Structural and functional relationships of natural and artificial dimeric bovine ribonucleases: New scaffolds for potential antitumor drugs

Gotte, Giovanni; Laurents, Douglas V.; Merlino, Antonello; Picone, Delia; Spadaccini, Roberta

doi:10.1016/j.febslet.2013.09.038

Cited by 29 publications

(23 citation statements)

References 99 publications

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“…In the present manuscript, we demonstrate that ONC strongly affects the proliferation of pancreatic adenocarcinoma cells, while it is significantly less active in normal and non-tumoral cells, as we previously observed for bovine seminal RNase (BS-RNase) [26,27]. This result can be explained with the higher capability of these RNases to interact with the very acidic membrane lipids of cancer cells than with those of normal cells [28][29][30].…”

Section: Discussionsupporting

confidence: 82%

Onconase induces autophagy sensitizing pancreatic cancer cells to gemcitabine and activates Akt/mTOR pathway in a ROS-dependent manner

Fiorini

Cordani

Gotte

et al. 2015

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

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Keywords: autophagy onconase pancreatic cancer reactive oxygen species (ROS) mammalian target of rapamycin (mTOR) gemcitabine Onconase® (ONC) is a member of the RNase super-family that is secreted in oocytes and early embryos of Rana pipiens. Over the last years, research interest about this small and basic frog RNase, also called ranpirnase, constantly increased because of its high cytotoxicity and anticancer properties. Onconase is currently used in clinical trials for cancer therapy; however, the precise mechanisms determining cytotoxicity in cancer cells have not yet been fully investigated. In the present manuscript, we evaluate the antitumoral property of onconase in pancreatic adenocarcinoma cells and in non-tumorigenic cells as a control. We demonstrate that ONC stimulates a strong antiproliferative and proapoptotic effect in cancer cells by reporting for the first time that ONC triggers Beclin1-mediated autophagic cancer cell death. In addition, ONC inhibits the expression of mitochondrial uncoupling protein 2 (UCP2) and of manganese-dependent superoxide dismutase (MnSOD) triggering mitochondrial superoxide ion production. ONC-induced reactive oxygen species (ROS) are responsible for Akt/ mTOR pathway stimulation determining the sensitivity of cancer cells to mTOR inhibitors and lessening autophagic stimulation. This indicates ROS/Akt/mTOR axis as a strategy adopted by cancer cells to reduce ONC-mediated cytotoxic autophagy stimulation. In addition, we demonstrate that ONC can sensitize pancreatic cancer cells to the standard chemotherapeutic agent gemcitabine allowing a reduction of drug concentration when used in combination settings, thus suggesting a lowering of chemotherapy-related side effects. Altogether, our results shed more light on the mechanisms lying at the basis of ONC antiproliferative effect in cancer cells and support its potential use to develop new anticancer strategies.

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Section: Discussionsupporting

confidence: 82%

Onconase induces autophagy sensitizing pancreatic cancer cells to gemcitabine and activates Akt/mTOR pathway in a ROS-dependent manner

Fiorini

Cordani

Gotte

et al. 2015

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research

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“…Brain ribonuclease is a member of the ribonuclease A superfamily of 10 to 28 kDa proteins [5–8] and was recently identified as the functional homolog of human ribonuclease-1 (RNase1) [9]. Ribonuclease A superfamily proteins are multifaceted and exhibit immuno-modulatory effects [9], antitumoral activity [10] and pro-apopototic activity [11]. Pancreatic RNase A is the better described RNase A superfamily member in cattle; this RNase is thought to function to breakdown the large amounts of RNA that accumulate in the ruminant gut [12, 13].…”

Section: Introductionmentioning

confidence: 99%

Changes in brain ribonuclease (BRB) messenger RNA in granulosa cells (GCs) of dominant vs subordinate ovarian follicles of cattle and the regulation of BRB gene expression in bovine GCs

Dentis

Schreiber

Gilliam

et al. 2016

Domestic Animal Endocrinology

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Brain ribonuclease (BRB) is a member of the ribonuclease A superfamily that is constitutively expressed in a range of tissues, and is the functional homolog of human ribonuclease 1. This study was designed to characterize BRB gene expression in granulosa cells (GC) during development of bovine dominant ovarian follicles, and to determine the hormonal regulation of BRB in GC. Estrous cycles of Holstein cows (n = 18) were synchronized and cows were ovariectomized on either day 3 to 4 or day 5 to 6 post-ovulation during dominant follicle growth and selection. Ovaries were collected, follicular fluid (FFL) was aspirated, and GC were collected for RNA isolation and quantitative PCR. Follicles were categorized as small (1 to 5 mm; pooled per ovary), medium (5 to 8 mm; individually collected) or large (8.1 to 17 mm; individually collected) based on surface diameter. Estradiol (E2) and progesterone (P4) levels were measured by RIA in FFL. Abundance of BRB mRNA in GC was 8.6- to 11.8-fold greater (P < 0.05) in small (n = 31), medium (n = 66) and large (n = 33) subordinate E2-inactive (FFL E2 < P4) follicles than in large (n = 16) dominant E2-active (FFL E2 > P4) follicles. In the largest 4 follicles, GC BRB mRNA abundance was negatively correlated (P < 0.01) with FFL E2 (r = −0.65) and E2/P4 ratio (r = −0.46). In Exp. 2, GC from large (8 to 22 mm diameter) and small (1 to 5 mm diameter) follicles were treated with IGF1 (0 or 30 ng/mL), and/or tumor necrosis factor α (TNFα) (0 or 30 ng/mL); IGF1 increased (P < 0.05) BRB mRNA abundance and TNFα decreased (P < 0.001) the IGF1-induced BRB mRNA abundance in large-follicle GC. In Exp. 3 to 6, E2, FSH, fibroblast growth factor 9 (FGF9), cortisol, wingless 3A (WNT3A), or Sonic hedgehog (SHH) did not affect (P > 0.10) abundance of BRB mRNA in GC; thyroxine and LH increased (P < 0.05) whereas prostaglandin E2 (PGE2) decreased (P < 0.05) BRB mRNA abundance in small-follicle GC. Treatment of small-follicle GC with recombinant human RNase1 increased (P < 0.05) GC numbers and estradiol production. In conclusion, BRB is a hormonally and developmentally regulated gene in bovine GC and may regulate estradiol production during follicular growth in cattle.

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“…BS-RNase exists as a mixture of two dimeric forms, M=M and MxM, each monomer being a structural homolog of RNase A [60]. The MxM dimer exchanges the N-terminal α-helices forming a 3D-swapped structure and is the form that even in the reducing conditions of the cell cytosol is cytotoxic [61] (for a comparative review on the RNase structures, see [62,63]). BS-RNase binds to the extracellular matrix, and this interaction seems to be important for its cytotoxic effect [64,65] even though it does not bind to cell membranes, suggesting an adsorption cell entry mechanism [66].…”

Section: What Makes a Ribonuclease Selectively Cytotoxic For Cancer Cmentioning

confidence: 99%

“…One of the first approaches to get cytotoxic dimeric RNase variants was to reproduce the structural determinants of BS-RNase swapping [62] in different members of the vertebrate-secreted RNase family. Thus, different combinations of those residues identified as responsible of dimer formation (Cys31, Cys32, Leu28, Gly16, Ser80) of BS-RNase were introduced in the sequence of either HP-RNase or RNase A. Alternatively, the full N-terminal hinge sequence (the peptide that links the N-terminal α-helix of V-shaped RNase structure with the rest of the protein body) of RNase A was replaced by that of BS-RNase [63] in order to endow RNase A with dimerization abilities. These changes resulted in the formation of different ratios of swapped and unswapped forms, which was critical for their cytotoxicity [63].…”

Section: Rnase Dimerization or Oligomerizationmentioning

confidence: 99%

“…Thus, different combinations of those residues identified as responsible of dimer formation (Cys31, Cys32, Leu28, Gly16, Ser80) of BS-RNase were introduced in the sequence of either HP-RNase or RNase A. Alternatively, the full N-terminal hinge sequence (the peptide that links the N-terminal α-helix of V-shaped RNase structure with the rest of the protein body) of RNase A was replaced by that of BS-RNase [63] in order to endow RNase A with dimerization abilities. These changes resulted in the formation of different ratios of swapped and unswapped forms, which was critical for their cytotoxicity [63]. Among these constructs it is remarkable that of a dimeric form of HP-RNase containing the mutations Glu111Gly, Gln28Leu, Arg31Cys, Arg32Cys, and Asn34Lys that was more cytotoxic and selective than BS-RNase for cancer cells [100].…”

Section: Rnase Dimerization or Oligomerizationmentioning

confidence: 99%

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Approaches to Endow Ribonucleases with Antitumor Activity: Lessons Learned from the Native Cytotoxic Ribonucleases

Castro¹,

Ribó²,

Benito³

et al. 2016

Anti-Cancer Drugs - Nature, Synthesis and Cell

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Typical antitumor drugs disrupt the flow of biochemical information from DNA to proteins with the aim of precluding uncontrolled cell proliferation and inducing cancer cell apoptosis. However, most of the currently used small antitumor drugs are genotoxic because they act over DNA. Pharmaceutical industry is now searching for a new line of cancer chemotherapeutics without genotoxic effects. Ribonucleases (RNases) are small basic proteins, present in all life forms, which belong to this kind of chemotherapeutics. Some of them present with remarkable selective antitumor activity linked to their ability to destroy RNA, a powerful way to control gene expression, leaving DNA unharmed. In the last two decades, the knowledge gained on the cytotoxic mechanism of these RNases has been used to engineer more powerful and selective variants to kill cancer cells. In this chapter, we describe the advances reached in endowing an RNase with antitumor abilities.

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Structural and functional relationships of natural and artificial dimeric bovine ribonucleases: New scaffolds for potential antitumor drugs

Cited by 29 publications

References 99 publications

Onconase induces autophagy sensitizing pancreatic cancer cells to gemcitabine and activates Akt/mTOR pathway in a ROS-dependent manner

Onconase induces autophagy sensitizing pancreatic cancer cells to gemcitabine and activates Akt/mTOR pathway in a ROS-dependent manner

Changes in brain ribonuclease (BRB) messenger RNA in granulosa cells (GCs) of dominant vs subordinate ovarian follicles of cattle and the regulation of BRB gene expression in bovine GCs

Approaches to Endow Ribonucleases with Antitumor Activity: Lessons Learned from the Native Cytotoxic Ribonucleases

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