Drug Repurposing in Rare Diseases: An Integrative Study of Drug Screening and Transcriptomic Analysis in Nephropathic Cystinosis.

Diagnosis and cure for rare diseases represent a great challenge for the scientific community who often comes up against the complexity and heterogeneity of clinical picture associated to a high cost and time-consuming drug development processes. Here we show a drug repurposing strategy applied to nephropathic cystinosis, a rare inherited disorder belonging to the lysosomal storage diseases. This approach consists in combining mechanism-based and cell-based screenings, coupled with an affordable computational analysis, which could result very useful to predict therapeutic responses at both molecular and system levels. Then, we identified potential drugs and metabolic pathways relevant for the pathophysiology of nephropathic cystinosis by comparing gene-expression signature of drugs that share common mechanisms of action or that involve similar pathways with the disease gene-expression signature achieved with RNA-seq.

Influence of dialysis techniques and alternate vitamin supplementation on homocysteine levels in patients with known MTHFR genotypes.

BACKGROUND: Treatment with folic acid and vitamin B12 appears capable of reducing total plasma homocysteine levels (tHcy), but it is unknown whether vitamin B12 alone reduces tHcy values. In this study we investigate the effects of alternate vitamins supplementation on homocysteine levels in patients treated by diffusive and convective dialysis techniques. METHODS: 74 patients were randomized blindly into two groups of 37 subjects each. The first group was treated initially with vitamin B12 for 2 months and with folic acid for the following 2 months. The second group was treated initially with folic acid. A wash out period of 2 months followed the treatment in both groups. RESULTS: Total homocysteine levels decreased in both groups following the alternate vitamins therapy and dialysis, without significant difference between diffusive and convective techniques. Surprisingly, after the wash-out period, tHcy increased remarkably, regardless of the dialysis procedure used. At the end of the study, folate levels showed a higher reduction with haemodialysis compared to haemodiafiltration. In contrast, vitamin B12 levels showed a significant increase using diffusive haemodialysis, confirming a decisive role of membrane performance. CONCLUSIONS: In conclusion we show for the first time that, even if total homocysteine levels decreased in both dialysis procedures, the convective techniques demonstrate a superior capacity on the reduction of tHcy levels compared to the diffusive method. Moreover, the lower depletion of vitamin B12 by diffusive techniques could determine a higher reduction of folate levels, demonstrating the decisive role of the membrane performance in the treatment of this patients.

Homocysteine, cysteine, folate and vitamin B12 status in type 2 diabetic patients with chronic kidney disease.

BACKGROUND: Hyperhomocysteinemia (hHcy) is a risk factor in the progression of chronic kidney disease (CKD). In type 2 diabetes (T2D), hHcy is strongly associated with increased risk of cardiovascular disease. Vitamin B12 and folic acid supplementation have been reported to lower homocysteine (tHcy) levels, but no data on plasma tHcy, cysteine (Cys), folate and vitamin B12 levels in T2D-CKD patients are reported. PROCEDURES: tHcy and Cys levels were analyzed in 178 T2D-CKD patients by high performance liquid chromatography (HPLC) with fluorescence detection. In addition, we determined folate and vitamin B12 levels using a chemiluminescence method. RESULTS: tHcy and Cys levels were increased in T2D patients, and this rise positively correlated with the CKD stage (P < 0.001). Folate levels were comparable to controls at various CKD stages, whereas vitamin B12 levels were lower, except at stage IV. We did not find any correlation between B-vitamins and levels of tHcy and Cys, regardless of the CKD stage. CONCLUSIONS: This is the first study reporting tHcy, Cys and B-vitamins status in T2D-CKD patients. Although limited to our cohort of 178 patients, our findings could be helpful in clarifying the conflicting literature regarding B-vitamins supplementation. Further studies are necessary before any Hcy-lowering therapy can be safely established in T2D-CKD subjects.

Pediatric reference intervals for muscle coenzyme Q(10).

Coenzyme Q(10) (CoQ(10)) is present in humans in both the reduced (ubiquinol, CoQ(10)H(2)) and oxidized (ubiquinone, CoQ(10)) forms. CoQ(10) is an essential cofactor in mitochondrial oxidative phosphorylation, and is necessary for ATP production. Total, reduced and oxidized CoQ(10) levels in skeletal muscle of 148 children were determined by HPLC coupled with electrochemical detection, and we established three level thresholds for total CoQ(10) in muscle. We defined as "severe deficiency", CoQ(10) levels falling in the range between 0.82 and 4.88 µmol/g tissue; as "intermediate deficiency", those ranging between 5.40 and 9.80 µmol/g tissue, and as "mild deficiency", the amount of CoQ(10) included between 10.21 and 19.10 µmol/g tissue. Early identification of CoQ(10) deficiency has important implications in children, not only for those with primary CoQ(10) defect, but also for patients with neurodegenerative disorders, in order to encourage earlier supplementation with this agent also in mild and intermediate deficiency.

Effects of folic acid before and after vitamin B12 on plasma homocysteine concentrations in hemodialysis patients with known MTHFR genotypes.

BACKGROUND: Treatment with folic acid and vitamin B12 appears to be effective in lowering total plasma homocysteine (tHcy) concentrations, but whether vitamin B12 alone lowers tHcy in patients with normal vitamin B12 status is unknown. The aims of the present study were to explore the effect of individual supplementation with folic acid or vitamin B12 on tHcy concentrations in hemodialysis (HD) patients and to compare changes in tHcy concentrations with MTHFR genotype. METHODS: We recruited 200 HD patients (119 men) from the "Umberto I" Hospital (Frosinone, Italy) and the Dialysis Unit of University Hospital "Tor Vergata". These patients were randomized blindly into 2 groups of 100 each. Unfortunately, during the study, 36 patients in the first group and 16 in the second group died. The first group was treated initially with vitamin B12 for 2 months and with folic acid for a following 2 months. The second group was treated initially with folic acid and then with vitamin B12. Samples were drawn before administration of either, after the first and second periods, and again 2 months after treatment. RESULTS: The concentrations of tHcy decreased in both groups after the consecutive vitamin therapies, and the decrease was genotype-dependent. The decrease was greater for the T/T genotype (P <0.05) and was more significant when the treatment was started with folic acid (P <0.01). CONCLUSION: The alternating vitamin treatment demonstrated for the first time the importance of folate therapy and the secondary contribution of vitamin B12 in lowering tHcy in HD patients.