miR-181a negatively regulates NF-κB signaling and affects activated B-cell-like diffuse large B-cell lymphoma pathogenesis.

Distinct subgroups of diffuse large B-cell lymphoma (DLBCL) genetically resemble specific mature B-cell populations that are blocked at different stages of the immune response in germinal centers (GCs). The activated B-cell (ABC)-like subgroup resembles post-GC plasmablasts undergoing constitutive survival signaling, yet knowledge of the mechanisms that negatively regulate this oncogenic signaling remains incomplete. In this study, we report that microRNA (miR)-181a is a negative regulator of nuclear factor κ-light-chain enhancer of activated B-cells (NF-κB) signaling. miR-181a overexpression significantly decreases the expression and activity of key NF-κB signaling components. Moreover, miR-181a decreases DLBCL tumor cell proliferation and survival, and anti-miR-181a abrogates these effects. Remarkably, these effects are augmented in the NF-κB dependent ABC-like subgroup compared with the GC B-cell (GCB)-like DLBCL subgroup. Concordantly, in vivo analyses of miR-181a induction in xenografts results in slower tumor growth rate and prolonged survival in the ABC-like DLBCL xenografts compared with the GCB-like DLBCL. We link these outcomes to relatively lower endogenous miR-181a expression and to NF-κB signaling dependency in the ABC-like DLBCL subgroup. Our findings indicate that miR-181a inhibits NF-κB activity, and that manipulation of miR-181a expression in the ABC-like DLBCL genetic background may result in a significant change in the proliferation and survival phenotype of this malignancy.

Active IKKß promotes the stability of GLI1 oncogene in diffuse large B-cell lymphoma.

GLI1 oncogene has been implicated in the pathobiology of several neoplasms including diffuse large B-cell lymphoma (DLBCL). However, mechanisms underlying GLI1-increased activity in DLBCL are poorly characterized. Herein, we demonstrate that IKKß phosphorylates GLI1 in DLBCL. IKKß activation increased GLI1 protein levels and transcriptional activity, whereas IKKß silencing decreased GLI1 levels and transcriptional activity. Tumor necrosis factor-α (TNFα) mediated IKKß activation-impaired GLI1 binding with the E3 ubiquitin ligase-ITCH, leading to decreased K48-linked ubiquitination/degradation of GLI1. We found 8 IKKß-dependent phosphorylation sites that mediate GLI1 stability. Mutating or deleting these residues facilitated GLI1-ITCH interaction and decreased the protective effect of TNFα on GLI1 stability. IKKß-GLI1 crosstalk is significant because combined inhibition of both molecules resulted in synergistic suppression of DLBCL viability in vivo and in vitro. By linking IKKß-mediated nuclear factor-κB activity with GLI1, we identified a crosstalk between these 2 pathways that can inform the design of novel therapeutic strategies in DLBCL.

LymphomiRs: microRNAs with regulatory roles in lymphomas.

PURPOSE OF REVIEW: This review provides current knowledge on the role of microRNAs (miRNAs) in lymphoma with an emphasis on mature B-cell lymphoma. RECENT FINDINGS: Although miRNAs were previously used to stratify lymphoma classification, prognosis, or treatment response, recent publications portray this class of small noncoding RNAs as critical players in the lymphomagenesis process. Although functional studies provide ample evidence for their role as lymphoma drivers or suppressors, genetic studies providing the underlying mechanisms for these phenotypes are still lacking. As more whole-genome sequencing of lymphoma cases become available, this gap may be soon filled. SUMMARY: Current findings highlight the potential role of miRNAs molecules as important factors in lymphomagenesis.

Potent antibacterial activity in surgical wounds with local administration of D-PLEX100.

Despite significant advances in infection control guidelines and practices, surgical site infections remain a substantial cause of morbidity, prolonged hospitalization, and mortality. The most effective component of SSI reduction strategies is the preoperative administration of intravenous antibiotics; however, systemic antibiotics drug exposure diminishes rapidly and may result in insufficient prophylactic activity against susceptible and resistant SSI pathogens at the wound. D-PLEX100 (D-PLEX) is an antibiotic-releasing drug (doxycycline) that is supplied as a sterile powder for paste reconstitution with sterile saline. D-PLEX paste is administered locally into the incision site along the entire length of soft tissue and sternal bone wound surfaces prior to skin closure. A single D-PLEX administration is intended for 30 days of constant antimicrobial prophylaxis in the prevention of incisional SSIs. We evaluated D-PLEX minimal bactericidal concentration (MBC) against a panel of bacteria that is prevalent in the abdominal wall and sternal surgical procedures including doxycycline susceptible and resistant strains. D-PLEX in vivo efficacy was assessed in incisional infection rabbit models (abdominal wall and sternal) challenged with a similar bacterial panel. The D-PLEX drug exposure profile was determined by in vitro release assay, and in vivo by quantitative pharmacokinetic parameters of local and systemic doxycycline concentrations released from D-PLEX after local administration in incisional rabbit models. Analyses of pathogens and variations in antibiotic resistance from wound isolates were determined from patients who participated in a previously reported prospective randomized trial that assessed the SSI rate in D-PLEX plus standard of care (SOC) versus SOC alone in colorectal resection surgery. The D-PLEX MBC values demonstrated >3- Log10 reduction in all the organisms tested relative to untreated controls, including doxycycline-resistant bacteria (i.e., Methicillin-resistant Staphylococcus aureus (MRSA), K. pneumoniae, and P. aeruginosa). In vivo, D-PLEX significantly reduced the bacterial loads in all the bacteria tested in both animal models (p=0.0001) with a marked impact observed in E. Coli (>6.5 Log10 reduction). D-PLEX exhibited a zero-order release kinetics profile in vitro for 30 days (R2 = 0.971) and the matched in vivo release profile indicated a constant local release of protein-unbound doxycycline for 30 days at 3-5 mcg/mL with significantly lower (>3 orders of magnitudes) systemic levels. In colorectal surgery patients, where significant SSI reduction was observed, analysis of the positive cultures in the overall population indicated similar pathogen diversity and antibiotic resistance rates in both treatment arms. However, almost all the patients with positive culture in the SOC arm were adjudicated as SSI (94%) compared to only 28% in the D-PLEX arm. The SSI-adjudicated D-PLEX patients also exhibited lower resistance rates to the SOC antibiotics and to MDRs compared to patients in the SOC arm. Thus, D-PLEX provides safe and effective prophylaxis activity against the most prevalent SSI pathogens including doxycycline-susceptible and resistant bacteria. Our findings suggest that D-PLEX is a promising addition to SSI prophylactic bundles and may address the gaps in current SSI prophylaxis. D-PLEX is now evaluated in Phase 3 clinical trial.

Reduction in surgical site infections by localized administration with D-PLEX100 in patients with multiple risk factors undergoing colorectal surgery.

BACKGROUND: D-PLEX100 is a novel drug-eluting lipid polymer matrix that supplies a high, local concentration of doxycycline for approximately 30 days. The objective of this post-hoc analysis was to assess the efficacy of D-PLEX100 in preventing superficial and deep SSIs in patients with ≥2 risk factors. PATIENTS AND METHODS: A post-hoc analysis of a previously reported prospective randomized trial assessing D-PLEX100 plus Standard of Care (SOC) versus SOC alone in colorectal surgery was performed to assess SSI rate in patients with ≥2 risk factors. RESULTS: The overall incidence of SSI was significantly lower for the D-PLEX100 arm (9.9%) versus SOC (21%), p = 0.033. Patients with ≥2 risk factors, SSI incidence was 37.5% for SOC and 15.8% in D-PLEX100 treated patients. CONCLUSIONS: D-PLEX100 reduces the incidence of SSIs beyond benefits associated with SOC treatment alone and including patients with ≥2 risk factors. D-PLEX100 may be a promising addition to established SSI prophylaxis bundles.

Mechanistic and signaling analysis of Muc4-ErbB2 signaling module: new insights into the mechanism of ligand-independent ErbB2 activity.

The membrane mucin Muc4 is aberrantly expressed in numerous epithelial carcinomas and is currently used as a cancer diagnostic and prognostic tool. Muc4 can also potentiate signal transduction by modulating differential ErbB2 phosphorylation in the absence and in the presence of the ErbB3 soluble ligand heregulin (HRG-beta1). These features of Muc4 suggest that Muc4 is not merely a cancer marker, but an oncogenic factor with a unique-binding/activation relationship with the receptor ErbB2. In the present study, we examined the signaling mechanisms that are associated with the Muc4-ErbB2 module by analyzing ErbB2 differential signaling in response to Muc4 expression. Our study was carried out in the A375 human melanoma and BT-474 breast cancer cell lines as our model systems. Quantitative and comparative signaling modulations were evaluated by immunoblot using phospho-specific antibodies, and densitometry analysis. Signaling complex components were identified by chemical cross-linking, fractionation by gel filtration, immunoprecipitation, and immunoblotting. Activated downstream signaling pathways were analyzed by an antibody microarray screen and immunoblot analyses. Our results indicate that Muc4 modulates ErbB2 signaling potential significantly by stabilizing and directly interacting with the ErbB2-ErbB3 heterodimer. Further analyses indicate that Muc4 promotes ErbB2 autocatalysis, but it has no effect on ErbB3 phosphorylation, although the chemical cross-linking data indicated that the signaling module is composed of Muc4, ErbB2, and ErbB3. Our microarray analysis indicates that Muc4 expression promotes cell migration by increasing the phosphorylation of the focal adhesion kinase and also through an increase in the levels of beta-catenin.

Muc4/MUC4 functions and regulation in cancer.

The membrane mucin MUC4 (human) is abundantly expressed in many epithelia, where it is proposed to play a protective role, and is overexpressed in some epithelial tumors. Studies on the rat homologue, Muc4, indicate that it acts through anti-adhesive or signaling mechanisms. In particular, Muc4/MUC4 can serve as a ligand/modulator of the receptor tyrosine kinase ErbB2, regulating its phosphorylation and the phosphorylation of its partner ErbB3, with or without the involvement of the ErbB3 ligand neuregulin. Muc4/MUC4 can also modulate cell apoptosis via multiple mechanisms, both ErbB2 dependent and independent. Muc4/MUC4 expression is regulated by multiple mechanisms, ranging from transcriptional to post-translational. The roles of MUC4 in tumors suggest that it may be valuable as a tumor marker or target for therapy.

MicroRNAs are independent predictors of outcome in diffuse large B-cell lymphoma patients treated with R-CHOP.

PURPOSE: Diffuse large B-cell lymphoma (DLBCL) heterogeneity has prompted investigations for new biomarkers that can accurately predict survival. A previously reported 6-gene model combined with the International Prognostic Index (IPI) could predict patients' outcome. However, even these predictors are not capable of unambiguously identifying outcome, suggesting that additional biomarkers might improve their predictive power. EXPERIMENTAL DESIGN: We studied expression of 11 microRNAs (miRNA) that had previously been reported to have variable expression in DLBCL tumors. We measured the expression of each miRNA by quantitative real-time PCR analyses in 176 samples from uniformly treated DLBCL patients and correlated the results to survival. RESULTS: In a univariate analysis, the expression of miR-18a correlated with overall survival (OS), whereas the expression of miR-181a and miR-222 correlated with progression-free survival (PFS). A multivariate Cox regression analysis including the IPI, the 6-gene model-derived mortality predictor score and expression of the miR-18a, miR-181a, and miR-222, revealed that all variables were independent predictors of survival except the expression of miR-222 for OS and the expression of miR-18a for PFS. CONCLUSION: The expression of specific miRNAs may be useful for DLBCL survival prediction and their role in the pathogenesis of this disease should be examined further.

Conformational changes in receptor tyrosine kinase signaling: an ErbB garden of delights.

The ErbB family of receptor tyrosine kinases plays important roles in cell proliferation, differentiation, and apoptosis. Recent structural studies of these receptors have demonstrated dramatic conformational effects that are critical to their ligand binding and activation, and have shown that these receptors provide levels of control beyond the classic dimerization/activation mechanism. These results indicate that this class of receptors has evolved subtle regulatory mechanisms via genetic and protein structural changes to influence their effects on cell behaviors.