Therapeutically targeting oncogenic CRCs facilitates induced differentiation of NB by RA and the BET bromodomain inhibitor.

Retinoic acids (RAs) are the most successful therapeutics for cancer differentiation therapy used in high-risk neuroblastoma (NB) maintenance therapy but are limited in effectiveness. This study identifies a strategy for improving efficacy through disruption of cancer cell identity via BET inhibitors. Mutations that block development are theorized to cause NB through retention of immature cell identities contributing to oncogenesis. NB has two interchangeable cell identities, maintained by two different core transcriptional regulatory circuitries (CRCs): a therapy-resistant mesenchymal/stem cell state and a proliferative adrenergic cell state. MYCN amplification is a common mutation of high-risk NB and recently found to block differentiation by driving high expression of the adrenergic CRC transcription factor ASCL1. We investigated whether disruption of immature CRCs can promote RA-induced differentiation since only a subset of NB patients responds to RA. We found that silencing ASCL1, a critical member of the adrenergic CRC, or global disruption of CRCs with the BET inhibitor JQ1, suppresses gene expression of multiple CRC factors, improving RA-mediated differentiation. Further, JQ1 and RA synergistically decrease proliferation and induce differentiation in NB cell lines. Our findings support preclinical studies of RA and BET inhibitors as a combination therapy in treating NB.

Inhibition of protein kinase C beta phosphorylation activates nuclear factor-kappa B and improves postischemic recovery in type 1 diabetes.

IMPACT STATEMENT: Diabetes worsens the outcomes of peripheral arterial disease (PAD) likely in part through inducing chronic inflammation. However, in PAD, recovery requires the nuclear factor-kappa B (NF-κB) activation, a known contributor to inflammation. Our study shows that individually, both ischemia and high glucose activate the canonical and non-canonical arms of the NF-κB pathways. We show for the first time that prolonged high glucose specifically impairs ischemia-induced activation of the canonical NF-κB pathway through activation of protein kinase C beta (PKCß). Accordingly, inhibition of PKCß restores the ischemia-induced NF-κB activity both in vitroin endothelial cells and in vivoin hind limbs of type 1 diabetic mice and improves perfusion recovery after experimental PAD. Thus, this study provides a mechanistic insight into how diabetes contributes to poor outcomes in PAD and a potential translational approach to improve PAD outcomes.

Dual specificity phosphatase 5 regulates perfusion recovery in experimental peripheral artery disease.

Peripheral artery disease (PAD) is caused by atherosclerotic occlusions of vessels outside the heart, particularly those of the lower extremities. Angiogenesis is one critical physiological response to vessel occlusion in PAD, but our understanding of the molecular mechanisms involved in angiogenesis is incomplete. Dual specificity phosphatase 5 (DUSP5) has been shown to play a key role in embryonic vascular development, but its role in post-ischemic angiogenesis is not known. We induced hind limb ischemia in mice and found robust upregulation of Dusp5 expression in ischemic hind limbs. Moreover, in vivo knockdown of Dusp5 resulted in impaired perfusion recovery in ischemic limbs and was associated with increased limb necrosis. In vitro studies showed upregulation of DUSP5 in human endothelial cells exposed to ischemia, and knockdown of DUSP5 in these ischemic endothelial cells resulted in impaired endothelial cell proliferation and angiogenesis, but did not alter apoptosis. Finally, we show that these effects of DUSP5 on post-ischemic angiogenesis are a result of DUSP5-dependent decrease in ERK1/2 phosphorylation and p21 protein expression. Thus, we have identified a role of DUSP5 in post-ischemic angiogenesis and implicated a DUSP5-ERK-p21 pathway that may serve as a therapeutic target for the modulation of post-ischemic angiogenesis in PAD.

Targeted Therapy for EBV-Associated B-cell Neoplasms.

Epstein-Barr virus (EBV) is directly implicated in several B-cell lymphoid malignancies. EBV-associated lymphomas are characterized by prominent activation of the NF-κB pathway and targeting this pathway establishes a rationale for a therapeutic approach. The ubiquitin/proteasome signaling plays an essential role in the regulation of the NF-κB pathway. Ixazomib is an FDA-approved, orally bioavailable proteasome inhibitor. Here we report the first preclinical evaluation of ixazomib-mediated growth-inhibitory effects on EBV-infected B-lymphoblastoid cell lines Raji and Daudi. Ixazomib induced apoptosis in these cell lines in a dose-dependent manner. Cell-cycle analysis demonstrated ixazomib treatment induced cell-cycle arrest at the G2-M phase with a concomitant decrease in G0-G1 and S phases. The results further revealed an increase in p53, p21, and p27 levels and a decrease in survivin and c-Myc protein levels. Mechanistically, ixazomib treatment resulted in the accumulation of polyubiquitinated proteins, including phosphorylated IκBα with a significant reduction of p65 subunit nuclear translocation. Altogether, our preclinical data support the rationale for in vivo testing of ixazomib in EBV-associated B-cell neoplasms. IMPLICATIONS: This preclinical study supports the use of oral proteasome inhibitor ixazomib for targeting NF-κB signaling in the treatment of EBV-associated B-cell neoplasms.Visual Overview: http://mcr.aacrjournals.org/content/molcanres/17/4/839/F1.large.jpg.

Type 1 diabetes alters ischemia-induced gene expression.

Peripheral Artery Disease (PAD) is a chronic, activity-limiting disease that is caused by atherosclerotic occlusion of blood vessels outside the heart. Type 1 Diabetes (T1D) not only increases an individual's likelihood of developing PAD, but also contributes to poor clinical outcomes after PAD manifestation. Although there is some evidence suggesting that hyperglycemia might alter expression of genes involved in regulating PAD severity or outcomes, our knowledge about the specific genes and pathways involved remains incomplete. We induced experimental PAD or hind limb ischemia in T1D and non-diabetic mice and subjected the ischemic gastrocnemius muscle tissues to genome-wide mRNA transcriptome and pathway analysis. We identified 513 probe sets that represented 443 different genes with highly significant expression differences (p < 0.005) between the ischemic diabetic and ischemic non-diabetic muscle tissues. Moreover, pathway analysis of the differentially expressed genes identified pathways involved in essential biological processes such as "cell cycle," "DNA replication," "metabolic pathways," "focal adhesion," "regulation of actin cytoskeleton," and "nucleotide excision repair". Taken together, our data offer the opportunity to test hypotheses on the roles played by the altered genes/molecular pathways in poor PAD outcomes in diabetes. Such studies may lead to the development of specific therapies to improve PAD outcomes in patients with comorbid diabetes.