Hepatitis E Virus ORF2 Protein Activates the Pro-Apoptotic Gene CHOP and Anti-Apoptotic Heat Shock Proteins

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Unraveling a tumor type-specific regulatory core underlying E2F1-mediated epithelial-mesenchymal transition to predict receptor protein signatures

Cancer is a disease of subverted regulatory pathways. In this paper, we reconstruct the regulatory network around E2F, a family of transcription factors whose deregulation has been associated to cancer progression, chemoresistance, invasiveness, and metastasis. We integrate gene expression profiles of cancer cell lines from two E2F1-driven highly aggressive bladder and breast tumors, and use network analysis methods to identify the tumor type-specific core of the network. By combining logic-based network modeling, in vitro experimentation, and gene expression profiles from patient cohorts displaying tumor aggressiveness, we identify and experimentally validate distinctive, tumor type-specific signatures of receptor proteins associated to epithelial-mesenchymal transition in bladder and breast cancer. Our integrative network-based methodology, exemplified in the case of E2F1-induced aggressive tumors, has the potential to support the design of cohort- as well as tumor type-specific treatments and ultimately, to fight metastasis and therapy resistance

STAT3 and STAT5 Targeting for Simultaneous Management of Melanoma and Autoimmune Diseases

Melanoma is a skin cancer which can become metastatic, drug-refractory, and lethal if managed late or inappropriately. An increasing number of melanoma patients exhibits autoimmune diseases, either as pre-existing conditions or as sequelae of immune-based anti-melanoma therapies, which complicate patient management and raise the need for more personalized treatments. STAT3 and/or STAT5 cascades are commonly activated during melanoma progression and mediate the metastatic effects of key oncogenic factors. Deactivation of these cascades enhances antitumor-immune responses, is efficient against metastatic melanoma in the preclinical setting and emerges as a promising targeting strategy, especially for patients resistant to immunotherapies. In the light of the recent realization that cancer and autoimmune diseases share common mechanisms of immune dysregulation, we suggest that the systemic delivery of STAT3 or STAT5 inhibitors could simultaneously target both, melanoma and associated autoimmune diseases, thereby decreasing the overall disease burden and improving quality of life of this patient subpopulation. Herein, we review the recent advances of STAT3 and STAT5 targeting in melanoma, explore which autoimmune diseases are causatively linked to STAT3 and/or STAT5 signaling, and propose that these patients may particularly benefit from treatment with STAT3/STAT5 inhibitors

Peptide-Based Technologies to Alter Adenoviral Vector Tropism: Ways and Means for Systemic Treatment of Cancer

Due to the fundamental progress in elucidating the molecular mechanisms of human diseases and the arrival of the post-genomic era, increasing numbers of therapeutic genes and cellular targets are available for gene therapy. Meanwhile, the most important challenge is to develop gene delivery vectors with high efficiency through target cell selectivity, in particular under in situ conditions. The most widely used vector system to transduce cells is based on adenovirus (Ad). Recent endeavors in the development of selective Ad vectors that target cells or tissues of interest and spare the alteration of all others have focused on the modification of the virus broad natural tropism. A popular way of Ad targeting is achieved by directing the vector towards distinct cellular receptors. Redirecting can be accomplished by linking custom-made peptides with specific affinity to cellular surface proteins via genetic integration, chemical coupling or bridging with dual-specific adapter molecules. Ideally, targeted vectors are incapable of entering cells via their native receptors. Such altered vectors offer new opportunities to delineate functional genomics in a natural environment and may enable efficient systemic therapeutic approaches. This review provides a summary of current state-of-the-art techniques to specifically target adenovirus-based gene delivery vectors

Antisense gapmers selectively suppress individual oncogenic p73 splice isoforms and inhibit tumor growth in vivo

BACKGROUND: Differential mRNA splicing and alternative promoter usage of the TP73 gene results in the expression of multiple NH2-truncated isoforms that act as oncogenes. Abundant levels of these p73 variants in a variety of human cancers correlated with adverse clinical prognosis and response failure to conventional therapies, underscoring their relevance as marker for disease severity and target for cancer intervention. With respect to an equally important role for amino-truncated p73 splice forms (ΔTAp73) and ΔNp73 (summarized as DNp73) in the tumorigenic process, we designed locked nucleic acid (LNA) antisense oligonucleotide (ASO) gapmers against individual species that were complementary to ΔEx2 and ΔEx2/3 splice junctions and a region in exon 3B unique for ΔN' and ΔN. RESULTS: Treatment of cancer cells with these ASOs resulted in a strong and specific reduction of tumorigenic p73 transcripts and proteins, importantly, without abolishing the wild-type p73 tumor suppressor form as observed with p73-shRNA. The specific antisense oligonucleotides rescued cells from apoptosis inhibition due to overexpression of their corresponding amino-truncated p73 isoform and decreased tumor cell proliferation. Furthermore, ASO-116 against ΔEx2/3 coupled to magnetic nanobead polyethyleneimine (MNB/PEI) carriers significantly inhibited malignant melanoma growth, which correlated with a shift in the balance between endogenous TAp73 and ΔEx2/3 towards apoptotic full-length p73. CONCLUSION: Our study demonstrates the successful development of LNA-ASOs that selectively differentiate between the closely related p73 oncoproteins, and provide new tools to further delineate their biological properties in different human malignancies and for therapeutic cancer targeting

Drug Repurposing at the Interface of Melanoma Immunotherapy and Autoimmune Disease

Cancer cells have a remarkable ability to evade recognition and destruction by the immune system. At the same time, cancer has been associated with chronic inflammation, while certain autoimmune diseases predispose to the development of neoplasia. Although cancer immunotherapy has revolutionized antitumor treatment, immune-related toxicities and adverse events detract from the clinical utility of even the most advanced drugs, especially in patients with both, metastatic cancer and pre-existing autoimmune diseases. Here, the combination of multi-omics, data-driven computational approaches with the application of network concepts enables in-depth analyses of the dynamic links between cancer, autoimmune diseases, and drugs. In this review, we focus on molecular and epigenetic metastasis-related processes within cancer cells and the immune microenvironment. With melanoma as a model, we uncover vulnerabilities for drug development to control cancer progression and immune responses. Thereby, drug repurposing allows taking advantage of existing safety profiles and established pharmacokinetic properties of approved agents. These procedures promise faster access and optimal management for cancer treatment. Together, these approaches provide new disease-based and data-driven opportunities for the prediction and application of targeted and clinically used drugs at the interface of immune-mediated diseases and cancer towards next-generation immunotherapies

Functional dissection of the alphavirus capsid protease: sequence requirements for activity

Abstract Background The alphavirus capsid is multifunctional and plays a key role in the viral life cycle. The nucleocapsid domain is released by the self-cleavage activity of the serine protease domain within the capsid. All alphaviruses analyzed to date show this autocatalytic cleavage. Here we have analyzed the sequence requirements for the cleavage activity of Chikungunya virus capsid protease of genus alphavirus. Results Amongst alphaviruses, the C-terminal amino acid tryptophan (W261) is conserved and found to be important for the cleavage. Mutating tryptophan to alanine (W261A) completely inactivated the protease. Other amino acids near W261 were not having any effect on the activity of this protease. However, serine protease inhibitor AEBSF did not inhibit the activity. Through error-prone PCR we found that isoleucine 227 is important for the effective activity. The loss of activity was analyzed further by molecular modelling and comparison of WT and mutant structures. It was found that lysine introduced at position 227 is spatially very close to the catalytic triad and may disrupt electrostatic interactions in the catalytic site and thus inactivate the enzyme. We are also examining other sequence requirements for this protease activity. Conclusions We analyzed various amino acid sequence requirements for the activity of ChikV capsid protease and found that amino acids outside the catalytic triads are important for the activity.</p

Development of Adenoviral Delivery Systems to Target Hepatic Stellate Cells In Vivo.

Hepatic stellate cells (HSCs) are known as initiator cells that induce liver fibrosis upon intoxication or other noxes. Deactivation of this ongoing remodeling process of liver parenchyma into fibrotic tissue induced by HSCs is an interesting goal to be achieved by targeted genetic modification of HSCs. The most widely applied approach in gene therapy is the utilization of specifically targeted vectors based on Adenovirus (Ad) serotype 5. To narrow down the otherwise ubiquitous tropism of parental Ad, two modifications are required: a) ablating the native tropism and b) redirecting the vector particles towards a specific entity solely present on the cells of interest. Therefore, we designed a peptide of the nerve growth factor (NGFp) with specific affinity for the p75 neurotrophin receptor (p75NTR) present on HSCs. Coupling of this NGFp to vector particles was done either via chemical conjugation using bifunctional polyethylene glycol (PEG) or, alternatively, by molecular bridging with a fusion protein specific for viral fiber knob and p75NTR. Both Ad vectors transmit the gene for the green fluorescent protein (GFP). GFP expression was monitored in vitro on primary murine HSCs as well as after systemic administration in mice with healthy and fibrotic livers using intravital fluorescence microscopy. Coupling of NGFp to Ad via S11 and/or PEGylation resulted in markedly reduced liver tropism and an enhanced adenoviral-mediated gene transfer to HSCs. Transduction efficiency of both specific Ads was uniformly higher in fibrotic livers, whereas Ad.GFP-S11-NGFp transduce activated HSCs better than Ad.GFP-PEG-NGFp. These experiments contribute to the development of a targeted gene transfer system to specifically deliver antifibrotic compounds into activated HSCs by systemically applied adenoviral vector modified with NGFp