Lyotropic Liquid-Crystalline Nanosystems as Drug Delivery Agents for 5-Fluorouracil: Structure and Cytotoxicity.

Lyotropic cubic liquid-crystalline systems have received increasing attention due to their unique microstructural and physicochemical properties as efficient nanocarriers for drug delivery. We report the preparation and characterization of bulk phases and cubosome dispersions of phytantriol loaded with the anticancer drug 5-fluorouracil, in neutral and anionic forms. In both cases, a Pn3m cubic phase was observed. The phytantriol phase behavior can be influenced by the addition of ionic agents, and, to this purpose, a positively charged lipid, such as N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride salt (DOTAP), was included in the studied formulations. It was found to induce a variation of the spontaneous membrane curvature of the phytantriol lipid bilayer, generating a transition from the Pn3m to the Im3m cubic phase. When 5-fluorouracil, in its anionic form (5-FUs), was encapsulated in these latter systems, a further transition to the HII hexagonal phase was observed as a consequence of the formation of a complex phytantriol/DOTAP/5-FUs. The physicochemical characterization was performed with various complementary techniques including synchrotron small-angle X-ray scattering, dynamic light scattering, and attenuated total reflection Fourier transform infrared and UV resonance Raman spectroscopies. Encapsulation of 5-fluorouracil in the corresponding nanodispersions was evaluated, and their in vitro cytotoxicity was assessed in MDA-MB-231 cell line. Phytantriol cubosomes containing 5-fluorouracil showed a higher toxicity compared with the bare drug solution, and hence they represent potential nanocarriers in the delivery of 5-fluorouracil for cancer therapy.

A chaperone enhances blood α-glucosidase activity in Pompe disease patients treated with enzyme replacement therapy.

Enzyme replacement therapy is currently the only approved treatment for Pompe disease, due to acid α-glucosidase deficiency. Clinical efficacy of this approach is variable, and more effective therapies are needed. We showed in preclinical studies that chaperones stabilize the recombinant enzyme used for enzyme replacement therapy. Here, we evaluated the effects of a combination of enzyme therapy and a chaperone on α-glucosidase activity in Pompe disease patients. α-Glucosidase activity was analyzed by tandem-mass spectrometry in dried blood spots from patients treated with enzyme replacement therapy, either alone or in combination with the chaperone N-butyldeoxynojirimycin given at the time of the enzyme infusion. Thirteen patients with different presentations (3 infantile-onset, 10 late-onset) were enrolled. In 11 patients, the combination treatment resulted in α-glucosidase activities greater than 1.85-fold the activities with enzyme replacement therapy alone. In the whole patient population, α-glucosidase activity was significantly increased at 12 hours (2.19-fold, P = 0.002), 24 hours (6.07-fold, P = 0.001), and 36 hours (3.95-fold, P = 0.003). The areas under the curve were also significantly increased (6.78-fold, P = 0.002). These results suggest improved stability of recombinant α-glucosidase in blood in the presence of the chaperone.

Severe spinal deformity and multiple vertebral collapses in juvenile Cushing Syndrome: a case report.

Cushing's Syndrome (CS) is rare in adolescence but the pathological effects of excessive circulating glucocorticoids concentration on bone during the developmental age still represent a challenge for orthopedists. Only few reports describe the gravity of early developed damages of spine in young affected by CS. A 18-years-old woman suffering from Cushing's Disease presented after many years treatment of the primary disease referring severe back pain and worsening back deformity. Radiological investigations showed vertebral collapses a devastating thoraco-lumbar scoliosis of 80° Cobb. Lumbar dual X-ray absorptiometry Z-score values were very low and consistent with severe osteoporosis. The patient was treated with bracing, antiresorptive therapy, calcium and vitamin D supplementation, and followed-up with imaging investigations to screen for further fractures. The bone mineral density will be monitored until its normalization will allow to plane surgical treatment in case of progression of spinal deformity and collapses. Early diagnosis and treatment of hypercortisolism, periodical clinical and radiographic follow-up, and treatment for the bone damage are mandatory to prevent the devastating sequelae of secondary osteoporosis.

The pharmacological chaperone N-butyldeoxynojirimycin enhances enzyme replacement therapy in Pompe disease fibroblasts.

In spite of the progress in the treatment of lysosomal storage diseases (LSDs), in some of these disorders the available therapies show limited efficacy and a need exists to identify novel therapeutic strategies. We studied the combination of enzyme replacement and enzyme enhancement by pharmacological chaperones in Pompe disease (PD), a metabolic myopathy caused by the deficiency of the lysosomal acid alpha-glucosidase. We showed that coincubation of Pompe fibroblasts with recombinant human alpha-glucosidase and the chaperone N-butyldeoxynojirimycin (NB-DNJ) resulted in more efficient correction of enzyme activity. The chaperone improved alpha-glucosidase delivery to lysosomes, enhanced enzyme maturation, and increased enzyme stability. Improved enzyme correction was also found in vivo in a mouse model of PD treated with coadministration of single infusions of recombinant human alpha-glucosidase and oral NB-DNJ. The enhancing effect of chaperones on recombinant enzymes was also observed in fibroblasts from another lysosomal disease, Fabry disease, treated with recombinant alpha-galactosidase A and the specific chaperone 1-deoxygalactonojirimycin (DGJ). These results have important clinical implications, as they demonstrate synergy between pharmacological chaperones and enzyme replacement. A synergistic effect of these treatments may result particularly useful in patients responding poorly to therapy and in tissues in which sufficient enzyme levels are difficult to obtain.

Homocysteinemia correlates with plasma thiol redox status in patients with end-stage renal disease.

BACKGROUND/AIMS: In end-stage renal disease (ESRD), hyperhomocysteinemia is a common finding associated with increased cardiovascular risk. However, the pathogenic role of homocysteine is still unclear. In vitro studies show that thiol redox status affects endothelial cell functions. We therefore investigated the possible association between homocysteinemia and plasma thiol redox status in ESRD patients. METHODS: Total plasma homocysteine (Hcy), cysteine (Cys) and free thiols (SH) were measured both before and after a dialytic session in 54 ESRD patients receiving (n = 15) or not receiving (n = 39) folate supplementation, and 17 control subjects. RESULTS: High predialysis levels of both Hcy and Cys were found to be negatively correlated with low SH levels both in supplemented (r = -0.680, p < 0.01 and r = -0.624, p < 0.02, respectively) and unsupplemented (r = -0.698, p < 0.001 and r = -0.445, p < 0.01, respectively) patients. Following dialysis, SH values returned to normal and the above correlations were no longer appreciable. CONCLUSION: A strong, folate therapy-insensitive association between homocysteinemia and plasma free thiol levels was found in ESRD patients. These results support a role for oxidative stress in ESRD-related hyperhomocysteinemia and suggest the plasma thiol redox status alteration as a possible pathogenic mechanism underlying the cardiovascular toxicity of hyperhomocysteinemia in these patients.