(557e) Use Of Trail To Deliver An Apoptotic Signal To Rolling Cancer Cells Under Shear

Tumor metastasis consists of a series of discrete steps that move the tumor cells from the primary site to a distal location. The tumor cells must invade the surrounding tissue and enter either the bloodstream or the lymphatic system, survive the circulation, adhesively interact with the vasculature that facilitates tethering and rolling and eventually arrest. Following arrest, the tumor cells extravasate into the distal site and grow into a secondary tumor. Although surgery, radiation therapy and chemotherapy are effective in controlling many cancers at the primary site, the development of metastatic cancer signals a poor prognosis. The survival rate for patients with such metastatic cancers is dramatically reduced, with most deaths being associated with the formation of secondary tumors. This makes metastatic tumors an important factor in determining the final outcome of the diseases. Tumor Necrosis Factor (TNF) Related Apoptosis Ligand (TRAIL) or Apo2L holds promise as a candidate for cancer specific therapeutic. TRAIL is known to bind five known receptors: two of which are death receptors (DR4 and DR5) that signal apoptosis while the other three are decoy receptors (DcR1, DcR2 and DcR3) that do not. Unlike other members of TNF family, such as Fas-ligand, upon binding to the death receptors TRAIL induces a death signal by apoptosis via the caspase pathway. We use this phenomenon to deliver a receptor-mediated apoptosis signal to leukemic cells adhesively rolling along a selectin-bearing surface. Our results show that cancer cells undergo selectin mediated rolling in parallel plate flow chambers, glass capillaries and polymeric microtubes. We have determined that the rolling velocities can be controlled to achieve optimal contact time with the surface and prevent saturation of the surface, by adjusting the surface density of the selectin molecule and the applied shear stress. We use the Annexin-V apoptosis assay to quantify cell death and identify the stage of cell death using a FACS Calibur Flowcytometer and cell proliferation assay to quantify the viability of cells after rolling experiments. When using soluble TRAIL we have found that there is a difference in sensitivity towards TRAIL between cell lines. The same effect was verified with immobilized TRAIL. This difference is believed to be due to differences in the relative expression of death receptors and decoy receptors. We have also verified that soluble TRAIL has no significant effect on the viability of CD34+ hematopoietic stem cells and other mononuclear blood cells, while much lower concentrations of TRAIL kill most cancer cells. The rolling experiments show a dramatic difference in the number of viable cells when cells are rolled on the combined TRAIL and selectin surface and the control surface of selectin only for short durations of time. Thus we have demonstrated that it is possible to deliver an apoptotic signal to rolling cancer cells. Current efforts are geared towards further improving the efficiency of the device and optimizing the device for long-term in-vivo usage.