The CDKN2A G500 Allele Is More Frequent in GBM Patients with No Defined Telomere Maintenance Mechanism Tumors and Is Associated with Poorer Survival

Prognostic markers for glioblastoma multiforme (GBM) are important for patient management. Recent advances have identified prognostic markers for GBMs that use telomerase or the alternative lengthening of telomeres (ALT) mechanism for telomere maintenance. Approximately 40% of GBMs have no defined telomere maintenance mechanism (NDTMM), with a mixed survival for affected individuals. This study examined genetic variants in the cyclin-dependent kinase inhibitor 2A (CDKN2A) gene that encodes the p16INK4a and p14ARF tumor suppressors, and the isocitrate dehydrogenase 1 (IDH1) gene as potential markers of survival for 40 individuals with NDTMM GBMs (telomerase negative and ALT negative by standard assays), 50 individuals with telomerase, and 17 individuals with ALT positive tumors. The analysis of CDKN2A showed NDTMM GBMs had an increased minor allele frequency for the C500G (rs11515) polymorphism compared to those with telomerase and ALT positive GBMs (p = 0.002). Patients with the G500 allele had reduced survival that was independent of age, extent of surgery, and treatment. In the NDTMM group G500 allele carriers had increased loss of CDKN2A gene dosage compared to C500 homozygotes. An analysis of IDH1 mutations showed the R132H mutation was associated with ALT positive tumors, and was largely absent in NDTMM and telomerase positive tumors. In the ALT positive tumors cohort, IDH1 mutations were associated with a younger age for the affected individual. In conclusion, the G500 CDKN2A allele was associated with NDTMM GBMs from older individuals with poorer survival. Mutations in IDH1 were not associated with NDTMM GBMs, and instead were a marker for ALT positive tumors in younger individuals

Δ133p53β isoform pro-invasive activity is regulated through an aggregation-dependent mechanism in cancer cells

International audienceAbstract The p53 isoform, Δ133p53β, is critical in promoting cancer. Here we report that Δ133p53β activity is regulated through an aggregation-dependent mechanism. Δ133p53β aggregates were observed in cancer cells and tumour biopsies. The Δ133p53β aggregation depends on association with interacting partners including p63 family members or the CCT chaperone complex. Depletion of the CCT complex promotes accumulation of Δ133p53β aggregates and loss of Δ133p53β dependent cancer cell invasion. In contrast, association with p63 family members recruits Δ133p53β from aggregates increasing its intracellular mobility. Our study reveals novel mechanisms of cancer progression for p53 isoforms which are regulated through sequestration in aggregates and recruitment upon association with specific partners like p63 isoforms or CCT chaperone complex, that critically influence cancer cell features like EMT, migration and invasion

Ascorbate content of clinical glioma tissues is related to tumour grade and to global levels of 5-hydroxymethyl cytosine.

peer reviewedGliomas are incurable brain cancers with poor prognosis, with epigenetic dysregulation being a distinctive feature. 5-hydroxymethylcytosine (5-hmC), an intermediate generated in the demethylation of 5-methylcytosine, is present at reduced levels in glioma tissue compared with normal brain, and that higher levels of 5-hmC are associated with improved patient survival. DNA demethylation is enzymatically driven by the ten-eleven translocation (TET) dioxygenases that require ascorbate as an essential cofactor. There is limited data on ascorbate in gliomas and the relationship between ascorbate and 5-hmC in gliomas has never been reported. Clinical glioma samples (11 low-grade, 26 high-grade) were analysed for ascorbate, global DNA methylation and hydroxymethylation, and methylation status of the O-6-methylguanine-DNA methyltransferase (MGMT) promoter. Low-grade gliomas contained significantly higher levels of ascorbate than high-grade gliomas (p = 0.026). Levels of 5-hmC were significantly higher in low-grade than high-grade glioma (p = 0.0013). There was a strong association between higher ascorbate and higher 5-hmC (p = 0.004). Gliomas with unmethylated and methylated MGMT promoters had similar ascorbate levels (p = 0.96). One mechanism by which epigenetic modifications could occur is through ascorbate-mediated optimisation of TET activity in gliomas. These findings open the door to clinical intervention trials in patients with glioma to provide both mechanistic information and potential avenues for adjuvant ascorbate therapy

Adaptive homeostasis and the p53 isoform network

All living organisms have developed processes to sense and address environmental changes to maintain a stable internal state (homeostasis). When activated, the p53 tumour suppressor maintains cell and organ integrity and functions in response to homeostasis disruptors (stresses) such as infection, metabolic alterations and cellular damage. Thus, p53 plays a fundamental physiological role in maintaining organismal homeostasis. The TP53 gene encodes a network of proteins (p53 isoforms) with similar and distinct biochemical functions. The p53 network carries out multiple biological activities enabling cooperation between individual cells required for long‐term survival of multicellular organisms (animals) in response to an ever‐changing environment caused by mutation, infection, metabolic alteration or damage. In this review, we suggest that the p53 network has evolved as an adaptive response to pathogen infections and other environmental selection pressures

Alpha-enolase is upregulated on the cell surface and responds to plasminogen activation in mice expressing a ∆133p53α mimic.

The p53 protein is a master regulator of the stress response. It acts as a tumor suppressor by inducing transcriptional activation of p53 target genes, with roles in apoptosis, cell cycle arrest and metabolism. The discovery of at least 12 isoforms of p53, some of which have tumor-promoting properties, has opened new avenues of research. Our previous work studied tumor phenotypes in four mouse models with different p53 backgrounds: wild-type p53, p53 null, mutant p53 lacking the proline domain (mΔpro), and a mimic for the human Δ133p53α p53 isoform (Δ122p53). To identify the major proteins affected by p53 function early in the response to DNA damage, the current study investigated the entire proteome of bone marrow, thymus, and lung in the four p53 models. Protein extracts from untreated controls and those treated with amsacrine were analyzed using two-dimensional fluorescence difference gel electrophoresis. In the bone marrow, reactive proteins were universally decreased by wild-type p53, including α-enolase. Further analysis of α-enolase in the p53 models revealed that it was instead increased in Δ122p53 hematopoietic and tumor cell cytosol and on the cell surface. Alpha-enolase on the surface of Δ122p53 cells acted as a plasminogen receptor, with tumor necrosis factor alpha induced upon plasminogen stimulation. Taken together, these data identified new proteins associated with p53 function. One of these proteins, α-enolase, is regulated differently by wild-type p53 and Δ122p53 cells, with reduced abundance as part of a wild-type p53 response and increased abundance with Δ122p53 function. Increased cell surface α-enolase on Δ122p53 cells provides a possible explanation for the model's pro-inflammatory features and suggests that p53 isoforms may direct an inflammatory response by increasing the amount of α-enolase on the cell surface

Antitumor cytotoxicity induced by bone-marrow-derived antigen-presenting cells is facilitated by the tumor suppressor protein p53 via regulation of IL-12

Activated antigen-presenting cells (APC) deliver the three signals cytotoxic T cells require to differentiate into effector cells that destroy the tumor. These comprise antigen, co-stimulatory signals and cytokines. Once these cells have carried out their function, they apoptose. We hypothesized that the tumor suppressor protein, p53, played an important role in generating the antitumor response facilitated by APC. CD11cC APC derived from p53 wild-type (wt) mouse (wt p53) GM-CSF bone marrow cultures (BMAPC) and activated had reduced survival compared to BMAPC from p53 null consistent with p53-mediated apoptosis following activation. There was a lower percentage of antigenic peptide/MHC I complexes on antigen-pulsed p53 null cells suggesting p53 played a role in antigen processing but there was no difference in antigen-specific T cell proliferative responses to these cells in vivo. In contrast, antigenspecific cytotoxicity in vivo was markedly reduced in response to p53 null BMAPC. When these cells were pulsed with a model tumor antigen and delivered as a prophylactic vaccination, they provided no protection against melanoma cell growth whereas wt BMAPC were very effective. This suggested that p53 might regulate the requisite third signal and, indeed, we found that p53 null BMAPC produced less IL-12 than wt p53 BMAPC and that p53 bound to the promoter region of IL-12. This work suggests that p53 in activated BMAPC is associated with the generation of IL-12 required for the differentiation of cytotoxic immune responses and an effective antitumor response. This is a completely new role for this protein that has implications for BMAPC-mediated immunotherapy

Placental Hypomethylation Is More Pronounced in Genomic Loci Devoid of Retroelements

The human placenta is hypomethylated compared to somatic tissues. However, the degree and specificity of placental hypomethylation across the genome is unclear. We assessed genome-wide methylation of the human placenta and compared it to that of the neutrophil, a representative homogeneous somatic cell. We observed global hypomethylation in placenta (relative reduction of 22%) compared to neutrophils. Placental hypomethylation was pronounced in intergenic regions and gene bodies, while the unmethylated state of the promoter remained conserved in both tissues. For every class of repeat elements, the placenta showed lower methylation but the degree of hypomethylation differed substantially between these classes. However, some retroelements, especially the evolutionarily younger Alu elements, retained high levels of placental methylation. Surprisingly, nonretrotransposon-containing sequences showed a greater degree of placental hypomethylation than retrotransposons in every genomic element (intergenic, introns, and exons) except promoters. The differentially methylated fragments (DMFs) in placenta and neutrophils were enriched in gene-poor and CpG-poor regions. The placentally hypomethylated DMFs were enriched in genomic regions that are usually inactive, whereas hypermethylated DMFs were enriched in active regions. Hypomethylation of the human placenta is not specific to retroelements, indicating that the evolutionary advantages of placental hypomethylation go beyond those provided by expression of retrotransposons and retrogenes

Increased Ascorbate Content of Glioblastoma Is Associated With a Suppressed Hypoxic Response and Improved Patient Survival.

peer reviewedGlioblastoma multiforme is a challenging disease with limited treatment options and poor survival. Glioblastoma tumours are characterised by hypoxia that activates the hypoxia inducible factor (HIF) pathway and controls a myriad of genes that drive cancer progression. HIF transcription factors are regulated at the post-translation level via HIF-hydroxylases. These hydroxylases require oxygen and 2-oxoglutarate as substrates, and ferrous iron and ascorbate as cofactors. In this retrospective observational study, we aimed to determine whether ascorbate played a role in the hypoxic response of glioblastoma, and whether this affected patient outcome. We measured the ascorbate content and members of the HIF-pathway of clinical glioblastoma samples, and assessed their association with clinicopathological features and patient survival. In 37 samples (37 patients), median ascorbate content was 7.6 μg ascorbate/100 mg tissue, range 0.8 - 20.4 μg ascorbate/100 mg tissue. In tumours with above median ascorbate content, HIF-pathway activity as a whole was significantly suppressed (p = 0.005), and several members of the pathway showed decreased expression (carbonic anhydrase-9 and glucose transporter-1, both p < 0.01). Patients with either lower tumour HIF-pathway activity or higher tumour ascorbate content survived significantly longer than patients with higher HIF-pathway or lower ascorbate levels (p = 0.011, p = 0.043, respectively). Median survival for the low HIF-pathway score group was 362 days compared to 203 days for the high HIF-pathway score group, and median survival for the above median ascorbate group was 390 days, compared to the below median ascorbate group with 219 days. The apparent survival advantage associated with higher tumour ascorbate was more prominent for the first 8 months following surgery. These associations are promising, suggesting an important role for ascorbate-regulated HIF-pathway activity in glioblastoma that may impact on patient survival