Quality of life predicts overall survival in women with platinum-resistant ovarian cancer: an AURELIA substudy.

BACKGROUND: Women with platinum-resistant ovarian cancer are a heterogeneous group whose median overall survival is 12 months. We hypothesized that their quality of life (QoL) scores would be prognostic. PATIENTS AND METHODS: Data from AURELIA (n = 326), a randomized trial of chemotherapy with or without bevacizumab, were used to identify baseline QoL domains [EORTC (European Organisation for Research and Treatment of Cancer) QLQ-C30 and OV28] that were significantly associated with overall survival in multivariable Cox regression analyses. Patients were classified as having good, medium, or poor risk. Cutpoints were validated in an independent dataset, CARTAXHY (n = 136). Multivariable analyses of significant QoL domains on survival were adjusted for clinicopathological prognostic factors. The additional QoL information was assessed using C statistic. RESULTS: In AURELIA, all domains, except cognitive function, predicted overall survival in univariable analyses. Physical function (P < 0.001) and abdominal/gastrointestinal symptom (P < 0.001) scores remained significant in multivariable models. In high (score <67), medium (67-93), and low (>93) risk categories for physical function, median overall survival was 11.0, 14.7, and 19.3 months, respectively (P < 0.001). In CARTAXHY, median overall survival was 7.9, 16.2, and 23.9 months (P < 0.001), respectively. For high- (>44), medium- (13-44), and low- (<13) risk categories for abdominal/gastrointestinal symptoms, median overall survival was 11.9, 14.3, and 19.7 months in AURELIA (P < 0.001) and 10.5, 19.6, and 24.1 months in CARTAXHY (P = 0.02). Physical function (P = 0.02) and abdominal/gastrointestinal symptoms (P = 0.03) remained independent prognostic factors after adjustment for clinicopathological factors. The C statistic of the full model was 0.71. For QoL factors alone, patient factors alone and disease factors alone, the C statistics were 0.61, 0.61, and 0.67 respectively. CONCLUSIONS: Physical function and abdominal/gastrointestinal symptom scores improved predictions of overall survival over clinicopathological factors alone in platinum-resistant ovarian cancer. This additional prognostic information could improve trial stratification, patient-doctor communication about prognosis, and clinical decision-making. CLINICAL TRIAL REGISTRATION: NCT00976911.

Clinical and biomarker profile of trauma-induced secondary cardiac injury.

BACKGROUND: Secondary cardiac injury has been demonstrated in critical illness and is associated with worse outcomes. The aim of this study was to establish the existence of trauma-induced secondary cardiac injury, and investigate its impact on outcomes in injured patients. METHODS: Injured adult patients eligible for enrolment in the Activation of Coagulation and Inflammation in Trauma 2 study, and admitted to the intensive care unit between January 2008 and January 2010, were selected retrospectively for the study. Markers of cardiac injury (brain natriuretic peptide (BNP), heart-type fatty acid binding protein (H-FABP) and troponin I) were measured on admission, and after 24 and 72 h in blood samples from injured patients. Individual records were reviewed for adverse cardiac events and death. RESULTS: During the study period, 135 patients were enrolled (106 male, 78·5 per cent) with a median age of 40 (range 16-89) years. Eighteen patients (13·3 per cent) had an adverse cardiac event during admission and these events were not associated with direct thoracic injury. The in-hospital mortality rate was higher among the adverse cardiac event cohort: 44 per cent (8 of 18) versus 17·1 per cent (20 of 117) (P = 0·008). Raised levels of H-FABP and BNP at 0, 24 and 72 h, and troponin I at 24 and 72 h, were associated with increased adverse cardiac events. BNP levels were higher in non-survivors on admission (median 550 versus 403 fmol/ml; P = 0·022), after 24 h (794 versus 567 fmol/ml; P = 0·033) and after 72 h (1043 versus 753 fmol/ml; P = 0·036), as were admission troponin I levels. CONCLUSION: Clinical and cardiac biomarker characteristics support the existence of trauma-induced secondary cardiac injury, which is associated with death, and unrelated to direct thoracic injury.

Constructed wetland treatment system in textile industry and sustainable development.

This study focuses on the evaluation of the adequacy and sustainability of a constructed wetland (CW), with vertical flow (VF) design to treat a strongly coloured textile wastewater.Secondly an accidental AO7 overloaded discharge (700 mg l(-1)) was studied. A set of three similar VFCW beds (3x1 m2), operating in series, allowed also the efficient treatment of the AO7 heavy loaded wastewaters. The treated effluent quality enables water reuse for irrigation purposes or within the process.

Vertical flow constructed wetland for textile effluent treatment.

A pulse feed vertical flow constructed wetland (VFCW) proved to be efficient in the treatment of a textile effluent being able to buffer, dilute and treat an Acid Orange (AO7) accidental discharge. The influence of the flooding level (FL) and pulse feed (PF) duration on the removal efficiencies of a VFCW was examined. Average AO7 removal efficiencies of 70% were achieved for an AO7 Inlet concentration of 700 mgl(-1) applied during 15 min cycle(-1) (every three hours) at a hydraulic load of 13 lm(-2) cycle(-1) and an FL of 21%. The VFCW was modelled by analogy with a combination of ideal reactors. The simplest combination that best reproduced the experimental results was an association of 2 reactors in series plus 1 reactor accounting the dead volumes. The model parameters helped to understand the hydrological and kinetic processes occurring in VFCW. Through the model simulation it was shown that 3 VFCW in series were enough to efficiently treat an organic mass load of 76 gAO7 m(-2) day(-1) in 9 hours and fulfil the discharge legislation. In this work it was possible to establish that the overall degradation kinetics was of first order.

Functional assessment of coronary artery disease by cardiac computed tomography.

INTRODUCTION: Rapid technological advances in computed tomography (CT) have allowed CT coronary angiography (CTCA) to be delivered at low radiation dose and high diagnostic accuracy. Due to its high negative predictive value for coronary artery disease, it has become a gatekeeper for the assessment of patients with chest pain of recent onset. Second line functional assessment of a detected coronary stenosis relies mostly on other imaging modalities. Functional assessment of coronary artery disease by CT is therefore an attractive addition to CTCA. Areas covered: This review will discuss the current evidence base and future development for CT perfusion imaging. Furthermore, this review will discuss CT-derived fractional flow reserve and CT coronary plaque characteristics as alternative approaches for functional evaluation of coronary artery disease. Finally, combining coronary anatomy and functional assessment of coronary flow with myocardial tissue characterization by CT may be attractive allowing triple assessment by CT. Expert commentary: The combined use of CTCA and functional assessment of coronary artery stenosis by CT perfusion or CT-derived fractional flow reserve is an attractive diagnostic pathway that requires further evaluation.

Aerobic degradation of acid orange 7 in a vertical-flow constructed wetland.

Biological, aerobic degradation of an azo dye and of the resultant, recalcitrant, aromatic amines in a constructed wetland (CW) was demonstrated for the first time. A vertical-flow CW, planted with Phragmites sp. was fed with 127 mgl(-1) of acid orange 7 (AO7) at hydraulic loads of 28, 40, 53 and 108 l m(-2) day(-1). Color removal efficiencies of up to 99% clearly demonstrate cleavage of the azo bond, also confirmed by the similar AO7 removal and SO(4)(2-) release rates revealing that adsorption onto the matrix was constant. The positive redox potential at the outlet demonstrates that aerobic conditions were present. Chemical oxygen demand and total organic carbon removal efficiencies of up to 93% were also indicative of AO7mineralization. The degradation of sulfanilic acid was confirmed by the presence of NO(3)(-), SO(4)(2-) and secondary metabolites, which suggest at least two degradation pathways leading to a common compound, 3-oxoadipate.

A quantitative proteomic approach to highlight Phragmites sp. adaptation mechanisms to chemical stress induced by a textile dyeing pollutant.

Phragmites sp. is present worldwide in treatment wetlands though the mechanisms involved in the phytoremediation remain unclear. In this study a quantitative proteomic approach was used to study the prompt response and adaptation of Phragmites to the textile dyeing pollutant, Acid Orange 7 (AO7). Previously, it was demonstrated that AO7 could be successfully removed from wastewater and mineralized in a constructed wetland planted with Phragmites sp. This azo dye is readily taken up by roots and transported to the plant aerial part by the xylem. Phragmites leaf samples were collected from a pilot scale vertical flow constructed wetland after 0.25, 3.25 and 24.25h exposure to AO7 (400mgL-1) immediately after a watering cycle used as control. Leaf soluble protein extraction yielded an average of 1560 proteins in a broad pI range (pH3-10) by two-dimensional gel electrophoresis. A time course comparative analysis of leaf proteome revealed that 40 proteins had a differential abundance compared to control (p<0.05) within a 3.25h period. After 24.25h in contact with AO7, leaf proteome was similar to control. Adaptation to AO7 involved proteins related with cellular signalling (calreticulin, Ras-related protein Rab11D and 20S proteasome), energy production and conversion (adenosine triphosphate synthase beta subunit) carbohydrate transport and metabolism (phosphoglucose isomerase, fructose-bisphosphate aldolase, monodehydroascorbate reductase, frutockinase-1 and Hypothetical protein POPTR_0003s12000g and the Uncharacterized protein LOC100272772) and photosynthesis (sedoheptulose-1,7-bisphosphatase and ferredoxin-NADP+ reductase). Therefore, the quantitative proteomic approach used in this work indicates that mechanisms associated with stress cell signalling, energy production, carbohydrate transport and metabolism as well as proteins related with photosynthesis are key players in the initial chemical stress response in the phytoremediation process of AO7.

Quantitative general theory for periodic breathing in chronic heart failure and its clinical implications.

BACKGROUND: In patients with chronic heart failure (CHF), periodic breathing (PB) predicts poor prognosis. Clinical studies have identified numerous risk factors for PB (which also includes Cheyne-Stokes respiration). Computer simulations have shown that oscillations can arise from delayed negative feedback. However, no simple general theory quantitatively explains PB and its mechanisms of treatment using widely-understood clinical concepts. Therefore, we introduce a new approach to the quantitative analysis of the dynamic physiology governing cardiorespiratory stability in CHF. METHODS AND RESULTS: An algebraic formula was derived (presented as a simple 2D plot), enabling prediction from easily acquired clinical data to determine whether respiration will be unstable. Clinical validation was performed in 20 patients with CHF (10 with PB and 10 without) and 10 healthy normal subjects. Measurements, including chemoreflex sensitivity (S) and delay (delta), alveolar volume (V(L)), and end-tidal CO(2) fraction (C), were applied to the stability formula. The breathing pattern was correctly predicted in 28 of the 30 subjects. The principal combined parameter (CS)x(delta/V(L)) was higher in patients with PB (14.2+/-3.0) than in those without PB (3.1+/-0.5; P:=0.0005) or in normal controls (2.4+/-0.5; P:=0.0003). This was because of differences in both chemoreflex sensitivity (1749+/-235 versus 620+/-103 and 526+/-104 L/min per atm CO(2); P:=0.0001 and P:<0.0001, respectively) and chemoreflex delay (0.53+/-0.06 vs 0.40+/-0.06 and 0.30+/-0.04 min; P:=NS and P:=0.02). CONCLUSION: This analytical approach identifies the physiological abnormalities that are important in the genesis of PB and explicitly defines the region of predicted instability. The clinical data identify chemoreflex gain and delay time (rather than hyperventilation or hypocapnia) as causes of PB.

Origin of oscillatory kinetics of respiratory gas exchange in chronic heart failure.

BACKGROUND: Respiratory gas exchange measurements in patients with chronic heart failure (CHF) at rest and during exercise commonly reveal prominent slow oscillations in ventilation (V(E)), measured oxygen uptake (VO(2)), and carbon dioxide production (VCO(2)), whose origin is not clear. Voluntary simulation of periodic breathing (PB) in normals has been reported to generate a different pattern of oscillations in gas exchange from that seen in spontaneous PB. This necessitates hypothesizing that PB is caused by a primary oscillation in tissue metabolism or in cardiac output. METHODS AND RESULTS: We developed an automated method by which normal controls could be guided to breathe according to a PB pattern. The resultant metabolic oscillations closely matched those seen in spontaneous PB and had several interesting properties. At low workloads (including rest), the oscillations in VO(2) were as prominent as those in V(E) in both spontaneous PB (alpha(VO2)/alpha(VE)=0.92+/-0.04) and voluntary PB (0.93+/-0.07). However, at increased workload, the oscillations in VO(2) because less prominent than those in V(E) in spontaneous PB (intermediate workload 0.63+/-0.05, high workload 0.57+/-0.04; P<0.001) and voluntary PB (intermediate 0.66+/-0.03, high 0.48+/-0.03; P<0.001). There was no difference in the relative size of metabolic oscillations between voluntary and spontaneous PB at matched workloads (P>0.05 at low, intermediate, and high workloads). Furthermore, VO(2) peaked before V(E) in both spontaneous and voluntary PB. This time delay varied from 6.4+/-0.4 s at low ventilation, to 11.3+/-0.9 s at high ventilation (P<0.0001). CONCLUSIONS: The magnitude and phase pattern of oscillations in gas exchange of spontaneous PB can be obtained by adequately matched voluntary PB. Therefore, the gas exchange features of PB are explicable by primary ventilatory oscillation.

Oxygenation in patients with a functionally univentricular circulation and complete mixing of blood: are saturation and flow interchangeable?

BACKGROUND: Perioperative management of patients with complete mixing of pulmonary and systemic blood centers on approximately equating pulmonary (Qp) and systemic (Qs) blood flow (Qp/Qs approximately 1). This empirically derived target is opposed by theoretical studies advocating a target Qp/Qs well below 1. We studied the cause of this persistent discrepancy. METHODS AND RESULTS: Classic theoretical studies have concentrated on maximizing 1 of many potential combination parameters of arterial oxygen content (CaO(2)) and systemic blood flow: total oxygen delivery (DO(2))=CaO(2)xQs. We defined "useful" oxygen delivery as the amount of oxygen above a notional saturation threshold (Sat(Thresh)): D(u)O(2)=carrying capacityx(SaO(2)-Sat(Thresh))xQs. Whereas DO(2) peaks at Qp/Qs ratios <1, D(u)O(2) peaks at higher Qp/Qs ratios, nearer to (or exceeding) 1. Systemic venous saturation (which mirrors tissue oxygen tension) peaks at Qp/Qs=1. CONCLUSIONS: First, the standard model of single-ventricle physiology can be reexpressed in a form allowing analysis by differential calculus, which allows broader conclusions to be drawn than does computer modeling alone. Second, the classic measure DO(2) fails to reflect the fact that proportional changes in saturation and flow are not clinically equivalent. Recognizing this asymmetry by using D(u)O(2) can give a target Qp:Qs balance that better represents clinical experience. Finally, to avoid an arbitrary choice of Sat(Thresh), systemic venous oxygen saturation (SsvO(2)) may be a useful parameter to maximize: this occurs at a Qp/Qs ratio of 1. Attempts to increase DO(2) by altering Qp/Qs away from this value will inevitably reduce SsvO(2) and therefore tissue oxygenation. Oxygen delivery is far from synonymous with tissue oxygen status.

Enhanced ventilatory response to exercise in patients with chronic heart failure and preserved exercise tolerance: marker of abnormal cardiorespiratory reflex control and predictor of poor prognosis.

BACKGROUND: In patients with chronic heart failure (CHF) and preserved exercise tolerance, the value of cardiopulmonary exercise testing for risk stratification is not known. Elevated slope of ventilatory response to exercise (VE/VCO(2)) predicts poor prognosis in advanced CHF. Derangement of cardiopulmonary reflexes may trigger exercise hyperpnea. We assessed the relationship between cardiopulmonary reflexes and VE/VCO(2)and investigated the prognostic value of (VE/VCO(2)) in CHF patients with preserved exercise tolerance. METHODS AND RESULTS: Among 344 consecutive CHF patients, we identified 123 with preserved exercise capacity, defined as a peak oxygen consumption (PEAK VO(2)) >/=18 mL. kg(-1). min(-1) (age 56 years; left ventricular ejection fraction 28%; peak VO(2) 23.5 mL. kg(-1). min(-1)). Hypoxic and hypercapnic chemosensitivity (n=38), heart rate variability (n=34), baroreflex sensitivity (n=20), and ergoreflex activity (n=20) were also assessed. We identified 40 patients (33%) with high VE/VCO(2) (ie, >34.0). During follow-up (49+/-22 months, >3 years in all survivors), 34 patients died (3-year survival 81%). High VE/VCO(2) (hazard ratio 4.3, P<0.0001) but not peak f1.gif" BORDER="0">O(2) (P=0.7) predicted mortality. In patients with high VE/VCO(2), 3-year survival was 57%, compared with 93% in patients with normal VE/VCO(2) P<0.0001). Patients with high VE/VCO(2) demonstrated impaired reflex control, as evidenced by augmented peripheral (P=0.01) and central (P=0.0006) chemosensitivity, depressed low-frequency component of heart rate variability (P<0.0001) and baroreflex sensitivity (P=0.03), and overactive ergoreceptors (P=0.003) compared with patients with normal VE/VCO(2). CONCLUSIONS: In CHF patients with preserved exercise capacity, enhanced ventilatory response to exercise is a simple marker of a widespread derangement of cardiovascular reflex control; it predicts poor prognosis, which VO(2) does not.

Exercise capacity and biochemical profile during exercise in patients with glycogen storage disease type I.

Glycogen storage disease type I (GSD-I) is an inherited disorder of carbohydrate metabolism. Hepatic glucose-6-phosphatase is deficient, leading to impaired gluconeogenesis and glycogenolysis. Patients prevent fasting hypoglycemia by frequent feeds of low glycemic index foods. Normal muscle does not contain glucose-6-phosphatase, and GSD-I is usually classified as a hepatic glycogenosis. However, clinical experience has suggested that patients have decreased cardiovascular fitness, but this had not been formally investigated. This paper reports the results of maximal treadmill cardiopulmonary exercise testing in adult patients with GSD-I. It documents a major reduction in exercise capacity in these patients and demonstrates biochemical aspects of exercise that are different from those of normal controls. All patients showed a reduction in exercise capacity, but there was a wide range of exercise tolerance. Additional work needs to address whether improved adherence to or intensification of therapy in adulthood will ameliorate exercise intolerance.

Effect of altering conditions of the sequence method on baroreflex sensitivity.

BACKGROUND: The sequence method is widely used as a simple, non-invasive measure of baroreflex sensitivity (BRS). This technique, originally described in anaesthetized cats, has been transferred virtually unchanged to humans, without evidence that the optimal values in cats are the same as those in patients with cardiovascular disease. OBJECTIVE: To study the effect of altering the components of the sequence method on the measured BRS in patients with chronic heart failure (CHF) and in normal individuals. METHODS: Eighty patients with CHF [aged 62 +/- 12 years (mean +/- SD)] and 40 normal control individuals [aged 38 +/- 15 years (mean +/- SD)] underwent measurement of heart rate and non-invasive blood pressure. Altering only the shift between blood pressure and R-R interval and the required correlation coefficient of the regression line had no effect on the value of BRS, but had a significant effect on the number of valid sequences. Alteration of the blood pressure or R-R interval thresholds, however, affected not only the number of valid sequences, but also the value of BRS in both groups. In normal controls, agreement with the bolus phenylephrine method was improved by increasing the blood pressure threshold, although this led to a reduction in the number of valid sequences. In patients with CHF, agreement was optimized by decreasing both the blood pressure and R-R interval thresholds. This also had the effect of increasing the number of valid sequences. CONCLUSION: Changes should be made to this technique, to optimize its validity in conscious humans, particularly when applied to patients with attenuated BRS.

Dinucleotide analogues as inhibitors of thymidine kinase, thymidylate kinase, and ribonucleotide reductase.

P1-(Adenosine-5')-Pn-(thymidine-5') tri-, tetra-, penta-, and hexaphosphates (ApnT) plus the analogues with a methylene group alpha, beta to the thymidine residue (ApncpT) were synthesized by coupling the appropriate two nucleotides, having activated one by morpholine. These were tested as potential dinucleotide inhibitors of thymidine kinase, thymidylate kinase, and ribonucleotide reductase. All three enzymes bind ATP and thymidine or its nucleotides and therefore might be inhibited by dinucleotides containing adenosine and thymidine. Ap5T and Ap6T strongly inhibited all three enzymes (IC50 = 2.4-20 microM). Ap4cpT and Ap5cpT also strongly inhibited the two kinases (IC50 = 4-20 microM) but were much weaker inhibitors of the reductase (IC50 = 130 and 230 microM).

A reappraisal of the effect upon thymidine kinase of thymidine derivatives carrying large groups at the 5'-position.

Several thymidine derivatives with hydrophobic 5'-substituents, linked by chemically stable amide and ether links, were synthesized as potential thymidine kinase inhibitors. None of these was active nor were several derivatives of thymidine 5'-acetate, which were previously reported to be inhibitors. It was shown that the apparent inhibition by the latter compounds was due to their facile hydrolysis in aqueous solution with release of thymidine. These results must cast doubt on any conclusions drawn from biological studies with 5'-esters of thymidine.

Quinazoline antifolates inhibiting thymidylate synthase: variation of the amino acid.

Five new analogues (1c-g) of the antifolate N10-propargyl-5,8-dideazafolic acid (1a) are described in which the benzoyl-L-glutamate moiety was replaced by benzoic acid (desglutamyl-N10-propargyl-5,8-dideazafolic acid), benzoyl-L-aspartate, 4-phenylbutyrate, benzoylglycine, and benzoyl-L-alanine. The esters of the appropriate 4-aminophenyl (benzoyl) starting materials were sequentially alkylated upon nitrogen, first with a propargyl halide and then with 2-amino-6-(bromomethyl)-4-hydroxyquinazoline hydrobromide. Saponification of the antifolate esters so produced gave the desired analogues. The new derivatives (1c-g) and also the known diethyl ester of 1a (1b) were tested for their inhibition of purified L1210 thymidylate synthase (TS) and for their inhibition of the growth of L1210 cells in culture. The TS inhibition of the analogues 1b-g was estimated by calculating the inverse relative potency, defined as the ratio IC50(compound)/IC50(1a). The results obtained were as follows: greater than 62, 84, 9, 333, 21, and 5, respectively. All were thus less inhibitory than 1a. None of the compounds improved upon 1a in inhibiting the growth of L1210 cells in culture.

Polymorphism of the TRAP gene of Plasmodium falciparum.

Natural sequence variation of the thrombospondin related anonymous protein (TRAP) gene of Plasmodium falciparum has been investigated by DNA analysis following the polymerase chain reaction amplification, and this shows the gene to be highly polymorphic. The region containing the sequence motif Trp-Ser-Pro-Cys-Ser-Val-Thr-Cys-Gly (WSPCSVTCG), common to TRAP, the circumsporozoite protein, properdin, and thrombospondin, was invariant. Elsewhere in the molecule, over 50 amino acid substitutions are described including the insertion of an in-frame, small-variable tandemly repeating motif between amino acid residues 352 and 353. Only one silent mutation was observed. Most nucleotide changes that occur in the first two codon positions result in conservative amino acid changes. Restriction fragment length polymorphism (RFLP) analysis was used to examine inheritance of TRAP in a cross between the HB3 and 3D7 clones of P. falciparum. Out of nine progeny examined, four possessed the HB3 gene and five the 3D7 gene. The TRAP gene hybridized to chromosome 13. Previous work has shown that a subtelomeric region of chromosome 13 from the 3D7 parent (marked by the HRP-III gene) was favoured strongly in this cross. The TRAP gene, however, is over 1 Mb away from this subtelomeric region and exhibits no such linkage because of chromosome crossovers. Five geographically separate isolates shared the same TRAP sequence as well as the same variant of the Th2R/Th3R region from the circumsporozoite protein. The correlation between independent markers in these isolates suggests that they have a common provenance.

Relation of heart rate and blood pressure turbulence following premature ventricular complexes to baroreflex sensitivity in chronic congestive heart failure.

Reduced heart rate variability (HRV) and attenuated baroreflex sensitivity (BS) after myocardial infarction and in patients with chronic congestive heart failure (CHF) are associated with poor prognosis. Recent studies have shown that a large proportion of the prognostic power from HRV measurements is localized in heart rate turbulence immediately after ventricular premature complexes. The mechanism of heart rate turbulence remains unknown. In the present study, we explore its relation to BS. In 45 patients with CHF and > or =3 ectopic beats in a 30-minute period, measurements of RR interval and continuous, noninvasive blood pressure (BP) were studied at rest. In response to an ectopic beat, average heart rate turbulence was 9.4 ms/beat (SD 6.1). Mean BP turbulence was 0.72 mm Hg/beat (SD 0.56). Using the ratio of heart rate and BP turbulence slopes to estimate BS showed good agreement (r = 0.67, p < 0.0001) with the alpha-index method (BSalpha). This relation was attributable to a marked correlation between heart rate turbulence and BSalpha (r = 0.70, p <0.0001); there was no correlation between BP turbulence and the BSalpha (r = 0.1, p = NS). Twenty-nine percent of patients had postectopic pulsus alternans, with a mean decay time of 1.4 beats (SD 0.5). The presence of pulsus alternans was associated with a significantly lower heart rate turbulence slope (6.3 [SEM 1.0] vs 10.7 [SEM 1.2] ms/beat, p = 0.03). Thus, heart rate turbulence is an effective measure of the baroreflex, correlating strongly with a standard measure. This is because it is the heart rate, rather than the BP, response to an ectopic beat that conveys the information relevant to BS measurement.

Identification of novel reduced pyridinium derivatives as synthetic co-factors for the enzyme DT diaphorase (NAD(P)H dehydrogenase (quinone), EC 1.6.99.2).

The enzyme DT diaphorase (NAD(P)H dehydrogenase (quinone), EC 1.6.99.2) is unusual in that it can utilize either NADH or NADPH as a co-factor for the reduction of its substrates. We have shown that the intact NAD(P)H molecule is not required and that other reduced pyridinium compounds can also act as co-factors for DT diaphorase. The entire adenine dinucleotide portion of NAD(P)H can be dispensed with entirely and the simplest quaternary (and therefore reducible) derivative of nicotinamide, 1-methylnicotinamide, was as effective as NAD(P)H as a co-factor for the reduction of the quinone, menadione. Nicotinamide 5'-O-benzoyl riboside was also as effective a co-factor as NAD(P)H, whilst nicotinamide ribotide and riboside have a higher Km, and decreased the kcat of DT diaphorase. Nicotinic acid derivatives had little activity. Kinetic analysis indicated that both nicotinamide ribotide and riboside may be interacting with the menadione binding site rather than the NAD(P)H site. Irrespective of the differences between the various reduced pyridinium derivatives in their ability to act as co-factors for the reduction of menadione by DT diaphorase, all the compounds that showed activity in this assay were equally effective co-factors for the reduction of the nitrobenzamide, CB 1954 (5-(aziridin-1-yl)-2,4-dinitrobenzamide). The apparent Km of DT diaphorase for all these co-factors approached zero. It was concluded that co-factor binding is not a rate-limiting step in the nitroreductase activity of DT diaphorase.

Virtual cofactors for an Escherichia coli nitroreductase enzyme: relevance to reductively activated prodrugs in antibody directed enzyme prodrug therapy (ADEPT).

A nitroreductase enzyme has been isolated from Escherichia coli that has the unusual property of being equally capable of using either NADH or NADPH as a cofactor for the reduction of its substrates which include menadione as well as 5-(aziridin-1-yl)-2,4-dinitrobenzamide (CB 1954). This property is shared with the mammalian enzyme, DT diaphorase. The nitroreductase can, like DT diaphorase, also use simple reduced pyridinium compounds as virtual cofactors. The intact NAD(P)H molecule is not required and the simplest quaternary (and therefore reducible) derivative of nicotinamide, 1-methylnicotinamide (reduced), is as effective as NAD(P)H in its ability to act as an electron donor for the nitroreductase. The structure-activity relationship is not identical to that of DT diaphorase and nicotinic acid riboside (reduced) is selective, being active only for the nitroreductase. Irrespective of the virtual cofactor used, the nitroreductase formed the same reduction products of CB 1954 (the 2- and 4-hydroxylamino derivatives in equal proportions). Nicotinic acid riboside (reduced), unlike NADH, was stable to metabolism by serum enzymes and had a plasma half-life of seven minutes in the mouse after an i.v. bolus administration. NADH had an unmeasurably short half-life. Nicotinic acid riboside (reduced) could also be produced in vivo by administration of nicotinic acid 5'-O-benzoyl riboside (reduced). These results demonstrate that the requirement for a cofactor need not be a limitation in the use of reductive enzymes in antibody directed enzyme prodrug therapy (ADEPT). It is proposed that the E. coli nitroreductase would be a suitable enzyme for ADEPT in combination with CB 1954 and a synthetic, enzyme-selective, virtual cofactor such as nicotinic acid riboside (reduced).