Platelet Count as a Prognostic Factor in Stage IV Non-Small Cell Lung Cancer.

Increase platelet count can accompany various cancers including lung cancer. This finding has recently been suggested to indicate poor prognosis. In patients with malignancies, thrombocytosis has previously been related disease stage, histological type and survival. In this study, the prevalence of thrombocytosis and the prognostic information provided by platelet count were analyzed in patients with stage IV Non-Small Cell Lung Cancer (NSCLC) with an aim to assess elevated platelet count as a prognostic factor in patients with stage IV NSCLC and to investigate whether there is relationship between thrombocytosis, other clinico-pathologic factors and median survival. This prospective observational study was conducted in National Institute of Cancer Research and Hospital (NICRH), Dhaka, Bangladesh from September 2019 to August 2020. A total of 108 patients were enrolled purposively. Detail history taking, thorough physical examination was done along with relevant investigations. Data were collected by semi structured questionnaire and analysis was done with the help of Statistical Package for Social Science (SPSS), version 21.0. The mean age of the patients was found 56.4±12.2 years with range from 35 to 75 years. Majority (79.6%) patients were male, 52.8% patients came from low income and 36.1% were farmer. Majority (40.7%) were symptomatic; in bed >50.0% of day. Almost two third (59.3%) had <5.0% weight loss. Almost three fourth (69.4%) had squamous cell carcinoma. At the time of first assessment 75(69.4%) patients had normal and 33(30.6%) had elevated platelet count level. Age, sex and histological type were statistically not significant between normal and elevated platelet count level groups. But performance status, weight loss were statistically significant (p<0.05) between two groups. According to univariate analysis, age, performance status at presentation, weight loss more than 10.0% for 3 months and platelet count prior the start of treatment were all significant predictors for the overall survival. In multivariate analysis age, performance status at presentation and initial thrombocytosis were independent prognostic determinants for overall survival. Median survival time was significantly higher for the normal platelet count group and elevated platelet count group (7.5 months versus 5.5 months) respectively (95% CI, 5.5-7.5), p<0.001. The frequency of thrombocytosis in patients with stage-IV NSCLC at first presentation was 30.6% and median survival time in these patients was significantly shorter compared in patients without thrombocytosis. These results concluded that an elevated platelet count could be a useful prognostic factor for survival in patients with stage-IV NSCLC.

Clinical Outcomes in Non-Small-Cell Lung Cancer Patients Treated With EGFR-Tyrosine Kinase Inhibitors and Other Targeted Therapies Based on Tumor Versus Plasma Genomic Profiling.

PURPOSE: To compare clinical outcomes in a cohort of patients with advanced non-small-cell lung cancer (NSCLC) with targetable genomic alterations detected using plasma-based circulating tumor DNA (ctDNA) or tumor-based next-generation sequencing (NGS) assays treated with US Food and Drug Administration-approved therapies at a large academic research cancer center. METHODS: A retrospective review from our MD Anderson GEMINI database identified 2,224 blood samples sent for ctDNA NGS testing from 1971 consecutive patients with a diagnosis of advanced NSCLC. Clinical, treatment, and outcome information were collected, reviewed, and analyzed. RESULTS: Overall, 27% of the ctDNA tests identified at least one targetable mutation and 73% of targetable mutations were EGFR-sensitizing mutations. Among patients treated with first-line epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor (TKI) therapies, there were no significant differences in progression-free survival of 379 days and 352 days (P value = .41) with treatment based on tissue (n = 40) or ctDNA (n = 40), respectively. Additionally, there were no differences in progression-free survival or objective response rate among those with low (n = 8, 0.01%-0.99%) versus high (n = 16, ≥ 1%) levels of ctDNA of the targetable mutation as measured by variant allele frequency (VAF). Overall, there was excellent testing concordance (n = 217 tests) of > 97%, sensitivity of 91.7%, and specificity of 99.7% between blood-based ctDNA NGS and tissue-based NGS assays. CONCLUSION: There were no significant differences in clinical outcomes among patients treated with approved EGFR-TKIs whose mutations were identified using either tumor- or plasma-based comprehensive profiling and those with very low VAF as compared with high VAF, supporting the use of plasma-based profiling to guide initial TKI use in patients with metastatic EGFR-mutant NSCLC.

Association of COPD with osteoporosis in male smokers: A case control study in a tertiary medical college hospital in Bangladesh.

OBJECTIVES: Chronic obstructive pulmonary disease (COPD) may increase the risk of osteoporosis and resulting fractures can contribute to disability and mortality of patients. We intended to evaluate the frequency of osteoporosis in male smokers with and without COPD and study whether any correlation existed between osteoporosis and COPD. MATERIALS AND METHODS: This case-control study was carried out in the Department of Medicine, Sylhet MAG Osmani Medical College Hospital, Sylhet, Bangladesh between July 2013 and June 2015. Seventy four male smokers with COPD and 66 age-matched male smokers without COPD were enrolled. All individuals underwent Bone Mass Densitometry (BMD) by Dual-Energy X-Ray Absorptiometry (DEXA). RESULTS: COPD and non-COPD groups did not differ regarding age and smoking pack-years. Osteoporosis at femoral neck (48.6% versus 16.7%; p< 0.001) and lumbar spine (68.9% versus 37.9%; p< 0.01) was significantly higher in COPD compared to controls. Osteopenia did not differ significantly. Patients with COPD were 4.5 times more likely to develop osteoporosis than controls after adjusting age, smoking-pack years and BMI (adjusted OR = 4.5; 95% CI = 1.8-11.5). CONCLUSIONS: Osteoporosis is more frequent in male smokers with COPD compared to smokers without COPD. COPD is a risk factor of osteoporosis independent of age, smoking and BMI.

Association of Low Level of High Density Lipoprotein cholesterol with Acute Coronary syndrome.

Incidence of coronary artery disease (CAD) is increasing in developing countries in Bangladesh with improvement of socioeconomic status, urbanization, changes of dietary habits and lifestyle. Dyslipidaemia is one of the major contributors increase CAD risk. This study was aimed to find out the association of low level HDL-C with acute coronary syndrome. This cross sectional study was conducted in the department of cardiology, Mymensingh Medical College Hospital, Mymensingh, Bangladesh from August 2009 to May 2010. Sociodemographic characteristics, smoking, hypertension, FBS, serum total cholesterol level, HDL-C, LDL-C, Triglyceride level were important variable considered. A total number of 100 respondents consisted of 50 cases (patient) Group I and 50 healthy people (control) Group II. Investigations included ECG, Troponin-I, FBS and Fasting Lipid Profile. The data was analyzed by computer with the help of SPSS. Chi-square test, T-test, ANOVA test used as test of significance. The mean level in cases of HDL-C 39.3±5.1 and in control level HDL-C 34.2±3.4 statistically significant (p<0.0001). In both group low concentration HDL-C (<40mg/dl) risk for CAD. Un-adjusted odds ratio 95% CI determinants of ACS, HDL-C of OR was 0.2. So, HDL-C is not protective factor. In multivariate logistic regression analysis that adjusted for confounders of HDL-C level (age, sex, smoking, hypertension, TC, LDL-C, TG) associated with ACS. HDL-C was strong predictor of ACS (RR in the highest) compared with lowest quarantile = 0.02; (95% CI=0.003-0.173; P for trend <0.0001). The study reflected that low HDL-C level associated with ACS. Categorization of patients with ACS on the basis HDL-C level may be helpful for risk stratification and management.

Admission Troponin I and Post Thrombolytic ST-Segment Resolution in Prediction of Early Risk Stratification in ST- Elevation Myocardial Infarction.

The prognostic value of admission troponin I (tnI) levels and the resolution of the ST-segment elevation in ST-elevation myocardial infarction (STEMI) is well established. However, the combination of these two early available markers for predicting the risk of early mortality has not been evaluated yet. This prospective analytical study conducted in the department of Cardiology, Sir Salimullah Medical College and Mitford Hospital, Dhaka and NICVD, Dhaka, Bangladesh from March 2004 to February 2005. We have evaluated 80 patients with streptokinase treated STEMI who had both admission troponin I and ST-monitoring. We used a prospectively defined cut-off value of troponin I of 0.1ng/ml. For ST-segment resolution, a cut-off of 50% measured after 90 minutes was used. Both a troponin I >0.1ng/ml and ST segment resolution <50% was related to higher early mortality; 16.7% vs. 14.3% (p<0.001) and 57.1% vs. 1.7% (p<0.001) respectively. In a multivariate analysis ST-segment resolution was and troponin I showed a strong trend to be independently related to early mortality. The combination of both further improved risk stratification. The early mortality in the group with elevation of troponin I and without ST-segment resolution compared to the group without troponin I elevation and with ST-segment resolution was 55.6% vs. 0%. Both troponin I on admission and ST-segment resolution after 90 minutes are strong predictors of early mortality. The combination of both gives additive early information about prognosis and further improves risk stratification.

Acute anti-obesity effects of intracerebroventricular 11ß-HSD1 inhibitor administration in diet-induced obese mice.

The hypothalamus is the regulatory centre of both appetite and energy balance and endoplasmic reticulum (ER) stress in the hypothalamus is involved in the pathogenesis of obesity. Recently, inhibition of 11 ß hydroxysteroid dehydrogenase type1 (11ß-HSD1) was reported to have an anti-obesity effect by reducing fat mass. However, the link between the role of 11ß-HSD1 in the hypothalamus and obesity has yet to be determined. In the present study, embryonal primary hypothalamic neurones and high-fat diet (HFD) fed mice were used to investigate the anorexigenic effects of 11ß-HSD1 inhibitors both in vitro and in vivo. In hypothalamic neurones, carbenoxolone (a non selecitve 11ß-HSD inhibitor) alleviated ER stress and ER stress-induced neuropeptide alterations. In HFD mice, i.c.v. administration of carbenoxolone or KR67500 (nonselective and selective 11ß-HSD1 inhibitors, respectively) was associated with less weight gain compared to control mice for 24 hours after treatment, presumably by reducing food intake. Furthermore, glucose regulated protein (Grp78), spliced X-box binding protein (Xbp-1s), c/EBP homologous protein (chop) and ER DnaJ homologue protein (Erdj4) expression was decreased in the hypothalami of mice administrated 11ß-HSD1 inhibitors compared to controls. Conversely, the phosphorylation of protein kinase B (PKB/Akt), signal transducer and activator of transcription 3 (Stat3), mitogen-activated protein kinase (MAPK/ERK) and S6 kinase1 (S6K1) in the hypothalamus was induced more in mice treated using the same regimes. In conclusion, acute 11ß-HSD1 inhibition in the hypothalamus could reduce food intake by decreasing ER stress and increasing insulin, leptin, and mammalian target of rapamycin complex 1 (mTORC1) signalling.

Dexamethasone-mediated oncogenicity in vitro and in an animal model of glioblastoma.

OBJECTIVEDexamethasone, a known regulator of mesenchymal programming in glioblastoma (GBM), is routinely used to manage edema in GBM patients. Dexamethasone also activates the expression of genes, such as CEBPB, in GBM stem cells (GSCs). However, the drug's impact on invasion, proliferation, and angiogenesis in GBM remains unclear. To determine whether dexamethasone induces invasion, proliferation, and angiogenesis in GBM, the authors investigated the drug's impact in vitro, in vivo, and in clinical information derived from The Cancer Genome Atlas (TCGA) cohort.METHODSExpression profiles of patients from the TCGA cohort with mesenchymal GBM (n = 155) were compared with patients with proneural GBM by comparative marker selection. To obtain robust data, GSCs with IDH1 wild-type (GSC3) and with IDH1 mutant (GSC6) status were exposed to dexamethasone in vitro and in vivo and analyzed for invasion (Boyden chamber, human-specific nucleolin), proliferation (Ki-67), and angiogenesis (CD31). Ex vivo tumor cells from dexamethasone-treated and control mice were isolated by fluorescence activated cell sorting and profiled using Affymetrix chips for mRNA (HTA 2.0) and microRNAs (miRNA 4.0). A pathway analysis was performed to identify a dexamethasone-regulated gene signature, and its relationship with overall survival (OS) was assessed using Kaplan-Meier analysis in the entire GBM TCGA cohort (n = 520).RESULTSThe mesenchymal subgroup, when compared with the proneural subgroup, had significant upregulation of a dexamethasone-regulated gene network, as well as canonical pathways of proliferation, invasion, and angiogenesis. Dexamethasone-treated GSC3 demonstrated a significant increase in invasion, both in vitro and in vivo, whereas GSC6 demonstrated a modest increase. Furthermore, dexamethasone treatment of both GSC3 and GSC6 lines resulted in significantly elevated cell proliferation and angiogenesis in vivo. Patients with mesenchymal GBM had significant upregulation of dexamethasone-regulated pathways when compared with patients with proneural GBM. A prognostic (p = 0.0007) 33-gene signature was derived from the ex vivo expression profile analyses and used to dichotomize the entire TCGA cohort by high (median OS 12.65 months) or low (median OS 14.91 months) dexamethasone signature.CONCLUSIONSThe authors present evidence that furthers the understanding of the complex effects of dexamethasone on biological characteristics of GBM. The results suggest that the drug increases invasion, proliferation, and angiogenesis in human GSC-derived orthotopic tumors, potentially worsening GBM patients' prognoses. The authors believe that careful investigation is needed to determine how to minimize these deleterious dexamethasone-associated side effects in GBM.

Mesoblastic Nephroma in a 22 Years Old Woman.

Mesoblastic nephroma is an uncommon renal tumor of infancy and rarely occurs in adults. We report a case of mesoblastic nephroma in adult. A 22-year-old woman, who presented with left flank pain, was found to have a left renal mass by abdominal ultrasonography. Computed tomography revealed a heterogeneous tumor. Left radical nephrectomy was performed. The tumor was a creamy white solid mass. Microscopically, the tumor was composed of spindle cell proliferation. Atypia and mitoses were not identified. Among the tumor cells, there were tubular arranged epithelial elements. The patient was free of recurrence 18 months postoperatively. Mesoblastic nephroma is classified as a benign tumor but recurrence and malignant transformation of this tumor have been reported, so regular postoperative follow up is required.

Detection of arsenic in water, herbal and soil samples by neutron activation analysis technique.

Arsenic contamination of ground water is well understood while other environmental systems are rarely considered to be contaminated by arsenic. A vital issue is whether or not appreciable arsenic transmits through the food chain. Reportedly, ayurvedic herbal medicine products (AHMPs) manufactured in Asia were found to be contaminated by harmful level of Arsenic. This study was aimed to quantify the arsenic levels in water, herbal and soil samples collected from the same origin using highly accurate neutron activation analysis (NAA) technique. Harmful level of arsenic was detected in most of the water and soil samples. Moreover, a considerably harmful level of Arsenic was detected in herbal samples collected from the same origin. As a result, AHMPs manufactured in Asia might be contaminated by arsenic through arsenic contaminated herb plants.

Redefinition of the cleavage sites of DNase I on the nucleosome core particle.

DNase I has been widely used for the footprinting of DNA-protein interactions including analyses of nucleosome core particle (NCP) structure. Our understanding of the relationship between the footprint and the structure of the nucleosome complex comes mainly from digestion studies of NCPs, since they have a well-defined quasi-symmetrical structure and have been widely investigated. However, several recent results suggest that the established consensus of opinion regarding the mode of digestion of NCPs by DNase I may be based on erroneous interpretation of results concerning the relationship between the NCP ends and the dyad axis. Here, we have used reconstituted NCPs with defined ends, bulk NCPs prepared with micrococcal nuclease and molecular modelling to reassess the mode of DNase I digestion. Our results indicate that DNase I cuts the two strands of the nucleosomal DNA independently with an average stagger of 4 nt with the 3'-ends protruding. The previously accepted value of 2 nt stagger is explained by the finding that micrococcal nuclease produces NCPs not with flush ends, but with approximately 1 nt 5'-recessed ends. Furthermore we explain why the DNA stagger is an even and not an odd number of nucleotides. These results are important for studies using DNase I to probe nucleosome structure in complex with other proteins or any DNA-protein complex containing B-form DNA. We also determine the origin of the 10n +/- 5 nt periodicity found in the internucleosomal ladder of DNase I digests of chromatin from various species. The explanation of the 10n +/- 5 nt ladder may have implications for the structure of the 30 nm fibre.

PML bodies associate specifically with the MHC gene cluster in interphase nuclei.

Promyelocytic leukemia (PML) bodies are nuclear multi-protein domains. The observations that viruses transcribe their genomes adjacent to PML bodies and that nascent RNA accumulates at their periphery suggest that PML bodies function in transcription. We have used immuno-FISH in primary human fibroblasts to determine the 3D spatial organisation of gene-rich and gene-poor chromosomal regions relative to PML bodies. We find a highly non-random association of the gene-rich major histocompatibilty complex (MHC) on chromosome 6 with PML bodies. This association is specific for the centromeric end of the MHC and extends over a genomic region of at least 1.6 megabases. We also show that PML association is maintained when a subsection of this region is integrated into another chromosomal location. This is the first demonstration that PML bodies have specific chromosomal associations and supports a model for PML bodies as part of a functional nuclear compartment.

Vpr is preferentially targeted by CTL during HIV-1 infection.

The HIV-1 accessory proteins Vpr, Vpu, and Vif are essential for viral replication, and their cytoplasmic production suggests that they should be processed for recognition by CTLs. However, the extent to which these proteins are targeted in natural infection, as well as precise CTL epitopes within them, remains to be defined. In this study, CTL responses against HIV-1 Vpr, Vpu, and Vif were analyzed in 60 HIV-1-infected individuals and 10 HIV-1-negative controls using overlapping peptides spanning the entire proteins. Peptide-specific IFN-gamma production was measured by ELISPOT assay and flow-based intracellular cytokine quantification. HLA class I restriction and cytotoxic activity were confirmed after isolation of peptide-specific CD8(+) T cell lines. CD8(+) T cell responses against Vpr, Vpu, and Vif were found in 45%, 2%, and 33% of HIV-1-infected individuals, respectively. Multiple CTL epitopes were identified in functionally important regions of HIV-1 Vpr and Vif. Moreover, in infected individuals in whom the breadth of HIV-1-specific responses was assessed comprehensively, Vpr and p17 were the most preferentially targeted proteins per unit length by CD8(+) T cells. These data indicate that despite the small size of these proteins Vif and Vpr are frequently targeted by CTL in natural HIV-1 infection and contribute importantly to the total HIV-1-specific CD8(+) T cell responses. These findings will be important in evaluating the specificity and breadth of immune responses during acute and chronic infection, and in the design and testing of candidate HIV vaccines.

Persistence of human immunodeficiency virus type 1-specific cytotoxic T-lymphocyte clones in a subject with rapid disease progression.

We longitudinally measured T-cell receptor transcript frequencies of human immunodeficiency virus type 1 (HIV-1) specific cytotoxic T lymphocytes (CTL) in an individual with rapidly progressive disease and high levels of viremia. CTL clones elicited during acute HIV-1 infection were present at the time of death, despite absent functional CTL responses, arguing against clonal deletion as a mechanism for the decline of CTL responses observed during HIV-1 infection.

Evolution of anthropoid jaw loading and kinematic patterns.

Major transformations in the skull and masticatory system characterized the evolution of crown anthropoids. To offer further insight into the phylogenetic and arguably adaptive significance of specific primate mandibular loading and kinematic patterns, allometric analyses of metric parameters linked to masticatory function are performed within and between 47 strepsirhine and 45 recent anthropoid species. When possible, basal anthropoids are considered. These results are subsequently integrated with prior experimental and morphological work on primate skull form. As compared to strepsirhines, crown anthropoids have a vertically longer ascending ramus linked to a glenoid and condyle positioned relatively higher above the occlusal plane. Interestingly, anthropoids and strepsirhines do not exhibit different mean ratios of condylar to glenoid height, which suggests that both clades are similar in their ability to evenly distribute occlusal contacts and perhaps forces along the postcanine teeth. Thus, given the considerable suborder differences in the scaling of both glenoid and condylar height, we argue that much of this variation in jaw-joint height is linked to suborder differences in relative facial height due in turn to increased encephalization, basicranial flexion, and facial kyphosis in anthropoids. Due to a more elongate ascending ramus, anthropoids evince more vertically oriented masseters than like-sized strepsirhines. Having a relatively longer ramus and a more medially displaced lateral pterygoid plate, crown anthropoids exhibit medial pterygoids oriented similar to those of strepsirhines, but with a variably longer lever arm. As anthropoid masseters are less advantageously placed to effect transverse movements/forces, we argue that balancing-side deep-masseter activity underlying a wishboning loading regime serves to increase, or at least maintain, transverse levels of jaw movement and occlusal force at the end of the masticatory power stroke. Crown anthropoids are also more isognathic and isodontic than strepsirhines. A consideration of early anthropoids suggests that the crown anthropoid masticatory pattern, i.e., more vertical masseters due to a high condyle as well as greater isognathy and isodonty, occurred stepwise during stem anthropoid evolution. This appears to correspond to a more transverse, and perhaps progressively larger, power stroke across oligopithecids, parapithecids, and propliopithecids.

Hemoglobin H disease in the eastern region of Saudi Arabia.

OBJECTIVE: Alpha-thalassemia is frequently encountered in eastern Saudi Arabia. We wanted to find out laboratory based incidence and laboratory features of Hemoglobin H disease in the Dammam region. METHODS: We retrospectively analyzed the results of Hemoglobin electrophoresis carried out during the last 5 years in our laboratory. Hemoglobin electrophoresis was performed on cellulose acetate, pH 8.6 using Helena or Biomidi kits. Hemoglobin S was confirmed by sickle solubility test. Variant hemoglobin if present, was confirmed by citrate agar (pH 6.0) electrophoresis. Helena rapid electrophoresis system was used for plate densitometry. The diagnosis of Hemoglobin H disease was made on the basis of the presence of Hemoglobin H on electrophoresis supplemented by demonstration of Hemoglobin H inclusions in red blood cells. RESULTS: Fifteen thousand, four hundred and ninety two blood samples were analyzed by Hemoglobin electrophoresis. We found 100 cases of Hemoglobin H disease, only one case was non-Saudi. The age ranged between 45 days to 85 years. There were 51 females and 49 males. Children (less than 12 years) were 35 and of adults there were 65. There were 35 adult females and 30 adult males. The mean +/- standard deviation of Hemoglobin H in children was 13.54 +/- 7, in adult females the mean +/- standard deviation of Hemoglobin H was 12 +/- 5.4, and in adult males it was 11.99 +/- 6.4. The Hemoglobin H inclusions seen in red blood cells ranged from 2.6-80 in children and 10-80 in adults. The sickle cell trait was co-existent in 7 cases. Hemoglobin Bart's along with Hemoglobin H was seen in 32 cases. Hemoglobin F was present, beyond first year of life in 34 cases. The Hemoglobin A2 as measured by densitometry was significantly low in all of the 3 age groups as compared to corresponding controls. The complete blood count results were available for analysis in only 26 cases of Hb H disease. The mean +/- SD values of Hb (g/dl), Hct (ratio), MCV (fl), MCH (pg) MCHC (g/dl), RDW-SD (fl) and RDW-CV (%) in these patients (all age groups together) were 8.15 +/- 1,.278 +/-.04, 59.4 +/- 5.8, 17.65 +/- 2.1, 29.4 +/- 1.7, 37.8 +/- 8.7 and 25.1 +/- 4.6. The mean Hb, Hct, MCV, MCH and MCHC were significantly reduced in all 3 age groups as compared to corresponding controls. RBC counts and RDW-CV were elevated in Hb H disease compared to corresponding controls. The blood film showed typical red cell morphology. CONCLUSION: Hb H disease is not infrequently encountered in the Dammam region. This condition should be kept in mind while evaluating patients for anemia. The genetic studies to determine the exact alpha-thalassemia determinants producing Hb H disease in eastern Saudi Arabia are needed.

HAD, a data bank of heavy-atom binding sites in protein crystals: a resource for use in multiple isomorphous replacement and anomalous scattering.

Information on the preparation and characterization of heavy-atom derivatives of protein crystals has been collected, either from the literature or directly from protein crystallographers, and assembled in the form of a heavy-atom data bank (HAD). The data bank contains coordinate data for the heavy-atom positions in a form that is compatible with the crystallographic data in the Brookhaven Protein Data Bank, together with a wealth of information on the crystallization conditions, the nature of the heavy-atom reagent and references to relevant publications. Some statistical information derived from the data bank, such as the most popular heavy-atom derivatives, is also included. The information can be directly accessed and should be useful to protein crystallographers seeking to improve their success in preparing heavy-atom derivatives for the methods of isomorphous replacement and anomalous dispersion. The World Wide Web address of HAD is http://www.icnet.uk/bmm/had.

Identification and analysis of domains in proteins.

An automatic algorithm based on inter-residue contacts is presented to identify domains in proteins. The results of the algorithm are compared to an assignment performed by inspection that was guided by the authors' description in the literature. The authors' and the algorithm's assignments for a chain were considered to agree if the same number of domains were identified and if the assignments were the same for at least 95% of the residues. With this criterion, the algorithm agreed with the authors' assignment for 78% of the 284 non-redundant chains considered. When some of the authors' assignments were re-evaluated based on the results of the algorithm, an agreement of 84% was obtained. The algorithm is therefore a useful tool for data validation in domain assignment. The authors assignments of domains were analysed for structural principles of domains. The number of chains forming one, two, three, four and five domains are 197, 67, 13, 6 and 1 respectively. Most domains in multidomain proteins are formed from continuous segments and adopt the same structural class. Distributions of the number of residues and the ellipticity of domains and chains are presented. The relationship between accessible surface area and molecular weight for domains and chains is examined.

Towards an intelligent system for the automatic assignment of domains in globular proteins.

The automatic identification of protein domains from coordinates is the first step in the classification of protein folds and hence is required for databases to guide structure prediction. Most algorithms encode a single concept based and sometimes do not yield assignments that are consistent with the generally accepted perception. Our development of an automatic approach to identify reliably domains from protein coordinates is described. The algorithm is benchmarked against a manual identification of the domains in 284 representative protein chains. The first step is the domain assignment by distance (DAD) algorithm that considers the density of inter-residue contacts represented in a contact matrix. The algorithm yields 85% agreement with the manual assignment. The paper then considers how the reliability of these assignments could be evaluated. Finally the use of structural comparisons using the STAMP algorithm to validate domain assignment is reported on a test case.

MacMolecular: a program for visualization of molecular structures on the Macintosh.

MacMolecular displays small- to medium-sized biomolecules, with particular emphasis on peptides. It has been developed to run on color Macintosh computers. The display can be stick, ball and stick, depth cued by thickness stick, or several types of space-filling representations. The program takes input from standard PDB files, simple Cartesian coordinate files, and, in addition, from Kinemage files in which atom information has been included. The program allows color changes of various types as well as the normal functions of translation, rotation, and zooming. In addition, animation files may be produced for subsequent display. Bonding of atoms is done by a distance algorithm (standard) or sequentially to properly display C alpha traces and traces of peptides containing simplified representations of amino acids. Stereo viewing is available, and manipulated structures which were drawn from PDB files can be written out to new PDB files. In addition, PICT files of the drawing window can be generated.

A three-dimensional molecular template for substrates of human cytochrome P450 involved in debrisoquine 4-hydroxylation.

A three-dimensional molecular template has been generated for substrates of human debrisoquine 4-hydroxylase cytochrome P450 (CYP2D6). This template defines the stereochemical requirements for CYP2D6 substrates in terms of the volume occupied and positions of key atoms. The modelling was based on the X-ray crystallographic coordinates of the location of the attacked C5 atom of camphor in relation to the haem in cytochrome P450 cam. Interactive molecular graphics combined with energy calculations were used to identify allowed conformers to superpose known CYP2D6 substrates to yield a molecular template. This model takes into account the site of attack of the known substrates and the requirement for a protonated nitrogen atom to interact with an anion site of the protein. A nitrogen-anion distance of between 2.5 and 4.5 A was allowed for the interaction. The substrates modelled were cardiovascular drugs (debrisoquine, sparteine, guanoxan and perhexiline), beta-adrenergic blocking agents (bufuralol and propranolol), tricyclic anti-depressants (desipramine, amitriptyline and nortriptyline) and other miscellaneous compounds (phenformin, methoxy-amphetamine, codeine and dextromethorphan). The template generated in this manner was then used to determine the likelihood that certain other compounds were substrates for CYP2D6. A carcinogenic protein pyrolysate product, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), did not fit the template and is therefore unlikely to be activated by this enzyme. A potent carcinogen in tobacco smoke, 4-(N-methyl-N-nitrosamino)-1-(3-pyridyl)-1-butanone (NNK), fitted the template but could not be modelled to form a favourable nitrogen-anion interaction. Experimental substrate competition studies also showed that NNK is unlikely to be a CYP2D6 substrate. It was also shown that the widely used drug for treatment of breast cancer, trans-1-(4-beta-dimethylaminoethoxyphenyl)-,2-diphenyl-1-ene (tamoxifen), did not fit the molecular template and is unlikely to be metabolized by CYP2D6. Coordinates of the template are available.