Why do results conflict regarding the prognostic value of the methylation status in colon cancers? the role of the preservation method

<p>Abstract</p> <p>Background</p> <p>In colorectal carcinoma, extensive gene promoter hypermethylation is called the CpG island methylator phenotype (CIMP). Explaining why studies on CIMP and survival yield conflicting results is essential. Most experiments to measure DNA methylation rely on the sodium bisulfite conversion of unmethylated cytosines into uracils. No study has evaluated the performance of bisulfite conversion and methylation levels from matched cryo-preserved and Formalin-Fixed Paraffin Embedded (FFPE) samples using pyrosequencing.</p> <p>Methods</p> <p>Couples of matched cryo-preserved and FFPE samples from 40 colon adenocarcinomas were analyzed. Rates of bisulfite conversion and levels of methylation of <it>LINE-1, MLH1 </it>and <it>MGMT </it>markers were measured.</p> <p>Results</p> <p>For the reproducibility of bisulfite conversion, the mean of bisulfite-to-bisulfite standard deviation (SD) was 1.3%. The mean of run-to-run SD of PCR/pyrosequencing was 0.9%. Of the 40 DNA couples, only 67.5%, 55.0%, and 57.5% of FFPE DNA were interpretable for <it>LINE-1, MLH1</it>, and <it>MGMT </it>markers, respectively, after the first analysis. On frozen samples the proportion of well converted samples was 95.0%, 97.4% and 87.2% respectively. For DNA showing a total bisulfite conversion, 8 couples (27.6%) for <it>LINE-1</it>, 4 couples (15.4%) for <it>MLH1 </it>and 8 couples (25.8%) for <it>MGMT </it>displayed significant differences in methylation levels.</p> <p>Conclusions</p> <p>Frozen samples gave reproducible results for bisulfite conversion and reliable methylation levels. FFPE samples gave unsatisfactory and non reproducible bisulfite conversions leading to random results for methylation levels. The use of FFPE collections to assess DNA methylation by bisulfite methods must not be recommended. This can partly explain the conflicting results on the prognosis of CIMP colon cancers.</p

Clinical spectrum of MTOR-related hypomelanosis of Ito with neurodevelopmental abnormalities

PURPOSE: Hypomelanosis of Ito (HI) is a skin marker of somatic mosaicism. Mosaic MTOR pathogenic variants have been reported in HI with brain overgrowth. We sought to delineate further the pigmentary skin phenotype and clinical spectrum of neurodevelopmental manifestations of MTOR-related HI. METHODS: From two cohorts totaling 71 patients with pigmentary mosaicism, we identified 14 patients with Blaschko-linear and one with flag-like pigmentation abnormalities, psychomotor impairment or seizures, and a postzygotic MTOR variant in skin. Patient records, including brain magnetic resonance image (MRI) were reviewed. Immunostaining (n = 3) for melanocyte markers and ultrastructural studies (n = 2) were performed on skin biopsies. RESULTS: MTOR variants were present in skin, but absent from blood in half of cases. In a patient (p.[Glu2419Lys] variant), phosphorylation of p70S6K was constitutively increased. In hypopigmented skin of two patients, we found a decrease in stage 4 melanosomes in melanocytes and keratinocytes. Most patients (80%) had macrocephaly or (hemi)megalencephaly on MRI. CONCLUSION: MTOR-related HI is a recognizable neurocutaneous phenotype of patterned dyspigmentation, epilepsy, intellectual deficiency, and brain overgrowth, and a distinct subtype of hypomelanosis related to somatic mosaicism. Hypopigmentation may be due to a defect in melanogenesis, through mTORC1 activation, similar to hypochromic patches in tuberous sclerosis complex

Clinical spectrum of MTOR-related hypomelanosis of Ito with neurodevelopmental abnormalities.

PURPOSE: Hypomelanosis of Ito (HI) is a skin marker of somatic mosaicism. Mosaic MTOR pathogenic variants have been reported in HI with brain overgrowth. We sought to delineate further the pigmentary skin phenotype and clinical spectrum of neurodevelopmental manifestations of MTOR-related HI. METHODS: From two cohorts totaling 71 patients with pigmentary mosaicism, we identified 14 patients with Blaschko-linear and one with flag-like pigmentation abnormalities, psychomotor impairment or seizures, and a postzygotic MTOR variant in skin. Patient records, including brain magnetic resonance image (MRI) were reviewed. Immunostaining (n = 3) for melanocyte markers and ultrastructural studies (n = 2) were performed on skin biopsies. RESULTS: MTOR variants were present in skin, but absent from blood in half of cases. In a patient (p.[Glu2419Lys] variant), phosphorylation of p70S6K was constitutively increased. In hypopigmented skin of two patients, we found a decrease in stage 4 melanosomes in melanocytes and keratinocytes. Most patients (80%) had macrocephaly or (hemi)megalencephaly on MRI. CONCLUSION: MTOR-related HI is a recognizable neurocutaneous phenotype of patterned dyspigmentation, epilepsy, intellectual deficiency, and brain overgrowth, and a distinct subtype of hypomelanosis related to somatic mosaicism. Hypopigmentation may be due to a defect in melanogenesis, through mTORC1 activation, similar to hypochromic patches in tuberous sclerosis complex

Spectres clinique et moléculaire des malformations rénales associées au syndrome de Kabuki

DIJON-BU Médecine Pharmacie (212312103) / SudocSudocFranceF

One-carbon cycle alterations induced by Dyrk1a dosage

© 2014 The Authors. Published by Elsevier Inc. Hyperhomocysteinemia due to cystathionine beta synthase deficiency confers diverse clinical manifestations. It is characterized by elevated plasma homocysteine levels, a common amino acid metabolized by remethylation to methionine or transsulfuration to cysteine.We recently found a relationship between hepaticDyrk1A protein expression, a serine/threonine kinase involved in signal transduction in biological processes, hepatic S-adenosylhomocysteine activity, and plasma homocysteine levels.We aimed to study whether there is also a relationship between Dyrk1a and cystathionine beta synthase activity. We used different murine models carrying altered gene coy numbers for Dyrk1a, and found a decreased cystathionine beta synthase activity in the liver of mice under-expressing Dyrk1a, and an increased in liver of mice over-expressing Dyrk1a. For each model, a positive correlation was found between cystathionine beta synthase activity and Dyrk1a protein expression in the liver of mice, which was confirmed in a non-modified genetic context. The positive correlation found between liver Dyrk1a protein expression and CBS activity in modified and non-modified genetic context strengthens the role of this kinase in one carbon metabolism.This work was supported by the Fondation Jérôme Lejeune (30CA1140087) and the Association Gaspard Félix (L'AGAFE) (30S1B07)Peer Reviewe