Phenotypic Characterization of Autoreactive B Cells—Checkpoints of B Cell Tolerance in Patients with Systemic Lupus Erythematosus

DNA-reactive B cells play a central role in systemic lupus erythematosus (SLE); DNA antibodies precede clinical disease and in established disease correlate with renal inflammation and contribute to dendritic cell activation and high levels of type 1 interferon. A number of central and peripheral B cell tolerance mechanisms designed to control the survival, differentiation and activation of autoreactive B cells are thought to be disturbed in patients with SLE. The characterization of DNA-reactive B cells has, however, been limited by their low frequency in peripheral blood. Using a tetrameric configuration of a peptide mimetope of DNA bound by pathogenic anti-DNA antibodies, we can identify B cells producing potentially pathogenic DNA-reactive antibodies. We, therefore, characterized the maturation and differentiation states of peptide, (ds) double stranded DNA cross-reactive B cells in the peripheral blood of lupus patients and correlated these with clinical disease activity. Flow cytometric analysis demonstrated a significantly higher frequency of tetramer-binding B cells in SLE patients compared to healthy controls. We demonstrated the existence of a novel tolerance checkpoint at the transition of antigen-naïve to antigen-experienced. We further demonstrate that patients with moderately active disease have more autoreactive B cells in both the antigen-naïve and antigen-experienced compartments consistent with greater impairment in B cell tolerance in both early and late checkpoints in these patients than in patients with quiescent disease. This methodology enables us to gain insight into the development and fate of DNA-reactive B cells in individual patients with SLE and paves the way ultimately to permit better and more customized therapies

A phase II study of the vitamin D analogue Seocalcitol in patients with inoperable hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is a common malignant tumour, which has a poor prognosis. Surgical resection can be curative but most patients are inoperable and most chemotherapy agents have minimal activity in this disease. Seocalcitol, a vitamin D analogue, induces differentiation and inhibits growth in cancer cell lines and in vivo. The vitamin D receptor is expressed in hepatocytes and more abundantly in HCC cells. In total, 56 patients with inoperable advanced HCC were included in an uncontrolled study of oral Seocalcitol treatment for up to 1 year ( with possible extension for responders). The dose was titrated according to serum calcium levels. The treatment effect was evaluated by regular CT scans. Out of 33 patients evaluable for tumour response, two had complete response (CR), 12 stable disease and 19 progressive disease. The CRs appeared after 6 and 24 months of treatment, and lasted for 29 and at least 36 months ( patient still in remission when data censored). Seocalcitol was well tolerated; the most frequent toxicity was hypercalcaemia and related symptoms. Most patients tolerated a daily dose of 10 mug of Seocalcitol. This is the first study showing activity, by reduction in tumour dimensions, of a differentiating agent in patients with an advanced bulky, solid tumour. Seocalcitol may have an effect in the treatment of HCC, especially in early disease when a prolonged treatment can be instituted. The survival benefit with or without tumour response should be determined in controlled studies

The relationship between vitamin D status and muscle strength in young healthy adults from sunny climate countries currently living in the northeast of Scotland

Summary: The current study examined the relationship between vitamin D status and muscle strength in young healthy adults: residents (>6 months) and newcomers (0–3 months), originally from sunny climate countries but currently living in the northeast of Scotland. Our longitudinal data found a positive, albeit small, relationship between vitamin D status and knee extensor isometric strength.  Introduction: Vitamin D has been suggested to play a role in muscle health and function, but studies so far have been primarily in older populations for falls prevention and subsequent risk of fractures.  Methods: Vitamin D status was assessed in a healthy young adults from sunny climate countries (n = 71, aged 19–42 years) with 56% seen within 3 months of arriving in Aberdeen [newcomers; median (range) time living in the UK = 2 months (9–105 days)] and the remainder resident for >6 months [residents; 23 months (6–121 months)]. Participants attended visits every 3 months for 15 months. At each visit, fasted blood samples were collected for analysis of serum 25-hydroxyvitamin D [25(OH)D], parathyroid hormone (PTH), carboxy-terminal collagen crosslinks (CTX) and N-terminal propeptide of type I collagen (P1NP). Maximal voluntary contractions (MVC) were performed for grip strength (both arms) and for maximal isometric strength of the knee extensors (right knee).  Results: There were small seasonal variations in 25(OH)D concentrations within the newcomers and residents, but no seasonal variation in bone turnover markers. There was a positive, albeit small, association between 25(OH)D and knee extensor maximal isometric strength. Mixed modelling predicted that for each 1 nmol/L increase in 25(OH)D, peak torque would increase by 1 Nm (p = 0.04).  Conclusions: This study suggests that vitamin D may be important for muscle health in young adults migrating from sunnier climates to high latitudes, yet the potential effect is small

Gene-specific repair of Pt/DNA lesions and induction of apoptosis by the oral platinum drug JM216 in three human ovarian carcinoma cell lines sensitive and resistant to cisplatin

JM216, an oral platinum drug entering into phase III clinical trial, exhibited comparable cytotoxicity to cisplatin in three human ovarian carcinoma cell lines: the sensitive (CH1), acquired resistant (CH1cisR) and intrinsically resistant (SKOV-3). Platinum accumulation and binding to DNA were similar in each of the three cell lines at equimolar doses, indicating that the resistant cell lines could tolerate higher intracellular platinum levels and platinum bound to DNA at IC50 concentrations of drug. Comparison with cisplatin demonstrated that intracellular platinum levels were marginally higher with JM216, but that platinum binding to DNA was similar for the two drugs in each of the cell lines. Each of the cell lines exhibited an ability to repair JM216 induced platinum/DNA lesions in the N-ras gene (gene-specific repair) at equitoxic concentrations of drug. However, this occurred to a greater extent in the two resistant cell lines such that by 24 h the CH1cisR and SKOV-3 had removed 72% and 67% respectively compared with approximately 32% for the CH1. Reduced gene-specific repair capacity in CH1 cells was also seen following incubation with 25 μM (or 5 μM – 2 × IC50) cisplatin, whereas the CH1cisR and SKOV-3 cell lines were repair proficient. JM216 induced apoptosis in the three cell lines following a 2h incubation with 2 × the IC50 of drug. Fluorescent microscopy of cells stained with propidium iodide showed that the detached cell population displayed typical apoptotic nuclei. Furthermore, field inversion gel electrophoresis demonstrated the presence of DNA fragments approximately 23–50 kb in size, indicative of apoptosis, in the detached cells. JM216 induced an S phase slow down in each of the three cell lines accompanied by a G2 block in the CH1 pair. Incubation with this concentration of JM216 also resulted in the induction of p53 in the CH1 and CH1cisR. These studies suggest that the relative sensitivity of the CH1 cell line to cisplatin and JM216 is at least partly attributable to a deficiency in gene-specific repair. The oral platinum drug, JM216, exerts its cytotoxic effects through the induction of apoptosis following a slow-down in S phase in both the sensitive and resistant lines. © 1999 Cancer Research Campaig

A prebiotic intervention study in children with autism spectrum disorders (ASDs)

Different dietary approaches, such as gluten and casein free diets, or the use of probiotics and prebiotics have been suggested in autistic spectrum disorders in order to reduce gastrointestinal (GI) disturbances. GI symptoms are of particular interest in this population due to prevalence and correlation with the severity of behavioural traits. Nowadays, there is lack of strong evidence about the effect of dietary interventions on these problems, particularly prebiotics. Therefore, we assessed the impact of exclusion diets and a 6-week Bimuno® galactooligosaccharide (B-GOS®) prebiotic intervention in 30 autistic children. RESULTS: The results showed that children on exclusion diets reported significantly lower scores of abdominal pain and bowel movement, as well as lower abundance of Bifidobacterium spp. and Veillonellaceae family, but higher presence of Faecalibacterium prausnitzii and Bacteroides spp. In addition, significant correlations were found between bacterial populations and faecal amino acids in this group, compared to children following an unrestricted diet. Following B-GOS® intervention, we observed improvements in anti-social behaviour, significant increase of Lachnospiraceae family, and significant changes in faecal and urine metabolites. CONCLUSIONS: To our knowledge, this is the first study where the effect of exclusion diets and prebiotics has been evaluated in autism, showing potential beneficial effects. A combined dietary approach resulted in significant changes in gut microbiota composition and metabolism suggesting that multiple interventions might be more relevant for the improvement of these aspects as well as psychological traits

FUTURE PERSPECTIVES IN MELANOMA RESEARCH. Meeting report from the “Melanoma Research: a bridge from Naples to the World. Napoli, December 5th–6 th2011”

After more than 30 years, landmark progress has been made in the treatment of cancer, and melanoma in particular, with the success of new molecules such as ipilimumab, vemurafenib and active specific immunization

Molecular markers of response and toxicity to FOLFOX chemotherapy in metastatic colorectal cancer

BACKGROUND: To investigate three genetic alterations (TP53 mutation, Kras mutation and microsatellite instability (MSI)) and three polymorphisms (methylene tetrahydrofolate reductase (MTHFR) C677T, excision repair cross complementing group 1 (ERCC1)-118 and X-ray repair cross complementing group 1 (XRCC1)-399) for their ability to predict response, survival and toxicity to FOLFOX first line chemotherapy in the treatment of metastatic colorectal cancer (mCRC). METHODS: Tumour tissues from 118 mCRC patients who underwent FOLFOX treatment from three successive phase II trials were evaluated for mutations in TP53 (exons 5–8) and Kras (codons 12 and 13) and for MSI using PCR-based analysis. Genotyping for common single nucleotide polymorphisms in the MTHFR (codon 677), ERCC1 (codon 118) and XRCC1 (codon 399) genes was also carried out using PCR techniques. These genetic markers were correlated with clinical response, survival and toxicity to treatment. RESULTS: Patients with the T allele of ERCC1-118 showed significantly worse progression-free survival in univariate analysis (HR 2.62; 95 % CI 1.14–6.02; P 0.02). None of the genetic alterations or polymorphisms showed significant association with clinical response to FOLFOX. The MTHFR, ERCC1 and XRCC1 polymorphisms showed no associations with overall haematological, gastrointestinal or neurological toxicity to FOLFOX, although MTHFR 677 TT genotype patients showed a significantly higher incidence of grade 3 or 4 diarrhoea (26%) compared with CC or CT genotype patients (6%, P 0.02). CONCLUSIONS: The ERCC1-118 and MTHFR C677T polymorphisms were associated with progression and severe diarrhoea

Copeptin for risk stratification in non-traumatic headache in the emergency setting: a prospective multicenter observational cohort study

In the emergency setting, non-traumatic headache is a benign symptom in 80% of cases, but serious underlying conditions need to be ruled out. Copeptin improves risk stratification in several acute diseases. Herein, we investigated the value of copeptin to discriminate between serious secondary headache and benign headache forms in the emergency setting.; Patients presenting with acute non-traumatic headache were prospectively enrolled into an observational cohort study. Copeptin was measured upon presentation to the emergency department. Primary endpoint was serious secondary headache defined by a neurologic cause requiring immediate treatment of the underlying disease. Secondary endpoint was the combination of mortality and hospitalization within 3 months. Two board-certified neurologist blinded to copeptin levels verified the endpoints after a structured 3-month-telephone interview.; Of the 391 patients included, 75 (19%) had a serious secondary headache. Copeptin was associated with serious secondary headache (OR 2.03, 95%CI 1.52-2.70, p < 0.0001). Area under the curve (AUC) for copeptin to identify the primary endpoint was 0.70 (0.63-0.76). After adjusting for age > 50, focal-neurological abnormalities, and thunderclap onset of symptoms, copeptin remained an independent predictive factor for serious secondary headache (OR 1.74, 95%CI 1.26-2.39, p = 0.001). Moreover, copeptin improved the AUC of the multivariate logistic clinical model (p-LR-test < 0.001). Even though copeptin values were higher in patients reaching the secondary endpoint, this association was not significant in multivariate logistic regression.; Copeptin was independently associated with serious secondary headache as compared to benign headaches forms. Copeptin may be a promising novel blood biomarker that should be further validated to rule out serious secondary headache in the emergency department.; Study Registration on 08/02/2010 as NCT01174901 at clinicaltrials.gov