ABSTRACT
8-arm PEG (polyethylene-glycol) is a highly promising nanoplatform due to its small size (<10 nm), ease-of-conjugation (many functionalized variants are readily available with "click-like" properties), biocompatibility, and optical inactivity. This study evaluates 8-arm PEG uptake into cells (in vitro) and localization and clearance in vasculature (in vivo) for targeting of choroidal neovascularization in mice, an animal model of macular degeneration. 8-arm PEG nanoparticles were labeled with fluorescein isothiocyanate (FITC) and functionalized in the absence or presence of pentameric Ar-Gly-Asp (RGD; 4 RGD motifs and a PGC linker), one of the most common peptide motifs used for active targeting. In vitro studies show that RGD-conjugated 8-arm PEG nanoparticles exhibit enhanced cellular uptake relative to non-RGD-conjugated control NPs at 34% ± 9%. Laser-induced choroidal neovascularization (CNV) was performed in a mouse model to measure 8-arm PEG localization and clearance to model macular degeneration lesions in vivo. It was determined that both RGD-conjugated and non-RGD-conjugated (nRGD) 8-arm PEG particles localized to CNV lesions, with a half-life around 24 h. In vivo experiments showed that RGD-conjugated nanoparticles exhibited enhanced localization by 15-20% relative to without RGD controls. Exhibiting a high rate of localization and fast clearance relative to larger nanoparticles, targeted 8-arm PEG nanoparticles with a conjugated RGD-peptide could be a promising modality for macular degeneration diagnosis and therapy.
ABSTRACT
: Almost all patients with EGFR-driven lung cancer who are treated with EGFR tyrosine kinase inhibitors (TKI) develop resistance to treatment. A single base (c.2369C>T) transition mutation, EGFR T790M, is the most frequent resistance event after first-generation exposure to EGFR TKIs. Whether T790M mutation is acquired or is selected from a preexisting clone has been a matter of significant debate. In this study, we show that treatment with EGFR TKIs leads to activation of the NFκB pathway, which in turn induces expression of activation-induced cytidine deaminase (AICDA). In turn, AICDA causes deamination of 5-methylcytosine to thymine at position c.2369 to generate the T790M mutation. Pharmacologic inhibition of the NFκB pathway or knockout of AICDA decreased the frequency or prevented the development of T790M mutation, respectively. In addition, patients treated with first-line EGFR TKI displayed increased expression of AICDA and detection of the T790M mutation upon progression. These results identify the mechanism of T790M acquisition and present an opportunity to target the process to delay or prevent it. SIGNIFICANCE: These findings identify the mechanism behind acquisition of a common resistance mutation to TKI treatment in lung cancer.
Subject(s)
5-Methylcytosine/chemistry , Cytidine Deaminase/metabolism , Lung Neoplasms/drug therapy , Lung Neoplasms/genetics , Protein-Tyrosine Kinases/antagonists & inhibitors , Aged , Cell Line, Tumor , Deamination , Disease Progression , ErbB Receptors/genetics , Female , Humans , Hydrolysis , Male , Mass Spectrometry , Methylation , Middle Aged , Mutation , NF-kappa B/metabolism , Polymerase Chain ReactionABSTRACT
CONTEXT: Pheochromocytoma/paraganglioma occurs almost exclusively after paternal transmission of succinate dehydrogenase D (SDHD) mutations. This parent-of-origin effect has not been fully explained but is accompanied by obligate loss of the maternal copy of chromosome 11. Loss of wild-type SDHD and an additional imprinted gene (hypothesized to be H19) appears necessary for tumor formation. Two previous reports suggested tumor formation after maternal transmission of SDHD mutation, but histological and molecular characterization was unavailable. OBJECTIVE: We report the first kindred in which histologically confirmed pheochromocytoma/paraganglioma occurred after maternal transmission of an SDHD mutation and investigate the molecular mechanism of tumor formation. DESIGN: The design of the investigation was the study of a three-generation family with SDHD c.242C>T (p.Pro81Leu) mutation. RESULTS: The index patient had a histologically confirmed pheochromocytoma and an identical SDHD germline mutation (p.Pro81Leu) to her mother (who had a glomus jugulare tumor) and paraganglioma tissue from her maternal grandfather. Tumor DNA from the index patient revealed loss of heterozygosity (LOH) at 11q23, causing loss of the wild-type paternal SDHD allele and LOH affecting maternal 11p15, including H19. These two regions of LOH were separated by a region exhibiting clearly retained heterozygosity, including SDHAF2, a recently reported paraganglioma susceptibility gene. CONCLUSIONS: Tumor formation can occur after maternal transmission of SDHD, a finding with important clinical implications for SDHD families. Tumor formation in SDHD mutation requires the loss of both the wild-type SDHD allele and maternal 11p15, leading to the predominant but now not exclusive pattern of disease inheritance after paternal SDHD transmission.
Subject(s)
Adrenal Gland Neoplasms/genetics , Germ-Line Mutation , Pheochromocytoma/genetics , Succinate Dehydrogenase/genetics , Alleles , Chromosomes, Human, Pair 11 , DNA Mutational Analysis , Female , Humans , Loss of Heterozygosity , Male , PedigreeABSTRACT
We describe the clinical characteristics of 4 singleton cases, 3 males and 1 female, with Myhre Syndrome (OMIM 139210), who were born to non-consanguineous parents. Three cases had no family history of similarly affected individuals but 1 male's mother had short stature, some facial features suggestive of Myhre syndrome and evidence of skewed X-chromosome inactivation in her blood DNA. Short stature, deafness, learning difficulties, skeletal anomalies and facial dysmorphisms were evident in all cases. Arthralgia and stiff joints with limited movement were also present. The facial appearance, thickened skin, a 'muscular' habitus are memorable features. The female patient was least affected: this patient and one affected male displayed streaky skin with areas of patchy thickening, suggestive of genetic mosaicism. One patient developed sleep apnoea, a restrictive ventilatory defect and died following a choking episode. Another affected male developed recurrent, progressive, proximal, tracheal stenosis requiring partial tracheal resection, laser treatment and eventually tracheotomy. Review of Myhre syndrome patients in the literature and syndromes in the differential diagnosis, suggests heterogeneity in Myhre syndrome and clinical overlap with Laryngotracheal stenosis, Arthropathy, Prognathism and Short stature syndrome.
Subject(s)
Abnormalities, Multiple/diagnosis , Cryptorchidism , Facies , Growth Disorders , Hand Deformities, Congenital , Hypertrophy , Intellectual Disability , Joint Diseases , Respiratory System/pathology , Adolescent , Adult , Cryptorchidism/diagnosis , Cryptorchidism/genetics , Cryptorchidism/pathology , DNA Mutational Analysis , Diagnosis, Differential , Female , Growth Disorders/diagnosis , Growth Disorders/genetics , Growth Disorders/pathology , Hand Deformities, Congenital/diagnosis , Hand Deformities, Congenital/genetics , Hand Deformities, Congenital/pathology , Humans , Hypercalcemia/diagnosis , Hypertrophy/diagnosis , Hypertrophy/genetics , Hypertrophy/pathology , Intellectual Disability/diagnosis , Intellectual Disability/genetics , Intellectual Disability/pathology , Joint Diseases/diagnosis , Joint Diseases/genetics , Joint Diseases/pathology , Laryngostenosis/diagnosis , Male , Metabolic Diseases/diagnosis , Nephrocalcinosis/diagnosis , Pedigree , Prognathism/diagnosis , Skin Abnormalities/pathology , Tracheal Stenosis/diagnosis , United Kingdom , X Chromosome InactivationABSTRACT
BACKGROUND: HFE-related genetic haemochromatosis (GH) is the commonest inherited genetic disorder in Caucasian populations with approximately one in 180 of individuals in the west of Scotland homozygous for the common C282Y mutation. The clinical diagnosis of GH, however, remains relatively uncommon - suggesting either under diagnosis or low clinical penetrance. AIM: We aimed to assess the biochemical and clinical penetrance of GH in first-degree relatives of patients with known GH, who subsequently themselves screened positive for the common GH mutations. METHODS: Individuals were identified from two large teaching hospitals in North Glasgow from July 1997 to July 2005 diagnosed with GH after predictive genetic testing after a relative was found to have GH. Details of patient history, biochemistry and known comorbidity at diagnosis and results of related further investigations were collected. RESULTS: Sixty-three individuals were identified, 31 (49%) of whom were males. Fifty-five individuals (87%) were C282Y homozygous and the remaining eight were compound heterozygotes for C282Y and H63D. All 31 male patients were found to have evidence of iron overload as opposed to 63% of females. Elevated liver enzyme levels were encountered in 15 patients (24%). All except one had evidence of iron overload. Four individuals underwent a liver biopsy, two of whom had hepatic fibrosis. Four patients were found to be diabetic. A full clinical history was obtained from 54 of 63 individuals, 38 (70%) of whom were entirely asymptomatic. Thirteen individuals complained of joint pains and a further nine complained of fatigue. CONCLUSION: This study suggests that although biochemical penetrance of GH is high, the clinical penetrance is low.
Subject(s)
Genetic Testing , Hemochromatosis/blood , Penetrance , Adult , Aged , Aging/blood , Arthralgia/etiology , Diabetes Mellitus, Type 2/etiology , Fatigue/etiology , Female , Ferritins/blood , Hemochromatosis/complications , Hemochromatosis/diagnosis , Hemochromatosis/genetics , Hemochromatosis Protein , Heterozygote , Histocompatibility Antigens Class I/genetics , Homozygote , Humans , Iron Overload/genetics , Liver/enzymology , Male , Membrane Proteins/genetics , Middle Aged , Mutation , Transferrin/metabolismABSTRACT
INTRODUCTION: Genetic variation in plasma fibrinogen and the platelet receptor GP IIIa locus has been independently associated with increased risks of ischaemic heart disease, but there have been few reports on the relationship with peripheral arterial disease. This study determined the risk of peripheral arterial disease and ischaemic heart disease associated with polymorphisms of fibrinogen T/G(+1689) and platelet glycoprotein Pl(A) genes and the effects of cigarette smoking and fibrinogen. MATERIALS AND METHODS: In the 5-year follow-up phase of the Edinburgh Artery Study, 939 subjects (60-79 years) had DNA extracted from a venous blood sample. One hundred sixteen subjects were identified as having angina, 87 a myocardial infarction, 104 had intermittent claudication and 663 subjects comprised a healthy group. RESULTS: Distribution of the fibrinogen genotype was similar across the disease and healthy groups. Logistic regression analyses found no significant association between fibrinogen genotype and ischaemic heart disease and peripheral arterial disease. A lower percentage of claudicants had the Pl(A2) allele (8.3% vs. 15.2%, p=0.025). After adjustment for age and sex, the risk of IC associated with the Pl(A2) was half that of the homozygous Pl(A1) genotype (OR 0.49, 95% CI 0.25, 0.88; p<==0.05). Adjustment for lifetime smoking and fibrinogen levels increased the odds slightly to nonsignificance. CONCLUSIONS: The Pl(A2) genotype was associated with a decreased risk of developing IC. There was no significant relationship between fibrinogen T/G(+1689) genotype and ischaemic and peripheral heart disease in this older population.