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J A Roth

Publications and source records attributed to J A Roth.

At least 127 records · Page 7Linked to original sources
PubMed1998

A multidisciplinary approach to therapy for unresectable malignant thymoma.

BACKGROUND: The therapeutic outcome for unresectable, locally advanced, malignant thymoma has been poor. OBJECTIVE: To improve tumor resectability and patient survival rates by studying a multimodal approach to therapy for unresectable malignant thymoma. DESIGN: Prospective cohort study. SETTING: Tertiary care cancer center. PARTICIPANTS: All eligible patients had newly diagnosed, histologically proven, unresectable malignant thymoma. INTERVENTION: The treatment regimen consisted of induction chemotherapy (three courses of cyclophosphamide, doxorubicin, cisplatin, and prednisone), surgical resection, postoperative radiation therapy, and consolidation chemotherapy (three courses of cyclophosphamide, doxorubicin, cisplatin, and prednisone). Tissue samples were taken at the time of surgical resection for assessment of tumor necrosis and Ki-67 expression. MEASUREMENTS: Tumor response and resectability (both overall and after induction chemotherapy) and disease-free survival rate in patients who received multimodal therapy. RESULTS: 13 patients were consecutively enrolled from February 1990 to December 1996, and 12 evaluable patients were assessed for response. Disease responded to induction chemotherapy completely in 3 patients (25%) and partially in 8 patients (67%); 1 patient had a minor response (8%). Eleven patients had surgical resection; 1 refused surgery. Tumors were removed completely in 9 (82%) and incompletely in 2 (18%) of 11 patients who had been receiving radiation therapy and consolidation chemotherapy. All 12 patients are alive (100% at 7 years), with a median follow-up of 43 months, and 10 patients are disease free (73% disease-free survival at 7 years). A high correlation was seen between tumor necrosis after induction chemotherapy and Ki-67 expression (r=-0.88). CONCLUSIONS: Aggressive multimodal treatment is highly effective and may cure locally advanced, unresectable malignant thymoma.

Adult↗
PubMed1998

Adenovirus-mediated wild-type p53 gene transfer down-regulates vascular endothelial growth factor expression and inhibits angiogenesis in human colon cancer.

Recent studies have indicated that angiogenesis may be regulated, in part, by p53 tumor suppressor gene function. We hypothesized that wild-type p53 replacement would down-regulate vascular endothelial growth factor (VEGF) expression and inhibit angiogenesis. KM12L4 and SW620, human colon cancer cell lines with p53 mutations, were transduced with a replication-defective adenoviral vector containing the wild-type p53 gene (Ad5/CMV/p53). Reverse transcription-PCR confirmed the presence of p53 in Ad5/CMV/p53-transduced cells. Transduction of colon cancer cells with wild-type p53 decreased VEGF RNA expression compared with that of controls. The decrease in VEGF expression in SW620 cells was dose dependent, with a 49% decrease observed at a multiplicity of infection of 50, and a 71% decrease observed at a multiplicity of infection of 100. Similar effects were seen in KM12L4 cells. VEGF supernatant protein levels were significantly reduced compared with those in nontransduced controls 48 h after the introduction of wild-type p53. Ad5/CMV/p53 inhibited tumor cell-induced angiogenesis in vivo. Restoration of wild-type p53 expression may decrease tumor growth by inhibiting the angiogenic response. These findings may explain, in part, the bystander effect seen with p53 tumor suppressor gene therapy.

Adenoviridae↗
PubMed1998

Evaluation of GAL4/TATA in vivo. Induction of transgene expression by adenovirally mediated gene codelivery.

A synthetic GAL4-responsive promoter consisting of five GAL4-binding sites and a TATA box (GAL4/TATA) was evaluated for its transcriptional activity in an adenoviral backbone using luciferase as the reporter. Basal luciferase activities in vitro were the same for cells infected with either adenovirus-containing luciferase cDNA driven by GAL4/TATA (Ad/GT-Luc) or adenovirus-containing luciferase cDNA not driven by a promoter (Ad/PO-Luc). In vitro induction of GAL4/TATA by coinfection of cells with adenovirus expressing the GAL4/VP16 fusion protein (Ad/3-phosphoglycerate kinase (PGK)-GV16) was dose-dependent and reached as high as 4 x 10(4)- to 9 x 10(4)-fold above basal levels when GAL4/TATA and GAL4/VP16 were delivered at a ratio of 10:1. In vivo studies in Balb/c mice showed no detectable luciferase activities in liver or other tissues examined in mice infused with either Ad/GT-Luc or Ad/PO-Luc. High levels of luciferase activity were, however, elicited when animals were infused with Ad/GT-Luc and Ad/PGK-GV16. Together, these results suggest that combination of the GAL4 gene regulatory system with adenovirally mediated in vivo gene delivery may be applicable to the in vivo evaluation of promoter activities and in vivo targeting of gene expression.

Adenoviridae↗
PubMed1998

Expression of p16 induces transcriptional downregulation of the RB gene.

The RB and p16(INK4A) tumor suppressor genes function in the same pathway of cell cycle control. Previous evidence indicates that the p16(INK4A) gene is transcriptionally repressed by the RB gene product, pRB. In this study using human ovarian cancer cell lines, we found that RB protein and mRNA were expressed at higher levels in cell lines lacking p16 than in those with normal p16. Since this suggests a potential role of p16 in regulating the cellular level of pRB, we studied the effect of wild-type p16(INK4A) on expression of the RB gene. Introduction of p16(INK4A), carried by an adenovirus vector, into p16-negative cell lines dramatically decreased expression of RB protein and mRNA. Nuclei run-off assays demonstrated that p16 expression induced transcriptional downregulation of the RB gene. These results indicate that expression of RB is inversely regulated by p16. The findings reveal a new dimension of pRB-p16 interaction and should have implications for p16(INK4A)-mediated gene therapy.

Adenoviridae↗
PubMed1998

Adenovirus-mediated wild-type p53 tumor suppressor gene therapy induces apoptosis and suppresses growth of human pancreatic cancer [seecomments].

BACKGROUND: The p53 tumor suppressor gene is mutated in up to 70% of pancreatic adenocarcinomas. We determined the effect of reintroduction of the wild-type p53 gene on proliferation and apoptosis in human pancreatic cancer cells using an adenoviral vector containing the wild-type p53 tumor suppressor gene. METHODS: Transduction efficiencies of six p53-mutant pancreatic cancer cell lines (AsPC-1, BxPC-3, Capan-1, CFPAC-1, MIA PaCa-2, and PANC-1) were determined using the reporter gene construct Ad5/CMV/beta-gal. Cell proliferation was monitored using a 3H-thymidine incorporation assay, Western blot analysis for p53 expression was performed, and DNA laddering and fluorescence-activated cell sorter analysis were used to assess apoptosis. p53 gene therapy was tested in vivo in a subcutaneous tumor model. RESULTS: The cell lines varied in transduction efficiency. The MIA PaCa-2 cells had the highest transduction efficiency, with 65% of pancreatic tumor cells staining positive for beta-galactosidase (beta-gal) at a multiplicity of infection (MOI) of 50. At the same MOI, only 15% of the CFPAC-1 cells expressed the beta-gal gene. Adenovirus-mediated p53 gene transfer suppressed growth of all human pancreatic cancer cell lines in a dose-dependent manner. Western blot analysis confirmed the presence of the p53 protein product at 48 hours after infection. DNA ladders demonstrated increased chromatin degradation, and fluorescence-activated cell sorter analysis demonstrated a four-fold increase in apoptotic cells at 48 and 72 hours following infection with Ad5/CMV/p53 in the MIA PaCa-2 and PANC-1 cells. Suppression of tumor growth mediated by induction of apoptosis was observed in vivo in an established nude mouse subcutaneous tumor model following intratumoral injections of Ad5/CMV/p53. CONCLUSIONS: Introduction of the wild-type p53 gene using an adenoviral vector in pancreatic cancer with p53 mutations induces apoptosis and inhibits cell growth. These data provide preliminary support for adenoviral mediated p53 tumor suppressor gene therapy of human pancreatic cancer.

Adenocarcinoma↗
PubMed1998

Long-term follow-up of patients enrolled in a randomized trial comparing perioperative chemotherapy and surgery with surgery alone in resectable stage IIIA non-small-cell lung cancer.

Our previously reported randomized study of patients with untreated, potentially resectable clinical stage IIIA non-small-cell lung cancer found that patients treated with perioperative chemotherapy and surgery had a significant increase in median survival compared to patients treated with surgery alone. We have now re-analyzed the results of the study with a median time from random allocation to analysis for all patients of 82 months. The increase in survival conferred by perioperative chemotherapy was maintained during the period of extended observation.

Antineoplastic Agents↗
PubMed1998

Immunology and prevention of infection in feedlot cattle.

This article reviews the basic types of immunity, the factors relevant to feedlot cattle that have been shown to suppress immune function, and what is known about the basis of protective acquired immunity against the common bovine respiratory pathogens. The characteristics and efficacy of vaccines for respiratory disease pathogens are also reviewed. Recommendations for optimizing vaccination are presented.

Animals↗
PubMed1998

Two-dimensional gel analysis of apoptosis-specific p53 isoforms induced by 2-methoxyestradiol in human lung cancer cells.

The natural metabolic byproduct of estradiol, 2-methoxyestradiol (2-MeOE2), induces apoptosis in human lung cancer cells by a p53-dependent mechanism. The expression of wild-type p53 isoforms was investigated in H1299 non-small cell lung carcinoma cells induced into apoptosis by 2-MeOE2. H1299 cells lack endogenous p53 and undergo predominantly a G1 arrest when infected with a recombinant wild-type p53 adenovirus. However, when H1299 cells transfected with p53 were treated with 2-MeOE2, they underwent rapid and extensive apoptosis. H1299 cells expressing mutant his273 p53 were unaffected by 2-MeOE2, indicating a dependence of 2-MeOE2-mediated apoptosis on the presence of a functional p53. Analysis of wild-type p53 phosphoisoforms in H1299 cells by two-dimensional gel electrophoresis revealed that 2-MeOE2 induced a unique group of acidic p53 isoforms. Although most of the wild-type p53 in untreated H1299 cells migrated as at least five diffuse species with isoelectric points from pH 5.5-6.3, as many as nine additional forms migrating toward the acidic region with pI values from 4.4-5.3 were detected in 2-MeOE2-treated apoptotic cells. Two other agents known to induce apoptosis, vinblastine and actinomycin D, induced a similar pattern of acidic p53 species as that observed for 2-MeOE2. The results indicated that the induction of apoptosis in H1299 cells by 2-MeOE2 is dependent on the upregulation of specific p53 isoforms. Identification of the specific p53 phosphoisoforms induced by MeOE2 will be an important step in understanding the regulation and function of p53 in apoptosis.

Journal Article↗
PubMed1998

Suppression of the immune response to an adenovirus vector and enhancement of intratumoral transgene expression by low-dose etoposide.

Adenoviral vectors are commonly used in gene therapy trials because of their efficiency in gene transfer. However, their use is limited by cellular and humoral immune responses that result in temporary transgene expression and reduced efficacy of repeated vector administration. We hypothesized that certain oncolytic agents commonly used to treat cancer patients could suppress the immune response to adenoviral vectors, and enable repeated adenovirus-mediated cancer gene therapy. Etoposide and cyclophosphamide were tested for their ability to suppress the humoral and cellular immune responses to an adenoviral vector in immunocompetent C3H mice. Intratumoral transgene expression was monitored in adenovirus-immunized animals treated with etoposide or cyclophosphamide. Neutralizing antibodies to adenovirus and cytotoxic T lymphocyte (CTL) lysis of virally transduced cells were significantly suppressed in mice treated with etoposide at 2 or 10 mg/kg/day or cyclophosphamide at 10 mg/kg/day compared with untreated mice (P < 0.05). Significantly larger areas of gene transduction were observed in treated animals compared with untreated mice or the mice treated with cyclophosphamide at 2 mg/kg/day (P < 0.05). Our results suggest that repeated adenovirally mediated gene therapy is achievable in cancer patients who are concurrently undergoing treatment with chemotherapy.

Adenoviridae↗
PubMed1998

Monitoring adenoviral p53 transduction efficiency by yeast functional assay.

Monitoring the transduction efficiency is of paramount importance in gene therapy. To monitor adenovirus-mediated wild-type p53 gene transfer, we have used a quantitative assay which tests the ability of human p53 to activate transcription in yeast. Selective amplification of cellular and viral p53 transcripts followed by quantitative assessment of mutant p53 content with the assay permits measurement of the wild-type p53 transduction efficiency into SF-188, U251MG and HUG31 glioblastoma cells. One reverse transcription primer tracks the wild-type/mutant ratio of endogenous p53 mRNA (P2), and the other the wild-type/mutant ratio of both endogenous and exogenous p53 mRNA (P1). Following infection of cell lines homozygous for mutant p53, the apparent transduction efficiency calculated (tau 0 = [P1-P2]/[1 + P2]) correlated with the level of p21 expression. Transduction efficiency in heterozygous wild-type/mutant HUG31 cells increased linearly with multiplicity of infection (MOI) for tau 0 values between 0.5 and 5.9, and admixture of normal cell-derived RNA produced only a modest reduction in tau 0 value, in keeping with theoretical predictions. These results suggest that the yeast p53 functional assay may be a useful tool for monitoring p53 gene therapy.

Adenoviridae↗
PubMed1998

The contribution of poly-L-lysine, epidermal growth factor and streptavidin to EGF/PLL/DNA polyplex formation.

High-level targeted gene delivery has been demonstrated by molecular conjugates in vitro; however, in vivo delivery has been limited. The complexity of the resulting protein/DNA polyplex and a lack of understanding of its formation are persistent limitations. In this report, we show the effect of the DNA-binding agent poly-L-lysine (PLL), the ligand epidermal growth factor (EGF), and the coupling protein streptavidin on particle size, charge and gene delivery. Smaller (< 80 nm) and more stable polyplexes were obtained with PLL1116 than with shorter versions of PLL, especially in 0.15 M NaCl. Stability was increased by adding streptavidin to the polyplex; however, EGF increased particle size (> 1000 nm) and decreased gene delivery when > 300 EGF molecules per polyplex were used, indicating that a critical number of EGF molecules was needed for efficient gene delivery. The correct combination of these components resulted in the most efficient gene delivery in vitro and now provide for testing a more stable protein/DNA polyplex to aid in enhancing gene delivery in vivo.

Biotinylation↗
PubMed1998

Intercellular trafficking of VP22-GFP fusion proteins is not observed in cultured mammalian cells.

Herpes simplex virus type 1 (HSV-1) VP22 was recently reported to mediate intercellular trafficking of a protein fused to the C-terminus of VP22. To explore the application of such trafficking, we constructed plasmids expressing green fluorescent protein (GFP) fused to the C-terminus of either wild-type VP22 or a 160 amino acid peptide from VP22. In vitro studies showed that the majority of both fused proteins were localized to the nuclei of transfected cells. Quantitative analysis of GFP-positive cells, however, showed no significant increase in intercellular protein trafficking for cells transfected with either fusion protein compared with a lacZ-expressing plasmid. Our results suggest that the use of HSV-1 VP22 for mediating intercellular trafficking of transgene products is limited.

Animals↗
PubMed1998

Viral and nonviral gene delivery vectors for cancer gene therapy.

The development of vectors that are capable of efficient gene delivery is crucial to the success of gene therapy. We have developed both recombinant viral and nonviral vectors with the goal of correcting genetic abnormalities in cancer cells that are responsible for malignant transformation. Infection of cancer cells by recombinant adenovirus (Adv) indicates that the level of transduction is variable and dependent on the virus-to-cell ratio. Infection of cells with Adv/p53 resulted in levels of tumor suppressor p53 gene expression that could mediate tumor cell growth suppression and apoptosis, both in vitro and in vivo. The treatment of cancer cells with cisplatin prior to Adv transduction resulted in a higher level of therapeutic gene expression. Epidermal growth factor (EGF)/DNA complexes targeted to cancer cells overexpressing the EGF receptor resulted in efficient transduction of several lung cancer cell lines in vitro. As a result, these vectors provide improved methods with which to treat cancer in the clinical setting with gene therapy.

Adenoviridae↗
PubMed1998

Gene replacement strategies for lung cancer.

Considerable evidence has accumulated that cancer has a genetic origin based on the development of somatic mutations in families of genes responsible for critical functions of cellular DNA repair, growth control, and division. Restoration of the function of a single pivotal gene product appears sufficient to mediate antitumor effects that are potentially clinically significant. For example, restoration of wild-type p53 function in the cancer cell by gene transfer is sufficient to cause either cell-cycle arrest or apoptosis. This effect is not restricted to p53 but has been observed for oncogenes and other tumor suppressor genes as well. Genes can be delivered with sufficient efficiency by direct intratumoral injection to mediate tumor regression as shown in preclinical studies and phase I clinical trials in non-small cell lung cancer. Although clinical trials of gene replacement are in the earliest stages, this treatment offers a unique mechanism of action with a potentially high therapeutic index.

Carcinoma, Non-Small-Cell Lung↗
PubMed1998

Adenovirus-mediated p53 gene transfer in patients with advanced recurrent head and neck squamous cell carcinoma.

PURPOSE: Standard therapies of head and neck squamous cell carcinoma (HNSCC) often cause profound morbidity and have not significantly improved survival over the last 30 years. Preclinical studies showed that adenoviral vector delivery of the wild-type p53 gene reduced tumor growth in mouse xenograft models. Our purpose was to ascertain the safety and therapeutic potential of adenoviral (Ad)-p53 in advanced HNSCC. PATIENTS AND METHODS: Patients with incurable recurrent local or regionally metastatic HNSCC received multiple intratumoral injections of Ad-p53, either with or without tumor resection. Patients were monitored for adverse events and antiadenoviral antibodies, tumors were monitored for response and p53 expression, and body fluids were analyzed for Ad-p53. RESULTS: Tumors of 33 patients were injected with doses of up to 1 x 10(11) plaque-forming units (pfu). No dose-limiting toxicity or serious adverse events were noted. p53 expression was detected in tumor biopsies despite antibody responses after Ad-p53 injections. Clinical efficacy could be evaluated in 17 patients with nonresectable tumors: two patients showed objective tumor regressions of greater than 50%, six patients showed stable disease for up to 3.5 months, and nine patients showed progressive disease. One resectable patient was considered a complete pathologic response. Ad-p53 was detected in blood and urine in a dose-dependent fashion, and in sputum. CONCLUSION: Patients were safely injected intratumorally with Ad-p53. Objective antitumor activity was detected in several patients. The infectious Ad-p53 in body fluids was asymptomatic, and suggests that systemic or regional treatment may be tolerable. These results suggest the further investigation of Ad-p53 as a therapeutic agent for patients with HNSCC.

Adenoviridae↗
PubMed1998

[2-methoxyestradiol induces p53 independent apoptosis in pancratic carcinoma and inhibits growth of lung metastasis].

2-methoxyestradiol (2-ME) is able to inhibit growth of certain tumors. We were interested in studying this effect of 2-ME on pancreatic cancer. In three out of four cell lines, 2-ME induced 50-90% growth inhibition after 48 hours treatment with 2 microM 2-ME. TUNEL staining for apoptosis revealed that after treatment with 2 microM 2-ME for 48 hours 37-97% of these cells underwent apoptotic cell death (p < 0.005-0.0001). Since all cell lines used harbor a p53 mutation, this apoptosis appears to be p53 independent. One of these cell lines (MIAPaCa-2) was used for the in vivo experiment in nude mice. After a daily oral administration of 1 mg 2-ME for three weeks, the number of lung metastases could be reduced by 59%, as compared to the control non treated group (p < 0.0005). No signs of toxicity were seen. It appears that 2-ME is a potent compound for the treatment of pancreatic cancer, which might be interesting for clinical application.

2-Methoxyestradiol↗