Publications
215 results found
Arcidiacono P, Ragonese F, Stabile A, et al., 2018, Antitumor activity and expression profiles of genes induced by sulforaphane in human melanoma cells, European Journal of Nutrition, Vol: 57, Pages: 2547-2569, ISSN: 0044-264X
PurposeHuman melanoma is a highly aggressive incurable cancer due to intrinsic cellular resistance to apoptosis, reprogramming, proliferation and survival during tumour progression. Sulforaphane (SFN), an isothiocyanate found in cruciferous vegetables, plays a role in carcinogenesis in many cancer types. However, the cytotoxic molecular mechanisms and gene expression profiles promoted by SFN in human melanoma remain unknown.MethodsThree different cell lines were used: two human melanoma A375 and 501MEL and human epidermal melanocytes (HEMa). Cell viability and proliferation, cell cycle analysis, cell migration and invasion and protein expression and phosphorylation status of Akt and p53 upon SFN treatment were determined. RNA-seq of A375 was performed at different time points after SFN treatment.ResultsWe demonstrated that SFN strongly decreased cell viability and proliferation, induced G2/M cell cycle arrest, promoted apoptosis through the activation of caspases 3, 8, 9 and hampered migration and invasion abilities in the melanoma cell lines. Remarkably, HEMa cells were not affected by SFN treatment. Transcriptomic analysis revealed regulation of genes involved in response to stress, apoptosis/cell death and metabolic processes. SFN upregulated the expression of pro-apoptotic genes, such as p53, BAX, PUMA, FAS and MDM2; promoted cell cycle inhibition and growth arrest by upregulating EGR1, GADD45B, ATF3 and CDKN1A; and simultaneously acted as a potent inhibitor of genotoxicity by launching the stress-inducible protein network (HMOX1, HSPA1A, HSPA6, SOD1).ConclusionOverall, the data show that SFN cytotoxicity in melanoma derives from complex and concurrent mechanisms during carcinogenesis, which makes it a promising cancer prevention agent.
Burt A, Crisanti A, 2018, Editorial: gene drive for vector control, Pathogens and Global Health, Vol: 111, Pages: 397-398, ISSN: 2047-7724
Arcidiacono P, Stabile AM, Ragonese F, et al., 2018, Anticarcinogenic activities of sulforaphane are influenced by Nerve Growth Factor in human melanoma A375 cells, FOOD AND CHEMICAL TOXICOLOGY, Vol: 113, Pages: 154-161, ISSN: 0278-6915
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- Citations: 6
Burt A, Crisanti A, 2018, Gene drive: evolved and synthetic, ACS Chemical Biology, Vol: 13, Pages: 343-346, ISSN: 1554-8929
Drive is a process of accelerated inheritance from one generation to the next that allows some genes to spread rapidly through populations even if they do not contribute to-or indeed even if they detract from-organismal survival and reproduction. Genetic elements that can spread by drive include gametic and zygotic killers, meiotic drivers, homing endonuclease genes, B chromosomes, and transposable elements. The fact that gene drive can lead to the spread of fitness-reducing traits (including lethality and sterility) makes it an attractive process to consider exploiting to control disease vectors and other pests. There are a number of efforts to develop synthetic gene drive systems, particularly focused on the mosquito-borne diseases that continue to plague us.
Burt A, Coulibaly M, Crisanti A, et al., 2018, Gene drive to reduce malaria transmission in sub-Saharan Africa, Journal of Responsible Innovation, Vol: 5, Pages: S66-S80, ISSN: 2329-9460
Despite impressive progress, malaria continues to impose a substantial burden of mortality and morbidity, particularly in sub-Saharan Africa, and new tools will be needed to achieve elimination. Gene drive is a natural process by which some genes are inherited at a greater-than-Mendelian rate and can spread through a population even if they cause harm to the organisms carrying them. Many different synthetic gene drive systems have been proposed to suppress the number of mosquitoes and/or reduce vector competence. As with any control measure, due attention should be paid to the possible evolution of resistance. No gene drive construct has yet been reported that is ‘field-ready’ for release, and when such constructs are developed, they should be assessed on a case-by-case basis. Gene drive approaches to vector control promise to have a number of key features that motivate their continued development, and scrutiny, by all concerned.
Simoes ML, Dong Y, Hammond AM, et al., 2017, ENGINEERED <i>ANOPHELES GAMBIAE</i> IMMUNITY TO <i>PLASMODIUM</i> INFECTION, 65th Annual Meeting of the American-Society-of-Tropical-Medicine-and-Hygiene (ASTMH), Publisher: AMER SOC TROP MED & HYGIENE, Pages: 444-444, ISSN: 0002-9637
Hammond AM, Kyrou K, Bruttini M, et al., 2017, The creation and selection of mutations resistant to a gene drive over multiple generations in the malaria mosquito, PLoS Genetics, Vol: 13, ISSN: 1553-7390
Gene drives have enormous potential for the control of insect populations of medical and agricultural relevance. By preferentially biasing their own inheritance, gene drives can rapidly introduce genetic traits even if these confer a negative fitness effect on the population. We have recently developed gene drives based on CRISPR nuclease constructs that are designed to disrupt key genes essential for female fertility in the malaria mosquito. The construct copies itself and the associated genetic disruption from one homologous chromosome to another during gamete formation, a process called homing that ensures the majority of offspring inherit the drive. Such drives have the potential to cause long-lasting, sustainable population suppression, though they are also expected to impose a large selection pressure for resistance in the mosquito. One of these population suppression gene drives showed rapid invasion of a caged population over 4 generations, establishing proof of principle for this technology. In order to assess the potential for the emergence of resistance to the gene drive in this population we allowed it to run for 25 generations and monitored the frequency of the gene drive over time. Following the initial increase of the gene drive we observed a gradual decrease in its frequency that was accompanied by the spread of small, nuclease-induced mutations at the target gene that are resistant to further cleavage and restore its functionality. Such mutations showed rates of increase consistent with positive selection in the face of the gene drive. Our findings represent the first documented example of selection for resistance to a synthetic gene drive and lead to important design recommendations and considerations in order to mitigate for resistance in future gene drive applications.
Bernardini F, Galizi R, Wunderlich M, et al., 2017, Cross-Species Y Chromosome Function Between Malaria Vectors of the Anopheles gambiae Species Complex., Genetics, ISSN: 0016-6731
Y chromosome function, structure and evolution is poorly understood in many species including the Anopheles genus of mosquitoes, an emerging model system for studying speciation that also represents the major vectors of malaria. While the Anopheline Y had previously been implicated in male mating behavior, recent data from the Anopheles gambiae complex suggests that, apart from the putative primary sex-determiner, no other genes are conserved on the Y. Studying the functional basis of the evolutionary divergence of the Y chromosome in the gambiae complex is complicated by complete F1 male hybrid sterility. Here we used an F1xF0 crossing scheme to overcome a severe bottleneck of male hybrid incompatibilities and enabled us to experimentally purify a genetically labelled A. gambiae Y chromosome in an A. arabiensis background. Whole genome sequencing confirmed that the A. gambiae Y retained its original sequence content in the A. arabiensis genomic background. In contrast to comparable experiments in Drosophila, we find that the presence of a heterospecific Y chromosome has no significant effect on the expression of A. arabiensis genes and transcriptional differences can be explained almost exclusively as a direct consequence of transcripts arising from sequence elements present on the A. gambiae Y chromosome itself. We find that Y hybrids show no obvious fertility defects and no substantial reduction in male competitiveness. Our results demonstrate that, despite their radically different structure, Y chromosomes of these two species of the gambiae complex that diverged an estimated 1.85Myr ago function interchangeably, thus indicating that the Y chromosome does not harbor loci contributing to hybrid incompatibility. Therefore, Y chromosome gene flow between members of the gambiae complex is possible even at their current level of divergence. Importantly, this also suggests that malaria control interventions based on sex-distorting Y drive would be transferable, whethe
Werther R, Hallinan JP, Lambert AR, et al., 2017, Crystallographic analyses illustrate significant plasticity and efficient recoding of meganuclease target specificity, Nucleic Acids Research, Vol: 45, Pages: 8621-8634, ISSN: 0305-1048
The retargeting of protein–DNA specificity, outsideof extremely modular DNA binding proteins suchas TAL effectors, has generally proved to be quitechallenging. Here, we describe structural analysesof five different extensively retargeted variants of asingle homing endonuclease, that have been shownto function efficiently in ex vivo and in vivo applications.The redesigned proteins harbor mutationsat up to 53 residues (18%) of their amino acid sequence,primarily distributed across the DNA bindingsurface, making them among the most signifi-cantly reengineered ligand-binding proteins to date.Specificity is derived from the combined contributionsof DNA-contacting residues and of neighboringresidues that influence local structural organization.Changes in specificity are facilitated by theability of all those residues to readily exchange bothform and function. The fidelity of recognition is notprecisely correlated with the fraction or total numberof residues in the protein–DNA interface that areactually involved in DNA contacts, including directionalhydrogen bonds. The plasticity of the DNArecognitionsurface of this protein, which allows substantialretargeting of recognition specificity withoutrequiring significant alteration of the surroundingprotein architecture, reflects the ability of the correspondinggenetic elements to maintain mobility andpersistence in the face of genetic drift within potentialhost target sites.
NOLAN T, CRISANTI A, 2017, Using gene drives to limit the spread of malaria, Scientist, Vol: 31, ISSN: 0890-3670
Beaghton A, Hammond A, Nolan T, et al., 2017, Requirements for Driving Antipathogen Effector Genes into Populations of Disease Vectors by Homing, Genetics, Vol: 205, Pages: 1587-1596
Simoes ML, Dong Y, Hammond A, et al., 2017, The Anopheles FBN9 immune factor mediates Plasmodium species-specific defense through transgenic fat body expression, Developmental and Comparative Immunology, Vol: 67, Pages: 257-265
Li X, Huang J, Zhang M, et al., 2016, Human CD8+T cells mediate protective immunity induced by a human malaria vaccine in human immune system mice, Vaccine, Vol: 34, Pages: 4501-4506, ISSN: 0264-410X
A number of studies have shown that CD8+ T cells mediate protective anti-malaria immunity in a mouse model. However, whether human CD8+ T cells play a role in protection against malaria remains unknown. We recently established human immune system (HIS) mice harboring functional human CD8+ T cells (HIS-CD8 mice) by transduction with HLA-A∗0201 and certain human cytokines using recombinant adeno-associated virus-based gene transfer technologies. These HIS-CD8 mice mount a potent, antigen-specific HLA-A∗0201-restricted human CD8+ T-cell response upon immunization with a recombinant adenovirus expressing a human malaria antigen, the Plasmodium falciparum circumsporozoite protein (PfCSP), termed AdPfCSP. In the present study, we challenged AdPfCSP-immunized HIS-CD8 mice with transgenic Plasmodium berghei sporozoites expressing full-length PfCSP and found that AdPfCSP-immunized (but not naïve) mice were protected against subsequent malaria challenge. The level of the HLA-A∗0201-restricted, PfCSP-specific human CD8+ T-cell response was closely correlated with the level of malaria protection. Furthermore, depletion of human CD8+ T cells from AdPfCSP-immunized HIS-CD8 mice almost completely abolished the anti-malaria immune response. Taken together, our data show that human CD8+ T cells mediate protective anti-malaria immunity in vivo.
Kohl A, Pondeville E, Schnettler E, et al., 2016, Advancing vector biology research: a community survey for future directions, research applications and infrastructure requirements., Pathogens and Global Health, Vol: 110, Pages: 164-172, ISSN: 2047-7732
Vector-borne pathogens impact public health, animal production, and animal welfare. Research on arthropod vectors such as mosquitoes, ticks, sandflies, and midges which transmit pathogens to humans and economically important animals is crucial for development of new control measures that target transmission by the vector. While insecticides are an important part of this arsenal, appearance of resistance mechanisms is increasingly common. Novel tools for genetic manipulation of vectors, use of Wolbachia endosymbiotic bacteria, and other biological control mechanisms to prevent pathogen transmission have led to promising new intervention strategies, adding to strong interest in vector biology and genetics as well as vector-pathogen interactions. Vector research is therefore at a crucial juncture, and strategic decisions on future research directions and research infrastructure investment should be informed by the research community. A survey initiated by the European Horizon 2020 INFRAVEC-2 consortium set out to canvass priorities in the vector biology research community and to determine key activities that are needed for researchers to efficiently study vectors, vector-pathogen interactions, as well as access the structures and services that allow such activities to be carried out. We summarize the most important findings of the survey which in particular reflect the priorities of researchers in European countries, and which will be of use to stakeholders that include researchers, government, and research organizations.
Galizi R, Hammond A, Kyrou K, et al., 2016, A CRISPR-Cas9 sex-ratio distortion system for genetic control., Scientific Reports, Vol: 6, ISSN: 2045-2322
Genetic control aims to reduce the ability of insect pest populations to cause harm via the release of modified insects. One strategy is to bias the reproductive sex ratio towards males so that a population decreases in size or is eliminated altogether due to a lack of females. We have shown previously that sex ratio distortion can be generated synthetically in the main human malaria vector Anopheles gambiae, by selectively destroying the X-chromosome during spermatogenesis, through the activity of a naturally-occurring endonuclease that targets a repetitive rDNA sequence highly-conserved in a wide range of organisms. Here we describe a CRISPR-Cas9 sex distortion system that targets ribosomal sequences restricted to the member species of the Anopheles gambiae complex. Expression of Cas9 during spermatogenesis resulted in RNA-guided shredding of the X-chromosome during male meiosis and produced extreme male bias among progeny in the absence of any significant reduction in fertility. The flexibility of CRISPR-Cas9 combined with the availability of genomic data for a range of insects renders this strategy broadly applicable for the species-specific control of any pest or vector species with an XY sex-determination system by targeting sequences exclusive to the female sex chromosome.
Hall AB, Papathanos P-A, Sharma A, et al., 2016, Radical remodeling of the Y chromosome in a recent radiation of malaria mosquitoes, Proceedings of the National Academy of Sciences of the United States of America, Vol: 113, Pages: E2114-E2123
Nolan T, Crisanti A, 2016, DRIVING OUT MALARIA, Scientist, Vol: 31, Pages: 24-31
Hammond A, Galizi R, Kyrou K, et al., 2016, A CRISPR-Cas9 gene drive system-targeting female reproduction in the malaria mosquito vector Anopheles gambiae, Nature Biotechnology, Vol: 34, Pages: 78-83, ISSN: 1087-0156
Gene drive systems that enable super-Mendelian inheritance of a transgene have the potential to modify insect populations over a timeframe of a few years. We describe CRISPR-Cas9 endonuclease constructs that function as gene drive systems in Anopheles gambiae, the main vector for malaria. We identified three genes (AGAP005958, AGAP011377 and AGAP007280) that confer a recessive female-sterility phenotype upon disruption, and inserted into each locus CRISPR-Cas9 gene drive constructs designed to target and edit each gene. For each targeted locus we observed a strong gene drive at the molecular level, with transmission rates to progeny of 91.4 to 99.6%. Population modeling and cage experiments indicate that a CRISPR-Cas9 construct targeting one of these loci, AGAP007280, meets the minimum requirement for a gene drive targeting female reproduction in an insect population. These findings could expedite the development of gene drives to suppress mosquito populations to levels that do not support malaria transmission.
Tetteh KK, Helb D, Verra F, et al., 2015, IMMUNE PROFILING OF <i>PLASMODIUM FALCIPARUM</i> ENDEMIC SERA USING BESPOKE PROTEIN MICROARRAYS, Publisher: AMER SOC TROP MED & HYGIENE, Pages: 488-488, ISSN: 0002-9637
Canavese M, Dottorini T, Crisanti A, 2015, VEGF and LPS synergistically silence inflammatory response to <i>Plasmodium berghei</i> infection and protect against cerebral malaria, PATHOGENS AND GLOBAL HEALTH, Vol: 109, Pages: 255-265, ISSN: 2047-7724
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- Citations: 9
Canavese M, Crisanti A, 2015, Vascular endothelial growth factor (VEGF) and lovastatin suppress the inflammatory response to <i>Plasmodium berghei</i> infection and protect against experimental cerebral malaria, PATHOGENS AND GLOBAL HEALTH, Vol: 109, Pages: 266-274, ISSN: 2047-7724
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- Citations: 4
Facchinelli L, Valerio L, Lees RS, et al., 2015, Stimulating <i>Anopheles gambiae</i> swarms in the laboratory: application for behavioural and fitness studies, MALARIA JOURNAL, Vol: 14, ISSN: 1475-2875
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- Citations: 20
Dritsou V, Topalis P, Windbichler N, et al., 2015, A draft genome sequence of an invasive mosquito: an Italian <i>Aedes</i> <i>albopictus</i>, PATHOGENS AND GLOBAL HEALTH, Vol: 109, Pages: 207-220, ISSN: 2047-7724
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- Citations: 25
Lin J-W, Spaccapelo R, Schwarzer E, et al., 2015, Replication of Plasmodium in reticulocytes can occur without hemozoin formation, resulting in chloroquine resistance, Journal of Experimental Medicine, Vol: 212, Pages: 893-903, ISSN: 1540-9538
Most studies on malaria-parasite digestion of hemoglobin (Hb) have been performed using P. falciparum maintained in mature erythrocytes, in vitro. In this study, we examine Plasmodium Hb degradation in vivo in mice, using the parasite P. berghei, and show that it is possible to create mutant parasites lacking enzymes involved in the initial steps of Hb proteolysis. These mutants only complete development in reticulocytes and mature into both schizonts and gametocytes. Hb degradation is severely impaired and large amounts of undigested Hb remains in the reticulocyte cytoplasm and in vesicles in the parasite. The mutants produce little or no hemozoin (Hz), the detoxification by-product of Hb degradation. Further, they are resistant to chloroquine, an antimalarial drug that interferes with Hz formation, but their sensitivity to artesunate, also thought to be dependent on Hb degradation, is retained. Survival in reticulocytes with reduced or absent Hb digestion may imply a novel mechanism of drug resistance. These findings have implications for drug development against human-malaria parasites, such as P. vivax and P. ovale, which develop inside reticulocytes.
Guiyedi V, Becavin C, Herbert F, et al., 2015, Asymptomatic Plasmodium falciparum infection in children is associated with increased auto-antibody production, high IL-10 plasma levels and antibodies to merozoite surface protein 3, Malaria Journal, Vol: 14, ISSN: 1475-2875
BackgroundMechanisms of acquired protection to malaria in asymptomatic Plasmodium falciparum carriers are only partially understood. Among them, the role plays by the self-reactive antibodies has not been clarified yet. In this study, the relationship between repertoires of circulating self-reactive and parasite-specific immunoglobulin G (IgG), their correlation with cytokine levels, and their association with protection against malaria was investigated in asymptomatic Plasmodium falciparum-infected Gabonese children.MethodsThe diversity of P. falciparum-specific antibody repertoire was analysed using a protein micro-array immunoassay, the total auto-antibody repertoire by quantitative immunoblotting and circulating cytokine levels were measured by ELISA in endemic controls (EC) and P. falciparum-infected children from Gabon with asymptomatic (AM) or mild malaria (MM). The association of self- and parasite-specific antibody repertoires with circulating cytokines was evaluated using single linkage hierarchical clustering, Kruskal – Wallis tests and Spearman’s rank correlation.ResultsChildren with AM exhibited an IgG response to merozoite surface protein 3 (MSP3) but not to MSP1-19, although their levels of total P. falciparum-specific IgG were similar to those in the MM group. Moreover, the asymptomatic children had increased levels of autoantibodies recognising brain antigens. In addition, a correlation between IL-10 levels and parasite load was found in AM and MM children. These two groups also exhibited significant correlations between plasma levels of IL-10 and IFN-γ with age and with total plasma IgG levels. IL-10 and IFN-γ levels were also associated with auto-antibody responses in AM.ConclusionsAltogether, these results indicate that a self-reactive polyclonal response associated with increased IgG to MSP3 and high plasma levels of IL-10 and IFN-γ may contribute to protective immune mechanisms triggered in asymptomatic P. falcipar
Simoni A, Siniscalchi C, Chan YS, et al., 2015, Development of synthetic selfish elements based on modular nucleases in Drosophila melanogaster (vol 42, pg 7461, 2014), NUCLEIC ACIDS RESEARCH, Vol: 43, Pages: 2991-2991, ISSN: 0305-1048
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- Citations: 1
Crisanti A, 2015, Untitled, PATHOGENS AND GLOBAL HEALTH, Vol: 109, Pages: 1-1, ISSN: 2047-7724
Castellano L, Rizzi E, Krell J, et al., 2015, The germline of the malaria mosquito produces abundant miRNAs, endo-siRNAs, piRNAs and 29-nt small RNAs, Bmc Genomics, Vol: 16
Neafsey DE, Waterhouse RM, Abai MR, et al., 2015, Highly evolvable malaria vectors: The genomes of 16 Anopheles mosquitoes, Science, Vol: 347
Schwenk R, Debot M, Porter M, et al., 2014, IgG2 Antibodies against a Clinical Grade Plasmodium falciparum CSP Vaccine Antigen Associate with Protection against Transgenic Sporozoite Challenge in Mice, PLOS One, Vol: 9, ISSN: 1932-6203
The availability of a highly purified and well characterized circumsporozoite protein (CSP) is essential to improve upon the partial success of recombinant CSP-based malaria vaccine candidates. Soluble, near full-length, Plasmodium falciparum CSP vaccine antigen (CS/D) was produced in E. coli under bio-production conditions that comply with current Good Manufacturing Practices (cGMP). A mouse immunogenicity study was conducted using a stable oil-in-water emulsion (SE) of CS/D in combination with the Toll-Like Receptor 4 (TLR4) agonist Glucopyranosyl Lipid A (GLA/SE), or one of two TLR7/8 agonists: R848 (un-conjugated) or 3M-051 (covalently conjugated). Compared to Alum and SE, GLA/SE induced higher CS/D specific antibody response in Balb/c mice. Subclass analysis showed higher IgG2:IgG1 ratio of GLA/SE induced antibodies as compared to Alum and SE. TLR synergy was not observed when soluble R848 was mixed with GLA/SE. Antibody response of 3M051 formulations in Balb/c was similar to GLA/SE, except for the higher IgG2:IgG1 ratio and a trend towards higher T cell responses in 3M051 containing groups. However, no synergistic enhancement of antibody and T cell response was evident when 3M051 conjugate was mixed with GLA/SE. In C57Bl/6 mice, CS/D adjuvanted with 3M051/SE or GLA/SE induced higher CSP repeat specific titers compared to SE. While, 3M051 induced antibodies had high IgG2c:IgG1 ratio, GLA/SE promoted high levels of both IgG1 and IgG2c. GLA/SE also induced more potent T-cell responses compared to SE in two independent C57/BL6 vaccination studies, suggesting a balanced and productive TH1/TH2 response. GLA and 3M-051 similarly enhanced the protective efficacy of CS/D against challenge with a transgenic P. berghei parasite and most importantly, high levels of cytophilic IgG2 antibodies were associated with protection in this model. Our data indicated that the cGMP-grade, soluble CS/D antigen combined with the TLR4-containing adjuvant GLA/SE warrants further evaluati
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