibitory effects of these PMTi compounds. Our data also indicate that the main target of these PMT-inhibitors in P. pastoris is Pmt2p, and that the F664 residue most likely interacts directly with the PMTi-compounds.

Community-acquired pneumonia episode costs by age and risk in commercially insured US adults aged ≥50 years.

BACKGROUND: Community-acquired pneumonia (CAP) causes substantial clinical and economic burden. While several studies have reported the cost to treat CAP, there is little information on the cost to treat by age, risk profile, and hospitalization in US adults aged ≥50 years. OBJECTIVE: To quantify the cost, from a payer perspective, of treating CAP at the episode level, stratified by age, risk profile, and hospitalization. METHODS: A retrospective study of claims data from a large US health plan (1 January 2006-31 December 2008) was conducted. Patients aged ≥50 years having at least one medical claim with a primary diagnosis for pneumonia were identified. A CAP episode was defined as the period between the first and last pneumonia ICD-9 code with a chest X-ray claim. Episode-level variables included risk stratum based on presence of an immunocompromising/chronic condition, age group, number and length of inpatient and outpatient CAP episodes, and all-cause and CAP-related healthcare costs (adjusted to 2011 costs). RESULTS: Among the 27,659 study patients, 28,575 CAP episodes (20,454 outpatient; 8,121 inpatient) occurred. Mean age of patients with a CAP episode was 62.6. Low-risk patients accounted for 44.4 % of all CAP episodes. Mean CAP episode length was 31.8 days for an inpatient episode and 10.2 days for an outpatient episode. Mean all-cause total healthcare cost for an inpatient CAP episode ranged from $11,148 to $51,219 depending on risk stratum and age group. Mean outpatient episode-related costs were much lower than inpatient episode-related costs. CONCLUSIONS: Cost to treat CAP requiring hospitalization is high regardless of age or the presence of underlying comorbidities. Given that almost half of the patients in this study did not have traditional risk factors for CAP, it is clear that better preventative strategies are needed.

Effect of colostral volume, interval between calving and first milking, and photoperiod on colostral IgG concentrations in dairy cows.

OBJECTIVE: To identify cow and management factors associated with colostral IgG concentration in dairy cows. DESIGN: Prospective observational study. ANIMALS: 81 multiparous Holstein-Friesian cows from a single herd. PROCEDURES: Serum was obtained at the start of the nonlactating period, and cows were assigned to 1 of 4 photoperiod groups: natural day length (n = 22 cows), long days (16 h of light/d [21]) or short days (8 h of light/d [20]) for the entire nonlactating period, or natural day length followed by short days for the last 21 days of the nonlactating period (18). Serum and colostrum were collected at the first milking after calving. Regression analysis was used to investigate associations between colostral IgG concentration and the interval between calving and first milking, colostral volume, photoperiod, length of the nonlactating period, and season of calving. RESULTS: Colostral IgG concentration decreased by 3.7% during each subsequent hour after calving because of postparturient secretion by the mammary glands. The interval between calving and first milking and the colostral volume were significantly and negatively associated with colostral IgG concentration, with the former effect predominating. Photoperiod had no effect on colostral IgG concentration or volume. Serum protein concentration at calving correlated poorly with colostral IgG concentration. CONCLUSIONS AND CLINICAL RELEVANCE: Dairy producers should harvest colostrum as soon as possible after calving to optimize transfer of passive immunity in neonatal calves. Photoperiod can be manipulated without adversely affecting colostral IgG concentration.

Tissue distribution and effects of fasting and obesity on the ghrelin axis in mice.

Ghrelin is a 28 amino acid peptide hormone derived from the 117 amino acid proghrelin, following cleavage by proprotein convertase 1 (PC1). In this study, we comprehensively assessed the tissue distribution and the effect of fasting and obesity on preproghrelin, Exon-4D, PC1 and GOAT expression and proghrelin-derived peptide (PGDP) secretion. The stomach was the major source of preproghrelin expression and PDGPs, followed by the small intestine. The remaining peripheral tissues (including the brain and pancreas) contained negligible expression levels. We detected obestatin in all stomach proghrelin cells, however, 22% of proghrelin cells in the small intestine did not express obestatin. There were strain differences in ghrelin secretion in response to fasting between CD1 and C57BL/6 mice. After a 24 hour-fast, CD1 mice had increased plasma levels of total ghrelin and obestatin with no change in preproghrelin mRNA or PGDP tissues levels. C57BL/6 mice showed a different response to a 24 hour-fast having increased proghrelin mRNA expression, stomach acylated ghrelin peptide and no change in plasma obestatin in C57BL/6 mice. In obese mice (ob/ob and diet-induced obesity (DIO)) there was a significant increase in preproghrelin mRNA levels while tissue and plasma PGDP levels were significantly reduced. Fasting did not affect PGDP in obese mice. Obese models displayed differences in GOAT expression, which was elevated in DIO mice, but reduced in ob/ob mice. We did not find co-localization of the leptin receptor in ghrelin expressing stomach cells, ruling out a direct effect of leptin on stomach ghrelin synthesis and secretion.

Precipitation of a monoclonal antibody by soluble tungsten.

Tungsten microparticles may be introduced into some pre-filled syringes during the creation of the needle hole. In turn, these microcontaminants may interact with protein therapeutics to produce visible particles. We found that soluble tungsten polyanions formed in acidic buffer below pH 6.0 can precipitate a monoclonal antibody within seconds. Soluble tungsten in pH 5.0 buffer at about 3 ppm was enough to cause precipitation of a mAb formulated at 0.02 mg/mL. The secondary structure of the protein was near-native in the collected precipitate. Our observations are consistent with the coagulation of a monoclonal antibody by tungsten polyanions. Tungsten-induced precipitation should only be a concern for proteins formulated below about pH 6.0 since tungsten polyanions are not formed at higher pHs. We speculate that the heterogenous nature of particle contamination within the poorly mixed syringe tip volume could mean that a specification for tungsten contamination based on the entire syringe volume is not appropriate. The potential potency of tungsten metal contamination is highlighted by the small number of particles that would be required to generate soluble tungsten levels needed to coagulate this antibody at pH 5.0.

Lipid levels and low-density lipoprotein cholesterol goal attainment in diabetic patients: rosuvastatin compared with other statins in usual care.

OBJECTIVE: To compare change in low-density lipoprotein cholesterol (LDL-C) levels and National Cholesterol Education Program (NCEP) Adult Treatment Panel III LDL-C goal attainment in diabetic patients treated with rosuvastatin versus other statins in a large, managed care health plan. RESEARCH DESIGN AND METHODS: This retrospective cohort analysis used medical and pharmacy claims linked to laboratory results from a commercial/MedicareAdvantage health plan. Study participants were >or= 18 years of age, had a diagnosis of diabetes, were newly treated with statins from 8/1/03 to 2/28/05, and were considered at high risk for cardiovascular events as defined by NCEP guidelines. Subjects were continuously enrolled for 12 months pre-index and >or= 30 days post-index, with variable follow-up until therapy discontinuation or end of health plan eligibility. MAIN OUTCOME MEASURES: Change in LDL-C from baseline, and attainment of NCEP LDL-C goal among patients not at goal before starting therapy. RESULTS: A total of 3337 adult patients with diabetes were identified with new use of statin therapy during the identification period. A total of 9% (n = 301) started on rosuvastatin, 49.4% (n = 1,649) on atorvastatin, 20.7% (n = 690) on simvastatin, 7.0% (n = 234) on pravastatin, 11.7% (n = 391) on lovastatin and 2.2% (n = 72) on fluvastatin. After controlling for covariates, rosuvastatin patients experienced a significantly greater decrease in LDL-C from baseline (38.7%) than patients taking atorvastatin (34.2%) (p = 0.05), simvastatin (31.5%), pravastatin (24.2%), fluvastatin (26.3%) or lovastatin (24.9%) (p < 0.0001). Rosuvastatin users were significantly more likely to attain LDL-C goal than those taking the other statins (odds ratio: 0.44, 0.28, 0.14, 0.14, 0.19, respectively; p < 0.001). Predicted percent attaining goal was significantly greater for those taking rosuvastatin (87.3%) than for those taking atorvastatin (76.9%), simvastatin (68.7%), pravastatin (55.0%), lovastatin (55.3%) or fluvastatin (61.3%) (p < 0.001). CONCLUSION: For diabetic patients, rosuvastatin is more effective at reducing LDL-C levels and attaining NCEP ATP III LDL-C goal than other statins in real-world clinical practice.

Low-density lipoprotein cholesterol (LDL-C) levels and LDL-C goal attainment among elderly patients treated with rosuvastatin compared with other statins in routine clinical practice.

BACKGROUND: Reducing low-density lipoprotein cholesterol (LDL-C) levels lowers the risk of consequences of cardiovascular disease. Research has confirmed these benefits in elderly patients. The 3-hydroxy-3-methylglutaryl coenzyme A inhibitors (ie, statins) have long-standing proven efficacy in reducing levels of LDL-C and total cholesterol. OBJECTIVE: The goal of this study was to compare change in LDL-C from baseline and National Cholesterol Education Program (NCEP) Adult Treatment Panel (ATP) III LDL-C goal attainment in a population of elderly patients (aged > or =65 years) treated with rosuvastatin versus other statins in routine clinical practice. METHODS: This was a retrospective cohort analysis using medical and pharmacy claims data linked to clinical laboratory results from a large managed care health plan of commercial and Medicare Advantage members in the United States. Included were members aged > or =65 years who were newly treated with statins (index date) from August 1, 2003, through February 28, 2005. All subjects were continuously enrolled for 12 months preindex and > or =30 days postindex, with variable follow-up until therapy discontinuation or end of health plan eligibility. Based on NCEP ATP III guidelines, patients were grouped into risk categories with associated LDL-C goals. The primary outcomes were change in LDL-C from baseline and attainment of NCEP ATP III LDL-C goal among patients not at goal before starting therapy. Generalized linear modeling was used to assess percent change in LDL-C from baseline, controlling for covariates (including age, sex, NCEP risk level, medication possession ratio, preindex LDL-C value, days from index date to postindex LDL-C value, and number of preindex office visits for dyslipidemia). In the subset of patients not at goal before starting therapy, logistic regression was used to estimate the odds of individual patients on other statins reaching goal as compared with rosuvastatin and to produce predicted percent attaining LDL-C goal on individual statins. RESULTS: Of the 2227 elderly new users of statin therapy, 8.0% started on rosuvastatin, 38.9% started on atorvastatin, 3.0% on fluvastatin, 31.0% on lovastatin, 5.5% on pravastatin, and 13.6% on simvastatin. Females comprised 57.7% of the population, and the mean (SD) age was 73 (5.8) years (range, 65-94 years). The mean (SD) doses of rosuvastatin, atorvastatin, fluvastatin, lovastatin, pravastatin, and simvastatin were 10.65 (4.59), 16.0 (12.78), 66.31 (23.56), 27.38 (14.07), 32.86 (16.46), and 28.1 (26.2) mg, respectively. After controlling for covariates, rosuvastatin-treated patients had a 35.8% decrease in LDL-C from baseline, which was significantly greater compared with patients in the atorvastatin, fluvastatin, lovastatin, pravastatin, and simvastatin (29.3%, 21.9%, 22.5%, 22.0%, and 24.9%, respectively; P < 0.05) groups. Atorvastatin (odds ratio [OR], 0.25; 95% CI, 0.12-0.52), fluvastatin (OR, 0.05; 95% CI, 0.02-0.14), lovastatin (OR, 0.10; 95% CI, 0.05-0.20), pravastatin (OR, 0.08; 95% CI, 0.03-0.20), and simvastatin (OR, 0.14; 95% CI, 0.06-0.30) were less likely to attain LDL-C goal compared with rosuvastatin (all, P < 0.001). Predicted percent attaining goal was 93.6% among rosuvastatin users, significantly greater than users of atorvastatin, fluvastatin, lovastatin, pravastatin, and simvastatin (81.2%, 55.8%, 66.8%, 64.1%, and 72.8%, respectively; P < 0.05). CONCLUSION: In this elderly patient population, rosuvastatin was a more effective treatment for reducing LDL-C levels and attaining NCEP ATP III LDL-C goals than the other statins.

Non-covalent ligand/DNA interactions: minor groove binding agents.

An understanding of the mechanism by which minor groove binding agents interact with DNA has led to the design of agents that can reversibly bind with high selectivity to extended DNA target sequences. Simple compounds, such as the polypyrroles and the bis-benzimidazoles, have been used as carriers for alkylating agents effectively directing alkylation to specific DNA sequences. The spectrum of DNA alkylation and mutation by classical alkylators, such as nitrogen mustards, has been profoundly modified by such attachment. The observed "side-by-side" binding of small polypyrrole antibiotics has led to the design of synthetic hairpin polyamides with programmable DNA sequence selectivity. These compounds are able to compete with natural substrates, such as specific transcription factors, and alter gene expression. They are being developed as artificial transcription factors, able to deliver activating peptides to specific recognition sequences, and as potential protein-DNA dimerization agents. Hairpin polyamides are also being used as carriers for the delivery of alkylators to defined DNA sites. The degree of control of gene expression thus offered by the hairpin polyamides suggests enormous promise for their clinical utility. Recent developments with other minor groove binding small molecules and technological advances are also discussed.

A standardized test battery for the study of synesthesia.

Synesthesia is an unusual condition in which stimulation of one modality evokes sensation or experience in another modality. Although discussed in the literature well over a century ago, synesthesia slipped out of the scientific spotlight for decades because of the difficulty in verifying and quantifying private perceptual experiences. In recent years, the study of synesthesia has enjoyed a renaissance due to the introduction of tests that demonstrate the reality of the condition, its automatic and involuntary nature, and its measurable perceptual consequences. However, while several research groups now study synesthesia, there is no single protocol for comparing, contrasting and pooling synesthetic subjects across these groups. There is no standard battery of tests, no quantifiable scoring system, and no standard phrasing of questions. Additionally, the tests that exist offer no means for data comparison. To remedy this deficit we have devised the Synesthesia Battery. This unified collection of tests is freely accessible online (http://www.synesthete.org). It consists of a questionnaire and several online software programs, and test results are immediately available for use by synesthetes and invited researchers. Performance on the tests is quantified with a standard scoring system. We introduce several novel tests here, and offer the software for running the tests. By presenting standardized procedures for testing and comparing subjects, this endeavor hopes to speed scientific progress in synesthesia research.

A case-control study of the HER2 Ile655Val polymorphism in relation to risk of invasive breast cancer.

BACKGROUND: Overexpression of the HER2 proto-oncogene in human cancer cells has been associated with a poor prognosis, and survival improves with therapy targeting the HER2 gene. Animal studies and protein modeling suggest that the Ile655Val polymorphism located in the transmembrane domain of the HER2 protein might influence breast cancer development by altering the efficiency of homodimerization. METHODS: To investigate this genetic polymorphism, incident cases of invasive breast cancer (N = 1,094) and population controls of a similar age (N = 976) were interviewed during 2001 to 2003 regarding their risk factors for breast cancer. By using DNA collected from buccal samples mailed by the participants, the HER2 Ile655Val polymorphism was evaluated with the Applied Biosystems allelic discrimination assay. Odds ratios (ORs) and 95% confidence intervals (95% CIs) were estimated by logistic regression adjusted for numerous breast cancer risk factors. Analysis was restricted to women with self-reported European descent. RESULTS: Prevalence of the Val/Val genotype was 5.6% in cases and 7.1% in controls. In comparison with the Ile/Ile genotype, the Ile/Val genotype was not significantly associated with breast cancer risk (OR 0.97, 95% CI 0.79 to 1.18), whereas the Val/Val genotype was associated with a reduced risk (OR 0.63, 95% CI 0.42 to 0.92). This inverse association seemed strongest in older women (OR 0.51, 95% CI 0.29 to 0.89 for women aged more than 55 years), women without a family history of breast cancer (OR 0.54, 95% CI 0.35 to 0.84), postmenopausal women with greater body mass index (OR 0.43, 95% CI 0.20 to 0.91 for a body mass index of 25.3 kg/m2 or more), and cases diagnosed with non-localized breast cancer (OR 0.49, 95% CI 0.26 to 0.90). CONCLUSION: Although results from our population-based case-control study show an inverse association between the HER2 Ile655Val polymorphism and risk of invasive breast cancer, most other studies of this single-nucleotide polymorphism suggest an overall null association. Any further study of this polymorphism should involve sample populations with complete risk factor information and sufficient power to evaluate gene-environment interactions between the HER2 polymorphism and factors such as age and family history of breast cancer.

Effects of anticancer drugs on transcription factor-DNA interactions.

DNA-interacting anticancer drugs are able to affect the propensity of DNA to interact with proteins through either reversible binding or covalent bond formation. The effect of the drugs on transcription factor interactions with DNA is reviewed. These effects can be classified as (i) competition between a drug and regulatory protein for target sequences; (ii) weakening of this interaction; (iii) enhancement of this interaction by chemical modification of the DNA and the creation of non-natural binding sites; and (iv) a 'suicide' mechanism, which is observed when a transcription factor induces changes in DNA structure, allowing a drug to bind to a target sequence. Several new strategies -- the antigene approach with oligonucleotides, peptide nucleic acids or locked nucleic acids, and sequence-specific polyamides -- are also reviewed.

Demonstration by real-time polymerase chain reaction that cellular DNA alkylation by novel aminoindoline compounds affects expression of the protooncogene c-myc.

Aminoindolines, analogues of the potent DNA alkylating agent seco-CBI-TMI, bind to and alkylate in the minor groove of AT-rich DNA in vitro. Here we extend the in vitro mechanism of action studies by treating cells in culture and examining the DNA binding patterns within AT-rich regions of the protooncogene locus c-myc, using a real-time polymerase chain reaction (PCR) stop assay. In addition, real-time reverse transcriptase (RT) PCR is used to examine the immediate effects of drug treatment on c-myc expression. These analyses demonstrate a concentration and time dependence for DNA alkylation at the chosen sites within the c-myc locus, as well as a prompt and significant downregulation of c-myc expression. While downregulation of this important growth regulator is likely not the only consequence of aminoindoline treatment, these studies begin to address the cellular pathways that are involved in the potent cytotoxic effects observed and provide insights for the future development of anticancer drugs of this class.

DNA and the chromosome - varied targets for chemotherapy.

The nucleus of the cell serves to maintain, regulate, and replicate the critical genetic information encoded by the genome. Genomic DNA is highly associated with proteins that enable simple nuclear structures such as nucleosomes to form higher-order organisation such as chromatin fibres. The temporal association of regulatory proteins with DNA creates a dynamic environment capable of quickly responding to cellular requirements and distress. The response is often mediated through alterations in the chromatin structure, resulting in changed accessibility of specific DNA sequences that are then recognized by specific proteins. Anti-cancer drugs that target cellular DNA have been used clinically for over four decades, but it is only recently that nuclease specific drugs have been developed to not only target the DNA but also other components of the nuclear structure and its regulation. In this review, we discuss some of the new drugs aimed at primary DNA sequences, DNA secondary structures, and associated proteins, keeping in mind that these agents are not only important from a clinical perspective but also as tools for understanding the nuclear environment in normal and cancer cells.

Transcription factors as targets for DNA-interacting drugs.

Gene expression, both tissue specific or inducible, is controlled at the level of transcription by various transcription factors interacting with specific sequences of DNA. Anticancer drugs and other potential therapeutic agents alter interactions of regulatory proteins with DNA by a variety of different mechanisms. The main ones, considered in the review, are: i) competition for the transcription factor DNA binding sequences by drugs that interact non-covalently with DNA (e.g. anthracyclines, acridines, actinomycin D, pyrrole antibiotics and their polyamide derivatives); ii) covalent modifications of DNA by alkylating agents (e.g. nitrogen mustards, cisplatin) that prevent transcription factors from recognizing their specific sequences, or that result in multiple "unnatural" binding sites in DNA which hijack the transcription factors, thus decreasing their availability in the nucleus; iii) competition with binding sites on the transcription factors by synthetic oligonucleotides or peptide nucleic acids in an antigene strategy. The latter compounds may also compete for binding sites on regulatory proteins, acting as decoys to lower their active concentration in the cell. In this review, we have summarized recent advances which have been made towards understanding the above mechanisms by which small molecules interfere with the function of transcription factors.

A new family of cyclophilins with an RNA recognition motif that interact with members of the trx/MLL protein family in Drosophila and human cells.

A new family of cyclophilins with an RNA recognition motif (RRM) has members in vertebrates, roundworms and flatworms. We have identified a Drosophilacyclophilin, Dcyp33, with a high degree of amino acid sequence identity and similarity with other members of the family. Dcyp33 interacts through its RRM domain with the third PHD finger of trithorax. This interaction is conserved in the human homologues of these proteins, Cyp33 and MLL. Over expression of Dcyp33 in DrosophilaSL1 cells results in down-regulation of AbdominalB Hoxgene expression, mirroring the effect of human Cyp33 on the expression of human HOXgenes.