Enhanced CCR9 expression levels in psoriatic skin are associated with poor clinical outcome to infliximab treatment

Abstract Introduction Philadelphia Chromosome positive acute lymphoblastic leukemia (ALL) is associated with particularly poor clinical outcome. Fusion of the ABL1 oncogene to a breakpoint cluster region (BCR) results in constitutively activated BCR-ABL1 tyrosine kinase activity that can be specifically targeted by small molecule tyrosine kinase inhibitors (TKI) such as Imatinib. After initially good clinical response to TKI, BCR-ABL1+ ALL patients invariably relapse into a more aggressive, TKI resistant disease. FOXM1 belongs to the forkhead box transcription factor family and is a key regulator of cell growth by promoting cell cycle progression. While a functional role of the transcription factor FOXM1 in terms of disease progression and drug resistance is well established for solid tumors, it remains elusive whether it has a similar function in ALL. Results We have identified high FOXM1 protein expression levels in patient-derived BCR-ABL1+ ALL samples compared to healthy B cells and B cell precursors. Consistent with this finding, the FOXM1 promoter region was de-methylated in 83 BCR-ABL1+ ALL patient samples compared to normal pre-B cells and Lymphoma samples. We used a mouse-model for BCR-ABL1+ ALL to characterize the role of BCR-ABL1 in the induction of FOXM1 expression: we cultured murine bone marrow B cell precursors in the presence of IL7 and induced transformation with a retroviral BCR-ABL1 expression vector. Consistent with findings for other oncogenes, i.e. activating mutation of Ras, BCR-ABL1 expression increased levels of FOXM1 compared to the normal IL7-dependent pre-B cells. In order to evaluate a potential clinical relevance of FOXM1 expression levels in ALL disease progression, we correlated FOXM1 mRNA levels with clinical outcome. High FOXM1 expression levels correlate with poor clinical outcome of ALL. In addition, the group of Sanders et al. (Genome Biol., 2013) has recently identified a set of 38 genes that are directly regulated by FOXM1 and associated with poor prognosis in breast cancer. 34 of the described 38 FOXM1 target genes were associated with high risk in ALL based on a data set collected by the German ALL-REZ BFM 2002 of the Berlin-Frankfurt-Münster study group (n=60). To study the functional role of FOXM1 in BCR-ABL1+ ALL, we performed experimental genetic deletion experiments using bone-marrow cells derived from Foxm1fl/fl mice. Deletion of Foxm1 decreases cell viability, colony formation, and proliferative capacity in vitro as well as leukemia formation in vivo of BCR-ABL1-driven leukemia. FOXM1-deleted ALL cells revealed a strikingly higher sensitivity towards TKI-treatment compared to the control cells in Imatinib dose-response curves (IC50EV: 420 nM vs IC50 CreERT2: 160 nM) as well as annexin V staining. As FOXM1 is a critical regulator of oxidative responses, we analyzed the intracellular reactive oxygen species (ROS) formation in the presence and absence of FOXM1 in BCR-ABL1 transduced ALL cells and observed consistently higher ROS levels after FOXM1 deletion. As a functional mediator of this effect, we analyzed the expression of the described FOXM1 target and ROS scavenger Catalase and found reduced levels after FOXM1 deletion. We determined Catalase expression in ALL cells after Imatinib treatment and found significant transcriptional upregulation. To evaluate whether this effect is FOXM1-dependent, we treated ALL cells with Imatinib and evaluated Catalase expression in the presence and absence of FOXM1. Only the FOXM1 expressing ALL cells were capable of upregulating Catalase after Imatinib treatment. Single locus chromatin immunoprecipitation (ChIP) analysis of a described binding site of FoxM1 in intron 1 of the catalase gene showed specific binding of FOXM1, albeit low enrichment compared to the positive control cyclin B1. As potential therapeutic agents to target FOXM1, we evaluated the effects of a previously described ARF peptide and the natural occurring antibiotic Thiostrepton. Both bind FOXM1 and inhibit its function and induced apoptosis in Ph+ ALL. In line with our observation of the genetic deletion, we observed induction of ROS with the treatment of either substance along with a decrease in Catalase expression. Conclusion Taken together, our data identify FOXM1 as a valid therapeutic target for the treatment of TKI sensitive and resistant BCR-ABL1+ ALL – either together with TKI or as a single agent. Disclosures: No relevant conflicts of interest to declare.

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3’UTR Polymorphism in ACSL1 Gene Correlates with Expression Levels and Poor Clinical Outcome in Colon Cancer Patients

PLoS ONE ◽

10.1371/journal.pone.0168423 ◽

2016 ◽

Vol 11 (12) ◽

pp. e0168423 ◽

Cited By ~ 17

Author(s):

Teodoro Vargas ◽

Juan Moreno-Rubio ◽

Jesús Herranz ◽

Paloma Cejas ◽

Susana Molina ◽

...

Keyword(s):

Colon Cancer ◽

Clinical Outcome ◽

Cancer Patients ◽

Poor Clinical Outcome ◽

Expression Levels

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Identification Of BCL6 As a Therapeutic Target In MLL-Rearranged ALL

Blood ◽

10.1182/blood.v122.21.72.72 ◽

2013 ◽

Vol 122 (21) ◽

pp. 72-72

Author(s):

Christian Hurtz ◽

Huimin Geng ◽

Erica Ballabio ◽

Gang Xiao ◽

Carina Ng ◽

...

Keyword(s):

Clinical Outcome ◽

B Cell ◽

Direct Consequence ◽

Poor Clinical Outcome ◽

Aberrant Expression ◽

Childhood All ◽

Expression Levels ◽

Protein Levels ◽

Bcl6 Expression ◽

Chip Analysis

Abstract Background BCL6 has been identified in diffuse large B cell lymphoma (DLBCL), where it is frequently translocated to immunoglobulin loci and acts as a protoncogene and transcriptional repressor. We found in Ph+ acute lymphoblastic leukemia (ALL) that BCL6 is involved in a novel form of drug resistance to tyrosine kinase inhibitors (TKI) by protecting cells from p53-mediated apoptosis (Duy et al., Nature 2011). Our current study is focusing on the function of BCL6 in Non-Ph+ALL based on the finding that high BCL6 levels represent a predictor of poor clinical outcome. Results We analyzed the gene expression data from 207 children with high-risk B cell precursor ALL enrolled in the pediatric clinical trial (COG P9906) and found that high expression levels of BCL6 at the time of diagnosis correlated with a poor overall and relapse-free survival (OS p=0.007; RFS p=0.02). Furthermore, multivariate analyses showed that patients with high BCL6 expression levels and established predictors of poor clinical outcome such as high white blood counts (WBC), a positive minimal residual disease (MRD) or mutations in the tumor suppressor IKZF1 had a significantly poor OS and RFS. Matched sample pairs from 49 patients at diagnosis and relapse showed that BCL6 levels were increased at relapse compared to diagnosis (p=0.003). We next studied BCL6 protein levels in Non-Ph+ cell lines and childhood ALL patient samples and found that BCL6 levels are particularly high in MLL-AF4 ALL (n=19). Surprisingly, the patients from the clinical trial that had high BCL6 levels and MLLR rearrangements were those with the worst clinical outcome (OS p= 0.0009 and RFS p= 0.000208). We next tested if the MLL-AF4 oncogene drives aberrant BCL6 expression. First, we performed a ChIP-analysis with the oncoprotein MLL-AF4 and found that the BCL6 promoter is directly bound by MLL-AF4, suggesting that MLL-AF4 may indeed drive BCL6 expression. We then performed a BCL6 Western Blot of inducible MLL-AF4-transgenic pro-B cells, demonstrating that activation of the MLL-AF4 transgene is sufficient to induce ∼10-fold upregulation of BCL6 protein levels. We conclude that aberrant expression of BCL6 in childhood ALL can be the direct consequence of MLL-AF4 activity. To further elucidate the BCL6 signaling pathway, we performed a ChIP-analysis with human MLL-AF4 leukemia cells and found that tumor suppressor genes (e.g. CDKN1B and BACH2) and B cell linage specific genes like PAX5 and CD19 are repressed by BCL6, potentially explaining the mixed lineage phenotype of MLLR driven B-ALL. We used a genetic mouse model of childhood ALL based on bone marrow precursor cells from BCL6-/- mice to decipher the function of BCL6. Since mutations in the RAS pathway are found in about 30% of childhood ALL cases, we transduced B cell progenitor cells from BCL6-/- and BCL6+/+ mice with Nras using an inducible oncogenic TET-NRASG12D system. Similar to MLL-AF4 driven cells, activation of NRASG12D results in higher BCL6 protein expression. Strikingly, BCL6-deficiency results in a failure of NRASG12DALL cells to initiate leukemia. Clinical relevance To verify if the high BCL6 expression levels in MLL-AF4 patients are important for the disease progression, we transduced primary human childhood ALL xenografts with a dominant-negative BCL6-mutant (BCL6-DN). Expression of BCL6-DN rapidly induced cell cycle arrest and cell death. To test if pharmacological inhibition of BCL6 is of potential use for children with MLL-AF4 leukemia, we treated human MLL-AF4 driven primary human xenograft cells with a recently developed retro-inverso BCL6 peptide inhibitor (RI-BPI). Strikingly, treatment with RI-BPI not only compromised colony formation in methylcellulose it also prevents leukemia-initiation in transplant recipient mice. RI-BPI also had a strong synergistic effect when combined with the chemotherapy drug Vincristine, which represents the backbone for most high risk regimen in pediatric ALL. Conclusions These findings identify BCL6 as an important factor in leukemia initiation and survival and its pharmacological inhibition as a novel strategy to treat childhood ALL. Aberrant expression of BCL6 in MLL-AF4 ALL is the direct consequence of MLL-AF4 activity in these cells. Based on these findings, we propose combinations of BCL6 inhibitors (e.g. RI-BPI) with currently used chemotherapeutics as potential approach to reduce the risk of ALL relapse and improve overall outcome. Disclosures: No relevant conflicts of interest to declare.

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