The ADAMs family of proteases: new biomarkers and therapeutic targets for cancer?

The ADAMs are transmembrane proteins implicated in proteolysis and cell adhesion. Forty gene members of the family have been identified, of which 21 are believed to be functional in humans. As proteases, their main substrates are the ectodomains of other transmembrane proteins. These substrates include precursor forms of growth factors, cytokines, growth factor receptors, cytokine receptors and several different types of adhesion molecules. Although altered expression of specific ADAMs has been implicated in different diseases, their best-documented role is in cancer formation and progression. ADAMs shown to play a role in cancer include ADAM9, ADAM10, ADAM12, ADAM15 and ADAM17. Two of the ADAMs, i.e., ADAM10 and 17 appear to promote cancer progression by releasing HER/EGFR ligands. The released ligands activate HER/EGFR signalling that culminates in increased cell proliferation, migration and survival. Consistent with a causative role in cancer, several ADAMs are emerging as potential cancer biomarkers for aiding cancer diagnosis and predicting patient outcome. Furthermore, a number of selective ADAM inhibitors, especially against ADAM10 and ADAM17, have been shown to have anti-cancer effects. At least one of these inhibitors is now undergoing clinical trials in patients with breast cancer.     

The “A Disintegrin And Metalloproteases” ADAM10 and ADAM17: Novel drug targets with therapeutic potential?

Proteolytic ectodomain release, a process known as "shedding", has been recognised as a key mechanism for regulating the function of a diversity of cell surface proteins. A Disintegrin And Metalloproteinases (ADAMs) have emerged as the major proteinase family that mediates ectodomain shedding. Dysregulation of ectodomain shedding is associated with autoimmune and cardiovascular diseases, neurodegeneration, infection, inflammation and cancer. Therefore, ADAMs are increasingly regarded as attractive targets for novel therapies. ADAM10 and its close relative ADAM17 (TNF-alpha converting enzyme (TACE)) have been studied in particular in the context of ectodomain shedding and have been demonstrated as key molecules in most of the shedding events characterised to date. Whereas the level of expression of ADAM10 may be of importance in cancer and neurodegenerative disorders, ADAM17 mainly coordinates pro- and anti-inflammatory activities during immune response. Despite the high therapeutical potential of ADAM inhibition, all clinical trials using broad-spectrum metalloprotease inhibitors have failed so far. This review will cover the emerging roles of both ADAM10 and ADAM17 in the regulation of major physiological and developmental pathways and will discuss the suitability of specifically modulating the activities of both proteases as a feasible way to inhibit inflammatory states, cancer and neurodegeneration.     

“Targeting the absence” and therapeutic engineering for cancer therapy

The Gotham Prize was awarded to Alex Varshavsky for “Targeting the absence”, a strategy employing negative targets of cancer therapy. This is a brilliant example of therapeutic engineering: designing a sequence of events that leads to the selective killing of one type of cell, while sparing all others. A complex molecular device (Varshavsky’s Demon) examines DNA, recognizes the present target in normal cells and kills cancer cells. The strategy is limited by the delivery (transfection or infection) of DNA-based devices into each cell of our body. How can we overcome this limitation? Can therapeutic engineering be applied to small drugs? Can each small molecule reach a cell separately and, once in a cell, exert orchestrated action governed by cellular context? Here I describe how a combination of small drugs can acquire a demonic power to check, choose and selectively kill. The cytotoxicity is restricted to cells lacking (or having) one of the targets. For example, in the presence of a normal target, one drug can cancel the cytotoxic action of another drug. And by increasing a number of targets, we can increase the precision and power of such ‘restrictive’ combinations. Here I discuss restrictive combinations of currently available drugs that could be tested in clinical trials. Could then these combinations cure cancer today? And what does ‘cure’ really mean? This article suggests the answer.     

Emerging roles for β-arrestin-1 in the control of the pancreatic β-cell function and mass: New therapeutic strategies and consequences for drug screening

Defective insulin secretion is a feature of type 2 diabetes that results from inadequate compensatory increase in β-cell mass, decreased β-cell survival and impaired glucose-dependent insulin release. Pancreatic β-cell proliferation, survival and secretion are thought to be regulated by signalling pathways linked to G-protein coupled receptors (GPCRs), such as the glucagon-like peptide-1 (GLP-1) and the pituitary adenylate cyclase-activating polypeptide (PACAP) receptors. β-arrestin-1 serves as a multifunctional adaptor protein that mediates receptor desensitization, receptor internalization, and links GPCRs to downstream pathways such as tyrosine kinase Src, ERK1/2 or Akt/PKB. Importantly, recent studies found that β-arrestin-1 mediates GLP-1 signalling to insulin secretion, GLP-1 antiapoptotic effect by phosphorylating the proapoptotic protein Bad through ERK1/2 activation, and PACAP potentiation of glucose-induced long-lasting ERK1/2 activation controlling IRS-2 expression. Together, these novel findings reveal an important functional role for β-arrestin-1 in the regulation of insulin secretion and β-cell survival by GPCRs.     

Purinergic receptors and neglected tropical diseases: why ignore purinergic signaling in the search for new molecular targets?

Purinergic receptors are widespread in the human organism and are involved in several physiological functions like neurotransmission, nociception, platelet aggregation, etc. In the immune system, they may regulate the expression and release of pro-inflammatory factors as well as the activation and death of several cell types. It is already described the participation of some purinergic receptors in the inflammation and pathological processes, such as a few neglected tropical diseases (NTDs) which affect more than 1 billion people in the world. Although the high social influence those diseases represent endemic countries, most of them do not have an efficient, safe or affordable drug treatment. In that way, this review aims to discuss the current literature involving purinergic receptor and immune response to NTDs pathogens, which may contribute in the search for new therapeutic possibilities.     

A step on the path in the discovery of new latent fingerprint development reagents: substituted Ruhemann’s purples and implications for the law

Energies corresponding to optimum geometries of ninhydrin, some of its analogs, the corresponding Ruhemann’s Purple analogs and some of the intermediates of the reaction between ninhydrin analogs and amino acids are calculated at Hartree–Fock/6-31G** level of theory. Such a study is significant from a forensic science point of view because of the strong interest in the forensic chemistry and law enforcement communities in developing alternatives to the current generation of ninhydrin-like chemicals for the detection and development of latent fingerprints. In examining our new predictions for the net energetics of the reactions in the formation of substituted Ruhemann’s Purples, we find that a fluorine-containing analog is the most thermodynamically feasible reaction. In light of this finding, we suggest further experimental studies to determine the kinetic feasibility of synthesizing the fluorine-containing analog, as well as other similar molecules and to determine their spectroscopic properties.     

Interleukin-10 and the regulation of mitogen-activated protein kinases: are these signalling modules targets for the anti-inflammatory action of this cytokine?

The many specific, yet overlapping and redundant activities of individual cytokines have been the basis for current concepts of therapeutical intervention. Cytokines are powerful two-edged weapons that can trigger a cascade of reactions and may show activities that often go beyond the single highly specific property that it is hoped they possess. Nevertheless, it can be stated that our new, though burgeoning, understanding of the biological mechanisms governing cytokine actions is an important contribution to medical knowledge. The crucial role of the anti-inflammatory cytokine, interleukin (IL)-10, in regulating potential molecular pathway mediating injury and cell death has attracted paramount attention in recent years. In this respect, the mitogen-activated protein kinase (MAPK) components have emerged as potential signalling cascades that regulate a plethora of cell functions, including inflammation and cell death. The biochemistry and molecular biology of cytokine actions, particularly IL-10, explain some well known and sometimes also some of the more obscure clinical aspects of the evolution of diseases.     

The “bone morphogenic proteins” pathways in bone and joint diseases: Translational perspectives from physiopathology to therapeutic targets

A large body of evidence supports an important role of bone morphogenic proteins (BMPs) pathways in skeletal development in the embryo. BMPs are also involved in skeletal homeostasis and diseases in the adult. They were first identified as major bone anabolic agents and recent advances indicate that they also regulate osteoclastogenesis and joint components via multiple cross-talks with other signaling pathways. This review attempts to integrate these data in the pathogenesis of bone and joints diseases, such as osteoporosis, fracture healing, osteoarthritis, inflammatory arthritis, or bone metastasis. The use of recombinant BMPs in bone tissue engineering and in the treatment of skeletal diseases, or future therapeutic strategies targeting BMPs signal and its regulators, will be discussed based on these considerations.     

Selection and mutation in the “new” genetics: an emerging hypothesis

It has been anticipated that new, much more sensitive, next generation sequencing (NGS) techniques, using massively parallel sequencing, will likely provide radical insights into the genetics of multifactorial diseases. While NGS has been used initially to analyze individual human genomes, and has revealed considerable differences between healthy individuals, we have used NGS to examine genetic variation within individuals, by sequencing tissues “in depth”, i.e., oversequencing many thousands of times. Initial studies have revealed intra-tissue genetic heterogeneity, in the form of multiple variants of a single gene that exist as distinct “majority and “minority” variants. This highly specialized form of somatic mosaicism has been found within both cancer and normal tissues. If such genetic variation within individual tissues is widespread, it will need to be considered as a significant factor in the ontogeny of many multifactorial diseases, including cancer. The discovery of majority and minority gene variants and the resulting somatic cell heterogeneity in both normal and diseased tissues suggests that selection, as opposed to mutation, might be the critical event in disease ontogeny. We, therefore, are proposing a hypothesis to explain multifactorial disease ontogeny in which pre-existing multiple somatic gene variants, which may arise at a very early stage of tissue development, are eventually selected due to changes in tissue microenvironments.     

Metabolic reprogramming of cancer-associated fibroblasts by TGF-β drives tumor growth: Connecting TGF-β signaling with “Warburg-like” cancer metabolism and L-lactate production

We have previously shown that a loss of stromal Cav-1 is a biomarker of poor prognosis in breast cancers. Mechanistically, a loss of Cav-1 induces the metabolic reprogramming of stromal cells, with increased autophagy/mitophagy, mitochondrial dysfunction and aerobic glycolysis. As a consequence, Cav-1-low CAFs generate nutrients (such as L-lactate) and chemical building blocks that fuel mitochondrial metabolism and the anabolic growth of adjacent breast cancer cells. It is also known that a loss of Cav-1 is associated with hyperactive TGF-β signaling. However, it remains unknown whether hyperactivation of the TGF-β signaling pathway contributes to the metabolic reprogramming of Cav-1-low CAFs. To address these issues, we overexpressed TGF-β ligands and the TGF-β receptor I (TGFβ-RI) in stromal fibroblasts and breast cancer cells. Here, we show that the role of TGF-β in tumorigenesis is compartment-specific, and that TGF-β promotes tumorigenesis by shifting cancer-associated fibroblasts toward catabolic metabolism. Importantly, the tumor-promoting effects of TGF-β are independent of the cell type generating TGF-β. Thus, stromal-derived TGF-β activates signaling in stromal cells in an autocrine fashion, leading to fibroblast activation, as judged by increased expression of myofibroblast markers, and metabolic reprogramming, with a shift toward catabolic metabolism and oxidative stress. We also show that TGF-β-activated fibroblasts promote the mitochondrial activity of adjacent cancer cells, and in a xenograft model, enhancing the growth of breast cancer cells, independently of angiogenesis. Conversely, activation of the TGF-β pathway in cancer cells does not influence tumor growth, but cancer cell-derived-TGF-β ligands affect stromal cells in a paracrine fashion, leading to fibroblast activation and enhanced tumor growth. In conclusion, ligand-dependent or cell-autonomous activation of the TGF-β pathway in stromal cells induces their metabolic reprogramming, with increased oxidative stress, autophagy/mitophagy and glycolysis, and downregulation of Cav-1. These metabolic alterations can spread among neighboring fibroblasts and greatly sustain the growth of breast cancer cells. Our data provide novel insights into the role of the TGF-β pathway in breast tumorigenesis, and establish a clear causative link between the tumor-promoting effects of TGF-β signaling and the metabolic reprogramming of the tumor microenvironment.     

Biosociality,biocitizenship and the new regime of hope and despair: interpreting “Portraits of Hope” and the “Mehmet Case”

The concepts of biocitizenship and biosociality, in many ways developed as a reaction to the former critique of genetification and fears of a return of eugenics, have gained a stronghold in much of the current debates on the social effects of modern-day genetics. In contrast to claims of a return to eugenics, the literature on biocitizenship highlights the new choice-enhancing possibilities involved in present-day biomedicine, underlining the break with past forms of biopower. In this analysis, hope becomes a life-inducing and vitalizing force, opening new avenues of civic participation and engagement. Most critics of this analysis have attacked the claims to novelty attributed to these concepts, arguing that more traditional forms of biopower remain as important as ever. In contrast, we argue that the biocitizenship literature underestimates the radical nature of this break with the past, ending up with a too narrow and one-sided interpretation of the ramifications of the new discourse of hope. On the basis of two different case stories, the “Portraits of Hope” campaign from California, USA and the “Mehmet Case” from Norway, we indicate an alternative “darker” reading of the new discourse of hope, arguing that its driving force is not so much future possibilities as present despair.     

Metabolic reprogramming of cancer-associated fibroblasts by TGF-β drives tumor growth: Connecting TGF-β signaling with “Warburg-like” cancer metabolism and L-lactate production

We have previously shown that a loss of stromal Cav-1 is a biomarker of poor prognosis in breast cancers. Mechanistically, a loss of Cav-1 induces the metabolic reprogramming of stromal cells, with increased autophagy/mitophagy, mitochondrial dysfunction and aerobic glycolysis. As a consequence, Cav-1-low CAFs generate nutrients (such as L-lactate) and chemical building blocks that fuel mitochondrial metabolism and the anabolic growth of adjacent breast cancer cells. It is also known that a loss of Cav-1 is associated with hyperactive TGF-β signaling. However, it remains unknown whether hyperactivation of the TGF-β signaling pathway contributes to the metabolic reprogramming of Cav-1-low CAFs. To address these issues, we overexpressed TGF-β ligands and the TGF-β receptor I (TGFβ-RI) in stromal fibroblasts and breast cancer cells. Here, we show that the role of TGF-β in tumorigenesis is compartment-specific, and that TGF-β promotes tumorigenesis by shifting cancer-associated fibroblasts toward catabolic metabolism. Importantly, the tumor-promoting effects of TGF-β are independent of the cell type generating TGF-β. Thus, stromal-derived TGF-β activates signaling in stromal cells in an autocrine fashion, leading to fibroblast activation, as judged by increased expression of myofibroblast markers, and metabolic reprogramming, with a shift toward catabolic metabolism and oxidative stress. We also show that TGF-β-activated fibroblasts promote the mitochondrial activity of adjacent cancer cells, and in a xenograft model, enhancing the growth of breast cancer cells, independently of angiogenesis. Conversely, activation of the TGF-β pathway in cancer cells does not influence tumor growth, but cancer cell-derived-TGF-β ligands affect stromal cells in a paracrine fashion, leading to fibroblast activation and enhanced tumor growth. In conclusion, ligand-dependent or cell-autonomous activation of the TGF-β pathway in stromal cells induces their metabolic reprogramming, with increased oxidative stress, autophagy/mitophagy and glycolysis, and downregulation of Cav-1. These metabolic alterations can spread among neighboring fibroblasts and greatly sustain the growth of breast cancer cells. Our data provide novel insights into the role of the TGF-β pathway in breast tumorigenesis, and establish a clear causative link between the tumor-promoting effects of TGF-β signaling and the metabolic reprogramming of the tumor microenvironment.     

The cycHJKL genes of Rhizobium meliloti involved in cytochrome c biogenesis are required for “respiratory” nitrate reduction ex planta and for nitrogen fixation during symbiosis

We report the genetic and biochemical analysis of Rhizobium meliloti mutants defective in symbiotic nitrogen fixation (Fix^?) and “respiratory” nitrate reduction (Rnr^?). The mutations were mapped close to the ade-1 and cys-46 chromosomal markers and the mutated locus proved to be identical to the previously described fix-14 locus. By directed Tn5 mutagenesis, a 4.5 kb segment of the chromosome was delimited in which all mutations resulted in Rnr^? and Fix^? phenotypes. Nucleotide sequence analysis of this region revealed the presence of four open reading frames coding for integral membrane and membrane-anchored proteins. Biochemical analysis of the mutants showed that the four proteins were necessary for the biogenesis of all cellular c-type cytochromes. In agreement with the nomenclature proposed for rhizobial genes involved in the formation of c-type cytochromes, the four genes were designated cycH, cycJ, cycK, and cycL, respectively. The predicted protein product of cycH exhibited a high degree of similarity to the Bradyrhizobium japonicum counterpart, while CycK and CycL shared more than 50% amino acid sequence identity with the Rhodobacter capsulatus Ccll and Cc12 proteins, respectively. cycJ encodes a novel membrane anchored protein of 150 amino acids. We suggest that this gene cluster codes for (parts of) a multi-subunit cytochrome c haem lyase. Moreover, our results indicate that in R. meliloti c-type cytochromes are required for respiratory nitrate reduction ex planta, as well as for symbiotic nitrogen fixation in root nodules.     

Post-genomic era and gene discovery for psychiatric diseases: there is a new art of the trade? The example of the HUMTH01 microsatellite in the Tyrosine Hydroxylase gene

The microsatellite HUMTH01, located in the first intron of the Tyrosine Hydroxylase (TH) gene (encoding the rate-limiting enzyme in the synthesis of catecholamines), is characterized by a TCAT repeated motif and has been used in genetic studies of neuropsychiatric and other complex diseases, in which catecholaminergic neurotransmission is implicated. After reporting a positive association between HUMTH01 and bipolar disorder as well as schizophrenia, the authors established that HUMTH01 alleles display the features of regulatory elements. Thereafter, they cloned two proteins (ZNF191 and HBP1), specifically binding to HUMTH01, and demonstrated that allelic variations of HUMTH01 have a quantitative silencing effect on TH gene expression in vitro, and correlate with quantitative and qualitative changes in the binding by ZNF191. The authors aim to characterize the transduction pathway impinging on the HUMTH01 microsatellite and establish its relevance for TH gene regulation in vivo. Since the TCAT repeated sequence is widespread throughout the genome, their approach may lead to the dissection of the mechanisms underlying the quantitative expression of several genes implicated in complex genetic traits, both normal and pathological. Thus, these investigations on the possible contribution and potential role of the HUMTH01 microsatellite in neuro-pathological conditions may represent an example of the different approaches needed to validate genetic targets in the "post-genomic era."     

A new method for examining the complexity and relationships of “timers” in developing systems

Simple methods are developed for analyzing the rate-limiting pathways, or “developmental timers,” for consecutive stages in a developing system. Two conditions are first defined for short and long timing to a developmental stage. Shifts are then performed at time intervals from short to long and long to short conditions. The total time to the stage (time under first condition plus time under second condition) is scored and plotted as a function of the time of shift, resulting in two plots, one for shifts from the short to long condition, and the other for shifts from the long to short condition. Each plot is then analyzed for the number of components, slopes of components, absolute times of origins and termini of components, and discontinuities between components. This information is then used (1) to distinguish between single- and multiple-component timers, (2) to assess the sensitivity of each timer component to the change in the environmental condition employed in the method, including reversibility, (3) to test for the addition of a new timer component under long conditions, and (4) to test for an identity change of a timer component between short and long conditions. These interpretations in turn provide a minimum estimate of the complexity of the rate-limiting pathway to a developmental stage, temporally define major transition points between timer components, and provide some insight into the nature of timer components. By characterizing the rate-limiting pathway from the origin of a developmental program for each consecutive stage in that program, distinctions can also be made between single, parallel, sequential, and branching timer relationships. From these interpretations, a detailed temporal “map” of the rate-limiting program can be generated for any developmental system in which consecutive stages can be reproducibly monitored with time.     

Generation of new transposons in vivo: an evolutionary role for the “staggered” head-to-head dimer and one-ended transposition

From a plasmid carrying the tnpA gene and one inverted repeat sequence (IR) of transposon Tn3, plasmids containing a structure characteristic of transposons, i.e., two IRs flanking a tnpA gene, were generated spontaneously in vivo. They appear to have arisen either through the formation of a “staggered” head-to-head dimer or by so-called one-ended transposition. These putative transposons could indeed transpose to, or form cointegrates with, a recipient plasmid. Based on these findings it is proposed that a primeval transposase gene and its target site evolved first, and subsequently gave rise to a “fully-fledged” transposon by head-to-head dimerization or one-ended transposition. Received: 30 October 1998 / Accepted: 1 April 1999     

“People think that Romanians and Roma are the same”: everyday bordering and the lifting of transitional controls

ABSTRACT

On 1 January 2014 the transitional controls on free movement adopted by the UK when Bulgaria and Romania joined the EU in 2007, ended. This paper demonstrates how the discourses of politicians relating to their removal, amplified via news media contributed to the extension of state bordering practices further into everyday life. Based on ethnographic research into everyday bordering during 2013–15 the paper uses an intersectional framework to explore how this homogenizing, bordering discourse was experienced and contested from differently situated perspectives of Roma and non-Roma social actors from established communities.