Inactivation of polycomb repressive complex 2 components in myeloproliferative and myelodysplastic/myeloproliferative neoplasms

The polycomb repressive complex 2 (PRC2) is a highly conserved histone H3 lysine 27 methyltransferase that regulates the expression of developmental genes. Inactivating mutations of the catalytic component of PRC2, EZH2, are seen in myeloid disorders. We reasoned that the other 2 core PRC2 components, SUZ12 and EED, may also be mutational targets in these diseases, as well as associated factors such as JARID2. SUZ12 mutations were identified in 1 of 2 patients with myelodysplastic syndrome/myeloproliferative neoplasms with 17q acquired uniparental disomy and in 2 of 2 myelofibrosis cases with focal 17q11 deletions. All 3 were missense mutations affecting the highly conserved VEFS domain. Analysis of a further 146 myelodysplastic syndrome/myeloproliferative neoplasm patients revealed an additional VEFS domain mutant, yielding a total mutation frequency of 1.4% (2 of 148). We did not find mutations of JARID2 or EED in association with acquired uniparental disomy for chromosome 6p or 11q, respectively; however, screening unselected cases identified missense mutations in EED (1 of 148; 1%) and JARID2 (3 of 148; 2%). All 3 SUZ12 mutations tested and the EED mutation reduced PRC2 histone methyltransferase activity in vitro, demonstrating that PRC2 function may be compromised in myeloid disorders by mutation of distinct genes.     

Mycobacterium tuberculosis--heterogeneity revealed through whole genome sequencing

The emergence of whole genome sequencing (WGS) technologies as primary research tools has allowed for the detection of genetic diversity in Mycobacterium tuberculosis (Mtb) with unprecedented resolution. WGS has been used to address a broad range of topics, including the dynamics of evolution, transmission and treatment. Here, we have analyzed 55 publically available genomes to reconstruct the phylogeny of Mtb, and we have addressed complications that arise during the analysis of publically available WGS data. Additionally, we have reviewed the application of WGS to the study of Mtb and discuss those areas still to be addressed, moving from global (phylogeography), to local (transmission chains and circulating strain diversity), to the single patient (clonal heterogeneity) and to the bacterium itself (evolutionary studies). Finally, we discuss the current WGS approaches, their strengths and limitations.     

GABAergic processes in the mesencephalic tegmentum modulate the occurrence of active (rapid eye movement) sleep in guinea pigs

The ventrolateral subdivision of the periaqueductal gray (vlPAG) and the adjacent dorsal mesencephalic reticular formation (dMRF) are involved in the modulation of active (rapid eye movement) sleep (AS). In order to determine the effects on AS of the suppression of neuronal activity in these regions, muscimol, a GABA receptor A (GABA(A)) receptor agonist, and bicuculline, a GABA(A) receptor antagonist, were microinjected bilaterally in guinea pigs and the states of sleep and wakefulness were examined. The main effect of muscimol was an increase in AS; this increase occurred in conjunction with a reduction in the time spent in wakefulness. The powerful effect of muscimol was striking especially when considering the small amount of naturally-occurring AS that is present in this species. Additional observable effects that were induced by muscimol were: 1) long lasting episodes of hypotonia/atonia during wakefulness and quiet sleep that included a lack of extensor tone in the hind limbs, and 2) frequently occurring cortical spindles, similar to those observed during naturally-occurring quiet sleep (sleep spindles), that were present during wakefulness. Conversely, bilateral microinjections of bicuculline induced a prolonged state of wakefulness and blocked the effect of subsequent injections of muscimol. These data suggest that endogenous GABA acts on GABA(A) receptors within the vlPAG and dMRF to promote AS in the guinea pig.     

Immunophenotypic characterization of anal gland carcinoma: loss of p63 and cytokeratin 5/6

Anal gland carcinoma (AGC) is a rare perianal invasive cancer composed of tubular glands lined by cuboidal epithelium. The clinical features and histogenesis of AGC are not well understood and its origin from anal glands is often difficult to prove. Little is known about immunophenotypic features of AGC that could be useful in establishing the diagnosis. This study evaluated the immunohistochemical profile of 2 cases of AGC in comparison to anal glands from 11 hemorrhoidectomy specimens. Sections from the specimens were routinely processed and immunostained using commercial antibodies to cytokeratin (CK) 7, CK20, CK5/ 6, p63, CDX2, smooth muscle actin, calponin, heavy chain smooth muscle myosin, p53, and p16. In case 1 of AGC, radiation and chemotherapy preceded an abdominoperineal resection. In biopsies from this case, the neoplastic anal glands had a tubular pattern, whereas most glands in the resection specimen exhibited mucinous features. The histologic pattern in case 2 was tubular. Normal anal glands showed immunoreactivity for myoepithelial and basal cell markers CK5/6 and p63 in basal and parabasal cell layers and for CK7 in superficial cell layers. In contrast, both cases of AGC were negative for CK5/6 and p63 and were diffusely positive for CK7. Normal glands and both cases of AGC were negative for the intestinal differentiation marker CDX2, CK20, smooth muscle actin, calponin, smooth muscle myosin heavy chain, p16, and p53. Our data suggest that loss of p63 and CK5/6 expression is a feature of AGC. Anal gland carcinoma shares negativity for CDX2 and CK7+/CK20- profile with normal anal glands. No evidence of myoepithelial cells was found in normal or malignant anal glands. These data may be useful in establishing the diagnosis of AGC.     

Differential responses to JNJ‐37822681, a specific and fast dissociating dopamine D2 receptor antagonist,in cynomolgus monkey and Sprague–Dawley rat general toxicology studies: clinical observations,prolactin levels,mammary histopathology findings and toxicokinetics

JNJ‐37822681 is a potent, specific and fast dissociating dopamine D₂ receptor antagonist intended for the treatment of schizophrenia. Its nonclinical toxicological profile was investigated in a series of general repeat dose toxicity studies in cynomolgus monkeys and Sprague–Dawley rats. The maximum duration of treatment was 9 and 6 months, respectively. Interspecies differences were noted in the response to JNJ‐37822681 in terms of extrapyramidal (EPS)‐like clinical signs and prolactin‐mediated tissue changes in the mammary gland. Monkeys showed severe EPS‐like clinical signs such as abnormal posture, abnormal eye movements and hallucination‐like behavior at relatively low exposures compared to those associated with EPS in patients with schizophrenia. The high sensitivity of the monkey to JNJ‐37822681‐induced EPS‐like signs was unexpected based on the fast dissociating properties of the compound. Rats, however, were not prone to EPS. Elevated serum prolactin levels were found in rats and monkeys. While rats showed slight to moderate prolactin‐related tissue changes upon histopathological examination in all studies, which among others affected the mammary gland, only minor mammary gland tissue changes were noted in monkeys. Prolactin levels were only slightly increased in patients with schizophrenia receiving relatively high dose levels of JNJ‐37822681. The monkey toxicology studies did not provide an exposure‐based safety margin, while in rats adverse effects were only noted at exposures considerably higher than those achieved at efficacious plasma concentrations in the clinic. Overall, the available data suggest that the cynomolgus monkey showed better predictivity towards the nature of JNJ‐37822681‐associated adverse events in humans than the Sprague–Dawley rat. Copyright © 2013 John Wiley & Sons, Ltd.     

Putative receptor binding sites on alphaviruses as visualized by cryoelectron microscopy.

The structures of Sindbis virus and Ross River virus complexed with Fab fragments from monoclonal antibodies have been determined from cryoelectron micrographs. Both antibodies chosen for this study bind to regions of the virions that have been implicated in cell-receptor recognition and recognize epitopes on the E2 glycoprotein. The two structures show that the Fab fragments bind to the outermost tip of the trimeric envelope spike protein. Hence, the same region of both the Sindbis virus and Ross River virus envelope spike is composed of E2 and is involved in recognition of the cellular receptor.