The NRG3/ERBB4 signaling cascade as a novel therapeutic target for canine glioma.

Canine glioma is a common brain tumor with poor prognosis despite surgery and/or radiation therapy. Therefore, newer and more effective treatment modalities are needed. Neuregulin 3 (NRG3) has known to be a ligand of ERBB4. This study aimed to investigate the usefulness of the NRG3/ERBB4 signaling cascade as a novel therapeutic target in canine glioma. We found out that microRNA (miR)-190a was downregulated in canine brain tumor tissues, including glioma and meningioma. miR-190a directly targeted NRG3 and inhibited the growth of canine glioma cells. The level of p-Akt, which is a downstream target of ERBB4 signaling, was decreased by transfection with miR-190a. NRG3 silencing also suppressed cell growth and decreased the levels of p-Akt and p-ERK1/2, and NRG3 overexpression exhibited opposed effects in canine glioma J3T-1 cells. The mRNA level of erbb4 was significantly upregulated in glioma tissues compared with that in normal brain tissues and meningioma tissues. Furthermore, compared with gefitinib and lapatinib, afatinib exerted a greater inhibitory effect on the growth of canine glioma cells. In conclusion, NRG3/ERBB4 signaling is negatively regulated by miR-190a and contributes to the growth of canine glioma cells, indicating that it may be a promising therapeutic target in canine glioma.

Interventions in WNT Signaling to Induce Cardiomyocyte Proliferation: Crosstalk with Other Pathways.

Myocardial infarction is a frequent cardiovascular event and a major cause for cardiomyocyte loss. In adult mammals, cardiomyocytes are traditionally considered to be terminally differentiated cells, unable to proliferate. Therefore, the wound-healing response in the infarct area typically yields scar tissue rather than newly formed cardiomyocytes. In the last decade, several lines of evidence have challenged the lack of proliferative capacity of the differentiated cardiomyocyte: studies in zebrafish and neonatal mammals have convincingly demonstrated the regenerative capacity of cardiomyocytes. Moreover, multiple signaling pathways have been identified in these models that-when activated in adult mammalian cardiomyocytes-can reactivate the cell cycle in these cells. However, cardiomyocytes frequently exit the cell cycle before symmetric division into daughter cells, leading to polyploidy and multinucleation. Now that there is more insight into the reactivation of the cell cycle machinery, other prerequisites for successful symmetric division of cardiomyocytes, such as the control of sarcomere disassembly to allow cytokinesis, require more investigation. This review aims to discuss the signaling pathways involved in cardiomyocyte proliferation, with a specific focus on wingless/int-1 protein signaling. Comparing the conflicting results from in vitro and in vivo studies on this pathway illustrates that the interaction with other cells and structures around the infarct is likely to be essential to determine the outcome of these interventions. The extensive crosstalk with other pathways implicated in cardiomyocyte proliferation calls for the identification of nodal points in the cell signaling before cardiomyocyte proliferation can be moved forward toward clinical application as a cure of cardiac disease. SIGNIFICANCE STATEMENT: Evidence is mounting that proliferation of pre-existing cardiomyocytes can be stimulated to repair injury of the heart. In this review article, an overview is provided of the different signaling pathways implicated in cardiomyocyte proliferation with emphasis on wingless/int-1 protein signaling, crosstalk between the pathways, and controversial results obtained in vitro and in vivo.

Slowing disease progression in the SOD1 mouse model of ALS by blocking neuregulin-induced microglial activation.

There are no effective treatments to slow disease progression in ALS. We previously reported that neuregulin (NRG) receptors are constitutively activated on microglia in the ventral horns in both ALS patients and SOD1 mice and in the corticospinal tracts of ALS patients, and that NRG receptor activation occurs prior to significant clinical disease onset in SOD1 mice. Here, we hypothesize that blocking NRG signaling on microglia would slow disease progression in SOD1 mice using a targeted NRG antagonist (HBD-S-H4). Recombinant HBD-S-H4 directly delivered into the central nervous system (CNS) through implanted intracerebroventricular cannulas showed no signs of toxicity and significantly inhibited NRG receptor activation on microglia resulting in reduced microglial activation and motor neuron loss. The treatment also resulted in a delay in disease onset and an increase in survival. The therapeutic effect was dose-dependent that varied as a function of genetic background in two different strains of SOD1 mice. As a complementary drug delivery approach, transgenic mice expressing HBD-S-H4 driven by an astrocytic promoter (GFAP) had slower disease progression in a dose dependent manner, based on the level of HBD-S-H4 expression. These studies provide mechanistic insights into how NRG signaling on microglia may lead to disease progression and demonstrate the utility of a humanized fusion protein that blocks NRG as a novel therapeutic for human ALS.

NRAGE is involved in homologous recombination repair to resist the DNA-damaging chemotherapy and composes a ternary complex with RNF8-BARD1 to promote cell survival in squamous esophageal tumorigenesis.

NRAGE, a neurotrophin receptor-interacting melanoma antigen-encoding gene homolog, is significantly increased in the nucleus of radioresistant esophageal tumor cell lines and is highly upregulated to promote cell proliferation in esophageal carcinomas (ECs). However, whether the overexpressed NRAGE promotes cell growth by participating in DNA-damage response (DDR) is still unclear. Here we show that NRAGE is required for efficient double-strand breaks (DSBs) repair via homologous recombination repair (HRR) and downregulation of NRAGE greatly sensitizes EC cells to DNA-damaging agents both in vitro and in vivo. Moreover, NRAGE not only regulates the stability of DDR factors, RNF8 and BARD1, in a ubiquitin-proteolytic pathway, but also chaperons the interaction between BARD1 and RNF8 via their RING domains to form a novel ternary complex. Additionally, the expression of NRAGE is closely correlated with RNF8 and BARD1 in esophageal tumor tissues. In summary, our findings reveal a novel function of NRAGE that will help to guide personalized esophageal cancer treatments by targeting NRAGE to increase cell sensitivity to DNA-damaging therapeutics in the long run.

Dual targeting of HER2-positive cancer with trastuzumab emtansine and pertuzumab: critical role for neuregulin blockade in antitumor response to combination therapy.

PURPOSE: Targeting HER2 with multiple HER2-directed therapies represents a promising area of treatment for HER2-positive cancers. We investigated combining the HER2-directed antibody-drug conjugate trastuzumab emtansine (T-DM1) with the HER2 dimerization inhibitor pertuzumab (Perjeta). EXPERIMENTAL DESIGN: Drug combination studies with T-DM1 and pertuzumab were performed on cultured tumor cells and in mouse xenograft models of HER2-amplified cancer. In patients with HER2-positive locally advanced or metastatic breast cancer (mBC), T-DM1 was dose-escalated with a fixed standard pertuzumab dose in a 3+3 phase Ib/II study design. RESULTS: Treatment of HER2-overexpressing tumor cells in vitro with T-DM1 plus pertuzumab resulted in synergistic inhibition of cell proliferation and induction of apoptotic cell death. The presence of the HER3 ligand, heregulin (NRG-1ß), reduced the cytotoxic activity of T-DM1 in a subset of breast cancer lines; this effect was reversed by the addition of pertuzumab. Results from mouse xenograft models showed enhanced antitumor efficacy with T-DM1 and pertuzumab resulting from the unique antitumor activities of each agent. In patients with mBC previously treated with trastuzumab, lapatinib, and chemotherapy, T-DM1 could be dosed at the maximum tolerated dose (MTD; 3.6 mg/kg every 3 weeks) with standard dose pertuzumab. Adverse events were mostly grade 1 and 2, with indications of clinical activity. CONCLUSIONS: Dual targeting of HER2 with the combination of T-DM1 and pertuzumab in cell culture and mouse xenograft models resulted in enhanced antitumor activity. In patients, this combination showed an encouraging safety and tolerability profile with preliminary evidence of efficacy.

Pertuzumab protects the achilles' heel of trastuzumab--emtansine.

Trastuzumab emtansine (T-DM1) represents a significant advancement in the treatment of HER2(+) breast cancers. Its clinical efficacy however will be limited by the development of therapeutic resistance. In this report, the HER3 ligand neuregulin is shown to mediate T-DM1 resistance, which was overcome by administration of pertuzumab, a steric inhibitor of HER2 dimerization.

Development of an autocrine neuregulin signaling loop with malignant transformation of human breast epithelial cells.

Neuregulin (NRG) is a heparin-binding factor that activates members of the epidermal growth factor family of tyrosine kinase receptors including erbB2 that is overexpressed in more aggressive types of breast cancer. The exact role that NRG plays in breast cancer is complicated by the fact that NRG has been shown to have both proliferative and antiproliferative effects, depending on the breast cancer cell line used. Using an isogenic series of breast epithelial cell lines (MCF10A) ranging from benign to malignant, we found that the actions of NRG changed from antiproliferative to proliferative as the cells progress to cancer. This correlated with a progressive inability of NRG to down-regulate a group of proliferation genes identified previously using cDNA microarrays. As the cells progress to malignancy, they expressed higher levels of erbB2 and lower levels of erbB3 and secreted high levels of NRG into the culture media, resulting in high basal levels of erbB receptor phosphorylation. Disruption of this autocrine signaling loop by blocking ligand-induced receptor activation inhibited cancer cell proliferation. These results demonstrate that the transition of MCF10A cells from normal to premalignant to malignant correlates with the development of a constitutively active autocrine NRG signaling loop that promotes cell proliferation and suggest that disrupting this autocrine loop may provide an important therapeutic measure to control breast cancer cell growth.

Intracellular signaling molecules involved in an inhibitory factor-induced decrease in fetal-type AChR expression.

The innervation-induced down-regulation of fetal-type acetylcholine receptor (AChR) expression in developing muscle fibers has largely been attributed to nerve-evoked muscle activity; however, there is increasing evidence that a neural trophic factor also contributes to this receptor down-regulation. Previous studies from this laboratory have shown that neural extracts contain a factor which decreases fetal-type AChR expression in skeletal muscle cell lines and therefore may account for the proposed inhibitory neurotrophic influence. The current study investigated possible intracellular signaling molecules involved in this receptor down-regulation and demonstrated that activation of protein kinase C and p70(S6k) appeared to be important in receptor down-regulation. Decreases in AChR density were independent of myogenin. In addition, the receptor down-regulation was independent of neuregulin, which also induces p70(S6k) activity. These studies demonstrate that neural extracts contain an inhibitory factor which can down-regulate fetal-type AChR expression independently of nerve-evoked muscle activity through intracellular signaling molecules which are known to regulate AChR expression.