Adjunctive everolimus therapy for tuberous sclerosis complex-associated refractory seizures: Results from the postextension phase of EXIST-3.

OBJECTIVE: Epilepsy is highly prevalent in patients with tuberous sclerosis complex (TSC). Everolimus showed higher efficacy than placebo for seizures in the primary analysis of the EXIST-3 study. Here, we present the long-term outcomes of everolimus at the end of the postextension phase (PEP; data cutoff date: October 25, 2017). METHODS: After completion of the extension phase, patients were invited to continue everolimus in the PEP with everolimus (targeted trough concentration = 5-15 ng/ml, investigator-judged). Efficacy assessments included changes in seizure status during the PEP collected at 12-week intervals as parent/caregiver-reported data through a structured questionnaire. RESULTS: Among 361 patients, 343 entered the extension phase and 249 entered the PEP. After 12 weeks in the PEP, 18.9% (46/244) of patients were seizure-free since the last visit of the extension phase and 64.8% (158/244) had a stable/improved seizure status. At 24 weeks, the corresponding percentages were 18.2% (42/231) and 64.5% (149/231). Among 244 patients, the response rate was 32.8% (80/244) during the 12-week maintenance period of the core phase and 63.9% (156/244) at the end of the extension phase. Of the 149 responders at the end of the extension phase, 70.5% were seizure-free or had stable/improved seizure status. Long-term efficacy data showed persistent responses were observed in 183 of 361 patients (50.7%); 63.9% of these patients had a response that lasted at least 48 weeks. The most frequent Grade 3-4 adverse events (≥2% incidence) reported throughout the study were pneumonia, status epilepticus, seizure, stomatitis, neutropenia, and gastroenteritis. Four patients died during the study. SIGNIFICANCE: The final analysis of EXIST-3 demonstrated the sustained efficacy of everolimus as adjunctive therapy in patients with TSC-associated treatment-refractory seizures, with a tolerable safety profile.

Adjunctive everolimus therapy for treatment-resistant focal-onset seizures associated with tuberous sclerosis (EXIST-3): a phase 3, randomised, double-blind, placebo-controlled study.

BACKGROUND: Everolimus, a mammalian target of rapamycin (mTOR) inhibitor, has been used for various benign tumours associated with tuberous sclerosis complex. We assessed the efficacy and safety of two trough exposure concentrations of everolimus, 3-7 ng/mL (low exposure) and 9-15 ng/mL (high exposure), compared with placebo as adjunctive therapy for treatment-resistant focal-onset seizures in tuberous sclerosis complex. METHODS: In this phase 3, randomised, double-blind, placebo-controlled study, eligible patients aged 2-65 years with tuberous sclerosis complex and treatment-resistant seizures (≥16 in an 8-week baseline phase) receiving one to three concomitant antiepileptic drugs were recruited from 99 centres across 25 countries. Participants were randomly assigned (1:1:1), via permuted-block randomisation (block size of six) implemented by Interactive Response Technology software, to receive placebo, low-exposure everolimus, or high-exposure everolimus. Randomisation was stratified by age subgroup (<6 years, 6 to <12 years, 12 to <18 years, and ≥18 years). Patients, investigators, site personnel, and the sponsor's study team were masked to treatment allocation. The starting dose of everolimus depended on age, body-surface area, and concomitant use of cytochrome 3A4/P-glycoprotein inducers. Dose adjustments were done to attain target trough ranges during a 6-week titration period, and as needed during a 12-week maintenance period of core phase. Patients or their caregivers recorded events in a seizure diary throughout the study. The primary endpoint was change from baseline in the frequency of seizures during the maintenance period, defined as response rate (the proportion of patients achieving ≥50% reduction in seizure frequency) and median percentage reduction in seizure frequency, in all randomised patients. This study is registered with ClinicalTrials.gov, number NCT01713946. FINDINGS: Between July 3, 2013, and May 29, 2015, 366 patients were enrolled and randomly assigned to placebo (n=119), low-exposure everolimus, (n=117), or high-exposure everolimus (n=130). The response rate was 15·1% with placebo (95% CI 9·2-22·8; 18 patients) compared with 28·2% for low-exposure everolimus (95% CI 20·3-37·3; 33 patients; p=0·0077) and 40·0% for high-exposure everolimus (95% CI 31·5-49·0; 52 patients; p<0·0001). The median percentage reduction in seizure frequency was 14·9% (95% CI 0·1-21·7) with placebo versus 29·3% with low-exposure everolimus (95% CI 18·8-41·9; p=0·0028) and 39·6% with high-exposure everolimus (95% CI 35·0-48·7; p<0·0001). Grade 3 or 4 adverse events occurred in 13 (11%) patients in the placebo group, 21 (18%) in the low-exposure group, and 31 (24%) in the high-exposure group. Serious adverse events were reported in three (3%) patients who received placebo, 16 (14%) who received low-exposure everolimus, and 18 (14%) who received high-exposure everolimus. Adverse events led to treatment discontinuation in two (2%) patients in the placebo group versus six (5%) in the low-exposure group and four (3%) in the high-exposure group. INTERPRETATION: Adjunctive everolimus treatment significantly reduced seizure frequency with a tolerable safety profile compared with placebo in patients with tuberous sclerosis complex and treatment-resistant seizures. FUNDING: Novartis Pharmaceuticals Corporation.

Everolimus for subependymal giant cell astrocytoma: 5-year final analysis.

OBJECTIVE: To analyze the cumulative efficacy and safety of everolimus in treating subependymal giant cell astrocytomas (SEGA) associated with tuberous sclerosis complex (TSC) from an open-label phase II study (NCT00411619). Updated data became available from the conclusion of the extension phase and are presented in this ≥5-year analysis. METHODS: Patients aged ≥ 3 years with a definite diagnosis of TSC and increasing SEGA lesion size (≥2 magnetic resonance imaging scans) received everolimus starting at 3mg/m(2) /day (titrated to target blood trough levels of 5-15ng/ml). The primary efficacy endpoint was reduction from baseline in primary SEGA volume. RESULTS: As of the study completion date (January 28, 2014), 22 of 28 (78.6%) initially enrolled patients finished the study per protocol. Median (range) duration of exposure to everolimus was 67.8 (4.7-83.2) months; 12 (52.2%) and 14 (60.9%) of 23 patients experienced SEGA volume reductions of ≥50% and ≥30% relative to baseline, respectively, after 60 months of treatment. The proportion of patients experiencing daily seizures was reduced from 7 of 26 (26.9%) patients at baseline to 2 of 18 (11.1%) patients at month 60. Most commonly reported adverse events (AEs) were upper respiratory tract infection and stomatitis of mostly grade 1 or 2 severity. No patient discontinued treatment due to AEs. The frequency of emergence of most AEs decreased over the course of the study. INTERPRETATION: Everolimus continues to demonstrate a sustained effect on SEGA tumor reduction over ≥5 years of treatment. Everolimus remained well-tolerated, and no new safety concerns were noted.

Everolimus for renal angiomyolipoma in patients with tuberous sclerosis complex or sporadic lymphangioleiomyomatosis: extension of a randomized controlled trial.

BACKGROUND: Mammalian target of rapamycin (mTOR) inhibitors are recommended as first-line treatment of renal angiomyolipoma associated with tuberous sclerosis complex (TSC) or sporadic lymphangioleiomyomatosis (sporadic LAM), but follow-up is limited. Longer term efficacy and tolerability data from a Phase 3, double-blind, placebo-controlled trial are presented. METHODS: Following favorable results from the primary analysis (data cutoff 30 June 2011) of the EXIST-2 trial, patients still receiving study treatment were allowed to enter an open-label extension. Everolimus was initiated at 10 mg once daily and titrated based on tolerability. The primary outcome was angiomyolipoma response rate (≥ 50% reduction from baseline in target lesion volumes). Safety was a secondary endpoint. RESULTS: As of the cutoff date (1 May 2013), 112 patients had received everolimus, and the response rate in 107 patients with angiomyolipoma (median duration of medication exposure of 28.9 months) was 54%. The proportion of patients achieving angiomyolipoma reductions of ≥ 30% and ≥ 50% increased over time, reaching 81.6% (62/76) and 64.5% (49/76), respectively, by Week 96. No everolimus-treated patients experienced renal bleeding. The long-term safety profile was consistent with previous reports; adverse events (AEs) were mostly Grade 1/2, and there were no new safety issues. The frequency of emerging AEs and severe AEs lessened over time. CONCLUSIONS: Longer term everolimus treatment appeared safe and effective in patients with TSC- or sporadic LAM-associated renal angiomyolipoma not requiring surgical intervention. Continued reduction in angiomyolipoma volume was demonstrated, and there was no angiomyolipoma-related bleeding; AEs were predictable and generally manageable. TRIAL REGISTRATION: clinicaltrialsgov identifier: NCT00790400 (http://clinicaltrials.gov/ct2/show/NCT00790400).

Forearm ischemia decreases endothelial colony-forming cell angiogenic potential.

BACKGROUND AIMS: Circulating endothelial progenitor cells and especially endothelial colony-forming cells (ECFCs) are promising candidate cells for endothelial regenerative medicine of ischemic diseases, but the conditions for an optimal collection from adult blood must be improved. METHODS: On the basis of a recently reported vascular niche of ECFCs, we hypothesized that a local ischemia could trigger ECFC mobilization from the vascular wall into peripheral blood to optimize their collection for autologous implantation in critical leg ischemia. Because the target population with critical leg ischemia is composed of elderly patients in whom a vascular impairment has been documented, we also analyzed the impact of aging on ECFC mobilization and vascular integrity. RESULTS: After having defined optimized ECFC culture conditions, we studied the effect of forearm ischemia on ECFC numbers and functions in 26 healthy volunteers (13 volunteers ages 20-30-years old versus 13 volunteers ages 60-70 years old). The results show that forearm ischemia induced an efficient local ischemia and a normal endothelial response but did not mobilize ECFCs regardless of the age group. Moreover, we report an alteration of angiogenic properties of ECFCs obtained after forearm ischemia, in vitro as well as in vivo in a hindlimb ischemia murine model. This impaired ECFC angiogenic potential was not associated with a quantitative modification of the circulating endothelial compartment. CONCLUSIONS: The procedure of local ischemia, although reulting in a preserved endothelial reactivity, did not mobilize ECFCs but altered their angiogenic potential.

Immune response to human embryonic stem cell-derived cardiac progenitors and adipose-derived stromal cells.

Transplantation of allogeneic human embryonic stem cell-derived cardiac progenitors triggers an immune response. We assessed whether this response could be modulated by the concomitant use of adipose-derived stromal cells (ADSC). Peripheral blood mononuclear cells were collected from 40 patients with coronary artery disease (CAD) and nine healthy controls. Cardiac progenitors (CD15(+) Mesp1(+)) were generated as already reported from the I6 cell line treated with bone morphogenetic protein (BMP)-2. Adipose-derived stromal cells were obtained from abdominal dermolipectomies. We assessed the proliferative response of peripheral lymphocytes from patients and controls to cardiac progenitors cultured on a monolayer of ADSC, to allogeneic lymphocytes in mixed lymphocyte culture and to the T cell mitogen phytohemaglutin A in presence or absence of ADSC. Cardiac progenitors cultured on a monolayer of ADSC triggered a proliferation of lymphocytes from both patients and controls albeit lower than that induced by allogeneic lymphocytes. When cultured alone, ADSC did not induce any proliferation of allogeneic lymphocytes. When added to cultures of lymphocytes, ADSC significantly inhibited the alloantigen or mitogen-induced proliferative response. Compared to healthy controls, lymphocytes from patients presenting CAD expressed a decreased proliferative capacity, in particular to mitogen-induced stimulation. Adipose-derived stromal cells express an immunomodulatory effect that limits both alloantigen and mitogen-induced lymphocyte responses. Furthermore, lymphocytes from patients with CAD are low responders to conventional stimuli, possibly because of their age and disease-associated treatment regimens. We propose that, in combination, these factors may limit the in vivo immunogenicity of cardiac progenitors co-implanted with ADSC in patients with CAD.

Composite cell sheets: a further step toward safe and effective myocardial regeneration by cardiac progenitors derived from embryonic stem cells.

BACKGROUND: The safety and efficacy of myocardial regeneration using embryonic stem cells are limited by the risk of teratoma and the high rate of cell death. METHODS AND RESULTS: To address these issues, we developed a composite construct made of a sheet of adipose tissue-derived stroma cells and embryonic stem cell-derived cardiac progenitors. Ten Rhesus monkeys underwent a transient coronary artery occlusion followed, 2 weeks later, by the open-chest delivery of the composite cell sheet over the infarcted area or a sham operation. The sheet was made of adipose tissue-derived stroma cells grown from a biopsy of autologous adipose tissue and cultured onto temperature-responsive dishes. Allogeneic Rhesus embryonic stem cells were committed to a cardiac lineage and immunomagnetically sorted to yield SSEA-1(+) cardiac progenitors, which were then deposited onto the cell sheet. Cyclosporine was given for 2 months until the animals were euthanized. Preimplantation studies showed that the SSEA-1(+) progenitors expressed cardiac markers and had lost pluripotency. After 2 months, there was no teratoma in any of the 5 cell-treated monkeys. Analysis of >1500 histological sections showed that the SSEA-1(+) cardiac progenitors had differentiated into cardiomyocytes, as evidenced by immunofluorescence and real-time polymerase chain reaction. There were also a robust engraftment of autologous adipose tissue-derived stroma cells and increased angiogenesis compared with the sham animals. CONCLUSIONS: These data collected in a clinically relevant nonhuman primate model show that developmentally restricted SSEA-1(+) cardiac progenitors appear to be safe and highlight the benefit of the epicardial delivery of a construct harboring cells with a cardiomyogenic differentiation potential and cells providing them the necessary trophic support.

Exhaled nitric oxide and clinical phenotypes of childhood asthma.

Whether exhaled NO helps to identify a specific phenotype of asthmatic patients remains debated. Our aims were to evaluate whether exhaled NO (FENO(0.05)) is independently associated (1) with underlying pathophysiological characteristics of asthma such as airway tone (bronchodilator response) and airway inflammation (inhaled corticosteroid [ICS]-dependant inflammation), and (2) with clinical phenotypes of asthma.We performed multivariate (exhaled NO as dependent variable) and k-means cluster analyses in a population of 169 asthmatic children (age ± SD: 10.5 ± 2.6 years) recruited in a monocenter cohort that was characterized in a cross-sectional design using 28 parameters describing potentially different asthma domains: atopy, environment (tobacco), control, exacerbations, treatment (inhaled corticosteroid and long-acting bronchodilator agonist), and lung function (airway architecture and tone). Two subject-related characteristics (height and atopy) and two disease-related characteristics (bronchodilator response and ICS dose > 200 µg/d) explained 36% of exhaled NO variance. Nine domains were isolated using principal component analysis. Four clusters were further identified: cluster 1 (47%): boys, unexposed to tobacco, with well-controlled asthma; cluster 2 (26%): girls, unexposed to tobacco, with well-controlled asthma; cluster 3 (6%): girls or boys, unexposed to tobacco, with uncontrolled asthma associated with increased airway tone, and cluster 4 (21%): girls or boys, exposed to parental smoking, with small airway to lung size ratio and uncontrolled asthma. FENO(0.05) was not different in these four clusters.In conclusion, FENO(0.05) is independently linked to two pathophysiological characteristics of asthma (ICS-dependant inflammation and bronchomotor tone) but does not help to identify a clinically relevant phenotype of asthmatic children.

Circulating endothelial cells: a new candidate biomarker of irreversible pulmonary hypertension secondary to congenital heart disease.

BACKGROUND: Congenital heart disease can be complicated by pulmonary arterial hypertension (PAH), the reversibility of which is often difficult to predict. We recently reported a lung biopsy study showing impaired apoptotic regulation of endothelial cells in irreversible PAH. The objective of the present study was to identify noninvasive biomarkers of endothelial turnover that could be used to identify congenital heart disease patients at risk of irreversible PAH. METHODS AND RESULTS: Circulating endothelial cells (CECs) isolated with CD146-coated beads and circulating CD34(+)CD133(+) progenitor cells (CPCs) were quantified in peripheral vein, pulmonary artery, and pulmonary vein blood samples from 26 patients with congenital heart disease (16 with reversible PAH [median age 2 years] and 10 with irreversible PAH [median age 9 years]) and 5 control patients. Surgical lung biopsy was performed in 19 cases. As expected, endothelial remodeling was observed in irreversible PAH but not in reversible PAH. CEC and CPC numbers were each similar in the 3 types of blood samples. CEC numbers were significantly higher in patients with irreversible PAH (median 57 CEC/mL) than in patients with reversible PAH and control subjects (median 3 CEC/mL in the 2 groups). In contrast, CPC numbers did not differ among patients with irreversible or reversible PAH and control subjects (median 84, 64, and 44 CPC/10(5) lymphocytes, respectively, in the 3 groups). CONCLUSIONS: Irreversible PAH in congenital heart disease is associated with endothelial damage and with increased circulating endothelial cell counts. The present study suggests that CECs could be a valuable tool to define therapeutic strategies in congenital heart disease patients with PAH.

Thromboangiitis obliterans and endothelial function.

BACKGROUND: Endothelial dysfunction may be involved in the pathophysiology of thromboangiitis obliterans (TAO). This study compares endothelial function and large artery stiffness between 10 TAO patients assessed during an exacerbation phase and 10 age- and sex-matched control subjects. MATERIAL AND METHODS: Flow-mediated vasodilation after gradual hand skin heating (from 28 degrees C to 44 degrees C) and endothelium-independent vasodilation after sublingual administration of 150 microg glyceryl trinitrate (GTN) were studied using a high-resolution echotracking system to simultaneously measure the brachial artery (BA) diameter and changes in wall shear stress. Aortic stiffness was assessed by carotid-to-femoral pulse wave velocity. RESULTS: The baseline BA diameter was significantly smaller in TAO patients (3599 +/- 668 microm) than in control subjects (4114 +/- 671, P = 0.04). Hand warming caused a linear increase in shear stress accompanied by a linear increase in BA diameter as a function of increasing temperature for TAO patients and control subjects. There was no significant difference between the two groups [relative increase in BA diameter: + 9.3% (-0.1 to 11.5) vs. + 4.8% (3.0 to 8.1), respectively; P = 0.63]. The slope of the BA diameter vs. shear stress relationship did not significantly differ between the two groups. The relative increase in the BA diameter after GTN was significantly greater in TAO patients than in controls [+ 30.8% (28.6 to 33.6) vs. + 16.2% (12.6 to 21.9) respectively; P = 0.02]. Finally, TAO patients had greater aortic stiffness than control subjects (9.81 +/- 1.72 m s(-1) vs. 7.82 +/- 0.84 m s(-1) respectively; P = 0.0052). CONCLUSIONS: Acute TAO is characterised by vasoconstriction and increased aortic stiffness in the absence of changes in flow-mediated dilation.

Does the human skeletal muscle harbor the murine equivalents of cardiac precursor cells?

The limited plasticity of adult muscle- or bone marrow- derived stem cells intended for cardiac regeneration impedes their conversion into cardiomyocytes. Since murine skeletal muscle was reported to harbor cardiac precursor cells, we assessed whether similar cells exist in man. Skeletal muscle biopsies obtained from 39 patients were sorted by flow cytometry which generated three populations (CD90+/CD34(-), CD34+/CD90(-), CD90(-)/CD34(-)) expressing similar levels of cardiac (Nkx2.5, cTn-T, cTn-I, Cx43) and skeletal muscle (Myf-5, MyoD, myogenin) mRNAs, as assessed by quantitative reverse transcriptase-PCR. However, compared to unpurified myoblasts, CD34+/CD90(-) cells expressed greater amounts of endothelium-specific mRNAs and were, therefore, selected for transplantation experiments. Thirty immunosuppressed rats then underwent coronary artery ligation and, 4 weeks later, were intramyocardially injected with culture medium, myoblasts, or CD34+/CD90(-) cells. After 1 month, left ventricular ejection fraction was significantly higher in the CD34+/CD90(-) group than in the control and myoblast-injected hearts, which was associated with smaller fibrosis and greater angiogenesis. The low engraftment rate suggested a paracrine mechanism supported by the greater release of growth factors by CD34+/CD90(-) cells than by unsorted myoblasts. In conclusion, the human skeletal muscle does not harbor cardiac-specified cells but contains a CD34+ fraction endowed with an angiogenic potential providing superior functional and structural benefits.

Comparison of endothelial biomarkers according to reversibility of pulmonary hypertension secondary to congenital heart disease.

The reversibility of pulmonary arterial hypertension (PAH) in children with congenital heart disease (CHD) is strongly associated with the degree of intimal proliferation, vessel narrowing, and number of circulating endothelial cells (CECs). Circulating endothelial cells may arise from either endothelial damage or accelerated turnover during vessel remodeling, but nothing is known about endothelial microparticles (EMPs) and other biomarkers reflecting endothelial alterations. This study aimed to document endothelial markers further according to the irreversibility of PAH secondary to CHD. The study investigated soluble markers of endothelial damage or activation (thrombomodulin, soluble endothelial protein C receptor, and soluble E-selectin), inflammation (interleukin-6), and angiogenic cytokine levels [vascular endothelial growth factor (VEGF) and placental growth factor (PlGF)] in 26 patients with CHD, 16 with reversible PAH (median age, 2 years) and 10 with irreversible PAH (median age, 9 years). Endothelial activation/apoptosis was evaluated by measuring EMP levels. Plasma procoagulant activity also was measured. The results show that the levels of soluble markers indicating endothelial activation were not predictors of PAH irreversibility. Lower levels of PlGF were observed in reversible compared with irreversible PAH but were not associated with the CEC level, the mean pulmonary artery pressure (mPAP), or age. No significant difference in procoagulant activity or EMP level was found between irreversible and reversible PAH. Among a large panel of biomarkers reflecting endothelial activation, regeneration, and injury, the high CEC levels previously described proved to be the only marker allowing discrimination between reversible and irreversible PAH secondary to CHD.

A role for succinate dehydrogenase genes in low chemoresponsiveness to hypoxia?

The detection of hypoxia by the carotid bodies elicits a ventilatory response of utmost importance for tolerance to high altitude. Germline mutations in three genes encoding subunit B, C and D of succinate dehydrogenase (SDHB, SDHC and SDHD) have been associated with paragangliomas of the carotid body. We hypothesized that SDH dysfunction within the carotid body could result in low chemoresponsiveness and intolerance to high altitude. The frequency of polymorphisms of SDHs, hypoxia-inducible factor type 1 (HIF1alpha) and angiotensin converting enzyme (ACE) genes was compared between 40 subjects with intolerance to high altitude and a low hypoxic ventilatory response at exercise (HVRe < or = 0.5 ml min(-1) kg(-1); HVR- group) and 41 subjects without intolerance to high altitude and a high HVRe (> or = 0.80 ml min(-1) kg(-1); HVR+). We found no significant association between low or high HVRe and (1) the allele frequencies for nine single nucleotide polymorphisms (SNPs) in the SDHD and SDHB genes, (2) the ACE insertion/deletion polymorphism and (3) four SNPs in the HIF1alpha gene. However, a marginal significant association was found between the synonymous polymorphism c.18A>C of the SDHB gene and chemoresponsiveness: 8/40 (20%) in the HVR- group and 3/41 (7%) in the HVR+ group (p = 0.12). A principal component analysis showed that no subject carrying the 18C allele had both high ventilatory and cardiac response to hypoxia. In conclusion, no clear association was found between gene variants involved in oxygen sensing and chemoresponsiveness, although some mutations in the SDHB and SDHD genes deserve further investigations in a larger population.

Blood pressure outcome of adrenalectomy in patients with primary hyperaldosteronism with or without unilateral adenoma.

OBJECTIVE: To assess blood pressure outcome in patients with primary aldosteronism, who were operated on the basis of a unilateral adenoma detected by computed tomography or a lateralized aldosterone hypersecretion detected by adrenal venous sampling, and to analyze the hormonal and nonhormonal factors associated with the outcome. METHODS: A retrospective study of 168 patients with primary aldosteronism undergoing surgery: 109 patients with a unilateral adenoma detected by computed tomography and 59 without a unilateral adenoma who underwent surgery because of an aldosterone to cortisol ratio at least five times higher on the dominant side than on the nondominant side. RESULTS: Patients with a unilateral adenoma were more likely to be women, had a shorter history of hypertension and had lower blood pressure levels and treatment scores than patients without a unilateral adenoma. The mean systolic blood pressures of patients with and without unilateral adenomas at follow-up were 133 +/- 16 and 137 +/- 16 mmHg, respectively. Hypertension cure or improvement was observed in 77% (95% confidence interval 69-85%) and 68% (95% confidence interval 56-80%) of patients, respectively. Using a linear regression model, baseline urinary aldosterone was positively associated, and baseline serum potassium was negatively associated, with decrease in systolic blood pressure. CONCLUSION: Adrenalectomy improves blood pressure control in patients with primary aldosteronism operated on the basis of either unilateral adenoma detected by computed tomography or a lateralized aldosterone hypersecretion. A high urinary aldosterone excretion and a low serum potassium level predict a more favorable outcome of surgery.

Genetic and dietary control of plasma tissue kallikrein secretion and urinary kinins exretion in man.

OBJECTIVES: Tissue kallikrein is a major kinin-forming enzyme involved in artery and kidney functions. Urinary tissue kallikrein activity (UKLKa) reflects renal tissue kallikrein activity and depends on Na and K intake, and genetic factors, especially the tissue kallikrein-inactivating polymorphism, R53H. The effect of these factors on the level of kinin peptides is, however, not known. Moreover, a circulating form of tissue kallikrein is present in human plasma but its origin and regulation are unknown. METHODS: We used a crossover study to investigate UKLKa, plasma tissue kallikrein (pTK), and urinary kallidin peptides and metabolites (Uki) in 10 R53H and 30 R53R normotensive male subjects randomly assigned to either a 1-week low Na-high K or a high Na-low K diet. RESULTS: UKLKa was 50-60% lower in R53H than R53R subjects and was increased by the low Na-high K diet. pTK was also 45-55% lower in R53H than R53R subjects and was increased by the low Na-high K diet. Uki was slightly but significantly higher under the low Na-high K than the high Na-low K diet, but did not differ between genotypes. CONCLUSION: These observations indicate that pTK levels are genetically determined and regulated by Na and K diet, in parallel with UKLKa; this suggests that circulating tissue kallikrein originates mainly from the kidney, and can contribute to circulatory adaptation to dietary ions. Uki is influenced by the Na and K diet, suggesting that kinins participate in renal adaptation to ion intake, but do not quantitatively reflect tissue kallikrein activity in urine, or presumably, in the kidney.

Effect of hydrochlorothiazide on urinary calcium excretion in dent disease: an uncontrolled trial.

BACKGROUND: Thiazide doses equivalent to 1 to 2 mg/kg/d of hydrochlorothiazide (HCTZ) have been proposed to correct hypercalciuria and prevent kidney failure in patients with Dent disease. However, they can cause adverse metabolic effects in the long term. In treating hypertension in children, lower thiazide doses have been shown to be as effective and well tolerated. STUDY DESIGN: Uncontrolled trial, with forced-titration sequential open-label study design. SETTING & PARTICIPANTS: 7 boys with genetically confirmed Dent disease and mild phenotype (neither overt sodium wasting nor kidney failure). INTERVENTION: After a 1-month run-in period, patients sequentially received amiloride (5 mg/d) alone (1 month) and then for 3 periods of 2 months in association with increasing doses of HCTZ (<0.2, 0.2 to 0.4, and 0.4 to 0.8 mg/kg/d). OUTCOMES: Urinary calcium excretion and extracellular volume indicators. MEASUREMENTS: At the end of each period, 2 daily 24-hour urinary collections were performed on the days preceding admission. Blood and spot urine samples also were collected. RESULTS: A greater HCTZ dose increased renin, aldosterone, and plasma protein concentrations. Amiloride alone had no effect on calcium excretion. The greatest HCTZ doses decreased spot urinary calcium excretion by 42% compared with baseline (median, 0.3; minimum, maximum, 0.2, 0.8 versus median, 0.8; minimum, maximum, 0.4, 1.1, respectively; P = 0.03). However, patients developed adverse reactions, including muscle cramps (n = 2), biological (n = 7) or symptomatic hypovolemia (n = 1), hypokalemia (n = 4), and hyponatremia (n = 1), which all corrected after treatment withdrawal. LIMITATION: Small sample size and absence of a control group. CONCLUSION: HCTZ doses greater than 0.4 mg/kg/d decreased calcium excretion, but were associated with significant adverse events. Thiazide diuretic therapy should be considered with caution in children with Dent disease.

Skeletal myoblast transplantation in ischemic heart failure: long-term follow-up of the first phase I cohort of patients.

BACKGROUND: Skeletal myoblast (SM) transplantation (Tx) in a post-myocardial infarction (MI) scar experimentally improves left ventricular (LV) ejection fraction (EF). Short-term follow-up (FU) studies have suggested that a similar benefit could clinically occur despite an increased risk of LV arrhythmias. METHODS AND RESULTS: We report the long-term FU of the first worldwide cohort of grafted patients (n = 9, 61.8+/-11.6 years, previous MI, EF < or = 35%) operated on (autologous SM Tx and bypass surgery) in 2000 to 2001 and evaluated before Tx, at 1 month (M1) and at a median FU of 52 (18 to 58) months after Tx (37 patient-years). NYHA class improved from 2.5+/-0.5 to 1.8+/-0.4 at M1 (P=0.004 versus baseline) and 1.7+/-0.5 at FU (P=not significant versus M1; P=0.0007 versus baseline). EF increased from 24.3+/-4% to 31+/-4.1% at M1 (+28%, P=0.001 versus baseline) and remained stable thereafter (28.7+/-8.1%, +18% versus baseline). There were 5 hospitalizations for heart failure in 3 patients at 28.6+/-9.9 months, allowing implant in 2 patients with a resynchronization pacemaker. An automatic cardiac defibrillator (ACD) was implanted in 5 patients for nonsustained (n =1) or sustained (n =4) ventricular tachycardia at 12.2+/-18.6 (1 to 45) months. Despite a beta-blocker/amiodarone combination therapy, there were 14 appropriate shocks for 3 arrhythmic storms in 3 patients at 6, 7, and 18 months after ACD implantation. CONCLUSIONS: In this cohort of severe heart failure patients both clinical status and EF stably improve over time with a strikingly low incidence of hospitalizations for heart failure (0.13/patient-years) and the arrhythmic risk can be controlled by medical therapy and/or on-request ACD implantation.

Succinate dehydrogenase B gene mutations predict survival in patients with malignant pheochromocytomas or paragangliomas.

CONTEXT: Pheochromocytomas and paragangliomas may be malignant either at presentation or during recurrence, but the clinical course of malignant tumors is unpredictable. OBJECTIVE: The objective was to analyze survival according to clinical characteristics at diagnosis of malignancy and the presence or absence of SDHB mutations. DESIGN: This was a retrospective cohort study. SETTING AND PARTICIPANTS: A total of 54 patients with malignant tumors were included. Malignancy was scored according to the presence of metastases or histologically documented lymph node invasion. MAIN OUTCOME MEASURES: The main outcome was the specific survival after the diagnosis of the first metastasis. RESULTS: Germline mutations were identified in SDHB (n = 23, including 21 patients with apparent sporadic tumors) and VHL (n = 1) genes, and two patients had neurofibromatosis 1. Patients were followed up from the diagnosis of primary tumor and from the diagnosis of the first metastasis to the present or to death with medians of 79 [interquartile range (IQR) 24; 190] and 39 [IQR 14; 94] months, respectively. The 5-yr probability of survival after the diagnosis of the first metastasis was 0.55 (95% confidence interval 0.39-0.69). Patients with SDHB mutations were younger, more frequently had extra-adrenal tumors, and had a shorter metanephrine excretion doubling time. The presence of SDHB mutations was significantly and independently associated with mortality (relative risk 2.7; 95% confidence interval 1.2, 6.4; P = 0.021). CONCLUSION: SDHB mutations, frequent in patients with malignant pheochromocytomas or paragangliomas, are associated with shorter survival. Therefore, SDHB genetic testing may be of prognostic value for such patients, even those with an apparent sporadic and/or benign presentation at diagnosis.

Is xenotransplantation of embryonic stem cells a realistic option?

To test the purported immune privilege of embryonic stem cells (ESC) in the challenging setting of xenotransplantation, 14 immunocompetent baboons were subjected to a coronary artery occlusion-reperfusion sequence and, two weeks later, randomized to receive in-scar injections of culture medium or cardiac-committed mouse ESC engineered to express fluorescent reporter genes driven by cardiac-specific promoters. Two months after transplantation, left ventricular function, as assessed by echocardiography, deteriorated to a similar extent in control and treated baboons. This correlated with failure to identify the grafted cells by X-gal histology and immunofluorescence. Rejection did not seem to be mediated by xenoantibodies, but rather by T lymphocytes and natural killer cells as suggested by positive immunostaining for CD3 and CD56 early after transplantation. There was no increase in circulating levels of regulatory T cells. These data raise a cautionary note about the immune privilege of ESC and suggest that from a mere immunologic standpoint, ESC xenotransplantation is likely to be an unrealistic challenge.

Prognostic value of tumor-infiltrating CD4+ T-cell subpopulations in head and neck cancers.

PURPOSE: CD4(+) T cells play a central role in initiating and maintaining anticancer immune responses. However, regulatory CD4(+)CD25(+) T cells which express Foxp3 have also been shown to inhibit antitumor effector T cells. In view of these heterogeneous CD4(+) T-cell populations, this study was designed to determine the prognostic value of various tumor-infiltrating CD4(+) T-cell populations in head and neck squamous cell carcinoma. EXPERIMENTAL DESIGN: Eighty-four newly diagnosed untreated patients with histologically proven primary head and neck squamous cell carcinoma were included in this study. Double or triple immunofluorescence staining was done to assess and quantify the activated CD4(+)CD69(+) T cells, regulatory CD4(+)Foxp3(+) T cells, and mixed CD4(+)CD25(+) T cells comprising both activated and regulatory T cells. RESULTS: On univariate analysis, high levels of tumor-infiltrating CD4(+)CD69(+) T cells were correlated with both better locoregional control (P = 0.01) and longer survival (P = 0.01). Infiltration by regulatory Foxp3(+)CD4(+) T cells was positively associated with a better locoregional control of the tumor. Multivariate analysis showed that the only significant prognostic factors related to locoregional control were T stage (P = 0.02) and CD4(+)Foxp3(+) T-cell infiltration of the tumor (P = 0.02). In the Cox multivariate analysis, only two variables influenced overall survival probability: T stage (P = 0.036) and CD4(+)CD69(+) T-cell infiltration (P = 0.017). CONCLUSION: This study shows that tumor-infiltrating activated CD4(+)CD69(+) T cells are associated with a good prognosis in head and neck squamous cell carcinoma. In addition, regulatory Foxp3(+)CD4(+) T cells are positively correlated with locoregional control may be through down-regulation of harmful inflammatory reaction, which could favor tumor progression.