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Although opioid analgesics are indispensable in treating pain, these drugs are accompanied by life-threatening side effects. While clinically relevant opioid drugs target the µ opioid receptor (MOR), a heterodimer between the MOR and the δ opioid receptor (DOR) has emerged as another target to develop safer analgesics. Although some heterodimer-preferring agonists have been reported so far, it is still difficult to activate the MOR/DOR heterodimer selectively in the presence of MOR or DOR monomers/homodimers. To gain insights to develop selective agonists for MOR/DOR, herein we prepared analogs of CYM51010, one of the reported heterodimer-preferring agonists, and collected structure-activity relationship information. We found that the ethoxycarbonyl group was needed for the activity for the heterodimer, although this group could be substituted with functional groups with similar sizes, such as an ethoxycarbonyl group. As for the acetylaminophenyl group, not a type of substituent, but rather a substituent located at a specific position (para-position) was essential for the activity. Changing the linker length between the acetylaminophenyl group and the piperidine moiety also had deleterious effects on the activity. On the other hand, the substitution of the acetylamino group with a trifluoroacetylamino group and the substitution of the phenethyl group with a benzyl group diminished the activities for the monomers/homodimers while keeping the activity for MOR/DOR, which enhanced the selectivity. Our findings herein will play an important role in developing selective agonists for MOR/DOR and for elucidating the physiological roles of this heterodimer in analgesic processes and in the establishment of side effects.
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Receptores Opioides delta , Receptores Opioides mu , Relação Estrutura-Atividade , Receptores Opioides delta/agonistas , Receptores Opioides delta/metabolismo , Receptores Opioides mu/agonistas , Receptores Opioides mu/metabolismo , Humanos , Estrutura Molecular , Animais , Analgésicos Opioides/química , Analgésicos Opioides/farmacologia , Analgésicos Opioides/síntese química , Relação Dose-Resposta a Droga , Cricetulus , Células CHORESUMO
Small synthetic oligodeoxynucleotides (ODNs) can mimic microbial nucleic acids by interacting with receptor systems and promoting immunostimulatory activities. Nevertheless, some ODNs can act differently on the plasmacytoid dendritic cell (pDC) subset, shaping their immunoregulatory properties and rendering them suitable immunotherapeutic tools in several clinical settings for treating overwhelming immune responses. We designed HIV-1-derived, DNA- and RNA-based oligonucleotides (gag, pol, and U5 regions) and assessed their activity in conferring a tolerogenic phenotype to pDCs in skin test experiments. RNA-but not DNA-oligonucleotides are capable of inducing tolerogenic features in pDCs. Interestingly, sensing the HIV-1-derived single-stranded RNA-gag oligonucleotide (RNA-gag) requires both TLR3 and TLR7 and the engagement of the TRIF adaptor molecule. Moreover, the induction of a suppressive phenotype in pDCs by RNA-gag is contingent upon the induction and activation of the immunosuppressive enzyme Arginase 1. Thus, our data suggest that sensing of the synthetic RNA-gag oligonucleotide in pDCs can induce a suppressive phenotype in pDCs, a property rendering RNA-gag a potential tool for therapeutic strategies in allergies and autoimmune diseases.
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Arginase , Células Dendríticas , HIV-1 , Arginase/metabolismo , Humanos , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Tolerância Imunológica , Oligonucleotídeos , RNA Viral/genética , RNA Viral/metabolismoRESUMO
Although survival from breast cancer has dramatically increased, many will develop recurrent, metastatic disease. Unfortunately, survival for this stage of disease remains very low. Activating the immune system has incredible promise since it has the potential to be curative. However, immune checkpoint blockade (ICB) which works through T cells has been largely disappointing for metastatic breast cancer. One reason for this is a suppressive myeloid immune compartment that is unaffected by ICB. Cholesterol metabolism and proteins involved in cholesterol homeostasis play important regulatory roles in myeloid cells. Here, we demonstrate that NR0B2, a nuclear receptor involved in negative feedback of cholesterol metabolism, works in several myeloid cell types to impair subsequent expansion of regulatory T cells (Tregs); Tregs being a subset known to be highly immune suppressive and associated with poor therapeutic response. Within myeloid cells, NR0B2 serves to decrease many aspects of the inflammasome, ultimately resulting in decreased IL1ß; IL1ß driving Treg expansion. Importantly, mice lacking NR0B2 exhibit accelerated tumor growth. Thus, NR0B2 represents an important node in myeloid cells dictating ensuing Treg expansion and tumor growth, thereby representing a novel therapeutic target to re-educate these cells, having impact across different solid tumor types. Indeed, a paper co-published in this issue demonstrates the therapeutic utility of targeting NR0B2.
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Neoplasias da Mama , Progressão da Doença , Células Mieloides , Linfócitos T Reguladores , Linfócitos T Reguladores/imunologia , Animais , Feminino , Camundongos , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Humanos , Células Mieloides/imunologia , Células Mieloides/metabolismo , Camundongos Knockout , Interleucina-1beta/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Inflamassomos/metabolismo , Inflamassomos/imunologiaRESUMO
Immune checkpoint blockade (ICB) has had limited utility in several solid tumors such as breast cancer, a major cause of cancer-related mortality in women. Therefore, there is considerable interest in alternate strategies to promote an anti-cancer immune response. A paper co-published in this issue describes how NR0B2, a protein involved in cholesterol homeostasis, functions within myeloid immune cells to modulate the inflammasome and reduce the expansion of immune-suppressive regulatory T cells (Treg). Here, we develop NR0B2 as a potential therapeutic target. NR0B2 in tumors is associated with improved survival for several cancer types including breast. Importantly, NR0B2 expression is also prognostic of ICB success. Within breast tumors, NR0B2 expression is inversely associated with FOXP3, a marker of Tregs. While a described agonist (DSHN) had some efficacy, it required high doses and long treatment times. Therefore, we designed and screened several derivatives. A methyl ester derivative (DSHN-OMe) emerged as superior in terms of (1) cellular uptake, (2) ability to regulate expected expression of genes, (3) suppression of Treg expansion using in vitro co-culture systems, and (4) efficacy against the growth of primary and metastatic tumors. This work identifies NR0B2 as a target to re-educate myeloid immune cells and a novel ligand with significant anti-tumor efficacy in preclinical models.
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Células Mieloides , Linfócitos T Reguladores , Humanos , Células Mieloides/imunologia , Células Mieloides/metabolismo , Células Mieloides/efeitos dos fármacos , Feminino , Animais , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Neoplasias da Mama/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/imunologia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Camundongos , Linhagem Celular Tumoral , Microambiente Tumoral , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêuticoRESUMO
Aureocin A53 is a peptide bacteriocin produced by an opportunistic pathogen Staphylococcus aureus strain A53. The spatial structure of aureocin, unlike its amino acid sequence, is similar to the bacteriocin BacSp222, which was recently found to have the ability to induce the inflammatory response in the host cells. The presented research aimed to verify such properties also for aureocin A53. We demonstrated that the synthetic aureocin has slight cytotoxic activity towards murine monocytic-macrophage cells. This molecule was also able to activate murine P388.D1 and RAW 264.7 cells to IFN-γ-dependent production of nitric oxide and to activate production of the pro-inflammatory cytokine - TNF. We also proved that the observed pro-inflammatory activity of the studied bacteriocin is related to the stimulation of the TLR2/TLR6 heterodimer and, consequently, activation of the NF-κB transcription factor. To sum up, A53 is the second bacteriocin described in the literature, showing the pro-inflammatory activity against murine macrophage-like cells.
Assuntos
Bacteriocinas , Macrófagos , Óxido Nítrico , Staphylococcus aureus , Camundongos , Animais , Bacteriocinas/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/imunologia , Staphylococcus aureus/efeitos dos fármacos , Células RAW 264.7 , Óxido Nítrico/metabolismo , NF-kappa B/metabolismo , Receptor 2 Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Linhagem Celular , Citocinas/metabolismo , Interferon gama/metabolismo , Inflamação/metabolismoRESUMO
Immune thrombocytopenia (ITP) is an autoimmune disease characterized by a low platelet (PLT) count and a high risk of bleeding, the clinical treatment for which still needs to be upgraded. Based on the critical role of human leukocyte antigen class II heterodimer ß5 (HLA-DRB5) in immune system, we herein investigated its effect on ITP. ITP murine models were established by the injection of guinea pig anti-mouse platelet serum (GP-APS), and the PLT of mouse peripheral blood was counted during the modeling. Quantitative real-time reverse transcription polymerase chain reaction, western blot and immunofluorescence assay was performed to quantify expressions of HLA-DRB5, major histocompatibility complex II (MHC-II) and co-stimulatory molecules (CD80, CD86). Flow cytometry was conducted to analyze the percentage of CD8+ T cells. As a result, the PLT count was decreased in mouse peripheral blood. Expressions of HLA-DRB5, MHC-II and co-stimulatory molecules, as well as the percentage of CD8+ T cells were elevated in peripheral blood of ITP mice. HLA-DRB5 knockdown mitigated ITP by increasing peripheral PLT level, downregulating expressions of MHC-II and co-stimulatory molecules and inactivating CD8+ T cells. Collectively, the downregulation of HLA-DRB5 restores the peripheral PLT count in ITP mice by reducing MHC-II-mediated antigen presentation of macrophages to inhibit the activation of CD8+ T cells.
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The deep-sea fungus Phomopsis lithocarpus FS508 produces tenellone-macrolide conjugated hetero-dimer lithocarpins A-G with anti-tumor activities. The deficiency of new intermolecular Diels-Alder (DA) enzymes hindered the development of new bioactive hetero-dimers. A novel single-function intermolecular DA enzyme, g7882, was initially discovered in this study. The deletion of g7882 led to the disappearance of lithocarpin A and an increase in precursor level . the overexpression of g7882 significantly improved lithocarpin A yield. The in vitro function of g7882DA was also confirmed by biochemical reaction using tenellone B as a substrate. Additionally, the knockout of KS modules of PKS in cluster 41 and cluster 81 (lit cluster) eliminated the production of lithocarpins, which firstly explains the biosynthetic process of hetero-dimer lithocarpins mediated by DA enzyme in FS508. Furthermore, the removal of a novel acetyltransferase GPAT in cluster 41 and the oxidoreductase, prenyltransferase in cluster81 resulted in the reduction of lithocarpin A in P. lithocarpus. The overexpression of gpat in P. lithocarpus FS508 improved the yield of lithocarpin A significantly and produced a new tenellone derivative lithocarol G. This study offers a new DA enzyme tool for the biosynthesis of novel hetero-dimer and biochemical clues for the biosynthetic logic elucidation of lithocarpins.
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Reação de Cicloadição , Ascomicetos/enzimologia , Família MultigênicaRESUMO
Ischemic stroke is the leading cause of death and disability worldwide. Nevertheless, there still lacks the effective therapies for ischemic stroke. Microglia are resident macrophages of the central nervous system (CNS) and can initiate immune responses and monitor the microenvironment. Microglia are activated and polarize into proinflammatory or antiinflammatory phenotype in response to various brain injuries, including ischemic stroke. Proinflammatory microglia could generate immunomodulatory mediators, containing cytokines and chemokines, these mediators are closely associated with secondary brain damage following ischemic stroke. On the contrary, anti-inflammatory microglia facilitate recovery following stroke. Regulating the activation and the function of microglia is crucial in exploring the novel treatments for ischemic stroke patients. Accumulating studies have revealed that RhoA/ROCK pathway and NF-κB are famous modulators in the process of microglia activation and polarization. Inhibiting these key modulators can promote the polarization of microglia to anti-inflammatory phenotype. In this review, we aimed to provide a comprehensive overview on the role of RhoA/ROCK pathway and NF-κB in the microglia activation and polarization, reveal the relationship between RhoA/ROCK pathway and NF-κB in the pathological process of ischemic stroke. In addition, we likewise discussed the drug modulators targeting microglia polarization.
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AVC Isquêmico , Microglia , NF-kappa B , Transdução de Sinais , Quinases Associadas a rho , Proteína rhoA de Ligação ao GTP , Microglia/metabolismo , NF-kappa B/metabolismo , Humanos , Quinases Associadas a rho/metabolismo , Animais , Proteína rhoA de Ligação ao GTP/metabolismo , AVC Isquêmico/metabolismo , AVC Isquêmico/imunologia , AVC Isquêmico/patologia , Transdução de Sinais/fisiologia , Polaridade Celular/fisiologia , Polaridade Celular/efeitos dos fármacosRESUMO
Optimal therapeutic and diagnostic efficacy is essential for healthcare's global mission of advancing oncologic drug development. Accurate diagnosis and detection are crucial prerequisites for effective risk stratification and personalized patient care in clinical oncology. A paradigm shift is emerging with the promise of multi-receptor-targeting compounds. While existing detection and staging methods have demonstrated some success, the traditional approach of monotherapy is being reevaluated to enhance therapeutic effectiveness. Heterodimeric site-specific agents are a versatile solution by targeting two distinct biomarkers with a single theranostic agent. This review describes the innovation of dual-targeting compounds, examining their design strategies, therapeutic implications, and the promising path they present for addressing complex diseases.
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Transcription factors BACH2 and IRF4 are both essential for antibody class-switch recombination (CSR) in activated B lymphocytes, while they oppositely regulate the differentiation of plasma cells (PCs). Here, we investigated how BACH2 and IRF4 interact during CSR and plasma-cell differentiation. We found that BACH2 organizes heterochromatin formation of target gene loci in mouse splenic B cells, including targets of IRF4 activation such as Aicda, an inducer of CSR, and Prdm1, a master plasma-cell regulator. Release of these gene loci from heterochromatin in response to B-cell receptor stimulation was coupled to AKT-mTOR pathway activation. In Bach2-deficient B cells, PC genes' activation depended on IRF4 protein accumulation, without an increase in Irf4 mRNA. Mechanistically, a PU.1-IRF4 heterodimer in activated B cells promoted BACH2 function by inducing gene expression of Bach2 and Pten, a negative regulator of AKT signaling. Elevated AKT activity in Bach2-deficient B cells resulted in IRF4 protein accumulation. Thus, BACH2 and IRF4 mutually modulate the activity of each other, and BACH2 inhibits PC differentiation by both the repression of PC genes and the restriction of IRF4 protein accumulation.
Assuntos
Fatores de Transcrição de Zíper de Leucina Básica , Diferenciação Celular , Fatores Reguladores de Interferon , Plasmócitos , Animais , Camundongos , Linfócitos B/metabolismo , Linfócitos B/imunologia , Linfócitos B/citologia , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/genética , Diferenciação Celular/genética , Heterocromatina/metabolismo , Heterocromatina/genética , Switching de Imunoglobulina/genética , Fatores Reguladores de Interferon/metabolismo , Fatores Reguladores de Interferon/genética , Camundongos Endogâmicos C57BL , Camundongos Knockout , Plasmócitos/metabolismo , Plasmócitos/imunologia , Plasmócitos/citologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , PTEN Fosfo-Hidrolase/genética , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Serina-Treonina Quinases TOR/genética , Transativadores/metabolismo , Transativadores/genéticaRESUMO
Heterodimers of metal nanocrystals (NCs) with tailored elemental distribution have emerged as promising candidates in the field of electrocatalysis, owing to their unique structures featuring heterogeneous interfaces with distinct components. Despite this, the rational synthesis of heterodimer NCs with similar elemental composition remains a formidable challenge, and their impact on electrocatalysis has remained largely elusive. In this study, Pd@Bi-PdBi heterodimer NCs are synthesized through an underpotential deposition (UPD)-directed growth pathway. In this pathway, the UPD of Bi promotes a Volmer-Weber growth mode, allowing for judicious modulation of core-satellite to heterodimer structures through careful control of supersaturation and growth kinetics. Significantly, the heterodimer NCs are employed in the electrocatalytic process of ethylene glycol (EG) with high activity and selectivity. Compared with pristine Pd octahedra and common PdBi alloy NC, the unique heterodimer structure of the Pd@Bi-PdBi heterodimer NCs endows them with the highest electrocatalytic performance of EG and the best selectivity (≈93%) in oxidizing EG to glycolic acid (GA). Taken together, this work not only heralds a new strategy for UPD-directed synthesis of bimetallic NCs, but also provides a new design paradigm for steering the selectivity of electrocatalysts.
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Directed structural modifications of natural products offer excellent opportunities to develop selectively acting drug candidates. Natural product hybrids represent a particular compound group. The components of hybrids constructed from different molecular entities may result in synergic action with diminished side effects. Steroidal homo- or heterodimers deserve special attention owing to their potentially high anticancer effect. Inspired by our recently described antiproliferative core-modified estrone derivatives, here, we combined them into heterodimers via Cu(I)-catalyzed azide-alkyne cycloaddition reactions. The two trans-16-azido-3-(O-benzyl)-17-hydroxy-13α-estrone derivatives were reacted with 3-O-propargyl-D-secoestrone alcohol or oxime. The antiproliferative activities of the four newly synthesized dimers were evaluated against a panel of human adherent gynecological cancer cell lines (cervical: Hela, SiHa, C33A; breast: MCF-7, T47D, MDA-MB-231, MDA-MB-361; ovarian: A2780). One heterodimer (12) exerted substantial antiproliferative activity against all investigated cell lines in the submicromolar or low micromolar range. A pronounced proapoptotic effect was observed by fluorescent double staining and flow cytometry on three cervical cell lines. Additionally, cell cycle blockade in the G2/M phase was detected, which might be a consequence of the effect of the dimer on tubulin polymerization. Computational calculations on the taxoid binding site of tubulin revealed potential binding of both steroidal building blocks, mainly with hydrophobic interactions and water bridges.
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Antineoplásicos , Proliferação de Células , Estrona , Humanos , Estrona/farmacologia , Estrona/análogos & derivados , Estrona/química , Estrona/síntese química , Proliferação de Células/efeitos dos fármacos , Antineoplásicos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Apoptose/efeitos dos fármacos , Dimerização , Simulação de Acoplamento Molecular , Feminino , Ensaios de Seleção de Medicamentos Antitumorais , Células HeLa , Tubulina (Proteína)/metabolismo , Tubulina (Proteína)/química , Células MCF-7RESUMO
Vascular endothelial growth factor 165 (VEGF165) is a prominent isoform of the VEGF-A protein that plays a crucial role in various angiogenesis-related diseases. It is homodimeric, and each of its monomers is composed of two domains connected by a flexible linker. DNA aptamers, which have emerged as potent therapeutic molecules for many proteins with high specificity and affinity, can also work for VEGF165. A DNA aptamer heterodimer composed of monomers of V7t1 and del5-1 connected by a flexible linker (V7t1:del5-1) exhibits a greater binding affinity with VEGF165 compared to either of the two monomers alone. Although the structure of the complex formed between the aptamer heterodimer and VEGF165 is unknown due to the highly flexible linkers, gaining structural information will still be valuable for future developments. Toward this end of accessing structural information, we adopt an ensemble docking approach here. We first obtain an ensemble of structures for both VEGF165 and the aptamer heterodimer by considering both small- and large-scale motions. We then proceed through an extraction process based on ensemble docking, molecular dynamics simulations, and binding free energy calculations to predict the structures of the VEGF165/V7t1:del5-1 complex. Through the same procedures, we reach a new aptamer heterodimer that bears a locked nucleic acid-modified counterpart of V7t1, namely RNV66:del5-1, which also binds well with VEGF165. We apply the same protocol to the monomeric units V7t1, RNV66, and del5-1 to target VEGF165. We observe that V7t1:del5-1 and RNV66:del5-1 show higher binding affinities with VEGF165 than any of the monomers, consistent with experiments that support the notion that aptamer heterodimers are more effective anti-VEGF165 aptamers than monomeric aptamers. Among the five different aptamers studied here, the newly designed RNV66:del5-1 shows the highest binding affinity with VEGF165. We expect that our ensemble docking approach can help in de novo designs of homo/heterodimeric anti-angiogenic drugs to target the homodimeric VEGF165.
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Aptâmeros de Nucleotídeos , Fator A de Crescimento do Endotélio Vascular , Inibidores da Angiogênese , Simulação de Dinâmica Molecular , Movimento (Física)RESUMO
Schistosoma japonicum glutathione-S-transferase (SjGST), the so-called GST-tag, is one of the most widely used protein tags for the purification of recombinant proteins by affinity chromatography. Attachment of SjGST enables the purification of a protein of interest (POI) using commercially available glutathione-immobilizing resins. Here we produced an SjGST mutant pair that forms heterodimers by adjusting the salt bridge pairs in the homodimer interface of SjGST. An MD study confirmed that the SjGST mutant pair did not disrupt the heterodimer formation. The modified SjGST protein pair coexpressed in Escherichia coli was purified by glutathione-immobilized resin. The stability of the heterodimeric form of the SjGST mutant pair was further confirmed by size exclusion chromatography. Surface plasmon resonance measurements unveiled the selective formation of heterodimers within the pair, accompanied by a significant suppression of homodimerization. The heterodimeric SjGST exhibited enzymatic activity in assays employing a commercially available fluorescent substrate. By fusing one member of the heterodimeric SjGST pair with a fluorescent protein and the other with the POI, we were able to conveniently and sensitively detect protein-protein interactions using fluorescence spectroscopy in the pull-down assays. Thus, utilization of the heterodimeric SjGST would be a useful tag for protein science.
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Cromatografia de Afinidade , Glutationa Transferase , Schistosoma japonicum , Schistosoma japonicum/enzimologia , Glutationa Transferase/metabolismo , Glutationa Transferase/química , Glutationa Transferase/isolamento & purificação , Glutationa Transferase/genética , Animais , Cromatografia de Afinidade/métodos , Multimerização Proteica , Proteínas de Helminto/metabolismo , Proteínas de Helminto/química , Proteínas de Helminto/genética , Proteínas de Helminto/isolamento & purificação , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Modelos MolecularesRESUMO
BACKGROUND: The androgen/androgen receptor (AR)-signaling axis plays a central role in prostate cancer (PCa). Upon androgen-binding the AR dimerizes with another AR, and translocates into the nucleus where the AR-dimer activates/inactivates androgen-dependent genes. Consequently, treatments for PCa are commonly based on androgen deprivation therapy (ADT). The clinical benefits of ADT are only transitory and most tumors develop mechanisms allowing the AR to bypass its need for physiological levels of circulating androgens. Clinical failure of ADT is often characterized by the synthesis of a constitutively active AR splice variant, termed AR-V7. AR-V7 mRNA expression is considered as a resistance mechanism following ADT. AR-V7 no longer needs androgenic stimuli for nuclear entry and/or dimerization. METHODS: Our goal was to mechanistically decipher the interaction between full-length AR (AR-FL) and AR-V7 in AR-null HEK-293 cells using the NanoLuc Binary Technology under androgen stimulation and deprivation conditions. RESULTS: Our data point toward a hypothesis that AR-FL/AR-FL homodimers form in the cytoplasm, whereas AR-V7/AR-V7 homodimers localize in the nucleus. However, after androgen stimulation, all the AR-FL/AR-FL, AR-FL/AR-V7 and AR-V7/AR-V7 dimers were localized in the nucleus. CONCLUSIONS: We showed that AR-FL and AR-V7 form heterodimers that localize to the nucleus, whereas AR-V7/AR-V7 dimers were found to localize in the absence of androgens in the nucleus.
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Luciferases , Neoplasias de Próstata Resistentes à Castração , Neoplasias da Próstata , Masculino , Humanos , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Androgênios , Neoplasias da Próstata/patologia , Antagonistas de Androgênios/farmacologia , Antagonistas de Androgênios/uso terapêutico , Células HEK293 , Neoplasias de Próstata Resistentes à Castração/patologia , Isoformas de Proteínas/genéticaRESUMO
BACKGROUND: v-RAF murine sarcoma viral homolog B1 (BRAF) is one of the most frequently mutated kinases in human cancers. BRAF exhibits three classes of mutations: Class I monomeric mutants (BRAFV600), class II BRAF homodimer mutants (non-V600), and class III BRAF heterodimers (non-V600). METHOD: In this manuscript, the protein-ligand interaction site of all three mutants: BRAF monomer, BRAF homodimer BRAF2:14-3-32, and BRAF heterodimer BRAF:14-3-32:MEK (Mitogen extracellular Kinase) has been discussed. FDA-approved drugs still have limitations against all three classes of mutants, especially against the second and third classes. Using the DesPot grid model, 1114 new compounds were designed. Using virtual screening, the three PDB Ids 4XV2 for monomers, 7MFF for homodimers, and 4MNE for heterodimers were used for 1114 newly designed compounds. RESULT: Dabrafenib, encorafenib, sorafenib and vemurafenib were included as standard drugs. The top 10 hit molecules were identified for each protein. Additional binding studies were performed using molecular docking studies on the protein-ligand site of each PDB identifier. Absorption, distribution, metabolism, excretion (ADME) and toxicity studies were also performed. CONCLUSION: It was identified that top-hit molecules had better binding and interaction activity than standard in all three classes of mutants.
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The occurrence of benign prostate hyperplasia (BPH) was related to disrupted sex steroid hormones, and metformin (Met) had a clinical response to sex steroid hormone-related gynaecological disease. However, whether Met exerts an antiproliferative effect on BPH via sex steroid hormones remains unclear. Here, our clinical study showed that along with prostatic epithelial cell (PEC) proliferation, sex steroid hormones were dysregulated in the serum and prostate of BPH patients. As the major contributor to dysregulated sex steroid hormones, elevated dihydrotestosterone (DHT) had a significant positive relationship with the clinical characteristics of BPH patients. Activation of adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) by Met restored dysregulated sex steroid hormone homeostasis and exerted antiproliferative effects against DHT-induced proliferation by inhibiting the formation of androgen receptor (AR)-mediated Yes-associated protein (YAP1)-TEA domain transcription factor (TEAD4) heterodimers. Met's anti-proliferative effects were blocked by AMPK inhibitor or YAP1 overexpression in DHT-cultured BPH-1 cells. Our findings indicated that Met would be a promising clinical therapeutic approach for BPH by inhibiting dysregulated steroid hormone-induced PEC proliferation.
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Integrin receptor αvß3 and gastrin-releasing peptide receptor (GRPR) expression of tumors could be detected using PET imaging with radiolabeled Arg-Gly-Asp (RGD) and the antagonistic bombesin analog RM26, respectively. The purpose of this study was to investigate the dual receptor-targeting property of the heterodimer RGD-RM26-03 (denoted as LNC1015), demonstrate the tumor diagnostic value of [68Ga]Ga-LNC1015 in preclinical experiments, and evaluate its preliminary clinical feasibility. METHODS: LNC1015 was designed and synthesized by linking cyclic RGD and the RM26 peptide. Preclinical pharmacokinetics were detected in a PC3 xenograft model using microPET and biodistribution studies. The clinical feasibility of [68Ga]Ga-LNC1015 PET/CT was performed in patients with breast cancer, and the results were compared with those of 18F-fluorodeoxyglucose (FDG). RESULTS: [68Ga]Ga-LNC1015 had good stability in saline for at least 2 h, and favorable binding affinity and specificity were demonstrated in vitro and in vivo. The tumor uptake and retention of [68Ga]Ga-LNC1015 during PET imaging were improved compared with its monomeric counterparts [68Ga]Ga-RGD and [68Ga]Ga-RM26 at all the time points examined. In our initial clinical studies, the tumor uptake and tumor-to-background ratio (TBR) of primary and metastatic lesions in [68Ga]Ga-LNC1015 PET/CT were significantly higher than those in [18F]FDG PET/CT, resulting in high lesion detection rate and tumor delineation. CONCLUSION: The dual targeting radiotracer [68Ga]Ga-LNC1015 showed significantly improved tumor uptake and retention, as well as lower liver uptake than [68Ga]Ga-RGD and [68Ga]Ga-RM26 monomer. The first-in-human study showed high TBRs in patients, suggesting favorable pharmacokinetics and high clinical feasibility for PET/CT imaging of cancer.
Assuntos
Radioisótopos de Gálio , Integrina alfaVbeta3 , Oligopeptídeos , Receptores da Bombesina , Receptores da Bombesina/metabolismo , Humanos , Animais , Camundongos , Feminino , Integrina alfaVbeta3/metabolismo , Oligopeptídeos/farmacocinética , Oligopeptídeos/química , Distribuição Tecidual , Masculino , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Radioquímica , Pessoa de Meia-Idade , Linhagem Celular Tumoral , Traçadores Radioativos , Compostos Radiofarmacêuticos/farmacocinética , Compostos Radiofarmacêuticos/síntese química , Compostos Radiofarmacêuticos/química , Técnicas de Química Sintética , Neoplasias da Mama/diagnóstico por imagem , Neoplasias da Mama/metabolismoRESUMO
BACKGROUND: In this study, [64Cu]Cu-NODAGA-RGD-BBN was prepared and its preclinical assessments were evaluated for PET imaging of GRPR overexpressing tumors. METHODS: NODAGA-RGD-BBN heterodimer peptide was successfully labeled with cyclotronproduced copper-64 at optimized conditions. The radiochemical purity of the radiotracer was checked by HPLC and RTLC methods. The stability of the radiolabeled compound was assessed in PBS (4°C) and in human blood serum (37°C). Binding affinity and internalization of [64Cu]Cu-NODAGA-RGD-BBN were studied on PC3, LNCaP, and CHO cell lines. The biodistribution of the radiotracer was evaluated in normal and tumor-bearing mice. RESULTS: [64Cu]Cu-NODAGA-RGD-BBN was prepared with radiochemical purity >99 ± 0.7% (HPLC/ITLC) and specific activity of 18.5 ± 2.2 TBq/mmol. The radiotracer showed high stability in PBS (95 ± 1.05%) and in human blood serum (96 ± 1.24%) and, high affinity to the GRP expressing tumor cells. [64Cu]Cu-NODAGA-RGD-BBN showed hydrophilic (log p = -1.14) and agonistic nature. The biodistribution and imaging studies demonstrated high uptake at the tumor site at all intervals post-injection and 3-4 h post-injection can be considered an appropriate time of imaging. CONCLUSION: The results indicated that [64Cu]Cu-NODAGA-RGD-BBN radiolabeled heterodimer peptide can be considered as a high-potential agent for PET imaging of GRPRoverexpressing tumors.
RESUMO
In the present study, the formation of a heterodimer involving both epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) has been explored as a potential therapeutic mechanism to inhibit the progression of breast cancer. Virtual screening using molecular docking resulted in the three hit compounds (ZINC08382411, ZINC08382438, and ZINC08382292) with minimum binding scores and commonly binding to both receptors. Further, MD simulation analysis of these complexes illustrated the high stability of these compounds with EGFR and HER2. RMSD showed that ZINC08382411 displayed the most stable RMSD of 2 - 3 Å when bound to both receptors, suggesting to have strong compatibility with the active site of the receptor. Hydrogen bond analysis showed that ZINC08382411 forms the maximum number of H-bonds (2 to 3) in both EGFR and HER2 bound complexes, with the highest occupancy of 62% and 79%, respectively. Binding free energy calculation showed that ZINC08382411 possesses maximum affinity towards both the receptors with ΔGbind = -129.628 and -164.063 kJ/mol, respectively. This approach recognizes the significance of EGFR and HER2 in breast cancer development and aims to disrupt their collaborative signaling, which is known to promote the antagonistic behavior of cancer cells. By focusing on this EGFR/HER2 heterodimer, the study offers a promising avenue for identifying a potential candidate (ZINC08382411) that may inhibit breast cancer cell growth and potentially improve patient outcomes. The study's findings may contribute to the ongoing efforts to advance breast cancer treatment strategies.Communicated by Ramaswamy H. Sarma.