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Chemotherapy is a powerful means of cancer treatment but its efficacy is compromised by the emergence of multidrug resistance (MDR), mainly linked to the efflux transporter ABCB1/P-glycoprotein (P-gp). Based on the chemical structure of betulin, identified in our previous work as an effective modulator of the P-gp function, a series of analogs were designed, synthesized and evaluated as a source of novel inhibitors. Compounds 6g and 6i inhibited rhodamine 123 efflux in the P-gp overexpressed leukemia cells, K562/Dox, at concentrations of 0.19 µM and 0.39 µM, respectively, and increased the intracellular accumulation of doxorubicin at the submicromolar concentration of 0.098 µM. Compounds 6g and 6i were able to restore the sensitivity of K562/Dox to Dox at 0.024 µM and 0.19 µM, respectively. Structure-activity relationship analysis and molecular modeling revealed important information about the structural features conferring activity. All the active compounds fitted in a specific region involving mainly transmembrane helices (TMH) 4-6 from one homologous half and TMH 7 and 12 from the other, also showing close contacts with TMH 6 and 12. Compounds that bound preferentially to another region were inactive, regardless of their free energy of binding. It should be noted that compounds 6g and 6i were devoid of toxic effects against peripheral blood mononuclear normal cells and erythrocytes. The data obtained indicates that both compounds might be proposed as scaffolds for obtaining promising P-gp inhibitors for overcoming MDR., (© 2024. The Author(s).)

The reactions of 2-naphthylamine and methyl 6-amino-2-naphthoate with formalin and paraformaldehyde were studied experimentally, spectrally, and by quantum chemical calculations. It was found that neither the corresponding aminals nor imines were formed under the described conditions but could be prepared and spectrally characterized at least in situ under modified conditions. Several of the previously undescribed intermediates and by-products were isolated or at least spectrally characterized. First principle density functional theory (DFT) calculations were performed to shed light on the key aspects of the thermochemistry of decomposition and further condensation of the corresponding aminals and imines. The calculations also revealed that the electrophilicity of methanal was significantly greater than that of ordinary oxo-compounds, except for perfluorinated ones. In summary, methanal was not behaving as the simplest aldehyde but as a very electron-deficient oxo-compound.

Background: Tumor angiogenesis is considered as a crucial pathologic feature of cancer with a key role in multidrug resistance (MDR). Adverse effects of the currently available drugs and the development of resistance to these remain as the hardest obstacles to defeat. Objetive: This work explores flora from Argentina as a source of new chemical entities with antiangiogenic activity. Methods: Tube formation assay using bovine aortic endothelial cells (BAECs) was the experiment of choice to assess antiangiogenic activity. The effect of the pure compound in cell invasiveness was investigated through the trans-well migration assay. The inhibitory effect of the pure compound on VEGFR-2 and PKC isozymes α and β2 activation was studied by molecular and massive dynamic simulations. Cytotoxicity on peripheral blood mononuclear cells and erythrocyte cells was evaluated by means of MTT and hemolysis assay, respectively. In silico prediction of pharmacological properties (ADME) and evaluation of drug-likeness features were performed using the SwissADME online tool. Results: Among the plants screened, T. minuta , showed an outstanding effect with an IC 50 of 33.6 ± 3.4 μg/ml. Bio-guided isolation yielded the terthiophene α-terthienylmethanol as its active metabolite. This compound inhibited VEGF-induced tube formation with an IC 50 of 2.7 ± 0.4 μM and significantly impaired the invasiveness of bovine aortic endothelial cells (BAECs) as well as of the highly aggressive breast cancer cells, MDA-MB-231, when tested at 10 μM. Direct VEGFR-2 and PKC inhibition were both explored by means of massive molecular dynamics simulations. The results obtained validated the inhibitory effect on protein kinase C (PKC) isozymes α and β2 as the main mechanism underlying its antiangiogenic activity. α-terthienylmethanol showed no evidence of toxicity against peripheral blood mononuclear and erythrocyte cells. Conclusion: These findings support this thiophene as a promising antiangiogenic phytochemical to fight against several types of cancer mainly those with MDR phenotype., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Llorens de los Ríos, Lanza, Barbieri, González, Chabán, Soria, Vera, Carpinella and Joray.)

P-gp-associated multidrug resistance is a major impediment to the success of chemotherapy. With the aim of finding non-toxic and effective P-gp inhibitors, we investigated a panel of quinolin-2-one-pyrimidine hybrids. Among the active compounds, two of them significantly increased intracellular doxorubicin and rhodamine 123 accumulation by inhibiting the efflux mediated by P-gp and restored doxorubicin toxicity at nanomolar range. Structure-activity relationships showed that the number of methoxy groups, an optimal length of the molecule in its extended conformation, and at least one flexible methylene group bridging the quinolinone to the moiety bearing the pyrimidine favored the inhibitory potency of P-gp. The best compounds showed a similar binding pattern and interactions to those of doxorubicin and tariquidar, as revealed by MD and hybrid QM/MM simulations performed with the recent experimental structure of P-gp co-crystallized with paclitaxel. Analysis of the molecular interactions stabilizing the different molecular complexes determined by MD and QTAIM showed that binding to key residues from TMH 4-7 and 12 is required for inhibition., (© 2021. The Author(s).)

Overexpression of P-glycoprotein (P-gp), which is linked to multidrug resistance (MDR), is one of the underlying obstacles to the success of chemotherapy as it reduces the efficacy of anticancer drugs and the side effects of these increase as a result of any increased dose to achieve the therapeutic effect. To identify agents with P-gp inhibitory properties, ethanol extracts from 80 plants were screened for their ability to increase intracellular doxorubicin-associated fluorescence, and the extract of Ligaria cuneifolia was found to be the most effective. Its bioassay-guided isolation yielded the pentacyclic triterpene betulin as active agent. This efficiently inhibited P-gp mediated efflux, as demonstrated by the enhancement of the intracellular accumulation of doxorubicin and rhodamine 123 from 1.56 μM in the P-gp overexpressing MDR leukemia cell, Lucena 1. Betulin was also able to render Lucena 1 sensitive to Dox from 0.39 μM. The docking studies revealed that betulin tightly binds to a key region of the TMDs, with a binding mode overlapping one main site of doxorubicin and, more interestingly, emulating the same contacts as tariquidar, as revealed by the per-residue energetic analysis from molecular dynamics simulations. MTT assay using peripheral blood mononuclear cells and hemolysis assay showed that betulin is devoid of toxicity. These findings provide important evidence that betulin may be a safe and promising entity to be further investigated to develop agents able to overcome P-gp-mediated MDR, resulting in a more effective and less toxic chemotherapy., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Hydroxyphenyl-azoles are among the most popular ESIPT (Excited State Intramolecular Proton Transfer) scaffolds and as such, they have been thoroughly studied. Nevertheless, some aspects regarding the interplay between the emissive properties of these fluorophores and the size of their π-conjugated framework remain controversial. Previous studies have demonstrated that benzannulation of 2'-hydroxyphenyl-oxazole at the phenol group of the molecule can lead to either red- or blue-shifted fluorescence emission, depending on the site where it occurs. In this report, benzannulation at the heterocyclic unit (the oxazole site) is analysed in order to get the whole picture. The extension of π-conjugation does not significantly affect the ESIPT emission wavelength, but it leads instead to higher energy barriers for proton transfer in the first excited singlet state, as a consequence of dramatic changes in the charge transfer character of excitation caused by successive benzannulation. Theoretical calculations revealed an interesting connection between intramolecular charge transfer and excited-state aromaticity in the S 1 state. The theoretical approach presented herein allows the behaviour of hydroxyphenyl-oxazoles in the excited state to be rationalized and, more generally, a deeper understanding of the factors governing the ESIPT process to be obtained, a crucial point in the design of new and efficient fluorophores., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

ESIPT (Excited State Intramolecular Proton Transfer) to C atom in 2-phenylphenol is known to be an intrinsically inefficient process. However, to the best of our knowledge, a structure-ESIPT efficiency relationship has not been elucidated yet. Here, we show that there exists a competitive interplay between photoacidity and ESIPT efficiency for the 2-phenylphenol system. The attachment of electron withdrawing groups to the phenol moiety promotes adiabatic deprotonation in the excited state and diminishes the charge transfer character of the excitations, and both these factors contribute in decreasing the ESIPT reaction yield. On the other hand, unfavorable conformational distribution in the ground state also appears as another important aspect responsible for the low ESIPT extent of 2-phenylphenol. A new derivative bearing electron donating, bulky substituents at ortho and para positions of the phenol ring shows an outstanding ESIPT performance, which demonstrates that the efficiency of the process can be significantly enhanced by modifying the substitution pattern. We anticipate that our results will help to guide the molecular designing of new compounds with high ESIPT efficiency.

Currently, only two drugs (i.e. benznidazole (BZN) and nifurtimox (NFX)) have been approved for the treatment of Trypanosoma cruzi (Tc) infection, the etiological agent causing Chagas disease. Since both drugs exhibit severe side effects, patients frequently abandon therapy, resulting in an inefficient pharmacotherapeutic treatment. In this context, there is an urgent need to develop new, safer and optimised anti-Tc agents. In this report, we present the synthesis and biological activity of 11 novel and 3 already reported N-arylsulfonyl-benzimidazole derivatives (NBSBZD,1-14) currently in development as potential anti-Tc compounds. These compounds were designed as part of a library of synthetic arylsulfonyl heterocycle derivatives constructed from privileged structures exhibiting drug-like properties. Based on bioactivity assays against Tc, (in both the extracellular and intracellular forms), we observed that 10 compounds exhibited bioactivity against the epimastigote form, while six of them exhibited activity against the amastigote counterpart. Also, the compounds showed less cytotoxicity compared to the reference drug BZN as measured in Vero cell culture. In order to elucidate the potential mechanism of action, metabolite excretion profiles studies were performed, and complemented with molecular modeling studies performed over known Tc druggable targets. Consistency was observed between experimental and theoretical findings, with metabolic profiles showing that compounds 1, 2, 9, 12 and 14 interfered with the normal glycolysis cycle of Tc, while molecular modeling studies were able to establish a solid structure-activity relationship towards the inhibition of 6-phospho-1-fructokinase, a key enzyme involved in the parasite glycolytic cascade. Overall, the present study constitutes a multidisciplinary contribution to the development of new anti-Chagas compounds., (Copyright © 2019 Elsevier Masson SAS. All rights reserved.)

To contribute enzymatic browning inhibitors to the food industry and also extend knowledge about the phytochemical profile of the anti-tyrosinase plant Lepechinia meyenii, its ethanol extract was subjected to bioguided fractionation. Three hydroxycinnamic acids, p-coumaric acid (1), caffeic acid (2) and rosmarinic acid (3), were isolated as mainly responsible for its activity. Compounds 1, 2 and 3 showed themselves highly effective for inhibiting tyrosinase with IC 50 values of 0.30, 1.50 and 4.14 μM, respectively, for monophenolase activity and 0.62, 2.30 and 8.59 μM, respectively for diphenolase activity. This is the first report describing the isolation of the compounds causing the tyrosinase inhibitory activity of L. meyenii extract. The inhibitory kinetics of 1-3 using both L-tyrosine and L-DOPA as substrates was investigated and the results obtained were discussed at molecular level by docking analysis. The resulting compounds 1-3 and a phenolic-enriched fraction of the extract, 2.9-fold more active than the starting material, may be suitable as non-toxic and inexpensive alternatives for the control of deleterious enzymatic darkening., (Copyright © 2019. Published by Elsevier Ltd.)

To find novel P-gp-inhibitors, a library of pregnane X receptor (PXR) ligands and the ZINC DrugsNow library were superimposed on the P-gp inhibitor (+)-pinoresinol ( 1 ) used as a query for a three-dimensional similarity search. After determining the TanimotoCombo index of similarity with 1 , eight compounds from the PXR library and two ZINC compounds were selected for biological evaluation. The P-gp inhibition study showed that compounds 7 , 8 , and 9 successfully increased intracellular doxorubicin (DOX) accumulation in the P-gp overexpressed Lucena 1 cells from 25, 12.5, and 6.25 μM, respectively. Among a series of analogues of 9 , compounds 26 - 30 were shown to be active, with 26 and 27 causing a significant increase in DOX accumulation from 1.56 μM and rendering Lucena 1 sensitive to DOX from 1.56 and 0.78 μM, respectively. Molecular modeling studies showed that both compounds bind to the P-gp at transmembrane helices (TMH) 4, 5, and 6, with 27 also showing contacts with TMH 3., Competing Interests: The authors declare no competing financial interest.

Background: Eqolisins are rare acid proteases found in archaea, bacteria and fungi. Certain fungi secrete acids as part of their lifestyle and interestingly these also have many eqolisin paralogs, up to nine paralogs have been recorded. This suggests a process of functional redundancy and diversification has occurred, which was the subject of the research we performed and describe here., Results: We identified eqolisin homologs by means of iterative HMMER analysis of the NR database. The identified sequences were scrutinized for which new hallmarks were identified by molecular dynamics simulations of mutants in highly conserved positions, using the structure of an eqolisin that was crystallized in the presence of a transition state inhibitor. Four conserved glycines were shown to be important for functionality. A substitution of W67F is shown to be accompanied by the L105W substitution. Molecular dynamics shows that the W67 binds to the substrate via a π-π stacking and a salt bridge, the latter being stronger in a virtual W67F/L105W double mutant of the resolved structure of Scytalido-carboxyl peptidase-B (PDB ID: 2IFW). Additional problematic mutations are discussed. Upon sequence scrutiny we obtained a set of 233 sequences that was used to reconstruct a Bayesian phylogenetic tree. We identified 14 putative specificity determining positions (SDPs) of which four are explained by mere structural explanations and nine seem to correspond to functional diversification related with substrate binding and specificity. A first sub-network of SDPs is related to substrate specificity whereas the second sub-network seems to affect the dynamics of three loops that are involved in substrate binding., Conclusion: The eqolisins form a small superfamily of acid proteases with nevertheless many paralogs in acidic fungi. Functional redundancy has resulted in diversification related to substrate specificity and substrate binding.

The serine-threonine checkpoint kinase 1 (Chk1) plays a critical role in the cell cycle arrest in response to DNA damage. In the last decade, Chk1 inhibitors have emerged as a novel therapeutic strategy to potentiate the anti-tumour efficacy of cytotoxic chemotherapeutic agents. In the search for new Chk1 inhibitors, a congeneric series of 2-aryl-2 H-pyrazolo[4,3-c]quinolin-3-one (PQ) was evaluated by in-vitro and in-silico approaches for the first time. A total of 30 PQ structures were synthesised in good to excellent yields using conventional or microwave heating, highlighting that 14 of them are new chemical entities. Noteworthy, in this preliminary study two compounds 4e 2 and 4h 2 have shown a modest but significant reduction in the basal activity of the Chk1 kinase. Starting from these preliminary results, we have designed the second generation of analogous in this class and further studies are in progress in our laboratories.

P-glycoprotein (P-gp) is a membrane protein associated with multidrug resistance (MDR) due to its key role in mediating the traffic of chemotherapeutic drugs outside cancer cells, leading to a cellular response that hinders efforts toward successful therapy. With the aim of finding agents that circumvent the MDR phenotype mediated by P-gp, 15 compounds isolated from native and naturalized plants of Argentina were screened. Among these, the non-cytotoxic lignan (±) pinoresinol successfully restored sensitivity to doxorubicin from 7 μM in the P-gp overexpressed human myelogenous leukemia cells, Lucena 1. This resistance-reversing effect was confirmed by competitively increasing the intracellular doxorubicin accumulation and by significantly inhibiting the efflux of doxorubicin and, to a lesser extent, that of rhodamine 123. The activity obtained was similar to that observed with verapamil. No such results were observed in the sensitive parental K562 cell line. To gain deeper insight into the mode of action of pinoresinol, its effect on P-gp function and expression was examined. The docking simulations indicated that the lignan bound to P-gp at the apex of the V-shaped transmembrane cavity, involving transmembrane helices 4, 5, and 6, and partially overlapped the binding region of tariquidar, which was used as a positive control. These results would shed some light on the nature of its interaction with P-gp at molecular level and merit further mechanistic and kinetic studies. In addition, it showed a maximum 29% activation of ATP hydrolysis and antagonized verapamil-stimulated ATPase activity with an IC 50 of 20.9 μM. On the other hand, pinoresinol decreased the presence of P-gp in the cell surface. Derivatives of pinoresinol with improved activity were identified by docking studies. The most promising one, the non-cytotoxic 1-acetoxypinoresinol, caused a reversion of doxorubicin resistance from 0.11 μM and thus higher activity than the lead compound. It also caused a significant increase in doxorubicin accumulation. Results were similar to those observed with verapamil. The results obtained positioned these compounds as potential candidates for effective agents to overcome P-gp-mediated MDR, leading to better outcomes for leukemia chemotherapy.