Peter Siesjö

Enhancer hijacking activates GFI1 family oncogenes in medulloblastoma.

Medulloblastoma is a highly malignant paediatric brain tumour currently treated with a combination of surgery, radiation and chemotherapy, posing a considerable burden of toxicity to the developing child. Genomics has illuminated the extensive intertumoral heterogeneity of medulloblastoma, identifying four distinct molecular subgroups. Group 3 and group 4 subgroup medulloblastomas account for most paediatric cases; yet, oncogenic drivers for these subtypes remain largely unidentified. Here we describe a series of prevalent, highly disparate genomic structural variants, restricted to groups 3 and 4, resulting in specific and mutually exclusive activation of the growth factor independent 1 family proto-oncogenes, GFI1 and GFI1B. Somatic structural variants juxtapose GFI1 or GFI1B coding sequences proximal to active enhancer elements, including super-enhancers, instigating oncogenic activity. Our results, supported by evidence from mouse models, identify GFI1 and GFI1B as prominent medulloblastoma oncogenes and implicate ‘enhancer hijacking’ as an efficient mechanism driving oncogene activation in a childhood cancer.

The immunobiology of aluminium adjuvants: how do they really work?

Aluminium adjuvants potentiate the immune response, thereby ensuring the potency and efficacy of typically sparingly available antigen. Their concomitant critical importance in mass vaccination programmes may have prompted recent intense interest in understanding how they work and their safety. Progress in these areas is stymied, however, by a lack of accessible knowledge pertaining to the bioinorganic chemistry of aluminium adjuvants, and, consequently, the inappropriate application and interpretation of experimental models of their mode of action. The objective herein is, therefore, to identify the many ways that aluminium chemistry contributes to the wide and versatile armoury of its adjuvants, such that future research might be guided towards a fuller understanding of their role in human vaccinations.

Detection of human cytomegalovirus in medulloblastomas reveals a potential therapeutic target

Medulloblastomas are the most common malignant brain tumors in children. They express high levels of COX-2 and produce PGE2, which stimulates tumor cell proliferation. Human cytomegalovirus (HCMV) is prevalent in the human population and encodes proteins that provide immune evasion strategies and promote oncogenic transformation and oncomodulation. In particular, HCMV induces COX-2 expression; STAT3 phosphorylation; production of PGE2, vascular endothelial growth factor, and IL-6; and tumor formation in vivo. Here, we show that a large proportion of primary medulloblastomas and medulloblastoma cell lines are infected with HCMV and that COX-2 expression, along with PGE2 levels, in tumors is directly modulated by the virus. Our analysis indicated that both HCMV immediate-early proteins and late proteins are expressed in the majority of primary medulloblastomas. Remarkably, all of the human medulloblastoma cell lines that we analyzed contained HCMV DNA and RNA and expressed HCMV proteins at various levels in vitro. When engrafted into immunocompromised mice, human medulloblastoma cells induced expression of HCMV proteins. HCMV and COX-2 expression correlated in primary tumors, cell lines, and medulloblastoma xenografts. The antiviral drug valganciclovir and the specific COX-2 inhibitor celecoxib prevented HCMV replication in vitro and inhibited PGE2 production and reduced medulloblastoma tumor cell growth both in vitro and in vivo. Ganciclovir did not affect the growth of HCMV-negative tumor cell lines. These findings imply an important role for HCMV in medulloblastoma and suggest HCMV as a novel therapeutic target for this tumor.

What has inflammation to do with traumatic brain injury

IntroductionInflammation is an stereotypical response to tissue damage and has been extensively documented in experimental and clinical traumatic brain injury (TBI), including children.DiscussionThe initiation and orchestration of inflammation in TBI, as in other tissues, is complex and multifactorial encompassing pro- and anti-inflammatory cytokines, chemokines, adhesion molecules, complement factors, reactive oxygen and nitrogen species, and other undefined factors. It is evident that inflammation can have both beneficial and detrimental effects in TBI, but the mechanisms underlying this dichotomy are mostly unknown. Modification of the inflammatory response may be neuroprotective. Monitoring inflammation is now possible with techniques such as microdialysis; however, the prognostic value of measuring inflammatory mediators in TBI is still unclear with conflicting reports. Except for corticosteroids, no anti-inflammatory agents have been tested in TBI, and the negative results with these may have been flawed by their multiple side effects. Clinical trials with anti-inflammatory agents that target multiple or central and downstream pathways are warranted in adult and pediatric TBI. This review examines the mechanisms of inflammation after TBI, monitoring, and possible routes of intervention.

Molecular Mechanisms of Acidosis-Mediated Damage

The present article is concerned with mechanisms which are responsible for the exaggerated brain damage observed in hyperglycemic animals subjected to transient global or forebrain ischemia. Since hyperglycemia enchances the production of lactate plus H+ during ischemia, it seems likely that aggravation of damage is due to exaggerated intra- and extracellular acidosis. This contention is supported by results showing a detrimental effect of extreme hypercapnia in normoglycemic rats subjected to transient ischemia or to hypoglycemic coma. Enhanced acidosis may exaggerate ischemic damage by one of three mechanisms: (i) accelerating free radical production via H(+)-dependent reactions, some of which are catalyzed by iron released from protein bindings by a lowering of pH, (ii) by perturbing the intracellular signal transduction pathway, leading to changes in gene expression or protein synthesis, or (iii) by activating endonucleases which cause DNA fragmentation. While activation of endonucleases must affect the nucleus, the targets of free radical attack are not known. Microvessels are considered likely targets of such attack in sustained ischemia and in trauma; however, enhanced acidosis is not known to aggravate microvascular dysfunction, or to induce inflammatory responses at the endothelial-blood interface. A more likely target is the mitochondrion. Thus, if the ischemia is of long duration (30 min) hyperglycemia triggers rapidly developing mitochondrial failure. It is speculated that this is because free radicals damage components of the respiratory chain, leading to a secondary deterioration of oxidative phosphorylation.

Regression of intracerebral rat glioma isografts by therapeutic subcutaneous immunization with interferon-γ, interleukin-7, or B7-1-transfected tumor cells

Progress in the definition of the roles of various costimulators and cytokines in determining the type and height of immune responses has made it important to explore genetically altered tumor cells expressing such molecules for therapeutic immunizations. We have studied the effect of therapeutic subcutaneous (s.c.) immunizations on the growth of preexisting intracerebral brain tumor isografts in the rat. Transfectant glioma cell clones expressing either rat interferon-γ (IFN-γ), rat interleukin-7 (IL-7), or rat B7-1 were selected. After irradiation (80 Gy) the clones were used for immunization (administered in up to four s.c. doses in a hind leg over 14-day intervals starting 1 day after the intracranial isografting of the parental tumor). Significant growth inhibition of the intracerebral parental tumors was induced by transfectants expressing IFN-γ and IL-7, respectively. The strongest effect was observed with IFN-γ-expressing cells, resulting in cures in 37% of the males and in 100% of the females. Immunization with IL-7 had a similar, strong initial effect, with significantly prolonged survival in the majority of the rats but a lower final cure rate (survival for >150 days). The B7-1-expressing tumor clones induced cures in seven of eight female rats; however, no cures were seen in the male rats. It was also shown that the B7-1-expressing cells were themselves strongly immunogenic in female rats, requiring high cell numbers to result in a progressively growing tumor upon s.c. isografting; this was not the case in male rats. As a whole, the results imply that despite the unfavorable location of intracerebral tumors, therapeutic s.c. immunizations with certain types of genetically altered tumor cells can induce complete regressions with permanent survival and without gross neurological or other apparent signs of brain damage. The present results demonstrate complete regressions when immunizations are initiated shortly after intracranial isografting, when the intracerebral tumor is small.

Transcranial Doppler pulsatility index is not a reliable indicator of intracranial pressure in children with severe traumatic brain injury

BACKGROUND The TCD-derived PI has been associated with ICP in adult studies but has not been well investigated in children. We examined the relationship between PI and ICP and CPP in children with severe TBI. METHODS Data were prospectively collected from consecutive TCD studies in children with severe TBI undergoing ICP monitoring. Ipsilateral ICP and CPP values were examined with Spearman correlation coefficient (mean values and raw observations), with a GEE, and as binary values (1 and 20 mm Hg, respectively). RESULTS Thirty-four children underwent 275 TCD studies. There was a weak relationship between mean values of ICP and PI (P = .04, r = 0.36), but not when raw observations (P = .54) or GEE (P = .23) were used. Pulsatility index was 0.76 when ICP was lower than 20 mm Hg and 0.86 when ICP was 20 mm Hg or higher. When PI was 1 or higher, ICP was lower than 20 mm Hg in 62.5% (25 of 40 studies), and when ICP was 20 mm Hg or higher, PI was lower than 1 in 75% (46 of 61 studies). The sensitivity and specificity of a PI threshold of 1 for examining the ICP threshold of 20 mm Hg were 25% and 88%, respectively. The relationship between CPP and PI was stronger (P = .001, r = -0.41), but there were too few observations below 50 mm Hg to examine PI at this threshold. CONCLUSION The absolute value of the PI is not a reliable noninvasive indicator of ICP in children with severe TBI. Further study is required to examine the relationship between PI and a CPP threshold of 50 mm Hg.

Cure of established, intracerebral rat gliomas induced by therapeutic immunizations with tumor cells and purified APC or adjuvant IFN-gamma treatment.

We have previously reported that immunizations with mutagen-induced immunogenic variants of a weakly immunogenic rat glioma could protect against isografts of the original tumor cells. In this study we show that prolonged survival and cures of rats with established gliomas in their brains can be achieved by therapeutic immunizations with tumor cell mutants, combined with in vitro and in vivo interferon (IFN)-gamma (adjuvant) treatment, or tumor cells admixed with semipurified syngeneic dendritic cells. Cure of rats with established intracerebral gliomas was possible when immunizations were initiated up to 5 days after intracerebral isografting of original tumor cells. Unexpectedly, immunizations combined with in vitro and in vivo IFN-gamma treatment or with admixed semipurified dendritic cells equalized the immunogenic potential of the original tumor cells and that of mutagen-induced immunogenic cell variants (tum-). This demonstrates that effective immunizations against a weakly immunogenic brain tumor can be achieved by different adjuvant concepts. The therapeutic effect of immunizations with tumor cells admixed with semipurified dendritic cells was highly significant in female rats, whereas only occasional cures and prolonged survival were recorded in male rats. The overall results show that therapeutic immunizations can indeed be effective against an established and growing intracerebral tumor.

PD-L1 Expression by Neurons Nearby Tumors Indicates Better Prognosis in Glioblastoma Patients

Glioblastoma multiforme (GBM) is the most aggressive form of brain tumor. In general, tumor growth requires disruption of the tissue microenvironment, yet how this affects glioma progression is unknown. We studied program death-ligand (PD-L)1 in neurons and gliomas in tumors from GBM patients and associated the findings with clinical outcome. Remarkably, we found that upregulation of PD-L1 by neurons in tumor-adjacent brain tissue (TABT) associated positively with GBM patient survival, whereas lack of neuronal PD-L1 expression was associated with high PD-L1 in tumors and unfavorable prognosis. To understand the molecular mechanism of PD-L1 signaling in neurons, we investigated PD-L1 function in cerebellar and cortical neurons and its impact on gliomas. We discovered that neuronal PD-L1-induced caspase-dependent apoptosis of glioma cells. Because interferon (IFN)-β induces PD-L1 expression, we studied the functional consequences of neuronal Ifnb gene deletion on PD-L1 signaling and function. Ifnb−/− neurons lacked PD-L1 and were defective in inducing glioma cell death; this effect was reversed on PD-L1 gene transfection. Ifnb−/− mice with intracerebral isografts survived poorly. Similar to the observations in GBM patients, better survival in wild-type mice was associated with high neuronal PD-L1 in TABT and downregulation of PD-L1 in tumors, which was defective in Ifnb−/− mice. Our data indicated that neuronal PD-L1 signaling in brain cells was important for GBM patient survival. Reciprocal PD-L1 regulation in TABT and tumor tissue could be a prognostic biomarker for GBM. Understanding the complex interactions between tumor and adjacent stromal tissue is important in designing targeted GBM therapies.

Wnt/β-catenin pathway regulates MGMT gene expression in cancer and inhibition of Wnt signalling prevents chemoresistance

The DNA repair enzyme O6-methylguanine-DNA methyltransferase (MGMT) is commonly overexpressed in cancers and is implicated in the development of chemoresistance. The use of drugs inhibiting MGMT has been hindered by their haematologic toxicity and inefficiency. As a different strategy to inhibit MGMT we investigated cellular regulators of MGMT expression in multiple cancers. Here we show a significant correlation between Wnt signalling and MGMT expression in cancers with different origin and confirm the findings by bioinformatic analysis and immunofluorescence. We demonstrate Wnt-dependent MGMT gene expression and cellular co-localization between active β-catenin and MGMT. Pharmacological or genetic inhibition of Wnt activity downregulates MGMT expression and restores chemosensitivity of DNA-alkylating drugs in mouse models. These findings have potential therapeutic implications for chemoresistant cancers, especially of brain tumours where the use of temozolomide is frequently used in treatment.