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At least 541 records · Page 30Linked to original sources
PubMed2006

HER-2/neu-gene engineered dendritic cell vaccine stimulates stronger HER-2/neu-specific immune responses compared to DNA vaccination.

HER-2/neu is a candidate for developing breast cancer-targeted immunotherapeutics. Although DNA-based and HER-2/neu transgene-modified dendritic cell (DC)-based vaccines are potent at eliciting HER-2/neu-specific antitumor immunity, there has been no side-by-side study comparing them directly. The present study utilizes an in vivo murine tumor model expressing HER-2/neu antigen to compare the efficacy between adenovirus (AdVneu)-transfected dendritic cells (DC(neu)) and plasmid DNA (pcDNAneu) vaccine. Our data showed that DC(neu) upregulated the expression of immunologically important molecules and inflammatory cytokines and partially converted regulatory T (Tr)-cell suppression through interleukin-6 (IL-6) secretion. Vaccination of DC(neu) induced stronger HER-2/neu-specific humoral and cellular immune responses than DNA vaccination, which downregulated HER-2/neu expression and lysed HER-2/neu-positive tumor cells in vitro, respectively. In two HER-2/neu-expressing tumor models, DC(neu) completely protected mice from tumor cell challenge compared to partial or no protection observed in DNA-immunized mice. In addition, DC(neu) significantly delayed breast cancer development in transgenic mice in comparison to DNA vaccine (P<0.05). Taken together, we have demonstrated that HER-2/neu-gene-modified DC vaccine is more potent than DNA vaccine in both protective and preventive animal tumor models. Therefore, DCs genetically engineered to express tumor antigens such as HER-2/neu represent a new direction in DC vaccine of breast cancer.

Adenoviridae↗
PubMed2001

A self-immunomodulating myoblast cell line for erythropoietin delivery.

The transplantation of genetically engineered cells faces limitations associated with host immunity. Allogeneic cells are typically rejected in response to inherent histo-incompatibility. Even autologous cells can induce an immune response toward antigenic molecules expressed following transfer of foreign genes. The goal of the present study was to investigate the ability of immunomodulating molecules co-expressed with biotherapeutic factors to overcome these limitations both in syngeneic and allogeneic cell transplantation. The C(2)C(12) mouse myoblast cell line was engineered to express CTLA4Ig, a soluble factor blocking T cell costimulation, in conjunction with erythropoietin (Epo), a reporter biotherapeutic protein. In syngeneic C3H mice, myoblasts expressing only mouse Epo were mostly rejected within 2 weeks, as indicated by the transient increase in host hematocrit. In allogeneic recipients, the same cells induced only a 1-week increase in the hematocrit reflecting an acute rejection process. CTLA4Ig expression significantly extended the survival of mouse Epo-secreting myoblasts in approximately half of syngeneic hosts, whereas it led only to a 1-week improvement effect in allogeneic recipients. When combined with a transient anti-CD154 treatment, CTLA4Ig expression prevented Epo-secreting C(2)C(12)myoblasts from being rejected in allogeneic DBA/2J recipients for at least 1 month. In contrast, the same anti-CD154 treatment alone induced only a 1 week improvement. These results demonstrate that CTLA4Ig co-expression associated with a transient anti-CD154 treatment can prolong the delivery of recombinant proteins via transfer of ex vivo modified cells in allogeneic recipients.

Abatacept↗
PubMed2002

Tumor suppression through angiogenesis inhibition by SUIT-2 pancreatic cancer cells genetically engineered to secrete NK4.

NK4, composed of the N-terminal hairpin and subsequent four-kringle domains of hepatocyte growth factor (HGF), acts not only as a competitive antagonist of HGF but also as an inhibitor of angiogenesis. By studying the antitumor effect of NK4, we evaluated the potential of gene therapy with NK4 as a treatment for pancreatic cancer. Expression vector pcDNA3-NK4 containing NK4 cDNA was used to transfect human pancreatic cancer cell line SUIT-2. Although the established NK4 transfectant continuously expressed NK4 protein, the expression was shown by migration assay to be insufficient to antagonize HGF in vitro. Proliferation of the NK4 transfectant did not differ significantly from that of a mock transfectant. In vivo, we used models of orthotopic implantation and liver metastasis to transplant NK4-transfected clone or mock-transfected clone into nude mice. Cell proliferation in vivo, evaluated by immunohistochemical staining of proliferating cell nuclear antigen, did not differ between NK4 and mock transfectants, and this was also the finding in the in vitro assay. However, the NK4-transfected clone showed significant inhibition of tumor progression in both the orthotopic implantation and liver metastasis models. The number of vessels within tumors was significantly decreased, and the apoptotic tumor cells were increased in number. The results of these experiments show that genetic modification of tumor cells with NK4 cDNA yields a significant antitumor effect and that this effect is mainly obtained by NK4's function as an angiogenesis inhibitor rather than as an HGF antagonist. We conclude that the potent angiogenesis inhibitor NK4 may be a promising molecule for gene therapy of pancreatic cancer.

Animals↗
PubMed2003

Recombinant immunotoxins and retargeted killer cells: employing engineered antibody fragments for tumor-specific targeting of cytotoxic effectors.

Over the past years, monoclonal antibodies have attracted enormous interest as targeted therapeutics, and a number of such reagents are in clinical use. However, responses could not be achieved in all patients with tumors expressing high levels of the respective target antigens, suggesting that other factors such as limited recruitment of endogenous immune effector mechanisms can also influence treatment outcome. This justifies the search for alternative, potentially more effective reagents. Antibody-toxins and cytolytic effector cells genetically modified to carry antibody-based receptors on the surface, represent such tailor-made targeting vehicles with the potential of improved tumor localization and enhanced efficacy. In this way, advances in recombinant antibody technology have made it possible to circumvent problems inherent in chemical coupling of antibodies and toxins, and have allowed construction via gene fusion of recombinant molecules which combine antibody-mediated recognition of tumor cells with specific delivery of potent protein toxins of bacterial or plant origin. Likewise, recombinant antibody fragments provide the basis for the construction of chimeric antigen receptors that, upon expression in cytotoxic T lymphocytes (CTLs) or natural killer (NK) cells, link antibody-mediated recognition of tumor antigens with these effector cells' potent cytolytic activities, thereby making them promising cellular therapeutics for adoptive cancer therapy. Here, general principles for the derivation of cytotoxic proteins and effector cells with antibody-dependent tumor specificity are summarized, and current strategies to employ these molecules and cells for directed cancer therapy are discussed, focusing mainly on the tumor-associated antigens epidermal growth factor receptor (EGFR) and the closely related ErbB2 (HER2) as targets.

ErbB Receptors↗
PubMed2000

Neuroprotective gene therapy for Huntington's disease using a polymer encapsulated BHK cell line engineered to secrete human CNTF.

Huntington's disease (HD) is an autosomal dominant genetic disease with devastating clinical effects on cognitive, psychological, and motor functions. These clinical symptoms primarily relate to the progressive loss of medium-spiny GABA-ergic neurons of the striatum. There is no known treatment to date. Several neurotrophic factors have, however, demonstrated the capacity to protect striatal neurons in various experimental models of HD. This includes the ciliary neurotrophic factor (CNTF), the substance examined in this protocol. An ex vivo gene therapy approach based on encapsulated genetically modified BHK cells will be used for the continuous and long-term intracerebral delivery of CNTF. A device, containing up to 106 human CNTF-producing BHK cells surrounded by a semipermeable membrane, will be implanted into the right lateral ventricle of 6 patients. Capsules releasing 0.15-0.5 microg CNTF/day will be used. In this phase I study, the principal goal will be the evaluation of the safety and tolerability of the procedure. As a secondary goal, HD symptoms will be analyzed using a large battery of neuropsychological, motor, neurological, and neurophysiological tests and the striatal pathology monitored using MRI and PET-scan imaging. It is expected that the gene therapy approach described in this protocol will mitigate the side effects associated with the peripheral administration of recombinant hCNTF and allow a well-tolerated, continuous intracerebroventricular delivery of the neuroprotective factor.

Animals↗
PubMed2004

Amniotic fluid and placental stem cells.

The amniotic fluid and the placenta are unique sources of different populations of stem cells--mesenchymal, hematopoietic, trophoblastic--and, possibly, of more primitive stem cells. Although much of the amniotic cavity/fluid and the placenta share a common embryonic origin, the specific origins of the stem cells found in these two compartments remain to be determined. Accordingly, it is not yet known whether all or part of these two stem-cell subsets are actually the same. The multilineage potential of the different stem cell populations from these two sources has begun to be described but still much remains to be learned. Thus, it is not surprising that clinical applications related to the use of these cells have yet to be reported. Nevertheless, fertile experimental work from many different groups has introduced a number of promising novel therapeutic concepts utilizing these cells, such as in tissue engineering, cell transplantation, and gene therapy.

Amnion↗
PubMed2004

[Embryonic and adult stem cells for tissue engineering in urology].

Stem cells are considered a possible alternative treatment for degenerative diseases. Parkinson's disease and diabetes mellitus may become curable using stem cells. The term "stem cell" is used very broadly, although it has been well defined. Undifferentiated cells (stem cells and progenitor cells) are isolated from prenatal and postnatal tissues. This review provides an overview of embryonic and adult stem cells and describes potential applications in the field of tissue engineering in urology.

Adult↗
PubMed2006

From hollow shells to artificial cells: biointerface engineering on polyelectrolyte capsules.

Biomimetic composites can be fabricated by coating hollow polyelectrolyte capsules with biological interfaces such as a phospholipid membrane and proteins. Polyelectrolyte capsules have been templated applying the Layer-by-Layer technique of polyelectrolyte assembled on decomposable cores, which are destroyed after the assembly of the polyelectrolyte multilayer. Phospholipid vesicles of 200-300 nm size are spreaded on the capsule wall forming a continuous lipid membrane. Further functionalisation of the outer capsule wall can be achieved with fused virions and recrystallised S-layers. Compartimentation of the capsule interior with lipid vesicles has been possible by using a solvent exchange method. The functionalisation of the outer capsule surface with biomolecules, together with the creation of internal compartments in the capsule, open new nanobiotechnological challenges towards the fabrication of artificial cells.

Bacillus↗
PubMed2004

Gene-engineered T cells as a superior adjuvant therapy for metastatic cancer.

The major limiting factor in the successful application of adjuvant therapy for metastatic disease is the lack of adjuvant specificity that leads to severe side effects. Reasoning that T cells of the immune system are highly specific, we generated tumor-specific T cells by genetic modification of mouse primary T cells with a chimeric receptor reactive with the human breast cancer-associated Ag erbB-2. These T cells killed breast cancer cells and secreted IFN-gamma in an Ag-specific manner in vitro. We investigated their use against metastatic breast cancer in mice in an adjuvant setting, and compared their effectiveness with the commonly applied adjuvants doxorubicin, 5-fluorouracil, and herceptin. Mice were inoculated orthotopically with the human erbB-2-expressing spontaneously metastatic mouse breast cancer 4T1.2 in mammary tissue, and the primary tumor was surgically removed 8 days later. Significant metastatic disease was demonstrated in lung and liver at the time of surgery on day 8 with increased tumor burden at later time points. T cell adjuvant treatment of day 8 metastatic disease resulted in dramatic increases in survival of mice, and this survival was significantly greater than that afforded by either doxorubicin, 5-fluorouracil, or herceptin.

Animals↗
PubMed2004

Harnessing the catabolic diversity of rhodococci for environmental and biotechnological applications.

The field of Rhodococcus cell engineering is rapidly advancing because of the availability of improved genetic tools and increased insights in their broad catabolic and biochemical diversity. Rhodococci harbor large linear plasmids that may contribute to their catabolic diversity. In addition, multiple pathways and gene homologs are often present, thus further increasing Rhodococcus catabolic versatility and efficiency. The recent development of effective genetic tools for Rhodococcus, such as unmarked gene deletion, transposon-based mutagenesis, and gene expression systems, now allows the construction of biocatalysts with desirable properties for industrial purposes. This is exemplified here by a description of cell engineering of biocatalysts for improved desulphurization and steroid biotransformation.

Androstenedione↗
PubMed2000

Organic transformations catalyzed by engineered yeast cells and related systems.

Asymmetric ketone reductions remain the most popular application of baker's yeast (Saccharomyces cerevisiae) in organic synthesis and data from the genome sequencing project is beginning to have an impact on improving the stereoselectivities of these reactions, augmenting traditional approaches based on selective inhibition. In addition, the catalytic repertoire of yeast has been expanded to include chiral ketone oxidations by overexpression of a bacterial Baeyer-Villiger monooxygenase.

Alcohol Oxidoreductases↗
PubMed2005

The specific delivery of proteins to human liver cells by engineered bio-nanocapsules.

A bio-nanocapsule (BNC), composed of the surface antigen (sAg) of the hepatitis B virus, is an efficient nanomachine with which to accomplish the liver-specific delivery of genes and drugs. Approximately 110 molecules of sAg are associated to form a BNC particle with an average diameter of 130 nm. The L protein is an sAg peptide composed mainly of preS and S regions. The preS region, with specific affinity for human hepatocytes, is localized in the N-terminus. The S region following the preS has two transmembrane regions responsible for the formation of particles. In this study, the fusion of emerald green fluorescent protein (EGFP) at the C-terminus of the S region was designed to deliver proteins to human hepatocytes. Truncation of the C-terminus of the S region was required to obtain sufficient expression levels in Cos7 cells. The nanoparticles that were produced delivered EGFP to human hepatoma cells, displaying the EGFP moiety outside, or enclosing it inside. However, only a single orientation characterizes the particle, so that either type of L fusion particle could be effectively and independently separated by an antibody affinity column. The dual C-terminal topologies of the L fusion particles designed in this study could be applied to various proteins for the C-terminal moiety of the L fusion proteins, depending on the character of the proteins, such as cytoplasmic proteins, as well as cytokines or ligands to cell surface receptors. We suggest that this fusion design is the most efficient way to prepare a BNC that delivers proteins to specific cells or tissues.

Amino Acid Sequence↗
PubMed1999

Hepatic metabolism of diclofenac: role of human CYP in the minor oxidative pathways.

The aim of this study was to re-examine the human hepatic metabolism of diclofenac, with special focus on the generation of minor hydroxylated metabolites implicated in the idiosyncratic hepatotoxicity of the drug. Different experimental approaches were used: human hepatocytes, human microsomes, and engineered cells expressing single human CYP (cytochromes P450). Human hepatocytes formed 3'-hydroxy-, 4'-hydroxy-, 5-hydroxy- 4',5-dihydroxy-, and N,5-dihydroxydiclofenac, as well as several lactams. Formation of 4'- and 5-hydroxydiclofenac by human liver microsomes followed a Michaelis-Menten kinetics (Km 9 +/- 1 microM; Vmax 432 +/- 15 pmol/min/mg and Km 43 +/- 5 microM; and Vmax 15.4 +/- 0.6 pmol/min/mg, respectively). Secondary metabolites were detected after incubation of 5-hydroxydiclofenac with human liver microsomes, yielding 4',5-dihydroxydiclofenac (Km 15 +/- 1 microM; Vmax 96 +/- 3 pmol/min/mg) and small amounts of N,5-dihydroxydiclofenac (non-Michaelis-Menten kinetics). Based on microsome studies and the incubations with human hepatocytes and engineered cells, we estimated that in vivo CYP2C9 would be exclusively responsible for the 4' hydroxylation of diclofenac (>99.5%) as well as 5-hydroxydiclofenac (>97%). CYP2C9 was exclusively responsible for the formation of 3'-hydroxydiclofenac. Multiple regression analysis evidenced that the rate of production of 5-hydroxydiclofenac in human microsomes followed the algorithm: 0.040 x S-mephenytoin 4'-hydroxylation + 0.083 x tolbutamide methylhydroxylation, (multiple correlation coefficient = 0.969). However, the incubation of diclofenac with cell lines expressing different human CYP suggested that 7 isoforms could be involved. Comparison of data obtained with CYP-expressing cells and human hepatocytes suggests that CYP2C8 > CYP2C19 approximately CYP2C18 >> CYP2B6 are the isoforms implicated in the 5-hydroxylation of diclofenac in vivo.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1991

Artificial cell adhesive proteins engineered by grafting the Arg-Gly-Asp cell recognition signal: factors modulating the cell adhesive activity of the grafted signal.

An artificial cell adhesive protein could be engineered by grafting the RGDS tetrapeptide, the core sequence of the major cell adhesive site of fibronectin, to a truncated form of Staphylococcal protein A (tSPA) via cassette mutagenesis of the tSPA expression vector pRIT2T [T. Maeda et al. (1989) J. Biol. Chem. 264, 15165-15168]. We synthesized a panel of tSPA derivatives grafted with various RGDS-containing oligopeptides to address the problem of how the cell adhesive activity of the resulting tSPA derivatives was affected by the length and amino acid sequence of the grafted oligopeptides and by the sites on tSPA where the extra oligopeptides were inserted. The results showed that (i) the amino acid residues flanking the RGDS core sequence played a key role in modulating the cell adhesive activity of the grafted RGDS signal; (ii) at least two sites on tSPA, each corresponding to on e of the two HindIII sites of pRIT2T, were competent in sustaining the cell adhesive activity of the grafted signal; and (iii) the divalent tSPA containing the RGDS signal at both sites was more active than monovalent derivatives containing only one signal at either site. These results provide a strategic basis for engineering of artificial cell adhesive proteins by grafting the RGDS signal.

Amino Acid Sequence↗
PubMed2003

Enhanced toxic metal accumulation in engineered bacterial cells expressing Arabidopsis thaliana phytochelatin synthase.

Phytochelatins (PCs) are metal-binding cysteine-rich peptides, enzymatically synthesized in plants and yeasts from glutathione in response to heavy metal stress by PC synthase (EC 2.3.2.15). In an attempt to increase the ability of bacterial cells to accumulate heavy metals, the Arabidopsis thaliana gene encoding PC synthase (AtPCS) was expressed in Escherichia coli. A marked accumulation of PCs was observed in vivo together with a decrease in the glutathione cellular content. When bacterial cells expressing AtPCS were placed in the presence of heavy metals such as cadmium or the metalloid arsenic, cellular metal contents were increased 20- and 50-fold, respectively. We discuss the possibility of using genes of the PC biosynthetic pathway to design bacterial strains or higher plants with increased abilities to accumulate toxic metals, and also arsenic, for use in bioremediation and/or phytoremediation processes.

Aminoacyltransferases↗
PubMed2006

[Cell culture, tissue engineering and regenerative medicine. Part I].

Tissue engineering is an interdisciplinary field that applies the principles and methods of engineering and the cell culture toward the development of biomaterials that restore, maintain or improve tissue function. The amalgamation of engineering and medicine has interested many scientists for at last two hundred years. What was the goal of cell culture? First, for progress in life sciences achievement and subsequent for virology and toxicology development. In vitro studies are done because of many problems with carrying out animal experiments. In this work the authors present the attempts of physicians, anatomopathologists, embryologists and biologists which contributed to fast development of new area in medicine--tissue engineering.

Animals↗
PubMed2006

Blood vessels engineered from human cells.

Human vascular tissue engineering has progressed substantially over the past decade. Issues remain as to the optimum culture scaffold and cell source that should be used for arterial regeneration. In addition, limitations in replicative life span of adult human cells as compared with young animal cells have made human vascular tissue engineering more challenging. Despite these obstacles, several strategies for generating replacement vessels have progressed to clinical trials.

Age Factors↗