ABSTRACT
Small-cell lung cancer (SCLC) is the most fatal form of lung cancer. Intratumoral heterogeneity, marked by neuroendocrine (NE) and non-neuroendocrine (non-NE) cell states, defines SCLC, but the cell-extrinsic drivers of SCLC plasticity are poorly understood. To map the landscape of SCLC tumor microenvironment (TME), we apply spatially resolved transcriptomics and quantitative mass spectrometry-based proteomics to metastatic SCLC tumors obtained via rapid autopsy. The phenotype and overall composition of non-malignant cells in the TME exhibit substantial variability, closely mirroring the tumor phenotype, suggesting TME-driven reprogramming of NE cell states. We identify cancer-associated fibroblasts (CAFs) as a crucial element of SCLC TME heterogeneity, contributing to immune exclusion, and predicting exceptionally poor prognosis. Our work provides a comprehensive map of SCLC tumor and TME ecosystems, emphasizing their pivotal role in SCLC's adaptable nature, opening possibilities for reprogramming the TME-tumor communications that shape SCLC tumor states.
Subject(s)
Lung Neoplasms , Small Cell Lung Carcinoma , Tumor Microenvironment , Humans , Small Cell Lung Carcinoma/pathology , Small Cell Lung Carcinoma/genetics , Small Cell Lung Carcinoma/metabolism , Lung Neoplasms/pathology , Lung Neoplasms/metabolism , Cancer-Associated Fibroblasts/metabolism , Cancer-Associated Fibroblasts/pathology , Neuroendocrine Tumors/pathology , Neuroendocrine Tumors/genetics , Neuroendocrine Tumors/metabolism , Neuroendocrine Cells/pathology , Neuroendocrine Cells/metabolism , Female , Male , PrognosisABSTRACT
PURPOSE: Tumoral programmed cell death ligand-1 (PD-L1) expression is common in human papillomavirus (HPV)-associated head and neck squamous cell carcinoma (HNSCC). We assessed whether a DNA vaccine targeting HPV-16/18 E6/E7 with IL12 adjuvant (MEDI0457) combined with the PD-L1 inhibitor durvalumab could enhance HPV-specific T-cell response and improve outcomes in recurrent/metastatic HPV-16/18-associated HNSCC. PATIENTS AND METHODS: In this phase Ib/IIa study, immunotherapy-naïve patients with ≥1 previous platinum-containing regimen (neoadjuvant/adjuvant therapy or for recurrent/metastatic disease) received MEDI0457 7 mg intramuscularly with electroporation on weeks 1, 3, 7, and 12, then every 8 weeks, plus durvalumab 1,500 mg intravenously on weeks 4, 8, and 12, then every 4 weeks, until confirmed progression and/or unacceptable toxicity. Coprimary objectives were safety and objective response rate (ORR; H0: ORR ≤ 15%); secondary objectives included 16-week disease control rate (DCR-16), overall survival (OS), and progression-free survival (PFS). RESULTS: Of 35 treated patients, 29 were response evaluable (confirmed HPV-associated disease; received both agents). ORR was 27.6% [95% confidence interval (CI), 12.7-47.2; four complete responses, four partial responses]; responses were independent of PD-L1 tumor-cell expression (≥25% vs. <25%). DCR-16 was 44.8% (95% CI, 26.5-64.3). Median PFS was 3.5 months (95% CI, 1.9-9.0); median OS was 29.2 months (15.2-not calculable). Twenty-eight (80.0%) patients had treatment-related adverse events [grade 3: 5 (14.3%); no grade 4/5], resulting in discontinuation in 2 (5.7%) patients. HPV-16/18-specific T cells increased on treatment; 4 of 8 evaluable patients had a >2-fold increase in tumor-infiltrating CD8+ T cells. CONCLUSIONS: MEDI0457 plus durvalumab was well tolerated. While the primary efficacy endpoint was not reached, clinical benefit was encouraging.
Subject(s)
Head and Neck Neoplasms , Papillomavirus Infections , Humans , Squamous Cell Carcinoma of Head and Neck/drug therapy , Human Papillomavirus Viruses , Head and Neck Neoplasms/drug therapy , Papillomavirus Infections/complications , Human papillomavirus 16/genetics , B7-H1 Antigen/genetics , Human papillomavirus 18ABSTRACT
Cellular senescence is an important contributor to aging and age-related diseases such as Alzheimer's disease (AD). Senescent cells are characterized by a durable cell proliferation arrest and the acquisition of a proinflammatory senescence-associated secretory phenotype (SASP), which participates in the progression of neurodegenerative disorders. Clearance of senescent glial cells in an AD mouse model prevented cognitive decline suggesting pharmacological agents targeting cellular senescence might provide novel therapeutic approaches for AD. Δ133p53α, a natural protein isoform of p53, was previously shown to be a negative regulator of cellular senescence in primary human astrocytes, with clinical implications from its diminished expression in brain tissues from AD patients. Here we show that treatment of proliferating human astrocytes in culture with amyloid-beta oligomers (Aß), an endogenous pathogenic agent of AD, results in reduced expression of Δ133p53α, as well as induces the cells to become senescent and express proinflammatory SASP cytokines such as IL-6, IL-1ß and TNFα. Our data suggest that Aß-induced astrocyte cellular senescence is associated with accelerated DNA damage, and upregulation of full-length p53 and its senescence-inducing target gene p21WAF1. We also show that exogenously enhanced expression of Δ133p53α rescues human astrocytes from Aß-induced cellular senescence and SASP through both protection from DNA damage and dominant-negative inhibition of full-length p53, leading to inhibition of Aß-induced, astrocyte-mediated neurotoxicity. The results presented here demonstrate that Δ133p53α manipulation could modulate cellular senescence in the context of AD, possibly opening new therapeutic avenues.
Subject(s)
Alzheimer Disease , Neurotoxicity Syndromes , Amyloid beta-Peptides , Astrocytes , Cellular Senescence , Humans , Tumor Suppressor Protein p53ABSTRACT
Molecular subtypes of small cell lung cancer (SCLC) defined by the expression of key transcription regulators have recently been proposed in cell lines and limited number of primary tumors. The clinical and biological implications of neuroendocrine (NE) subtypes in metastatic SCLC, and the extent to which they vary within and between patient tumors and in patient-derived models is not known. We integrate histology, transcriptome, exome, and treatment outcomes of SCLC from a range of metastatic sites, revealing complex intra- and intertumoral heterogeneity of NE differentiation. Transcriptomic analysis confirms previously described subtypes based on ASCL1, NEUROD1, POU2F3, YAP1, and ATOH1 expression, and reveal a clinical subtype with hybrid NE and non-NE phenotypes, marked by chemotherapy-resistance and exceedingly poor outcomes. NE tumors are more likely to have RB1, NOTCH, and chromatin modifier gene mutations, upregulation of DNA damage response genes, and are more likely to respond to replication stress targeted therapies. In contrast, patients preferentially benefited from immunotherapy if their tumors were non-NE. Transcriptional phenotypes strongly skew towards the NE state in patient-derived model systems, an observation that was confirmed in paired patient-matched tumors and xenografts. We provide a framework that unifies transcriptomic and genomic dimensions of metastatic SCLC. The marked differences in transcriptional diversity between patient tumors and model systems are likely to have implications in development of novel therapeutic agents.
Subject(s)
Lung Neoplasms , Neuroendocrine Tumors , Small Cell Lung Carcinoma , Gene Expression Regulation, Neoplastic , Humans , Lung Neoplasms/pathology , Neuroendocrine Tumors/genetics , Small Cell Lung Carcinoma/pathology , Transcription Factors/metabolismABSTRACT
Tau accumulation is a core component of Alzheimer's disease and other neurodegenerative tauopathies. While tau's impact on neurons is a major area of research, the effect of extracellular tau on astrocytes is largely unknown. This article summarizes our recent studies showing that astrocyte senescence plays a critical role in neurodegenerative diseases and integrates extracellular tau into the regulatory loop of senescent astrocyte-mediated neurotoxicity. Human astrocytes in vitro undergoing senescence were shown to acquire the inflammatory senescence-associated secretory phenotype (SASP) and toxicity to neurons, which may recapitulate aging- and disease-associated neurodegeneration. Here, we show that human astrocytes exposed to extracellular tau in vitro also undergo cellular senescence and acquire a neurotoxic SASP (e.g. IL-6 secretion), with oxidative stress response (indicated by upregulated NRF2 target genes) and a possible activation of inflammasome (indicated by upregulated ASC and IL-1ß). These findings suggest that senescent astrocytes induced by various conditions and insults, including tau exposure, may represent a therapeutic target to inhibit or delay the progression of neurodegenerative diseases. We also discuss the pathological activity of extracellular tau in microglia and astrocytes, the disease relevance and diversity of tau forms, therapeutics targeting senescence in neurodegeneration, and the roles of p53 and its isoforms in astrocyte-mediated neurotoxicity and neuroprotection.
Subject(s)
Astrocytes/metabolism , Cellular Senescence/physiology , Neurodegenerative Diseases/metabolism , Senescence-Associated Secretory Phenotype/physiology , tau Proteins/toxicity , Astrocytes/drug effects , Astrocytes/pathology , Cells, Cultured , Cellular Senescence/drug effects , Humans , Neurodegenerative Diseases/chemically induced , Neurodegenerative Diseases/pathology , Senescence-Associated Secretory Phenotype/drug effectsABSTRACT
BACKGROUND: Chromosomal inversions involving anaplastic lymphoma kinase (ALK) and echinoderm microtubule associated protein like 4 (EML4) generate a fusion protein EML4-ALK in non-small cell lung cancer (NSCLC). The understanding of EML4-ALK function can be improved by a functional study using normal human cells. METHODS: Here we for the first time conduct such study to examine the effects of EML4-ALK on cell proliferation, cellular senescence, DNA damage, gene expression profiles and transformed phenotypes. RESULTS: The lentiviral expression of EML4-ALK in mortal, normal human fibroblasts caused, through its constitutive ALK kinase activity, an early induction of cellular senescence with accumulated DNA damage, upregulation of p16INK4A and p21WAF1, and senescence-associated ß-galactosidase (SA-ß-gal) activity. In contrast, when EML4-ALK was expressed in normal human fibroblasts transduced with telomerase reverse transcriptase (hTERT), which is activated in the vast majority of NSCLC, the cells showed accelerated proliferation and acquired anchorage-independent growth ability in soft-agar medium, without accumulated DNA damage, chromosome aberration, nor p53 mutation. EML4-ALK induced the phosphorylation of STAT3 in both mortal and hTERT-transduced cells, but RNA sequencing analysis suggested that the different signaling pathways contributed to the different phenotypic outcomes in these cells. While EML4-ALK also induced anchorage-independent growth in hTERT-immortalized human bronchial epithelial cells in vitro, the expression of EML4-ALK alone did not cause detectable in vivo tumorigenicity in immunodeficient mice. CONCLUSIONS: Our data indicate that the expression of hTERT is critical for EML4-ALK to manifest its in vitro transforming activity in human cells. This study provides the isogenic pairs of human cells with and without EML4-ALK expression.
Subject(s)
Carcinogenesis/genetics , Carcinoma, Non-Small-Cell Lung/genetics , Lung Neoplasms/genetics , Oncogene Proteins, Fusion/metabolism , Telomerase/metabolism , Animals , Carcinogenesis/pathology , Carcinoma, Non-Small-Cell Lung/pathology , Cell Line , Cell Proliferation/genetics , Cellular Senescence/genetics , DNA Damage , Disease Models, Animal , Epithelial Cells , Female , Fibroblasts , Gene Expression Regulation, Neoplastic , Genetic Vectors/genetics , Humans , Lentivirus/genetics , Lung Neoplasms/pathology , Mice , Oncogene Proteins, Fusion/genetics , RNA-Seq , Telomerase/genetics , Telomere Homeostasis/genetics , TransfectionABSTRACT
OBJECTIVES: Surgery with curative intent is the standard treatment for stage I lung adenocarcinoma. However, disease recurrence occurs in a third of patients. Prognostic biomarkers are needed to improve postoperative management. Here, we evaluate the utility of Homeobox A9 (HOXA9) promoter methylation, alone or in combination with Blood Vessel Invasion (BVI) assessment, for prognostic stratification of stage I lung adenocarcinoma patients. MATERIALS AND METHODS: We developed a Droplet Digital PCR (ddPCR) assay to measure HOXA9 promoter methylation in formalin-fixed paraffin-embedded (FFPE) biospecimens generated during routine pathology. The prognostic value of HOXA9 promoter methylation and BVI, alone and in combination, was evaluated by Kaplan-Meier survival and Cox regression analyses in a cohort of 177 stage I lung adenocarcinoma patients from the NCI-MD study. RESULTS: The ddPCR assay showed linearity, sensitivity and specificity for measuring HOXA9 promoter methylation down to 0.1% methylated DNA input. The HOXA9 promoter was methylated de novo in FFPE tumors (Pâ¯<â¯0.0001). High methylation was independently associated with worse cancer-specific survival (Hazard Ratio [HR], 3.37; Pâ¯=â¯0.0002) and identified high-risk stage IA and IB patients. Addition of this molecular marker improved a risk model comprised of clinical and pathologic parameters (age, gender, race, stage, and smoking history; nested likelihood ratio test; Pâ¯=â¯0.0004) and increased the C-index from 0.60 (95% CI 0.51-0.69) to 0.68 (0.60-0.76). High methylation tumors displayed high frequency of TP53 mutations and other molecular characteristics associated with aggressiveness. BVI was independently associated with poor outcome (HR, 2.62; Pâ¯=â¯0.054). A score that combined BVI with HOXA9 promoter methylation further stratified high-risk patients (trend Pâ¯=â¯0.0001 comparing 0, 1 or 2 positive markers). CONCLUSIONS: ddPCR can be used to quantify HOXA9 promoter methylation in FFPE samples. Alone or combined with BVI in a prognostic classifier, HOXA9 promoter methylation could potentially inform the clinical management of patients with early-stage lung adenocarcinoma.
Subject(s)
Adenocarcinoma/genetics , Biomarkers, Tumor/metabolism , Blood Vessels/pathology , Homeodomain Proteins/genetics , Lung Neoplasms/genetics , Promoter Regions, Genetic/genetics , Adenocarcinoma/diagnosis , Adenocarcinoma/mortality , Aged , Case-Control Studies , Cohort Studies , DNA Methylation , Female , Humans , Lung Neoplasms/diagnosis , Lung Neoplasms/mortality , Male , Middle Aged , Neoplasm Invasiveness , Neoplasm Recurrence, Local , Neoplasm Staging , Prognosis , Survival AnalysisABSTRACT
Cellular senescence is a hallmark of normal aging and aging-related syndromes, including the premature aging disorder Hutchinson-Gilford Progeria Syndrome (HGPS), a rare genetic disorder caused by a single mutation in the LMNA gene that results in the constitutive expression of a truncated splicing mutant of lamin A known as progerin. Progerin accumulation leads to increased cellular stresses including unrepaired DNA damage, activation of the p53 signaling pathway and accelerated senescence. We previously established that the p53 isoforms ∆133p53 and p53ß regulate senescence in normal human cells. However, their role in premature aging is unknown. Here we report that p53 isoforms are expressed in primary fibroblasts derived from HGPS patients, are associated with their accelerated senescence and that their manipulation can restore the replication capacity of HGPS fibroblasts. We found that in near-senescent HGPS fibroblasts, which exhibit low levels of ∆133p53 and high levels of p53ß, restoration of Δ133p53 expression was sufficient to extend replicative lifespan and delay senescence, despite progerin levels and abnormal nuclear morphology remaining unchanged. Conversely, Δ133p53 depletion or p53ß overexpression accelerated the onset of senescence in otherwise proliferative HGPS fibroblasts. Our data indicate that Δ133p53 exerts its role by modulating full-length p53 (FLp53) signaling to extend the replicative lifespan and promotes the repair of spontaneous progerin-induced DNA double-strand breaks (DSBs). We showed that Δ133p53 dominant-negative inhibition of FLp53 occurs directly at the p21/CDKN1A and miR-34a promoters, two p53 senescence-associated genes. In addition, Δ133p53 expression increased the expression of DNA repair RAD51, likely through upregulation of E2F1, a transcription factor that activates RAD51, to promote repair of DSBs. In summary, our data indicate that Δ133p53 modulates p53 signaling to repress progerin-induced early onset of senescence in HGPS cells. Therefore, restoration of ∆133p53 expression may be a novel therapeutic strategy to treat aging-associated phenotypes of HGPS in vivo.
Subject(s)
Aging, Premature/genetics , Cellular Senescence/genetics , Fibroblasts/physiology , Tumor Suppressor Protein p53/physiology , Aging, Premature/pathology , Cells, Cultured , DNA Damage/genetics , Fibroblasts/pathology , Humans , Progeria/genetics , Progeria/pathology , Protein Isoforms/physiology , Time Factors , Tumor Suppressor Protein p53/geneticsABSTRACT
BACKGROUND: Phase I experts recommend revisiting dose-limiting toxicity (DLT) definition to include chronic and cumulative toxicities induced by new molecularly targeted therapies. Patient's assessment of late toxicities' tolerability is, however, unknown. MATERIALS AND METHODS: A prospective survey on adverse events (AEs) tolerability on 23 National Cancer InstituteCommon Terminology Criteria for Adverse Event, Version 4 (NCI-CTCAE.v4) items was conducted at Gustave Roussy's Phase I department. Patients' maximum tolerability duration was recorded at baseline, during trial and at trial completion. Results were compared with the corresponding physicians' survey. RESULTS: 52 patients enrolled in 27 Phase I trials between May 2014 and November 2015 completed 102 forms. At baseline, the most feared G2/G3 AEs were haematuria (74%), vomiting (71%) and hyperglycemia (64%)/dry mouth (94%), hyperglycemia (92%) and vomiting (92%). At trial completion, the most feared G2/G3 AEs were personality change (83.3%), haematuria (82%) and fever (80%)/dry mouth, fever and dizziness (100% each). Tolerability score did not differ over time. More previous treatments and occurrence of severe AEs were associated with better tolerability at study completion (p=0.0234 and p=0.0153, respectively, in multivariate analysis). Patient's tolerability differed from physician's assessment. CONCLUSION: AEs considered intolerable by patients are toxicities that directly impact their quality of life and differ from those feared by physicians or included in DLT definition. Patient-reported tolerability of AEs may help in optimising drug development.
ABSTRACT
GPR139 is an orphan class A G protein-coupled receptor found mainly in the central nervous system. It has its highest expression levels in the hypothalamus and striatum, regions regulating metabolism and locomotion, respectively, and has therefore been suggested as a potential target for obesity and Parkinson's disease. The two aromatic amino acids L-Trp and L-Phe have been proposed as putative endogenous agonists, and three structurally related benzohydrazide, glycine benzamide, and benzotriazine surrogate agonist series have been published. Herein, we assayed 158 new analogues selected from a pharmacophore model, and identified 12 new GPR139 agonists, containing previously untested bioisosteres. Furthermore, we present the first combined structure-activity relationships, and a refined pharmacophore model to serve as a rationale for future ligand identification and optimization.
Subject(s)
Hydrazines/chemistry , Models, Chemical , Models, Molecular , Nerve Tissue Proteins/agonists , Nerve Tissue Proteins/chemistry , Receptors, G-Protein-Coupled/agonists , Receptors, G-Protein-Coupled/chemistry , Triazines/chemistry , Animals , CHO Cells , Cricetulus , Humans , Nerve Tissue Proteins/genetics , Receptors, G-Protein-Coupled/genetics , Structure-Activity RelationshipABSTRACT
Lung cancer is the leading cause of cancer-related deaths worldwide. Recent implementation of low-dose computed tomography (LDCT) screening is predicted to lead to diagnosis of lung cancer at an earlier stage, with survival benefit. However, there is still a pressing need for biomarkers that will identify individuals eligible for screening, as well as improve the diagnostic accuracy of LDCT. In addition, biomarkers for prognostic stratification of patients with early stage disease, and those that can be used as surrogates to monitor tumor evolution, will greatly improve clinical management. Molecular alterations found in the DNA of tumor cells, such as mutations, translocations and methylation, are reflected in DNA that is released from the tumor into the bloodstream. Thus, in recent years, circulating tumor DNA (ctDNA) has gained increasing attention as a noninvasive alternative to tissue biopsies and potential surrogate for the entire tumor genome. Activating gene mutations found in ctDNA have been proven effective in predicting response to targeted therapy. Analysis of ctDNA is also a valuable tool for longitudinal follow-up of cancer patients that does not require serial biopsies and may anticipate the acquisition of resistance. DNA methylation has also emerged as a promising marker for early detection, prognosis and real-time follow-up of tumor dynamics that is independent of the genomic composition of the primary tumor. This review summarizes the various investigational applications of methylated ctDNA in lung cancer reported to date. It also provides a brief overview of the technologies for analysis of DNA methylation in liquid biopsies, and the challenges that befall the implementation of methylated ctDNA into routine clinical practice.
ABSTRACT
Several lines of evidence indicate that whole-genome duplication resulting in tetraploidy facilitates carcinogenesis by providing an intermediate and metastable state more prone to generate oncogenic aneuploidy. Here, we report a novel strategy to preferentially kill tetraploid cells based on the abrogation of the spindle assembly checkpoint (SAC) via the targeting of TTK protein kinase (better known as monopolar spindle 1, MPS1). The pharmacological inhibition as well as the knockdown of MPS1 kills more efficiently tetraploid cells than their diploid counterparts. By using time-lapse videomicroscopy, we show that tetraploid cells do not survive the aborted mitosis due to SAC abrogation upon MPS1 depletion. On the contrary diploid cells are able to survive up to at least two more cell cycles upon the same treatment. This effect might reflect the enhanced difficulty of cells with whole-genome doubling to tolerate a further increase in ploidy and/or an elevated level of chromosome instability in the absence of SAC functions. We further show that MPS1-inhibited tetraploid cells promote mitotic catastrophe executed by the intrinsic pathway of apoptosis, as indicated by the loss of mitochondrial potential, the release of the pro-apoptotic cytochrome c from mitochondria, and the activation of caspases. Altogether, our results suggest that MPS1 inhibition could be used as a therapeutic strategy for targeting tetraploid cancer cells.
Subject(s)
Cell Cycle Proteins/metabolism , Protein Serine-Threonine Kinases/metabolism , Protein-Tyrosine Kinases/metabolism , Tetraploidy , Apoptosis/drug effects , Apoptosis/genetics , Cell Cycle Proteins/antagonists & inhibitors , Cell Cycle Proteins/genetics , Cell Line, Tumor , Cell Survival/drug effects , Cell Survival/genetics , Diploidy , HCT116 Cells , Humans , Immunoblotting , M Phase Cell Cycle Checkpoints/drug effects , M Phase Cell Cycle Checkpoints/genetics , Microscopy, Fluorescence , Mitosis/drug effects , Mitosis/genetics , Morpholines/pharmacology , Neoplasms/genetics , Neoplasms/metabolism , Neoplasms/pathology , Nocodazole/pharmacology , Paclitaxel/pharmacology , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Protein Serine-Threonine Kinases/genetics , Protein-Tyrosine Kinases/antagonists & inhibitors , Protein-Tyrosine Kinases/genetics , Purines/pharmacology , RNA Interference , Time-Lapse Imaging/methods , Tubulin Modulators/pharmacologyABSTRACT
The selection of human cancer cell lines in cis-diamminedichloroplatinum(II) (CDDP, best known as cisplatin) is accompanied by stereotyped alterations that contribute to the acquisition of a CDDP-resistant state. Thus, CDDP resistance often leads to the upregulation of the DNA repair enzyme poly (ADP-ribose) polymerase-1 (PARP1) with the consequent intracellular accumulation of poly (ADP-ribose) (PAR)-modified proteins. Here we report another frequent alteration accompanying CDDP resistance, namely upregulation of the antiapoptotic BCL-2 family protein MCL-1. Six out of 8 CDDP resistant cancer cell lines manifested an increase in MCL-1 protein expression level, while only a minority of cell lines overexpressed BCL-2 or BCL-XL. BCL-XL was decreased in six out of 8 cancer cell lines. Importantly, MCL-1 overexpressing, CDDP resistant cells appear to be 'addicted' to MCL-1 because they died upon depletion of MCL-1 by RNA interference or pharmacological inhibition of MCL-1 expression by the BH3 mimetic obatoclax. Knockdown of PARP1 did not succeed in reducing MCL-1 expression, while depletion or inhibition of MCL-1 failed to affect the activity of PARP1. Hence, the two resistance mechanisms are not linked to each other by a direct cause-effect relationship. Importantly, CDDP-resistant, MCL-1 overexpressing human non-small cell lung cancers responded to monotherapy with obatoclax in vivo, in xenotransplanted mice, underscoring the probable therapeutic relevance of these findings.
Subject(s)
Antineoplastic Agents/therapeutic use , Carcinoma, Non-Small-Cell Lung/drug therapy , Cisplatin/therapeutic use , Drug Resistance, Neoplasm/physiology , Gene Expression Regulation, Neoplastic/physiology , Myeloid Cell Leukemia Sequence 1 Protein/metabolism , Animals , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cisplatin/pharmacology , Humans , Indoles , Mice , Myeloid Cell Leukemia Sequence 1 Protein/genetics , Poly (ADP-Ribose) Polymerase-1 , Poly(ADP-ribose) Polymerases/genetics , Poly(ADP-ribose) Polymerases/metabolism , Pyrroles/pharmacology , Pyrroles/therapeutic use , RNA InterferenceABSTRACT
Tetraploidy constitutes a genomically metastable state that can lead to aneuploidy and genomic instability. Tetraploid cells are frequently found in preneoplastic lesions, including intestinal cancers arising due to the inactivation of the tumor suppressor adenomatous polyposis coli (APC). Using a phenotypic screen, we identified resveratrol as an agent that selectively reduces the fitness of tetraploid cells by slowing down their cell cycle progression and by stimulating the intrinsic pathway of apoptosis. Selective killing of tetraploid cells was observed for a series of additional agents that indirectly or directly stimulate AMP-activated protein kinase (AMPK) including salicylate, whose chemopreventive action has been established by epidemiological studies and clinical trials. Both resveratrol and salicylate reduced the formation of tetraploid or higher-order polyploid cells resulting from the culture of human colon carcinoma cell lines or primary mouse epithelial cells lacking tumor protein p53 (TP53, best known as p53) in the presence of antimitotic agents, as determined by cytofluorometric and videomicroscopic assays. Moreover, oral treatment with either resveratrol or aspirin, the prodrug of salicylate, repressed the accumulation of tetraploid intestinal epithelial cells in the Apc(Min/+) mouse model of colon cancer. Collectively, our results suggest that the chemopreventive action of resveratrol and aspirin involves the elimination of tetraploid cancer cell precursors.
Subject(s)
Adenomatous Polyposis Coli/prevention & control , Aspirin/therapeutic use , Cell Death/drug effects , Epithelial Cells/drug effects , Stilbenes/therapeutic use , Tetraploidy , Animals , Aspirin/pharmacology , Cell Line, Tumor , Epithelial Cells/chemistry , Flow Cytometry , Image Processing, Computer-Assisted , In Situ Hybridization, Fluorescence , Mice , Mice, Inbred C57BL , Microscopy, Video , Resveratrol , Stilbenes/pharmacologyABSTRACT
Non-small cell lung carcinoma patients are frequently treated with cisplatin (CDDP), most often yielding temporary clinical responses. Here, we show that PARP1 is highly expressed and constitutively hyperactivated in a majority of human CDDP-resistant cancer cells of distinct histologic origin. Cells manifesting elevated intracellular levels of poly(ADP-ribosyl)ated proteins (PAR(high)) responded to pharmacologic PARP inhibitors as well as to PARP1-targeting siRNAs by initiating a DNA damage response that translated into cell death following the activation of the intrinsic pathway of apoptosis. Moreover, PARP1-overexpressing tumor cells and xenografts displayed elevated levels of PAR, which predicted the response to PARP inhibitors in vitro and in vivo more accurately than PARP1 expression itself. Thus, a majority of CDDP-resistant cancer cells appear to develop a dependency to PARP1, becoming susceptible to PARP inhibitor-induced apoptosis.
Subject(s)
Antineoplastic Agents/pharmacology , Carcinoma, Non-Small-Cell Lung/pathology , Cisplatin/pharmacology , Drug Resistance, Neoplasm , Lung Neoplasms/pathology , Poly(ADP-ribose) Polymerase Inhibitors , Animals , Apoptosis/drug effects , Blotting, Western , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/metabolism , Cell Proliferation/drug effects , Female , Humans , Lung Neoplasms/drug therapy , Lung Neoplasms/metabolism , Mice , Mice, Nude , Phenanthrenes/pharmacology , Poly (ADP-Ribose) Polymerase-1 , Poly(ADP-ribose) Polymerases/genetics , Poly(ADP-ribose) Polymerases/metabolism , RNA, Messenger/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
The BCL-2 homolog BCL-XL, one of the two protein products of BCL2L1, has originally been characterized for its prominent prosurvival functions. Similar to BCL-2, BCL-XL binds to its multidomain proapoptotic counterparts BAX and BAK, hence preventing the formation of lethal pores in the mitochondrial outer membrane, as well as to multiple BH3-only proteins, thus interrupting apical proapoptotic signals. In addition, BCL-XL has been suggested to exert cytoprotective functions by sequestering a cytosolic pool of the pro-apoptotic transcription factor p53 and by binding to the voltage-dependent anion channel 1 (VDAC1), thereby inhibiting the so-called mitochondrial permeability transition (MPT). Thus, BCL-XL appears to play a prominent role in the regulation of multiple distinct types of cell death, including apoptosis and regulated necrosis. More recently, great attention has been given to the cell death-unrelated functions of BCL-2-like proteins. In particular, BCL-XL has been shown to modulate a number of pathophysiological processes, including-but not limited to-mitochondrial ATP synthesis, protein acetylation, autophagy and mitosis. In this short review article, we will discuss the functions of BCL-XL at the interface between cell death and metabolism.
ABSTRACT
The antineoplastic agent cis-diammineplatinum(II) dichloride (cisplatin, CDDP) is part of the poorly effective standard treatment of non-small cell lung carcinoma (NSCLC). Here, we report a novel strategy to improve the efficacy of CDDP. In conditions in which CDDP alone or either of two PARP inhibitors, PJ34 hydrochloride hydrate or CEP 8983, used as standalone treatments were inefficient in killing NSCLC cells, the combination of CDDP plus PJ34 or that of CDDP plus CEP 8983 were found to kill a substantial fraction of the cells. This cytotoxic synergy could be recapitulated by combining CDDP and the siRNA-mediated depletion of the principal PARP isoform, PARP1, indicating that it is mediated by on-target effects of PJ34 or CEP 8983. CDDP and PARP inhibitors synergized in inducing DNA damage foci, mitochondrial membrane permeabilization leading to cytochrome c release, and dissipation of the inner transmembrane potential, caspase activation, plasma membrane rupture and loss of clonogenic potential in NSCLC cells. Collectively, our results indicate that CDDP can be advantageously combined with PARP inhibitors to kill several NSCLC cell lines, independently from their p53 status. Combined treatment with CDDP and PARP inhibitors elicits the intrinsic pathway of apoptosis.
Subject(s)
Carcinoma, Non-Small-Cell Lung/drug therapy , Cisplatin/pharmacology , Enzyme Inhibitors/pharmacology , Lung Neoplasms/drug therapy , Poly(ADP-ribose) Polymerase Inhibitors , Antineoplastic Agents/pharmacology , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Apoptosis/drug effects , Carbazoles/pharmacology , Carcinoma, Non-Small-Cell Lung/metabolism , Cell Line, Tumor , Cell Membrane/drug effects , Cytochromes c/metabolism , DNA Damage/drug effects , Drug Synergism , Humans , Lung Neoplasms/metabolism , Mitochondrial Membranes/drug effects , Phenanthrenes/pharmacology , Phthalimides/pharmacology , Poly (ADP-Ribose) Polymerase-1 , Poly(ADP-ribose) Polymerases/genetics , RNA Interference , RNA, Small InterferingABSTRACT
Circumstantial evidence suggests that colon carcinogenesis can ensue the transient tetraploidization of (pre-)malignant cells. In line with this notion, the tumor suppressors APC and TP53, both of which are frequently inactivated in colon cancer, inhibit tetraploidization in vitro and in vivo. Here, we show that-contrarily to their wild-type counterparts- Tp53 (-/-) colonocytes are susceptible to drug-induced or spontaneous tetraploidization in vitro. Colon organoids generated from tetraploid Tp53 (-/-) cells exhibit a close-to-normal morphology as compared to their diploid Tp53 (-/-) counterparts, yet the colonocytes constituting these organoids are characterized by an increased cell size and an elevated expression of the immunostimulatory protein calreticulin on the cell surface. The subcutaneous injection of tetraploid Tp53 (-/-) colon organoids led to the generation of proliferating tumors in immunodeficient, but not immunocompetent, mice. Thus, tetraploid Tp53 (-/-) colonocytes fail to survive in immunocompetent mice and develop neoplastic lesions in immunocompromised settings only. These results suggest that tetraploidy is particularly oncogenic in the context of deficient immunosurveillance.