Deregulation of microRNA expression in monocytes and CD4+ T lymphocytes from patients with axial spondyloarthritis.

BACKGROUND: MicroRNAs (MiRs) play an important role in the pathogenesis of chronic inflammatory diseases. This study is the first to investigate miR expression profiles in purified CD4+ T lymphocytes and CD14+ monocytes from patients with axial spondyloarthritis (axSpA) using a high-throughput qPCR approach. METHODS: A total of 81 axSpA patients fulfilling the 2009 ASAS classification criteria, and 55 controls were recruited from October 2014 to July 2017. CD14+ monocytes and CD4+ T lymphocytes were isolated from peripheral blood mononuclear cells. MiR expression was investigated by qPCR using the Exiqon Human MiRnome panel I analyzing 372 miRNAs. Differentially expressed miRNAs identified in the discovery cohort were validated in the replication cohort. RESULTS: We found a major difference in miR expression patterns between T lymphocytes and monocytes regardless of the patient or control status. Comparing disease-specific differentially expressed miRs, 13 miRs were found consistently deregulated in CD14+ cells in both cohorts with miR-361-3p, miR-223-3p, miR-484, and miR-16-5p being the most differentially expressed. In CD4+ T cells, 11 miRs were differentially expressed between patients and controls with miR-16-1-3p, miR-28-5p, miR-199a-5p, and miR-126-3p were the most strongly upregulated miRs among patients. These miRs are involved in disease relevant pathways such as inflammation, intestinal permeability or bone formation. Mir-146a-5p levels correlated inversely with the degree of inflammation in axSpA patients. CONCLUSIONS: We demonstrate a consistent deregulation of miRs in both monocytes and CD4+ T cells from axSpA patients, which could contribute to the pathophysiology of the disease with potential interest from a therapeutic perspective.

Comparison of the quantification of KRAS mutations by digital PCR and E-ice-COLD-PCR in circulating-cell-free DNA from metastatic colorectal cancer patients.

Circulating cell-free DNA (ccfDNA) bears great promise as biomarker for personalized medicine, but ccfDNA is present only at low levels in the plasma or serum of cancer patients. E-ice-COLD-PCR is a recently developed enrichment method to detect and identify mutations present at low-abundance in clinical samples. However, recent studies have shown the importance to accurately quantify low-abundance mutations as clinically important decisions will depend on certain mutation thresholds. The possibility for an enrichment method to accurately quantify the mutation levels remains a point of concern and might limit its clinical applicability. In the present study, we compared the quantification of KRAS mutations in ccfDNA from metastatic colorectal cancer patients by E-ice-COLD-PCR with two digital PCR approaches. For the quantification of mutations by E-ice-COLD-PCR, cell lines with known mutations diluted into WT genomic DNA were used for calibration. E-ice-COLD-PCR and the two digital PCR approaches showed the same range of the mutation level and were concordant for mutation levels below the clinical relevant threshold. E-ice-COLD-PCR can accurately detect and quantify low-abundant mutations in ccfDNA and has a shorter time to results making it compatible with the requirements of analyses in a clinical setting without the loss of quantitative accuracy.

Twelve weeks posttreatment follow-up is as relevant as 24 weeks to determine the sustained virologic response in patients with hepatitis C virus receiving pegylated interferon and ribavirin.

UNLABELLED: A sustained virologic response (SVR) in patients with chronic hepatitis C receiving pegylated interferon (PEG-IFN) plus ribavirin is defined as undetectable serum HCV-RNA at 24 weeks (W+24) posttreatment follow-up. Viral load outcome in patients with virological relapse (VR) has not been explored. This study evaluated whether the assessment of serum HCV-RNA 12 weeks (W+12) after the end of treatment was as relevant as W+24 to evaluate SVR in 573 patients who received combination PEG-IFN and ribavirin and had a virological response at the end of treatment. Serum HCV-RNA was measured, using a new assay based on transcription-mediated amplification (TMA) with a lowest detection limit of 5-10 IU/mL, at W+12 and W+24 after the end of treatment. VR was defined as reappearance of detectable HCV-RNA at W+24 posttreatment follow-up. The positive predictive value (PPV) of undetectable serum HCV-RNA at W+12 was evaluated to identify patients with SVR, and the viral load outcome was measured in relapse patients. At the W+24 posttreatment follow-up, 408 (71%) patients had an SVR, 181 (71.2%) were treated with PEG-IFNalpha-2a and ribavirin, and 227 (71.1%) were treated with PEG-IFNalpha-2b and ribavirin. At W+12, serum HCV-RNA was undetectable in 409 patients, and 408 patients were SVR (PPV 99.7%, 95% confidence interval 99.1-100). In relapse patients, serum HCV-RNA levels were 5.623 +/- 0.748, 4.979 +/- 0.870, and 5.216 +/- 0.758 log(10) IU/mL at baseline, W+12, and W+24, respectively. CONCLUSION: Our results show that the assessment of serum HCV-RNA 12 weeks after the end of treatment, using the highly sensitive TMA assay (PPV 99.7%), is as relevant as after 24 weeks to predict SVR and make decisions on the management of treated patients, suggesting a new definition for SVR.

Influence of genotype on hepatitis B surface antigen kinetics in hepatitis B e antigen-negative patients treated with pegylated interferon-alpha2a.

BACKGROUND: The aim of this study was to assess the influence of hepatitis B virus (HBV) genotypes on serum hepatitis B surface antigen (HBsAg) kinetics in hepatitis B e antigen (HBeAg)-negative patients treated with pegylated interferon-alpha2a (PEG-IFN-alpha2a). METHODS: A total of 48 consecutive patients treated with PEG-IFN-alpha2a (180 microg/week) for 48 weeks were assessed. HBV genotype was determined. Serum HBV DNA and HBsAg were assessed at baseline, during treatment (weeks 12, 24 and 48) and during follow-up (weeks 72 and 96). RESULTS: The distribution of HBV genotype was A 27%, B 17%, C 12%, D 29% and E 14%. Mean +/-sd pretreatment serum HBV DNA (6.9 +/-1.5 log(10) copies/ml) was not different between genotypes and decreased under treatment in all genotypes without significant difference. Mean +/-sd pretreatment serum HBsAg (3.6 +/-0.6 log(10) IU/ml) was significantly different between genotypes (P<0.001), with high levels in genotypes A and C, intermediate levels in genotypes D and E, and low levels in genotype B (4.0 +/-0.3, 4.1 +/-0.7, 3.6 +/-0.5, 3.6 +/-0.4 and 2.7 +/-0.6 log(10) IU/ml, respectively). Serum HBsAg decreased under treatment in all genotypes with a significant difference. At the end of treatment, mean +/-sd decrease was high in genotype A, intermediate in genotypes B and D, and low in genotypes C and E (1.3 +/-1.6, 0.7 +/-0.7, 0.6 +/-0.9, 0.4 +/-1.0 and 0.4 +/-0.9 log(10) IU/ml, respectively; P<0.001). During follow-up, serum HBsAg continued to decrease in genotypes A and D, whereas rebound was observed in genotypes B, C and E. CONCLUSIONS: HBV genotype has a strong influence on serum HBsAg kinetics during PEG-IFN-alpha2a therapy in HBeAg-negative patients.

Clinical performances of two real-time PCR assays and bDNA/TMA to early monitor treatment outcome in patients with chronic hepatitis C.

BACKGROUND: Early viral monitoring is essential for the management of treatment outcome in patients with chronic hepatitis C. A variety of commercially available assays are now available to quantify HCV-RNA in routine clinical practice. OBJECTIVES: Compare the clinical results of 3 commercially available assays to evaluate the positive predictive value (PPV) and the negative predictive value (NPV) of rapid virological response (RVR) at week 4 and early virological response (EVR) at week 12. STUDY DESIGN: 287 patients treated with standard care regimen combination therapy were studied. HCV-RNA values measured at baseline, week 4, week 12 with VERSANT HCV 3.0 Assay (bDNA), and VERSANT HCV-RNA Qualitative Assay (TMA) (bDNA/TMA); COBAS Ampliprep/COBAS/TaqMan (CAP/CTM) and Abbott m2000sp extraction/m2000rt amplification system (ART). RVR was defined as undetectable serum HCV-RNA and EVR as a > OR =2 log decline in baseline viral load (BLV). RESULTS: Median (range) BVLs were: 5.585(2.585-6.816), 5.189(2.792-7.747) and 4.804(2.380-6.580) log(10)IU/ml, with bDNA/TMA, CAP/CTM and ART, respectively (p<0.01); RVR was observed in 22%, 30% and 27% of the patients and PPVs were 97%, 91% and 94% with bDNA/TMA, CAP/CTM and ART, respectively (p=0.317). EVR was observed in 76%, 73% and 67% of the patients and NPVs were 93%, 83% and 79% with bDNA/TMA, CAP/CTM and ART, respectively (p=0.09). CONCLUSIONS: Treatment monitoring should include both detection of serum HCV-RNA at week 4 to predict SVR and at week 12 to predict non-SVR. The value of all 3 assays was similar for evaluating RVR or EVR. Because of viral load discrepancies the same assay should be used throughout patient treatment follow-up.

A clinical, neuropsychological and olfactory evaluation of a large family with LRRK2 mutations.

We evaluated the neurological and neuropsychological profiles and olfaction as presymptomatic markers in a large family with Parkinson disease (PD) caused by the G2019S mutation in the LRRK2 gene. Five affected family members, 14 asymptomatic mutation carriers and 15 non-carriers were compared. Patients had typical dopa-responsive PD, frequently associated with cognitive impairment. Asymptomatic carriers and non-carriers could not be distinguished because of their neuropsychological status, the presence of depression or olfactory impairment. We were therefore unable to identify a presymptomatic marker of LRRK2-related PD.

Frequency of the LRRK2 G2019S mutation in siblings with Parkinson's disease.

BACKGROUND: Mutations in the LRRK2 gene, the most frequent of which is the G2019S mutation in exon 41, cause familial and sporadic Parkinson's disease (PD) with reduced penetrance. OBJECTIVES: To assess the frequency of the LRRK2 G2019S mutation in families thought to have autosomal recessive PD (siblings but not their parents were affected) and to determine the clinical features of LRRK2 mutation carriers. METHODS: We sequenced both strands of exon 41 of the LRKK2 gene in 90 index cases from French and North African families in which PD might have been inherited as a recessive trait. Patients with mutations underwent detailed clinical evaluations. RESULTS: We found heterozygous G2019S mutations in 1 Algerian and 1 French family (overall relative frequency = 2.2%). Four LRRK2-positive patients had typical PD, but their Mini Mental State Examination (MMSE) scores were lower than those of patients without this mutation. CONCLUSION: The LRRK2 G2019S mutation is as frequent in families with possible autosomal recessive PD (2.2%) as in the sporadic cases published elsewhere (1.9%). The clinical features in the LRRK2-positive patients were those of typical PD, except for lower MMSE scores.

LRRK2 exon 41 mutations in sporadic Parkinson disease in Europeans.

BACKGROUND: Mutations in leucine-rich repeat kinase 2 gene (LRRK2), particularly the G2019S mutation in exon 41, have been detected in familial and sporadic Parkinson disease (PD) cases. OBJECTIVES: To assess the frequency of LRRK2 exon 41 mutations in a series of sporadic PD cases from Europe and to determine the clinical features of LRRK2 mutation carriers. DESIGN: We analyzed European cases of sporadic PD for the presence of LRRK2 exon 41 mutations. These mutations were screened by denaturing high-performance liquid chromatography, and abnormal chromatograph traces were investigated by direct sequencing to determine the exact nature of the variants. Early-onset sporadic PD cases were also screened for parkin mutations. The haplotypes associated with the G2019S mutation were determined. The clinical characteristics of patients carrying LRRK2 mutations were detailed. SETTING: French Network for the Study of Parkinson Disease Genetics. Patients Three hundred twenty patients with apparently sporadic PD from Europe. MAIN OUTCOME MEASURES: Results of genetic analyses. RESULTS: We found the G2019S mutation in 6 patients and identified 2 new variants (Y2006H and T2031S) in 1 patient each. Their clinical features were similar to those of typical PD. All G2019S mutation carriers shared a common haplotype. CONCLUSIONS: The G2019S mutation is almost as frequent in sporadic cases (1.9%) as in previously reported familial cases (2.9%) in Europe and occurs in the same common founder. We identified 2 novel variants. Although the phenotype of LRRK2 mutation carriers closely resembles that of typical PD, the age at onset was younger (29 years in 1 patient) than previously described, and 3 patients were improved by deep brain stimulation.