RESUMEN
Despite recent therapeutic advances, the 5-year survival rate for adults with acute myeloid leukemia (AML) is poor and standard-of-care chemotherapy is associated with significant toxicity, highlighting the need for new therapeutic approaches. Recent work from our group and others established that the G protein-coupled estrogen receptor (GPER) is tumor suppressive in melanoma and other solid tumors. We performed a preliminary screen of human cancer cell lines from multiple malignancies and found that LNS8801, a synthetic pharmacologic agonist of GPER currently in early phase clinical trials, promoted apoptosis in human AML cells. Using human AML cell lines and primary cells, we show that LNS8801 inhibits human AML in preclinical in vitro models, while not affecting normal mononuclear cells. Although GPER is broadly expressed in normal and malignant myeloid cells, this cancer-specific LNS8801-induced inhibition appeared to be independent of GPER signaling. LNS8801 induced AML cell death primarily through a caspase-dependent apoptosis pathway. This was independent of secreted classical death receptor ligands, and instead required induction of reactive oxygen species (ROS) and activation of endoplasmic reticulum (ER) stress response pathways including IRE1α. These studies demonstrate a novel activity of LNS8801 in AML cells and show that targeting ER stress with LNS8801 may be a useful therapeutic approach for AML. Significance: Previous work demonstrated that LNS8801 inhibits cancer via GPER activation, especially in solid tumors. Here we show that LNS8801 inhibits AML via GPER-independent mechanisms that include ROS induction and ER activation.
Asunto(s)
Endorribonucleasas , Leucemia Mieloide Aguda , Adulto , Humanos , Especies Reactivas de Oxígeno , Proteínas Serina-Treonina Quinasas , Leucemia Mieloide Aguda/tratamiento farmacológico , Estrógenos , Estrés del Retículo EndoplásmicoRESUMEN
Human melanocytic nevi (moles) result from a brief period of clonal expansion of melanocytes. As a cellular defensive mechanism against oncogene-induced hyperplasia, nevus-resident melanocytes enter a senescent state of stable cell cycle arrest. Senescent melanocytes can persist for months in mice and years in humans with a risk to escape the senescent state and progress to melanoma. The mechanisms providing prolonged survival of senescent melanocytes remain poorly understood. Here, we show that senescent melanocytes in culture and in nevi express high level of the anti-apoptotic BCL-2 family member BCL-W but remain insensitive to the pan-BCL-2 inhibitor ABT-263. We demonstrate that resistance to ABT-263 is driven by mTOR-mediated enhanced translation of another anti-apoptotic member, MCL-1. Strikingly, the combination of ABT-263 and MCL-1 inhibitors results in synthetic lethality to senescent melanocytes, and its topical application sufficient to eliminate nevi in male mice. These data highlight the important role of redundant anti-apoptotic mechanisms for the survival advantage of senescent melanocytes, and the proof-of-concept for a non-invasive combination therapy for nevi removal.
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Nevo Pigmentado , Nevo , Neoplasias Cutáneas , Masculino , Humanos , Animales , Ratones , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Melanocitos/metabolismo , Nevo/metabolismo , Neoplasias Cutáneas/metabolismoRESUMEN
Melanoma risk is 30 times higher in people with lightly pigmented skin versus darkly pigmented skin. Using primary human melanocytes representing the full human skin pigment continuum and preclinical melanoma models, we show that cell-intrinsic differences between dark and light melanocytes regulate melanocyte proliferative capacity and susceptibility to malignant transformation, independent of melanin and ultraviolet exposure. These differences result from dihydroxyphenylalanine (DOPA), a melanin precursor synthesized at higher levels in melanocytes from darkly pigmented skin. We used both high-throughput pharmacologic and genetic in vivo CRISPR screens to determine that DOPA limits melanocyte and melanoma cell proliferation by inhibiting the muscarinic acetylcholine receptor M1 (CHRM1) signaling. Pharmacologic CHRM1 antagonism in melanoma leads to depletion of c-Myc and FOXM1, both of which are proliferation drivers associated with aggressive melanoma. In preclinical mouse melanoma models, pharmacologic inhibition of CHRM1 or FOXM1 inhibited tumor growth. CHRM1 and FOXM1 may be new therapeutic targets for melanoma.
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Solid tumors possess heterogeneous metabolic microenvironments where oxygen and nutrient availability are plentiful (fertile regions) or scarce (arid regions). While cancer cells residing in fertile regions proliferate rapidly, most cancer cells in vivo reside in arid regions and exhibit a slow-cycling state that renders them chemoresistant. Here, we developed an in vitro system enabling systematic comparison between these populations via transcriptome analysis, metabolomic profiling, and whole-genome CRISPR screening. Metabolic deprivation led to pronounced transcriptional and metabolic reprogramming, resulting in decreased anabolic activities and distinct vulnerabilities. Reductions in anabolic, energy-consuming activities, particularly cell proliferation, were not simply byproducts of the metabolic challenge, but rather essential adaptations. Mechanistically, Bcl-xL played a central role in the adaptation to nutrient and oxygen deprivation. In this setting, Bcl-xL protected quiescent cells from the lethal effects of cell-cycle entry in the absence of adequate nutrients. Moreover, inhibition of Bcl-xL combined with traditional chemotherapy had a synergistic antitumor effect that targeted cycling cells. Bcl-xL expression was strongly associated with poor patient survival despite being confined to the slow-cycling fraction of human pancreatic cancer cells. These findings provide a rationale for combining traditional cancer therapies that target rapidly cycling cells with those that target quiescent, chemoresistant cells associated with nutrient and oxygen deprivation. SIGNIFICANCE: The majority of pancreatic cancer cells inhabit nutrient- and oxygen-poor tumor regions and require Bcl-xL for their survival, providing a compelling antitumor metabolic strategy.
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Neoplasias Pancreáticas , Proteína bcl-X , Apoptosis , Ciclo Celular , Línea Celular Tumoral , Humanos , Nutrientes , Oxígeno/metabolismo , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Microambiente Tumoral , Proteína bcl-X/metabolismoRESUMEN
NRAS-driven melanomas frequently develop resistance to MAPK/extracellular signal-regulated kinase kinase inhibitors (MEKis), which limits their therapeutic utility. Nguyen et al. (2020) show that MEKi-resistant cells upregulate phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme in serine synthesis. Suppression of PHGDH rendered cells sensitive to MEKis, suggesting that PHGDH may be a therapeutic target for melanoma.
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Melanoma , Preparaciones Farmacéuticas , GTP Fosfohidrolasas , Glutatión , Humanos , Melanoma/tratamiento farmacológico , Melanoma/genética , Proteínas de la Membrana , Quinasas de Proteína Quinasa Activadas por Mitógenos , Fosfoglicerato-Deshidrogenasa/genética , Fosfoglicerato-Deshidrogenasa/metabolismo , Serina/metabolismo , Regulación hacia ArribaRESUMEN
Neuroblastoma is a malignancy of the developing sympathetic nervous system that accounts for 12% of childhood cancer deaths. Like many childhood cancers, neuroblastoma shows a relative paucity of somatic single-nucleotide variants (SNVs) and small insertions and deletions (indels) compared to adult cancers. Here, we assessed the contribution of somatic structural variation (SV) in neuroblastoma using a combination of whole-genome sequencing (WGS) of tumor-normal pairs (n = 135) and single-nucleotide polymorphism (SNP) genotyping of primary tumors (n = 914). Our study design allowed for orthogonal validation and replication across platforms. SV frequency, type, and localization varied significantly among high-risk tumors. MYCN nonamplified high-risk tumors harbored an increased SV burden overall, including a significant excess of tandem duplication events across the genome. Genes disrupted by SV breakpoints were enriched in neuronal lineages and associated with phenotypes such as autism spectrum disorder (ASD). The postsynaptic adapter protein-coding gene, SHANK2, located on Chromosome 11q13, was disrupted by SVs in 14% of MYCN nonamplified high-risk tumors based on WGS and 10% in the SNP array cohort. Expression of SHANK2 was low across human-derived neuroblastoma cell lines and high-risk neuroblastoma tumors. Forced expression of SHANK2 in neuroblastoma cells resulted in significant growth inhibition (P = 2.6 × 10-2 to 3.4 × 10-5) and accelerated neuronal differentiation following treatment with all-trans retinoic acid (P = 3.1 × 10-13 to 2.4 × 10-30). These data further define the complex landscape of somatic structural variation in neuroblastoma and suggest that events leading to deregulation of neurodevelopmental processes, such as inactivation of SHANK2, are key mediators of tumorigenesis in this childhood cancer.
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Genes Supresores de Tumor , Variación Estructural del Genoma , Proteínas del Tejido Nervioso/genética , Neuroblastoma/genética , Neurogénesis/genética , Línea Celular Tumoral , Cromotripsis , Estudios de Cohortes , Roturas del ADN , Variaciones en el Número de Copia de ADN , Femenino , Humanos , Masculino , Proteína Proto-Oncogénica N-Myc/genética , Neuroblastoma/patología , Polimorfismo de Nucleótido Simple , Sitios de Carácter Cuantitativo , ARN Neoplásico , RNA-Seq , Medición de Riesgo , Telomerasa/genética , Células Tumorales Cultivadas , Secuenciación Completa del GenomaRESUMEN
Menin, a protein encoded by the MEN1 gene, suppresses cancers associated with multiple endocrine neoplasia type 1 (MEN1), but promotes the development of a subset of leukemia induced by mixed lineage leukemia (MLL)-derived fusion proteins (MLL-FPs). The crystal structure of menin indicates that it acts as a scaffold protein to bind the N-terminus of MLL via a central pocket. Small molecule menin-MLL inhibitors (MIs) bind the menin pocket to disrupt the menin/MLL interaction, resulting in suppression of MLL-FP-transformed acute myeoloid leukemia (AML). It is thought that MIs suppress the MLL-FP-induced leukemia by blocking the menin/MLL interaction and menin/MLL-induced HOX gene transcription. However, it is not clear whether MIs also affect other aspects of menin biology beyond disruption of the menin/MLL interaction. Here we show for the first time that MIs reduced menin protein levels and decreased the half-life of menin protein but have no effect on mRNA level in MLL-FP-expressing leukemia cells, and proteasome or E1 ligase inhibitor rescued the MI-induced menin degradation. Notably, the MI-induced reduction of H3K4m3 and HOXA9 expression was rescued with a proteasome inhibitor that blocks MI-induced menin protein degradation. Mechanistically, MIs promote the interaction of menin with Hsp70-associated ubiquitin ligase CHIP, resulting in increased menin ubiquitination, leading to increased menin degradation. Together, these findings uncover a novel mechanism whereby small molecule MIs increase menin degradation by triggering the Hsp70/CHIP-mediated ubiquitin-proteasome pathway that ultimately leads to the reduction in HOXA9 gene expression and leukemia suppression.
RESUMEN
Self-renewing somatic tissues depend upon the proper balance of chromatin-modifying enzymes to coordinate progenitor cell maintenance and differentiation, disruption of which can promote carcinogenesis. As a result, drugs targeting the epigenome hold significant therapeutic potential. The histone demethylase, LSD1 (KDM1A), is overexpressed in numerous cancers, including epithelial cancers; however, its role in the skin is virtually unknown. Here we show that LSD1 directly represses master epithelial transcription factors that promote differentiation. LSD1 inhibitors block both LSD1 binding to chromatin and its catalytic activity, driving significant increases in H3K4 methylation and gene transcription of these fate-determining transcription factors. This leads to both premature epidermal differentiation and the repression of squamous cell carcinoma. Together these data highlight both LSD1's role in maintaining the epidermal progenitor state and the potential of LSD1 inhibitors for the treatment of keratinocyte cancers, which collectively outnumber all other cancers combined.
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Diferenciación Celular , Linaje de la Célula , Células Epiteliales/citología , Histona Demetilasas/antagonistas & inhibidores , Factores de Transcripción/metabolismo , Células 3T3 , Adulto , Animales , Sitios de Unión , Carcinoma de Células Escamosas/patología , Diferenciación Celular/genética , Línea Celular Tumoral , Linaje de la Célula/genética , Epidermis/metabolismo , Células Epiteliales/metabolismo , Regulación de la Expresión Génica , Genoma Humano , Histona Demetilasas/metabolismo , Histonas/metabolismo , Humanos , Lisina/metabolismo , Metilación , Ratones , Unión Proteica , Factores de Transcripción de la Familia Snail/metabolismo , Transcripción GenéticaRESUMEN
Hematopoietic stem cell (HSC) homeostasis is controlled by cytokine receptor-mediated Janus kinase 2 (JAK2) signaling. We previously found that JAK2 is promptly ubiquitinated upon cytokine stimulation. Whether a competing JAK2 deubiquitination activity exists is unknown. LNK is an essential adaptor protein that constrains HSC expansion through dampening thrombopoietin (TPO)-induced JAK2 signaling. We show here that a LNK-associated lysine-63 (K63)-deubiquitinating enzyme complex, Brcc36 isopeptidase complex (BRISC), attenuates HSC expansion through control of JAK2 signaling. We pinpoint a direct interaction between the LNK SH2 domain and a phosphorylated tyrosine residue in KIAA0157 (Abraxas2), a unique and defining BRISC component. Kiaa0157 deficiency in mice led to an expansion of phenotypic and functional HSCs. Endogenous JAK2 and phospho-JAK2 were rapidly K63-ubiquitinated upon TPO stimulation, and this action was augmented in cells depleted of the BRISC core components KIAA0157, MERIT40, or BRCC36. This increase in JAK2 ubiquitination after BRISC knockdown was associated with increased TPO-mediated JAK2 activation and protein levels, and increased MPL receptor presence at the cell surface. In addition, BRISC depletion promoted membrane proximal association between the MPL receptor and pJAK2/JAK2, thus enhancing activated JAK2/MPL at the cell membrane. These findings define a novel pathway by which K63-ubiquitination promotes JAK2 stability and activation in a proteasome-independent manner. Moreover, mutations in BRCC36 are found in clonal hematopoiesis in humans. This research may shed light on the mechanistic understanding of a potential role of BRCC36 in human HSCs.