RESUMO
Estrogen receptor-positive (ER+) breast cancers frequently remain dependent on ER signaling even after acquiring resistance to endocrine agents, prompting the development of optimized ER antagonists. Fulvestrant is unique among approved ER therapeutics due to its capacity for full ER antagonism, thought to be achieved through ER degradation. The clinical potential of fulvestrant is limited by poor physicochemical features, spurring attempts to generate ER degraders with improved drug-like properties. We show that optimization of ER degradation does not guarantee full ER antagonism in breast cancer cells; ER "degraders" exhibit a spectrum of transcriptional activities and anti-proliferative potential. Mechanistically, we find that fulvestrant-like antagonists suppress ER transcriptional activity not by ER elimination, but by markedly slowing the intra-nuclear mobility of ER. Increased ER turnover occurs as a consequence of ER immobilization. These findings provide proof-of-concept that small molecule perturbation of transcription factor mobility may enable therapeutic targeting of this challenging target class.
Assuntos
Neoplasias da Mama/metabolismo , Antagonistas do Receptor de Estrogênio/farmacologia , Fulvestranto/farmacologia , Receptores de Estrogênio/antagonistas & inibidores , Receptores de Estrogênio/metabolismo , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , Cinamatos/farmacologia , Resistencia a Medicamentos Antineoplásicos , Antagonistas do Receptor de Estrogênio/uso terapêutico , Feminino , Fulvestranto/uso terapêutico , Células HEK293 , Xenoenxertos , Humanos , Indazóis/farmacologia , Ligantes , Células MCF-7 , Camundongos , Camundongos Endogâmicos NOD , Camundongos Nus , Camundongos SCID , Polimorfismo de Nucleotídeo Único , Proteólise/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacosRESUMO
Fulvestrant is an FDA-approved drug with a dual mechanism of action (MOA), acting as a full antagonist and degrader of the estrogen receptor protein. A significant limitation of fulvestrant is the dosing regimen required for efficacy. Due to its high lipophilicity and poor pharmacokinetic profile, fulvestrant needs to be administered through intramuscular injections which leads to injection site soreness. This route of administration also limits the dose and target occupancy in patients. We envisioned a best-in-class molecule that would function with the same dual MOA as fulvestrant, but with improved physicochemical properties and would be orally bioavailable. Herein we report our progress toward that goal, resulting in a new lead GNE-502 which addressed some of the liabilities of our previously reported lead molecule GNE-149.
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Antineoplásicos/farmacologia , Antineoplásicos/farmacocinética , Neoplasias da Mama/tratamento farmacológico , Descoberta de Drogas , Receptores de Estrogênio/metabolismo , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Relação Dose-Resposta a Droga , Feminino , Humanos , Células MCF-7 , Camundongos , Estrutura Molecular , Conformação Proteica , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: The purpose of the presented investigation is to evaluate the resulting torque on loaded 3D printed springs using different coil thickness and length. METHODS: Specimens were designed and printed using the 3D printer MAX (Asiga, Sydney, Australia) with 3D printable, experimental, flexible material (Code:BM2008, GC, Tokyo, Japan). The specimens were divided into three groups according to spring coil design. Control group (n = 18), length group (n = 19) and thickness group (n = 22). Groups were tested using a Sauter Machine for torque calculation (DB, Grindelwald, Switzerland) in conjunction with a universal testing machine (Zwick Z010, Ulm, Germany) for clock-wise and anti-clockwise testing. Statistical analysis was performed using the Steel-Dwass test to compare median values of the three groups in both testing directions (p < 0.001). RESULTS: The highest torque value was determined in the thickness group for both clockwise and anti-clockwise testing directions, achieving 44.00 N/mm and 39.62 N/mm respectively. The length group ranged from 21.65 to 11.04 N/mm in clockwise direction and from 18.04 to 11.38 N/mm in counter-clockwise testing. The control group ranged from 22.72 to 17.18 N/mm in the clock-wise direction while in the anti-clock wise testing it ranged from 21.34 to 16.02 N/mm. CONCLUSIONS: The amount of torque produced from the computer aided designing/computer aided manufacturing (CAD/CAM) springs is being affected by diameter more than the length design parameter in comparison to the control group. The values of the thickness group are significantly higher than those of the length group (P < 0.001).
Assuntos
Desenho Assistido por Computador , Impressão Tridimensional , Austrália , Estudos de Viabilidade , Alemanha , Humanos , Japão , Teste de Materiais , Suíça , TorqueRESUMO
Chimeric molecules which effect intracellular degradation of target proteins via E3 ligase-mediated ubiquitination (e.g., PROTACs) are currently of high interest in medicinal chemistry. However, these entities are relatively large compounds that often possess molecular characteristics which may compromise oral bioavailability, solubility, and/or in vivo pharmacokinetic properties. Accordingly, we explored whether conjugation of chimeric degraders to monoclonal antibodies using technologies originally developed for cytotoxic payloads might provide alternate delivery options for these novel agents. In this report we describe the construction of several degrader-antibody conjugates comprised of two distinct ERα-targeting degrader entities and three independent ADC linker modalities. We subsequently demonstrate the antigen-dependent delivery to MCF7-neo/HER2 cells of the degrader payloads that are incorporated into these conjugates. We also provide evidence for efficient intracellular degrader release from one of the employed linkers. In addition, preliminary data are described which suggest that reasonably favorable in vivo stability properties are associated with the linkers utilized to construct the degrader conjugates.
Assuntos
Anticorpos Monoclonais/imunologia , Portadores de Fármacos/química , Receptor alfa de Estrogênio/imunologia , Anticorpos Monoclonais/química , Antineoplásicos/química , Antineoplásicos/imunologia , Antineoplásicos/farmacologia , Desenho de Fármacos , Receptor alfa de Estrogênio/metabolismo , Humanos , Imunoconjugados/química , Imunoconjugados/imunologia , Imunoconjugados/farmacologia , Células MCF-7 , Proteólise/efeitos dos fármacos , Receptor ErbB-2/metabolismoRESUMO
Despite tremendous progress made in the understanding of the ERα signaling pathway and the approval of many therapeutic agents, ER+â¯breast cancer continues to be a leading cause of cancer death in women. We set out to discover compounds with a dual mechanism of action in which they not only compete with estradiol for binding with ERα, but also can induce the degradation of the ERα protein itself. We were attracted to the constrained chromenes containing a tetracyclic benzopyranobenzoxepine scaffold, which were reported as potent selective estrogen receptor modulators (SERMs). Incorporation of a fluoromethyl azetidine side chain yielded highly potent and efficacious selective estrogen receptor degraders (SERDs), such as 16aa and surprisingly, also its enantiomeric pair 16ab. Co-crystal structures of the enantiomeric pair 16aa and 16ab in complex with ERα revealed default (mimics the A-D rings of endogenous ligand estradiol) and core-flipped binding modes, rationalizing the equivalent potency observed for these enantiomers in the ERα degradation and MCF-7 anti-proliferation assays.
Assuntos
Antineoplásicos/farmacologia , Benzopiranos/farmacologia , Receptor alfa de Estrogênio/química , Antineoplásicos/química , Benzopiranos/química , Cristalização , Humanos , Células MCF-7 , Modelos Moleculares , Estrutura Molecular , Conformação Proteica , Transdução de Sinais , Relação Estrutura-AtividadeRESUMO
Phenolic groups are responsible for the high clearance and low oral bioavailability of the estrogen receptor alpha (ERα) clinical candidate GDC-0927. An exhaustive search for a backup molecule with improved pharmacokinetic (PK) properties identified several metabolically stable analogs, although in general at the expense of the desired potency and degradation efficiency. C-8 hydroxychromene 30 is the first example of a phenol-containing chromene that not only maintained excellent potency but also exhibited 10-fold higher oral exposure in rats. The improved in vivo clearance in rat was hypothesized to be the result of C-8 hydroxy group being sterically protected from glucuronide conjugation. The excellent potency underscores the possibility of replacing the presumed indispensable phenolic group at C-6 or C-7 of the chromene core. Co-crystal structures were obtained to highlight the change in key interactions and rationalize the retained potency.
Assuntos
Azetidinas/farmacologia , Receptor alfa de Estrogênio/metabolismo , Flavonoides/farmacologia , Administração Oral , Animais , Azetidinas/administração & dosagem , Azetidinas/metabolismo , Azetidinas/farmacocinética , Cristalografia por Raios X , Descoberta de Drogas , Estabilidade de Medicamentos , Flavonoides/administração & dosagem , Flavonoides/metabolismo , Flavonoides/farmacocinética , Humanos , Células MCF-7 , Microssomos Hepáticos/metabolismo , Ratos , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
A novel strategy to attach indole-containing payloads to antibodies through a carbamate moiety and a self-immolating, disulfide-based linker is described. This new strategy was employed to connect a selective estrogen receptor down-regulator (SERD) to various antibodies in a site-selective manner. The resulting conjugates displayed potent, antigen-dependent down-regulation of estrogen receptor levels in MCF7-neo/HER2 and MCF7-hB7H4 cells. They also exhibited similar antigen-dependent modulation of the estrogen receptor in tumors when administered intravenously to mice bearing MCF7-neo/HER2 tumor xenografts. The indole-carbamate moiety present in the new linker was stable in whole blood from various species and also exhibited good inâ vivo stability properties in mice.
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Indóis/química , Animais , Anticorpos Monoclonais/química , Linhagem Celular Tumoral , Humanos , Imunoconjugados/administração & dosagem , Células MCF-7 , CamundongosRESUMO
Multiferroic materials with simultaneous magnetic and ferroelectric ordering that persist above room temperature are rare. Using first-principles density functional theory calculations, we demonstrate fluorination of oxygen-deficient AA'Fe2O5 perovskites, where A and A' are cations with +3 and +2 oxidation states, respectively, and have a layered ordering, as an effective strategy to obtain room-temperature multiferroics. We show that by controlling the size of the A and A' cations, it is possible to stabilize a noncentrosymmetric phase arising due to the hybrid improper ferroelectricity mechanism, with polarization as high as 13 µC/cm2. The fluorination also stabilizes Fe in +3 oxidation state, which results in superexchange interactions that are strong enough to sustain magnetic order well above room temperature. We also show the presence of a magnetoelectric coupling wherein the switching mode that reverses the direction of the spontaneous polarization also affects the strength of the magnetic interactions. The results show that low-temperature fluorination of anion-deficient perovskites with layered cation ordering can be an effective approach to design new multiferroics.
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BACKGROUND: Globally, stunting affects â¼150 million children under five, while wasting affects nearly 50 million. Current interventions have had limited effectiveness in ameliorating long-term sequelae of undernutrition including stunting, cognitive deficits and immune dysfunction. Disrupted development of the gut microbiota has been linked to the pathogenesis of undernutrition, providing potentially new treatment approaches. METHODS: 124 Bangladeshi children with moderate acute malnutrition (MAM) enrolled (at 12-18 months) in a previously reported 3-month RCT of a microbiota-directed complementary food (MDCF-2) were followed for two years. Weight and length were monitored by anthropometry, the abundances of bacterial strains were assessed by quantifying metagenome-assembled genomes (MAGs) in serially collected fecal samples and levels of growth-associated proteins were measured in plasma. FINDINGS: Children who had received MDCF-2 were significantly less stunted during follow-up than those who received a standard ready-to-use supplementary food (RUSF) [linear mixed-effects model, ßtreatment group x study week (95% CI) = 0.002 (0.001, 0.003); P = 0.004]. They also had elevated fecal abundances of Agathobacter faecis, Blautia massiliensis, Lachnospira and Dialister, plus increased levels of a group of 37 plasma proteins (linear model; FDR-adjusted P < 0.1), including IGF-1, neurotrophin receptor NTRK2 and multiple proteins linked to musculoskeletal and CNS development, that persisted for 6-months post-intervention. INTERPRETATION: MDCF-2 treatment of Bangladeshi children with MAM, which produced significant improvements in wasting during intervention, also reduced stunting during follow-up. These results suggest that the effectiveness of supplementary foods for undernutrition may be improved by including ingredients that sponsor healthy microbiota-host co-development. FUNDING: This work was supported by the BMGF (Grants OPP1134649/INV-000247).
Assuntos
Microbioma Gastrointestinal , Humanos , Lactente , Feminino , Masculino , Bangladesh/epidemiologia , Fezes/microbiologia , Metagenoma , Transtornos do Crescimento/etiologiaRESUMO
Severe acute malnutrition (SAM), defined anthropometrically as a weight-for-length z-score more than 3 standard deviations below the mean (WLZ<-3), affects 19 million children under 5-years-old worldwide. Complete anthropometric recovery after standard inventions is rare with children often left with moderate acute malnutrition (MAM; WLZ -2 to -3). Here we conduct a randomized controlled trial (RCT), involving 12-18-month-old Bangladeshi children from urban and rural sites, who after hospital-based treatment for SAM received a 3-month intervention with a microbiota-directed complementary food (MDCF-2) or a ready-to-use supplementary food (RUSF) as they transitioned to MAM. The rate of WLZ improvement was significantly greater with MDCF-2 than the more calorically-dense RUSF, as we observed in a previous RCT of Bangladeshi children with MAM without antecedent SAM. A correlated meta-analysis of aptamer-based measurements of 4,520 plasma proteins in this and the prior RCT revealed 215 proteins positively-associated with WLZ (prominently those involved in musculoskeletal and CNS development) and 44 negatively-associated proteins (related to immune activation), with a significant enrichment in levels of the positively WLZ-associated proteins in the MDCF-2 arm. Characterizing changes in 754 bacterial metagenome-assembled genomes in serially collected fecal samples disclosed the effects of acute rehabilitation for SAM on the microbiome, its transition as each child achieves a state of MAM, and how specific strains of Prevotella copri function at the intersection between MDCF-2 glycan metabolism and the rescue of growth faltering. These results provide a rationale for further testing the generalizability of the efficacy of MDCF and identify biomarkers for defining treatment responses.
RESUMO
Developments in the field of nanoplasmonics have the potential to advance applications from information processing and telecommunications to light-based sensing. Traditionally, nanoscale noble metals such as gold and silver have been used to achieve the targeted enhancements in light-matter interactions that result from the presence of localized surface plasmons (LSPs). However, interest has recently shifted to intrinsically doped semiconductor nanocrystals (NCs) for their ability to display LSP resonances (LSPRs) over a much broader spectral range, including the infrared (IR). Among semiconducting plasmonic NCs, spinel metal oxides (sp-MOs) are an emerging class of materials with distinct advantages in accessing the telecommunications bands in the IR and affording useful environmental stability. Here, we report the plasmonic properties of Fe3O4 sp-MO NCs, known previously only for their magnetic functionality, and demonstrate their ability to modify the light-emission properties of telecom-emitting quantum dots (QDs). We establish the synthetic conditions for tuning sp-MO NC size, composition and doping characteristics, resulting in unprecedented tunability of electronic behavior and plasmonic response over 450 nm. In particular, with diameter-dependent variations in free-electron concentration across the Fe3O4 NC series, we introduce a strong NC size dependency onto the optical response. In addition, our observation of plasmonics-enhanced decay rates from telecom-emitting QDs reveals Purcell enhancement factors for simple plasmonic-spacer-emitter sandwich structures up to 51-fold, which are comparable to values achieved previously only for emitters in the visible range coupled with conventional noble metal NCs.
RESUMO
A 53-year-old male with no significant past medical history presented with an acute traumatic fracture of his thumb. Preoperative chest radiograph before K-wire fixation demonstrated an incidental 9 cm opacity of the left lung. Chest computed tomography revealed a 6.3 cm aggressive appearing pleural-based mass with erosion and destruction of the underlying rib. The patient underwent percutaneous biopsy with interventional radiology, and pathology revealed a small round blue cell tumor with positive CD99 staining and a FUS-ERG chromosomal translocation. The patient was diagnosed with Askin tumor, a peripheral primitive neuroectodermal tumor of the thoracopulmonary region belonging to the Ewing sarcoma tumor family. Computed tomography and magnetic resonance imaging of Askin tumors may show features such as a heterogeneous soft tissue mass, pleural effusion, rib destruction, hemorrhage, necrosis, and cystic degeneration. Askin tumors typically exhibit the EWS-FLI1 fusion mutation, although FUS-ERG chromosomal translocation has been described. Both rarity and variability of Askin tumors present a diagnostic challenge for clinicians. Collaborative effort amongst radiologists and pathologists is essential for diagnosis.
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The biological and medicinal impacts of proteolysis-targeting chimeras (PROTACs) and related chimeric molecules that effect intracellular degradation of target proteins via ubiquitin ligase-mediated ubiquitination continue to grow. However, these chimeric entities are relatively large compounds that often possess molecular characteristics, which may compromise oral bioavailability, solubility, and/or in vivo pharmacokinetic properties. We therefore explored the conjugation of such molecules to monoclonal antibodies using technologies originally developed for cytotoxic payloads so as to provide alternate delivery options for these novel agents. In this report, we describe the first phase of our systematic development of antibody-drug conjugates (ADCs) derived from bromodomain-containing protein 4 (BRD4)-targeting chimeric degrader entities. We demonstrate the antigen-dependent delivery of the degrader payloads to PC3-S1 prostate cancer cells along with related impacts on MYC transcription and intracellular BRD4 levels. These experiments culminate with the identification of one degrader conjugate, which exhibits antigen-dependent antiproliferation effects in LNCaP prostate cancer cells.
Assuntos
Proteínas de Ciclo Celular/antagonistas & inibidores , Dipeptídeos/farmacologia , Compostos Heterocíclicos com 3 Anéis/farmacologia , Imunoconjugados/farmacologia , Proteólise/efeitos dos fármacos , Fatores de Transcrição/antagonistas & inibidores , Anticorpos Monoclonais/imunologia , Antígenos de Neoplasias/imunologia , Proteínas de Ciclo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Dipeptídeos/síntese química , Dipeptídeos/farmacocinética , Compostos Heterocíclicos com 3 Anéis/síntese química , Compostos Heterocíclicos com 3 Anéis/farmacocinética , Humanos , Imunoconjugados/química , Imunoconjugados/imunologia , Oxirredutases/imunologia , Células PC-3 , Fatores de Transcrição/metabolismo , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismoRESUMO
Breast cancer remains a leading cause of cancer death in women, representing a significant unmet medical need. Here, we disclose our discovery efforts culminating in a clinical candidate, 35 (GDC-9545 or giredestrant). 35 is an efficient and potent selective estrogen receptor degrader (SERD) and a full antagonist, which translates into better antiproliferation activity than known SERDs (1, 6, 7, and 9) across multiple cell lines. Fine-tuning the physiochemical properties enabled once daily oral dosing of 35 in preclinical species and humans. 35 exhibits low drug-drug interaction liability and demonstrates excellent in vitro and in vivo safety profiles. At low doses, 35 induces tumor regressions either as a single agent or in combination with a CDK4/6 inhibitor in an ESR1Y537S mutant PDX or a wild-type ERα tumor model. Currently, 35 is being evaluated in Phase III clinical trials.
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Antineoplásicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Carbolinas/uso terapêutico , Antagonistas do Receptor de Estrogênio/uso terapêutico , Receptor alfa de Estrogênio/metabolismo , Animais , Antineoplásicos/química , Antineoplásicos/farmacocinética , Carbolinas/química , Carbolinas/farmacocinética , Cães , Antagonistas do Receptor de Estrogênio/química , Antagonistas do Receptor de Estrogênio/farmacocinética , Feminino , Humanos , Células MCF-7 , Macaca fascicularis , Camundongos , Estrutura Molecular , Ratos , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Heterobifunctional compounds that direct the ubiquitination of intracellular proteins in a targeted manner via co-opted ubiquitin ligases have enormous potential to transform the field of medicinal chemistry. These chimeric molecules, often termed proteolysis-targeting chimeras (PROTACs) in the chemical literature, enable the controlled degradation of specific proteins via their direction to the cellular proteasome. In this report, we describe the second phase of our research focused on exploring antibody-drug conjugates (ADCs), which incorporate BRD4-targeting chimeric degrader entities. We employ a new BRD4-binding fragment in the construction of the chimeric ADC payloads that is significantly more potent than the corresponding entity utilized in our initial studies. The resulting BRD4-degrader antibody conjugates exhibit potent and antigen-dependent BRD4 degradation and antiproliferation activities in cell-based experiments. Multiple ADCs bearing chimeric BRD4-degrader payloads also exhibit strong, antigen-dependent antitumor efficacy in mouse xenograft assessments that employ several different tumor models.
Assuntos
Antineoplásicos/uso terapêutico , Proteínas de Ciclo Celular/antagonistas & inibidores , Proliferação de Células/efeitos dos fármacos , Imunoconjugados/uso terapêutico , Neoplasias/tratamento farmacológico , Proteólise/efeitos dos fármacos , Fatores de Transcrição/antagonistas & inibidores , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais/uso terapêutico , Antígenos de Neoplasias/imunologia , Antineoplásicos/síntese química , Antineoplásicos/farmacocinética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Dipeptídeos/síntese química , Dipeptídeos/farmacocinética , Dipeptídeos/uso terapêutico , Feminino , Compostos Heterocíclicos com 3 Anéis/síntese química , Compostos Heterocíclicos com 3 Anéis/farmacocinética , Compostos Heterocíclicos com 3 Anéis/uso terapêutico , Humanos , Imunoconjugados/imunologia , Imunoconjugados/farmacocinética , Camundongos SCID , Oxirredutases/imunologia , Fatores de Transcrição/metabolismo , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
It is well known that in the high-temperature superconductor YBa2Cu3O7-x (YBCO), oxygen vacancies (VO) control the carrier concentration, its critical current density and transition temperature. In this work, it is revealed that VO also allows the accommodation of local strain fields caused by large-scale defects within the crystal. We show that the nanoscale strain associated with Y2Ba4Cu8O16 (Y124) intergrowths-that are common defects in YBCO-strongly affect the venue and concentration of VO. Local probe measurements in conjunction with density-functional-theory calculations indicate a strain-driven reordering of VO from the commonly observed CuO chains towards the bridging apical sites located in the BaO plane and bind directly to the superconducting CuO2 planes. Our findings have strong implications on the physical properties of the YBCO, as the presence of apical VO alters the transfer of carriers to the CuO2 planes, confirmed by changes in the Cu and O core-loss edge probed using electron energy loss spectroscopy, and creates structural changes that affect the Cu-O bonds in the superconducting planes. In addition, the revelation of apical VO also has implications on modulating critical current densities and enhancing vortex pinning.
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Estrogen receptor alpha (ERα) is a well-validated drug target for ER-positive (ER+) breast cancer. Fulvestrant is FDA-approved to treat ER+ breast cancer and works through two mechanisms-as a full antagonist and selective estrogen receptor degrader (SERD)-but lacks oral bioavailability. Thus, we envisioned a "best-in-class" molecule with the same dual mechanisms as fulvestrant, but with significant oral exposure. Through lead optimization, we discovered a tool molecule 12 (GNE-149) with improved degradation and antiproliferative activity in both MCF7 and T47D cells. To illustrate the binding mode and key interactions of this scaffold with ERα, we obtained a cocrystal structure of 6 that showed ionic interaction of azetidine with Asp351 residue. Importantly, 12 showed favorable metabolic stability and good oral exposure. 12 exhibited antagonist effect in the uterus and demonstrated robust dose-dependent efficacy in xenograft models.
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AIMS: Peroxiredoxins (Prxs) are ubiquitous cysteine-based peroxidases involved in oxidant defense and signal transduction. Despite much study, the precise roles of conserved residues remain poorly defined. In this study, we carried out extensive functional and structural characterization of 10 variants of such residues in a model decameric bacterial Prx. RESULTS: Three active site proximal mutations of Salmonella typhimurium AhpC, T43V, R119A, and E49Q, lowered catalytic efficiency with hydrogen peroxide by 4-5 orders of magnitude, but did not affect reactivity toward their reductant, AhpF. pKa values of the peroxidatic cysteine were also shifted up by 1-1.3 pH units for these and a decamer disruption mutant, T77I. Except for the decamer-stabilizing T77V, all mutations destabilized decamers in the reduced form. In the oxidized form, three mutants-T77V, T43A, and T43S-exhibited stabilized decamers and were more efficiently reduced by AhpF than wild-type AhpC. Crystal structures of most mutants were solved and many showed alterations in stability of the fully folded active site loop. INNOVATION: This is the first study of Prx mutants to comprehensively assess the effects of mutations on catalytic activities, the active site cysteine pKa, and the protein structure and oligomeric status. CONCLUSION: The Arg119 side chain must be properly situated for efficient catalysis, but for other debilitating variants, the functional defects could be explained by structural perturbations and/or associated decamer destabilization rather than direct effects. This underscores the importance of our comprehensive approach. A remarkable new finding was the preference of the reductant for decamers. Antioxid. Redox Signal. 28, 521-536.
Assuntos
Catálise , Peróxido de Hidrogênio/química , Peroxidases/química , Peroxirredoxinas/química , Sequência de Aminoácidos/genética , Domínio Catalítico , Cristalografia por Raios X , Cisteína/química , Peróxido de Hidrogênio/metabolismo , Cinética , Modelos Moleculares , Mutação , Oxirredução , Peroxidases/genética , Peroxidases/metabolismo , Peroxirredoxinas/genética , Peroxirredoxinas/metabolismo , Salmonella typhimurium/enzimologia , Salmonella typhimurium/genéticaRESUMO
Purpose Therapies with novel mechanisms of action are needed for multiple myeloma (MM). T cells can be genetically modified to express chimeric antigen receptors (CARs), which are artificial proteins that target T cells to antigens. B-cell maturation antigen (BCMA) is expressed by normal and malignant plasma cells but not normal essential cells. We conducted the first-in-humans clinical trial, to our knowledge, of T cells expressing a CAR targeting BCMA (CAR-BCMA). Patients and Methods Sixteen patients received 9 × 106 CAR-BCMA T cells/kg at the highest dose level of the trial; we are reporting results of these 16 patients. The patients had a median of 9.5 prior lines of MM therapy. Sixty-three percent of patients had MM refractory to the last treatment regimen before protocol enrollment. T cells were transduced with a γ-retroviral vector encoding CAR-BCMA. Patients received CAR-BCMA T cells after a conditioning chemotherapy regimen of cyclophosphamide and fludarabine. Results The overall response rate was 81%, with 63% very good partial response or complete response. Median event-free survival was 31 weeks. Responses included eradication of extensive bone marrow myeloma and resolution of soft-tissue plasmacytomas. All 11 patients who obtained an anti-MM response of partial response or better and had MM evaluable for minimal residual disease obtained bone marrow minimal residual disease-negative status. High peak blood CAR+ cell levels were associated with anti-MM responses. Cytokine-release syndrome toxicities were severe in some cases but were reversible. Blood CAR-BCMA T cells were predominantly highly differentiated CD8+ T cells 6 to 9 days after infusion. BCMA antigen loss from MM was observed. Conclusion CAR-BCMA T cells had substantial activity against heavily treated relapsed/refractory MM. Our results should encourage additional development of CAR T-cell therapies for MM.
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Antígeno de Maturação de Linfócitos B/imunologia , Imunoterapia Adotiva/métodos , Mieloma Múltiplo/terapia , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos Quiméricos/imunologia , Linfócitos T/transplante , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Antígeno de Maturação de Linfócitos B/genética , Ciclofosfamida/administração & dosagem , Citocinas/sangue , Citocinas/imunologia , Humanos , Mieloma Múltiplo/sangue , Mieloma Múltiplo/imunologia , Prognóstico , Receptores de Antígenos Quiméricos/sangue , Linfócitos T/imunologia , Condicionamento Pré-Transplante , Vidarabina/administração & dosagem , Vidarabina/análogos & derivadosRESUMO
Zika virus (ZIKV) has been of major international public health concern following large outbreaks in the Americas occurring in 2015-2016. Most notably, ZIKV has been seen to pose dangers in pregnancy due to its association with congenital abnormalities such as microcephaly. Numerous experimental approaches have been taken to address how the virus can cross the placenta, alter normal fetal development, and disrupt specific cellular functions. Many areas concerning the mechanisms of transmission, especially from mother to fetus, are largely unknown but demand further research. Several promising new studies are presented that provide insight into possible mechanisms of transmission, different cell types affected, and immune responses towards the virus. By aiming to better understand the processes behind altered fetal neuronal development due to ZIKV infection, the hope is to find ways to increase protection of the fetus and prevent congenital abnormalities such as microcephaly. As ZIKV infection is spreading to increasingly more areas and bringing harmful outcomes and birth defects with it, it is imperative to identify the mechanisms of transmitting this infectious agent, consider different genetic backgrounds of hosts and strain types, and navigate methods to protect those affected from the detrimental effects of this newly emerging virus.