Synthesis and Leishmanicidal Activity of Novel Urea, Thiourea, and Selenourea Derivatives of Diselenides.

A novel series of thirty-one N-substituted urea, thiourea, and selenourea derivatives containing diphenyldiselenide entities were synthesized, fully characterized by spectroscopic and analytical methods, and screened for their in vitro leishmanicidal activities. The cytotoxic activity of these derivatives was tested against Leishmania infantum axenic amastigotes, and selectivity was assessed in human THP-1 cells. Thirteen of the synthesized compounds showed a significant antileishmanial activity, with 50% effective concentration (EC50) values lower than that for the reference drug miltefosine (EC50, 2.84 µM). In addition, the derivatives 9, 11, 42, and 47, with EC50 between 1.1 and 1.95 µM, also displayed excellent selectivity (selectivity index ranged from 12.4 to 22.7) and were tested against infected macrophages. Compound 11, a derivative with a cyclohexyl chain, exhibited the highest activity against intracellular amastigotes, with EC50 values similar to those observed for the standard drug edelfosine. Structure-activity relationship analyses revealed that N-aliphatic substitution in urea and selenourea is recommended for the leishmanicidal activity of these analogs. Preliminary studies of the mechanism of action for the hit compounds was carried out by measuring their ability to inhibit trypanothione reductase. Even though the obtained results suggest that this enzyme is not the target for most of these derivatives, their activity comparable to that of the standards and lack of toxicity in THP-1 cells highlight the potential of these compounds to be optimized for leishmaniasis treatment.

Identification of a Novel Quinoxaline-Isoselenourea Targeting the STAT3 Pathway as a Potential Melanoma Therapeutic.

The prognosis for patients with metastatic melanoma remains very poor. Constitutive signal transducer and activator of transcription 3 (STAT3) activation has been correlated to metastasis, poor patient survival, larger tumor size, and acquired resistance against vemurafenib (PLX-4032), suggesting its potential as a molecular target. We recently designed a series of isoseleno- and isothio-urea derivatives of several biologically active heterocyclic scaffolds. The cytotoxic effects of lead isoseleno- and isothio-urea derivatives (compounds 1 and 3) were studied in a panel of five melanoma cell lines, including B-RAFV600E-mutant and wild-type (WT) cells. Compound 1 (IC50 range 0.8⁻3.8 µM) showed lower IC50 values than compound 3 (IC50 range 8.1⁻38.7 µM) and the mutant B-RAF specific inhibitor PLX-4032 (IC50 ranging from 0.4 to >50 µM), especially at a short treatment time (24 h). These effects were long-lasting, since melanoma cells did not recover their proliferative potential after 14 days of treatment. In addition, we confirmed that compound 1 induced cell death by apoptosis using Live-and-Dead, Annexin V, and Caspase3/7 apoptosis assays. Furthermore, compound 1 reduced the protein levels of STAT3 and its phosphorylation, as well as decreased the expression of STAT3-regulated genes involved in metastasis and survival, such as survivin and c-myc. Compound 1 also upregulated the cell cycle inhibitor p21. Docking studies further revealed the favorable binding of compound 1 with the SH2 domain of STAT3, suggesting it acts through STAT3 inhibition. Taken together, our results suggest that compound 1 induces apoptosis by means of the inhibition of the STAT3 pathway, non-specifically targeting both B-RAF-mutant and WT melanoma cells, with much higher cytotoxicity than the current therapeutic drug PLX-4032.

A diphenyldiselenide derivative induces autophagy via JNK in HTB-54 lung cancer cells.

Symmetric aromatic diselenides are potential anticancer agents with strong cytotoxic activity. In this study, the in vitro anticancer activities of a novel series of diarylseleno derivatives from the diphenyldiselenide (DPDS) scaffold were evaluated. Most of the compounds exhibited high efficacy for inducing cytotoxicity against different human cancer cell lines. DPDS 2, the compound with the lowest mean GI50 value, induced both caspase-dependent apoptosis and arrest at the G0 /G1 phase in acute lymphoblastic leucemia CCRF-CEM cells. Consistent with this, PARP cleavage; enhanced caspase-2, -3, -8 and -9 activity; reduced CDK4 expression and increased levels of p53 were detected in these cells upon DPDS 2 treatment. Mutated p53 expressed in CCRF-CEM cells retains its transactivating activity. Therefore, increased levels of p21CIP1 and BAX proteins were also detected. On the other hand, DPDS 6, the compound with the highest selectivity index for cancer cells, resulted in G2 /M cell cycle arrest and caspase-independent cell death in p53 deficient HTB-54 lung cancer cells. Autophagy inhibitors 3-methyladenine, wortmannin and chloroquine inhibited DPDS 6-induced cell death. Consistent with autophagy, increased LC3-II and decreased SQSTM1/p62 levels were detected in HTB-54 cells in response to DPDS 6. Induction of JNK phosphorylation and a reduction in phospho-p38 MAPK were also detected. Moreover, the JNK inhibitor SP600125-protected HTB-54 cells from DPDS 6-induced cell death indicating that JNK activation is involved in DPDS 6-induced autophagy. These results highlight the anticancer effects of these derivatives and warrant future studies examining their clinical potential.

Novel Methylselenoesters as Antiproliferative Agents.

Selenium (Se) compounds are potential therapeutic agents in cancer. Importantly, the biological effects of Se compounds are exerted by their metabolites, with methylselenol (CH3SeH) being one of the key executors. In this study, we developed a new series of methylselenoesters with different scaffolds aiming to modulate the release of CH3SeH. The fifteen compounds follow Lipinski's Rule of Five and with exception of compounds 1 and 14, present better drug-likeness values than the positive control methylseleninic acid. The compounds were evaluated to determine their radical scavenging activity. Compound 11 reduced both DPPH and ABTS radicals. The cytotoxicity of the compounds was evaluated in a panel of five cancer cell lines (prostate, colon and lung carcinoma, mammary adenocarcinoma and chronic myelogenous leukemia) and two non-malignant (lung and mammary epithelial) cell lines. Ten compounds had GI50 values below 10 µM at 72 h in four cancer cell lines. Compounds 5 and 15 were chosen for further characterization of their mechanism of action in the mammary adenocarcinoma cell line due to their similarity with methylseleninic acid. Both compounds induced G2/M arrest whereas cell death was partially executed by caspases. The reduction and metabolism were also investigated, and both compounds were shown to be substrates for redox active enzyme thioredoxin reductase.

Novel Heteroaryl Selenocyanates and Diselenides as Potent Antileishmanial Agents.

A series of new selenocyanates and diselenides bearing interesting bioactive scaffolds (quinoline, quinoxaline, acridine, chromene, furane, isosazole, etc.) was synthesized, and their in vitro leishmanicidal activities against Leishmania infantum amastigotes along with their cytotoxicities in human THP-1 cells were determined. Interestingly, most tested compounds were active in the low micromolar range and led us to identify four lead compounds (1h, 2d, 2e, and 2f) with 50% effective dose (ED50) values ranging from 0.45 to 1.27 µM and selectivity indexes of >25 for all of them, much higher than those observed for the reference drugs. These active derivatives were evaluated against infected macrophages, and in order to gain preliminary knowledge about their possible mechanism of action, the inhibition of trypanothione reductase (TryR) was measured. Among these novel structures, compounds 1h (3,5-dimethyl-4-isoxazolyl selenocyanate) and 2d [3,3'-(diselenodiyldimethanediyl)bis(2-bromothiophene)] exhibited good association between TryR inhibitory activity and antileishmanial potency, pointing to 1h, for its excellent theoretical ADME (absorption, distribution, metabolism, and excretion) properties, as the most promising lead molecule for leishmancidal drug design.

Identification of selenocompounds with promising properties to reverse cancer multidrug resistance.

In previous studies, 56 novel selenoesters and one cyclic selenoanhydride with chemopreventive, antiproliferative and cytotoxic activity were described. Herein, the selenoanhydride and selected selenoesters were evaluated for their ability to reverse the cancer multidrug resistance (MDR) using the ABCB1 efflux pump inhibition assay in mouse MDR T-lymphoma cells. Results showed that the selenoanhydride (1) and the selenoesters with ketone terminal fragments (9-11) exerted (1.7-3.6)-fold stronger efflux pump inhibitory action than the reference verapamil. In addition, those four derivatives triggered apoptotic events in more than 80% of the examined MDR mouse cells.

Leishmanicidal activities of novel methylseleno-imidocarbamates.

The generation of new antileishmanial drugs has become a priority. Selenium and its derivatives stand out as having promising leishmanicidal activity. In fact, some parasites express selenoproteins and metabolize selenium. Recently, selenium derivatives have shown the potential to reduce parasitemia, clinical manifestations, and mortality in parasite-infected mice. In this paper, after selecting four candidates according to drug similarity parameters, we observed that two of them, called compounds 2b [methyl-N,N'-di(thien-2-ylcarbonyl)-imidoselenocarbamate] and 4b [methyl-N,N'-di(5-nitrothien-3-ylcarbonyl)-imidoselenocarbamate], exhibit low 50% inhibitory concentrations (IC50s) (<3 µM) and good selectivity indexes (SIs) (>5) in Leishmania major promastigotes and lack toxicity on macrophages. In addition, in analysis of their therapeutic potential against L. major in vitro infection, both compounds display a dramatic reduction of amastigote burden (∼80%) with sublethal concentrations. Furthermore, in macrophages, these selenocompounds induce nitric oxide production, which has been described to be critical for defense against intracellular pathogens. Compounds 2b and 4b were demonstrated to cause cell cycle arrest in G1. Interestingly, evaluation of expression of genes related to proliferation (PCNA), treatment resistance (ABC transporter and alpha-tubulin), and virulence (quinonoid dihydropteridine reductase [QDPR]) showed several alterations in gene expression profiling. All these results prompt us to propose both compounds as candidates to treat leishmanial infections.

In vitro radical scavenging and cytotoxic activities of novel hybrid selenocarbamates.

Novel selenocyanate and diselenide derivatives containing a carbamate moiety were synthesised and evaluated in vitro to determine their cytotoxic and radical scavenging properties. Cytotoxic activity was tested against a panel of human cell lines including CCRF-CEM (lymphoblastic leukaemia), HT-29 (colon carcinoma), HTB-54 (lung carcinoma), PC-3 (prostate carcinoma), MCF-7 (breast adenocarcinoma), 184B5 (non-malignant, mammary gland derived) and BEAS-2B (non-malignant, derived from bronchial epithelium). Most of the compounds displayed high antiproliferative activity with GI50 values below 10µM in MCF-7, CCRF-CEM and PC-3 cells. Radical scavenging properties of the new selenocompounds were confirmed testing their ability to scavenge DPPH and ABTS radicals. Based on the activity of selenium-based glutathione peroxidases (GPxs), compounds 1a, 2e and 2h were further screened for their capacity to reduce hydrogen peroxide under thiol presence. Results suggest that compound 1a mimics GPxs activity. Cytotoxic parameters, radical scavenging activity and ADME profile point to 1a as promising drug candidate.

Novel structural insights for imidoselenocarbamates with antitumoral activity related to their ability to generate methylselenol.

In the search for molecules with potential antiangiogenic activity we found that several imidoselenocarbamate derivatives, which have pro-apoptotic and antiproliferative activities, under hypoxic conditions release methylselenol, a volatile and highly reactive gas that was considered to be responsible for the observed biological activity. The kinetic for the liberation of methylselenol is highly dependent on the nature of the overall structure and correlate with their proven pro-apoptotic activity in lung cancer cell line H157. The preliminary structure-activity relationships allow us to select as the basic structural element a scaffold constructed with an imidoselenocarbamate fragment decorated with a methyl residue on the Se central atom and two heteroaromatic lateral rings. These imidoselenocarbamate derivatives may be of interest both for their antitumoral activities and because they have a structure that can be considered as a template for the design of new derivatives with apoptotic activity. This activity is related to the controlled delivery of methylselenol and makes this an interesting approach to develop new antitumoral agents.

Selenium compounds, apoptosis and other types of cell death: an overview for cancer therapy.

Selenium (Se) is an essential trace element involved in different physiological functions of the human body and plays a role in cancer prevention and treatment. Induction of apoptosis is considered an important cellular event that can account for the cancer preventive effects of Se. The mechanisms of Se-induced apoptosis are associated with the chemical forms of Se and their metabolism as well as the type of cancer studied. So, some selenocompounds, such as SeO(2) involve the activation of caspase-3 while sodium selenite induces apoptosis in the absence of the activation of caspases. Modulation of mitochondrial functions has been reported to play a key role in the regulation of apoptosis and also to be one of the targets of Se compounds. Other mechanisms for apoptosis induction are the modulation of glutathione and reactive oxygen species levels, which may function as intracellular messengers to regulate signaling pathways, or the regulation of kinase, among others. Emerging evidence indicates the overlaps between the apoptosis and other types of cell death such as autophagy. In this review we report different processes of cell death induced by Se compounds in cancer treatment and prevention.

Antileishmanial activity of imidothiocarbamates and imidoselenocarbamates.

In the present study, a family of 15 imidothio- and imidoselenocarbamates (1-15) analogs have been synthesized and screened for their in vitro antileishmanial potential against Leishmania infantum promastigotes. The six most active ones (2, 4, 7, 13, 14, and 15) were also tested in an axenic amastigote model. In order to establish their selectivity indexes (SI) the cytotoxic effect of each compound was also assayed against Jurkat and THP-1 cell lines. Compounds 2 and 4, both with a pyridine moiety, showed a moderate antileishmanial activity with an IC(50) value of 4.68 ± 0.46 and 3.03 ± 0.24 µM, respectively, in the amastigote model. The activity was compared with that of standard drugs, edelfosine (IC50 = 0.82 ± 0.13 µM) and miltefosine (IC50 = 2.84 ± 0.10 µM). Related to selectivity, the SI of both compounds are similar to those of the standard drugs when compared against the THP-1 cell line. Moreover, compound 4 was able to reduce the number of amastigote-infected THP-1 cells to 40% of that observed in untreated controls after a 96-h period of treatment. These derivatives thus represent two new leads for further studies aimed at establishing their mechanism of action.

Novel library of selenocompounds as kinase modulators.

Although the causes of cancer lie in mutations or epigenic changes at the genetic level, their molecular manifestation is the dysfunction of biochemical pathways at the protein level. The 518 protein kinases encoded by the human genome play a central role in various diseases, a fact that has encouraged extensive investigations on their biological function and three dimensional structures. Selenium (Se) is an important nutritional trace element involved in different physiological functions with antioxidative, antitumoral and chemopreventive properties. The mechanisms of action for selenocompounds as anticancer agents are not fully understood, but kinase modulation seems to be a possible pathway. Various organosulfur compounds have shown antitumoral and kinase inhibition effects but, in many cases, the replacement of sulfur by selenium improves the antitumoral effect of compounds. Although Se atom possesses a larger atomic volume and nucleophilic character than sulfur, Se can also formed interactions with aminoacids of the catalytic centers of proteins. So, we propose a novel chemical library that includes organoselenium compounds as kinase modulators. In this study thirteen selenocompounds have been evaluated at a concentration of 3 or 10 µM in a 24 kinase panel using a Caliper LabChip 3000 Drug Discover Platform. Several receptor (EGFR, IGFR1, FGFR1…) and non-receptor (Abl) kinases have been selected, as well as serine/threonine/lipid kinases (AurA, Akt, CDKs, MAPKs…) implicated in main cancer pathways: cell cycle regulation, signal transduction, angiogenesis regulation among them. The obtained results showed that two compounds presented inhibition values higher than 50% in at least four kinases and seven derivatives selectively inhibited one or two kinases. Furthermore, three compounds selectively activated IGF-1R kinase with values ranging from -98% to -211%. In conclusion, we propose that the replacement of sulfur by selenium seems to be a potential and useful strategy in the search of novel chemical compound libraries against cancer as kinase modulators.

Synthesis and in vitro anticancer activities of some selenadiazole derivatives.

A novel series of fourteen substituted selenadiazoles has been synthesized and the compounds tested for their in vitro antiproliferative and cytotoxic activities. The tests were carried out against leukemia (CCRF-CEM), colon (HT-29), lung (HTB-54), and breast (MCF-7) cancer cells. In order to assess the selectivity of the compounds under investigation the assays were also carried out on two non-tumoral lines - one mammary (184B5) and one bronchial epithelium (BEAS-2B) cell line. Assay-based antiproliferative activity studies revealed that seven derivatives (2a, 2c, 2e, 2f, 2g, 3a, and 3b) exhibited good activity against MCF-7 cells: for instance, 2c and 2f inhibited cell growth with nanomolar GI50 values. Compound 2f had a better antitumoral profile than vinorelbine and paclitaxel, two drugs that are used as first-line treatments in advanced, recurrent, and/or metastatic cancer. In the other cell lines the compounds showed moderate activity or were inactive - with the exception of 2a, which was also found to have antiproliferative activity. Modulation of the cell cycle and apoptotic effects of active compounds were further evaluated in MCF-7 cells. Of these, 6-bromo[1,2,5]selenadiazolo[3,4-b]pyridine (2a) was the most active, with an apoptogenic effect 3.9 times higher than that of camptothecin, which was used as a positive control. Compound 2a also provoked cell cycle arrest with a significant decrease in the G0/G1 phase cell population and an increase in S and G2/M cells, thus suggesting mitotic arrest prior to metaphase.

Antioxidant-prooxidant properties of a new organoselenium compound library.

The present study describes the biological evaluation of a library of 59 organo-selenium compounds as superoxide (O2⁻) generators and cytotoxic agents in human prostate cancer cells (PC-3) and in breast adenocarcinoma (MCF-7). In order to corroborate that the biological activity for selenium compounds depends on the chemical form, a broad structural variety is presented. These structures include selenocyanates, diselenides, selenoalkyl functional moieties and eight newly synthesized symmetrically substituted dithioselenites and selenylureas. Eleven of the derivatives tested showed high levels of superoxide generation in vitro via oxidation of reduced glutathione (GSH) and nine of them were more catalytic than the reference compound, diselenodipropionic acid. Eighteen of the library compounds inhibited cell growth more than or similar to reference chemotherapeutic drugs in PC-3 and eleven were more potent cytotoxic agents than etoposide in the MCF-7 cell line. Considering both parameters (superoxide generation and cell cytotoxicity) compounds B1, C6 and C9 displayed the best therapeutic profiles. Considering that many diselenide compounds can generate superoxide (O2⁻) in vitro via oxidation of GSH and other thiols, the analogue B1, that contains a diselenide moiety, was selected for a preliminary mechanistic investigation, which revealed that B1 has apoptogenic effects similar to camptothecin mediated by reactive oxygen species (ROS) in lymphocytic leukemia cells (CCRF-CEM) and affected the MCF-7 cell-cycle in G2/M and S-phases.

Synthesis and pharmacological screening of several aroyl and heteroaroyl selenylacetic acid derivatives as cytotoxic and antiproliferative agents.

The synthesis and cytotoxic activity of a series of twenty six aroyl and heteroaroyl selenylacetic acid derivatives of general formula Ar-CO-Se-CH(2)-COOH or Heterar-CO-Se-CH(2)-COOH are reported. The synthesis was carried out by reaction of acyl chlorides with sodium hydrogen selenide, prepared in situ, and this led to the formation of sodium aroylselenides that subsequently reacted with alpha-bromoacetic acid to produce the corresponding selenylacetic acid derivatives. All of the compounds were tested against a prostate cancer cell line (PC-3) and some of the more active compounds were assessed against a panel of four human cancer cell lines (CCRF-CEM, HTB-54, HT-29, MCF-7) and one mammary gland-derived non-malignant cell line (184B5). Some of the compounds exhibited remarkable cytotoxic and antiproliferative activities against MCF-7 and PC-3 that were higher than those of the reference compounds doxorubicin and etoposide, respectively. For example, in MCF-7 when Ar = phenyl, 3,5-dimethoxyphenyl or benzyl the TGI values were 3.69, 4.18 and 6.19 microM. On the other hand, in PC-3 these compounds showed values of 6.8, 4.0 and 2.9 microM. Furthermore, benzoylselenylacetic acid did not provoke apoptosis nor did it perturb the cell cycle in MCF-7.

Selenium compounds and apoptotic modulation: a new perspective in cancer therapy.

Recent epidemiological studies have demonstrated that selenium may be an effective chemopreventive and anticancer agent with a broad spectrum against several human cancer cells (prostate, colon, bladder, lung, liver, ovarian, leukemia). A wide range of potential mechanisms have been proposed for the antitumorigenic effects of selenium and these include antiandrogen activity, growth inhibitory effects by regulation of p53 and antioxidant function, and through DNA damage. However, apoptosis is one of the most plausible mechanisms for the anticancer activity. The regulating mechanisms of apoptosis are extremely complex and for selenium compounds they mainly involve a mitochondrial pathway, protein kinases, tumor necrosis factor, activation of caspases and reactive oxygen species. The aim of this review is to summarize the current knowledge about more than twenty eight selenium-containing molecules and to discuss the implications for apoptosis and the impact in cancer therapy.

Novel potent organoselenium compounds as cytotoxic agents in prostate cancer cells.

A series of 17 symmetrical substituted imidothiocarbamate and imidoselenocarbamate derivatives has been synthesized by reacting appropriately substituted acyl chlorides with alkyl imidothiocarbamates and alkyl imidoselenocarbamates. The antitumoral activities of the compounds were evaluated in vitro by examining their cytotoxic effects against human prostate cancer cells (PC-3). Five compounds showed interesting activity levels and 3p (IC(50)=1.85 microM) was 7.3 times more active than the standard etoposide used in the treatment of prostate cancer and emerges as the most interesting compound.

Intraocular lens opacification after nonpenetrating glaucoma surgery with mitomycin-C.

A 58-year-old woman had successful phacoemulsification with intraocular lens (IOL) implantation in January 2001. Two years later, nonpenetrating glaucoma surgery with mitomycin-C (MMC) 0.02% was performed for uncontrolled glaucoma. Two months later, opacification of the anterior IOL surface was observed. The IOL was removed and a hydrophobic acrylic AcrySof IOL (Alcon) implanted. The opacified IOL was studied by flame atomic absorption spectrometry, which showed the presence of calcium carbonate. A new IOL of the same model was placed in an aqueous solution with calcium carbonate and basic pH, and the same opacification developed. We hypothesize that the change in aqueous humor pH after glaucoma surgery and the characteristics of the IOL precipitated deposition of calcium.

Topological and quantum molecular descriptors as effective tools for analyzing cytotoxic activity achieved by a series of (diselanediyldibenzene-4,1-diylnide)biscarbamate derivatives.

A molecular modeling study has been carried out on a previously reported series of (diselanediyldibenzene-4,1-diylnide)biscarbamate derivatives that show cytotoxic and antiproliferative in vitro activity against MCF-7 human cell line; radical scavenging properties were also confirmed when these compounds were tested for their ability to scavenge DPPH and ABTS radicals. The data obtained allowed us to classify the compounds into two different groups: (a) aliphatic carbamates for which the activity could be related with a first nucleophilic attack (mediated by H2O, for example) on the selenium atoms of the central scaffold, followed by the release of the alkyl N-(4-selanylphenyl) and N-(4-selenenophenyl)carbamate moieties. Then, a second nucleophilic attack on the carbamate moiety, to yield 4-aminobenzeneselenol and 4-selenenoaniline respectively, which can ultimately be responsible for the activity of the compounds; (b) aromatic carbamates, for which we propose a preferred nucleophilic attack on the carbamate moiety, yielding 4-[(4-aminophenyl)diselanyl]aniline, the common structural fragment for this series, for which we have previously demonstrated its cytotoxic profile. Then, selenium atoms of the central fragment may later undergo a new nucleophilic attack, to yield 4-selenenoaniline and 4-aminobenzeneselenol. The phenolic moieties released in this process may also have a synergistic cytotoxic and redox activity. The data that support this connection include the conformational behavior and the molecular topography of the derivatives which can influence the accessibility of the hydrolysis points, and some quantum descriptors (bond order, atomic charges, total valences, ionization potential, electron affinity, HOMO 0 and LUMO 0 location, etc.) that have been related to the biological activity of the compounds.

New symmetrical quinazoline derivatives selectively induce apoptosis in human cancer cells.

In the search of new symmetrical derivatives with anticancer activity, we have looked for novel compounds able to induce a selective proapoptotic mechanism in cancer cells. The potential antitumoral activity of several quinazoline derivatives was evaluated in vitro examining their cytotoxic effects against human breast, colon and bladder cancer cell lines. The IC(50) value of the compounds that showed cytotoxic activity was calculated. These compounds were tested for their ability to induce caspase-3 activation and nuclear chromatin degradation. Non-tumoral human cell lines were used to test the selectivity of the cytotoxic compounds against cancer cells. Several compounds showed no cytotoxicity in these cell lines. Finally, JRF12 (2,4-dibenzylaminoquinazoline) was chosen as the best candidate and its mechanism of action was studied in more detail. A time dependent evaluation of apoptosis was performed in the three cancer cell lines, followed by an evaluation of the cell cycle regulation involvement that showed a decrease of cells in G(1) phase and increase of cells in G(2) phase before cell death. 2,4-dibenzylaminoquinazoline treatment produces few changes in the expression of genes as evaluated by using oligonucleotide microarrays and Q-RT-PCR assays. In conclusion, 2,4-dibenzylaminoquinazoline is a promising anticancer drug showing cytostatic and apoptotic effects mainly in a transcription independent manner.