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Wei-Hong Yang

Publications and source records attributed to Wei-Hong Yang.

12 recordsLinked to original sources
PubMed2006

RNA-associated protein 55 (RAP55) localizes to mRNA processing bodies and stress granules.

The mRNA processing body (P-body) is a cellular structure that has an important role in mRNA degradation. P-bodies have also been implicated in RNAi-mediated post-transcriptional gene silencing. The objective of this study was to identify and characterize novel components of the mammalian P-body. Approximately 5% of patients with the autoimmune disease primary biliary cirrhosis have antibodies directed against this structure. Serum from one of these patients was used to identify a cDNA encoding RAP55, a 463-amino acid protein. RAP55 colocalized with previously identified P-body components DCP1a and Ge-1. RAP55 contains an N-terminal Sm-like domain and two C-terminal RGG-rich domains separated by an FDF motif. The two RGG domains and the FDF domain were necessary and sufficient to target the protein to P-bodies. A fragment of RAP55 consisting of the FDF and the second RGG domains did not localize to P-bodies, but was able to displace other P-body components from this structure. After cells were subjected to arsenite-induced stress, RAP55 was detected in TIA-containing stress granules. The second RGG domain was necessary and sufficient for stress granule localization. siRNA-mediated knock-down of RAP55 resulted in loss of P-bodies, suggesting that RAP55 acts prior to the 5'-decapping step in mRNA degradation. The results of this study show that RAP55 is a component of P-bodies in cells at rest and localizes in stress granules in arsenite-treated cells. RAP55 may serve to shuttle mRNAs between P-bodies and stress granules.

Blotting, Western↗
PubMed2006

Processing body autoantibodies reconsidered.

Processing bodies (P-bodies) are cellular structures that have critical roles in mRNA degradation and post-transcriptional gene silencing. Some patients with autoimmune disease have high titer antibodies directed against P-bodies, and certain sera have been used as markers for the GW182 component of these structures. This study shows that available reference sera are unreliable markers for GW182 because the sera contain antibodies directed against Ge-1, a second P-body autoantigen.

Argonaute Proteins↗
PubMed2005

Mediation of Epstein-Barr virus EBNA-LP transcriptional coactivation by Sp100.

The Epstein-Barr virus (EBV) EBNA-LP protein is important for EBV-mediated B-cell immortalization and is a potent gene-specific coactivator of the viral transcriptional activator, EBNA2. The mechanism(s) by which EBNA-LP functions as a coactivator remains an important question in the biology of EBV-induced B-cell immortalization. In this study, we found that EBNA-LP interacts with the promyelocytic leukemia nuclear body (PML NB)-associated protein Sp100 and displaces Sp100 and heterochromatin protein 1alpha (HP1alpha) from PML NBs. Interaction between EBNA-LP and Sp100 was mediated through conserved region 3 in EBNA-LP and the PML NB targeting domain in Sp100. Overexpression of Sp100 lacking the N-terminal PML NB targeting domain, but not a mutant form of Sp100 lacking the HP1alpha interaction domain, was sufficient to coactivate EBNA2 in a gene-specific manner independent of EBNA-LP. These findings suggest that Sp100 is a major mediator of EBNA-LP coactivation. These studies indicate that modulation of PML NB-associated proteins may be important for establishment of latent viral infections, and also identify a convenient model system to investigate the functions of Sp100.

Antigens, Nuclear↗
PubMed2005

Engineering lipid vesicles of enhanced intratumoral transport capabilities: correlating liposome characteristics with penetration into human prostate tumor spheroids.

Liposomes have been widely used delivery systems, particularly relevant to the development of cancer therapeutics. Numerous liposome-based drugs are in the clinic or in clinical trials today against multiple tumor types; however, systematic studies of liposome interactions with solid or metastatic tumor nodules are scarce. This study is describing the in vitro interaction between liposomes and avascular human prostate (LNCaP-LN3) tumor spheroids. The ability of fluorescently labelled liposomal delivery systems of varying physicochemical characteristics to penetrate within multicellular tumor spheroids has been investigated by confocal laser scanning microscopy. A variety of liposome characteristics and experimental parameters were investigated, including lipid bilayer composition, duration of liposome-spheroid interaction, mean liposome size, steric stabilization of liposomes. Electrostatic binding between cationic liposomes and spheroids was very efficient; however, it impeded any significant penetration of the vesicles within deeper layers of the tumor spheroid. Small unilamellar liposomes of neutral surface character did not bind as efficiently but exhibited enhanced penetrative transport capabilities closer to the tumor core. Polymer-coated (sterically stabilised) liposomes exhibited almost no interaction with the spheroid, indicating that their limited diffusion within avascular tissues may be a limiting step for their use against micrometastases. Multicellular tumor spheroids were used as models of solid tumor interstitium relevant to delivery systems able to extravasate from the microcapillaries or as models of prevascularized micrometastases. This study illustrates that interactions between liposomes and other drug delivery systems with multicellular tumor spheroids can offer critically important information with respect to optimizing solid or micrometastatic tumor delivery and targeting strategies.

Humans↗
PubMed2005

Ge-1 is a central component of the mammalian cytoplasmic mRNA processing body.

The mRNA processing body (P-body) is a cellular structure that regulates gene expression by degrading cytoplasmic mRNA. The objective of this study was to identify and characterize novel components of the mammalian P-body. Approximately 5% of patients with the autoimmune disease primary biliary cirrhosis have antibodies directed against this structure. Serum from one of these patients was used to identify a cDNA encoding Ge-1, a 1,401-amino-acid protein. Ge-1 contains an N-terminal WD 40 motif and C-terminal domains characterized by a repeating psi(X(2-3)) motif. Ge-1 co-localized with previously identified P-body components, including proteins involved in mRNA decapping (DCP1a and DCP2) and the autoantigen GW 182. The Ge-1 C-terminal domain was necessary and sufficient to target the protein to P-bodies. Following exposure of cells to oxidative stress, Ge-1-containing P-bodies were found adjacent to TIA-containing stress granules. During the recovery period, TIA returned to the nucleus while Ge-1-containing P-bodies localized to the perinuclear region. siRNA-mediated knock-down of Ge-1 resulted in loss of P-bodies containing Ge-1, DCP1a, and DCP2. In contrast, Ge-1-containing P-bodies persisted despite knock-down of DCP2. Taken together, the results of this study show that Ge-1 is a central component of P-bodies and suggest that Ge-1 may act prior to the 5(')-decapping step in mRNA degradation.

Amino Acid Motifs↗
PubMed2005

Liposome-mediated radiotherapeutics within avascular tumor spheroids: comparative dosimetry study for various radionuclides, liposome systems, and a targeting antibody.

UNLABELLED: Absorbed dose profiles within tumor spheroids simulating avascular micrometastases have been calculated for a variety of liposome- and antibody-radionuclide combinations to assess the anticipated therapeutic efficacy based on the intratumoral distribution of the carrier systems within the spheroid model. METHODS: Experiments studying the targeting and diffusion capability of the most clinically relevant liposome systems and the anti-PSMA (prostate-specific membrane antigen) antibody J591 within spheroids of the prostate cancer cell line LNCaP (diameter, 150-200 mum) have been performed. The intratumoral biodistribution data were then used as the input to obtain absorbed dose profiles within the tumor spheroid mass. The dosimetric analysis was performed for a variety of medium- and high-energy beta-emitting radionuclides ((32)P, (90)Y, (188)Re, (67)Cu, (131)I) and 2 low-energy Auger or conversion electron emitters ((123)I, (125)I) following the point-kernel convolution method in the continuous slowing-down approximation. RESULTS: Relative absorbed dose distribution calculations as a function of the distance from the rim of the spheroids are presented. For all liposome systems studied, the SUV-DMPC-chol (small unilamellar vesicle-dimyristoyl-phosphatidylcholine-cholesterol) was most efficient in penetrating deeper within the spheroids. For the beta-emitters it delivered its maximum absorbed dose (D(max)) at 40- to 50-microm depth, exhibiting an almost flat absorbed dose profile beyond that point, as is evident by the high absorbed dose value at the center of the spheroid (D(core)), D(core)/D(max) > 0.9; the respective values for the J591 antibody were 20 mum and 0.85. The Auger or conversion emitters resulted in the most heterogeneous absorbed dose distribution; the ratio D(core)/D(max) fell to 0.4 for the SUV-DMPC-chol and to 0.4-0.5 for the antibody. In general, a 2- to 10-fold "cross-fire"-related increase of the core absorbed dose was observed. For liposomes exhibiting high binding capacity (3beta-[N-(N',N']-dimethylaminoethane)carbamoyl]cholesterol [DC-chol]), however, the low-energy emitters deliver up to a 40% higher D(max) relative to the beta-emitters. The surface characteristics of liposomes appear to have a noticeable influence on the absorbed dose profiles. The use of neutral (DMPC-chol) versus cationic (DC-chol) lipids resulted in up to a 10-fold increase of D(core)/D(max) depending on the radionuclide. Changing the cationic lipid used to N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium methyl sulfate also had a notable influence (up to a 6-fold increase), whereas the effect of fusogenic lipids (dioleoylphosphatidylcholine) was found to be much smaller. CONCLUSION: It is possible to engineer liposome systems that are particularly effective in delivering an almost uniform absorbed dose profile at the central region of micrometastatic tumors, provided that conjugates with the appropriate radionuclides are constructed. In view of the passive means of diffusion of liposomes within solid tumors, it is suggested that they may effectively complement an antibody-based therapeutic regime against micrometastatic tumors, leading to cytotoxic absorbed dose levels throughout the entire tumor volume--thus, hindering tumor recurrence.

Antibodies, Monoclonal↗
PubMed2005

The cytoplasmic dot staining pattern is detected in a subgroup of patients with primary biliary cirrhosis.

OBJECTIVE: To determine the clinical significance of the cytoplasmic dot anti-"nuclear" antibody (ANA) staining pattern. METHODS: We describe a patient with fatigue, arthralgias, elevated serum transaminase, and antibodies staining 5-20 cytoplasmic dots in HEp-2 cells. A liver biopsy revealed the presence of Stage III primary biliary cirrhosis (PBC). Using 2-color immunofluorescence, we determined the relationship between the cytoplasmic dot staining pattern and that produced by antibodies directed against the GW182 component of mRNA processing bodies. To determine the prevalence of the cytoplasmic dot staining pattern in patients with PBC, sera from 493 patients were tested for antibodies producing this staining pattern. RESULTS: Antibodies in our patient's serum colocalized with anti-GW182 antibodies in cytoplasmic dots, but did not react with recombinant GW182, suggesting that they were directed against an additional component(s) of these structures. The cytoplasmic dot staining pattern was observed in 21 of 493 (4.3%) patients with PBC. In comparison, this staining pattern was not produced by serum from 248 patients with other autoimmune diseases. CONCLUSION: A subset of patients with PBC have autoantibodies that produce the cytoplasmic dot staining pattern. These antibodies react with one or more as yet unidentified components of the mRNA processing body. Appreciation of the clinical significance of the cytoplasmic dot staining pattern may assist in appropriate diagnosis and treatment of patients with PBC.

Animals↗
PubMed2004

Binding and interstitial penetration of liposomes within avascular tumor spheroids.

The liposomal delivery of cancer therapeutics, including gene therapy vectors, is an area of intense study. Poor penetration of liposomes into interstitial tumor spaces remains a problem, however. In this work, the penetration of different liposomal formulations into prostate carcinoma spheroids was examined. Spheroid penetration was assessed by confocal microscopy of fluorescently labeled liposomes. The impact of liposomal surface charge, mean diameter, lipid bilayer fluidity and fusogenicity on spheroid penetration was examined. A variety of different liposome systems relevant to clinical or preclinical protocols have been studied, including classical zwitterionic (DMPC:chol) and sterically stabilized liposomes (DMPC:chol:DOPE-PEG2000), both used clinically, and cationic liposomes (DMPC:DOPE:DC-chol and DOTAP), forming the basis of the vast majority of nonviral gene transfer vectors tested in various cancer trials. Surface interactions between strongly cationic vesicles and the tumor cells led to an electrostatically derived binding-site barrier effect, inhibiting further association of the delivery systems with the tumor spheroids (DMPC:DC-chol). However, inclusion of the fusogenic lipid DOPE and use of a cationic lipid of lower surface charge density (DOTAP instead of DC-chol) led to improvements in the observed intratumoral distribution characteristics. Sterically stabilized liposomes did not interact with the tumor spheroids, whereas small unilamellar classical liposomes exhibit extensive distribution deeper into the tumor volume. Engineering liposomal delivery systems with a relatively low charge molar ratio and enhanced fusogenicity, or electrostatically neutral liposomes with fluid bilayers, offered enhanced intratumoral penetration. This study shows that a delicate balance exists between the strong affinity of delivery systems for the tumor cells and the efficient penetration and distribution within the tumor mass, similar to previous work studying targeted delivery by ligand-receptor interactions of monoclonal antibodies. Structure-function relationships from the interaction of different liposome systems with 3-dimensional tumor spheroids can lead to construction of delivery systems able to target efficiently and penetrate deeper within the tumor interstitium and act as a screening tool for a variety of therapeutics against cancer.

Antibodies, Monoclonal↗
PubMed2004

Alpha-particle emitting atomic generator (Actinium-225)-labeled trastuzumab (herceptin) targeting of breast cancer spheroids: efficacy versus HER2/neu expression.

PURPOSE: The humanized monoclonal antibody, trastuzumab (Herceptin), directed against HER2/neu, has been effective in the treatment of breast cancer malignancies. However, clinical activity has depended on HER2/neu expression. Radiolabeled trastuzumab has been considered previously as a potential agent for radioimmunotherapy. The objective of this study was to investigate the efficacy of trastuzumab labeled with the alpha-particle emitting atomic generator, actinium-225 ((225)Ac), against breast cancer spheroids with different HER2/neu expression levels. (225)Ac has a 10-day half-life and a decay scheme yielding four alpha-particles. EXPERIMENTAL DESIGN: The breast carcinoma cell lines MCF7, MDA-MB-361 (MDA), and BT-474 (BT) with relative HER2/neu expression (by flow cytometry) of 1:4:18 were used. Spheroids of these cell lines were incubated with different concentrations of (225)Ac-trastuzumab, and spheroid growth was measured by light microscopy over a 50-day period. RESULTS: The activity concentration required to yield a 50% reduction in spheroid volume at day 35 was 18.1, 1.9, and 0.6 kBq/ml (490, 52, 14 nCi/ml) for MCF7, MDA, and BT spheroids, respectively. MCF7 spheroids continued growing but with a 20-30 day growth delay at 18.5 kBq/ml. MDA spheroid growth was delayed by 30-40 days at 3.7 kBq/ml; at 18.5 kBq/ml, 12 of 12 spheroids disaggregated after 70, days and cells remaining from each spheroid failed to form colonies within 2 weeks of being transferred to adherent dishes. Eight of 10 BT spheroids failed to regrow at 1.85 kBq/ml. All of the BT spheroids at activity concentrations 3.7 kBq/ml failed to regrow and to form colonies. The radiosensitivity of these three lines as spheroids was evaluated as the activity concentration required to reduce the treated to untreated spheroid volume ratio to 0.37, denoted DVR(37). An external beam radiosensitivity of 2 Gy was found for spheroids of all three of the cell lines. After alpha-particle irradiation a DVR(37) of 1.5, 3.0, and 2.0 kBq/ml was determined for MCF7, MDA, and BT, respectively. CONCLUSION: These studies suggest that (225)Ac-labeled trastuzumab may be a potent therapeutic agent against metastatic breast cancer cells exhibiting intermediate to high HER2/neu expression.

Actinium↗
PubMed2004

Do antinuclear antibodies in primary biliary cirrhosis patients identify increased risk for liver failure?

BACKGROUND & AIMS: Primary biliary cirrhosis (PBC) is a chronic cholestatic liver disease with a variable clinical course. Identification of serologic markers associated with increased risk of liver failure would assist in management of PBC patients. The objective of this study was to identify antinuclear antibody (ANA) markers that may be used to predict PBC outcome. METHODS: Indirect immunofluorescence was used to identify ANAs in 492 PBC patients. chi2 and Kaplan-Meier analyses were used to examine the association between ANAs and liver failure. RESULTS: A greater percentage of ANA-positive, compared to ANA-negative, PBC patients developed liver failure (41% vs 25%, P = .005). The presence of anti-centromere antibodies was associated with liver failure (anti-centromere antibody positive vs negative, 58% vs 33%, P = .001). The time to liver failure was shorter in ANA-positive, compared with ANA-negative, patients (log rank score 5.8, P = .02). After 8.9 years (the median follow-up for patients without liver failure), 68% of ANA-positive and 81% of ANA-negative patients were free of liver failure. Anti-centromere antibodies were also associated with a shorter time to liver failure (log rank score 8.4, P = .004). After 8.9 years, 52% of anti-centromere antibody positive and 74% of anti-centromere antibody negative patients were without liver failure. CONCLUSIONS: ANAs in general, and anti-centromere antibodies in particular, are associated with liver failure in PBC. PBC patients with ANAs may be candidates for treatment with experimental therapies to prolong the interval between diagnosis and liver failure. ANA-negative patients, who appear to have a relatively benign clinical course, should perhaps be treated with ursodeoxycholic acid alone.

Antibodies, Antinuclear↗
PubMed2003

Combination treatment with 17-N-allylamino-17-demethoxy geldanamycin and acute irradiation produces supra-additive growth suppression in human prostate carcinoma spheroids.

Failure to control localized prostate cancer can result not only in localized disease progression but also distant metastatic spread. Whereas significant advances in both surgical technique and radiation therapy have improved local control rates with decreased morbidity, consistent long-term control remains elusive. This study investigates the potential of 17-N-allylamino-17-demethoxy geldanamycin (17AAG), a geldanamycin derivative, to sensitize tumor cells to ionizing radiation, permitting a significant improvement to targeted radiotherapies of prostate carcinoma. As a monotherapeutic, 17AAG functions to modulate the action of heat shock protein 90, ultimately affecting a multitude of cellular signaling pathways. It is in Phase I trial and has shown promise in controlling prostate cancer progression. Human prostate tumor cells (LNCaP and CWR22Rv1) were grown as spheroids and incubated for 96 h with increasing doses of 17AAG immediately before and after 2 or 6 Gy low linear energy transfer (LET), high dose-rate irradiation (Cs-137 irradiator). Twelve or 24 spheroids (initial diameter, 150-200 microm) were used per experiment. Response was determined by spheroid volume measurements taken over at least 40 days, after treatment. Incubation of either cell line with 17AAG (<or=1000 nM) or irradiation (<or=6 Gy) alone resulted in transient median growth delays ranging from 2 to 9 days (relative to controls). Combining treatments produced dose- and cell line-dependent supra-additive responses. For LNCaP spheroids, the combination of 2 Gy and 100 nM 17AAG resulted in growth delays additive of the treatments individually; however, increasing either the radiation to 6 Gy or the 17AAG concentration to 1000 nM led to synergistic interactions. Similarly, synergy was noted in CWR22Rv1 studies at only 6 Gy and 1000 nM 17AAG. Terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) and Ki67 staining of spheroid sections revealed the increased growth control to be a function of spheroids failing to re-enter the cell cycle. For all 6 Gy experiments, cells remaining from each of the spheroids that failed to regrow were transferred to adherent dishes to evaluate clonogenicity; growth-controlled spheroids also failed to form colonies within 2 weeks of being plated. These results suggest that significant gains in treatment effectiveness may be obtained by combining these treatment modalities, warranting additional preclinical investigation.

Benzoquinones↗