antioxidants Article Novel N,N0-Disubstituted Selenoureas as Potential Antioxidant and Cytotoxic Agents Gorka Calvo-Martín 1,2,†, Daniel Plano 1,2,† , Ignacio Encío 2,3 and Carmen Sanmartín 1,2,* 1 Departamento de Tecnología y Química Farmacéuticas, Universidad de Navarra, Irunlarrea 1, E-31008 Pamplona, Spain; [email protected] (G.C.-M.); [email protected] (D.P.) 2 Instituto de Investigación Sanitaria de Navarra (IdiSNA), Irunlarrea, 3, E-31008 Pamplona, Spain; [email protected] 3 Departamento de Ciencias de la Salud, Universidad Pública de Navarra, Avda. Barañain s/n, E-31008 Pamplona, Spain * Correspondence: [email protected]; Tel.: +34-948425600 (ext. 806388) † These authors have contributed equally to the manuscript. Abstract: A series of 30 novel N,N disubstituted selenoureas were synthesized, characterized, and their antioxidant ability was tested using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,20-azino- bis(3-ethylbenzthiazoline-6-sulfonic acid (ABTS) assays. Additionally, their cytotoxic activity was tested in vitro in a panel of three different cancer (breast, lung and colon) and two normal cell lines. Each selenourea entity contains a para-substituted phenyl ring with different electron-withdrawing and electron-donating groups, and different aliphatic and aromatic nuclei. All of the synthesized selenoureas present antioxidant capacity at high concentrations in the DPPH assay, and three of them (2b, 2c and 2d) showed greater radical scavenging capacity than ascorbic acid at lower concentrations. Citation: Calvo-Martín, G.; Plano, D.; These results were confirmed by the ABTS assay, where these novel selenoureas present even higher Encío, I.; Sanmartín, C. Novel antioxidant capacity than the reference compound Trolox. On the other hand, 10 selenoureas present N,N0-Disubstituted Selenoureas as IC50 values below 10 µM in at least one cancer cell line, resulting in the adamantyl nucleus (6a– Potential Antioxidant and Cytotoxic 6e), the most interesting in terms of activity and selectivity. Outstanding results were found for Agents. Antioxidants 2021, 10, 777. µ https://doi.org/10.3390/ selenourea 6c, tested in the NCI60 cell line panel and showing an average GI50 of 1.49 M for the antiox10050777 60 cell lines, and LC50 values ranging from 9.33 µM to 4.27 µM against 10 of these cancer cell lines. To gain insight into its anticancer activity mechanism, we investigated the cell cycle progression Academic Editors: Jose of the promising compound 6c, as well as the type of programmed-cell death in a colon cancer Manuel Martinez-Martos and cell line it provokes (HT-29). Compound 6c provoked S phase cell cycle arrest and the induction Holger Steinbrenner of cell death was independent of caspase activation, suggesting autophagy, though this assertion requires additional studies. Overall, we envision that this compound can be further developed for Received: 12 April 2021 the potential treatment of colon cancer. Accepted: 11 May 2021 Published: 14 May 2021 Keywords: selenoureas; antioxidant; cytotoxicity; radical scavenging; selenium Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- 1. Introduction iations. Cancer is a group of complex and multifactorial diseases and is considered the third leading cause of death globally. The World Health Organization (WHO) estimates that one-sixth of deaths can be attributed to cancer [1]. Lung (12.3%), breast (12.3%), colorectal (10.6%), and prostate (7.5%) are the types of cancer with the highest incidence worldwide, Copyright: © 2021 by the authors. according to the statistics [2,3]. Several chemotherapy regimens have demonstrated efficacy, Licensee MDPI, Basel, Switzerland. but these cancers’ heterogeneity and evolvement of resistance have generated a growing This article is an open access article distributed under the terms and need for new anticancer agents that would sort out these problems. conditions of the Creative Commons Selenium (Se) is an essential micronutrient; trace amounts are necessary for the func- Attribution (CC BY) license (https:// tion of the human body and it is mainly obtained through diet and/or nutritional sup- creativecommons.org/licenses/by/ plements [4]. Se is a component of the selenoproteins (mostly in the form of amino acid 4.0/). selenocysteine) that participate in a wide range of cellular physiological processes [5]. In the Antioxidants 2021, 10, 777. https://doi.org/10.3390/antiox10050777 https://www.mdpi.com/journal/antioxidants Antioxidants 2021, 10, x FOR PEER REVIEW 2 of 15 Antioxidants 2021, 10, 777 2 of 14 [5].human In the organism, human Se organism, acts as an antioxidantSe acts as (e.g.,an antioxidant glutathione (e.g., peroxidase glutathione (GPX)), peroxidase protecting (GPX)),against theprotecting harmful against effects the offree harmful radicals. effects Depending of free radicals. on the SeDepending dose and chemicalon the Se form,dose anddiverse chemical effects form, of this diverse element effects have beenof this observed element on have cellular been functions observed such on ascellular immunitary func- tionsresponse such and as immunitary energy metabolism response [6 ].and A marginalenergy metabolism problem of [6]. selenium A marginal use is problem its narrow of seleniumrange between use is theits toxicnarrow dose range and between the dose the necessary toxic dose for theand proper the dose functioning necessary of for living the properorganisms functioning and, for theseof living reasons, organisms each selenium and, for these derivative reasons, must each be studiedselenium individually. derivative mustFor example, be studied sodium individually. selenite For exhibited example, excellent sodium activity selenite in exhibited the prevention excellent [7] activity and treat- in thement prevention of different [7] types and oftreatment cancers, whileof different other derivatives types of cancers, with other while chemical other forms,derivatives such withas sodium other selenate,chemical didforms, not such exert as these sodium properties selenate, [8]. did On not the exert other these hand, properties different studies[8]. 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Se Forinto these small reasons, molecules, the both incorporation organic and of inorganic, Se into small has attracted molecules, special both attention organic amongand inor- the ganic,researchers, has attracted and it is special now considered attention aamong promising the researchers, candidate in and the fieldit is now of drug considered discovery a promisingfor cancer therapycandidate [11 in,12 the]. field of drug discovery for cancer therapy [11,12]. The anticanceranticancer effecteffect of of selenium selenium compounds compounds may may be assertedbe asserted through through various various path- pathwaysways in the in cell the [ 13cell]. They[13]. They have have been reportedbeen reported to act to protectively act protectively against against oxidative oxidative injury injuryby stimulation by stimulation of DNA of repair, DNA by repair, the regulation by the regulation of inflammatory of inflammatory and immune and responses, immune responses,by the induction by the ofinduction cell cycle of arrest, cell cycle and arre byst, provoking and by provoking cell death bycell different death by processes different processessuch as apoptosis, such as apoptosis, necrosis, autophagy,necrosis, autophagy, ferroptosis, ferroptosis, necroptosis, necroptosis, entosis, anoikis, entosis, NETo- anoi- kis,sis, orNETosis, mitotic or catastrophe mitotic catastrophe inhibition inhibition of local invasion of local andinvasion migration, and migration, as well as as blocking well as blockingangiogenesis, angiogenesis, modulating modulating the cell proliferation, the cell proliferation, etc. [10,14 etc.]. [10,14]. Several organoselenium agents agents have been developed for the inhibition of different typestypes of cancer cell growth. Among Among them, them, selenoureas selenoureas represent represent a a group of simple but well-studied moleculesmolecules withwith anticancer anticancer and/or and/or antioxidant antioxidant properties properties [15 [15–20].–20]. In addition,In addi- tion,our research our research team hasteam reported has reported the preparation the preparation and cytotoxic and cytotoxic and antioxidant and antioxidant activities ac- for tivitiesdifferent for selenourea different seleno derivativesurea derivatives [21–24]. [21–24]. With thethe aboveabove factsfacts in in mind, mind, and and using using the th fragment-basede fragment-based approach, approach, we we designed designed the thecompounds compounds presented presented in this in paper. this paper. Herein, He werein, pursue we thepursue development the development of new molecules, of new molecules,with the selenourea with the functioningselenourea functioning as cytotoxic agentsas cytotoxic decorated agents in decorated the nitrogen in atomsthe nitrogen with a atomssmall setwith of fragmenta small set hits. of fragment Therefore,