Minimal methylation classifier (MIMIC): A novel method for derivation and rapid diagnostic detection of disease-associated DNA methylation signatures.

Rapid and reliable detection of disease-associated DNA methylation patterns has major potential to advance molecular diagnostics and underpin research investigations. We describe the development and validation of minimal methylation classifier (MIMIC), combining CpG signature design from genome-wide datasets, multiplex-PCR and detection by single-base extension and MALDI-TOF mass spectrometry, in a novel method to assess multi-locus DNA methylation profiles within routine clinically-applicable assays. We illustrate the application of MIMIC to successfully identify the methylation-dependent diagnostic molecular subgroups of medulloblastoma (the most common malignant childhood brain tumour), using scant/low-quality samples remaining from the most recently completed pan-European medulloblastoma clinical trial, refractory to analysis by conventional genome-wide DNA methylation analysis. Using this approach, we identify critical DNA methylation patterns from previously inaccessible cohorts, and reveal novel survival differences between the medulloblastoma disease subgroups with significant potential for clinical exploitation.

Residual postoperative tumour volume predicts outcome after high-dose radiotherapy for chordoma and chondrosarcoma of the skull base and spine.

AIMS: High-dose radiotherapy after surgical debulking is the treatment of choice for chordomas and chondrosarcomas. This study reviewed our outcomes, in relation to residual tumour volume and radiation dose, in order to inform our future practice. PATIENTS AND METHODS: Nineteen patients referred to the Neuro-Oncology Unit at Addenbrooke's Hospital (Cambridge, UK) between 1996 and 2009 and treated with photon radiotherapy were reviewed. Seventeen of the 19 were treated with curative intent. The median follow-up was 53 months. The tumours in the study had a mean gross tumour volume (GTV) of 17.2 cm(3) (median 10.5 cm(3)) and a range of 0-76.3 cm(3). The median dose was 65Gy in 39 fractions. RESULTS: The 5 year cause-specific survival for radically treated patients with chordomas was 92% and the 5 year local control rate was 83%. The 5 year cause-specific survival and local control rates with chondrosarcomas were both 100%. A planning target volume (PTV) below 90 cm(3) is predictive of local control, but volumes above this are not. The GTV seems to be a better predictor of outcome: among the 17 of 19 patients treated curatively, a GTV threshold of 30 cm(3) distinguished local failures from the 15 patients with local control, with sensitivity to detect local control of 100% (95% confidence interval 78-100%), specificity 100% (95% confidence interval 16-100%) and positive predictive value 100% (95% confidence interval 78-100%). CONCLUSIONS: Our results show a high level of efficacy for fractionated photon radiotherapy after surgery, in keeping with other series. In addition, we found that although surgical debulking is essential, a small residual tumour volume may still be controlled with high-dose photon radiotherapy. This information may be relevant during neurosurgical planning, possibly allowing a reduction in risk of serious neurological deficits. This should encourage the further development of sophisticated photon radiotherapy, for patients unsuitable for proton therapy.

Morphometric analysis of neuromuscular topography in the serratus anterior muscle.

Groups of neurons form ordered topographic maps on their targets, and defining the mechanisms that develop such maps, and re-connect them after disruption, has biological as well as clinical importance. The neuromuscular system is an accessible and well-studied model for defining the principles that guide map formation, both during its development and its reformation after motor nerve damage. We present evidence for the expression of this map at the level of nerve terminal morphology and muscle fiber type in the serratus anterior muscle. Morphometric analyses indicate, first, a rostrocaudal difference in nerve terminal size depending on the ventral root of origin of the axons. Second, motor endplates are larger on type IIB than type IIA muscle fibers. Third, whereas IIB muscle fibers are distributed rather evenly along the rostrocaudal axis of the muscle, the more rostral type IIB fibers are preferentially innervated by anteriorly derived (C6) motor neurons, and more caudal IIB fibers are preferentially innervated by posteriorly derived (C7) motor neurons. This inference is supported by analysis of the size of nerve terminals formed in each muscle sector by rostral and caudal roots, and by evidence that the larger terminals are on IIB fibers. These results demonstrate a subcellular expression of neuromuscular topography in the serratus anterior muscle (SA) muscle in the form of differences in nerve terminal size. These results provide deeper insights into the organization of a neuromuscular system. They also offer a rationale for a topographic map, that is, to allow spinal motor centers to activate selectively different compartments within a muscle.

Secretion of an anti-inflammatory, immunomodulatory factor by Schistosomulae of Schistosoma mansoni.

Penetration and migration of schistosomulae of Schistosoma mansoni through the skin of mice is associated with reduced inflammatory responses especially after a moderate infection. Previous studies to identify the mechanisms by which schistosomulae of S. mansoni suppress inflammatory responses in human skin showed that the excretory/secretory (ES) products of schistosomulae of s. mansoni contain activities that induce production of the antiinflammatory cytokine IL-1ra from human keratinocytes. In the present study we have characterized the ES products of the schistosomulae of S. mansoni to identify the IL-1ra inducing activity. We demonstrate that this anti-inflammatory activity is associated with a protein of molecular mass 16.8 kDa (Sm 16.8). Depletion studies confirm that Sm 16.8 is the major IL-1ra inducing activity in the ES products. A comparison of the proteins in the ES products of schistosomulae of S. mansoni with those of Trichobilharzia ocellata, a bird schistosome that induces an acute inflammation in human skin, shows that Sm 16.8 is absent in the ES products of T. ocellata. Interestingly, ES products of the schistosomulae of T. ocellata were potent inducers of IL-1 alpha from human keratinocytes, whereas ES products from schistosomulae of s. mansoni induce little or no IL-1 alpha secretion from keratinocytes. Functional studies show that Sm 16.8 suppresses antigen induced lymphoproliferative responses in vitro. Addition of Sm 16.8 to spleen or axillary lymph node cell cultures resulted in a significant reduction in antigen induced IL-2 secretion. These studies show that schistosomulae of S. mansoni elaborate an anti-inflammatory, immunomodulatory factor that may help the parasite to evade host immune responses in the skin. Given the capabilities of Sm 16.8 to induce IL-ira and suppress lymphoproliferation, this protein may also have a potential use as a therapeutic agent for inflammatory skin disorders.