Glycosidic intermediates identified in 1H MR spectra of intact tumour cells may contribute to the clarification of aspects of glycosylation pathways

The glycosylation process, through the addition of carbohydrates, is a major post‐translational modification of proteins and glycolipids. Proteins may be glycosylated in either the secretory pathway leading to N‐linked or O‐linked glycoproteins or as nucleocytoplasmic glycosylation that targets only single proteins involving a single β‐linked N‐acetylglucosamine. In both cases, the key precursors are the uridine diphospho‐N‐acetylhexosamines synthesised by the hexosamine biosynthetic pathway. Furthermore, uridine diphospho‐N‐acetylglucosamine participates in the biosynthesis of sialic acid. In this work, we propose MRS for the detection of uridine diphospho‐N‐acetylhexosamines visible in high‐resolution MR spectra of intact cells from different human tumours. Signals from the nucleotide and amino sugar moieties, including amide signals observed for the first time in whole cells, are assigned, also taking advantage of spectral changes that follow cell treatment with ammonium chloride. Finally, parallel changes in uridine diphospho‐N‐acetylhexosamines and glutamine pools, observed after pH changes induced by ammonium chloride in the different tumour cell lines, may provide more details on the glycosylation processes. Copyright © 2010 John Wiley & Sons, Ltd.     

Genomic damage induced by 1‐MHz ultrasound in vitro

Genotoxic effects of therapeutic ultrasound are poorly documented, when compared with the wide use of this physical agent. The aim of this work was to investigate the clastogenic and aneugenic potential of 1 MHz ultrasound, employing intensities (200 and 300 mW/cm²) above the cavitational threshold, but in the range of those normally used in therapeutics. Both normal fibroblasts (AG01522) and tumoral cells (MCF‐7) were sonicated. While no effects on viability were noted, significant increases of CREST‐negative micronuclei (indicative of clastogenesis) and CREST‐positive micronuclei (indicative of aneuploidy) were detected. Clastogenesis was confirmed by increases of γ‐H2AX foci, while increases of spindle anomalies confirmed the induction of aneuploidy. Our results confirm previous works that showed ultrasound‐induced DNA breakage. Moreover, our experiments show that the known effect of ultrasound‐induced damage to microtubules is also able to damage the mitotic spindle and induce aneuploidy. On the overall, this work highlights the importance to further investigate the potential risks related to therapeutics US. Environ. Mol. Mutagen. 59:60–68, 2018. © 2017 Wiley Periodicals, Inc.     

Outcome prediction by immunophenotypic minimal residual disease detection in adult T-cell acute lymphoblastic leukaemia

Flow-cytometric detection of minimal residual disease (MRD) identifies patients with high relapse risk in childhood acute lymphoblastic leukaemia (ALL). We studied the efficacy of this method in adult T-ALL treated with the Italian co-operative GIMEMA (Gruppo Italiano Malattie Ematologiche dell'Adulto) LAL0496 protocol. Bone marrow samples from 53 patients were taken at fixed treatment time points and MRD was analysed using a leukaemia-specific immunophenotype (cytoplasmic-CD3/nuclear-terminal desoxynucleotidyl transferase). The median follow-up was 17 months (range 3-61) and a median of 4.5 analyses/patient was performed (range 3-12). Six out of 53 (11.3%) patients were refractory to treatment, 30/53 (56.6%) relapsed and 17/53 (32.1%) remain in continuous complete remission. The probability of relapse at 2 years for MRD-positive patients at preconsolidation was 81.5%vs 38.9% for MRD-negative patients (P = 0.00078). This risk was still 54.5% for MRD-positive vs 15.8% for MRD-negative patients pre-third reinduction (P = 0.0098) and 50.0% for MRD-positive vs 16.4% for MRD-negative patients pre-sixth reinduction (P = 0.032). The relapse-predicting value of MRD did not depend on features at diagnosis such as age, sex and leucocyte count. Our data suggest that immunophenotypic MRD monitoring in the first year of treatment is a useful outcome predictor for adult T-ALL patients.     

Radiofrequency ablation of hepatic tissue: a new experimental animal model

BACKGROUND/AIMS: Experimental radiofrequency ablation has already been performed in healthy livers of porcine models, but not in less expensive and easy-to-manage rats, with devices capable of delivering radiofrequency ablation in the 20-30 g liver of such small animals being so far unavailable. METHODOLOGY: We experimented with a modified system of radiofrequency ablation of liver tissue in rat models developing a custom-made needle-microelectrode of very small dimensions (0.3x2 mm) and an electrode-tip cooling technique, based on saline solution infusion. We adjusted duration (seconds) and power (watts) of radiofrequency ablation letting them range between 5-50 seconds and 5-25 W, respectively, to obtain the greatest lesions with the least side effects. After sacrificing the animals, an accurate histological examination of the liver was made. RESULTS: It is possible to establish beforehand the diameter of thermal liver lesion on the basis of joules of applied energy. The greatest increase of liver thermal lesion diameter (8 mm) is obtained with a 250-joule (10 W for 25 seconds) thermal energy cooling the electrode-tissue interface. CONCLUSIONS: Experimental radiofrequency ablation in rat liver is an effective and cheap way to study its effects on healthy hepatic tissues. It might be the first step to treat experimentally caused liver tumors.     

Begelomab for severe refractory dermatomyositis: A case report

Rationale:Severe refractory idiopathic inflammatory myopathy (IIM) represents a challenge for the clinician. The lack of efficacy of available tools reflects our incomplete insight into the molecular events sustaining the inflammatory tissue damage in these patients. We present the first case of refractory IIM treated with anti-dipeptidyl peptidase-4 (DPP-4)/cluster of differentiation 26 (CD26) monoclonal antibody.Patient concerns:A 55-year old man presented with proximal muscle weakness, diffuse erythematous skin lesions which rapidly evolved into ulcerations, dysphagia and dysphonia.Diagnosis:Increased serum creatine kinase levels and histological findings at muscle and skin biopsies were compatible with the diagnosis of dermatomyositis (DM). Several lines of treatment failed to control the disease including steroids, mycophenolate mofetil, tacrolimus, intravenous immunoglobulins and rituximab. Despite therapy, the patient also had recurrent intestinal vasculitis causing bowel perforation. Concurrently, DPP-4/CD26 expression in the patient''s skin and skeletal muscle was observed.Interventions:The patient was treated with begelomab, a murine immunoglobulin G2b monoclonal antibody against DPP-4/CD26.Outcomes:Dysphagia, skin lesions and intestinal vasculitis resolved and the patient experienced a significant improvement of his quality of life.Conclusion:Blockade of DPP-4/CD26, which is expressed on T cells and mediates T cell activation and function, is safe and might be effective in patients with refractory DM.     

Thyroglobulin measurement before rhTSH-aided 131I ablation in detecting metastases from differentiated thyroid carcinoma

Aim Thyroidectomy followed by administration of large activities of 131‐iodine (¹³¹I) is the treatment of choice for differentiated thyroid carcinoma (DTC). The serum thyroglobulin (Tg) measurement during hypothyroidism (offT4‐Tg), just before radioiodine thyroid ablation, has proved to be effective for predicting persistent/recurrent disease. However, the Tg measurement cannot be used as a corresponding value for pre‐ablative offT4‐Tg when recombinant human TSH (rhTSH) is used as stimulus before treatment. The present study was undertaken to evaluate if post‐thyroidectomy Tg values, measured before rhTSH‐stimulated radioiodine ablation is of prognostic value in patients affected by DTC. Methods We enrolled 126 patients with DTC submitted to total thyroidectomy. T4 treatment was started just after surgery to suppress TSH levels and Tg levels (onT4‐Tg) were measured just before rhTSH‐aided thyroid ablation by ¹³¹I (3700 MBq). Neck radioiodine uptake (RAIU) was measured just before ablation and a post‐treatment whole body scan (PT‐WBS) was performed. Results A significant relationship was found between thyroid remnants’ RAIU and onT4‐Tg levels (P < 0·001). The 1·10 ng/ml onT4‐Tg threshold selected by ROC curve analysis identifies patients with positive PT‐WBS with 83·3% sensitivity, 65·7% specificity, 44·5% positive predictive value (PPV) and 93·6% negative predictive value (NPV). The 0·65 ng/ml cut‐off level recognizes metastatic patients with 82·9% sensitivity, 55·2% specificity, 43·3% PPV and 97·8% NPV when compared with 12 months restaging results. Among 63 patients with initially undetectable onT4‐Tg (i.e. ≤ 0·2 ng/ml) none had positive PT‐WBS nor DTC relapse at 12‐month restaging (NPV 100%). Conclusions Based on our data we conclude that pre‐ablative onT4‐Tg is a prognostic marker and should be used instead of pre‐ablative TSH‐stimulated Tg measurement when rhTSH‐aided radioiodine ablation is done.