María Cecilia Carpinella

Screening novel artemisinin derivatives identified A16 and A20 as potent Rho123 efflux inhibitors (MEC: 49 nM), enhancing doxorubicin (Dox) cytotoxicity in K562/Dox cells at 50 and 5 nM, respectively.

The increasing prevalence of multidrug resistance (MDR) in Candida species poses a major obstacle to effective antifungal therapy. Since the efflux pumps Cdr1 and Mdr1 are key mediators of the MDR phenotype, particularly by conferring resistance to azoles, their inhibition has emerged as a promising strategy to restore antifungal susceptibility. Bioassay-guided fractionation of Solanum atriplicifolium, selected based on its ability to inhibit Mdr1 and Cdr1, led to the isolation of six previously undescribed acylhexoses, including four acylglucoses, named atriplicifolin A (1), atriplicifolin B (2), atriplicifolin C (3) and atriplicifolin D (4) and two acylinositols, named atriplicifolin E (5) and atriplicifolin F (6). Among these, compounds 3 and 4 successfully restored fluconazole sensitivity in resistant Saccharomyces cerevisiae strains overexpressing Mdr1 (AD/CaMDR1) and Cdr1 (AD/CaCDR1), with greater activity against the latter, with minimum effective concentrations (MECs) of 6.2 and 7.5 µM, respectively. Both compounds also significantly increased the intracellular accumulation of Nile red and rhodamine 6G in AD/CaMDR1 and AD/CaCDR1 strains, again showing a stronger effect in the Cdr1-overexpressing strain, where MEC values ranged from 3.1 to 15.5 µM. Molecular dynamics simulations supported these results by revealing that compounds 3 and 4 displayed higher binding affinity for Cdr1 than fluconazole, Nile red or rhodamine 6G. Furthermore, analysis of the persistence and energetic contribution of individual ligand–transporter interactions over time, particularly within specific regions of the transporter, provided valuable insights for the rational design of novel inhibitors. Overall, these findings identify compounds 3 and 4 as promising scaffolds for the development of Mdr1 and Cdr1 modulators aimed at overcoming fungal multidrug resistance.

Desde tiempos inmemoriales, la humanidad ha recurrido a la naturaleza para satisfacer sus necesidades. La utilización de medicinas herbales, a veces conocidas como fitomedicina o medicina botánica, ha sido una de las estrategias más destacada para tratar y prevenir muchas enfermedades. El conocimiento acerca de la terapéutica basada en plantas se remonta a tiempos milenarios, tal como ha sido documentado por diversas culturas, entre las más notables la egipcia, griega y romana, así como en la medicina tradicional china o en el sistema Ayurveda de India. Se estima que hay al menos 250.000 especies de plantas que coexisten en ecosistemas con otros seres vivos y en donde la química juega un rol fundamental1. Este último argumento se sustenta en la aptitud de las plantas de sintetizar metabolitos secundarios cuya principal función es la defensa frente a organismos indeseados, propiedad que ha sido aprovechada por el hombre para nutrir un vasto arsenal de sustancias bioactivas. Estos compuestos son capaces de interactuar con una importante variedad de proteínas y otros blancos biológicos, habilidad que los posiciona como efectivos moduladores de diversos procesos o componentes celulares afectando a organismos vivos perjudiciales o interfiriendo con diversas patologías. Esta propiedad ha dado lugar a diferentes aplicaciones en el campo de la farmacología. De esta manera las plantas se han constituido en la principal fuente de drogas terapéuticas siendo la base de reconocidos sistemas de medicina tradicional, muchos de los cuales son aún utilizados. Ha sido descripto que alrededor del 80% de la población mundial utiliza productos fitoterápicos como tratamiento primario o alternativo de la salud. En la actualidad, las drogas derivadas de plantas comprenden un alto porcentaje del total de entidades aprobadas y aproximadamente un tercio de los fármacos más vendidos provienen de especies vegetales. Las novedosas estructuras de las moléculas de productos naturales superan a las de otras fuentes: a través de comparaciones con las bases de datos, se ha podido determinar que un 40% de las características químicas estructurales de los metabolitos naturales están ausentes en los compuestos sintéticos. Este importante número de posibles farmacóforos es fundamental cuando se buscan nuevos agentes terapéuticos.

Continuing our search for bioactive compounds in species from the Juncaceae family, Juncus articulatus was investigated. Ten previously undescribed phenanthrenes─articulins A-J (1-10)─and ten known compounds─juncuenin B, dehydrojuncuenin B, juncatrin B, ensifolins E, F, H, I, K, juncuenin D, and luzulin A (11-20)─along with other compounds, have been isolated and identified. The isolated compounds were evaluated for antibacterial activity against Escherichia coli, Pseudomonas aeruginosa, methicillin-susceptible Staphylococcus aureus (MSSA), and methicillin-resistant Staphylococcus aureus (MRSA). Compounds 12 and 14 exhibited the most potent activity against planktonic and sessile MSSA and MRSA with minimum inhibitory concentration (MIC) values of 15.1 μM (12 for both bacterial strains) and 15.3 μM (14 for both bacterial strains). Compounds 15, 17, and 18 also exhibited activity against both strains, although to a lower extent, with MIC values ranging from 30.0 to 56.8 μM. The inhibition of biofilm formation of these compounds was observed at 15.1-114.3 μM. This study elucidates the phenanthrene composition of J. articulatus and the antibacterial effect of these compounds.

Giardiasis is a parasitosis caused by Giardia lamblia with significant epidemiological and clinical importance due to its high prevalence and pathogenicity. The lack of optimal therapies for treating this parasite makes the development of new effective chemical entities an urgent need. In the search for new inhibitors of the adenylyl cyclase gNC1 obtained from G. lamblia, 14 extracts from Argentinian native plants were screened. Lepechinia floribunda and L. meyenii extracts exhibited the highest gNC1 inhibitory activity, with IC₅₀ values of 9 and 31 µg/mL, respectively. In silico studies showed rosmarinic acid, a hydroxycinnamic acid present in both mentioned species, to be a promising anti-gNC1 compound. This result was confirmed experimentally, with rosmarinic acid showing an IC₅₀ value of 10.1 µM. Theoretical and experimental findings elucidate the molecular-level mechanism of rosmarinic acid, pinpointing the key interactions stabilizing the compound–enzyme complex and the binding site. These results strongly support that rosmarinic acid is a promising scaffold for developing novel compounds with inhibitory activity against gNC1, which could serve as potential therapeutic agents to treat giardiasis.

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.

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 P-glycoprotein (P-gp). Based on the chemical structure of the P-gp reverser betulin identified in our previous work, a series of analogs were designed, synthesized and evaluated as a source of novel inhibitors. Compounds 6g and 6i efficiently inhibited rhodamine 123 efflux in the P-gp overexpressed leukemia cells, Lucena 1, at concentrations of 0.19 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 Lucena 1 to Dox at 0.024 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.

Editorial: Considering plant metabolites and their synthetic derivatives as candidates for the development of drugs against multidrug resistant (MDR) tumors

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 IC₅₀ 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 IC₅₀ 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.

Background: The development of multidrug resistance (MDR) associated with the overexpression of the efflux transporters Mdr1 and Cdr1 in Candida species impedes antifungal therapies. The urgent need for novel agents able to inhibit the function of both pumps, led us to evaluate this property in 137 extracts obtained from Argentinian plants. Methods: The ability of the extracts to reverse efflux pump-mediated MDR was determined with an agar chemosensitization assay using fluconazole (FCZ) resistant Mdr1- and Cdr1-overexpressing clinical isolates of Candida albicans and Candida glabrata as well as Saccharomyces cerevisiae strains selectively expressing Mdr1 (AD/CaMDR1) or Cdr1 (AD/CaCDR1). The resistance-reversing activity of the most potent extracts was further confirmed using a Nile Red accumulation assay. Results: Fifteen plant extracts overcame the FCZ resistance of Candida albicans 1114, which overexpresses CaMdr1 and CaCdr1, and AD/CaMDR1, with those from Acalypha communis and Solanum atriplicifolium being the most effective showing 4- to 16-fold reversal of resistance at concentrations ≥ 25 µg/mL. Both extracts, and to a lesser extent that from Pterocaulon alopecuroides, also restored FCZ sensitivity in CgCdr1-overexpressing C. glabrata 109 and in AD/CaCDR1 with fold reversal values ranging from 4 to 32 and therefore demonstrating a dual effect against Mdr1 and Cdr1. Both, A. communis and S. atriplicifolium extracts at concentrations ≥ 12.5 and ≥ 25 µg/mL, respectively, increased the intracellular Nile Red accumulation in all yeast strains overexpressing efflux pumps. Conclusions: The non-toxic and highly active extracts from A. communis and S. atripicifolium, provide promising sources of compounds for potentiating the antifungal effect of FCZ by blocking the efflux function of Mdr1 and Cdr1 transporters.