RESUMO
Few approaches have been made toward exploring autologous NK cells in settings of cancer immunotherapy. Here, we demonstrate the feasibility of infusing multiple doses of ex vivo activated and expanded autologous NK cells in patients with multiple myeloma (MM) post-autologous stem cell transplantation. Infused NK cells were detected in circulation up to 4 weeks after the last infusion. Elevations in plasma granzyme B levels were observed following each consecutive NK cell infusion. Moreover, increased granzyme B levels were detected in bone marrow 4 weeks after the last infusion. All measurable patients had objective, detectable responses after NK cell infusions in terms of reduction in M-component and/or minimal residual disease. The present study demonstrates that autologous NK cell-based immunotherapy is feasible in a setting of MM consolidation therapy. It opens up the possibility for usage of autologous NK cells in clinical settings where patients are not readily eligible for allogeneic NK cell-based immunotherapies.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Mieloma Múltiplo , Quimioterapia de Consolidação , Granzimas , Humanos , Células Matadoras Naturais , Mieloma Múltiplo/terapia , Transplante de Células-Tronco , Transplante AutólogoRESUMO
Atherosclerotic cardiovascular disease is a major problem despite the availability of drugs that influence major risk factors. New treatments are needed, and there is growing interest in therapies that may have multiple actions. Thyroid hormone modulates several cardiovascular risk factors and delays atherosclerosis progression in humans. However, use of thyroid hormone is limited by side effects, especially in the heart. To overcome this limitation, pharmacologically selective thyromimetics that mimic metabolic effects of thyroid hormone and bypass side effects are under development. In animal models, such thyromimetics have been shown to stimulate cholesterol elimination through LDL and HDL pathways and decrease body weight without eliciting side effects. We report here studies on a selective thyromimetic [KB2115; (3-[[3,5-dibromo-4-[4-hydroxy-3-(1-methylethyl)-phenoxy]-phenyl]-amino]-3-oxopropanoic acid)] in humans. In moderately overweight and hypercholesterolemic subjects KB2115 was found to be safe and well tolerated and elicited up to a 40% lowering of total and LDL cholesterol after 14 days of treatment. Bile acid synthesis was stimulated without evidence of increased cholesterol production, indicating that KB2115 induced net cholesterol excretion. KB2115 did not provoke detectable effects on the heart, suggesting that the pharmacological selectivity observed in animal models translates to humans. Thus, selective thyromimetics deserve further study as agents to treat dyslipidemia and other risk factors for atherosclerosis.
Assuntos
Anilidas/farmacologia , Ácidos e Sais Biliares/metabolismo , LDL-Colesterol/metabolismo , Coração/efeitos dos fármacos , Glândula Tireoide/efeitos dos fármacos , Hormônios Tireóideos/metabolismo , Adolescente , Adulto , Anilidas/química , Método Duplo-Cego , Feminino , Humanos , Metabolismo dos Lipídeos , Masculino , Pessoa de Meia-Idade , Placebos , Fatores de Risco , Tiazolidinedionas/farmacologiaRESUMO
Obesity and metabolic syndrome are increasing dramatically worldwide, contributing to cardiovascular morbidity and mortality. There are currently few safe and efficacious therapeutics for obesity and most strategies are focused on appetite suppression. Thyroid hormones reduce adiposity via increased metabolic rate, but unfortunately they cause large changes in metabolic rate and direct cardiac acceleration, making them useless for treating obesity. Thyroid hormone receptors (TRs) work as transcription factors and two subtypes exist: TRalpha and TRbeta. TRalpha mediates tachycardia and much of the metabolic rate effect, while TRbeta mediates cholesterol and TSH lowering effects of thyroid hormones. TRbeta activation modestly increases metabolic rate such that a therapeutic window of 5-10 fold increases in metabolic rate can be seen without tachycardia. This was initially studied in TRalpha(1)(-/-) mice. Recent structure activity work has resulted in the discovery of several TRbeta selective thyromimetics such as KB-141. Studies with KB-141 show that it has a 10-fold window in which therapeutic increases in metabolic rate are seen without tachycardia or cardiac hypertrophy. This agent lowers cholesterol in rats and primates. In primates, KB-141 causes significant weight and cholesterol reduction in addition to the independent risk factor Lp(a). These effects were seen without any effect on heart rate, unlike thyroid hormone (T(3)). Further work with TRbeta selective agents is warranted and recent work suggests the possibility of developing compounds that selectively penetrate different tissues which may have an even more desirable therapeutic window. Selective thyromimetics, therefore, may be useful as adjunctive therapy to appetite suppressants along with exercise and diet restriction.
Assuntos
Diabetes Mellitus/tratamento farmacológico , Hiperlipidemias/tratamento farmacológico , Obesidade/tratamento farmacológico , Receptores beta dos Hormônios Tireóideos/agonistas , Animais , Fármacos Antiobesidade/farmacologia , Humanos , Hipoglicemiantes/farmacologia , Hipolipemiantes/farmacologia , Camundongos , Camundongos Knockout , Éteres Fenílicos/farmacologia , Fenilacetatos/farmacologia , Primatas , Ratos , Doenças da Glândula Tireoide/fisiopatologia , Receptores beta dos Hormônios Tireóideos/genética , Hormônios Tireóideos/fisiologiaRESUMO
Antagonists have been developed for several nuclear receptors but not for others, including TRs. TR antagonists may have significant clinical utility for treating hormone excess states and other conditions. A structure derived "extension hypothesis" was applied to synthesize a TR antagonist. The principal design feature was to attach an extension group to a TR agonist whose structure would perturb formation of the TR coactivator-binding surface. The compound, 3,5-dibromo-4-(3',5'-diisopropyl-4'-hydroxyphenoxy)benzoic acid, has no (TRalpha) or very weak partial (TRbeta) TR agonist activity and blocks TR binding of T3, formation of the coactivator-binding surface, and both a positive T3 response on a thyroid hormone response element and a negative T3 response on the TSHbeta promoter in cultured cells. The results suggest that 3,5-dibromo-4-(3',5'-diisopropyl-4'-hydroxyphenoxy)benzoic acid is a TR antagonist for thyroid hormone response element-mediated responses, this approach can be used more generally to generate nuclear receptor antagonists, and this compound or analogues may have medical and research utility.
Assuntos
Benzoatos/síntese química , Benzoatos/farmacologia , Antagonistas de Hormônios/síntese química , Antagonistas de Hormônios/farmacologia , Receptores dos Hormônios Tireóideos/antagonistas & inibidores , Fosfatase Alcalina/metabolismo , Animais , Bromobenzoatos , Células CHO , Linhagem Celular , Clonagem Molecular , Cricetinae , Cristalografia , Desenho de Fármacos , Eletroforese em Gel de Poliacrilamida , Genes Reporter , Vetores Genéticos , Humanos , Éteres Fenílicos , Placenta/enzimologia , Receptores dos Hormônios Tireóideos/química , Receptores dos Hormônios Tireóideos/genética , Relação Estrutura-Atividade , TransfecçãoRESUMO
Endogenous thyroid receptor hormones 3,5,3',5'-tetraiodo-l-thyronine (T(4), 1) and 3,5,3'-triiodo-l-thyronine (T(3), 2) exert a significant effects on growth, development, and homeostasis in mammals. They regulate important genes in intestinal, skeletal, and cardiac muscles, the liver, and the central nervous system, influence overall metabolic rate, cholesterol and triglyceride levels, and heart rate, and affect mood and overall sense of well being. The literature suggests many or most effects of thyroid hormones on the heart, in particular on the heart rate and rhythm, are mediated through the TRalpha(1) isoform, while most actions of the hormones on the liver and other tissues are mediated more through the TRbeta(1) isoform of the receptor. Some effects of thyroid hormones may be therapeutically useful in nonthyroid disorders if adverse effects can be minimized or eliminated. These potentially useful features include weight reduction for the treatment of obesity, cholesterol lowering for treating hyperlipidemia, amelioration of depression, and stimulation of bone formation in osteoporosis. Prior attempts to utilize thyroid hormones pharmacologically to treat these disorders have been limited by manifestations of hyperthyroidism and, in particular, cardiovascular toxicity. Consequently, development of thyroid hormone receptor agonists that are selective for the beta-isoform could lead to safe therapies for these common disorders while avoiding cardiotoxicity. We describe here the synthesis and evaluation of a series of novel TR ligands, which are selective for TRbeta(1) over TRalpha(1). These ligands could potentially be useful for treatment of various disorders as outlined above. From a series of homologous R(1)-substituted carboxylic acid derivatives, increasing chain length was found to have a profound effect on affinity and selectivity in a radioreceptor binding assay for the human thyroid hormone receptors alpha(1) and beta(1) (TRalpha(1) and TRbeta(2)) as well as a reporter cell assay employing CHOK1-cells (Chinese hamster ovary cells) stably transfected with hTRalpha(1) or hTRbeta(1) and an alkaline phosphatase reporter-gene downstream thyroid response element (TRAFalpha(1) and TRAFbeta(1)). Affinity increases in the order formic, acetic, and propionic acid, while beta-selectivity is highest when the R(1) position is substituted with acetic acid. Within this series 3,5-dibromo-4-[(4-hydroxy-3-isopropylphenoxy)phenyl]acetic acid (11a) and 3,5-dichloro-4-[(4-hydroxy-3-isopropylphenoxy)phenyl]acetic acid (15) were found to reveal the most promising in vitro data based on isoform selectivity and were selected for further in vivo studies. The effect of 2, 11a, and 15 in a cholesterol-fed rat model was monitored including potencies for heart rate (ED(15)), cholesterol (ED(50)), and TSH (ED(50)). Potency for tachycardia was significantly reduced for the TRbeta selective compounds 11a and 15 compared with 2, while both 11a and 15 retained the cholesterol-lowering potency of 2. This left an approximately 10-fold therapeutic window between heart rate and cholesterol, which is consistent with the action of ligands that are approximately 10-fold more selective for TRbeta(1). We also report the X-ray crystallographic structures of the ligand binding domains of TRalpha and TRbeta in complex with 15. These structures reveal that the single amino acid difference in the ligand binding pocket (Ser277 in TRalpha or Asn331 in TRbeta) results in a slightly different hydrogen bonding pattern that may explain the increased beta-selectivity of 15.
Assuntos
Fenilacetatos/síntese química , Receptores beta dos Hormônios Tireóideos/agonistas , Fosfatase Alcalina/genética , Animais , Sítios de Ligação , Células CHO , Colesterol/administração & dosagem , Colesterol/sangue , Cricetinae , Cristalografia por Raios X , Genes Reporter , Frequência Cardíaca/efeitos dos fármacos , Humanos , Ligantes , Masculino , Fenilacetatos/química , Fenilacetatos/farmacologia , Isoformas de Proteínas , Ensaio Radioligante , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Receptores alfa dos Hormônios Tireóideos/agonistas , Receptores alfa dos Hormônios Tireóideos/genética , Receptores alfa dos Hormônios Tireóideos/metabolismo , Receptores beta dos Hormônios Tireóideos/genética , Receptores beta dos Hormônios Tireóideos/metabolismo , Tireotropina/sangue , Tri-Iodotironina/farmacologiaRESUMO
It is desirable to obtain TR antagonists for treatment of hyperthyroidism and other conditions. We have designed TR antagonists from first principles based on TR crystal structures. Since agonist ligands are buried in the fold of the TR ligand binding domain (LBD), we reasoned that ligands that resemble agonists with large extensions should bind the LBD, but would prevent its folding into an active conformation. In particular, we predicted that extensions at the 5' aryl position of ligand should reposition helix (H) 12, which forms part of the co-activator binding surface, and thereby inhibit TR activity. We have found that some synthetic ligands with 5' aryl ring extensions behave as antagonists (DIBRT, NH-3), or partial antagonists (GC-14, NH-4). Moreover, one compound (NH-3) represents the first potent TR antagonist with nanomolar affinity that also inhibits TR action in an animal model. However, the properties of the ligands also reveal unexpected aspects of TR behavior. While nuclear receptor antagonists generally promote binding of co-repressors, NH-3 blocks co-activator binding and also prevents co-repressor binding. More surprisingly, many compounds with extensions behave as full or partial agonists. We present hypotheses to explain both behaviors in terms of dynamic equilibrium of H12 position.
Assuntos
Receptores dos Hormônios Tireóideos/antagonistas & inibidores , Receptores dos Hormônios Tireóideos/química , Animais , Sítios de Ligação , Núcleo Celular/metabolismo , Dimerização , Desenho de Fármacos , Humanos , Ligantes , Modelos Biológicos , Modelos Químicos , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Isoformas de Proteínas , Estrutura Terciária de ProteínaRESUMO
Based on the scaffold of the pharmacologically selective thyromimetic 2b, structurally a close analog to KB-141 (2a), a number of novel N-acylated-alpha-amino acid derivatives were synthesized and tested in a TR radioligand binding assay as well as in a reporter cell assay. On the basis of TRbeta(1)-isoform selectivity and affinity, as well as affinity to the reporter cell assay, 3d was selected for further studies in the cholesterol-fed rat model. In this model 3d revealed an improved therapeutic window between cholesterol and TSH lowering but decreased margins versus tachycardia compared with 2a.
Assuntos
Aminoácidos/farmacologia , Mimetismo Molecular , Éteres Fenílicos/farmacologia , Fenilacetatos/farmacologia , Receptores dos Hormônios Tireóideos/efeitos dos fármacos , Aminoácidos/química , Animais , Células CHO , Colesterol na Dieta/administração & dosagem , Cricetinae , Cricetulus , Ligantes , Ensaio Radioligante , Ratos , Receptores dos Hormônios Tireóideos/metabolismoRESUMO
Based on the examination of the X-ray crystallographic structures of the LBD of TRalpha and TRbeta in complex with KB-141 (2), a number of novel 4'-hydroxy bioisosteric thyromimetics were prepared. Optimal affinity and beta-selectivity (33 times), was found with a medium-sized alkyl-substituted amido group; iso-butyl (12c). It can be concluded that bioisosteric replacements of the 4'-hydroxy position represent a new promising class of TRbeta-selective synthetic thyromimetics.
Assuntos
Amidas/farmacologia , Receptores beta dos Hormônios Tireóideos/efeitos dos fármacos , Amidas/síntese química , Amidas/química , Cristalografia por Raios X , Desenho de Fármacos , Humanos , Ligantes , Modelos Moleculares , Conformação Molecular , Estrutura Molecular , Éteres Fenílicos/química , Fenilacetatos/química , Conformação Proteica , Estrutura Terciária de Proteína , Relação Estrutura-Atividade , Receptores alfa dos Hormônios Tireóideos/efeitos dos fármacos , Hormônios Tireóideos/química , Tri-Iodotironina/químicaRESUMO
Based on the examination of the crystal structure of rat TRbeta complexed with 3,5,3'-triiodo-l-thyronine (2) a novel TRbeta-selective indole derivative 6b was prepared and tested in vitro. This compound was found to be 14 times selective for TRbeta over TRalpha in binding and its beta-selectivity could be rationalized through the comparison of the X-ray crystallographic structures of 6b complexed with TRalpha and TRbeta.
Assuntos
Indóis/química , Indóis/farmacologia , Receptores beta dos Hormônios Tireóideos/agonistas , Receptores beta dos Hormônios Tireóideos/metabolismo , Animais , Cristalografia por Raios X , Ciclização , Humanos , Indóis/metabolismo , Concentração Inibidora 50 , Ligantes , Estrutura Molecular , Ratos , Especificidade por Substrato , Receptores beta dos Hormônios Tireóideos/química , Tiroxina/síntese química , Tiroxina/químicaRESUMO
Few treatments for obesity exist and improvements for treatment of hyperlipidemia are still desirable. Thyroid hormone receptors (TRs) regulate body weight, adiposity, and cholesterol levels. However, thyroid hormones can have deleterious effects, particularly cardiac acceleration, that limits the use of hormones in the treatment of obesity. There is evidence that the TRbeta subtype mediates lowering of blood cholesterol levels and possibly elevation of metabolic rate, whereas TRalpha appears to control heart rate. In studies, described in this review article, we examined the effects of selective TRbeta activation on metabolic rate and heart rate in mice, rats and monkeys. T3 had a greater effect on increasing heart rate in wild type (WT) than in TRalpha-/- mice (ED15 values of 34 and 469 nmol/kg/day, respectively). T3 increased metabolic rate (MVO2) in both WT and TRalpha-/- mice, but the effect on TRalpha-/- mice was less pronounced compared to WT mice. Stimulation of MVO2 is mediated by both TRalpha and TRbeta, but with different profiles. In cholesterol-fed rats, KB-141, a selective TRbeta agonist, increased MVO2 with a 10-fold selectivity and lowered cholesterol with a 27-fold selectivity vs. tachycardia. In primates, KB-141 caused significant, cholesterol, Lp(a) and body weight reduction after 1 week of treatment with no effect on heart rate. These data suggest that selective TRbeta agonists may represent a novel class of drugs for the treatment of obesity, hypercholesterolemia and elevated Lp(a), which may make them useful therapeutics for patients with metabolic syndrome.
Assuntos
Hipercolesterolemia/tratamento farmacológico , Obesidade/tratamento farmacológico , Éteres Fenílicos/uso terapêutico , Fenilacetatos/uso terapêutico , Receptores beta dos Hormônios Tireóideos/agonistas , Animais , Colesterol/administração & dosagem , Colesterol/metabolismo , Coração/efeitos dos fármacos , Coração/fisiologia , Frequência Cardíaca/efeitos dos fármacos , Humanos , Hipercolesterolemia/metabolismo , Obesidade/metabolismo , Éteres Fenílicos/farmacocinética , Éteres Fenílicos/farmacologia , Fenilacetatos/farmacocinética , Fenilacetatos/farmacologia , Relação Estrutura-Atividade , Distribuição Tecidual , Redução de Peso/efeitos dos fármacosRESUMO
High affinity thyromimetics containing a novel phenyl-naphthylene core are reported. The functionalized core is readily accessible via a Suzuki coupling protocol. Examples of this new class of TR ligands have sub-nanomolar binding affinities for the TRbeta receptor and low to modest selectivity for TRbeta. They also exhibit an SAR that diverges from other thyromimetics that are based on the diaryl ether core found in 3,5,3'-triiodothyronine.
Assuntos
Naftalenos/química , Receptores dos Hormônios Tireóideos/metabolismo , Hormônios Tireóideos/metabolismo , Modelos Moleculares , Relação Estrutura-Atividade , Hormônios Tireóideos/químicaRESUMO
Few treatments for obesity exist and, whereas efficacious therapeutics for hyperlipidemia are available, further improvements are desirable. Thyroid hormone receptors (TRs) regulate both body weight and cholesterol levels. However, thyroid hormones also have deleterious effects, particularly on the heart. The TR beta subtype is involved in cholesterol lowering and possibly elevating metabolic rate, whereas TR alpha appears to be more important for control of heart rate (HR). In the current studies, we examined the effect of TR beta activation on metabolic rate and HR with either TR alpha 1-/- mice or the selective TR beta agonist KB-141 in mice, rats, and monkeys. 3,5,3'-triiodi-l-thyronine (T3) had a greater effect on increasing HR in WT than in TR alpha-/- mice (ED15 values of 34 and 469 nmol/kg/day, respectively). T3 increased metabolic rate [whole body oxygen consumption (MVO2)] in both WT and TR alpha-/- mice, but the effect in the TR alpha 1-/- mice at the highest dose was half that of the WT mice. Thus, stimulation of MVO2 is likely due to both TR alpha and -beta. T3 had equivalent potency for cholesterol reduction in WT and TR alpha-/- mice. KB-141 increased MVO2 with selectivities of 16.5- and 11.2-fold vs. HR in WT and TR alpha 1-/- mice, respectively. KB-141 also increased MVO2 with a 10-fold selectivity and lowered cholesterol with a 27-fold selectivity vs. HR in rats. In primates, KB-141 caused significant cholesterol, lipoprotein (a), and body-weight reduction (up to 7% after 1 wk) with no effect on HR. TR beta-selective agonists may constitute a previously uncharacterized class of drugs to treat obesity, hypercholesterolemia, and elevated lipoprotein (a).
Assuntos
Colesterol/sangue , Lipoproteína(a)/sangue , Éteres Fenílicos/farmacologia , Fenilacetatos/farmacologia , Receptores dos Hormônios Tireóideos/agonistas , Redução de Peso/efeitos dos fármacos , Animais , Anticolesterolemiantes/farmacologia , Sistema Cardiovascular/efeitos dos fármacos , Colesterol na Dieta/administração & dosagem , Humanos , Técnicas In Vitro , Cinética , Macaca fascicularis , Camundongos , Camundongos Knockout , Ratos , Ratos Sprague-Dawley , Receptores dos Hormônios Tireóideos/metabolismo , Receptores alfa dos Hormônios Tireóideos/deficiência , Receptores alfa dos Hormônios Tireóideos/genética , Receptores beta dos Hormônios Tireóideos , Tri-Iodotironina/farmacologiaRESUMO
A set of thyromimetics having improved selectivity for TR-beta1 were prepared by replacing the 3'-isopropyl group of 2 and 3 with substituents having increased steric bulk. From this limited SAR study, the most potent and selective compounds identified were derived from 2 and contained a 3'-phenyl moiety bearing small hydrophobic groups meta to the biphenyl link. X-ray crystal data of 15c complexed with TR-beta1 LBD shows methionine 442 to be displaced by the bulky R3' phenyl ethyl amide side chain. Movement of this amino acid side chain provides an expanded pocket for the bulky side chain while the ligand-receptor complex retains full agonist activity.