Eric P. Brass

Trends in Antimicrobial Drug Development: Implications for the Future

The need for new antimicrobial agents is greater than ever because of the emergence of multidrug resistance in common pathogens, the rapid emergence of new infections, and the potential for use of multidrug-resistant agents in bioweapons. Paradoxically, some pharmaceutical companies have indicated that they are curtailing anti-infective research programs. We evaluated the United States Food and Drug Administration (FDA) databases of approved drugs and the research and development programs of the worlds largest pharmaceutical and biotechnology companies to document trends in the development of new antimicrobial agents. FDA approval of new antibacterial agents decreased by 56% over the past 20 years (1998-2002 vs. 1983-1987). Projecting future development, new antibacterial agents constitute 6 of 506 drugs disclosed in the developmental programs of the largest pharmaceutical and biotechnology companies. Despite the critical need for new antimicrobial agents, the development of these agents is declining. Solutions encouraging and facilitating the development of new antimicrobial agents are needed.

Benefit of exercise conditioning for patients with peripheral arterial disease.

Patients with atherosclerotic peripheral arterial disease (PAD) of the lower extremities have impaired walking ability due to exercise-induced muscle ischemia and the resultant pain of intermittent claudication. To evaluate the benefit of exercise training as a treatment for patients with PAD, as well as possible mechanisms associated with improvement, we randomly assigned 19 men with disabling claudication to treated and control groups. Treatment consisted of supervised treadmill walking (1 hr/day, 3 days/wk, for 12 weeks) with progressive increases in speed and grade as tolerated. Graded treadmill testing was performed to maximal toleration of claudication pain on entry and after 12 weeks of training to define changes in peak exercise performance. After 12 weeks, treated subjects had increased their peak walking time 123%, peak oxygen consumption 30%, and pain-free walking time 165% (all p less than 0.05). Control subjects had no change in peak oxygen consumption, but after 12 weeks, peak walking time increased 20% (p less than 0.05). In treated subjects, maximal calf blood flow (measured by a plethysmograph) increased 38 +/- 45% (p less than 0.05), but the change in flow was not correlated to the increase in peak walking time. Elevated plasma concentrations of acylcarnitines have been associated with the functional impairment of PAD and may reflect the metabolic state of ischemic skeletal muscle. In treated subjects, a 26% decrease in resting plasma short-chain acylcarnitine concentration was correlated with improvement in peak walking time (r = -0.78, p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Clinical Trials for Claudication Assessment of Exercise Performance, Functional Status, and Clinical End Points

Peripheral arterial disease (PAD) affects a large proportion of the general population, with an age-adjusted prevalence of approximately 12% and a prevalence of intermittent claudication of 3% to 7%.1 2 In symptomatic persons, the limited lower extremity arterial supply cannot meet the dynamic metabolic demand of the muscles during ambulatory activities, resulting in the symptom of claudication. Claudication is associated with a severe limitation in walking ability,3 which may adversely affect social, leisure, and occupational activities in many patients.4 The treatment of all patients with PAD is initially directed at cardiovascular risk factor modification, since these individuals have a high future risk of cardiovascular mortality.5 Severely affected patients who have ischemic rest pain or tissue loss are candidates for interventional therapy (bypass surgery or angioplasty) to maintain limb viability.6 7 However, since the majority of patients with claudication are not at short-term risk of limb loss, the primary therapeutic goal is to improve exercise performance and community-based functional status. The past decade has witnessed a marked increase in the evaluation and utilization of therapies to treat patients with claudication.8 Percutaneous transluminal angioplasty is considered an appropriate intervention for patients with “earlier stages of symptomatic disability” due to claudication,9 and the American Heart Association has recently recommended that invasive interventions are appropriate for patients with incapacitating claudication.6 In addition, there is increased interest in medical therapies for claudication. Exercise training elicits well-established and clinically important changes in treadmill exercise performance and community-based walking ability.3 10 11 Recent pharmacological advances have led to a greater use of drugs to treat claudication, with new agents in clinical development. Examples include drugs that alter blood rheology12 and drugs that improve ischemic skeletal muscle metabolism.13 A clinical classification to evaluate therapies for PAD has …

Chronic changes in skeletal muscle histology and function in peripheral arterial disease.

BackgroundPeripheral arterial disease (PAD) is associated with an impairment in exercise performance and muscle function that is not fully explained by the reduced leg blood flow during exercise. This study characterized the effects of PAD on muscle function, histology, and metabolism. Methods and ResultsTwenty-six patients with PAD and six age-matched control subjects were studied. Ten of the PAD patients had unilateral disease, which permitted paired comparisons between their diseased and nonsymptomatic legs. All PAD patients had a lower peak treadmill walking time and peak oxygen consumption than controls. Vascular disease (diseased leg in unilateral patients and the most severely diseased leg in bilateral patients) was associated with decreased calf muscle strength compared with control values. In patients with unilateral disease, the diseased legs had a greater percentage of angular fibers (indicating chronic denervation) and a decreased type H fiber cross-sectional area (expressed as percent of total fiber area) compared with the nonsymptomatic, or control, legs. In diseased legs, gastrocnemius muscle strength was correlated with the total calf cross-sectional area (r=0.78, p<0.05) and type II fiber cross-sectional area (r=0.63, p<0.05). Activities of citrate synthase, phosphofructokinase, and lactate dehydrogenase in all 26 PAD patients (most diseased leg) did not differ from control values. Despite a wide range in citrate synthase activity in PAD patients, activity of this enzyme was not correlated with muscle strength or treadmill exercise performance. ConclusionIn patients with PAD, gastrocnemius muscle weakness is associated with muscle fiber denervation and a decreased type II fiber cross-sectional area. In contrast, the PAD patients displayed substantial heterogeneity in muscle enzyme activities that was not associated with exercise performance. Denervation and type H fiber atrophy may contribute to the muscle dysfunction in patients with PAD and further confirm that the pathophysiology of chronic PAD extends beyond arterial obstruction.

The Deferasirox–AmBisome Therapy for Mucormycosis (DEFEAT Mucor) study: a randomized, double-blinded, placebo-controlled trial

OBJECTIVES Host iron availability is fundamental to mucormycosis pathogenesis. The combination of liposomal amphotericin B (LAmB) and deferasirox iron chelation therapy synergistically improved survival in diabetic mice with mucormycosis. To determine the safety of combination deferasirox plus LAmB therapy for mucormycosis, a multicentred, placebo-controlled, double-blinded clinical trial was conducted. METHODS Twenty patients with proven or probable mucormycosis were randomized to receive treatment with LAmB plus deferasirox (20 mg/kg/day for 14 days) or LAmB plus placebo (NCT00419770, clinicaltrials.gov). The primary analyses were for safety and exploratory efficacy. RESULTS Patients in the deferasirox arm (n=11) were more likely than those in the placebo arm (n=9) to have active malignancy, neutropenia and corticosteroid therapy, and were less likely to receive concurrent non-study antifungal therapy. Reported adverse events and serious adverse events were similar between the groups. However, death was more frequent in the deferasirox than in the placebo arm at 30 days (45% versus 11%, P=0.1) and 90 days (82% versus 22%, P=0.01). Global success (alive, clinically stable, radiographically improved) for the deferasirox arm versus the placebo arm at 30 and 90 days, respectively, was 18% (2/11) versus 67% (6/9) (P=0.06) and 18% (2/11) versus 56% (5/9) (P=0.2). CONCLUSIONS Patients with mucormycosis treated with deferasirox had a higher mortality rate at 90 days. Population imbalances in this small Phase II study make generalizable conclusions difficult. Nevertheless, these data do not support a role for initial, adjunctive deferasirox therapy for mucormycosis.

Acquired skeletal muscle metabolic myopathy in atherosclerotic peripheral arterial disease

Peripheral arterial disease (PAD) is associated with an increased risk of overall cardiovascular mortality, and substantial morbidity resulting from claudication. While the initial disease process is clearly the result of atherosclerosis in the arterial circulation of the limb, altered hemodynamics do not completely explain the pathophysiology of claudication. Work from several laboratories has demonstrated secondary changes in the skeletal muscle of patients with PAD which are consistent with the presence of an acquired metabolic myopathy in these patients. Key findings include an alteration in the expression of mitochondrial enzymes, the accumulation of metabolic intermediates, altered regulation of mitochondrial respiration, increased oxidative stress, and the presence of somatic mutations in the mitochondrial genome. Understanding the metabolic changes associated with PAD is important in understanding the pathophysiology of claudication and in the development of novel therapeutic strategies.

Carnitine and acylcarnitine metabolism during exercise in humans. Dependence on skeletal muscle metabolic state.

Carnitine metabolism has been previously shown to change with exercise in normal subjects, and in patients with ischemic muscle diseases. To characterize carnitine metabolism further during exercise, six normal male subjects performed constant-load exercise on a bicycle ergometer on two separate occasions. Low-intensity exercise was performed for 60 min at a work load equal to 50% of the lactate threshold, and high-intensity exercise was performed for 30 min at a work load between the lactate threshold and maximal work capacity for the individual. Low-intensity exercise was not associated with a change in muscle (vastus lateralis) carnitine metabolism. In contrast, from rest to 10 min of high-intensity exercise, muscle short-chain acylcarnitine content increased 5.5-fold while free carnitine content decreased 66%, and muscle total carnitine content decreased by 19% (all P less than 0.01). These changes in skeletal muscle carnitine metabolism were present at the completion of 30 min of high-intensity exercise, and persisted through a 60-min recovery period. With 30 min of high-intensity exercise, plasma short-chain and long-chain acylcarnitine concentrations increased by 46% and 23%, respectively. Neither exercise state was associated with a change in the urine excretion rates of free carnitine or acylcarnitines. Thus, alterations in skeletal muscle carnitine metabolism, characterized by an increase in acylcarnitines and a decrease in free and total carnitine, are dependent on the work load and, therefore, the metabolic state associated with the exercise, and are poorly reflected in the plasma and urine carnitine pools.

Skeletal Muscle Mitochondrial DNA Injury in Patients With Unilateral Peripheral Arterial Disease

BACKGROUND Patients with peripheral arterial disease (PAD) have exercise limitation due to claudication-limited pain and metabolic alterations in skeletal muscle. PAD is also associated with oxidative stress, which is a known cause of mitochondrial DNA (mtDNA) injury. The present study was designed to test the hypothesis that PAD is associated with mtDNA injury, as reflected by an increased frequency of a specific 4977-base pair (bp) mtDNA deletion mutation. METHODS AND RESULTS The deletion frequency was quantified in gastrocnemius muscle of 8 patients with unilateral PAD and 10 age-matched control subjects with the use of polymerase chain reaction methodologies. Muscle from the hemodynamically unaffected (less affected) PAD limb showed an 8-fold increased deletion frequency and the hemodynamically affected (worse affected) PAD limb had a 17-fold increased deletion frequency compared with muscle from control subjects. The frequency of the 4977-bp deletion in the worse-affected limb was positively correlated with the age of the patients but not the claudication-limited exercise performance of the patients. Total mtDNA content, citrate synthase activity, and cytochrome c oxidase activity were not different in the muscle from the 3 limb populations. However, the ratio of citrate synthase to cytochrome c oxidase was higher in the worse- versus less-affected limbs of PAD patients. CONCLUSIONS The present study demonstrates a large increase in the frequency of the mtDNA 4977-bp deletion in patients with PAD but in a distribution not limited to the hemodynamically affected limb.

Pivalate-Generating Prodrugs and Carnitine Homeostasis in Man

Prodrugs that liberate pivalate (trimethylacetic acid) after hydrolysis have been developed to improve the bioavailability of therapeutic candidates. Catabolism of pivalate released by activation of a prodrug is limited in mammalian tissues. Pivalate can be activated to a coenzyme A thioester in cells. In humans, formation and urinary excretion of pivaloylcarnitine generated from pivaloyl-CoA is the major route of pivalate elimination. Because the total body carnitine pool is limited and can only slowly be replenished through normal diet or biosynthesis, treatment with large doses of pivalate prodrugs may deplete tissue carnitine content. Animal models and long-term treatment of patients with pivalate prodrugs have resulted in toxicity consistent with carnitine depletion. However, low plasma carnitine concentrations after pivalate prodrug exposure may not reflect tissue carnitine content and, thus, cannot be used as a surrogate for potential toxicity. The extent of tissue carnitine depletion will be dependent on the dose of pivalate, because carnitine losses may approximate the pivalate exposure on a stoichiometric basis. These concepts, combined with estimates of carnitine dietary intake and biosynthetic rates, can be used to estimate the impact of pivalate exposure on carnitine homeostasis. Thus, even in populations with altered carnitine homeostasis due to underlying conditions, the use of pivalate prodrugs for short periods of time is unlikely to result in clinically significant carnitine depletion. In contrast, long-term treatment with substantial doses of pivalate prodrugs may require administration of carnitine supplementation to avoid carnitine depletion.

Relationships between muscle mitochondrial DNA content, mitochondrial enzyme activity and oxidative capacity in man: alterations with disease.

Abstract Muscle mitochondrial content is tightly regulated, and requires the expression of both nuclear and mitochondrial genes. In addition, muscle mitochondrial content is a major determinant of aerobic exercise capacity in healthy subjects. The current study was designed to test the hypothesis that in healthy humans, muscle mitochondrial DNA (mtDNA) content is correlated with citrate synthase activity (a representative nuclear-encoded mitochondrial enzyme) and aerobic exercise capacity as defined by whole-body peak oxygen consumption (V˙O2). Furthermore, it was postulated that these relationships might be altered with disease. Twelve healthy and five paraplegic subjects underwent exercise testing and vastus lateralis muscle biopsy sampling. An additional ten healthy subjects and eight patients with unilateral peripheral arterial disease (PAD) underwent exercise testing and gastrocnemius muscle biopsy sampling. Citrate synthase activity and mtDNA content were positively correlated in the vastus lateralis muscles from the healthy subjects. This relationship was similar in muscle from paraplegic subjects. mtDNA content was positively correlated with peak V˙O2 in the healthy subjects and in the paraplegic subjects in whom peak V˙O2 had been elicited by functional electrical stimulation of the muscle. In contrast, the PAD subjects demonstrated higher mtDNA contents than would have been predicted based on their claudication-limited peak V˙O2. Thus, in healthy humans there are strong relationships between muscle mtDNA content and both muscle citrate synthase activity and peak V˙O2. These relationships are consistent with coordinant nuclear DNA and mtDNA expression, and with mitochondrial content being a determinant of aerobic exercise capacity. The relationships seen in healthy humans are quantitatively similar in paraplegic patients, but not in patients with PAD, a disease which is associated with a metabolic myopathy. The relationships between mtDNA content, mitochondrial enzyme activities and exercise capacity provide insight into the physiologic and pathophysiologic regulation of muscle mitochondrial expression.