Transamidase site-targeted agents alter the conformation of the transglutaminase cancer stem cell survival protein to reduce GTP binding activity and cancer stem cell survival.

Type 2 transglutaminase (TG2) is an important cancer stem cell survival protein that exists in open and closed conformations. The major intracellular form is the closed conformation that functions as a GTP-binding GTPase and is required for cancer stem cell survival. However, at a finite rate, TG2 transitions to an open conformation that exposes the transamidase catalytic site involved in protein-protein crosslinking. The activities are mutually exclusive, as the closed conformation has GTP binding/GTPase activity, and the open conformation transamidase activity. We recently showed that GTP binding, but not transamidase activity, is required for TG2-dependent cancer stem cell invasion, migration and tumour formation. However, we were surprised that transamidase site-specific inhibitors reduce cancer stem cell survival. We now show that compounds NC9, VA4 and VA5, which react exclusively at the TG2 transamidase site, inhibit both transamidase and GTP-binding activities. Transamidase activity is inhibited by direct inhibitor binding at the transamidase site, and GTP binding is blocked because inhibitor interaction at the transamidase site locks the protein in the extended/open conformation to disorganize/inactivate the GTP binding/GTPase site. These findings suggest that transamidase site-specific inhibitors can inhibit GTP binding/signalling by driving a conformation change that disorganizes the TG2 GTP binding to reduce TG2-dependent signalling, and that drugs designed to target this site may be potent anti-cancer agents.

Different patterns of gene expression of topoisomerase II isoforms in differentiated tissues during murine development.

The expression of DNA topoisomerase II alpha and beta genes was studied in murine normal tissues. Northern blot analysis using probes specific for the two genes showed that the patterns of expression were different among 22 tissues of adult mice. Expression levels of topoisomerase II alpha gene were high in proliferating tissues, such as bone marrow and spleen, and undetectable or low in 17 other tissues. In contrast, high or intermediate expression of topoisomerase II beta gene was found in a variety of tissues (15) of adult mice, including those with no proliferating cells. Topoisomerase II gene expression was also studied during murine development. In whole embryos both genes were expressed at higher levels in early than late stages of embryogenesis. Heart, brain and liver of embryos two days before delivery, and these same tissues plus lung and thymus of newborn (1-day-old) mice expressed appreciable levels of the two genes. Interestingly, a post-natal induction of the beta gene expression was observed in the brain but not in the liver; conversely, the expression of the alpha gene was increased 1 day after birth in the liver but not in the brain. However, gene expression of a proliferation-associated enzyme, thymidylate synthase, was similar in these tissues between embryos and newborns. Thus, the two genes were differentially regulated in the post-natal period, and a tissue-specific role may be suggested for the two isoenzymes in the development of differentiated tissues such as the brain and liver. Based on the differential patterns of expression of the two isoforms, this analysis indicates that topoisomerase II alpha may be a specific marker of cell proliferation, whereas topoisomerase II beta may be implicated in functions of DNA metabolism other than replication.

A practical approach to exercise echocardiography: immediate postexercise echocardiography.

Echocardiographic measurements of left ventricular end-systolic dimension and fractional shortening obtained in the supine position before and immediately after maximal upright exercise were evaluated in 11 normal volunteers, 35 patients with coronary artery disease and 17 patients without coronary artery disease. The time course of recovery from acute exercise-induced changes in echocardiographic dimensions was analyzed using serial postexercise recordings from normal subjects. An exercise-induced decrease in end-systolic dimension (greater than or equal to 3 mm) and increase in fractional shortening (greater than or equal to 5%) persisted for 3 minutes or longer in the immediate postexercise period in each of the normal volunteers. With these criteria to separate normal from abnormal responses, abnormal responses were observed in 16 (94%) of 17 patients with coronary artery disease and in only 2 (6%) of 35 patients without coronary artery disease. Immediate postexercise echocardiography appears to be a practical and potentially valuable adjunct in the detection of coronary artery disease.

Dual-chamber pacing with a short atrioventricular delay in congestive heart failure: a randomized study.

OBJECTIVES: This prospective study assessed the initial hemodynamic effects and long-term clinical benefits of dual-chamber pacing with a short atrioventricular (AV) delay in patients with chronic heart failure who had no traditional indication for pacemaker implantation. BACKGROUND: Dual-chamber pacing with a short AV delay has been proposed as a nonpharmacologic treatment for drug-refractory heart failure. Both initial and long-term hemodynamic as well as functional benefits have been reported. All previous studies have used an AV delay of 100 ms. Despite encouraging results, these previous studies have been anecdotal and uncontrolled. METHODS: This double-blind, randomized, crossover trial included 12 subjects with chronic congestive heart failure despite optimal medical therapy. Patients were required to be in sinus rhythm with no evidence of significant bradyarrhythmias. On the day after implantation of a dual-chamber pacemaker, invasive hemodynamic measurements were made at varying AV delays between 100 and 200 ms. Patients were then randomized to either dual-chamber pacing with a 100-ms AV delay or backup mode (VVI at 40 beats/min). After 4 to 6 weeks, crossover to the other pacing mode was programmed. RESULTS: Hemodynamic measurements on the day after pacemaker implantation demonstrated no benefit of pacing with any AV delay compared with intrinsic conduction. At the optimal AV interval for each patient, neither cardiac output (4.5 +/- 1.5 vs 4.7 +/- 1.6 liters/min [mean +/- SD]) nor wedge pressure (16 +/- 10 vs 17 +/- 8 mm Hg) improved significantly from baseline measurements during intrinsic conduction. The long-term pacing protocol was completed in nine patients. Ejection fraction was 16 +/- 6% with dual-chamber (VDD mode) pacing and 18 +/- 4% in backup mode (p = NS). No patient had an increase in ejection fraction by > or = 5% with VDD pacing, nor did any patient improve in New York Heart Association functional class with short AV delay dual-chamber pacing. Also, there were no significant reductions in body weight or diuretic requirements during this pacing period. CONCLUSIONS: Dual-chamber pacing with a short AV delay does not improve hemodynamic and clinical status or ejection fraction measured on the day after pacemaker implantation in patients with chronic congestive heart failure. Routine use of pacemaker therapy with a short AV delay aas a primary treatment of heart failure in patients without standard arrhythmic indications is unwarranted.

Effect of postural changes, nitroglycerin and verapamil on diastolic ventricular function as determined by radionuclide angiography in normal subjects.

Whereas improvement in diastolic function indexes in response to therapeutic interventions has been attributed to a beneficial effect of the intervention, measurements of diastolic function appear to be influenced by changes in loading conditions, heart rate and sympathetic tone. To determine the effect of body position and short-term pharmacologic intervention on radionuclide angiographically determined left ventricular peak filling rate, high temporal resolution time-activity curves and absolute left ventricular volumes obtained by equilibrium-gated blood pool scans were evaluated in 12 normal subjects in the supine position at rest and in response to several postural and pharmacologic manipulations. This study confirmed the reproducibility of the technique and demonstrated that in normal subjects, peak filling rate varies in response to changes in body position and to short-term administration of sublingual nitroglycerin and intravenous verapamil. Peak filling rate ranged from 3.3 to 5.1 end-diastolic volumes (EDV)/s with a variability of 13.7% during five baseline supine measurements in the 12 subjects. Compared with values in the supine position (mean +/- SEM = 4.38 +/- 0.24 EDV/s), peak filling rate increased +16 +/- 6% to 4.75 +/- 0.27 EDV/s in the upright position (p less than 0.05) but did not change significantly with leg elevation. Peak filling rate at baseline and during postural changes correlated significantly with ejection fraction (r = +0.49), with stroke volume (r = +0.26) and inversely with end-systolic volume (r = -0.41), but did not correlate with heart rate or blood pressure.(ABSTRACT TRUNCATED AT 250 WORDS)

Prognostic importance of the serum magnesium concentration in patients with congestive heart failure.

Magnesium abnormalities are common in patients with congestive heart failure but the clinical and prognostic significance of an abnormal serum magnesium concentration in this disorder has not been investigated. Therefore, the relation between serum magnesium concentration and the clinical characteristics and long-term outcome of 199 patients with chronic heart failure was evaluated. The serum magnesium concentration was less than 1.6 mEq/liter in 38 patients (19%), within the normal range in 134 patients (67%) and greater than 2.1 mEq/liter in 27 patients (14%). Patients with hypomagnesemia had more frequent ventricular premature complexes and episodes of ventricular tachycardia than did patients with a normal serum magnesium concentration (p less than 0.05). Even though the two groups were similar with respect to severity of heart failure and neurohormonal variables, patients with a low serum magnesium concentration had a significantly worse prognosis during long-term follow-up (45% versus 71% 1 year survival, p less than 0.05). Patients with hypermagnesemia had more severe symptoms, greater neurohormonal activation and worse renal function than did patients with a normal serum magnesium concentration but tended to have fewer ventricular arrhythmias. Hypermagnesemic patients had a worse prognosis than did those with a normal magnesium concentration (37% versus 71% 1 year survival, p less than 0.05). In conclusion, the measurement of serum magnesium concentration provides important clinical and prognostic information in patients with chronic heart failure.

Effects of BG9719 (CVT-124), an A1-adenosine receptor antagonist, and furosemide on glomerular filtration rate and natriuresis in patients with congestive heart failure.

OBJECTIVES: To determine the effects of furosemide and the selective A1 adenosine receptor BG9719 on renal function in patients with congestive heart failure (CHF). BACKGROUND: Studies suggest that adenosine may affect renal function by various mechanisms, but the effects of blockade of this system in humans is unknown. In addition, the effects of a therapeutic dose of furosemide on glomerular filtration rate (GFR) and renal plasma flow (RPF) in heart failure patients are controversial. METHODS: On different days, 12 patients received placebo, BG9719 and furosemide. Glomerular filtration rate, RPF and sodium and water excretion were assessed immediately following drug administration. RESULTS: Glomerular filtration rate was 84 +/- 23 ml/min/1.73m2 after receiving placebo, 82 +/- 24 following BG9719 administration and a decreased (p < 0.005) 63 +/- 18 following furosemide. Renal plasma flow was unchanged at 293 +/- 124 ml/min/1.73m2 on placebo, 334 +/- 155 after receiving BG9719 and 374 +/- 231 after receiving furosemide. Sodium excretion increased from 8 +/- 8 mEq following placebo administration to 37 +/- 26 mEq following BG9719 administration. In the six patients in whom it was measured, sodium excretion was 104 +/- 78 mEq following furosemide administration. CONCLUSIONS: Natriuresis is effectively induced by both furosemide and the adenosine A1 antagonist BG9719 in patients with CHF. Doses of the two drugs used in this study did not cause equivalent sodium and water excretion but only furosemide decreased GFR. These data suggest that adenosine is an important determinant of renal function in patients with heart failure.

Beneficial effects of metoprolol in heart failure associated with coronary artery disease: a randomized trial.

OBJECTIVES: This clinical trial was performed to determine the safety and clinical impact of titrated metoprolol therapy in patients with heart failure, documented coronary artery disease and a low ejection fraction. BACKGROUND: Despite known cardiodepressant effects, long-term use of beta-adrenergic antagonists appears to be beneficial in patients with idiopathic dilated cardiomyopathy. However, this therapy has not been critically evaluated in patients with heart failure and coronary artery disease. METHODS: In 50 patients with heart failure, known coronary artery disease and an ejection fraction < or = 0.40, we examined the impact of metoprolol therapy in a 6-month double-blind, placebo-controlled randomized trial, assessing the frequency of heart failure exacerbations and changes in symptoms (New York Heart Association functional class), ejection fraction and exercise duration. Placebo-treated patients who completed 6-month follow-up studies then underwent a trial with metoprolol therapy (crossover group). RESULTS: Metoprolol was titrated to a mean maximal dose of 87 mg/day (range 25 to 100) without serious adverse reactions. During double-blind therapy, use of a beta-blocker was associated with a significant reduction in the number of hospital admissions (4% vs. 32%, p < 0.05), overall improved functional class (p = 0.02), increased ejection fraction (4 +/- 7% [mean +/- SD] compared with 0 +/- 6%, p < 0.05) and a greater increase in exercise duration (193 +/- 276 vs. 38 +/- 213 s with placebo, p < 0.01). Crossover outcome paralleled the favorable impact seen during randomized metoprolol therapy. CONCLUSIONS: Cautious use of titrated metoprolol appears to be safe and beneficial when added to standard heart failure therapy in patients with dilated cardiomyopathy associated with coronary artery disease.

Sustained hemodynamic response to flosequinan in patients with heart failure receiving angiotensin-converting enzyme inhibitors.

OBJECTIVES: We evaluated the short- and long-term effects of flosequinan in 47 patients with severe heart failure despite ongoing captopril treatment. BACKGROUND: There have been no previous evaluations of the long-term hemodynamic effects of any direct-acting vasodilator in patients with heart failure receiving an angiotensin-converting enzyme inhibitor. Flosequinan is an arterial and venous vasodilator with actions similar to those of the hydralazine-isosorbide dinitrate combination. METHODS: After baseline hemodynamic measurements using balloon-tipped pulmonary artery and radial arterial catheters, patients were randomized to receive 50, 100 or 150 mg of flosequinan daily. Hemodynamic variables were measured immediately before and after short-term flosequinan administration and after 8 weeks of therapy. RESULTS: With short-term flosequinan administration, mean arterial, right atrial and left ventricular filling pressures decreased by 6.4 +/- 1.1, 3.8 +/- 0.5 and 7.3 +/- 0.7 mm Hg, respectively (all p < 0.001). Cardiac index increased by 0.5 +/- 0.1 liters/min per m2, systemic vascular resistance decreased by 616 +/- 105 dynes.s.cm-5 and heart rate increased by 4 +/- 1 beats/min (all p < 0.001). After 8 weeks of long-term flosequinan administration, the vasodilator effect of a dose of flosequinan persisted. Compared with pretreatment baseline values, mean arterial, right atrial and left ventricular filling pressures at the peak effect of flosequinan were decreased by 3.5 +/- 1.3, 2.8 +/- 0.7 and 5.1 +/- 1.3 mm Hg, respectively (all p < 0.01). Systemic vascular resistance had decreased by 585 +/- 95 dynes.s.cm-5, cardiac index had increased by 0.5 +/- 0.1 liters/min per m2 and heart rate had increased by 10 +/- 2 beats/min (all p < 0.001). CONCLUSIONS: The arterial and venous vasodilator flosequinan exerts both short- and long-term sustained hemodynamic effects in patients with heart failure receiving angiotensin-converting enzyme inhibitors.

Assessment of severity of mitral stenosis by echocardiographic leaflet separation.

Mitral valve area (MVA) determined at cardiac catheterization was compared with M mode echocardiographic measurements in 44 patients with mitral stenosis and no substantial mitral regurgitation. Despite statistically significant correlations, measurements of anterior leaflet motion, including rate of diastolic closure (EF slope) were not useful in predicting severity of stenosis. In contrast, maximal diastolic separation of anterior and posterior leaflets (SEP) was more closely correlated with MVA and appears to have some predictive value. Narrow separation was associated with severe mitral stenosis. Wide separation was associated with relatively mild stenosis. Intermediate values in 16 of 44 patients (36%) were not of predictive value. Recognizing this limitation, measurement of maximal diastolic mitral leaflet separation from M mode echocardiograms is proposed as a simple and useful method for assessing severity of mitral stenosis.

Routine serum enzyme tests in the diagnosis of acute myocardial infarction. Cost-effectiveness.

To determine the cost-effectiveness of routine use of serial SGOT, lactic dehydrogenase (LDH), and LDH isoenzyme determinations in patients with suspected acute myocardial infarction (AMI), 166 consecutive patients admitted to a coronary care unit were prospectively identified and clinical findings analyzed independently using predetermined criteria. Based on chest pain characteristics, ECG, and creatine kinase--MB (CK-MB) results, patients were placed in categories of definite AMI (31%), possible AMI (34%), or AMI excluded (36%). The SGOT and/or LDH patterns were considered positive (ie, suggestive of AMI) in 82% of the patients with definite AMI but only confirmed CK-MB results. Positive SGOT/LDH results yielded new clinically relevant information in only 14 patients (8%). Total charges for SGOT/LDH determinations in these 166 patients totaled $10,938 or approximately $780 for each additional clinically important positive result. When serial ECG and CK-MB results are available, routine serial SGOT/LDH determinations are not justified.

Lidocaine and its active metabolites.

It has been shown that the antiarrhythmic and toxic effects of lidocaine may be in part dependent on its two active metabolites, monoethylglycylxylidide (MEGX) and glycylxylidide (GX). Presently available gas liquid chromatographic analytic methods require long and tedious steps or sophisticated equipment such as gas liquid chromatography-mass spectrometry. The assay method reported here with the use of high-performance liquid chromatography is rapid and allows accurate, precise determination of lidocaine, MEGX, and GX in biologic fluids. On the 3 patients studied extensively with the use of this assay, one patient had MEGX concentrations almost twice those of lidocaine. At 83% lidocaine potency, the contribution of MEGX in this patient was about 1.5 times that of lidocaine. The second patient studied on two consecutive days had a 20% increase in serum lidocaine concentration and an equivalent decrease in MEGX concentration on the second day. In the third patient lidocaine was stopped with a resulting half-life of 3.8 hr, which is consistent with previously reported values for patients on long-term lidocaine infusion. Urinary excretion of lidocaine and its metabolites is in agreement with previous work. These data suggest that much information still remains to be learned about the active metabolites of lidocaine as well as of lidocaine.

Quinidine saliva concentrations: absence of correlation with serum concentrations at steady state.

Serum and saliva quinidine concentrations were measured in eight subjects with cardiac arrhythmias on various dosage regimens. There was good correlation between serum and saliva quinidine concentration after a single dose, but there was no such relationship after repeated dosing. Comparison of the area under a hysteresis loop, obtained by plotting the saliva/serum quinidine concentration ratio as a function of serum quinidine concentration over a dosing interval, indicated an exponential increase with increasing mean serum quinidine concentration. Salivary quinidine concentration predictions based on the Henderson-Hasselbalch equation did not correlate with the serum quinidine concentration under the steady-state conditions. These data suggest that quinidine concentration in saliva is not a direct reflection of its serum concentration in cardiac patients on maintenance (steady-state) therapy and hence not useful for therapeutic drug monitoring.

Fiber optic probe cytometer.

An optical fiber probe is used to both excite and collect fluorescence from a suspension of cells. The configuration of the probe is such that one or a few cells are sensed at a time, with a convenient cell concentration. With fluorescently labeled antibodies to cellular antigens, the fiber optic cytometer is able to identify the presence of a specific set of cells with high sensitivity.

Improvement in depressed cardiac function in hypertensive patients during pindolol treatment.

To assess changes in left ventricular function during antihypertensive treatment using pindolol, a beta-adrenocepter blocking drug with potent intrinsic sympathomimetic activity, serial echocardiographic measurements were obtained in 70 hypertensive patients before and during 15 weeks of treatment with pindolol. For analysis, the patients were separated into three groups on the basis of their baseline left ventricular fractional shortening (Group I, 35 patients with normal fractional shortening of 28 percent or more; Group II, 16 patients with abnormal fractional shortening of 21 to 27 percent; and Group III, 19 patients with markedly abnormal fractional shortening of 20 percent or less). More than half of the patients in Group I and Group II had decreases in mean blood pressure of 10 percent or more in response to pindolol, but only one fourth of Group III patients had similar responses (p less than 0.05). Patients with normal pretreatment fractional shortening had a mild decrease in fractional shortening during pindolol treatment, whereas patients with either abnormal or markedly abnormal fractional shortening had an increase in fractional shortening. This increase in fractional shortening suggests the possibility that the partial agonist or intrinsic sympathomimetic activity of pindolol may play a role in preserving left ventricular function in patients with borderline or impaired function.

Rest angina with transient S-T segment elevation. Correlation of clinical features with coronary anatomy.

In 44 consecutive patients with angina at rest associated with transient S-T segment elevation, clinical features were correlated with angiographic coronary anatomy. Patients were divided into three groups depending on the number of major vessels having greater than or equal to 70 per cent luminal narrowing: Group I = no or minimal disease (six patients); group II = single vessel disease (13 patients); and group III = multiple vessel disease (25 patients). The following features did not differ significantly among groups I, II or III: age, sex, risk factors, time from onset of episodes of pain at rest to study or arrhythmias during ischemic episodes. Patients in group III were more likely to have angina on effort (p less than 0.001) and an abnormal base line electrocardiogram (p less than 0.001) than patients in groups I or II. However, the absence of these features did not separate patients in group I from those in group II. In patients with angina at rest associated with transient S-T segment elevation, clinical features identify patients with multiple vessel disease but do not allow differentiation of patients with no or minimal coronary disease from patients with single vessel disease.

The effects of diuresis on the pharmacokinetics of the loop diuretics furosemide and torsemide in patients with heart failure.

PURPOSE: To evaluate the pharmacokinetics of furosemide and torsemide before and after diuresis in patients presenting with marked fluid overload. SUBJECTS AND METHODS: We studied 44 patients with New York Heart Association class III or IV heart failure, ejection fraction < or =40%, and an estimated excess fluid body weight > or =6.8 kg. Oral furosemide or torsemide was administered before and after diuresis. Pharmacokinetic parameters were assessed before and after diuresis. RESULTS: Following diuresis, maximum plasma concentration increased from 11.0+/-5.0 microg/mL to 13.9+/-6.8 with torsemide (P <0.05) and from 3.1< or =1.5 to 3.9+/-1.9 with furosemide (P=0.16). Maximum concentration increased by more than 30% in only one third of the patients. Total absorption (by area under the curve method) increased 6% among patients on torsemide (P=0.38) and 7% among patients on furosemide (P=0.63) and increased >30% in only 1 torsemide and 2 furosemide patients. The time to maximum concentration decreased from 1.40+/-.82 h to 0.81+/-0.36 with torsemide (P <0.01). There were no differences between furosemide and torsemide in the effects of edema on absorption. CONCLUSION: Marked diuresis altered the pharmacokinetics of both furosemide and torsemide in only a small percentage of patients. The use of adequate doses of oral diuretics in edematous patients may be successful, thereby permitting home treatment with oral diuretics and avoiding the cost of hospitalizations or home intravenous administration services.

Intrinsic sympathomimetic activity of pindolol. Evidence for interaction with pretreatment sympathetic tone.

Eighty-six hypertensive patients were treated with pindolol, a beta-adrenergic blocking drug with intrinsic sympathomimetic activity. To evaluate the interactions between beta-blockade, intrinsic sympathomimetic activity, and the level of pretreatment sympathetic activity, blood pressure, heart rate, and forced expiratory volume in one second (FEV1) were examined before and during a 15-week treatment program. The response of patients with a relatively higher pretreatment sympathetic tone reflected by a resting heart rate equal to or greater than 80 beats per minute (Group I) was compared with the response of patients with a lower pretreatment heart rate (less than 80 beats per minute) (Group II). Decreases in mean blood pressure were similar in the two groups. In patients in Group I, the net effect of pindolol was a modest decrease in heart rate and FEV1, supporting the concept that when sympathetic tone is relatively high, the beta-blocking effect is dominant. In contrast, patients in Group II showed little change in heart rate or FEV1 during pindolol treatment, reflecting a balance between the intrinsic sympathomimetic activity and beta-blocking effects of pindolol. Thus, the intrinsic sympathomimetic activity of pindolol is physiologically evident, and relative impact is dependent on the pretreatment level of sympathetic tone.

Beta-blockers in congestive cardiomyopathy. Conceptual advance or contraindication?

The precise role of adrenergic activity in congestive cardiomyopathy has not been established. A number of mechanisms through which increased catecholamine levels may be harmful, along with the clinical and experimental evidence supporting this concept, are summarized in this review. In this context, the suggestion that beta blockers may be beneficial for patients with severe heart failure, despite their well-known propensity to decrease cardiac contractility, can be better understood. Published reports on the use of beta blocker therapy for congestive cardiomyopathy now include approximately 200 patients, but have yielded inconsistent results. Non-randomized trials in Sweden have suggested increased survival, with most patients having improved functional status while receiving beta blockade, although improvement may take three to six months to become evident. The Swedish group also reported clinical deterioration after discontinuation of beta blockade. Two recent randomized trials in America yielded promising results, but the unexpectedly low mortality in the placebo groups emphasizes the critical importance of concurrent controls. Unfavorable reports have involved small groups with short-duration therapy. Even in these reports, overt aggravation of clinical heart failure has been quite infrequent but sometimes profound. As large scale trials are undertaken, an obvious goal is the development of methods to differentiate the patients with congestive cardiomyopathy who will benefit in response to beta blocker therapy from the few patients who will have a serious adverse response.