Morena d'Avenia

IKKγ protein is a target of BAG3 regulatory activity in human tumor growth

BAG3, a member of the BAG family of heat shock protein (HSP) 70 cochaperones, is expressed in response to stressful stimuli in a number of normal cell types and constitutively in a variety of tumors, including pancreas carcinomas, lymphocytic and myeloblastic leukemias, and thyroid carcinomas. Down-regulation of BAG3 results in cell death, but the underlying molecular mechanisms are still elusive. Here, we investigated the molecular mechanism of BAG3-dependent survival in human osteosarcoma (SAOS-2) and melanoma (M14) cells. We show that bag3 overexpression in tumors promotes survival through the NF-κB pathway. Indeed, we demonstrate that BAG3 alters the interaction between HSP70 and IKKγ, increasing availability of IKKγ and protecting it from proteasome-dependent degradation; this, in turn, results in increased NF-κB activity and survival. These results identify bag3 as a potential target for anticancer therapies in those tumors in which this gene is constitutively expressed. As a proof of principle, we show that treatment of a mouse xenograft tumor model with bag3siRNA-adenovirus that down-regulates bag3 results in reduced tumor growth and increased animal survival.

BAG3 promotes pancreatic ductal adenocarcinoma growth by activating stromal macrophages

The incidence and death rate of pancreatic ductal adenocarcinoma (PDAC) have increased in recent years, therefore the identification of novel targets for treatment is extremely important. Interactions between cancer and stromal cells are critically involved in tumour formation and development of metastasis. Here we report that PDAC cells secrete BAG3, which binds and activates macrophages, inducing their activation and the secretion of PDAC supporting factors. We also identify IFITM-2 as a BAG3 receptor and show that it signals through PI3K and the p38 MAPK pathways. Finally, we show that the use of an anti-BAG3 antibody results in reduced tumour growth and prevents metastasis formation in three different mouse models. In conclusion, we identify a paracrine loop involved in PDAC growth and metastatic spreading, and show that an anti-BAG3 antibody has therapeutic potential.

A novel miR-371a-5p-mediated pathway, leading to BAG3 upregulation in cardiomyocytes in response to epinephrine, is lost in Takotsubo cardiomyopathy

Molecular mechanisms protecting cardiomyocytes from stress-induced death, including tension stress, are essential for cardiac physiology and defects in these protective mechanisms can result in pathological alterations. Bcl2-associated athanogene 3 (BAG3) is expressed in cardiomyocytes and is a component of the chaperone-assisted autophagy pathway, essential for homeostasis of mechanically altered cells. BAG3 ablation in mice results in a lethal cardiomyopathy soon after birth and mutations of this gene have been associated with different cardiomyopathies including stress-induced Takotsubo cardiomyopathy (TTC). The pathogenic mechanism leading to TTC has not been defined, but it has been suggested that the heart can be damaged by excessive epinephrine (epi) spillover in the absence of a protective mechanism. The aim of this study was to provide more evidence for a role of BAG3 in the pathogenesis of TTC. Therefore, we sequenced BAG3 gene in 70 TTC patients and in 81 healthy donors with the absence of evaluable cardiovascular disease. Mutations and polymorphisms detected in the BAG3 gene included a frequent nucleotide change g2252c in the BAG3 3′-untranslated region (3′-UTR) of Takotsubo patients (P<0.05), resulting in loss of binding of microRNA-371a-5p (miR-371a-5p) as evidenced by dual-luciferase reporter assays and argonaute RNA-induced silencing complex catalytic component 2/pull-down assays. Moreover, we describe a novel signaling pathway in cardiomyocytes that leads to BAG3 upregulation on exposure to epi through an ERK-dependent upregulation of miR-371a-5p. In conclusion, the presence of a g2252c polymorphism in the BAG3 3′-UTR determines loss of miR-371a-5p binding and results in an altered response to epi, potentially representing a new molecular mechanism that contributes to TTC pathogenesis.

BAG3 is a novel serum biomarker for pancreatic adenocarcinomas.

to the small number of patients (62.5 vs. 37.5 % , P = 0.11). However, when comparing median cumulative exposure to RBV between groups as measured by the area under the drug exposure curve from week 0 to 12 based on biweekly measurements of RBV plasma levels rather than RBV dosage per se , cumulative exposure to RBV above ≥ 224.3 μ g / dl / day was signifi cantly associated with SVR (odds ratio 8.8; confi dence interval 1.35 – 57.43, P = 0.02) ( Figure 1 ). Anemia in group A was more severe than in group B (mean hemoglobin 99.6 vs. 106.3 g / l; P < 0.001), but well manageable with erythropoietin beta at doses between 9,000 and 30,000 IU per week according to a recently proposed consensus ( 10 ) Except for anemia, adverse events were similar in both groups. In conclusion — and in accordance with the study by Jin et al. ( 7 ) — optimal exposure to RBV guided by therapeutic drug monitoring signifi cantly improves SVR in patients with CHC genotype 1. Th erefore, regular RBV plasma level measurements at least for the fi rst 12 weeks of therapy and RBV dose adjustment may be advocated.

BAG3 Protein Is Over-Expressed in Endometrioid Endometrial Adenocarcinomas

Endometrioid endometrial cancer is the most common gynaecological tumor in developed countries, and its incidence is increasing. The definition of subtypes, based on clinical and endocrine features or on histopathological characteristics, correlate to some extent with patients prognosis, but there is substantial heterogeneity within tumor types. The search for molecules and mechanisms implied in determining the progression and the response to therapy for this cancer is still ongoing. BAG3 protein, a member of BAG family of co‐chaperones, has a pro‐survival role in several tumor types. BAG3 anti‐apoptotic properties rely on its characteristic to bind several intracellular partners, thereby, modulating crucial events such as apoptosis, differentiation, cell motility, and autophagy. BAG3 expression in human endometrial cancer tissues was not investigated so far. Here, we show that BAG3 protein levels are elevated in tumoral and hyperplastic cells in respect to normal glands. Furthermore, BAG3 subcellular localization appears to be changed in tumoral compared to normal cells. Our results indicate a possible role for BAG3 protein in the maintenance of cell survival in endometrioid endometrial cancer and suggest that this field of studies is worthy of further investigations. J. Cell. Physiol. 232: 309–311, 2017.

Abstract 2393: PDAC cells release BAG3 that activates a paracrine loop with stromal macrophages

Interactions between cancer cells and stromal cells are critically involved in tumor formation, progression and development of metastasis. Here we report for the first time that human pancreatic adenocarcinoma (PDAC) cells secrete BAG3 that binds and activates macrophages, inducing their activation and the secretion of pancreatic cancer- supporting factors. We also identify IFITM-2 as a BAG3 receptor, by its co-precipitation with BAG3 from macrophage cell membrane and by impairment of BAG3- induced activation in IFITM-2- silenced macrophages. Finally we show the importance of this pathway in vivo since in nu/nu mice carrying orthotopic xenografts of human PDAC (MIA PaCa-2) cells, BAG3 sequestration by an anti-BAG3 antibody results in reduced tumor growth and prevents metastastasis formation. In conclusion we have identified a novel paracrine loop involved in PDAC growth and metastatic spreading and shown that its pharmacological blockage with an anti-BAG3 antibody has therapeutic potential. Citation Format: Alessandra Rosati, Anna Basile, Raffaella D9Auria, Morena d9Avenia, Margot De Marco, Antonia Falco, Michelina Festa, Luana Guerriero, Vittoria Iorio, Maria Pascale, Renato Franco, Claudio Arra, Antonio Barbieri, Domenica Rea, Giulio Menichini, Michael Hahne, Marteen Bijlsma, Daniela Barcaroli, David A. Tuveson, Jelena Todoric, Michael Karin, Vincenzo De Laurenzi, Maria Caterina Turco. PDAC cells release BAG3 that activates a paracrine loop with stromal macrophages. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2393. doi:10.1158/1538-7445.AM2015-2393

Abstract 3034: air (apoptosis-induced regulator) expression and its sequence localization in human genome

The aim of this study was to characterize a newly identified transcript named air (AX538681) whose genomic sequence is missing in human genome draft. air was identified as a differentially expressed cDNA in TNFα-stimulated Jurkat cells, transfected with a IkBα-hyperexpressing vector, versus control cells (Turco et al, 2007). Indeed air expression appeared to be induced by a pro-apoptotic stimulus and repressed by NF-κB activity. Air cDNA was isolated by differential screening analysis of an expression library from Jurkat cells; its sequence (3.4kb long, 79% A/T rich) didn9t show any similarity with annotated sequences except the homology with an EST (U74659) from human neuroblastoma cells and a 98% identity with traces from chimpanzee WGS database. This evidence suggested that air could belong to an un-sequenced region, whose secondary structure results in cloning instability. Air sequence is detectable by Southern blot and PCR, providing the evidence of its presence in human genome (in press). air genomic 3’ and 5’ flanking regions (each one ∼2.2kb long) were isolated by vectorette PCR and sequenced to allow the synthesis of a probe (∼8kb long) suitable to perform a FISH assay. Preliminary experiments, performed on a thyroid cancer cell line, showed that air localize on Chr. 22. Moreover, we found that air transcript is longer than 3.4kb, since air 5’ flanking region is detectable by RT-PCR on total RNA (unpublished). Four main putative ORFs were identified by ORF-Finder tool, but the aminoacidic sequence of three of them did not show any homology with conserved structural motifs while the fourth, localized on the new isolated 5’ transcript end, showed a transmembrane-like motif. Anyway, we don9t exclude that air could be a long ncRNA (but it is not the Air described by Nagano et al, 2008), in fact, it is rich in stop codons, it9s strongly predisposed to form stem loop secondary structures and its expression coincides with pro-apoptotic events (submitted). Air role in modulating cell apoptosis was investigated: air silencing in lymphoma, neuroblastoma and AML cells, treated with stressful stimuli, showed a significant (p These findings identify air as a novel potential tool for enhancing cancer response to therapeutics. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3034. doi:10.1158/1538-7445.AM2011-3034

Physiology of Immune System: Regulation of Stem Cell Survival

The immune system is a complex defense mechanism, able to protect the body against pathogens. It consists of a network of cells, tissues, and organs that work together to protect the body. Leukocytes are key operatives of the immune system. Cells destined to become immune cells, like all blood cells, arise in your bodys bone marrow from stem cells (HSC). A large body of evidence show the transcription factors play critical roles in the homeostasis of T cells, B cells, neutrophils and other non-lymphoid leneages. This review discusses the role of the Smek (Suppressor of mek null) gene, that acts in the stress response pathway of animals by binding to and enhancing the transcription of FoxO transcription factors. Furthermore, the review deals with tachykinins, involved in neurotransmission and immune/hematopoietic modulation. Both molecules, objects of recent patents, may have real therapeutic potential.

Novel Targets for Apoptosis Modulation: BAG3 Protein and Other Co- Chaperones

Deregulation of apoptosis is responsible for diseases that involve defects in the death pathway, such as neoplasias, or in its inhibition, like degenerative processes. Among apoptosis- regulating molecules, a role is emerging for BAG3, a member of the BAG proteins family (BAG1L, BAG1, BAG2, BAG3, BAG4. BAG5 and BAG6) involved in cochaperoning of heat shock proteins. Through its BAG, WW and prolix-rich domains, BAG3 protein can interact with a variety of molecular partners, including Hsc70/Hsp70, phospholipase C-γ and others. It has been recently shown that, in human primary lymphoid and myeloblastic leukemias, thyroid carcinoma and other human tumors, BAG3 expression sustains cell survival and impairs cell response to therapy. This review discusses two patents concerning, respectively, BAG3 and other Hsc70/Hsp70 co-chaperones, namely HspBP-1 and HspBP-2.