Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 22
Filter
1.
Ann Surg ; 2024 Jul 08.
Article in English | MEDLINE | ID: mdl-38975668

ABSTRACT

OBJECTIVE: This study focuses on dose-response investigation using a codon-optimized and de novo-synthesized E-Selectin/AAV2 (E-Sel/AAV2) vector in preparation for Investigational New Drug (IND)-enabling of subsequent clinical studies. BACKGROUND: Gene therapy is a potential solution for patients suffering from chronic limb-threatening ischemia (CLTI). Understanding the dose for effective gene delivery is crucial for future IND-enabling studies. METHODS: Expression of the codon-optimized E-Selectin gene was assessed by flow cytometry following in vitro cell transfection assay and RT-qPCR for murine limbs injected in vivo with AAV-m-E-Selectin (E-Sel/AAV2). Dose-response studies involved three cohorts of FVB/NJ mice (n=6/group) with escalating log doses of E-Selectin/AAV2 injected intramuscularly (IM) in divided aliquots, ranging from 2×109 VG to 2×1011 VG, into ischemic limbs created by left femoral artery/vein ligation/excision and administration of nitric oxide synthase inhibitor, L-NAME. Limb perfusion, extent of gangrene free limb, functional limb recovery and therapeutic angiogenesis were assessed. RESULTS: Codon-optimized E-Sel/AAV2 gene therapy exhibits superior expression level than WT E-Sel/AAV2 gene therapy both in vitro and in vivo. Mice treated with a high dose (2×1011 VG) of E-Sel/AAV2 showed significantly improved perfusion indices, lower Faber's scores, increased running stamina and neovascularization compared with lower doses tested with control groups, indicating a distinct dose-dependent response. No toxicity was detected in any of the animal groups studied. CONCLUSION: E-Sel/AAV2 Vascular Regeneration Gene Therapy (VRGT) holds promise for enhancing the recovery of ischemic hindlimb perfusion and function, with the effective dose identified in this study as 2×1011 VG aliquots injected IM.

2.
Int J Mol Sci ; 25(4)2024 Feb 11.
Article in English | MEDLINE | ID: mdl-38396848

ABSTRACT

Fibroblasts are stromal cells ubiquitously distributed in the body of nearly every organ tissue. These cells were previously considered to be "passive cells", solely responsible for ensuring the turnover of the extracellular matrix (ECM). However, their versatility, including their ability to switch phenotypes in response to tissue injury and dynamic activity in the maintenance of tissue specific homeostasis and integrity have been recently revealed by the innovation of technological tools such as genetically modified mouse models and single cell analysis. These highly plastic and heterogeneous cells equipped with multifaceted functions including the regulation of angiogenesis, inflammation as well as their innate stemness characteristics, play a central role in the delicately regulated process of wound healing. Fibroblast dysregulation underlies many chronic conditions, including cardiovascular diseases, cancer, inflammatory diseases, and diabetes mellitus (DM), which represent the current major causes of morbidity and mortality worldwide. Diabetic foot ulcer (DFU), one of the most severe complications of DM affects 40 to 60 million people. Chronic non-healing DFU wounds expose patients to substantial sequelae including infections, gangrene, amputation, and death. A complete understanding of the pathophysiology of DFU and targeting pathways involved in the dysregulation of fibroblasts are required for the development of innovative new therapeutic treatments, critically needed for these patients.


Subject(s)
Diabetes Mellitus , Diabetic Foot , Animals , Mice , Humans , Diabetic Foot/therapy , Fibroblasts/metabolism , Extracellular Matrix/metabolism , Chronic Disease , Disease Progression , Diabetes Mellitus/metabolism
3.
Front Cardiovasc Med ; 10: 1236345, 2023.
Article in English | MEDLINE | ID: mdl-37600026

ABSTRACT

Cardiovascular diseases (CVD) continue to be the leading cause of morbidity and mortality globally and claim the lives of over 17 million people annually. Current management of CVD includes risk factor modification and preventative strategies including dietary and lifestyle changes, smoking cessation, medical management of hypertension and cholesterol lipid levels, and even surgical revascularization procedures if needed. Although these strategies have shown therapeutic efficacy in reducing major adverse cardiovascular events such as heart attack, stroke, symptoms of chronic limb-threatening ischemia (CLTI), and major limb amputation significant compliance by patients and caregivers is required and off-target effects from systemic medications can still result in organ dysfunction. Stem cell therapy holds major potential for CVD applications but is limited by the low quantities of cells that are able to traffic to and engraft at diseased tissue sites. New preclinical investigations have been undertaken to modify mesenchymal stem cells (MSCs) to achieve targeted cell delivery after systemic administration. Although previous reviews have focused broadly on the modification of MSCs for numerous local or intracoronary administration strategies, here we review recent preclinical advances related to overcoming challenges imposed by the high velocity and dynamic flow of the circulatory system to specifically deliver MSCs to ischemic cardiac and peripheral tissue sites. Many of these technologies can also be applied for the targeted delivery of other types of therapeutic cells for treating various diseases.

4.
J Surg Res ; 291: 496-506, 2023 11.
Article in English | MEDLINE | ID: mdl-37536191

ABSTRACT

INTRODUCTION: The utility of incidental appendectomy (IA) during many ovarian operations has not been evaluated in the pediatric population. This study sought to compare outcomes after ovarian surgery with IA in the pediatric population. METHODS: Females (≤20 y old) undergoing ovarian surgeries (oophorectomy, detorsion and/or drainage) were identified from the Nationwide Readmissions Database (2016-2018). Those with appendicitis were excluded. A propensity score-matched analysis (PSMA) with 46 covariates (demographics, comorbidities, hospitalization factors, etc.) was performed between those receiving ovarian surgery with or without IA. RESULTS: There were 13,202 females (median age 17 [IQR 14-20] y old) who underwent oophorectomy (90%), detorsion (26%), and/or ovarian drainage (13%). There were more episodes of torsion in the PSMA cohort receiving ovarian surgery alone (17% versus 10% IA; P = 0.016), while other indications (ovarian mass, cyst) were similar. Open (66% versus 34% laparoscopic) IAs were more frequent. Length of stay (LOS) was longer for those undergoing IA (3 [2-4] versus 2 [2-4] days ovarian surgery alone; P < 0.001). There was a higher rate of postoperative GI complications in the IA cohort. Subgroup analysis of those undergoing laparoscopic operations demonstrated no difference in LOS or postoperative complications between patients undergoing IA or not. CONCLUSIONS: These data indicate that IA in pediatric ovarian operations is associated with longer LOS and higher GI postoperative complications. However, laparoscopic IA was not associated with higher cost, complications, LOS, or readmissions. This suggests that IA performed during ovarian surgeries in select patients may be cost-effective and worthy of future study.


Subject(s)
Appendicitis , Laparoscopy , Female , Humans , Child , Adolescent , Appendectomy/adverse effects , Retrospective Studies , Appendicitis/surgery , Appendicitis/complications , Postoperative Complications/epidemiology , Postoperative Complications/etiology , Postoperative Complications/surgery , Hospitalization , Length of Stay , Laparoscopy/adverse effects
5.
Ann Surg ; 278(3): 383-395, 2023 09 01.
Article in English | MEDLINE | ID: mdl-37334717

ABSTRACT

OBJECTIVE: Here, we report a new method to increase the therapeutic potential of mesenchymal stem/stromal cells (MSCs) for ischemic wound healing. We tested biological effects of MSCs modified with E-selectin, a cell adhesion molecule capable of inducing postnatal neovascularization, on a translational murine model. BACKGROUND: Tissue loss significantly worsens the risk of extremity amputation for patients with chronic limb-threatening ischemia. MSC-based therapeutics hold major promise for wound healing and therapeutic angiogenesis, but unmodified MSCs demonstrate only modest benefits. METHODS: Bone marrow cells harvested from FVB/ROSA26Sor mTmG donor mice were transduced with E-selectin-green fluorescent protein (GFP)/AAV-DJ or GFP/AAV-DJ (control). Ischemic wounds were created via a 4 mm punch biopsy in the ipsilateral limb after femoral artery ligation in recipient FVB mice and subsequently injected with phosphate-buffered saline or 1×10 6 donor MSC GFP or MSC E-selectin-GFP . Wound closure was monitored daily for 7 postoperative days, and tissues were harvested for molecular and histologic analysis and immunofluorescence. Whole-body DiI perfusion and confocal microscopy were utilized to evaluate wound angiogenesis. RESULTS: Unmodified MSCs do not express E-selectin, and MSC E-selectin-GFP gain stronger MSC phenotype yet maintain trilineage differentiation and colony-forming capability. MSC E-selectin-GFP therapy accelerates wound healing compared with MSC GFP and phosphate-buffered saline treatment. Engrafted MSC E-selectin-GFP manifest stronger survival and viability in wounds at postoperative day 7. Ischemic wounds treated with MSC E-selectin-GFP exhibit more abundant collagen deposition and enhanced angiogenic response. CONCLUSIONS: We establish a novel method to potentiate regenerative and proangiogenic capability of MSCs by modification with E-selectin/adeno-associated virus. This innovative therapy carries the potential as a platform worthy of future clinical studies.


Subject(s)
E-Selectin , Mesenchymal Stem Cell Transplantation , Mice , Animals , Wound Healing/physiology , Extremities , Phosphates/pharmacology
6.
Cardiovasc Ther ; 2023: 6679390, 2023.
Article in English | MEDLINE | ID: mdl-37251271

ABSTRACT

The response to ischemia in peripheral artery disease (PAD) depends on compensatory neovascularization and coordination of tissue regeneration. Identifying novel mechanisms regulating these processes is critical to the development of nonsurgical treatments for PAD. E-selectin is an adhesion molecule that mediates cell recruitment during neovascularization. Therapeutic priming of ischemic limb tissues with intramuscular E-selectin gene therapy promotes angiogenesis and reduces tissue loss in a murine hindlimb gangrene model. In this study, we evaluated the effects of E-selectin gene therapy on skeletal muscle recovery, specifically focusing on exercise performance and myofiber regeneration. C57BL/6J mice were treated with intramuscular E-selectin/adeno-associated virus serotype 2/2 gene therapy (E-sel/AAV) or LacZ/AAV2/2 (LacZ/AAV) as control and then subjected to femoral artery coagulation. Recovery of hindlimb perfusion was assessed by laser Doppler perfusion imaging and muscle function by treadmill exhaustion and grip strength testing. After three postoperative weeks, hindlimb muscle was harvested for immunofluorescence analysis. At all postoperative time points, mice treated with E-sel/AAV had improved hindlimb perfusion and exercise capacity. E-sel/AAV gene therapy also increased the coexpression of MyoD and Ki-67 in skeletal muscle progenitors and the proportion of Myh7+ myofibers. Altogether, our findings demonstrate that in addition to improving reperfusion, intramuscular E-sel/AAV gene therapy enhances the regeneration of ischemic skeletal muscle with a corresponding benefit on exercise performance. These results suggest a potential role for E-sel/AAV gene therapy as a nonsurgical adjunct in patients with life-limiting PAD.


Subject(s)
Neovascularization, Physiologic , Peripheral Arterial Disease , Mice , Animals , E-Selectin/genetics , Mice, Inbred C57BL , Muscle, Skeletal/blood supply , Ischemia/genetics , Ischemia/therapy , Genetic Therapy/methods , Peripheral Arterial Disease/genetics , Peripheral Arterial Disease/therapy , Hindlimb/blood supply , Muscle Development , Disease Models, Animal
7.
Front Cardiovasc Med ; 10: 1113982, 2023.
Article in English | MEDLINE | ID: mdl-36818343

ABSTRACT

Progressive peripheral arterial disease (PAD) can result in chronic limb-threatening ischemia (CLTI) characterized by clinical complications including rest pain, gangrene and tissue loss. These complications can propagate even more precipitously in the setting of common concomitant diseases in patients with CLTI such as diabetes mellitus (DM). CLTI ulcers are cutaneous, non-healing wounds that persist due to the reduced perfusion and dysfunctional neovascularization associated with severe PAD. Existing therapies for CLTI are primarily limited to anatomic revascularization and medical management of contributing factors such as atherosclerosis and glycemic control. However, many patients fail these treatment strategies and are considered "no-option," thereby requiring extremity amputation, particularly if non-healing wounds become infected or fulminant gangrene develops. Given the high economic burden imposed on patients, decreased quality of life, and poor survival of no-option CLTI patients, regenerative therapies aimed at neovascularization to improve wound healing and limb salvage hold significant promise. Cell-based therapy, specifically utilizing mesenchymal stem/stromal cells (MSCs), is one such regenerative strategy to stimulate therapeutic angiogenesis and tissue regeneration. Although previous reviews have focused primarily on revascularization outcomes after MSC treatments of CLTI with less attention given to their effects on wound healing, here we review advances in pre-clinical and clinical studies related to specific effects of MSC-based therapeutics upon ischemic non-healing wounds associated with CLTI.

8.
J Thorac Cardiovasc Surg ; 164(2): 378-385, 2022 08.
Article in English | MEDLINE | ID: mdl-35459540

ABSTRACT

BACKGROUND: The influence of SARS-CoV-2 on surgery for non-small cell lung cancer needs to be understood to inform clinical decision making during and after the COVID-19 pandemic. OBJECTIVE: This study reports on the 90-day rate of infection as well as the morbidity and mortality of lung surgery for cancer in a tertiary care hospital located in a pandemic epicenter. METHODS: We conducted a retrospective review of a prospective database to identify consecutive patients who underwent lung cancer resection before (January 1, 2020-March 10, 2020, group 1; 57 patients) and during the COVID-19 pandemic (March 11, 2020-June 10, 2020, group 2; 41 patients). The primary end point was the occurrence of SARS-CoV-2 infection during the first 90-days after surgery. The secondary outcome measure was 90-day perioperative morbidity and mortality. RESULTS: Patient characteristics were not significantly different between the groups. Ninety-day COVID-19 infection rates was 7.3% (3 out of 41) for patients undergoing an operation during the pandemic and 3.5% (2 out of 57) in patients operated on immediately before the pandemic. All patients tested positive 10 to 62 days after the index surgical procedure following hospital discharge. Four COVID-19-positive patients were symptomatic and 4 out of 5 patients required hospitalization, were men, previous or current smokers with hyperlipidemia, and underwent a sublobar resection. Univariate analysis did not identify any differences in postoperative complications before or during the COVID-19 pandemic. Ninety-day mortality was 5% (2 out of 41) for lung cancer surgery performed during the pandemic, with all deaths occurring due to COVID-19, compared with 0% (0 out of 57) mortality in patients who underwent an operation before the pandemic. CONCLUSIONS: During the COVID-19 pandemic, COVID-19 infections occurred in 7.3% of patients who underwent surgery for non-small cell lung cancer. In this series all infections occurred after hospital discharge. Our results suggest that COVID-19 infections occurring within 90 days of surgery portend a 40% mortality, warranting close postoperative surveillance.


Subject(s)
COVID-19 , Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , COVID-19/epidemiology , Carcinoma, Non-Small-Cell Lung/epidemiology , Carcinoma, Non-Small-Cell Lung/surgery , Female , Humans , Lung Neoplasms/epidemiology , Lung Neoplasms/surgery , Male , Pandemics , Retrospective Studies , SARS-CoV-2
9.
Am J Disaster Med ; 17(4): 273-276, 2022.
Article in English | MEDLINE | ID: mdl-37551895

ABSTRACT

OBJECTIVE: Targeting of healthcare infrastructures in Ukraine has served as a tactical warfare strategy. The goal of the Global Surgical and Medical Support Group (GSMSG) is to provide sustainable, high quality medical and surgical care in war zones. INTERVENTIONS: GSMSG deployed United States (US) Special Operations veterans and highly specialized licensed medical experts. Training of Ukrainian civilians was implemented. RESULTS: Over 20,000 Ukrainians trained and over 100 major surgeries performed with assistance of world experts. Specialized medical equipment was also provided. CONCLUSION: GSMSG lays the groundwork for effective, high quality, and sustainable surgical interventions into a nation in conflict.

11.
J Clin Invest ; 128(9): 4086-4097, 2018 08 31.
Article in English | MEDLINE | ID: mdl-29990309

ABSTRACT

Anaplastic thyroid carcinomas (ATCs) have a high prevalence of BRAF and TP53 mutations. A trial of vemurafenib in nonmelanoma BRAFV600E-mutant cancers showed significant, although short-lived, responses in ATCs, indicating that these virulent tumors remain addicted to BRAF despite their high mutation burden. To explore the mechanisms mediating acquired resistance to BRAF blockade, we generated mice with thyroid-specific deletion of p53 and dox-dependent expression of BRAFV600E, 50% of which developed ATCs after dox treatment. Upon dox withdrawal there was complete regression in all mice, although recurrences were later detected in 85% of animals. The relapsed tumors had elevated MAPK transcriptional output, and retained responses to the MEK/RAF inhibitor CH5126766 in vivo and in vitro. Whole-exome sequencing identified recurrent focal amplifications of chromosome 6, with a minimal region of overlap that included Met. Met-amplified recurrences overexpressed the receptor as well as its ligand Hgf. Growth, signaling, and viability of Met-amplified tumor cells were suppressed in vitro and in vivo by the Met kinase inhibitors PF-04217903 and crizotinib, whereas primary ATCs and Met-diploid relapses were resistant. Hence, recurrences are the rule after BRAF suppression in murine ATCs, most commonly due to activation of HGF/MET signaling, which generates exquisite dependency to MET kinase inhibitors.


Subject(s)
Proto-Oncogene Proteins B-raf/antagonists & inhibitors , Proto-Oncogene Proteins c-met/metabolism , Thyroid Carcinoma, Anaplastic/drug therapy , Thyroid Carcinoma, Anaplastic/metabolism , Thyroid Neoplasms/drug therapy , Thyroid Neoplasms/metabolism , Amino Acid Substitution , Animals , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Coumarins/pharmacology , Crizotinib/pharmacology , Disease Models, Animal , Drug Resistance, Neoplasm/genetics , Genes, p53 , Humans , Indoles/pharmacology , MAP Kinase Signaling System , Mice , Mice, Transgenic , Mutation, Missense , Protein Kinase Inhibitors/pharmacology , Proto-Oncogene Proteins B-raf/genetics , Proto-Oncogene Proteins c-met/antagonists & inhibitors , Pyrazines/pharmacology , Sulfonamides/pharmacology , Thyroid Carcinoma, Anaplastic/genetics , Thyroid Neoplasms/genetics , Triazoles/pharmacology
12.
J Exp Med ; 213(1): 25-34, 2016 Jan 11.
Article in English | MEDLINE | ID: mdl-26666262

ABSTRACT

t(8;21) is one of the most frequent chromosomal abnormalities observed in acute myeloid leukemia (AML). However, expression of AML1-ETO is not sufficient to induce transformation in vivo. Consistent with this observation, patients with this translocation harbor additional genetic abnormalities, suggesting a requirement for cooperating mutations. To better define the genetic landscape in AML and distinguish driver from passenger mutations, we compared the mutational profiles of AML1-ETO-driven mouse models of leukemia with the mutational profiles of human AML patients. We identified TET2 and PTPN11 mutations in both mouse and human AML and then demonstrated the ability of Tet2 loss and PTPN11 D61Y to initiate leukemogenesis in concert with expression of AML1-ETO in vivo. This integrative genetic profiling approach allowed us to accurately predict cooperating events in t(8;21)(+) AML in a robust and unbiased manner, while also revealing functional convergence in mouse and human AML.


Subject(s)
Alleles , Epistasis, Genetic , Genomics/methods , Leukemia, Myeloid, Acute/genetics , Animals , Cell Transformation, Neoplastic/genetics , Chromosomes, Human, Pair 21 , Chromosomes, Human, Pair 8 , Core Binding Factor Alpha 2 Subunit/genetics , Disease Models, Animal , Gene Expression Regulation, Leukemic , Gene Knockout Techniques , Humans , Mice , Mutation , Oncogene Proteins, Fusion/genetics , Phenotype , Protein Tyrosine Phosphatase, Non-Receptor Type 11/genetics , RUNX1 Translocation Partner 1 Protein , Translocation, Genetic
13.
Cancer Discov ; 5(11): 1178-93, 2015 Nov.
Article in English | MEDLINE | ID: mdl-26359368

ABSTRACT

UNLABELLED: Ch22q LOH is preferentially associated with RAS mutations in papillary and in poorly differentiated thyroid cancer (PDTC). The 22q tumor suppressor NF2, encoding merlin, is implicated in this interaction because of its frequent loss of function in human thyroid cancer cell lines. Nf2 deletion or Hras mutation is insufficient for transformation, whereas their combined disruption leads to murine PDTC with increased MAPK signaling. Merlin loss induces RAS signaling in part through inactivation of Hippo, which activates a YAP-TEAD transcriptional program. We find that the three RAS genes are themselves YAP-TEAD1 transcriptional targets, providing a novel mechanism of promotion of RAS-induced tumorigenesis. Moreover, pharmacologic disruption of YAP-TEAD with verteporfin blocks RAS transcription and signaling and inhibits cell growth. The increased MAPK output generated by NF2 loss in RAS-mutant cancers may inform therapeutic strategies, as it generates greater dependency on the MAPK pathway for viability. SIGNIFICANCE: Intensification of mutant RAS signaling through copy-number imbalances is commonly associated with transformation. We show that NF2/merlin inactivation augments mutant RAS signaling by promoting YAP/TEAD-driven transcription of oncogenic and wild-type RAS, resulting in greater MAPK output and increased sensitivity to MEK inhibitors.


Subject(s)
Gene Deletion , Genes, ras , Mitogen-Activated Protein Kinases/antagonists & inhibitors , Neurofibromin 2/genetics , Nuclear Proteins/metabolism , Thyroid Neoplasms/genetics , Thyroid Neoplasms/metabolism , Transcription Factors/metabolism , Animals , Binding Sites , Cell Cycle Proteins , Cell Line, Tumor , Cell Transformation, Neoplastic/genetics , Cell Transformation, Neoplastic/metabolism , Chromosome Deletion , Chromosomes, Human, Pair 22 , DNA Copy Number Variations , Disease Models, Animal , Drug Resistance, Neoplasm/genetics , Gene Expression Regulation, Neoplastic/drug effects , Gene Order , Gene Targeting , Humans , Mice , Mice, Transgenic , Models, Biological , Neoplasm Staging , Nucleotide Motifs , Position-Specific Scoring Matrices , Promoter Regions, Genetic , Protein Binding , Protein Kinase Inhibitors/pharmacology , Signal Transduction/drug effects , Thyroid Neoplasms/drug therapy , Thyroid Neoplasms/pathology , Transcriptional Activation
14.
J Clin Endocrinol Metab ; 99(10): E1976-87, 2014 Oct.
Article in English | MEDLINE | ID: mdl-25029414

ABSTRACT

CONTEXT: Thyroid growth is regulated by TSH and requires mammalian target of rapamycin (mTOR). Thyroid cancers frequently exhibit mutations in MAPK and/or phosphoinositol-3-kinase-related kinase effectors. OBJECTIVE: The objective of the study was to explore the contribution of RET/PTC, RAS, and BRAF to mTOR regulation and response to mTOR inhibitors. METHODS: PCCL3 cells conditionally expressing RET/PTC3, HRAS(G12V), or BRAF(V600E) and human thyroid cancer cells harboring mutations of these genes were used to test pathways controlling mTOR and its requirement for growth. RESULTS: TSH/cAMP-induced growth of PCCL3 cells requires mTOR, which is stimulated via protein kinase A in a MAPK kinase (MEK)- and AKT-independent manner. Expression of RET/PTC3, HRAS(G12V), or BRAF(V600E) in PCCL3 cells induces mTOR but does not entirely abrogate the cAMP-mediated control of its activity. Acute oncoprotein-induced mTOR activity is regulated by MEK and AKT, albeit to differing degrees. By contrast, mTOR was not activated by TSH/cAMP in human thyroid cancer cells. Tumor genotype did not predict the effects of rapamycin or the mTOR kinase inhibitor AZD8055 on growth, with the exception of a PTEN-null cell line. Selective blockade of MEK did not influence mTOR activity of BRAF or RAS mutant cells. Combined MEK and mTOR kinase inhibition was synergistic on growth of BRAF- and RAS-mutant thyroid cancer cells in vitro and in vivo. CONCLUSION: Thyroid cancer cells lose TSH/cAMP dependency of mTOR signaling and cell growth. mTOR activity is not decreased by the MEK or AKT inhibitors in the RAS or BRAF human thyroid cancer cell lines. This may account for the augmented effects of combining the mTOR inhibitors with selective antagonists of these oncogenic drivers.


Subject(s)
MAP Kinase Signaling System/physiology , Multiprotein Complexes/metabolism , TOR Serine-Threonine Kinases/metabolism , Thyroid Neoplasms/metabolism , Animals , Antibiotics, Antineoplastic/pharmacology , Benzimidazoles/pharmacology , Cell Line, Tumor , Cyclic AMP/metabolism , Humans , MAP Kinase Signaling System/drug effects , Mechanistic Target of Rapamycin Complex 1 , Mice, Nude , Morpholines/pharmacology , Neoplasm Transplantation , Oncogene Proteins/genetics , Oncogene Proteins/metabolism , Proto-Oncogene Proteins B-raf/genetics , Proto-Oncogene Proteins B-raf/metabolism , Proto-Oncogene Proteins c-ret/genetics , Proto-Oncogene Proteins c-ret/metabolism , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Rats , Sirolimus/pharmacology , Thyroid Neoplasms/pathology , Thyrotropin/metabolism , ras Proteins/genetics , ras Proteins/metabolism
15.
Cell Rep ; 5(6): 1625-38, 2013 Dec 26.
Article in English | MEDLINE | ID: mdl-24332853

ABSTRACT

Defining the role of epigenetic regulators in hematopoiesis has become critically important, because recurrent mutations or aberrant expression of these genes has been identified in both myeloid and lymphoid hematological malignancies. We found that PRMT4, a type I arginine methyltransferase whose function in normal and malignant hematopoiesis is unknown, is overexpressed in acute myelogenous leukemia patient samples. Overexpression of PRMT4 blocks the myeloid differentiation of human stem/progenitor cells (HSPCs), whereas its knockdown is sufficient to induce myeloid differentiation of HSPCs. We demonstrated that PRMT4 represses the expression of miR-223 in HSPCs via the methylation of RUNX1, which triggers the assembly of a multiprotein repressor complex that includes DPF2. As part of the feedback loop, PRMT4 expression is repressed posttranscriptionally by miR-223. Depletion of PRMT4 results in differentiation of myeloid leukemia cells in vitro and their decreased proliferation in vivo. Thus, targeting PRMT4 holds potential as a novel therapy for acute myelogenous leukemia.


Subject(s)
Core Binding Factor Alpha 2 Subunit/metabolism , Epigenetic Repression , Hematopoiesis , Myeloid Progenitor Cells/metabolism , Protein-Arginine N-Methyltransferases/metabolism , Animals , Cell Line, Tumor , DNA-Binding Proteins/metabolism , Humans , Methylation , Mice , MicroRNAs/genetics , MicroRNAs/metabolism , Myeloid Progenitor Cells/cytology , Protein Binding , Protein-Arginine N-Methyltransferases/genetics , RNA Processing, Post-Transcriptional , Transcription Factors
16.
J Clin Invest ; 123(11): 4935-44, 2013 Nov.
Article in English | MEDLINE | ID: mdl-24135138

ABSTRACT

Exposure to ionizing radiation during childhood markedly increases the risk of developing papillary thyroid cancer. We examined tissues from 26 Ukrainian patients with thyroid cancer who were younger than 10 years of age and living in contaminated areas during the time of the Chernobyl nuclear reactor accident. We identified nonoverlapping somatic driver mutations in all 26 cases through candidate gene assays and next-generation RNA sequencing. We found that 22 tumors harbored fusion oncogenes that arose primarily through intrachromosomal rearrangements. Altogether, 23 of the oncogenic drivers identified in this cohort aberrantly activate MAPK signaling, including the 2 somatic rearrangements resulting in fusion of transcription factor ETS variant 6 (ETV6) with neurotrophic tyrosine kinase receptor, type 3 (NTRK3) and fusion of acylglycerol kinase (AGK) with BRAF. Two other tumors harbored distinct fusions leading to overexpression of the nuclear receptor PPARγ. Fusion oncogenes were less prevalent in tumors from a cohort of children with pediatric thyroid cancers that had not been exposed to radiation but were from the same geographical regions. Radiation-induced thyroid cancers provide a paradigm of tumorigenesis driven by fusion oncogenes that activate MAPK signaling or, less frequently, a PPARγ-driven transcriptional program.


Subject(s)
Carcinoma/genetics , Chernobyl Nuclear Accident , Mutation , Neoplasms, Radiation-Induced/genetics , Oncogene Fusion , Thyroid Neoplasms/genetics , Adolescent , Animals , Base Sequence , Carcinoma, Papillary , Child , Child, Preschool , Cohort Studies , DNA, Neoplasm/genetics , Female , Gene Rearrangement , Humans , MAP Kinase Signaling System/genetics , Male , Mice , Molecular Sequence Data , NIH 3T3 Cells , PPAR gamma/genetics , Phosphatidylinositol 3-Kinases/genetics , Phosphotransferases (Alcohol Group Acceptor)/genetics , Proto-Oncogene Proteins B-raf/genetics , Proto-Oncogene Proteins c-ets/genetics , Proto-Oncogene Proteins c-ret/genetics , Receptor, trkC/genetics , Receptors, Thyrotropin/genetics , Repressor Proteins/genetics , Thyroid Cancer, Papillary , Ukraine , Young Adult , ETS Translocation Variant 6 Protein
17.
Cell Stem Cell ; 11(6): 836-44, 2012 Dec 07.
Article in English | MEDLINE | ID: mdl-23217424

ABSTRACT

High-grade gliomas are aggressive and uniformly fatal tumors, composed of a heterogeneous population of cells that include many with stem-cell-like properties. The acquisition of stem-like traits might contribute to glioma initiation, growth, and recurrence. Here we investigated the role of the transcription factor myeloid Elf-1 like factor (MEF, also known as ELF4) in gliomas. We found that MEF is highly expressed in both human and mouse glioblastomas and its absence impairs gliomagenesis in a PDGF-driven glioma mouse model. We show that modulation of MEF levels in both mouse neural stem cells and human glioblastoma cells has a significant impact on neurosphere formation. Moreover, we identify Sox2 as a direct downstream target of MEF. Taken together, our studies implicate MEF as a previously unrecognized gatekeeper gene in gliomagenesis that promotes stem cell characteristics through Sox2 activation.


Subject(s)
Brain Neoplasms/metabolism , Brain Neoplasms/pathology , DNA-Binding Proteins/metabolism , Glioma/metabolism , Glioma/pathology , Neoplastic Stem Cells/pathology , Transcription Factors/metabolism , Animals , Brain Neoplasms/genetics , Cell Line, Tumor , Cell Proliferation , DNA-Binding Proteins/genetics , Gene Expression Regulation, Neoplastic , Glioma/genetics , Humans , Mice , Neoplastic Stem Cells/metabolism , Neural Stem Cells/metabolism , Neural Stem Cells/pathology , SOXB1 Transcription Factors/metabolism , Spheroids, Cellular/metabolism , Spheroids, Cellular/pathology , Tissue Culture Techniques , Transcription Factors/genetics
18.
Science ; 333(6043): 765-9, 2011 Aug 05.
Article in English | MEDLINE | ID: mdl-21764752

ABSTRACT

The chromosomal translocations found in acute myelogenous leukemia (AML) generate oncogenic fusion transcription factors with aberrant transcriptional regulatory properties. Although therapeutic targeting of most leukemia fusion proteins remains elusive, the posttranslational modifications that control their function could be targetable. We found that AML1-ETO, the fusion protein generated by the t(8;21) translocation, is acetylated by the transcriptional coactivator p300 in leukemia cells isolated from t(8;21) AML patients, and that this acetylation is essential for its self-renewal-promoting effects in human cord blood CD34(+) cells and its leukemogenicity in mouse models. Inhibition of p300 abrogates the acetylation of AML1-ETO and impairs its ability to promote leukemic transformation. Thus, lysine acetyltransferases represent a potential therapeutic target in AML.


Subject(s)
Cell Transformation, Neoplastic , Core Binding Factor Alpha 2 Subunit/metabolism , E1A-Associated p300 Protein/metabolism , Hematopoietic Stem Cells/cytology , Leukemia, Myeloid, Acute/metabolism , Lysine/metabolism , Oncogene Proteins, Fusion/metabolism , Acetylation , Animals , Cell Line , Cell Line, Tumor , Core Binding Factor Alpha 2 Subunit/chemistry , E1A-Associated p300 Protein/antagonists & inhibitors , Fetal Blood/cytology , Gene Expression Profiling , Hematopoietic Stem Cells/physiology , Humans , Leukemia, Myeloid, Acute/pathology , Mice , Mice, Inbred C57BL , Mutant Proteins/metabolism , Oncogene Proteins, Fusion/chemistry , Preleukemia/metabolism , Preleukemia/pathology , Protein Binding , Protein Interaction Domains and Motifs , Protein Processing, Post-Translational , RUNX1 Translocation Partner 1 Protein , Transcriptional Activation , Tumor Cells, Cultured
19.
Stem Cells Dev ; 20(11): 1889-900, 2011 Nov.
Article in English | MEDLINE | ID: mdl-21341991

ABSTRACT

Human embryonic stem cells (hESCs) can be used to study the early events in human development and, hopefully, to understand how to differentiate human pluripotent cells for clinical use. To define how L3MBTL1, a chromatin-associated polycomb group protein with transcriptional repressive activities, regulates early events in embryonic cell differentiation, we created hESC lines that constitutively express shRNAs directed against L3MBTL1. The L3MBTL1 knockdown (KD) hESCs maintained normal morphology, proliferation, cell cycle kinetics, cell surface markers, and karyotype after 40 passages. However, under conditions that promote spontaneous differentiation, the L3MBTL1 KD cells differentiated into a relatively homogeneous population of large, flat trophoblast-like cells, unlike the multilineage differentiation seen with the control cells. The differentiated L3MBTL1 KD cells expressed numerous trophoblast markers and secreted placental hormones. Although the L3MBTL1 KD cells could be induced to differentiate into various embryonic lineages, they adopted an exclusive trophoblast fate during spontaneous differentiation. Our data demonstrate that depletion of L3MBTL1 does not affect hESC self-renewal, rather it enhances differentiation toward extra-embryonic trophoblast tissues.


Subject(s)
Cell Differentiation , Chromosomal Proteins, Non-Histone/deficiency , Ectoderm/cytology , Embryonic Stem Cells/physiology , Trophoblasts/cytology , Antigens, Differentiation/genetics , Antigens, Differentiation/metabolism , Bone Morphogenetic Protein 4/pharmacology , Bone Morphogenetic Protein 4/physiology , Cell Differentiation/drug effects , Cell Line , Cell Proliferation , Cell Shape , Chromosomal Proteins, Non-Histone/genetics , Coculture Techniques , Gene Expression , Gene Knockdown Techniques , Green Fluorescent Proteins/metabolism , Humans , Microscopy, Fluorescence , RNA Interference , Recombinant Proteins/metabolism , Repressor Proteins , Tumor Suppressor Proteins
20.
Proc Natl Acad Sci U S A ; 107(52): 22552-7, 2010 Dec 28.
Article in English | MEDLINE | ID: mdl-21149733

ABSTRACT

The l3mbtl1 gene is located on the long arm of chromosome 20 (q12), within a region commonly deleted in several myeloid malignancies. L3MBTL1 is a human homolog of the Drosophila polycomb L(3)MBT tumor suppressor protein and thus a candidate tumor suppressor in del(20q12) myeloid disorders. We used the loss-of-function approach to explore the possible tumor suppressive mechanism of L3MBTL1 and found that depletion of L3MBTL1 from human cells causes replicative stress, DNA breaks, activation of the DNA damage response, and genomic instability. L3MBTL1 interacts with Cdc45, MCM2-7 and PCNA, components of the DNA replication machinery, and is required for normal replication fork progression, suggesting that L3MBTL1 causes DNA damage, at least in part, by perturbing DNA replication. An activated DNA damage response and genomic instability are common features in tumorigenesis and a consequence of overexpression of many oncogenes. We propose that the loss of L3MBTL1 contributes to the development of 20q(-) hematopoietic malignancies by inducing replicative stress, DNA damage, and genomic instability.


Subject(s)
Genomic Instability , Myeloproliferative Disorders/metabolism , Neoplasm Proteins/metabolism , Tumor Suppressor Proteins/metabolism , Blotting, Western , Cell Cycle , Cell Line , Cell Line, Tumor , Cell Proliferation , Chromosomal Proteins, Non-Histone , Chromosome Deletion , Chromosomes, Human, Pair 20/genetics , DNA Damage , DNA Replication , HEK293 Cells , Histones/metabolism , Humans , Immunoprecipitation , K562 Cells , Lysine/metabolism , Methylation , Myeloproliferative Disorders/genetics , Neoplasm Proteins/genetics , Protein Binding , RNA Interference , Repressor Proteins , Retinoblastoma Protein/metabolism , Tumor Suppressor Proteins/genetics
SELECTION OF CITATIONS
SEARCH DETAIL
...