Camila Vicencio Esguerra

• Banono, Nancy Saana & Esguerra, Camila (2020). Pharmacological validation of the prepulse inhibition of startle response in larval zebrafish using a commercial automated system and software. Journal of Visualized Experiments.  ISSN 1940-087X.  161 . doi: 10.3791/61423 Full text in Research Archive. Show summary

  Tamoxifen is the most prescribed selective estrogen receptor (ER) modulator in patients with ER-positive breast cancers. Tamoxifen requires the transcription factor paired box 2 protein (PAX2) to repress the transcription of ERBB2/HER2. Now, we identified that PAX2 inhibits cell growth of ER+/HER2− tumor cells in a dose-dependent manner. Moreover, we have identified that cell growth inhibition can be achieved by expressing moderate levels of PAX2 in combination with tamoxifen treatment. Global run-on sequencing of cells overexpressing PAX2, when coupled with PAX2 ChIP-seq, identified common targets regulated by both PAX2 and tamoxifen. The data revealed that PAX2 can inhibit estrogen-induced gene transcription and this effect is enhanced by tamoxifen, suggesting that they converge on repression of the same targets. Moreover, PAX2 and tamoxifen have an additive effect and both induce coding genes and enhancer RNAs (eRNAs). PAX2–tamoxifen upregulated genes are also enriched with PAX2 eRNAs. The enrichment of eRNAs is associated with the highest expression of genes that positivity regulate apoptotic processes. In luminal tumors, the expression of a subset of these proapoptotic genes predicts good outcome and their expression are significantly reduced in tumors of patients with relapse to tamoxifen treatment. Mechanistically, PAX2 and tamoxifen coexert an antitumoral effect by maintaining high levels of transcription of tumor suppressors that promote cell death. The apoptotic effect is mediated in large part by the gene interferon regulatory factor 1. Altogether, we conclude that PAX2 contributes to better clinical outcome in tamoxifen treated ER-positive breast cancer patients by repressing estrogen signaling and inducing cell death related pathways.

• Banono, Nancy Saana; Gawel, Kinga Aurelia; De Witte, Linus & Esguerra, Camila (2020). Zebrafish Larvae Carrying a Splice Variant Mutation in cacna1d: A New Model for Schizophrenia-Like Behaviours?. Molecular Neurobiology.  ISSN 0893-7648.  58, s 877- 894 . doi: 10.1007/s12035-020-02160-5 Full text in Research Archive. Show summary

  Persons with certain single nucleotide polymorphisms (SNPs) in the CACNA1D gene (encoding voltage-gated calcium channel subunit alpha 1-D) have increased risk of developing neuropsychiatric disorders such as bipolar, schizophrenia and autism. The molecular consequences of SNPs on gene expression and protein function are not well understood. Thus, the use of animal models to determine genotype-phenotype correlations is critical to understanding disease pathogenesis. Here, we describe the behavioural changes in larval zebrafish carrying an essential splice site mutation (sa17298) in cacna1da. Heterozygous mutation resulted in 50% reduction of splice variants 201 and 202 (haploinsufficiency), while homozygosity increased transcript levels of variant 201 above wild type (WT; gain-of-function, GOF). Due to low homozygote viability, we focused primarily on performing the phenotypic analysis on heterozygotes. Indeed, cacna1dasa17298/WT larvae displayed hyperlocomotion—a behaviour characterised in zebrafish as a surrogate phenotype for epilepsy, anxiety or psychosis-like behaviour. Follow-up tests ruled out anxiety or seizures, however, as neither thigmotaxis defects nor epileptiform-like discharges in larval brains were observed. We therefore focused on testing for potential “psychosis-like” behaviour by assaying cacna1dasa17298/WT larval locomotor activity under constant light, during light-dark transition and in startle response to dark flashes. Furthermore, exposure of larvae to the antipsychotics, risperidone and haloperidol reversed cacna1da-induced hyperactivity to WT levels while valproate decreased but did not reverse hyperactivity. Together, these findings demonstrate that cacna1da haploinsufficiency induces behaviours in larval zebrafish analogous to those observed in rodent models of psychosis. Future studies on homozygous mutants will determine how cacna1d GOF alters behaviour in this context.

• Gawel, Kinga Aurelia; Kukula-Koch, Wirginia; Nieoczym, Dorota; Stępnik, Katarzyna; van der Ent, Wietske; Banono, Nancy Saana; Tarabasz, Dominik; Turski, Waldemar A. & Esguerra, Camila (2020). The Influence of Palmatine Isolated from Berberis sibirica Radix on Pentylenetetrazole-Induced Seizures in Zebrafish. Cells.  ISSN 2073-4409.  9(5) . doi: 10.3390/cells9051233 Full text in Research Archive. Show summary

  Palmatine (PALM) and berberine (BERB) are widely identified isoquinoline alkaloids among the representatives of the Berberidaceae botanical family. The antiseizure activity of BERB was shown previously in experimental epilepsy models. We assessed the effect of PALM in a pentylenetetrazole (PTZ)-induced seizure assay in zebrafish, with BERB as an active reference compound. Both alkaloids were isolated from the methanolic root extract of Berberis sibirica by counter-current chromatography, and their ability to cross the blood-brain barrier was determined via quantitative structure-activity relationship assay. PALM exerted antiseizure activity, as confirmed by electroencephalographic analysis, and decreased c-fos and bdnf levels in PTZ-treated larvae. In a behavioral assay, PALM dose-dependently decreased PTZ-induced hyperlocomotion. The combination of PALM and BERB in ED16 doses revealed hyperadditive activity towards PTZ-induced hyperlocomotion. Notably, we have indicated that both alkaloids may exert their anticonvulsant activity through different mechanisms of action. Additionally, the combination of both alkaloids in a 1:2.17 ratio (PALM: BERB) mimicked the activity of the pure extract, which indicates that these two active compounds are responsible for its anticonvulsive activity. In conclusion, our study reveals for the first time the anticonvulsant activity of PALM and suggests the combination of PALM and BERB may have higher therapeutic value than separate usage of these compounds.

• Tiraboschi, Ettore; Martina, Silvia; van der Ent, Wietske; Grzyb, Kamil; Gawel, Kinga Aurelia; Cordero-Maldonado, Maria Lorena; Poovathingal, Suresh K.; Heintz, Sarah; Somisetty, Venkata Satheesh; Brattespe, Jarle; Xu, Ju; Suster, Maximiliano; Skupin, Alexander & Esguerra, Camila (2020). New insights into the early mechanisms of epileptogenesis in a zebrafish model of Dravet syndrome. Epilepsia.  ISSN 0013-9580.  61(3), s 549- 560 . doi: 10.1111/epi.16456 Full text in Research Archive. Show summary

  Abstract Objective: To pinpoint the earliest cellular defects underlying seizure onset (epileptogenic period) during perinatal brain development in a new zebrafish model of Dravet syndrome (DS) and to investigate potential disease-modifying activity of the 5HT2 receptor agonist fenfluramine. Methods: We used CRISPR/Cas9 mutagenesis to introduce a missense mutation, designed to perturb ion transport function in all channel isoforms, into scn1lab, the zebrafish orthologue of SCN1A (encoding voltage-gated sodium channel alpha subunit 1). We performed behavioral analysis and electroencephalographic recordings to measure convulsions and epileptiform discharges, followed by single-cell RNA-Seq, morphometric analysis of transgenic reporter-labeled γ-aminobutyric acidergic (GABAergic) neurons, and pharmacological profiling of mutant larvae. Results: Homozygous mutant (scn1labmut/mut ) larvae displayed spontaneous seizures with interictal, preictal, and ictal discharges (mean = 7.5 per 20-minute recording; P < .0001; one-way analysis of variance). Drop-Seq analysis revealed a 2:1 shift in the ratio of glutamatergic to GABAergic neurons in scn1labmut/mut larval brains versus wild type (WT), with dynamic changes in neuronal, glial, and progenitor cell populations. To explore disease pathophysiology further, we quantified dendritic arborization in GABAergic neurons and observed a 40% reduction in arbor number compared to WT (P < .001; n = 15 mutant, n = 16 WT). We postulate that the significant reduction in inhibitory arbors causes an inhibitory to excitatory neurotransmitter imbalance that contributes to seizures and enhanced electrical brain activity in scn1labmut/mut larvae (high-frequency range), with subsequent GABAergic neuronal loss and astrogliosis. Chronic fenfluramine administration completely restored dendritic arbor numbers to normal in scn1labmut/mut larvae, whereas similar treatment with the benzodiazepine diazepam attenuated seizures, but was ineffective in restoring neuronal cytoarchitecture. BrdU labeling revealed cell overproliferation in scn1labmut/mut larval brains that were rescued by fenfluramine but not diazepam. Significance: Our findings provide novel insights into early mechanisms of DS pathogenesis, describe dynamic cell population changes in the scn1labmut/mut brain, and present first-time evidence for potential disease modification by fenfluramine. Keywords: Dravet syndrome; epileptogenesis; fenfluramine; sodium channel; zebrafish.

• Copmans, Daniëlle; Kildgaard, Sara; Rasmussen, Silas A.; Ślęzak, Monika; Dirkx, Nina; Partoens, Michèle; Esguerra, Camila Vicencio; Crawford, Alexander Dettmar; Larsen, Thomas O. & de Witte, Peter A.M. (2019). Zebrafish-based discovery of antiseizure compounds from the north sea: Isoquinoline alkaloids TMC-120A and TMC-120B. Marine Drugs.  ISSN 1660-3397.  17:607(11), s 1- 20 . doi: 10.3390/md17110607 Full text in Research Archive.
• Abudu, Yakubu Princely; Pankiv, Serhiy; Mathai, Benan John; Lystad, Alf Håkon; Bindesbøll, Christian; Brenne, Hanne Britt; Ng, Matthew Yoke Wui; Thiede, Bernd; Yamamoto, Ai; Nthiga, Thaddaeus Mutugi; Lamark, Trond; Esguerra, Camila Vicencio; Johansen, Terje & Simonsen, Anne (2019). NIPSNAP1 and NIPSNAP2 act as “eat-me signal” for mitophagy. Developmental Cell.  ISSN 1534-5807.  49(4), s 509- 525.e12 . doi: 10.1016/j.devcel.2019.03.013 Full text in Research Archive.
• Gawel, Kinga Aurelia; Banono, Nancy Saana; Michalak, Agnieszka & Esguerra, Camila Vicencio (2019). A critical review of zebrafish schizophrenia models: Time for validation?. Neuroscience and Biobehavioral Reviews.  ISSN 0149-7634.  107, s 6- 22 . doi: 10.1016/j.neubiorev.2019.08.001
• Nieoczym, Dorota; Socala, Katarzyna; Gawel, Kinga Aurelia; Esguerra, Camila Vicencio; Wyska, Elzbieta & Wlaz, Piotr (2019). Anticonvulsant Activity of Pterostilbene in Zebrafish and Mouse Acute Seizure Tests. Neurochemical Research.  ISSN 0364-3190.  44(5), s 1043- 1055 . doi: 10.1007/s11064-019-02735-2 Full text in Research Archive.
• Sisodiya, Sanjay M.; Fowler, Hayley J.; Lake, Iain; Nanji, Rosemary Ogwen; Gawel, Kinga Aurelia; Esguerra, Camila Vicencio; Newton, Charles & Foley, Aideen (2019). Climate change and epilepsy: Time to take action. Epilepsia Open.  ISSN 2470-9239.  4(4), s 524- 536 . doi: 10.1002/epi4.12359 Full text in Research Archive.
• Leclercq, K; Afrikanova, Tatiana; Langlois, M.; De Prins, A; Buenafe, OE; Rospo, CC; Van Eeckhaut, A; deWitte, P; Crawford, AD; Smolders, I; Esguerra, Camila Vicencio & Kaminski, RM (2015). Cross-species pharmacological characterization of the allylglycine seizure model in mice and larval zebrafish. Epilepsy & Behavior.  ISSN 1525-5050.  45, s 53- 63 . doi: 10.1016/j.yebeh.2015.03.019
• Schubert, Julian; Siekierska, Aleksandra; Langlois, Melanie; May, Patrick; Huneau, Clément; Becker, Felicitas; Muhle, Hiltrud; Suls, Arvid; Lemke, Johannes R.; de Kovel, Carolien G.F.; Thiele, Holger; Konrad, Kathryn; Kawalia, Amit; Toliat, Mohammad Reza; Sander, Thomas; Ruschendorf, Franz; Caliebe, Almuth; Nagel, Inga; Kohl, Bernard; Kecskés, Angéla; Jacmin, Maxime; Hardies, Katia; Weckhuysen, Sarah; Riesch, Erik; Dorn, Thomas; Brilstra, Eva H.; Baulac, Stephanie; Møller, Rikke S.; Hjalgrim, Helle; Koeleman, Bobby P.C.; Jurkat-Rott, Karin; Lehmann-Horn, Frank; Roach, Jared C.; Glusman, Gustavo; Hood, Leroy; Galas, David J.; Martin, Benoit; De Witte, Peter A.M.; Biskup, Saskia; De Jonghe, Peter; Helbig, Ingo; Balling, Rudi; Nürnberg, Peter; Crawford, Alexander D.; Esguerra, Camila Vicencio; Weber, Yvonne G. & Lerche, Holger (2014). Mutations in STX1B, encoding a presynaptic protein, cause fever-associated epilepsy syndromes. Nature Genetics.  ISSN 1061-4036.  46, s 1327- 1332 . doi: 10.1038/ng.3130

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• Banono, Nancy Saana; Gawel, Kinga Aurelia; De Witte, Linus & Esguerra, Camila (2020). Zebrafish larvae carrying a mutation in cacna1d display schizophrenia-like behaviours.
• Song, You; Hultman, Maria Thérése; Moe, Jannicke; Petersen, Karina; Rundberget, Jan Thomas; Xie, Li; Cao, Yang; Esguerra, Camila Vicencio; De Bin, Riccardo; Lee, YeonKyeong; Solhaug, Knut Asbjørn; Kamstra, Jorke H; Legler, Juliette; Iguchi, Taisen; Lillicrap, Adam David; Andersen, Sjur; Villeneuve, Daniel L. & Tollefsen, Knut-Erik (2020). The RiskAOP project - Quantitative Adverse Outcome Pathway assisted risk assessment of mitochondrial toxicants.

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