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1.
Cell ; 175(1): 101-116.e25, 2018 09 20.
Artículo en Inglés | MEDLINE | ID: mdl-30220459

RESUMEN

IDH1 mutations are common in low-grade gliomas and secondary glioblastomas and cause overproduction of (R)-2HG. (R)-2HG modulates the activity of many enzymes, including some that are linked to transformation and some that are probably bystanders. Although prior work on (R)-2HG targets focused on 2OG-dependent dioxygenases, we found that (R)-2HG potently inhibits the 2OG-dependent transaminases BCAT1 and BCAT2, likely as a bystander effect, thereby decreasing glutamate levels and increasing dependence on glutaminase for the biosynthesis of glutamate and one of its products, glutathione. Inhibiting glutaminase specifically sensitized IDH mutant glioma cells to oxidative stress in vitro and to radiation in vitro and in vivo. These findings highlight the complementary roles for BCATs and glutaminase in glutamate biosynthesis, explain the sensitivity of IDH mutant cells to glutaminase inhibitors, and suggest a strategy for maximizing the effectiveness of such inhibitors against IDH mutant gliomas.


Asunto(s)
Glioma/metabolismo , Ácido Glutámico/biosíntesis , Transaminasas/fisiología , Línea Celular Tumoral , Glioma/fisiopatología , Ácido Glutámico/efectos de los fármacos , Glutaratos/metabolismo , Glutaratos/farmacología , Homeostasis/efectos de los fármacos , Humanos , Isocitrato Deshidrogenasa/genética , Isocitrato Deshidrogenasa/fisiología , Antígenos de Histocompatibilidad Menor/genética , Antígenos de Histocompatibilidad Menor/fisiología , Mutación , Oxidación-Reducción/efectos de los fármacos , Proteínas Gestacionales/genética , Proteínas Gestacionales/fisiología , Transaminasas/antagonistas & inhibidores , Transaminasas/genética
2.
Cell ; 160(5): 977-989, 2015 Feb 26.
Artículo en Inglés | MEDLINE | ID: mdl-25723171

RESUMEN

There is a lack of effective predictive biomarkers to precisely assign optimal therapy to cancer patients. While most efforts are directed at inferring drug response phenotype based on genotype, there is very focused and useful phenotypic information to be gained from directly perturbing the patient's living cancer cell with the drug(s) in question. To satisfy this unmet need, we developed the Dynamic BH3 Profiling technique to measure early changes in net pro-apoptotic signaling at the mitochondrion ("priming") induced by chemotherapeutic agents in cancer cells, not requiring prolonged ex vivo culture. We find in cell line and clinical experiments that early drug-induced death signaling measured by Dynamic BH3 Profiling predicts chemotherapy response across many cancer types and many agents, including combinations of chemotherapies. We propose that Dynamic BH3 Profiling can be used as a broadly applicable predictive biomarker to predict cytotoxic response of cancers to chemotherapeutics in vivo.


Asunto(s)
Muerte Celular , Neoplasias/tratamiento farmacológico , Transducción de Señal , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular , Femenino , Humanos , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Mitocondrias/metabolismo , Neoplasias/patología , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/patología , Medicina de Precisión
3.
Nature ; 615(7950): 158-167, 2023 03.
Artículo en Inglés | MEDLINE | ID: mdl-36634707

RESUMEN

Despite the success of PD-1 blockade in melanoma and other cancers, effective treatment strategies to overcome resistance to cancer immunotherapy are lacking1,2. Here we identify the innate immune kinase TANK-binding kinase 1 (TBK1)3 as a candidate immune-evasion gene in a pooled genetic screen4. Using a suite of genetic and pharmacological tools across multiple experimental model systems, we confirm a role for TBK1 as an immune-evasion gene. Targeting TBK1 enhances responses to PD-1 blockade by decreasing the cytotoxicity threshold to effector cytokines (TNF and IFNγ). TBK1 inhibition in combination with PD-1 blockade also demonstrated efficacy using patient-derived tumour models, with concordant findings in matched patient-derived organotypic tumour spheroids and matched patient-derived organoids. Tumour cells lacking TBK1 are primed to undergo RIPK- and caspase-dependent cell death in response to TNF and IFNγ in a JAK-STAT-dependent manner. Taken together, our results demonstrate that targeting TBK1 is an effective strategy to overcome resistance to cancer immunotherapy.


Asunto(s)
Resistencia a Antineoplásicos , Evasión Inmune , Inmunoterapia , Proteínas Serina-Treonina Quinasas , Humanos , Evasión Inmune/genética , Evasión Inmune/inmunología , Inmunoterapia/métodos , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/genética , Organoides , Factores de Necrosis Tumoral/inmunología , Interferón gamma/inmunología , Esferoides Celulares , Caspasas , Quinasas Janus , Factores de Transcripción STAT
4.
EMBO J ; 43(3): 391-413, 2024 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-38225406

RESUMEN

Cristae membrane state plays a central role in regulating mitochondrial function and cellular metabolism. The protein Optic atrophy 1 (Opa1) is an important crista remodeler that exists as two forms in the mitochondrion, a membrane-anchored long form (l-Opa1) and a processed short form (s-Opa1). The mechanisms for how Opa1 influences cristae shape have remained unclear due to lack of native three-dimensional views of cristae. We perform in situ cryo-electron tomography of cryo-focused ion beam milled mouse embryonic fibroblasts with defined Opa1 states to understand how each form of Opa1 influences cristae architecture. In our tomograms, we observe a variety of cristae shapes with distinct trends dependent on s-Opa1:l-Opa1 balance. Increased l-Opa1 levels promote cristae stacking and elongated mitochondria, while increased s-Opa1 levels correlated with irregular cristae packing and round mitochondria shape. Functional assays indicate a role for l-Opa1 in wild-type apoptotic and calcium handling responses, and show a compromised respiratory function under Opa1 imbalance. In summary, we provide three-dimensional visualization of cristae architecture to reveal relationships between mitochondrial ultrastructure and cellular function dependent on Opa1-mediated membrane remodeling.


Asunto(s)
Fibroblastos , Membranas Mitocondriales , Animales , Ratones , Fibroblastos/metabolismo , Membranas Mitocondriales/metabolismo , Mitocondrias/metabolismo , Proteínas Mitocondriales/metabolismo
5.
Nat Chem Biol ; 2024 Mar 13.
Artículo en Inglés | MEDLINE | ID: mdl-38480981

RESUMEN

A common approach for understanding how drugs induce their therapeutic effects is to identify the genetic determinants of drug sensitivity. Because 'chemo-genetic profiles' are performed in a pooled format, inference of gene function is subject to several confounding influences related to variation in growth rates between clones. In this study, we developed Method for Evaluating Death Using a Simulation-assisted Approach (MEDUSA), which uses time-resolved measurements, along with model-driven constraints, to reveal the combination of growth and death rates that generated the observed drug response. MEDUSA is uniquely effective at identifying death regulatory genes. We apply MEDUSA to characterize DNA damage-induced lethality in the presence and absence of p53. Loss of p53 switches the mechanism of DNA damage-induced death from apoptosis to a non-apoptotic death that requires high respiration. These findings demonstrate the utility of MEDUSA both for determining the genetic dependencies of lethality and for revealing opportunities to potentiate chemo-efficacy in a cancer-specific manner.

6.
Br J Cancer ; 130(9): 1463-1476, 2024 May.
Artículo en Inglés | MEDLINE | ID: mdl-38438589

RESUMEN

BACKGROUND: Uterine serous cancer (USC) comprises around 10% of all uterine cancers. However, USC accounts for approximately 40% of uterine cancer deaths, which is attributed to tumor aggressiveness and limited effective treatment. Galectin 3 (Gal3) has been implicated in promoting aggressive features in some malignancies. However, Gal3's role in promoting USC pathology is lacking. METHODS: We explored the relationship between LGALS3 levels and prognosis in USC patients using TCGA database, and examined the association between Gal3 levels in primary USC tumors and clinical-pathological features. CRISPR/Cas9-mediated Gal3-knockout (KO) and GB1107, inhibitor of Gal3, were employed to evaluate Gal3's impact on cell function. RESULTS: TCGA analysis revealed a worse prognosis for USC patients with high LGALS3. Patients with no-to-low Gal3 expression in primary tumors exhibited reduced clinical-pathological tumor progression. Gal3-KO and GB1107 reduced cell proliferation, stemness, adhesion, migration, and or invasion properties of USC lines. Furthermore, Gal3-positive conditioned media (CM) stimulated vascular tubal formation and branching and transition of fibroblast to cancer-associated fibroblast compared to Gal3-negative CM. Xenograft models emphasized the significance of Gal3 loss with fewer and smaller tumors compared to controls. Moreover, GB1107 impeded the growth of USC patient-derived organoids. CONCLUSION: These findings suggest inhibiting Gal3 may benefit USC patients.


Asunto(s)
Proteínas Sanguíneas , Cistadenocarcinoma Seroso , Galectina 3 , Neoplasias Uterinas , Humanos , Femenino , Neoplasias Uterinas/patología , Neoplasias Uterinas/genética , Neoplasias Uterinas/metabolismo , Galectina 3/genética , Galectina 3/metabolismo , Cistadenocarcinoma Seroso/patología , Cistadenocarcinoma Seroso/genética , Cistadenocarcinoma Seroso/metabolismo , Proliferación Celular , Línea Celular Tumoral , Pronóstico , Animales , Ratones , Galectinas/genética , Galectinas/metabolismo , Movimiento Celular
7.
Mol Cell ; 51(6): 751-65, 2013 Sep 26.
Artículo en Inglés | MEDLINE | ID: mdl-24074954

RESUMEN

Apoptosis is a highly regulated form of cell death that controls normal homeostasis as well as the antitumor activity of many chemotherapeutic agents. Commitment to death via the mitochondrial apoptotic pathway requires activation of the mitochondrial pore-forming proteins BAK or BAX. Activation can be effected by the activator BH3-only proteins BID or BIM, which have been considered to be functionally redundant in this role. Herein, we show that significant activation preferences exist between these proteins: BID preferentially activates BAK while BIM preferentially activates BAX. Furthermore, we find that cells lacking BAK are relatively resistant to agents that require BID activation for maximal induction of apoptosis, including topoisomerase inhibitors and TRAIL. Consequently, patients with tumors that harbor a loss of BAK1 exhibit an inferior response to topoisomerase inhibitor treatment in the clinic. Therefore, BID and BIM have nonoverlapping roles in the induction of apoptosis via BAK and BAX, affecting chemotherapy response.


Asunto(s)
Proteínas Reguladoras de la Apoptosis/metabolismo , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/metabolismo , Proteínas de la Membrana/metabolismo , Neoplasias Ováricas/genética , Proteínas Proto-Oncogénicas/metabolismo , Proteína Destructora del Antagonista Homólogo bcl-2/metabolismo , Proteína X Asociada a bcl-2/metabolismo , Apoptosis/efectos de los fármacos , Proteínas Reguladoras de la Apoptosis/genética , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/genética , Proteína 11 Similar a Bcl2 , Femenino , Regulación Neoplásica de la Expresión Génica , Células HeLa , Humanos , Proteínas de la Membrana/genética , Mitocondrias/efectos de los fármacos , Mitocondrias/genética , Mitocondrias/metabolismo , Estadificación de Neoplasias , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/patología , Proteínas Proto-Oncogénicas/genética , Inhibidores de Topoisomerasa/administración & dosificación , Activación Transcripcional/efectos de los fármacos , Proteína Destructora del Antagonista Homólogo bcl-2/genética , Proteína X Asociada a bcl-2/genética
8.
Part Fibre Toxicol ; 16(1): 40, 2019 10 29.
Artículo en Inglés | MEDLINE | ID: mdl-31665028

RESUMEN

BACKGROUND: Amorphous silica nanoparticles (SiO2 NPs) have been regarded as relatively benign nanomaterials, however, this widely held opinion has been questioned in recent years by several reports on in vitro and in vivo toxicity. Surface chemistry, more specifically the surface silanol content, has been identified as an important toxicity modulator for SiO2 NPs. Here, quantitative relationships between the silanol content on SiO2 NPs, free radical generation and toxicity have been identified, with the purpose of synthesizing safer-by-design fumed silica nanoparticles. RESULTS: Consistent and statistically significant trends were seen between the total silanol content, cell membrane damage, and cell viability, but not with intracellular reactive oxygen species (ROS), in the macrophages RAW264.7. SiO2 NPs with lower total silanol content exhibited larger adverse cellular effects. The SAEC epithelial cell line did not show any sign of toxicity by any of the nanoparticles. Free radical generation and surface reactivity of these nanoparticles were also influenced by the temperature of combustion and total silanol content. CONCLUSION: Surface silanol content plays an important role in cellular toxicity and surface reactivity, although it might not be the sole factor influencing fumed silica NP toxicity. It was demonstrated that synthesis conditions for SiO2 NPs influence the type and quantity of free radicals, oxidative stress, nanoparticle interaction with the biological milieu they come in contact with, and determine the specific mechanisms of toxicity. We demonstrate here that it is possible to produce much less toxic fumed silicas by modulating the synthesis conditions.


Asunto(s)
Macrófagos/efectos de los fármacos , Nanopartículas/toxicidad , Silanos/toxicidad , Dióxido de Silicio/toxicidad , Animales , Técnicas de Cultivo de Célula , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Membrana Celular/patología , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Macrófagos/metabolismo , Macrófagos/patología , Ratones , Nanopartículas/química , Estrés Oxidativo/efectos de los fármacos , Células RAW 264.7 , Especies Reactivas de Oxígeno , Silanos/química , Dióxido de Silicio/química , Propiedades de Superficie
9.
Blood ; 126(13): 1555-64, 2015 Sep 24.
Artículo en Inglés | MEDLINE | ID: mdl-26194763

RESUMEN

Mantle cell lymphoma (MCL) is a distinct subtype of non-Hodgkin lymphoma characterized by overexpression of cyclin D1 in 95% of patients. MCL patients experience frequent relapses resulting in median survival of 3 to 5 years, requiring more efficient therapeutic regimens. Interleukin (IL)-21, a member of the IL-2 cytokine family, possesses potent antitumor activity against a variety of cancers not expressing the IL-21 receptor (IL-21R) through immune activation. Previously, we established that IL-21 exerts direct cytotoxicity on IL-21R-expressing diffuse large B-cell lymphoma cells. Herein, we demonstrate that IL-21 possesses potent cytotoxicity against MCL cell lines and primary tumors. We identify that IL-21-induced direct cytotoxicity is mediated through signal transducer and activator of transcription 3-dependent cMyc upregulation, resulting in activation of Bax and inhibition of Bcl-2 and Bcl-XL. IL-21-mediated cMyc upregulation is only observed in IL-21-sensitive cells. Further, we demonstrate that IL-21 leads to natural killer (NK)-cell-dependent lysis of MCL cell lines that were resistant to direct cytotoxicity. In vivo treatment with IL-21 results in complete FC-muMCL1 tumor regression in syngeneic mice via NK- and T-cell-dependent mechanisms. Together, these data indicate that IL-21 has potent antitumor activity against MCL cells via direct cytotoxic and indirect, immune-mediated effects.


Asunto(s)
Factores Inmunológicos/inmunología , Factores Inmunológicos/uso terapéutico , Interleucinas/inmunología , Interleucinas/uso terapéutico , Linfoma de Células del Manto/inmunología , Linfoma de Células del Manto/terapia , Animales , Apoptosis/efectos de los fármacos , Linfocitos T CD4-Positivos/inmunología , Línea Celular Tumoral , Femenino , Humanos , Inmunoterapia , Células Asesinas Naturales/inmunología , Linfoma de Células del Manto/patología , Ratones Endogámicos C57BL , Proteínas Proto-Oncogénicas c-myc/inmunología , Receptores de Interleucina-21/inmunología , Factor de Transcripción STAT3/inmunología , Transducción de Señal/efectos de los fármacos , Proteína X Asociada a bcl-2/inmunología
10.
Proc Natl Acad Sci U S A ; 110(42): 17041-6, 2013 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-24085845

RESUMEN

Breast Cancer Type 1 Susceptibility Protein (BRCA1)-deficient cells have compromised DNA repair and are sensitive to poly(ADP-ribose) polymerase (PARP) inhibitors. Despite initial responses, the development of resistance limits clinical efficacy. Mutations in the BRCA C-terminal (BRCT) domain of BRCA1 frequently create protein products unable to fold that are subject to protease-mediated degradation. Here, we show HSP90-mediated stabilization of a BRCT domain mutant BRCA1 protein under PARP inhibitor selection pressure. The stabilized mutant BRCA1 protein interacted with PALB2-BRCA2-RAD51, was essential for RAD51 focus formation, and conferred PARP inhibitor as well as cisplatin resistance. Treatment of resistant cells with the HSP90 inhibitor 17-dimethylaminoethylamino-17-demethoxygeldanamycin reduced mutant BRCA1 protein levels and restored their sensitivity to PARP inhibition. Resistant cells also acquired a TP53BP1 mutation that facilitated DNA end resection in the absence of a BRCA1 protein capable of binding CtIP. Finally, concomitant increased mutant BRCA1 and decreased 53BP1 protein expression occur in clinical samples of BRCA1-mutated recurrent ovarian carcinomas that have developed resistance to platinum. These results provide evidence for a two-event mechanism by which BRCA1-mutant tumors acquire anticancer therapy resistance.


Asunto(s)
Antineoplásicos/farmacología , Proteína BRCA1/metabolismo , Cisplatino/farmacología , Resistencia a Antineoplásicos/efectos de los fármacos , Mutación , Neoplasias Ováricas/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Proteína BRCA1/genética , Proteína BRCA2/genética , Proteína BRCA2/metabolismo , Benzoquinonas/farmacología , Línea Celular Tumoral , Resistencia a Antineoplásicos/genética , Proteína del Grupo de Complementación N de la Anemia de Fanconi , Femenino , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Proteínas HSP90 de Choque Térmico/genética , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , Lactamas Macrocíclicas/farmacología , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Platino (Metal)/farmacología , Poli(ADP-Ribosa) Polimerasas/genética , Poli(ADP-Ribosa) Polimerasas/metabolismo , Estructura Terciaria de Proteína , Recombinasa Rad51/genética , Recombinasa Rad51/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo
11.
Trends Mol Med ; 30(1): 56-73, 2024 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-38057226

RESUMEN

Environmental exposures are linked to diseases of high public health concern, including cancer, neurodegenerative disorders, and autoimmunity. These diseases are caused by excessive or insufficient cell death, prompting investigation of mechanistic links between environmental toxicants and dysregulation of cell death pathways, including apoptosis. This review describes how legacy and emerging environmental exposures target the intrinsic apoptosis pathway to potentially drive pathogenesis. Recent discoveries reveal that dynamic regulation of apoptosis may heighten the vulnerability of healthy tissues to exposures in children, and that apoptotic signaling can guide immune responses, tissue repair, and tumorigenesis. Understanding how environmental toxicants dysregulate apoptosis will uncover opportunities to deploy apoptosis-modulating agents for the treatment or prevention of exposure-linked diseases.


Asunto(s)
Apoptosis , Neoplasias , Niño , Humanos , Apoptosis/fisiología , Exposición a Riesgos Ambientales/efectos adversos , Neoplasias/etiología , Evaluación de Resultado en la Atención de Salud , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo
12.
Proc Natl Acad Sci U S A ; 107(29): 13069-74, 2010 Jul 20.
Artículo en Inglés | MEDLINE | ID: mdl-20615981

RESUMEN

Primary effusion lymphoma (PEL) is an aggressive B-cell lymphoma most commonly diagnosed in HIV-positive patients and universally associated with Kaposi's sarcoma-associated herpesvirus (KSHV). Chemotherapy treatment of PEL yields only short-term remissions in the vast majority of patients, but efforts to develop superior therapeutic approaches have been impeded by lack of animal models that accurately mimic human disease. To address this issue, we developed a direct xenograft model, UM-PEL-1, by transferring freshly isolated human PEL cells into the peritoneal cavities of NOD/SCID mice without in vitro cell growth to avoid the changes in KSHV gene expression evident in cultured cells. We used this model to show that bortezomib induces PEL remission and extends overall survival of mice bearing lymphomatous effusions. The proapoptotic effects of bortezomib are not mediated by inhibition of the prosurvival NF-kappaB pathway or by induction of a terminal unfolded protein response. Transcriptome analysis by genomic arrays revealed that bortezomib down-regulated cell-cycle progression, DNA replication, and Myc-target genes. Furthermore, we demonstrate that in vivo treatment with either bortezomib or doxorubicin induces KSHV lytic reactivation. These reactivations were temporally distinct, and this difference may help elucidate the therapeutic window for use of antivirals concurrently with chemotherapy. Our findings show that this direct xenograft model can be used for testing novel PEL therapeutic strategies and also can provide a rational basis for evaluation of bortezomib in clinical trials.


Asunto(s)
Ácidos Borónicos/uso terapéutico , Linfoma de Efusión Primaria/tratamiento farmacológico , Pirazinas/uso terapéutico , Ensayos Antitumor por Modelo de Xenoinjerto , Anciano de 80 o más Años , Animales , Apoptosis/efectos de los fármacos , Ácidos Borónicos/farmacología , Bortezomib , Ciclo Celular/efectos de los fármacos , Ciclo Celular/genética , Chlorocebus aethiops , Replicación del ADN/efectos de los fármacos , Replicación del ADN/genética , Regulación hacia Abajo/efectos de los fármacos , Factor de Transcripción E2F3/metabolismo , Resultado Fatal , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Herpesvirus Humano 8/efectos de los fármacos , Herpesvirus Humano 8/fisiología , Humanos , Linfoma de Efusión Primaria/virología , Masculino , Ratones , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas c-myc/metabolismo , Pirazinas/farmacología , Análisis de Supervivencia , Resultado del Tratamiento , Respuesta de Proteína Desplegada/efectos de los fármacos , Células Vero , Virión/efectos de los fármacos , Virión/metabolismo
13.
Trends Cancer ; 9(2): 96-110, 2023 02.
Artículo en Inglés | MEDLINE | ID: mdl-37284233

RESUMEN

The intrinsic apoptosis pathway is controlled by the BCL-2 family of proteins. Although the pro-survival members of this family can help cancer cells evade apoptosis, they may also produce apoptotic vulnerabilities that can potentially be exploited therapeutically. Apoptotic vulnerabilities can be driven by endogenous factors including altered genetics, signaling, metabolism, structure and lineage or differentiation state as well as imposed factors, the most prominent being exposure to anti-cancer agents. The recent development of BH3 mimetics that inhibit pro-survival BCL-2 family proteins has allowed these apoptotic vulnerabilities to be targeted with demonstrable clinical success. Here, we review the key concepts that are vital for understanding, uncovering, and exploiting apoptotic vulnerabilities in cancer for the potential improvement of patient outcomes.


Asunto(s)
Antineoplásicos , Neoplasias , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Apoptosis , Proteínas Reguladoras de la Apoptosis/metabolismo , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico
14.
Structure ; 31(3): 265-281.e7, 2023 03 02.
Artículo en Inglés | MEDLINE | ID: mdl-36706751

RESUMEN

Apoptosis is important for development and tissue homeostasis, and its dysregulation can lead to diseases, including cancer. As an apoptotic effector, BAK undergoes conformational changes that promote mitochondrial outer membrane disruption, leading to cell death. This is termed "activation" and can be induced by peptides from the human proteins BID, BIM, and PUMA. To identify additional peptides that can regulate BAK, we used computational protein design, yeast surface display screening, and structure-based energy scoring to identify 10 diverse new binders. We discovered peptides from the human proteins BNIP5 and PXT1 and three non-native peptides that activate BAK in liposome assays and induce cytochrome c release from mitochondria. Crystal structures and binding studies reveal a high degree of similarity among peptide activators and inhibitors, ruling out a simple function-determining property. Our results shed light on the vast peptide sequence space that can regulate BAK function and will guide the design of BAK-modulating tools and therapeutics.


Asunto(s)
Proteínas Reguladoras de la Apoptosis , Proteínas Proto-Oncogénicas , Humanos , Proteínas Proto-Oncogénicas/química , Proteínas Reguladoras de la Apoptosis/química , Proteína 11 Similar a Bcl2 , Proteína bcl-X/metabolismo , Proteína Destructora del Antagonista Homólogo bcl-2/química , Proteína Destructora del Antagonista Homólogo bcl-2/metabolismo , Apoptosis/fisiología , Péptidos , Proteína X Asociada a bcl-2/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/química
15.
bioRxiv ; 2023 May 16.
Artículo en Inglés | MEDLINE | ID: mdl-36712034

RESUMEN

DNA damage can activate apoptotic and non-apoptotic forms of cell death; however, it remains unclear what features dictate which type of cell death is activated. We report that p53 controls the choice between apoptotic and non-apoptotic death following exposure to DNA damage. In contrast to the conventional model, which suggests that p53-deficient cells should be resistant to DNA damage-induced cell death, we find that p53-deficient cells die at high rates following DNA damage, but exclusively using non-apoptotic mechanisms. Our experimental data and computational modeling reveal that non-apoptotic death in p53-deficient cells has not been observed due to use of assays that are either insensitive to cell death, or that specifically score apoptotic cells. Using functional genetic screening - with an analysis that enables computational inference of the drug-induced death rate - we find in p53-deficient cells that DNA damage activates a mitochondrial respiration-dependent form of cell death, called MPT-driven necrosis. Cells deficient for p53 have high basal respiration, which primes MPT-driven necrosis. Finally, using metabolite profiling, we identified mitochondrial activity-dependent metabolic vulnerabilities that can be targeted to potentiate the lethality of DNA damage specifically in p53-deficient cells. Our findings reveal how the dual functions of p53 in regulating mitochondrial activity and the DNA damage response combine to facilitate the choice between apoptotic and non-apoptotic death.

16.
bioRxiv ; 2023 Nov 09.
Artículo en Inglés | MEDLINE | ID: mdl-36711707

RESUMEN

Cristae membrane state plays a central role in regulating mitochondrial function and cellular metabolism. The protein Optic atrophy 1 (Opa1) is an important crista remodeler that exists as two forms in the mitochondrion, a membrane-anchored long form (l-Opa1) and a processed short form (s-Opa1). The mechanisms for how Opa1 influences cristae shape have remained unclear due to lack of native three-dimensional views of cristae. We perform in situ cryo-electron tomography of cryo-focused ion beam milled mouse embryonic fibroblasts with defined Opa1 states to understand how each form of Opa1 influences cristae architecture. In our tomograms, we observe a variety of cristae shapes with distinct trends dependent on s-Opa1:l-Opa1 balance. Increased l-Opa1 levels promote cristae stacking and elongated mitochondria while increased s-Opa1 levels correlated with irregular cristae packing and round mitochondria shape. Functional assays indicate a role for l-Opa1 in wild-type apoptotic and calcium handling responses, and compromised respiratory function under Opa1 imbalance. In summary, we provide three-dimensional visualization of cristae architecture to reveal relationships between mitochondrial ultrastructure and cellular function dependent on Opa1-mediated membrane remodeling.

17.
Cancer Res ; 83(20): 3442-3461, 2023 10 13.
Artículo en Inglés | MEDLINE | ID: mdl-37470810

RESUMEN

Although external beam radiotherapy (xRT) is commonly used to treat central nervous system (CNS) tumors in patients of all ages, young children treated with xRT frequently experience life-altering and dose-limiting neurocognitive impairment (NI) while adults do not. The lack of understanding of mechanisms responsible for these differences has impeded the development of neuroprotective treatments. Using a newly developed mouse model of xRT-induced NI, we found that neurocognitive function is impaired by ionizing radiation in a dose- and age-dependent manner, with the youngest animals being most affected. Histologic analysis revealed xRT-driven neuronal degeneration and cell death in neurogenic brain regions in young animals but not adults. BH3 profiling showed that neural stem and progenitor cells, neurons, and astrocytes in young mice are highly primed for apoptosis, rendering them hypersensitive to genotoxic damage. Analysis of single-cell RNA sequencing data revealed that neural cell vulnerability stems from heightened expression of proapoptotic genes including BAX, which is associated with developmental and mitogenic signaling by MYC. xRT induced apoptosis in primed neural cells by triggering a p53- and PUMA-initiated, proapoptotic feedback loop requiring cleavage of BID and culminating in BAX oligomerization and caspase activation. Notably, loss of BAX protected against apoptosis induced by proapoptotic signaling in vitro and prevented xRT-induced apoptosis in neural cells in vivo as well as neurocognitive sequelae. On the basis of these findings, preventing xRT-induced apoptosis specifically in immature neural cells by blocking BAX, BIM, or BID via direct or upstream mechanisms is expected to ameliorate NI in pediatric patients with CNS tumor. SIGNIFICANCE: Age- and differentiation-dependent apoptotic priming plays a pivotal role in driving radiotherapy-induced neurocognitive impairment and can be targeted for neuroprotection in pediatric patients.


Asunto(s)
Proteínas Reguladoras de la Apoptosis , Apoptosis , Animales , Niño , Preescolar , Humanos , Ratones , Apoptosis/fisiología , Proteínas Reguladoras de la Apoptosis/metabolismo , Proteína X Asociada a bcl-2/metabolismo , Muerte Celular , Transducción de Señal , Proteína p53 Supresora de Tumor/genética
18.
Theranostics ; 13(14): 4711-4729, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37771768

RESUMEN

Background: The introduction of magnetic resonance (MR)-guided radiation treatment planning has opened a new space for theranostic nanoparticles to reduce acute toxicity while improving local control. In this work, second-generation AGuIX® nanoparticles (AGuIX-Bi) are synthesized and validated. AGuIX-Bi are shown to maintain MR positive contrast while further amplifying the radiation dose by the replacement of some Gd3+ cations with higher Z Bi3+. These next-generation nanoparticles are based on the AGuIX® platform, which is currently being evaluated in multiple Phase II clinical trials in combination with radiotherapy. Methods: In this clinically scalable methodology, AGuIX® is used as an initial chelation platform to exchange Gd3+ for Bi3+. AGuIX-Bi nanoparticles are synthesized with three ratios of Gd/Bi, each maintaining MR contrast while further amplifying radiation dose relative to Bi3+. Safety, efficacy, and theranostic potential of the nanoparticles were evaluated in vitro and in vivo in a human non-small cell lung cancer model. Results: We demonstrated that increasing Bi3+ in the nanoparticles is associated with more DNA damage and improves in vivo efficacy with a statistically significant delay in tumor growth and 33% complete regression for the largest Bi/Gd ratio tested. The addition of Bi3+ by our synthetic method leads to nanoparticles that present slightly altered pharmacokinetics and lengthening of the period of high tumor accumulation with no observed evidence of toxicity. Conclusions: We confirmed the safety and enhanced efficacy of AGuIX-Bi with radiation therapy at the selected ratio of 30Gd/70Bi. These results provide crucial evidence towards patient translation.


Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Nanopartículas , Humanos , Medicina de Precisión , Medios de Contraste , Imagen por Resonancia Magnética/métodos , Dosis de Radiación , Nanomedicina Teranóstica/métodos
19.
Blood ; 115(3): 570-80, 2010 Jan 21.
Artículo en Inglés | MEDLINE | ID: mdl-19965678

RESUMEN

Interleukin-21 (IL-21), a member of the IL-2 cytokine family, has diverse regulatory effects on natural killer (NK), T, and B cells. In contrast to other cytokines that are usually immunostimulatory, IL-21 can induce apoptosis of murine B cells at specific activation-differentiation stages. This effect may be used for treatment of B-cell malignancies. Herein we report that diffuse large B-cell lymphoma (DLBCL) cell lines exhibit widespread expression of the IL-21 receptor (IL-21R) and that IL-21 stimulation leads to cell-cycle arrest and caspase-dependent apoptosis. IL-21 also induces apoptosis in de novo DLBCL primary tumors but does not affect viability of human healthy B cells. Furthermore, IL-21 promotes tumor regression and prolongs survival of mice harboring xenograft DLBCL tumors. The antilymphoma effects of this cytokine are dependent on a mechanism involving IL-21-activated signal transducer and activator of transcription 3 (STAT3) up-regulating expression of c-Myc. This up-regulation promotes a decrease in expression of antiapoptotic Bcl-2 and Bcl-X(L) proteins triggering cell death. Our results represent one of the first examples in which the STAT3-c-Myc signaling pathway, which can promote survival and oncogenesis, can induce apoptosis in neoplastic cells. Moreover, based on IL-21's potency in vitro and in animal models, our findings indicate that this cytokine should be examined in clinical studies of DLBCL.


Asunto(s)
Apoptosis/genética , Genes myc , Interleucinas/fisiología , Linfoma de Células B Grandes Difuso/genética , Linfoma de Células B Grandes Difuso/patología , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Línea Celular Tumoral , Doxiciclina/administración & dosificación , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células HeLa , Humanos , Interleucinas/administración & dosificación , Interleucinas/farmacología , Linfoma de Células B Grandes Difuso/tratamiento farmacológico , Ratones , Ratones SCID , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Transducción de Señal/fisiología , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/fisiología , Ensayos Antitumor por Modelo de Xenoinjerto
20.
Blood ; 116(24): 5217-27, 2010 Dec 09.
Artículo en Inglés | MEDLINE | ID: mdl-20844236

RESUMEN

HGAL is a germinal center (GC)-specific gene that negatively regulates lymphocyte motility and whose expression predicts improved survival of patients with diffuse large B-cell lymphoma (DLBCL) and classical Hodgkin lymphoma (cHL). We demonstrate that HGAL serves as a regulator of the RhoA signaling pathway. HGAL enhances activation of RhoA and its down-stream effectors by a novel mechanism - direct binding to the catalytic DH-domain of the RhoA-specific guanine nucleotide exchange factors (RhoGEFs) PDZ-RhoGEF and LARG that stimulate the GDP-GTP exchange rate of RhoA. We delineate the structural domain of HGAL that mediates its interaction with the PDZ-RhoGEF protein. These observations reveal a novel molecular mechanism underlying the inhibitory effects of GC-specific HGAL protein on the motility of GC-derived lymphoma cells. This mechanism may underlie the limited dissemination and better outcome of patients with HGAL-expressing DLBCL and cHL.


Asunto(s)
Movimiento Celular , Linfoma/patología , Proteínas de Neoplasias/fisiología , Proteína de Unión al GTP rhoA/metabolismo , Sitios de Unión , Centro Germinal/patología , Factores de Intercambio de Guanina Nucleótido/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intracelular , Proteínas de Microfilamentos , Unión Proteica , Factores de Intercambio de Guanina Nucleótido Rho , Transducción de Señal
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