In vivo CAR T-cell generation in nonhuman primates using lentiviral vectors displaying a multidomain fusion ligand.

Chimeric antigen receptor (CAR) T-cell therapies have demonstrated transformative efficacy in treating B-cell malignancies. However, high costs and manufacturing complexities hinder their widespread use. To overcome these hurdles, we have developed the VivoVec platform, a lentiviral vector capable of generating CAR T cells in vivo.

Preclinical proof of concept for VivoVec, a lentiviral-based platform for in vivo CAR T-cell engineering.

Background: Chimeric antigen receptor (CAR) T-cell therapies have demonstrated transformational outcomes in the treatment of B-cell malignancies, but their widespread use is hindered by technical and logistical challenges associated with ex vivo cell manufacturing. To overcome these challenges, we developed VivoVec, a lentiviral vector-based platform for in vivo engineering of T cells. UB-VV100, a VivoVec clinical candidate for the treatment of B-cell malignancies, displays an anti-CD3 single-chain variable fragment (scFv) on the surface and delivers a genetic payload that encodes a second-generation CD19-targeted CAR along with a rapamycin-activated cytokine receptor (RACR) system designed to overcome the need for lymphodepleting chemotherapy in supporting successful CAR T-cell expansion and persistence.

Botulinum Toxin A Ameliorates Neuroinflammation in the MPTP and 6-OHDA-Induced Parkinson's Disease Models.

Recently, increasing evidence suggests that neuroinflammation may be a critical factor in the development of Parkinson's disease (PD) in addition to the ratio of acetylcholine/dopamine because dopaminergic neurons are particularly vulnerable to inflammatory attack. In this study, we investigated whether botulinum neurotoxin A (BoNT-A) was effective for the treatment of PD through its anti-neuroinflammatory effects and the modulation of acetylcholine and dopamine release. We found that BoNT-A ameliorated MPTP and 6-OHDA-induced PD progression, reduced acetylcholine release, levels of IL-1β, IL-6 and TNF-α as well as GFAP expression, but enhanced dopamine release and tyrosine hydroxylase expression.

Systems analysis of dynamic transcription factor activity identifies targets for treatment in Olaparib resistant cancer cells.

The development of resistance to targeted therapeutics is a challenging issue for the treatment of cancer. Cancers that have mutations in BRCA, a DNA repair protein, have been treated with poly(ADP-ribose) polymerase (PARP) inhibitors, which target a second DNA repair mechanism with the aim of inducing synthetic lethality. While these inhibitors have shown promise clinically, the development of resistance can limit their effectiveness as a therapy.

Assembly of Bak homodimers into higher order homooligomers in the mitochondrial apoptotic pore.

In mitochondrial apoptosis, Bak is activated by death signals to form pores of unknown structure on the mitochondrial outer membrane via homooligomerization. Cytochrome c and other apoptotic factors are released from the intermembrane space through these pores, initiating downstream apoptosis events. Using chemical crosslinking and double electron electron resonance (DEER)-derived distance measurements between specific structural elements in Bak, here we clarify how the Bak pore is assembled.

Dynamic transcription factor activity networks in response to independently altered mechanical and adhesive microenvironmental cues.

Multiple aspects of the local extracellular environment profoundly affect cell phenotype and function. Physical and chemical cues in the environment trigger intracellular signaling cascades that ultimately activate transcription factors (TFs) - powerful regulators of the cell phenotype. TRACER (TRanscriptional Activity CEll aRrays) was employed for large-scale, dynamic quantification of TF activity in human fibroblasts cultured on hydrogels with a controlled elastic modulus and integrin ligand density.

Localized lentivirus delivery via peptide interactions.

Gene delivery from biomaterial scaffolds has been employed to induce the expression of tissue inductive factors for applications in regenerative medicine. The delivery of viral vectors has been described as reflecting a balance between vector retention and release. Herein, we investigated the design of hydrogels in order to retain the vector at the material in order to enhance transgene expression.

Betacellulin ameliorates hyperglycemia in obese diabetic db/db mice.

Unlabelled: We found that administration of a recombinant adenovirus (rAd) expressing betacellulin (BTC) into obese diabetic db/db mice ameliorated hyperglycemia. Exogenous glucose clearance was significantly improved, and serum insulin levels were significantly higher in rAd-BTC-treated mice than rAd-β-gal-treated control mice. rAd-BTC treatment increased insulin/bromodeoxyuridine double-positive cells in the islets, and islets from rAd-BTC-treated mice exhibited a significant increase in the level of G1-S phase-related cyclins as compared with control mice.

Dynamic transcription factor activity profiles reveal key regulatory interactions during megakaryocytic and erythroid differentiation.

The directed differentiation toward erythroid (E) or megakaryocytic (MK) lineages by the MK-E progenitor (MEP) could enhance the ex vivo generation of red blood cells and platelets for therapeutic transfusions. The lineage choice at the MEP bifurcation is controlled in large part by activity within the intracellular signal transduction network, the output of which determines the activity of transcription factors (TFs) and ultimately gene expression. Although many TFs have been implicated, E or MK differentiation is a complex process requiring multiple days, and the dynamics of TF activities during commitment and terminal maturation are relatively unexplored.

Differential Expression of Gene Profiles in MRGX-treated Lung Cancer.

Objectives: Modified regular ginseng extract (MRGX) has stronger anti-cancer activity-possessing gensenoside profiles.

Methods: To investigate changes in gene expression in the MRGX-treated lung cancer cells (A549), we examined genomic data with cDNA microarray results. After completing the gene-ontology-based analysis, we grouped the genes into up-and down-regulated profiles and into ontology-related regulated genes and proteins through their interaction network.

Dynamic transcription factor networks in epithelial-mesenchymal transition in breast cancer models.

The epithelial-mesenchymal transition (EMT) is a complex change in cell differentiation that allows breast carcinoma cells to acquire invasive properties. EMT involves a cascade of regulatory changes that destabilize the epithelial phenotype and allow mesenchymal features to manifest. As transcription factors (TFs) are upstream effectors of the genome-wide expression changes that result in phenotypic change, understanding the sequential changes in TF activity during EMT provides rich information on the mechanism of this process.

Sustained, localized transgene expression mediated from lentivirus-loaded biodegradable polyester elastomers.

The study of biomaterials for gene delivery in tissue engineering and regenerative medicine is a growing area, necessitating the investigation of new biomaterials and gene delivery vectors. Poly(1,8-octanediol citrate) (POC) and poly(glycerol-sebacate) (PGS) are biodegradable, biocompatible elastomers that have tunable mechanical properties, surface characteristics, and degradation rate. The objective of this work was to investigate whether POC and PGS would support the immobilization and release of lentivirus to allow sustained and localized transgene expression.

Dynamic transcription factor activity profiling in 2D and 3D cell cultures.

Live-cell assays to measure cellular function performed within 3D cultures have the potential to elucidate the underlying processes behind disease progression and tissue formation. Cells cultured in 3D interact and remodel their microenvironment and can develop into complex structures. We have developed a transcription factor (TF) activity array that uses bioluminescence imaging (BLI) of lentiviral delivered luminescent reporter constructs that allows for the non-invasive imaging of TF activity in both 2D and 3D culture.

Hydrogel macroporosity and the prolongation of transgene expression and the enhancement of angiogenesis.

The utility of hydrogels for regenerative medicine can be improved through localized gene delivery to enhance their bioactivity. However, current systems typically lead to low-level transgene expression located in host tissue surrounding the implant. Herein, we investigated the inclusion of macropores into hydrogels to facilitate cell ingrowth and enhance gene delivery within the macropores in vivo.

The impact of adhesion peptides within hydrogels on the phenotype and signaling of normal and cancerous mammary epithelial cells.

The microenviroment contributes to directing mammary epithelial cell (MEC) development and the progression of breast cancer. Three-dimensional culture models have been used to support formation of structures that display varying degrees of disorganization that parallel the degree of cancer. Synthetic hydrogels were employed to investigate the mechanisms by which specific adhesion signals in the microenvironment directed development.

Multifunctional, multichannel bridges that deliver neurotrophin encoding lentivirus for regeneration following spinal cord injury.

Therapeutic strategies following spinal cord injury must address the multiple barriers that limit regeneration. Multiple channel bridges have been developed that stabilize the injury following implantation and provide physical guidance for regenerating axons. These bridges have now been employed as a vehicle for localized delivery of lentivirus.

Fibrin hydrogels for lentiviral gene delivery in vitro and in vivo.

Gene delivery from hydrogels represents a versatile approach for localized expression of tissue inductive factors that can promote cellular processes that lead to regeneration. Lentiviral gene therapy vectors were entrapped within fibrin hydrogels, either alone or complexed with hydroxylapatite (HA) nanoparticles. The inclusion of HA into the hydrogel led to the formation of small aggregates distributed throughout the hydrogel, with no obvious alteration of the pore structure outside the aggregates.

Betacellulin-induced beta cell proliferation and regeneration is mediated by activation of ErbB-1 and ErbB-2 receptors.

Background: Betacellulin (BTC), a member of the epidermal growth factor family, is known to play an important role in regulating growth and differentiation of pancreatic beta cells. Growth-promoting actions of BTC are mediated by epidermal growth factor receptors (ErbBs), namely ErbB-1, ErbB-2, ErbB-3 and ErbB-4; however, the exact mechanism for beta cell proliferation has not been elucidated. Therefore, we investigated which ErbBs are involved and some molecular mechanisms by which BTC regulates beta cell proliferation.

Phosphatidylserine immobilization of lentivirus for localized gene transfer.

Localized and efficient gene transfer can be promoted by exploiting the interaction between the vector and biomaterial. Regulation of the vector-material interaction was investigated by capitalizing on the binding between lentivirus and phosphatidylserine (PS), a component of the plasma membrane. PS was incorporated into microspheres composed of the copolymers of lactide and glycolide (PLG) using an emulsion process.

Lentivirus immobilization to nanoparticles for enhanced and localized delivery from hydrogels.

Hydrogels can provide a controllable cell microenvironment for numerous applications in regenerative medicine, and delivery of gene therapy vectors can be employed to enhance their bioactivity. We investigated the delivery of lentiviral vectors from hydrogels, and employed the immobilization of lentivirus to hydroxylapatite (HA) nanoparticles as a means to retain and stabilize vectors within hydrogels, and thereby increase delivery efficiency. Entrapment of the vector alone within the hydrogel maintained the activity of the virus more effectively compared to the absence of a hydrogel, and release was slowed with an increasing solid content of the hydrogel.

Amelioration of hyperglycemia by intestinal overexpression of glucagon-like peptide-1 in mice.

To investigate whether the local production of glucagon-like peptide-1 (GLP-1) in the intestine can differentiate intestinal stem/progenitor cells into insulin-producing cells, we intra-intestinally injected a recombinant adenovirus expressing GLP-1 (rAd-GLP-1) into diabetic mice. There were no significant differences in body weight or food intake between rAd-GLP-1- and rAd-betaGAL-treated control mice. rAd-GLP-1-treated mice showed intestinal insulin mRNA expression, insulin- and glucagon-positive cells in the intestine, and significantly increased serum insulin, but not glucagon.

Lentivirus delivery by adsorption to tissue engineering scaffolds.

Biomaterial scaffolds capable of localized gene delivery are being investigated for numerous regenerative medicine applications and as model systems for fundamental studies of tissue formation. In this manuscript, we investigate the delivery of lentivirus from a tissue engineering scaffold using a surface immobilization strategy. Poly(lactide-co-glycolide) (PLG) was employed as the biomaterial for delivery, which has been widely used for a number of tissue engineering applications.

Role of nitric oxide in the pathogenesis of encephalomyocarditis virus-induced diabetes in mice.

The D variant of encephalomyocarditis virus (EMC-D virus) causes diabetes in mice by destroying pancreatic beta cells. In mice infected with a low dose of EMC-D virus, macrophages play an important role in beta-cell destruction by producing soluble mediators such as interleukin-1beta (IL-1beta), tumor necrosis factor alpha (TNF-alpha), and nitric oxide (NO). To investigate the role of NO and inducible NO synthase (iNOS) in the development of diabetes in EMC-D virus-infected mice, we infected iNOS-deficient DBA/2 mice with EMC-D virus (2 x 10(2) PFU/mouse).

Remission of Diabetes by β-Cell Regeneration in Diabetic Mice Treated With a Recombinant Adenovirus Expressing Betacellulin.

Type 1 diabetes results from destruction of the majority of the pancreatic β cells by β cell-specific autoimmune responses; therefore, expansion of the β-cell mass in vivo Is a possible approach to its treatment. Betacellulin (BTC) is known to promote β-cell growth and differentiation. We investigated whether transient, constitutive expression, and secretion of BTC would regenerate sufficient numbers of pancreatic β cells to restore normoglycemia in diabetic animals.

Remission of diabetes by beta-cell regeneration in diabetic mice treated with a recombinant adenovirus expressing betacellulin.

Type 1 diabetes results from destruction of the majority of the pancreatic beta cells by beta cell-specific autoimmune responses; therefore, expansion of the beta-cell mass in vivo is a possible approach to its treatment. Betacellulin (BTC) is known to promote beta-cell growth and differentiation. We investigated whether transient, constitutive expression, and secretion of BTC would regenerate sufficient numbers of pancreatic beta cells to restore normoglycemia in diabetic animals.