Metabolic adaptation is essential for cell survival during nutrient deprivation. We report that eukaryotic elongation factor 2 kinase (eEF2K), which is activated by AMP-kinase (AMPK), confers cell ...survival under acute nutrient depletion by blocking translation elongation. Tumor cells exploit this pathway to adapt to nutrient deprivation by reactivating the AMPK-eEF2K axis. Adaptation of transformed cells to nutrient withdrawal is severely compromised in cells lacking eEF2K. Moreover, eEF2K knockdown restored sensitivity to acute nutrient deprivation in highly resistant human tumor cell lines. In vivo, overexpression of eEF2K rendered murine tumors remarkably resistant to caloric restriction. Expression of eEF2K strongly correlated with overall survival in human medulloblastoma and glioblastoma multiforme. Finally, C. elegans strains deficient in efk-1, the eEF2K ortholog, were severely compromised in their response to nutrient depletion. Our data highlight a conserved role for eEF2K in protecting cells from nutrient deprivation and in conferring tumor cell adaptation to metabolic stress.
Display omitted
Display omitted
•eEF2K is required for cell survival under acute nutrient deprivation•AMPK-eEF2K reactivation supports adaptation of transformed cells to nutrient stress•eEF2K expression predicts poor prognosis in aggressive brain tumors•Efk1 (eEF2K ortholog) promotes survival of C. elegans under nutrient deprivation
Tumor cells adapt to the stress of nutrient deprivation by increasing the activity of translation elongation factor 2 kinase (eEF2K), which protects cells from apoptosis by inhibiting the elongation step of mRNA translation.
We report that human secretory breast carcinoma (SBC), a rare subtype of infiltrating ductal carcinoma, expresses the
ETV6-NTRK3 gene fusion previously cloned in pediatric mesenchymal cancers. This ...gene fusion encodes a chimeric tyrosine kinase with potent transforming activity in fibroblasts.
ETV6-NTRK3 expression was confirmed in 12 (92%) of 13 SBC cases, but not in other ductal carcinomas. Retroviral transfer of ETV6-NTRK3 (EN) into murine mammary epithelial cells resulted in transformed cells that readily formed tumors in nude mice. Phenotypically, tumors produced glands and expressed epithelial antigens, confirming that EN transformation is compatible with epithelial differentiation. This represents a recurrent chromosomal rearrangement and expression of a dominantly acting oncogene as a primary event in human breast carcinoma.
Oxidative stress plays a key role in late onset diseases including cancer and neurodegenerative diseases such as Huntington disease. Therefore, uncovering regulators of the antioxidant stress ...responses is important for understanding the course of these diseases. Indeed, the nuclear factor erythroid 2-related factor 2 (NRF2), a master regulator of the cellular antioxidative stress response, is deregulated in both cancer and neurodegeneration. Similar to NRF2, the tumor suppressor Homologous to the E6-AP Carboxyl Terminus (HECT) domain and Ankyrin repeat containing E3 ubiquitin–protein ligase 1 (HACE1) plays a protective role against stress-induced tumorigenesis in mice, but its roles in the antioxidative stress response or its involvement in neurodegeneration have not been investigated. To this end we examined Hace1 WT and KO mice and found that Hace1 KO animals exhibited increased oxidative stress in brain and that the antioxidative stress response was impaired. Moreover, HACE1 was found to be essential for optimal NRF2 activation in cells challenged with oxidative stress, as HACE1 depletion resulted in reduced NRF2 activity, stability, and protein synthesis, leading to lower tolerance against oxidative stress triggers. Strikingly, we found a reduction of HACE1 levels in the striatum of Huntington disease patients, implicating HACE1 in the pathology of Huntington disease. Moreover, ectopic expression of HACE1 in striatal neuronal progenitor cells provided protection against mutant Huntingtin-induced redox imbalance and hypersensitivity to oxidative stress, by augmenting NRF2 functions. These findings reveal that the tumor suppressor HACE1 plays a role in the NRF2 antioxidative stress response pathway and in neurodegeneration.
Array genomic hybridization is being used clinically to detect pathogenic copy number variants in children with intellectual disability and other birth defects. However, there is no agreement ...regarding the kind of array, the distribution of probes across the genome, or the resolution that is most appropriate for clinical use.
We performed 500 K Affymetrix GeneChip array genomic hybridization in 100 idiopathic intellectual disability trios, each comprised of a child with intellectual disability of unknown cause and both unaffected parents. We found pathogenic genomic imbalance in 16 of these 100 individuals with idiopathic intellectual disability. In comparison, we had found pathogenic genomic imbalance in 11 of 100 children with idiopathic intellectual disability in a previous cohort who had been studied by 100 K GeneChip array genomic hybridization. Among 54 intellectual disability trios selected from the previous cohort who were re-tested with 500 K GeneChip array genomic hybridization, we identified all 10 previously-detected pathogenic genomic alterations and at least one additional pathogenic copy number variant that had not been detected with 100 K GeneChip array genomic hybridization. Many benign copy number variants, including one that was de novo, were also detected with 500 K array genomic hybridization, but it was possible to distinguish the benign and pathogenic copy number variants with confidence in all but 3 (1.9%) of the 154 intellectual disability trios studied.
Affymetrix GeneChip 500 K array genomic hybridization detected pathogenic genomic imbalance in 10 of 10 patients with idiopathic developmental disability in whom 100 K GeneChip array genomic hybridization had found genomic imbalance, 1 of 44 patients in whom 100 K GeneChip array genomic hybridization had found no abnormality, and 16 of 100 patients who had not previously been tested. Effective clinical interpretation of these studies requires considerable skill and experience.
Metastatic dissemination is the leading cause of death in cancer patients, which is particularly evident for high-risk sarcomas such as Ewing sarcoma, osteosarcoma, and rhabdomyosarcoma. Previous ...research identified a crucial role for YB-1 in the epithelial-to-mesenchymal transition (EMT) and metastasis of epithelial malignancies. Based on clinical data and two distinct animal models, we now report that YB-1 is also a major metastatic driver in high-risk sarcomas. Our data establish YB-1 as a critical regulator of hypoxia-inducible factor 1α (HIF1α) expression in sarcoma cells. YB-1 enhances HIF1α protein expression by directly binding to and activating translation of HIF1A messages. This leads to HIF1α-mediated sarcoma cell invasion and enhanced metastatic capacity in vivo, highlighting a translationally regulated YB-1-HIF1α axis in sarcoma metastasis.
Display omitted
•YB-1 expression is elevated in high-risk human sarcomas•YB-1 promotes sarcoma invasion and metastasis•YB-1 regulates HIF1α expression by directly promoting its mRNA translation•YB-1 effects on sarcoma invasion and metastasis are mediated by HIF1α
YB-1 binds DNA and RNA and has been shown to promote epithelial-to-mesenchymal transition and metastasis of carcinomas. El-Naggar et al. show that YB-1 also contributes to metastasis of high-risk sarcomas by binding to HIF1A mRNA and enhancing its translation.
Ewing sarcoma and other peripheral primitive neuroectodermal tumors (pPNETs) display limited neural differentiation and are thought to have a neural crest origin. Greater than 95% of these tumors ...share common t(11;22) (q24;q12) or t(21;22) (q22;q12) chromosomal translocations leading to
EWS/FLII or
EWS/ERG gene fusions, respectively. The resulting chimeric oncoproteins seem to function as aberrant transcription factors. However, whether these molecules contribute to the limited neural differentiation observed in pPNETs or actually inhibit differentiation remains unclear. We report a Ewing sarcoma case from the forearm of a 10-year-old girl which expressed
EWS/FLI1 fusion transcripts. The tumor was treated with surgery, chemotherapy, and local radiation, but residual tumor was detected within a year as a well-differentiated peripheral neural tumor lacking detectable
EWS/FLI1 expression. Further studies suggested that the primary and residual tumors were clonally related. This association between apparent therapy-induced differentiation in Ewing sarcoma and absence of detectable fusion transcripts in the residual tumor provides presumptive evidence that
EWS/FLI1 expression may inhibit differentiation in tumour cells.
Pediatric pre-B acute lymphoblastic leukemia is the most common childhood cancer. Although current therapies achieve a high rate of remission, relapse of pre-B ALL remains a significant clinical ...challenge and new forms of therapy are needed. The graft-vs-leukemia (GVL) effect after bone marrow transplantation has shown that the immune system is capable of producing an effective anti-tumor response, which suggests that immune-mediated therapies may provide a complementary treatment strategy. Toll-like receptors (TLR), found on many immune cells, including B cells, have been shown to be important molecules in both innate and adaptive immune responses. Ligation of TLRs with their respective ligands results in an increase in the immunogenicity of antigen presenting cells (APC), through upregulation of MHC antigens and costimulatory molecules and production of cytokines and chemokines. We have previously reported that stimulation of pre-B ALL cells with the TLR9 ligand, CpG DNA, enhances the induction of Th1 immune responses by allogeneic T cells. In this study we examined the expression profile of TLRs 1-8 in precursor-B ALL cells and the effects of TLR ligation on the immune stimulatory capacity of precursor-B ALL cell lines. Eight precursor-B ALL cell lines were used in this study (Nalm6, REH, Sup-B15, KOPN-57bi, 380, 697, OP-1 and RS4:11). Standard RT-PCR analysis was used to examine the expression of TLRs 1-8 by the cell lines. The cell lines were stimulated with ligands for TLR2 (peptidoglycan), TLR3 (poly I:C) and TLR4 (LPS) and evaluated for changes in costimulatory molecule expression and allogeneic T cell stimulation. Non-quantatative PCR detected each TLR, with the exception of TLR8, in the majority of the cell lines. TLR8 was only detected, at low level, in RS4:11 cells. Despite the broad expression profile of the TLRs, significant differences in the effect of TLR ligation were observed between cell lines. In general, only modest increases in CD40 and CD86 expression were observed on responsive cell lines, with the majority of the lines showing no significant changes in response to TLR2, 3 or 4 ligation, despite receptor detection by PCR. Changes in allogeneic T cell proliferation in response to TLR stimulated ALL cell lines were observed, with the largest increases occurring with peptidoglycan and LPS. As was the case with costimulatory molecule expression, no T cell proliferative response change was common to all cell lines. However, analysis of cytokine production by T cells revealed a consistent increase in IFN-gamma and reduction in IL-5 levels in response to peptidoglycan stimulated ALL cells. The results reported in this study indicate that precursor-B ALL cell lines express several TLR molecules and that TLR ligation alters the immunogenicity of the majority of these lines. Interestingly, ligation of TLR2 with peptidoglycan induced a profound shift towards Th1 cytokine production. These observatios suggest that TLR ligation, most notably TLR2 ligation, may provide a strategy to influence anti-ALL immune responses and enhance immune mediated control of this disease.
Synovial sarcomas (SS) are characterized by the t(X;18)(p11;q11) translocation and its resultant fusion gene,
SYT-SSX. Two homologues of the
SSX gene (ie,
SSX1 and
SSX2) are involved in the vast ...majority of SS and the
SYT-SSX1 type of fusion has been associated with inferior clinical outcome. Thus, detection of the presence and type of
SYT-SSX fusion is critical for diagnosis and prognosis in SS. Identification of
SYT-SSX fusion type is typically accomplished by reverse-transcription polymerase chain reaction (RT-PCR) followed by a post-PCR analytic method. As mRNA nucleotide sequences of the SSX1 and SSX2 segments involved in the
SYT-SSX fusion are nearly identical, post-PCR methods must be highly discriminatory. We describe a novel method to identify and differentiate these two chimeric transcripts using RT-PCR followed by fluorescent thermostable ligase detection reaction (f-LDR), microparticle bead capture and flow cytometric detection. Evaluation of this unique approach in 11 cases of SS without prior knowledge of
SYT-SSX status, six cases of control sarcomas (CS) and three hematopoietic cell lines, revealed that the f-LDR technique was rapid, unambiguous, and highly specific. The f-LDR results were compared to
XmnI enzyme digestion patterns and sequencing of PCR products, revealing a 100% concordance for all cases of SS with regards to SYT-SSX transcript type. In addition, there was a strong association of transcript type detected by f-LDR and morphological subclassification of SS, as previously reported. We conclude that this f-LDR method with flow-based detection is a robust approach to post-PCR detection of specific nucleotide sequences in SS and may be more broadly applicable in molecular oncology.
We report two cases of intra-abdominal desmoplastic small round cell tumor with characteristic clinical, histological, immunohistochemical, and ultrastructural features. Fusion of the EWS gene on ...chromosome 22 and the WT1 gene on chromosome 11, resulting from the chromosomal translocation t(11;22)(p13;q12), was detected by reverse transcriptase polymerase chain reaction (RT-PCR) in both cases. This translocation has been previously reported in this type of tumor using either cytogenetic or molecular biological techniques. Tumor tissue from both cases revealed no chimeric fusion transcripts characteristic of the Ewing sarcoma family of peripheral primitive neuroectodermal tumors or of alveolar rhabdomyosarcoma, two tumors in the differential diagnosis of intra-abdominal desmoplastic small round cell tumor. This report demonstrates the utility of molecular studies as an adjunct in the diagnosis of this rare and aggressive tumor.
PDF
