The Study Of Secretory Type TRAIL Gene Modificaed Human Adipose Derived Stem Cells In Targeting Theraphy To Glioma

BackgroundGliomas are the most frequent primary malignant tumors in the central nervoussystem and have a variety of histologic types as well as a different degree of malignancy.Of adult tumors, glioblastoma multiform (GBM) represents50%of all gliomas and isattributed as the most aggressive form of the disease. Surgical resection remains quitedifficult due to the diffuse nature of tumor cells and local invasion/infiltration. Thecurrent life expectancy of patients with GBM is seen to be14.6months even throughcurrent methods of multimodal therapy, with total resection and a combination of chemo-and radiotherapy. Therefore, how to improve the survival and quality of life in GBMpatients have been challenges facing by medical profession.In early1990s, the upsurge of gene therapy has offered hopes to the treatment ofmalignant glioma, and HSV-TK GCV has been one of representatives. Although theHSV-TK GCV system has achieved good results in animal models, the results from clinicaltrials was disappointed, which might be due to instability of viral vectors, short-termexpression of target gene, antilogous immunity, lack of tumor targeting and incapability totrack infiltrative tumor cells. Many current studies have shown that bone marrowmesenchymal stem cells, human umbilical cord blood stem cells, neural stem cells have thecapability of tumor tracking. The tracking capability is associated with secretion ofcytokines and chemokines, and stem cell can pass through the damaged blood-brain barrier,gathered around the tumor of high concentrations. This feature has been proved in variousin vivo and in vitro experiments. This feature is currently considered that mesenchymalstem cells can do as a carrier of enzyme/prodrug gene in combinated targeting therapy fortumor treatment, which is one of ideal strategies. Adipose-derived stem cells (ADSCs) alsohave been proved having the capability of tumor tracking.In vitro experiment, the TRAIL induce the apoptosis of the tumor in glioma, neuroblastoma, cervix uteri cancer, non small cell lung cancer, renal cell carcinoma, livercancer, thyroid cancer, melanoma. TRAIL has special lethal effect on tumor cells, such asHela cervix uteri cancer cells and A549lung cancer cells, human liver cancer cells andhuman breast cancer cells, progranulocyte. In preliminary pharmacodynamic study showedthat TRAIL can significantly inhibit the growth of human hepatocellular carcinoma cells inmice, but non-toxic side effects on the normal mice. A tumor-burdened animal applicationrestructuring TRAIL of protein inhibit tumor cell growth, even make tumor regression, andwithout apparent damage to the host. Many experiments show that the TRAIL combinedwith chemotherapy drugs, have good antitumor effect. So TRAIL gene, as an ideal goalgene of straightforward tumor suppression effect, have the prospect for further research.Objective: To study TRAIL modified ADSCs’ effect on targeted therapy to gliomacombined the characters of ADSCs target tumor migration with TRAIL selectively cancercells kill.Methods:1. To isolate ADSCs from healthy adult subsutaneous adipose tissue sampelsusing collagenase digestion followed by centrifugal separation, and certify the ADSCsdifferentiative potential.2. To investigate whether ADSCs have migration ability toward glioma cells in vivo and invitro. GFP labeled ADSCs were cocultured with U87MG glioma cells, and one week afterU87MG glioma mouse model was established, LacZ labeled ADSCs was intravenouslyinjected.3. Secreted TRAIL (sTRAIL) gene fragments was obtained from pROF-sTrail plasmid, andreconstructed it into rAAV-GFP plasmid to generate high-titer AAV-sTRAIL virus particles,then transfected ADSCs with AAV-sTRAIL virus, to detect the expression.4. To establishe a nude mouse glioma model and intravenously and intratumrally injectesTRAIL modified ADSCs and observe the model’s lifetime.Results:1. ADSCs were typically expanded in monolayer culture on standard tissueculture plastics. When cultured with condition media, they could differentiate into matureadiopocytes, osteoblasts and chondrocytes.2. GFP positive ADSCs accumulated in the U87MG glioma cell clones, whereas no GFP positive cells were observed in the absence of U87MG glioma cells. a large amount of blueLacZ was distributed throughout the brain tumor tissue. Furthmore, the level of LacZpositive ADSCs in glioma (117.52±36.13/section) was much higher than in thecontralateral normal brain tissue (34.82±7.14/section) and other organs (heart:0/section;lungs:28.54±7.56/section: liver:18.63±5.27/section: spleen:7.67±5.73/section:.kidney:3g.78±12.g4/section)(p <0.001). We also observed that LacZ labeled ADSCsinoculated into the contralateral hemisphere migrated away from the initial injection sitetowards the glioma cells along the white matter fibers.3. High-titer AAV-sTRAIL virus particles was generated. When transfected ADSCs withAAV-sTRAIL virus, we found that AAV-sTRAIL virus did not show toxicity to ADSCs.4. Of all tumor-burden mice,36%of mice in ADSCs-sTRAIL treatment group survivedmore than45days. We also found that co-culture of ADSCs-sTRAIL with U251gliomacells improved the U251glioma cells apoptosis.Conclusion: Research data indicate that a cellular carrier like ADSC is effective at stalbyexpressing and delivering TRAIL virus towards brain tumor cells, thus opening noveltherapeutic opportunities for still incurable cancers.