RESUMEN
Cellular lipid requirements are achieved through a combination of biosynthesis and import programs. Using isotope tracer analysis, we show that type I interferon (IFN) signaling shifts the balance of these programs by decreasing synthesis and increasing import of cholesterol and long chain fatty acids. Genetically enforcing this metabolic shift in macrophages is sufficient to render mice resistant to viral challenge, demonstrating the importance of reprogramming the balance of these two metabolic pathways in vivo. Unexpectedly, mechanistic studies reveal that limiting flux through the cholesterol biosynthetic pathway spontaneously engages a type I IFN response in a STING-dependent manner. The upregulation of type I IFNs was traced to a decrease in the pool size of synthesized cholesterol and could be inhibited by replenishing cells with free cholesterol. Taken together, these studies delineate a metabolic-inflammatory circuit that links perturbations in cholesterol biosynthesis with activation of innate immunity.
Asunto(s)
Colesterol/metabolismo , Inmunidad Innata , Interferón gamma/metabolismo , Transducción de Señal , Animales , Línea Celular Tumoral , Humanos , Interferon beta-1b , Proteínas de la Membrana/metabolismo , Ácido Mevalónico/metabolismo , Ratones , Ratones Endogámicos C57BL , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/metabolismo , Proteína 2 de Unión a Elementos Reguladores de Esteroles/metabolismoRESUMEN
Depending on endoplasmic reticulum (ER) stress levels, the ER transmembrane multidomain protein IRE1α promotes either adaptation or apoptosis. Unfolded ER proteins cause IRE1α lumenal domain homo-oligomerization, inducing trans autophosphorylation that further drives homo-oligomerization of its cytosolic kinase/endoribonuclease (RNase) domains to activate mRNA splicing of adaptive XBP1 transcription factor. However, under high/chronic ER stress, IRE1α surpasses an oligomerization threshold that expands RNase substrate repertoire to many ER-localized mRNAs, leading to apoptosis. To modulate these effects, we developed ATP-competitive IRE1α Kinase-Inhibiting RNase Attenuators-KIRAs-that allosterically inhibit IRE1α's RNase by breaking oligomers. One optimized KIRA, KIRA6, inhibits IRE1α in vivo and promotes cell survival under ER stress. Intravitreally, KIRA6 preserves photoreceptor functional viability in rat models of ER stress-induced retinal degeneration. Systemically, KIRA6 preserves pancreatic ß cells, increases insulin, and reduces hyperglycemia in Akita diabetic mice. Thus, IRE1α powerfully controls cell fate but can itself be controlled with small molecules to reduce cell degeneration.
Asunto(s)
Estrés del Retículo Endoplásmico , Endorribonucleasas/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Regulación Alostérica , Animales , Apoptosis/efectos de los fármacos , Línea Celular , Endorribonucleasas/química , Endorribonucleasas/metabolismo , Activación Enzimática/efectos de los fármacos , Humanos , Islotes Pancreáticos/metabolismo , Masculino , Ratones , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Ratas , Retina/metabolismo , Ribonucleasas/antagonistas & inhibidoresRESUMEN
The zinc-finger transcription factor Helios is critical for maintaining the identity, anergic phenotype and suppressive activity of regulatory T (Treg) cells. While it is an attractive target to enhance the efficacy of currently approved immunotherapies, no existing approaches can directly modulate Helios activity or abundance. Here, we report the structure-guided development of small molecules that recruit the E3 ubiquitin ligase substrate receptor cereblon to Helios, thereby promoting its degradation. Pharmacological Helios degradation destabilized the anergic phenotype and reduced the suppressive activity of Treg cells, establishing a route towards Helios-targeting therapeutics. More generally, this study provides a framework for the development of small-molecule degraders for previously unligandable targets by reprogramming E3 ligase substrate specificity.
Asunto(s)
Proteínas de Unión al ADN/efectos de los fármacos , Factor de Transcripción Ikaros/efectos de los fármacos , Linfocitos T Reguladores/efectos de los fármacos , Factores de Transcripción/efectos de los fármacos , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Línea Celular , Proteínas de Unión al ADN/genética , Humanos , Factor de Transcripción Ikaros/genética , Células Jurkat , Ratones , Modelos Moleculares , Estructura Molecular , Mutación/genética , Bibliotecas de Moléculas Pequeñas , Especificidad por Sustrato , Factores de Transcripción/genética , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
Phosphatidylinositol 5-phosphate 4-kinase, type II, gamma (PIP4K2C) remains a poorly understood lipid kinase with minimal enzymatic activity but potential scaffolding roles in immune modulation and autophagy-dependent catabolism. Achieving potent and selective agents for PIP4K2C while sparing other lipid and non-lipid kinases has been challenging. Here, we report the discovery of the highly potent PIP4K2C binder TMX-4102, which shows exclusive binding selectivity for PIP4K2C. Furthermore, we elaborated the PIP4K2C binder into TMX-4153, a bivalent degrader capable of rapidly and selectively degrading endogenous PIP4K2C. Collectively, our work demonstrates that PIP4K2C is a tractable and degradable target, and that TMX-4102 and TMX-4153 are useful leads to further interrogate the biological roles and therapeutic potential of PIP4K2C.
Asunto(s)
AutofagiaRESUMEN
The covalent Bruton tyrosine kinase (BTK) inhibitor ibrutinib is highly efficacious against multiple B-cell malignancies. However, it is not selective for BTK, and multiple mechanisms of resistance, including the C481S-BTK mutation, can compromise its efficacy. We hypothesized that small-molecule-induced BTK degradation may overcome some of the limitations of traditional enzymatic inhibitors. Here, we demonstrate that BTK degradation results in potent suppression of signaling and proliferation in cancer cells and that BTK degraders efficiently degrade C481S-BTK. Moreover, we discovered DD-03-171, an optimized lead compound that exhibits enhanced antiproliferative effects on mantle cell lymphoma (MCL) cells in vitro by degrading BTK, IKFZ1, and IKFZ3 as well as efficacy against patient-derived xenografts in vivo. Thus, "triple degradation" may be an effective therapeutic approach for treating MCL and overcoming ibrutinib resistance, thereby addressing a major unmet need in the treatment of MCL and other B-cell lymphomas.
Asunto(s)
Agammaglobulinemia Tirosina Quinasa/metabolismo , Antineoplásicos/farmacología , Proliferación Celular/efectos de los fármacos , Linfoma de Células del Manto/tratamiento farmacológico , Bibliotecas de Moléculas Pequeñas/farmacología , Adenina/análogos & derivados , Animales , Humanos , Factor de Transcripción Ikaros/metabolismo , Linfoma de Células del Manto/enzimología , Linfoma de Células del Manto/patología , Ratones , Ratones Endogámicos NOD , Ratones SCID , Piperidinas , Proteolisis , Pirazoles/farmacología , Pirimidinas/farmacología , Transducción de Señal , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Cyclin-dependent kinases 4 and 6 (CDK4/6) are key regulators of the cell cycle, and there are FDA-approved CDK4/6 inhibitors for treating patients with metastatic breast cancer. However, due to conservation of their ATP-binding sites, development of selective agents has remained elusive. Here, we report imide-based degrader molecules capable of degrading both CDK4/6, or selectively degrading either CDK4 or CDK6. We were also able to tune the activity of these molecules against Ikaros (IKZF1) and Aiolos (IKZF3), which are well-established targets of imide-based degraders. We found that in mantle cell lymphoma cell lines, combined IKZF1/3 degradation with dual CDK4/6 degradation produced enhanced anti-proliferative effects compared to CDK4/6 inhibition, CDK4/6 degradation, or IKZF1/3 degradation. In summary, we report here the first compounds capable of inducing selective degradation of CDK4 and CDK6 as tools to pharmacologically dissect their distinct biological functions.
Asunto(s)
Quinasa 4 Dependiente de la Ciclina/metabolismo , Quinasa 6 Dependiente de la Ciclina/metabolismo , Inhibidores de Proteínas Quinasas/química , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Quinasa 4 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 6 Dependiente de la Ciclina/antagonistas & inhibidores , Humanos , Factor de Transcripción Ikaros/metabolismo , Imidas/química , Piperazinas/química , Piperazinas/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Piridinas/química , Piridinas/farmacologíaRESUMEN
Stable isotope labeling has become an important methodology for determining lipid metabolic parameters of normal and neoplastic cells. Conventional methods for fatty acid and cholesterol analysis have one or more issues that limit their utility for in vitro stable isotope-labeling studies. To address this, we developed a method optimized for measuring both fatty acids and cholesterol from small numbers of stable isotope-labeled cultured cells. We demonstrate quantitative derivatization and extraction of fatty acids from a wide range of lipid classes using this approach. Importantly, cholesterol is also recovered, albeit at a modestly lower yield, affording the opportunity to quantitate both cholesterol and fatty acids from the same sample. Although we find that background contamination can interfere with quantitation of certain fatty acids in low amounts of starting material, our data indicate that this optimized method can be used to accurately measure mass isotopomer distributions for cholesterol and many fatty acids isolated from small numbers of cultured cells. Application of this method will facilitate acquisition of lipid parameters required for quantifying flux and provide a better understanding of how lipid metabolism influences cellular function.
Asunto(s)
Colesterol/aislamiento & purificación , Ácidos Grasos/aislamiento & purificación , Marcaje Isotópico/métodos , Metabolismo de los Lípidos , Línea Celular , Colesterol/metabolismo , Ácidos Grasos/metabolismo , Humanos , Isótopos/farmacologíaRESUMEN
Mandated child abuse reporters may judge specific disciplinary practices as unacceptable for young children, whereas child law professionals arbitrating allegations may be less inclusive. Do the views of these groups diverge, by child age, regarding discipline? Judgments of community norms across a wide range of children's ages were obtained from 380 medical and legal professionals. Because the Parent-Child Conflict Tactics Scale (PC-CTS) can be used to assess the epidemiology of child disciplinary behaviors and as a proxy to examine the incidence or prevalence of child abuse, the disciplinary practices described on the PC-CTS were presented as triggers for questions. Significant child age effects were found for disciplinary practices classified as "harsh." The consistencies between legal and medical professionals were striking. Both groups reflected changes in United States norms, as non-physical approaches were the most approved. We conclude that instruments estimating the prevalence of child maltreatment by parent-report should consider modifying how specific disciplinary practices are classified. Copyright © 2016 John Wiley & Sons, Ltd.
Asunto(s)
Crianza del Niño/psicología , Derecho Penal/métodos , Castigo/psicología , Adulto , Actitud , Actitud del Personal de Salud , Niño , Maltrato a los Niños , Derecho Penal/normas , Relaciones Familiares , Femenino , Personal de Salud/psicología , Humanos , Abogados/psicología , Masculino , Persona de Mediana Edad , Percepción , Características de la Residencia , Estados UnidosRESUMEN
Over 95% of pancreatic ductal adenocarcinomas (PDAC) harbor oncogenic mutations in K-Ras. Upon treatment with K-Ras inhibitors, PDAC cancer cells undergo metabolic reprogramming towards an oxidative phosphorylation-dependent, drug-resistant state. However, direct inhibition of complex I is poorly tolerated in patients due to on-target induction of peripheral neuropathy. In this work, we develop molecular glue degraders against ZBTB11, a C2H2 zinc finger transcription factor that regulates the nuclear transcription of components of the mitoribosome and electron transport chain. Our ZBTB11 degraders leverage the differences in demand for biogenesis of mitochondrial components between human neurons and rapidly-dividing pancreatic cancer cells, to selectively target the K-Ras inhibitor resistant state in PDAC. Combination treatment of both K-Ras inhibitor-resistant cell lines and multidrug resistant patient-derived organoids resulted in superior anti-cancer activity compared to single agent treatment, while sparing hiPSC-derived neurons. Proteomic and stable isotope tracing studies revealed mitoribosome depletion and impairment of the TCA cycle as key events that mediate this response. Together, this work validates ZBTB11 as a vulnerability in K-Ras inhibitor-resistant PDAC and provides a suite of molecular glue degrader tool compounds to investigate its function.
RESUMEN
Augmenting the natural melanocortin pathway in mouse eyes with uveitis or diabetes protects the retinas from degeneration. The retinal cells are protected from oxidative and apoptotic signals of death. Therefore, we investigated the effects of a therapeutic application of the melanocortin alpha-melanocyte-stimulating hormone (α-MSH) on an ischemia and reperfusion (I/R) model of retinal degenerative disease. Eyes were subjected to an I/R procedure and were treated with α-MSH. Retinal sections were histopathologically scored. Also, the retinal sections were immunostained for viable ganglion cells, activated Muller cells, microglial cells, and apoptosis. The I/R caused retinal deformation and ganglion cell loss that was significantly reduced in I/R eyes treated with α-MSH. While α-MSH treatment marginally reduced the number of GFAP-positive Muller cells, it significantly suppressed the density of Iba1-positive microglial cells in the I/R retinas. Within one hour after I/R, there was apoptosis in the ganglion cell layer, and by 48 h, there was apoptosis in all layers of the neuroretina. The α-MSH treatment significantly reduced and delayed the onset of apoptosis in the retinas of I/R eyes. The results demonstrate that therapeutically augmenting the melanocortin pathways preserves retinal structure and cell survival in eyes with progressive neuroretinal degenerative disease.
Asunto(s)
Apoptosis , Homeostasis , Daño por Reperfusión , Retina , Células Ganglionares de la Retina , alfa-MSH , Animales , alfa-MSH/farmacología , alfa-MSH/metabolismo , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Ratones , Apoptosis/efectos de los fármacos , Retina/metabolismo , Retina/efectos de los fármacos , Retina/patología , Homeostasis/efectos de los fármacos , Células Ganglionares de la Retina/metabolismo , Células Ganglionares de la Retina/efectos de los fármacos , Células Ganglionares de la Retina/patología , Ratones Endogámicos C57BL , Microglía/metabolismo , Microglía/efectos de los fármacos , Masculino , Células Ependimogliales/metabolismo , Células Ependimogliales/efectos de los fármacos , Células Ependimogliales/patología , Modelos Animales de Enfermedad , Degeneración Retiniana/metabolismo , Degeneración Retiniana/patología , Degeneración Retiniana/tratamiento farmacológicoRESUMEN
In this issue of Cell Chemical Biology, Bonazzi et al. demonstrate that pharmacologically degrading the transcription factor Helios (IKZF2) results in destabilization of regulatory T cells, which normally restrain anti-tumor immunity. These results highlight how molecular glue degraders can selectively target previously undruggable proteins with potential applications in the clinic.
Asunto(s)
Neoplasias , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Linfocitos T Reguladores/metabolismo , Factores de Transcripción/metabolismo , Regulación de la Expresión GénicaRESUMEN
Phosphatidylinositol 5-phosphate 4-kinases (PI5P4Ks), a family of three members in mammals (α, ß and γ), have emerged as potential therapeutic targets due to their role in regulating many important cellular signaling pathways. In comparison to the PI5P4Kα and PI5P4Kß, which usually have similar expression profiles across cancer cells, PI5P4Kγ exhibits distinct expression patterns, and pathological functions for PI5P4Kγ have been proposed in the context of cancer and neurodegenerative diseases. PI5P4Kγ has very low kinase activity and has been proposed to inhibit the PI4P5Ks through scaffolding function, providing a rationale for developing a selective PI5P4Kγ degrader. Here, we report the development and characterization of JWZ-1-80, a first-in-class PI5P4Kγ degrader. JWZ-1-80 potently degrades PI5P4Kγ via the ubiquitin-proteasome system and exhibits proteome-wide selectivity and is therefore a useful tool compound for further dissecting the biological functions of PI5P4Kγ.
Asunto(s)
Mamíferos , Animales , Citoplasma , Fosforilación , ProteolisisRESUMEN
PSGL-1 (P-selectin glycoprotein-1) is a T cell-intrinsic checkpoint regulator of exhaustion with an unknown mechanism of action. Here, we show that PSGL-1 acts upstream of PD-1 and requires co-ligation with the T cell receptor (TCR) to attenuate activation of mouse and human CD8+ T cells and drive terminal T cell exhaustion. PSGL-1 directly restrains TCR signaling via Zap70 and maintains expression of the Zap70 inhibitor Sts-1. PSGL-1 deficiency empowers CD8+ T cells to respond to low-affinity TCR ligands and inhibit growth of PD-1-blockade-resistant melanoma by enabling tumor-infiltrating T cells to sustain an elevated metabolic gene signature supportive of increased glycolysis and glucose uptake to promote effector function. This outcome is coupled to an increased abundance of CD8+ T cell stem cell-like progenitors that maintain effector functions. Additionally, pharmacologic blockade of PSGL-1 curtails T cell exhaustion, indicating that PSGL-1 represents an immunotherapeutic target for PD-1-blockade-resistant tumors.
Asunto(s)
Linfocitos T CD8-positivos , Receptor de Muerte Celular Programada 1 , Humanos , Linfocitos T CD8-positivos/metabolismo , Diferenciación Celular , Receptor de Muerte Celular Programada 1/metabolismo , Receptores de Antígenos de Linfocitos T/metabolismo , Agotamiento de Células TRESUMEN
Respiratory tract (RT) infections by members of the enterovirus (EV) genus of the Picornaviridae family are the most frequent cause for the common cold and a major factor in the exacerbation of chronic pulmonary diseases. The lack of a practical small-animal model for these infections has obstructed insight into pathogenic mechanisms of the common cold and their role in chronic RT illness and has hampered preclinical evaluation of antiviral strategies. Despite significant efforts, it has been difficult to devise rodent models that exhibit viral replication in the RT. This is due mainly to well-known intracellular host restrictions of EVs with RT tropism in rodent cells. We report the evolution of variants of the common-cold-causing coxsackievirus A21, an EV with tropism for the human intercellular adhesion molecule 1 (hICAM-1), through serial passage in the lungs of mice transgenic for the hICAM-1 gene. This process was accompanied by multiple changes in the viral genome, suggesting exquisite adaptation of hICAM-1-tropic enteroviruses to the specific growth conditions within the RT. In vivo mouse RT-adapted, variant coxsackievirus A21 exhibited replication competence in the lungs of hICAM-1 transgenic mice, providing a basis for unraveling EV-host interactions in the mouse RT.
Asunto(s)
Adaptación Biológica , Enterovirus/fisiología , Molécula 1 de Adhesión Intercelular/metabolismo , Receptores Virales/metabolismo , Sistema Respiratorio/virología , Tropismo Viral , Animales , Enterovirus/patogenicidad , Genoma Viral , Humanos , Molécula 1 de Adhesión Intercelular/genética , Ratones , Ratones Transgénicos , ARN Viral/genética , Receptores Virales/genéticaRESUMEN
Recent interest in the role that extracellular signal-regulated kinase 5 (ERK5) plays in various diseases, particularly cancer and inflammation, has grown. Phenotypes observed from genetic knockdown or deletion of ERK5 suggested that targeting ERK5 could have therapeutic potential in various disease settings, motivating the development ATP-competitive ERK5 inhibitors. However, these inhibitors were unable to recapitulate the effects of genetic loss of ERK5, suggesting that ERK5 may have key kinase-independent roles. To investigate potential non-catalytic functions of ERK5, we report the development of INY-06-061, a potent and selective heterobifunctional degrader of ERK5. In contrast to results reported through genetic knockdown of ERK5, INY-06-061-induced ERK5 degradation did not induce anti-proliferative effects in multiple cancer cell lines or suppress inflammatory responses in primary endothelial cells. Thus, we developed and characterized a chemical tool useful for validating phenotypes reported to be associated with genetic ERK5 ablation and for guiding future ERK5-directed drug discovery efforts.
Asunto(s)
Células Endoteliales , Proteína Quinasa 7 Activada por Mitógenos , Humanos , Proteína Quinasa 7 Activada por Mitógenos/genética , Proteína Quinasa 7 Activada por Mitógenos/metabolismo , Células Endoteliales/metabolismo , Inmunidad Celular , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Proliferación CelularRESUMEN
Immunomodulatory imide drugs (IMiDs), such as thalidomide and its analogues, are some of the most commonly utilized E3 ligase ligands for the development of proteolysis targeting chimeras (PROTACs). While the canonical neo-substrates of IMiDs (i.e., Ikaros and Aiolos) are often considered to be unwanted targets of PROTACs, maintaining the degradation of these neo-substrates also provides the opportunity to synergistically degrade multiple proteins with a single compound. Here, we report the development of ALV-07-082-03, a CDK4/CDK6/Helios triple degrader that consists of palbociclib, an FDA-approved CDK4/6 inhibitor, conjugated to DKY709, a novel IMiD-based Helios degrader. Pharmacological codegradation of CDK4/6 and Helios resulted in potent suppression of downstream signaling and proliferation in cancer cells, as well as enhanced derepression of IL-2 secretion. Thus, not only do we demonstrate the possibility of rationally redirecting the neo-substrate specificity of PROTACs by incorporating alternative molecular glue molecules as E3 ligase ligands but our findings also suggest that cotargeting CDK4/6 and Helios may have synergistic effects.
Asunto(s)
Quimera , Talidomida , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Quimera/metabolismo , Péptidos y Proteínas de Señalización Intercelular , Interleucina-2/metabolismo , Ligandos , Proteolisis , Especificidad por Sustrato , Talidomida/farmacología , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
CDK4/6 inhibitors are approved to treat breast cancer and are in trials for other malignancies. We examined CDK4/6 inhibition in mouse and human CD8+ T cells during early stages of activation. Mice receiving tumor-specific CD8+ T cells treated with CDK4/6 inhibitors displayed increased T-cell persistence and immunologic memory. CDK4/6 inhibition upregulated MXD4, a negative regulator of MYC, in both mouse and human CD8+ T cells. Silencing of Mxd4 or Myc in mouse CD8+ T cells demonstrated the importance of this axis for memory formation. We used single-cell transcriptional profiling and T-cell receptor clonotype tracking to evaluate recently activated human CD8+ T cells in patients with breast cancer before and during treatment with either palbociclib or abemaciclib. CDK4/6 inhibitor therapy in humans increases the frequency of CD8+ memory precursors and downregulates their expression of MYC target genes, suggesting that CDK4/6 inhibitors in patients with cancer may augment long-term protective immunity. SIGNIFICANCE: CDK4/6 inhibition skews newly activated CD8+ T cells toward a memory phenotype in mice and humans with breast cancer. CDK4/6 inhibitors may have broad utility outside breast cancer, particularly in the neoadjuvant setting to augment CD8+ T-cell priming to tumor antigens prior to dosing with checkpoint blockade.This article is highlighted in the In This Issue feature, p. 2355.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Quinasa 4 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 6 Dependiente de la Ciclina/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/uso terapéutico , Adulto , Anciano , Aminopiridinas/uso terapéutico , Animales , Bencimidazoles/uso terapéutico , Neoplasias de la Mama/patología , Neoplasias de la Mama Masculina/tratamiento farmacológico , Neoplasias de la Mama Masculina/patología , Linfocitos T CD8-positivos/efectos de los fármacos , Línea Celular Tumoral , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Piperazinas/uso terapéutico , Inhibidores de Proteínas Quinasas/farmacología , Piridinas/uso terapéuticoRESUMEN
Cereblon (CRBN) is an E3 ligase adapter protein that can be reprogrammed by imide-class compounds such as thalidomide, lenalidomide, and pomalidomide to induce the degradation of neo-substrate proteins. In order to identify additional small molecule CRBN modulators, we implemented a focused combinatorial library approach where we fused an imide-based CRBN-binding pharmacophore to a heterocyclic scaffold, which could be further elaborated. We screened the library for CRBN-dependent antiproliferative activity in the multiple myeloma cell line MM1.S and identified five hit compounds. Quantitative chemical proteomics of hit compounds revealed that they induced selective degradation of GSPT1, a translation termination factor that is currently being explored as a therapeutic target for the treatment of acute myeloid leukemia. Molecular docking studies with CRBN and GSPT1 followed by analogue synthesis identified a possible hydrogen bond interaction with the central pyrimidine ring as a molecular determinant of hit compounds' selectivity. This study demonstrates that a focused combinatorial library design, phenotypic screening, and chemical proteomics can provide a suitable workflow to efficiently identify novel CRBN modulators.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Factores de Terminación de Péptidos/metabolismo , Proteolisis/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/farmacología , Talidomida/análogos & derivados , Talidomida/farmacología , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas Adaptadoras Transductoras de Señales/química , Antineoplásicos/metabolismo , Antineoplásicos/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Humanos , Enlace de Hidrógeno , Simulación del Acoplamiento Molecular , Factores de Terminación de Péptidos/química , Unión Proteica , Bibliotecas de Moléculas Pequeñas/metabolismo , Talidomida/metabolismo , Ubiquitina-Proteína Ligasas/químicaRESUMEN
Cyclin-dependent kinase 7 (CDK7) is a central regulator of the cell cycle and gene transcription. However, little is known about its impact on genomic instability and cancer immunity. Using a selective CDK7 inhibitor, YKL-5-124, we demonstrated that CDK7 inhibition predominately disrupts cell-cycle progression and induces DNA replication stress and genome instability in small cell lung cancer (SCLC) while simultaneously triggering immune-response signaling. These tumor-intrinsic events provoke a robust immune surveillance program elicited by T cells, which is further enhanced by the addition of immune-checkpoint blockade. Combining YKL-5-124 with anti-PD-1 offers significant survival benefit in multiple highly aggressive murine models of SCLC, providing a rationale for new combination regimens consisting of CDK7 inhibitors and immunotherapies.
Asunto(s)
Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Quinasas Ciclina-Dependientes/genética , Inestabilidad Genómica , Neoplasias Pulmonares/genética , Carcinoma Pulmonar de Células Pequeñas/genética , Animales , Antineoplásicos/farmacología , Linfocitos T CD4-Positivos/citología , Linfocitos T CD8-positivos/citología , Quimiocina CXCL9/metabolismo , Daño del ADN , Femenino , Humanos , Sistema Inmunológico , Inflamación , Interferón gamma/metabolismo , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/inmunología , Masculino , Ratones , Pruebas de Micronúcleos , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Pirazoles/farmacología , Pirroles/farmacología , Transducción de Señal , Carcinoma Pulmonar de Células Pequeñas/tratamiento farmacológico , Carcinoma Pulmonar de Células Pequeñas/inmunología , Factor de Necrosis Tumoral alfa/metabolismo , Quinasa Activadora de Quinasas Ciclina-DependientesRESUMEN
The design of selective small molecules is often stymied by similar ligand binding pockets. Here, we report BSJ-03-123, a phthalimide-based degrader that exploits protein-interface determinants to achieve proteome-wide selectivity for the degradation of cyclin-dependent kinase 6 (CDK6). Pharmacologic CDK6 degradation targets a selective dependency of acute myeloid leukemia cells, and transcriptomics and phosphoproteomics profiling of acute degradation of CDK6 enabled dynamic mapping of its immediate role in coordinating signaling and transcription.