Genetic Engineering of T Cells Resistant to Oxidative Stress

Provided is a method to genetically engineer human lymphocytes, to enable expression of an antioxidant enzyme, which can render the cells resistant to immune suppression mediated by reactive oxygen species (ROS).

Adoptive immunotherapy of patients with advanced cancer or viral infections has so far met with positive, but limited success. This limited therapeutic effect of adoptively transferred lymphocytes may be attributable in part to elimination of Natural Killer cell and T cell activities by oxidative stress. Systemic administration of drugs that strengthen the patient's resistance to ROS is limited by several factors: Drugs have difficulty penetrating into tumors, can cause toxicity due to the relatively high dosages needed to reach the necessary concentration in the tumor or virus infected tissue, and most drugs have a limited half-life.

Dr. Nishimura (Medical University of South Carolina) and his collaborators at the Karolinksa Institutet have developed a method to overcome these problems through "arming" the patients lympocytes (the so called T-cells and NK-cells) through genetic modification with an antioxidant enzyme (catalase) which can neutralize ROS. A patient's own lymphocytes are transduced with a viral construct coding for catalase. This procedure is done ex vivo in a test tube. The modified lymphocytes can then be transferred back to the patient.

Because the modified lymphocytes are now more resistant to ROS, they are protected from cell death and have an enhanced capacity to recognize virus-expressing target cells. They will be able to actively migrate into tumor tissues and have the potential of replicating and remaining in the host for prolonged periods of time.

Novel Acid Ceramidase Modifying Compounds to Treat Cancer

Sphingolipid metabolism is recognized as a key factor in tumor cell survival and metastasis. Ceramide is a "tumor suppressor sphingolipid" involved in regulation of cell growth, differentiation, senescence and programmed cell death. Drs. Norris and Hannun, and their colleagues at the Medical University of South Carolina, have developed and tested a portfolio of novel small molecule therapeutics that target ceramide metabolism and result in cancer cell apoptosis.

The novel small molecule therapeutics were rationally designed and proven to inhibit enzymes (called acid ceramidase and sphingosine kinase) that create resistance to conventional cancer therapies. The compounds sensitize cancer cells to undergo programmed cell death, therefore they can be administered either as a stand alone treatment or in combination with "standard of care" chemotherapy or radiation therapy. Chemo-additive therapy will permit the oncologist to improve morbidity and clinical outcomes at lower, less toxic, drug or radiation doses.

Substantial preclinical data is available and the researchers continue to actively test these compounds.

Cancer-specific adenovirus expressing an inducible shRNA

The present invention slectivly target elongation factor 2 (EF2) in cancer cells by generating an adenovirus with two novel features.  The virus can only replicate in cancer cells, and it expresses an inducible shRNA against EF2.  Preliminary studies in vitro show that shRNA against EF2 are promising.  The resulting invention would likely result in a novel approach to targeting specific cancers in the human body.

Bacterial Toxin Delivery Vehicle

This technology uses a plasmid carrying a lethal-but inactivated-marker and a phage lytic replicon. The plasmid is maintained in a lysogenic state via a temperature sensitive repressor mutation. Induction by temperature shift will result in plasmid replication and packaging into phage virions. Virions carrying the lethal toxin gene are used to deliver the antimicrobial agent to a pathogen host that cannot inactivate the toxin.

Production Increase of natural and Hybrid Natural Products Produced by a Fermentable organism such as Bacteria or Fungi through Cofermentation with a Selected Second Natural Organism, such as a Bacterium or a Fungus

Investigators disclose a method of increasing the fermentation yield of certain compounds from natural or genetically modified bacteria or fungi without requiring additional energy inputs. The ideal embodiment of the invention may be when the desired product is an antibiotic.

An Autologous Upregulation Mechanism Allowing Optimized Cell Type-Specific and Regulated Gene Expression in Eukaryotic Cells

Prostate cancer is now the second leading cause of cancer death among men in the United States and currently available treatment options are often ineffective. Thus, there is a great need for new treatment strategies to improve these outcomes. Here, we describe an Ad vector that delivers an alternative transgene expression regulation strategy, one that induces high levels of FasL expression in prostate cancer cells without any expression in the cells of other origins. This strategy involves the incorporation of the TRE upstream of the ARR2PB promoter to enhance its activity with Tet-regulation. The expressions of both FasL-GFP and tTA were placed under the control of these TRE-ARR2PB promoters, so that in the cells of prostate origin, a positive feedback loop is generated. This design greatly amplified tTA and FasL-GFP expression in a prostate-specific manner, with virtually no expression in non-prostate cells.

The implications of this new regulation system, however, go beyond that of prostate cancer gene therapy as this complex Ad vector could be modified to incorporate any tissue-specific promoter in the place of ARR2PB to deliver high levels of expression of any gene of interest in a highly specific manner.

Recombinant fusion proteins consisting of complement inhibitors and fragments of soluble P and E-selectin glycoprotein ligand-1 that taret to cell adhesion molecules

Disclosed are methods to produce novel fusion proteins, which rely on different targeting mechanisms, to bind to E-selectin and P-selectin with higher affinity and specificity then any prior construct. Historically the proteins were difficult to express recombinantly, however inventors at MUSC found that coexpression of enzymes allows them to produce the functional protein in reasonable quantities.

Phage-Based Therapeutic System

The disclosed invention relates to the discovery, identification and characterization of toxic agents with are lethal to drug-resistant pathogens and drug-resistant diseased cells (such as cancerous cells). Further, the invention provides a novel system by which multiple targets may be simultaneously targeted to cause the death of a diseased cell.

This application relates to multi-ribozymes and their use to target RNA in a tissue-specific or target-specific manner, for the treatment of cancers and bacterial, viral and parasitic infections.

DNA and Protein sequences of Human Alkaline Cermidase Gene: an important target for chemotherapy

This invention provides the DNA sequence of a human alkaline ceramidase gene and the protein sequence of the gene product. The human alkaline ceramidase catalyzes hydrolysis of ceramide to yield free fatty acid and sphingosine which is phosphorylated by sphingosine kinase to form sphingosine-1-phosphate. Ceramide induces the programmed cell death of many cancer cells, whereas sphingosine-1-phosphate triggers cell proliferation. Therefore, the alkaline ceramidase is a key enzyme in regulation of intracellular levels of ceramide and sphingosine-1-phosphate. Inhibitors of this ceramidase will cause an increase in levels of ceramide and a decrease in levels of sphingosine-1-phosphate, thus leading to cell death. Therefore, these inhibitors will be important candidates as chemotherapeutic agents.

Moreover, this ceramidase has the reverse hydrolysis activity of catalyzing ceramide formation from sphingosine and a fatty acid. This reverse activity can compensate for the loss of the activity of a CoA-dependent ceramide synthase. In other words, this enzyme is also important for cell survival when cells are stressed. Therefore, this alkaline ceramidase has applications in other diseases in addition to cancer.

Also included is an assay for identifying inhibitors of this enzyme.

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