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Fyrir marga lyfjaflokka þykir staðbundin lyfjagjöf í augndropaformi ákjósanlegasta leiðin vegna einfaldleika við lyfjagjöf. Eru augndropar algengasta aðferðin til að meðhöndla augnsjúkdóma í fremri hluta augans. Staðbundin lyfjagjöf í augndropaformi yfir hornhimnu er hinsvegar óhagkvæm þegar meðhöndla þarf aftari hluta augans. Hefðbundin lyfjameðferð fyrir aftari hluta augans felur í sér að lyfinu er dælt í glerhlaup augans en með tilheyrandi áhættu á fylgikvillum ásamt tilheyrandi óþægindum fyrir sjúklinga. Sýnt hefur verið fram á að leiðin í gegnum augnslímhúð og augnhvítu hefur hlutfallslega hærra gegndræpi fyrir risasameindir, stærra aðgengilegt yfirborð og meiri nálægð við sjónhimnu samanborið við hornhimnu. Fyrir vikið hafa verið kenningar um að augnslímhúð og augnhvíta sé mögulega leið lyfja í augndropaformi yfir í aftari hluta augans. Meginmarkmið þessarar ritgerðar eru (1) að kanna hversu vel ákveðin lyf í sýklódextrín augndropaformi flytjast yfir í fremri og aftari hluta augans, (2) að ákvarða lyfjafræðileg áhrif angíótensínviðtaka-hamlara á augnþrýsting eftir augndropagjöf, (3) að meta og bera saman in vitro gegndræpi í gegnum hornhimnu og í gegnum augnslímhúð og augnhvítu fyrir staðbundna augndropalyfjagjöf í aftari hluta augans og (4) að bera kennsl á helstu eðlisefnafræðilega eiginleika sem hafa áhrif á gegndræpi lyfja yfir hornhimnu í fullri þykkt og leiðina frá augnslímhúð, í gegnum augnhvítu og æðahimnu að sjónhimnu (augnslímhúð-augnhvítu-æðahimnu-sjónhimnu leið). Aðferðir 1. Framkvæmdar voru tvær in vivo rannsóknir á kanínum þar sem mældur var lyfjastyrkur í augnvefjum eftir stakan skammt af augnlyfjum í sýklódextrín nanóögnum. Í fyrri rannsókninni voru tveir mismunandi angíótensínviðtakahamlarar, 1.5% irbesartan og 0.15% candesartan, gefnir 49 kanínum. Styrkur lyfjanna var mældur í hornhimnu og augnvökva eftir 1, 2, 3, 6 og 12 klukkustundir frá lyfjagjöf. Í seinni rannsókninni var ný γ-sýklódextrín öragna augnlyfjaferja með cediranib maleate (öflugur VEGF hamlari) prófuð á 26 kanínum. Styrkur cediranib maleate var mældur í vefjum fremri og aftari hluta augans eftir 1, 3, og klukkustundir frá lyfjagjöf. 2. Augnþrýstingslækkandi áhrif irbesartan og candesartan voru mæld í 10 kanínum. Algengt glákuaugndropalyf, 0.5% timolol, var síðan notað til samanburðar. Augnþrýstingur var mældur með sérstökum augnþrýstingsmæli á tíma 0, 2 og 4 klukkustundum eftir stakan lyfjaskammt. 3. Framkvæmd var in vitro rannsókn á augnvef úr svínum með Franz-sveimklefa til að bera saman gegndræpi 27 mismunandi efnasambanda yfir hornhimnu og augnslímhúð – augnhvítu – æðahimnu og sjónhimnu. Reiknað var út flæði lyfja og gegndræpisstuðlar fyrir hvert efnasamband og vef. Lýsing á 39 sameindum fyrir efnasamböndin sem voru notuð, voru fengin frá ákveðnum gagnagrunnum á alnetinu og notuð til að reikna út fylgni við gegndræpisstuðul með Spearmans rho fylgnistuðul. Niðurstöður 1. Styrkur irbesartan í augnvökva meðferðarauga fór upp í 282 ± 159 ng/g (meðaltal ± SD, n = 26) og fyrir candesartan 70 ± 73 ng/g. Hæsti styrkur í hornhimnu var 3662,6 ± 987,7 ng/g fyrir irbesartan og 3503,9 ± 801,5 ng/g fyrir candesartan. Cediranib maleate mældist í sjónhimnu í styrkleika 1070 ± 1404 nM og í glerhlaupi 19 ± 9 nM (n = 6). 2. Candesartan hafði mest lækkandi áhrif á augnþrýsting, með breytingu upp á 5,6 mmHg fjórum klukkustundum eftir lyfjagjöf samanborið við 4,0 mmHg fyrir irbesartan og 4,6 mmHg fyrir timolol. 3. 22 af 27 efnasamböndum (81%) sýndu hærra gegndræpi í gegnum augnslímhúð-augnhvítu-æðahimnu-sjónhimnu leiðina, samanborið við hornhimnu. Meðalgegndræpi fyrir augnslímhúð-augnhvítu-æðahimnu-sjónhimnu vefina var 2,9 sinnum hærra en gegndræpi hornhimnunnar. Fylgni var á milli aukins vatnsleysnis og meira gegndræpis yfir bæði hornhimnu í fullri þykkt og augnslímhúð-augnhvítu-æðahimnu-sjónhimnu-leiðina, á meðan hærri fitusækni, mólmassi og fjöldi hringa leiddi til lægri gegndræpisstuðla. Umræða Irbesartan, candesartan og cediranib mældust í styrk af tilætluðum lyfjafræðilegum áhrifum í fremri og aftari hluta augans eftir lyfjagjöf með sýklódextrin augnlyfjaferjum. Niðurstöðurnar gefa til kynna staðbundna dreifingu lyfjanna sem gæti komið í veg fyrir þörf á inngripsmiklum lyfjagjöfum. Irbesartan og candesartan í augndropaformi lækkuðu augnþrýsting hjá kanínum með góðum árangri, sem gæti bent til nýs flokks glákulyfja. In vitro gögn okkar benda til þess að fyrir mörg efnasambönd gæti augnslímhúðaugnhvítu leiðin verið áhrifameiri og mögulega eina leiðin til að skila lyfjum í vefi í aftari hluta augans með staðbundinni augndropalyfjagjöf., Purpose: Topical ocular administration is the preferred route for many classes of drugs because of its ease of administration. It is the most common way of treating eye diseases in the anterior segment. Topical administration through the cornea, however, is inefficient for drug delivery to the back of the eye. Intravitreal injections are the standard drug treatment for posterior segment disorders but are associated with the risk of complications and patient discomfort. The conjunctiva-sclera route has shown a relatively higher permeability to macromolecules, a larger surface of exposed tissue and a closer location to the retina when compared to the cornea. Consequently, it has been suggested as an alternative route for topical drug delivery to the posterior segment. The main goals of this thesis are (1) to test the efficacy of cyclodextrin-based eye drop formulations in delivering certain drugs to the anterior and posterior segments of the eye, (2) to determine the pharmacological effect of angiotensin receptor blockers on intraocular pressure after topical administration, (3) to evaluate and compare the in vitro permeabilities of the corneal and noncorneal routes for topical drug delivery to the posterior segment and (4) to identify the main physicochemical properties affecting the permeation of drugs across full-thickness cornea and conjunctiva-sclera-choroid-retina. Methods: 1. Two separate in vivo studies were performed in rabbits to measure the drug concentration in the intraocular tissues after a single dose administration of cyclodextrin-based nanosuspension eye drops. In the first study, two different angiotensin receptor blockers, 1.5% irbesartan and 0.15% candesartan, were administered to 49 rabbits and the drug concentrations in the cornea and aqueous humour were quantified at 1, 2, 3, 6 and 12 hours after administration. In the second study, a novel γ-cyclodextrin nanoparticle eye drops formulation containing cediranib maleate, was applied to 26 rabbits. The concentration of cediranib maleate in the tissues of both anterior and posterior segments was quantified at 1, 3 and 6 hours after administration. 2. The potential lowering effect of irbesartan and candesartan on intraocular pressure was investigated in 10 rabbits. The IOP was measured at 0, 2 and 4 hours after a single-dose administration using a rebound tonometer and compared with a commercial 0.5% timolol solution. 3. In vitro studies were performed using porcine ocular tissues mounted on Franzdiffusion cells to compare the permeability of 27 different compounds across vi full-thickness cornea and conjunctiva-sclera-choroid-retina. The drug fluxes and permeability coefficients were calculated for each compound and tissue. 39 molecular descriptors were obtained for our compounds from different online databases and correlated with their permeability coefficient using Spearman’s rho. Results: 1. Irbesartan and candesartan reached maximum concentrations in the aqueous humour of the treated eyes three hours after administration. Cediranib maleate was found in all studied tissues, and the highest concentrations in the retina and vitreous were found 1 hour after the administration. 2. Irbesartan and candesartan showed an IOP lowering effect that was equivalent to that of timolol four hours after the administration. 3. Regarding the in vitro permeability data, 22 out of 27 compounds (81%) showed a higher permeability across conjunctiva-sclera-choroid-retina compared with the full-thickness cornea. The mean permeability of the noncorneal route was 2.9 times higher than the cornea. The correlation analysis showed that charge, mass and size, lipophilicity, and the ability to form hydrogen bonds were the most relevant properties for corneal permeability, while lipophilicity and the number of rings and rotatable bonds were more important for the noncorneal route. Discussion: The drugs were found at pharmacologically relevant concentrations in both anterior and posterior segments of the eye after topical administration of cyclodextrinbased formulations. These results indicate a local delivery that could contribute to minimizing systemic drug absorption while replacing the need for invasive drug delivery techniques. Irbesartan and candesartan successfully lowered the IOP in rabbits when topically administered, suggesting the potential for a new class of glaucoma drugs. Our in vitro data suggest that, for many compounds, the noncorneal route could represent a more effective, and possibly the only way of drug delivery to the posterior segment following topical administration.

Cancer patients (CPs) have been identified as particularly vulnerable to SARS-CoV-2 infection, and therefore are a priority group for receiving COVID-19 vaccination. From the patients with advanced solid tumors, about 20% respond very efficiently to immunotherapy with anti-PD1/PD-L1 antibodies and achieve long lasting cancer responses. It is unclear whether an efficient cancer-specific immune response may also correlate with an efficient response upon COVID-19 vaccination. Here, we explored the antiviral immune response to the mRNA-based COVID-19 vaccine BNT162b2 in a group of 11 long-lasting cancer immunotherapy responders. We analysed the development of SARS-CoV-2-specific IgG serum antibodies, virus neutralizing capacities and T cell responses. Control groups included patients treated with adjuvant cancer immunotherapy (IMT, cohort B), CPs not treated with immunotherapy (no-IMT, cohort C) and healthy controls (cohort A). The median ELISA IgG titers significantly increased after the prime-boost COVID vaccine regimen in all cohorts (Cohort A: pre-vaccine = 900 (100-2700), 3 weeks (w) post-boost = 24300 (2700-72900); Cohort B: pre-vaccine = 300 (100-2700), 3 w post-boost = 8100 (300-72900); Cohort C: pre-vaccine = 500 (100-2700), 3 w post-boost = 24300 (300-72900)). However, at the 3 w post-prime time-point, only the healthy control group showed a statistically significant increase in antibody levels (Cohort A = 8100 (900-8100); Cohort B = 900 (300-8100); Cohort C = 900 (300-8100)) (P < 0.05). Strikingly, while all healthy controls generated high-level antibody responses after the complete prime-boost regimen (Cohort A = 15/15 (100%), not all CPs behaved alike [Cohort B= 12/14 (84'6%); Cohort C= 5/6 (83%)]. Their responses, including those of the long-lasting immunotherapy responders, were more variable (Cohort A: 3 w post-boost (median nAb titers = 95.32 (84.09-96.93), median Spike-specific IFN-γ response = 64 (24-150); Cohort B: 3 w post-boost (median nAb titers = 85.62 (8.22-97.19), median Spike-specific IFN-γ response (28 (1-372); Cohort C: 3 w post-boost (median nAb titers = 95.87 (11.8-97.3), median Spike-specific IFN-γ response = 67 (20-84)). Two long-lasting cancer responders did not respond properly to the prime-boost vaccination and did not generate S-specific IgGs, neutralizing antibodies or virus-specific T cells, although their cancer immune control persisted for years. Thus, although mRNA-based vaccines can induce both antibody and T cell responses in CPs, the immune response to COVID vaccination is independent of the capacity to develop an efficient anti-cancer immune response to anti PD-1/PD-L1 antibodies. The authors are supported by Spanish Melanoma Group Grant (III Beca GEM para Grupos Emergentes), the Spanish Ministry of Science and Innovation grant no. PID2019-106323RB-I00 AEI//10.13039/501100011033, the “Unidad de Excelencia María de Maeztu”, funded by the MCIN and the AEI (DOI: 10.13039/501100011033); Ref: CEX2018-000792-M.

International audience; Zearalenone (ZEN) is a mycotoxin classified as an endocrine disruptor. Many endocrine disruptors are also metabolic disruptors able to modulate energy balance and inflammatory processes in a process often involving a family of protein hormones known as adipokines. The aim of our study was to elucidate the role of ZEN as metabolic disruptor in pigs by investigating the changes in energy balance and adipokines levels in response to different treatment diets. To this end, weaned piglets (n = 10/group) were exposed to either basal feed or feed contaminated with 680 and 1620 µg/kg ZEN for 28 days. Serum samples collected at days 7 and 21 were subjected to biochemistry analysis, followed by determination of adipokine levels using a combined approach of protein array and ELISA. Results indicate that ZEN has an impact on lipid and glucose metabolism that was different depending on the dose and time of exposure. In agreement with these changes, ZEN altered circulating adipokines concentrations, inducing significant changes in adiponectin, resistin, and fetuin B. Our results suggest that ZEN may function as a natural metabolism-disrupting chemical.

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International audience; The molecular basis of male fertility remains unclear, especially in chickens, where decades of genetic selection increased male fertility variability as a side effect. As transcription and translation are highly limited in sperm, proteins are key molecules defining their functionality, making proteomic approaches one of the most adequate methods to investigate sperm capacity. In this context, it is interesting to combine complementary proteomic approaches to maximize the identification of proteins related to sperm-fertilizing ability. In the present study, we aimed at identifying proteins related to fertility in meat-type roosters, showing fertility variability. Fertile roosters (fertility rates higher than 70% after artificial insemination) differed from subfertile roosters (fertility rates lower than 40%) in their sperm mass motility. Fertile and subfertile sperm protein contents were compared using two complementary label-free quantitative proteomic methods: Intact Cell MALDI-TOF-Mass Spectrometry and GeLC-MS/MS. Combining the two strategies, 57 proteins were identified as differentially abundant. Most of them were described for the first time as differentially abundant according to fertility in this species. These proteins were involved in various molecular pathways including flagellum integrity and movement, mitochondrial functions, sperm maturation, and storage in female tract as well as oocyte–sperm interaction. Collectively, our data improved our understanding of chicken sperm biology by revealing new actors involved in the complexity of male fertility that depends on multiple cell functions to reach optimal rates. This explains the inability of reductionist in vitro fertility testing in predicting male fertility and suggests that the use of a combination of markers is a promising approach.

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Trabajo de fin de Máster. Máster en Tecnologías de la Información y la Comunicación Aplicadas a la Enseñanza y el Tratamiento de Lenguas

Stability of meat juice samples during two years storage at -20ºC for Hp and CRP measurements. 8. European Colloquium on Acute Phase Proteins

Affinity comparison of four new specific antibodies towards porcine serum amyloid A. 8. European Colloquium on Acute Phase Proteins

Canine CRP measurement in whole-blood and saliva: An alternative to serum quantification. 16. ECVIM-CA Congress

Use of C-reactive protein and total sialic acid to detect malignancies in canine effusions. ESVCP Congress