RESUMEN
The development of inhibitors of key biological mechanisms involved in multidrug resistance (MDR) burden meets an important medical need but still represents a challenging task. Major MDR targets in both bacterial and cancer cells are multidrug efflux systems. Several aspects should be considered in the attempt to design efficient inhibitors of these systems such as toxicity, stability, permeability as a few examples. In order to successfully design promising new compounds, a full understanding of the efflux mechanism is required, from both biological and structural points of view. It is nowadays well established that the success rate in classical drug design and biological evaluation improves when combined with in silico methodologies. In this review, we focus on the biological evaluation and molecular mechanistic insights of inhibitors of the drug efflux activity of the Hedgehog receptor Patched1 (Ptch1). Ptch1 is known to be over-expressed in many types of cancers, but its activity and role in the resistance to chemotherapy of cancer cells have been highlighted only recently. Remarkably, due to its peculiar efflux mechanism, inhibition of Ptch1 was shown to be particularly relevant for improving the efficacy of chemotherapy without concomitant toxicity for healthy cells or potential side effects. To date, three compounds have been identified as efficient Ptch1 inhibitors, namely astemizole, methiothepin and panicein A hydroquinone. Due to the chemical and structural differences of these molecules, the hit-to-lead drug design is not straightforward. This review describes how the merging of in vitro, in vivo and in silico studies provides molecular details that could contribute to the rational design of new Ptch1 inhibitors.
Asunto(s)
Proteínas Hedgehog , Neoplasias , Receptor Patched-1 , Diseño de Fármacos , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Humanos , Neoplasias/tratamiento farmacológicoRESUMEN
We previously reported that methiothepin, a small molecule known as a nonselective serotonin 5-HT receptor antagonist, inhibited the doxorubicin efflux activity of the Hedgehog receptor Ptch1 and enhanced the cytotoxic, pro-apoptotic, anti-proliferative, and anti-clonogenic effects of doxorubicin on adrenocortical carcinoma cells. Here, we show that methiothepin also inhibits doxorubicin efflux and increases doxorubicin cytotoxicity in melanoma cells which endogenously overexpress Ptch1. Melanoma patients having the BRAFV600E mutation are treated with vemurafenib, an inhibitor of BRAFV600E, often in combination with trametinib, an inhibitor of MEK. Almost all patients ultimately acquire resistance to the treatment leading to disease progression. Here, we report that methiothepin overcomes the resistance of BRAFV600E melanoma cells by enhancing the cytotoxicity of vemurafenib and trametinib on these cells leading to melanoma cells death. We observe that the addition of methiothepin to vemurafenib prevents migration of resistant melanoma cells more efficiently than vemurafenib alone. Our results provide an additional proof that Ptch1 drug efflux inhibition increases the effectiveness of anti-cancer treatments and overcomes resistance of melanoma cells expressing Ptch1.
Asunto(s)
Antineoplásicos , Resistencia a Antineoplásicos/efectos de los fármacos , Melanoma/tratamiento farmacológico , Metiotepina , Neoplasias Cutáneas/tratamiento farmacológico , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Doxorrubicina/administración & dosificación , Humanos , Metiotepina/farmacología , Metiotepina/uso terapéutico , Receptor Patched-1/metabolismo , Piridonas/administración & dosificación , Pirimidinonas/administración & dosificación , Vemurafenib/administración & dosificaciónRESUMEN
Resistance-nodulation-cell division family proteins are transmembrane proteins identified as large spectrum drug transporters involved in multidrug resistance. A prototypical case in this superfamily, responsible for antibiotic resistance in selected gram-negative bacteria, is AcrB. AcrB forms a trimer using the proton motive force to efflux drugs, implementing a functional rotation mechanism. Unfortunately, the size of the system (1049 amino acid per monomer and membrane) has prevented a systematic dynamical exploration, so that the mild understanding of this coupled transport jeopardizes our ability to counter it. The large number of crystal structures of AcrB prompts studies to further our understanding of the mechanism. To this end, we present a novel strategy based on two key ingredients, which are to study dynamics by exploiting information embodied in the numerous crystal structures obtained to date, and to systematically consider subdomains, their dynamics, and their interactions. Along the way, we identify the subdomains responsible for dynamic events, refine the states (A, B, E) of the functional rotation mechanism, and analyze the evolution of intramonomer and intermonomer interfaces along the functional cycle. Our analysis shows the relevance of AcrB's efflux mechanism as a template within the HAE1 family but not beyond. It also paves the way to targeted simulations exploiting the most relevant degrees of freedom at certain steps, and to a targeting of specific interfaces to block the drug efflux. Our work shows that complex dynamics can be unveiled from static snapshots, a strategy that may be used on a variety of molecular machines of large size.
Asunto(s)
Proteínas de Escherichia coli , Proteínas Asociadas a Resistencia a Múltiples Medicamentos , Sitio Alostérico , Antibacterianos/química , Antibacterianos/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Simulación de Dinámica Molecular , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/química , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Unión Proteica , Conformación ProteicaRESUMEN
Adrenocortical carcinoma (ACC) presents a high risk of relapse and metastases with outcomes not improving despite extensive research and new targeted therapies. We recently showed that the Hedgehog receptor Patched is expressed in ACC, where it strongly contributes to doxorubicin efflux and treatment resistance. Here, we report the identification of a new inhibitor of Patched drug efflux, the anti-histaminergic drug astemizole. We show that astemizole enhances the cytotoxic, proapoptotic, antiproliferative and anticlonogenic effects of doxorubicin on ACC cells at concentrations of astemizole or doxorubicin that are not effective by themselves. Our results suggest that a low concentration of astemizole sensitizes ACC cells to doxorubicin, which is a component of the standard treatment for ACC composed of etoposide, doxorubicin, cisplatin and mitotane (EDPM). Patched uses the proton motive force to efflux drugs. This makes its function specific to cancer cells, thereby avoiding toxicity issues that are commonly observed with inhibitors of ABC multidrug transporters. Our data provide strong evidence that the use of astemizole or a derivative in combination with EDPM could be a promising therapeutic option for ACC by increasing the treatment effectiveness at lower doses of EDPM, which would reduce the severe side effects of this regimen.
RESUMEN
Melanoma patients harboring the BRAFV600E mutation are treated with vemurafenib. Almost all of them ultimately acquire resistance, leading to disease progression. Here, we find that a small molecule from a marine sponge, panicein A hydroquinone (PAH), overcomes resistance of BRAFV600E melanoma cells to vemurafenib, leading to tumor elimination in corresponding human xenograft models in mice. We report the synthesis of PAH and demonstrate that this compound inhibits the drug efflux activity of the Hedgehog receptor, Patched. Our SAR study allowed identifying a key pharmacophore responsible for this activity. We showed that Patched is strongly expressed in metastatic samples from a cohort of melanoma patients and is correlated with decreased overall survival. Patched is a multidrug transporter that uses the proton motive force to efflux drugs. This makes its function specific to cancer cells, thereby avoiding toxicity issues that are commonly observed with inhibitors of ABC multidrug transporters. Our data provide strong evidence that PAH is a highly promising lead for the treatment of vemurafenib resistant BRAFV600E melanoma.
RESUMEN
Knowledge of the bioactive conformations of small molecules or the ability to predict them with theoretical methods is of key importance to the design of bioactive compounds such as drugs, agrochemicals, and cosmetics. Using an elaborate cheminformatics pipeline, which also evaluates the support of individual atom coordinates by the measured electron density, we compiled a complete set ("Sperrylite Dataset") of high-quality structures of protein-bound ligand conformations from the PDB. The Sperrylite Dataset consists of a total of 10,936 high-quality structures of 4,548 unique ligands. Based on this dataset, we assessed the variability of the bioactive conformations of 91 small molecules-each represented by a minimum of ten structures-and found it to be largely independent of the number of rotatable bonds. Sixty-nine molecules had at least two distinct conformations (defined by an RMSD greater than 1 Å). For a representative subset of 17 approved drugs and cofactors we observed a clear trend for the formation of few clusters of highly similar conformers. Even for proteins that share a very low sequence identity, ligands were regularly found to adopt similar conformations. For cofactors, a clear trend for extended conformations was measured, although in few cases also coiled conformers were observed. The Sperrylite Dataset is available for download from http://www.zbh.uni-hamburg.de/sperrylite_dataset.