C-myc Dysregulation Research Articles

Obesity under the moonlight of c-MYC.

The moonlighting protein c-Myc is a master regulator of multiple biological processes including cell proliferation, differentiation, angiogenesis, apoptosis and metabolism. It is constitutively and aberrantly expressed in more than 70% of human cancers. Overwhelming evidence suggests that c-Myc dysregulation is involved in several inflammatory, autoimmune, metabolic and other non-cancerous diseases. In this review, we addressed the role of c-Myc in obesity. Obesity is a systemic disease, accompanied by multi-organ dysfunction apart from white adipose tissue (WAT), such as the liver, the pancreas, and the intestine. c-Myc plays a big diversity of functions regulating cellular proliferation, the maturation of progenitor cells, fatty acids (FAs) metabolism, and extracellular matrix (ECM) remodeling. Moreover, c-Myc drives the expression of a wide range of metabolic genes, modulates the inflammatory response, induces insulin resistance (IR), and contributes to the regulation of intestinal dysbiosis. Altogether, c-Myc is an interesting diagnostic tool and/or therapeutic target in order to mitigate obesity and its consequences.

Network and Computational Drug Repurposing Analysis for c-Myc Inhibition in Burkitt Lymphoma.

The treatment rate of Burkitt lymphoma (BL) is still low in low-income countries and among elderly patients. The c-Myc dysregulation induced by mutations is one of the characteristics of BL. However, studies on the downstream signaling pathways of c-Myc are still lacking. This study aimed to identify the signaling pathways regulated by c-Myc. Network and gene set analyses using c-Myc inhibition (i.e., c-Myc knock-down and c-Myc inhibitor treatment) transcriptome datasets for BL cell lines were performed to determine the pathways regulated by c-Myc. In addition, computational drug repurposing was used to identify drugs that can regulate c-Myc downstream signaling pathway. Computational drug repurposing revealed that the ERK/MAPK signaling pathway is regulated by c-Myc in BL and that this pathway can be modulated by vorinostat. Furthermore, in the pharmacogenomics database, vorinostat showed a cell viability half-maximal inhibitory concentration of less than 2 μM in the BL cell lines. The downstream signaling pathway regulated by c-Myc and the drug that can modulate this pathway is presented for the first time.

Hnrnpk Overexpression Drives Nucleolar Aberrancies Causing Ribosomopathies

Background: Protein biogenesis is a complex process involving nucleoli and ribosomes. Alterations in any step could lead to alterations in ribosome functionality and protein synthesis. Hnrnpk is an RNA-binding protein (RBP) involve in these processes, finding that an overexpression (OE) produces nucleus and nucleolar stress (NS), decreases transcription, and drives an imbalance in ribosome biogenesis, causing a reduced translation. Aims: To elucidate how hnRNP K dysregulation affects the hematopoietic stem cell (HSCs) biology. Methods: To study the impact of Hnrnpk OE in vivo, we developed an inducible tamoxifen mouse model, HnrnpkTg/hUbc-CreERT2. Survival was evaluated by Kaplan-Meier, phenotype was described by symptoms/signs, CBC, bone marrow (BM) H/E, IHC and FCM analysis, and serum IL-6 ELISA. HSCs were cultured to study the impact of Hnrnpk OE in the HSCs dynamics. Hnrnpk OE was established in vitro using CRISPR/SAM. RNA-seq analysis was performed in a single read 85-base format and analyzed with DESeq2. TMT-based deep proteome profiling was also performed. Both were GSEA preranked. Transcription and translation were tested using Click-it RNA and HPG kit respectively, and translation efficiency by polysome assay. NS were analyzed by confocal microscopy and transmission electron microscopy (TEM). Protein-protein interaction between Hnrnpk and Ncl was studied by IP. Possible phenotype rescue was carried using HnrnpkTg/hUbc-CreERT2/ c-Myclox/wt, HnrnpkTg/hUbc-CreERT2/ Tp53lox/wt and HnrnpkTg/hUBC-CreERT2/ NclKDin vitro and in vivo models. Cell cycle FACS, senescence assays and karyotyping were performed. Molecular mechanism was elucidated by qRT-PCR and WB. Results: Hnrnpk Tg/hUbc-CreERT2 mice had widespread Hnrnpk OE and lifespan's reduction. By CBC, we found the development of leukopenia, lymphopenia, anaemia and thrombocytopenia (Fig.A). BM H/E, IHC and FCM showed a reduction of B220 + and CD34 + and Sca1 + HSCs, and an increment in myeloid cells (Fig. B). Also, we found higher senescent β-galactosidase expression in BM and IL-6 in vivo (Fig.C). Then, we found a decay in viability and an exhaustion in HSCs (Fig.D). To understand Hnrnpk implication in BM failure phenotype in vivo, we generated Hnrnpk OE cells (Fig.E). RNA-seq showed an upregulation in G2/M-checkpoint pathway related molecules (Fig.F), confirmed by FACS analysis, showing and increment of arrested G2/M phase-cells (Fig.G). Moreover, we showed a rise in β-galactosidase activity, polyploidy and genomic instability (Fig. H), linked to an increment in p21 and p16 (Fig.I). Then, TEM revealed nucleolar alterations in the in vitro model, including segregation, anormal accumulations or fragmentation of nucleolar components, alterations validated by confocal microscopy (Fig.J-K). Also, we found a Ncl increment in vitro, consistent with the protein-protein interaction between Hnrnpk and Ncl. Then, proteomics showed that Hnrnpk OE correlates with ribosome biogenesis regulators, c-Myc and mTOR dysregulation (Fig.L). Moreover, we found a decrease in transcription (Fig.M), consistent with rRNAs reduction, driving translation and protein synthesis deficiency (Fig.N). There was a partial reversion of NS hallmarks in HnrnpkTg/hUbc-CreERT2/ c-Myclox/wt, HnrnpkTg/hUbc-CreERT2/ Tp53lox/wt and HnrnpkTg/hUBC-CreERT2/ NclKD MEFs in vitro models and partially rescue phenotype in vivo (Fig.O). Finally, we focused on the nucleolus. Thus, Hnrnpk OE cells showed a reduction in Fbl, increase in Ncl and Ncl diffusion in nucleoplasm. All these data suggest the existence of a ribosompathy-like phenotype. Conclusions: this work found that RBPs dysregulation such as Hnrnpk OEdrives BM failure phenotype, promoting the exhaustion of HSCs by nucleoulus/ribosome dysregulation that triggers cell cycle arrest and apoptosis, dependent of p53. We therefore suggest that Hnrnpk induces ribosome dysfunction consistent with some types of ribosomopathies. This work was financially supported by CRIS contra el Cancer Association (NGO) AES ISCIII (PI18/00295, PI21/00191), ISCIII Miguel Servet (CP19/00140), Cancer Research UK [C355/A26819], Sociedad Española de Hematología y Hemoterapia (SEHH), FC AECC and AIRC under the Accelerator Award Program and National Cancer Institutes of Health Award (R01CA207204, SMP) Leukemia and Lymphoma Society (6577-19, SMP).

LncRNA BCAN-AS1 stabilizes c-Myc via N6-methyladenosine-mediated binding with SNIP1 to promote pancreatic cancer.

C-Myc overexpression contributes to multiple hallmarks of human cancer but directly targeting c-Myc is challenging. Identification of key factors involved in c-Myc dysregulation is of great significance to develop potential indirect targets for c-Myc. Herein, a collection of long non-coding RNAs (lncRNAs) interacted with c-Myc is detected in pancreatic ductal adenocarcinoma (PDAC) cells. Among them, lncRNA BCAN-AS1 is identified as the one with highest c-Myc binding enrichment. BCAN-AS1 was abnormally elevated in PDAC tumors and high BCAN-AS1 level was significantly associated with poor prognosis. Mechanistically, Smad nuclear-interacting protein 1 (SNIP1) was characterized as a new N6-methyladenosine (m6A) mediator binding to BCAN-AS1 via recognizing its m6A modification. m6A-modified BCAN-AS1 acts as a scaffold to facilitate the formation of a ternary complex together with c-Myc and SNIP1, thereby blocking S phase kinase-associated protein 2 (SKP2)-mediated c-Myc ubiquitination and degradation. Biologically, BCAN-AS1 promotes malignant phenotypes of PDAC in vitro and in vivo. Treatment of metastasis xenograft and patient-derived xenograft mouse models with in vivo-optimized antisense oligonucleotide of BCAN-AS1 effectively represses tumor growth and metastasis. These findings shed light on the pro-tumorigenic role of BCAN-AS1 and provide an innovant insight into c-Myc-interacted lncRNA in PDAC.

The Bivalent Bromodomain Inhibitor MT-1 Inhibits Prostate Cancer Growth.

Bromodomains (BD) are epigenetic readers of histone acetylation involved in chromatin remodeling and transcriptional regulation of several genes including protooncogene cellular myelocytomatosis (c-Myc). c-Myc is difficult to target directly by agents due to its disordered alpha helical protein structure and predominant nuclear localization. The epigenetic targeting of c-Myc by BD inhibitors is an attractive therapeutic strategy for prostate cancer (PC) associated with increased c-Myc upregulation with advancing disease. MT-1 is a bivalent BD inhibitor that is 100-fold more potent than the first-in-class BD inhibitor JQ1. MT-1 decreased cell viability and causes cell cycle arrest in G0/G1 phase in castration-sensitive and resistant PC cell lines in a dose-dependent fashion. The inhibition of c-Myc function by MT-1 was molecularly corroborated by the de-repression of Protein Kinase D1 (PrKD) and increased phosphorylation of PrKD substrate proteins: threonine 120, serine 11, and serine 216 amino acid residues in β-Catenin, snail, and cell division cycle 25c (CDC25c) proteins, respectively. The treatment of 3D cell cultures derived from three unique clinically annotated heavily pretreated patient-derived PC xenografts (PDX) mice models with increasing doses of MT-1 demonstrated the lowest IC50 in tumors with c-Myc amplification and clinically resistant to Docetaxel, Cabazitaxel, Abiraterone, and Enzalutamide. An intraperitoneal injection of either MT-1 or in combination with 3jc48-3, an inhibitor of obligate heterodimerization with MYC-associated protein X (MAX), in mice implanted with orthotopic PC PDX, decreased tumor growth. This is the first pre-clinical study demonstrating potential utility of MT-1 in the treatment of PC with c-Myc dysregulation.

P1285: IN VIVO IMPACT OF ABERRANT OVEREXPRESSION OF C-REL NF-ΚB SUBUNIT IN GERMINAL CENTER B-LYMPHOCYTES: A TUMOR ONCOGENIC EVENT IN GCB-DLBCLS

Background: Diffuse large B-cell lymphomas (DLBCLs) are the most NF-κB-related aggressive non-Hodgkin’s lymphomas. DLBCLs are clinically, molecularly and genetically heterogeneous, suggesting different oncogenic mechanisms. Gene-expression profiling studies led to the proposal of two main cell-of-origin molecular subtypes known as germinal center B-cell-like (GCB) DLBCLs, and activated B-cell like (ABC) DLBCLs. With the Lymphoma Study Association LYSA, we previously published that strong c-Rel DNA-binding activity was mostly found in GCBs and was associated with increased REL mRNA expression. These cases with both overexpression of REL mRNA and c-Rel related gene expression profile were correlated with various recurrent GCB-DLBCLs genetic events, including REL gains, BCL2 translocation, MEF2B, EZH2, CREBBP, and TNFRSF14 mutations and with the EZB GCB genetic subtype. Aims: We propose that c-Rel is a major oncogene of GCB-DLBCLs. We speculate that c-Rel confers a selective advantage in the germinal center (GC)-specific environment. We wish to study the consequences of aberrant overexpression and activation of c-Rel in vivo that could promote tumorigenesis in DLBCL precursor cells by disrupting the normal dynamics of B-cell development in the GC. Methods: We have generated the fl_STOP-REL mouse model: the murine REL transgene preceded by a STOP-Neomycin cassette flanked of loxP sites was introduced in Rosa26 locus on chromosome 6. The REL transgene is followed by an IRES to allow YFP marker concomitant expression. To reach GC B-cell specific expression, we used AID_Cre-ert2 mice: the Cre-ert2 transgene was at the AICDA gene locus on chromosome 6. On the same chromosome, the tdTomato marker preceded by a STOP cassette flanked by loxP sites was introduced at the Rosa26 locus. A pool of REL overexpressing GC B-cell was generated after tamoxifen administration and T-cell-dependent antigen stimulation with sheep red blood cells (SRBC). The fate of these YFP and tdTomato positive GC B-cells was followed over-time until 8 months. The first four months, a SRBC immune booster was performed to maintain the GCs. Results: As expected, our model fl_STOP-REL | AID_Cre-ert2 (i.e. REL mice) allowed for in vivo competition between c-Rel overexpressing (tdTomato+/YFP+) and normal (tdTomato+/YFP-) GC B-cells. In blood, the percentage of YFP positive B-lymphocytes among the tdTomato positive cells was increased over-time: 30% at 14 days, 65% at 2 months and about 80% at 8 months. From blood, spleen and lymph nodes, phenotyping analysis by flow cytometry of REL/YFP positive B-cells indicated an activated profile compared to normal GC-derived B-cells: CD19+, IgM+, IgD-, CD21+, CD80+, and B220low. At 8 months after generation of GC B-cells pool, approximatively 3-5% of B-cells were tdTomato and YFP positives in spleen of REL mice, whereas tdTomato positives cells in control mice were rare hardly detectable (less than 0.8%). Altogether, our result indicated that c-Rel reprograms B-cell to be activated and to better survive in vivo for very long time. Image:Summary/Conclusion: We bring strong experimental arguments showing that increased c-Rel expression in GC B-cell favors their survival for several months, and would thus be a primary event. We suspect that additional oncogenic events are probably needed for tumor clonal expansion such as BCL2, EZH2, CREBBP, TNFRSF14 and/or c-Myc dysregulation.

Prognostic significance of C-MYC dysregulation in esophageal squamous cell carcinoma: a meta-analysis.

Aim: The clinicopathological and prognostic significance of C-MYC dysregulation (amplification or overexpression) in esophageal squamous cell carcinoma(ESCC)remains controversial. Therefore, we performed this meta-analysis to elucidate this relationship. Materials & methods: Available studies were retrieved from PubMed, Web of Science, EMBASE and the Cochrane Library, and ten studies with a total of 1432 patients were included in this meta-analysis. Results: Pooled results showed that C-MYC dysregulation was significantly associated with poor overall survival(hazard ratio: 1.405 [95% CI: 1.170-1.639]; p < 0.001) and lymph node metastasis (odds ratio: 1.798 [95% CI: 1.125-2.873]; p = 0.014). Subgroup analysis confirmed the results and more prominent predictive effects were observed in the C-MYC amplification group. Conclusion: C-MYC dysregulation is a promising biomarker for ESCC prognosis.

Anticancer response to disulfiram may be enhanced by co-treatment with MEK inhibitor or oxaliplatin: modulation by tetrathiomolybdate, KRAS/BRAF mutations and c-MYC/p53 status.

Ammonium tetrathiomolybdate (TTM) and disulfiram (DSF) are copper (Cu) chelators in cancer clinical trials partly because Cu chelation: a) restricts the activity of Cu-binding MEK1/2 enzymes which drive tumourigenesis by KRAS or BRAF oncogenic mutations and b) enhances uptake of oxaliplatin (OxPt), clinically used in advanced KRAS-mutant colorectal carcinomas (CRC). Whereas TTM decreases intracellular Cu trafficking, DSF can reach other Cu-dependent intracellular proteins. Since the use of individual or combined Cu chelation may help or interfere with anti-cancer therapy, this study investigated whether TTM modifies the response to DSF supplemented with: 1) UO126, a known MEK1/2 inhibitor; 2) other Cu chelators like neocuproine (NC) or 1, 10-o-phenanthroline (OPT) in wt p53 melanoma cells differing in BRAF or KRAS mutations; 3) OxPt in mutant p53 CRC cells devoid of KRAS and BRAF mutations or harbouring either KRAS or BRAF mutations. TTM was not toxic against V600E-mut-BRAF A375 and G12D-mut-KRAS/high c-myc C8161 melanoma cells. Moreover, TTM protected both melanoma types from toxicity induced by DSF, NC and co-treatment with sub-lethal levels of DSF and the MEK inhibitor, UO126. Toxicity by co-treatment with DSF+OPT was poorly reversed by TTM in C8161 melanoma cells. In contrast to the greater toxicity of 0.1 μM DSF against mutant p53 CRC cells irrespective of their KRAS mutation, TTM did not protect G12V-mut-KRAS/high c-myc SW620 CRC from DSF+OxPt compared to KRAS-WT/BRAF-WT Caco-2 CRC. Our results show that DSF co-treatment with: a) MEK inhibitors may enhance tumour suppression; b) OxPt in CRC may counteract impaired response to cetuximab by KRAS/BRAF mutations and c) as a single treatment, TTM may be less effective than DSF and decreases the efficacy of the latter.HighlightsPotentiation of melanoma antitumour toxicity of DSF by MEK inhibitor is reversed by TTM.KRAS/c-MYC dysregulation attenuates TTM reversion of melanoma toxicity by DSF + OPT.KRAS/c-MYC dysregulation increases melanoma NC toxicity reversed by TTM.BRAF mutation and lower c-MYC may attenuate toxicity by DSF ± OxPt in colorectal cancer cells

The Role of c-MYC in B-Cell Lymphomas: Diagnostic and Molecular Aspects.

c-MYC is one of the most essential transcriptional factors, regulating a diverse array of cellular functions, including proliferation, growth, and apoptosis. Dysregulation of c-MYC is essential in the pathogenesis of a number of B-cell lymphomas, but is rarely reported in T-cell lymphomas. c-MYC dysregulation induces lymphomagenesis by loss of the tight control of c-MYC expression, leading to overexpression of intact c-MYC protein, in contrast to the somatic mutations or fusion proteins seen in many other oncogenes. Dysregulation of c-MYC in B-cell lymphomas occurs either as a primary event in Burkitt lymphoma, or secondarily in aggressive lymphomas such as diffuse large B-cell lymphoma, plasmablastic lymphoma, mantle cell lymphoma, or double-hit lymphoma. Secondary c-MYC changes include gene translocation and gene amplification, occurring against a background of complex karyotype, and most often confer aggressive clinical behavior, as evidenced in the double-hit lymphomas. In low-grade B-cell lymphomas, acquisition of c-MYC rearrangement usually results in transformation into highly aggressive lymphomas, with some exceptions. In this review, we discuss the role that c-MYC plays in the pathogenesis of B-cell lymphomas, the molecular alterations that lead to c-MYC dysregulation, and their effect on prognosis and diagnosis in specific types of B-cell lymphoma.

A c-Myc-regulated stem cell-like signature in high-risk neuroblastoma: A systematic discovery (Target neuroblastoma ESC-like signature)

c-Myc dysregulation is hypothesized to account for the ‘stemness’ – self-renewal and pluripotency – shared between embryonic stem cells (ESCs) and adult aggressive tumours. High-risk neuroblastoma (HR-NB) is the most frequent, aggressive, extracranial solid tumour in childhood. Using HR-NB as a platform, we performed a network analysis of transcriptome data and presented a c-Myc subnetwork enriched for genes previously reported as ESC-like cancer signatures. A subsequent drug-gene interaction analysis identified a pharmacogenomic agent that preferentially interacted with this HR-NB-specific, ESC-like signature. This agent, Roniciclib (BAY 1000394), inhibited neuroblastoma cell growth and induced apoptosis in vitro. It also repressed the expression of the oncogene c-Myc and the neural ESC marker CDK2 in vitro, which was accompanied by altered expression of the c-Myc-targeted cell cycle regulators CCND1, CDKN1A and CDKN2D in a time-dependent manner. Further investigation into this HR-NB-specific ESC-like signature in 295 and 243 independent patients revealed and validated the general prognostic index of CDK2 and CDKN3 compared with CDKN2D and CDKN1B. These findings highlight the very potent therapeutic benefits of Roniciclib in HR-NB through the targeting of c-Myc-regulated, ESC-like tumorigenesis. This work provides a hypothesis-driven systems computational model that facilitates the translation of genomic and transcriptomic signatures to molecular mechanisms underlying high-risk tumours.

C-Myc dysregulation is a co-transforming event for nuclear factor-κB activated B cells.

While c-Myc dysregulation is constantly associated with highly proliferating B-cell tumors, nuclear factor (NF)-κB addiction is found in indolent lymphomas as well as diffuse large B-cell lymphomas, either with an activated B-cell like phenotype or associated with the Epstein-Barr virus. We raised the question of the effect of c-Myc in B cells with NF-κB activated by three different inducers: Epstein-Barr virus-latency III program, TLR9 and CD40. Induction of c-Myc overexpression increased proliferation of Epstein-Barr virus-latency III immortalized B cells, an effect that was dependent on NF-κB. Results from transcriptomic signatures and functional studies showed that c-Myc overexpression increased Epstein-Barr virus-latency III-driven proliferation depending on NF-κB. In vitro, induction of c-Myc increased proliferation of B cells with TLR9-dependant activation of MyD88, with decreased apoptosis. In the transgenic λc-Myc mouse model with c-Myc overexpression in B cells, in vivo activation of MyD88 by TLR9 induced splenomegaly related to an increased synthesis phase (S-phase) entry of B cells. Transgenic mice with both continuous CD40 signaling in B cells and the λc-Myc transgene developed very aggressive lymphomas with characteristics of activated diffuse large B-cell lymphomas. The main characteristic gene expression profile signatures of these tumors were those of proliferation and energetic metabolism. These results suggest that c-Myc is an NF-κB co-transforming event in aggressive lymphomas with an activated phenotype, activated B-cell like diffuse large B-cell lymphomas. This would explain why NF-κB is associated with both indolent and aggressive lymphomas, and opens new perspectives on the possibility of combinatory therapies targeting both the c-Myc proliferating program and NF-κB activation pathways in diffuse large B-cell lymphomas.

Renal Cell Carcinomas in Vinylidene Chloride-exposed Male B6C3F1 Mice Are Characterized by Oxidative Stress and TP53 Pathway Dysregulation.

Vinylidene chloride (VDC) has been widely used in the production of plastics and flame retardants. Exposure of B6C3F1 mice to VDC in the 2-year National Toxicology Program carcinogenicity bioassay resulted in a dose-dependent increases in renal cell hyperplasia, renal cell adenoma, and renal cell carcinomas (RCCs). Among those differentially expressed genes from controls and RCC of VDC-exposed mice, there was an overrepresentation of genes from pathways associated with chronic xenobiotic and oxidative stress as well as c-Myc overexpression and dysregulation of TP53 cell cycle checkpoint and DNA damage repair pathways in RCC. Trend analysis comparing RCC, VDC-exposed kidney, and chamber control kidney showed a conservation of pathway dysregulation in terms of overrepresentation of xenobiotic and oxidative stress, and DNA damage and cell cycle checkpoint pathways in both VDC-exposed kidney and RCC, suggesting that these mechanisms play a role in the pathogenesis of RCC in VDC-exposed mice.

A Phase 2 Trial of AZD1152 in Relapsed/Refractory Diffuse Large B-Cell Lymphoma

IntroductionDiffuse large B-cell lymphoma (DLBCL) is the most common non-Hodgkin lymphoma (NHL) and has an aggressive natural history. R-CHOP has improved patient (pt) survival and established a standard of first line care. In the rituximab era the prognosis for relapsed DLBCL is poor, with salvage chemotherapy and autologous stem cell transplantation (ASCT) only curing a small proportion (<25% of all relapses) of those with adequate performance status. Relapsed DLBCL in those unfit for ASCT or post-ASCT is rarely curable, and remains an unmet clinical need.AZD1152 (barasertib) is a highly selective and potent Aurora B kinase inhibitor. Aurora B kinase has an important function at the spindle assembly check-point in mitosis. The kinase is overexpressed in 48% of DLBCL. Exposure of high grade lymphoma B cells to AZD1152-HQPA in vitropotently induced growth arrest and apoptosis via caspase-dependent and independent mechanisms.AZD1152 was well tolerated in a phase I study in relapsed/refractory acute myeloid leukaemia. We performed a phase II single arm open-label trial to determine the efficacy and tolerability of AZD1152 in relapsed/refractory DLBCL pts who had failed or were ineligible for salvage/high dose chemotherapy and had received at least one line of potentially curative immunochemotherapy.MethodsPts with relapsed/refractory (de novoor transformed indolent NHL) DLBCL with measurable disease (at least a non-irradiated single lesion (>1.5cm)) were treated with up to six cycles of 96-hour continuous intravenous (iv) infusions of AZD1152 as an inpt every 3 weeks.Due to concerns over possible precipitation of drug in the line, regular 4 hourly saline flushes were required via each lumen to maintain patency. Prophylactic G-CSF was permissible per ASCO guidelines. Ongoing cycles were commenced when neutrophils recovered to ≥1.0x109/L and platelets to ≥50x109/L.The primary endpoint was overall response rate (ORR). Secondary end-points were progression-free survival (PFS) at one year, % change in tumour size, and safety measured by the incidence and severity of adverse events (AEs). Response assessment using FDG-PET was completed at day 15 of cycle 2. Pts with stable disease (SD), minimal, partial (PR) or complete response (CR) according to Cheson criteria remained on protocol, but those with progressive disease (PD) were taken off study.As data that suggests MYC overexpression may enhance responses to aurora B kinase inhibition, MYC by IHC and FISH for c-MYC translocation was performed. All diagnostic specimens were reviewed by specialist haemato-pathologists.ResultsFifteen pts were enrolled in 20 months, and received 1-6 cycles. Overall, 42 cycles were administered (41 at full dose). One pt completed all six planned cycles. The cohort included 8 males and 7 females, with a median age of 65 (35-74 years). 13% had a low International Prognostic Index (IPI) (0-1), 53% an intermediate IPI (2-3) and 33% a high IPI (4-5). The best ORR was 20% (see Fig 1 for tumour percentage reduction) with no cases of CR and 3 PR. 33% obtained SD for between 2-6 cycles. The median PFS was 60 days (95% CI 36-84 days) (see Fig 2). IPI (p=0.584) (See Fig 3), gender (p=0.113), disease bulk at diagnosis (p= 0.654) or c-MYC dysregulation (p=0.331) did not predict PFS. Early discontinuation occurred in 14 pts from PD.SafetyMost AEs were grade 1-2. 18 episodes of neutropenic fever occurred, including one grade 4 bacteraemia. There were no fatal SAEs and each episode resolved with iv antibiotics and G-CSF. All other toxicities were transient, manageable and expected. 13% of pts developed a total of 2 episodes of mucositis following 42 total doses. Neutropenia (grade 3-4), diarrhoea and nausea were common but manageable and expected. No Suspected Unexpected Serious Adverse Reactions occurred.ConclusionsThree-weekly AZD1152 was safely administered to pts with relapsed DLBCL. Responses were seen, providing a proof of concept that Aurora B kinase is a valid target in DLBCL. However, efficacy was modest. The infusate delivery was cumbersome. Dose intensity was maintained during treatment. Important but manageable AEs included neutropenic fever and mucositis. Aurora B kinase is a rational target for further investigation. NCT01354392. [Display omitted] [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

A statin‐regulated microRNA represses human c‐Myc expression and function

c-Myc dysregulation is one of the most common abnormalities found in human cancer. MicroRNAs (miRNAs) are functionally intertwined with the c-Myc network as multiple miRNAs are regulated by c-Myc, while others directly suppress c-Myc expression. In this work, we identified miR-33b as a primate-specific negative regulator of c-Myc. The human miR-33b gene is located at 17p11.2, a genomic locus frequently lost in medulloblastomas, of which a subset displays c-Myc overproduction. Through a small-scale screening with drugs approved by the US Food and Drug Administration (FDA), we found that lovastatin upregulated miR-33b expression, reduced cell proliferation and impaired c-Myc expression and function in miR-33b-positive medulloblastoma cells. In addition, a low dose of lovastatin treatment at a level comparable to approved human oral use reduced tumour growth in mice orthotopically xenografted with cells carrying miR-33b, but not with cells lacking miR-33b. This work presents a highly promising therapeutic option, using drug repurposing and a miRNA as a biomarker, against cancers that overexpress c-Myc.

Oncogenesis of T-ALL and nonmalignant consequences of overexpressing intracellular NOTCH1

Mutations resulting in overexpression of intracellular Notch1 (ICN1) are frequently observed in human T cell acute lymphoblastic leukemia (T-ALL). We have determined the consequences of ICN1 overexpression from retroviral vectors introduced into bone marrow cells. Early consequences are the generation of polyclonal nontumorigenic CD4+8+ T cell receptor (TCR)-αβ+ cells that do not qualify as tumor precursors despite the observation that they overexpress Notch 1 and c-Myc and degrade the tumor suppressor E2A by posttranslational modification. The first tumorigenic cells are detected among more immature CD4−8+TCR-αβ− cells that give rise to monoclonal tumors with a single, unique TCR-β chain and diverse TCR-α chains, pinpointing malignant transformation to a stage after pre-TCR signaling and before completion of TCR-α rearrangement. In T-ALL, E2A deficiency is accompanied by further transcriptional up-regulation of c-Myc and concomitant dysregulation of the c-Myc-p53 axis at the transcriptional level. Even though the tumors consist of phenotypically heterogeneous cells, no evidence for tumor stem cells was found. As judged by array-based comparative genomic hybridization (array CGH) and spectral karyotype (SKY) analysis, none of the tumors arise because of genomic instability.

C-Myc Overexpression Causes Anaplasia in Medulloblastoma

Both anaplasia and increased c-myc gene expression have been shown to be negative prognostic indicators for survival in medulloblastoma patients. myc gene amplification has been identified in many large cell/anaplastic medulloblastoma, but no causative link between c-myc and anaplastic changes has been established. To address this, we stably overexpressed c-myc in two medulloblastoma cell lines, DAOY and UW228, and examined the changes in growth characteristics. When analyzed in vitro, cell lines with increased levels of c-myc had higher rates of growth and apoptosis as well as significantly improved ability to form colonies in soft agar compared with control. When injected s.c. into nu/nu mice, flank xenograft tumors with high levels of c-myc in DAOY cell line background were 75% larger than those derived from control. Overexpression of c-myc was required for tumor formation by UW228 cells. Most remarkably, the histopathology of the Myc tumors was severely anaplastic, with large areas of necrosis/apoptosis, increased nuclear size, and macronucleoli. Indices of proliferation and apoptosis were also significantly higher in Myc xenografts. Thus, c-myc seems to play a causal role in inducing anaplasia in medulloblastoma. Because anaplastic changes are often observed in recurrent medulloblastoma, we propose that c-myc dysregulation is involved in the progression of these malignant embryonal neoplasms.

C-Myc inhibition negatively impacts lymphoma growth

Background Proto-oncogene c-Myc dysregulation is commonly found in aggressive tumors. Dysregulation is central to lymphomagenesis in Burkitt lymphoma and other non-Hodgkin's lymphomas. This suggests targeting c-Myc as a treatment for myc-associated malignancies. Methods Microarrays showed c-Myc dysregulation in a B-lymphoblastoid line, TIB-215. This was confirmed by real-time reverse transcriptase–polymerase chain reaction (RT-PCR) and extended to 3 additional Burkitt lymphoma lines. Growth effects of a c-Myc inhibitor, compound 10058-F4, were determined in these 4 lines using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide analyses and direct cell counts. Drug effects on c-Myc gene expression levels were measured using minor groove binding–TaqMan real-time reverse transcriptase–polymerase chain reaction. Drug specificity was analyzed in rat c-Myc knockout (−/−) and Myc-transfected cells. Results c-Myc dysregulation was shown to be cell-cycle independent without rapid decay of c-Myc mRNA levels in all 4 lines. Using a c-Myc inhibitor, we found that growth inhibition was time- and dose-dependent. This inhibition caused unexpected downregulation (≥65%) of c-Myc mRNAs. Conclusions The inhibition of c-Myc decreased growth in aggressive lymphoma cells. This mechanism included c-Myc mRNA downregulation and dissociation of c-Myc/Max protein heterodimer. These results support targeting c-Myc in tumors with high morbidity and mortality.

Telomerase activity and differential expression of telomerase genes and c-myc in chicken cells in vitro.

This study examined telomerase activity and gene expression profiles for three genes in Gallus gallus domesticus: telomerase reverse transcriptase (chTERT), telomerase RNA (chTR), and c-myc. Expression of these genes was studied in chicken embryonic stem (chES) cells, chicken embryo fibroblasts (CEFs), and DT40 cells using quantitative real-time polymerase chain reaction. Our results establish that, relative to transcription levels in telomerase-negative CEFs, chTERT and chTR are up-regulated in telomerase-positive chES cells. Transcription levels of chTERT, chTR, and c-myc are dramatically up-regulated in telomerase-positive DT40 cells, relative to CEFs and chES cells. These results are consistent with a model in which telomerase activity is up-regulated in proliferating embryonic stem cells requiring stable telomeres to endure multiple rounds of cell division; down-regulated in differentiated, lifespan-limited cells; and dramatically up-regulated in immortalized, transformed cells for which uncontrolled proliferation is correlated with c-myc dysregulation and telomerase activity.

Burkitt's and Burkitt-like lymphoma.

Burkitt's and Burkitt-like lymphoma (BL/BLL) are aggressive B-cell malignancies with a high proliferative rate that may be fatal within months if not treated promptly. Furthermore, treatment of BL/BLL requires comprehensive supportive care to avoid disease-related complications such as acute renal failure secondary to tumor lysis syndrome. Improvements in our understanding of the biology of BL and BLL have led to more effective therapeutic protocols. Clinical trials have demonstrated that short duration, multi-agent, dose-intensive chemotherapy regimens combined with aggressive central nervous system therapy results in long-term survival rates in children and young adults near 70% to 80%, whereas long-term disease-free survival rates in older adults remains suboptimal at 15% to 25%. Outcomes in HIV-associated BL/BLL are improved because of more effective chemotherapy regimens and enhanced HIV care. Autologous bone marrow transplantation has proven feasible in many patient populations with BL/BLL and may lead to cure in selected patients. Improved therapeutic strategies are warranted, such as integrating agents such as monoclonal antibodies to combination dose-intensive chemotherapy. Moreover, further study into the molecular biology of BL/BLL with attention to the role of c-myc dysregulation is needed to help predict prognostic factors and for the development of molecular targeted therapies. Clinical trials remain critical to determine the most effective treatment regimens that will continue to improve cure rates in this aggressive but treatable disease.

P14ARF homozygous deletion or MDM2 overexpression in Burkitt lymphoma lines carrying wild type p53.

The hallmark of Burkitt lymphoma (BL) is a constitutively activated c-myc gene that drives tumor cell growth. A majority of BL-derived cell lines also carry mutant p53. In addition, the p16INK4a promoter is hypermethylated in most BL biopsies and BL cell lines, leading to silencing of this gene. Activation of c-myc and/or cell cycle dysregulation can induce ARF expression and p53-dependent apoptosis. We therefore investigated the p14ARF-MDM2-p53 pathway in BL cell lines. p14ARF was expressed and localized to nucleoli in all BL carrying mutant p53. Three out of seven BL carrying wt p53 had a homozygous deletion of the CDKN2A locus that encodes both p14ARF and p16INK4a. Three BL carrying wild type p53 retained the CDKN2A locus and overexpressed MDM2. DNA sequencing revealed a point mutation in CDKN2A exon 2 in one of these BL, Seraphine. However, this point mutation did not affect p14ARF's nucleolar localization or ability to induce p53. The Bmi-1 protein that negatively regulates the p14ARF promoter and co-operates with c-myc in tumorigenesis was expressed at low to moderate levels in all BL analysed. Our results indicate that inactivation of the ARF-MDM2-p53 pathway is an essential step during the development of Burkitt lymphoma, presumably as a mechanism to escape c-myc induced apoptosis.