I. Our first innovation is selecting the most appropriate sample: It is well known and widely accepted in the scientific community that the primary tumor consists of the stroma [fibroblast, monocytes, lymphocytes, vessels, etc.] and the malignant cells which [are in-homogenous] since they are composed from different subgroups and subclones with different features and abilities. Only very few populations will develop metastatic ability which will allow them to invade the surrounding tissue and pass into the circulation and perform the epithelial to mesenchymal transition (EMT). These are known as the circulating tumor cells [CTC’s]. A large proportion of the CTC’s are actually cancer stem cells. These cells have all the necessary information and ability for micro colonization and micrometastases and will later develop into potent micrometastases. For this reason RGCC has selected blood samples as the most appropriate form for analysis since it includes the cancer cells with the relevant information for both calculating the risk and for the potent metastases.
II. Innovation in finding and isolating the CTC’s: RGCC uses powerful sorters and flow cytometers as well as negative selection based interrogation. We are able to actually isolate the relevant CTC’s and not enrich them. Hence, we manage to have a pure sample of CTC’s and simultaneously harvest all the group of CTC’s from a single blood sample.
III. Appropriate expansion of the CTC’s: The CTC’s will expand as the cancer stem cell like cell and then enter in exponential phase of growth which will generate a respectful number of circulating stem cells [CSC’s] in a very short period of time. At the same time we manage to keep intact the both genotype and phenotype of the cells and avoid any changes to the primary [CSC]. Therefore, after the final expansion we have maintained the identical genotype.
IV. Gene-expression profile of the CSC’s: The expanded cells will be analyzed for one expression profile in a hall genome micro-array analysis. Hence, we now have all the information concerning epigenetic screening of CSC’s. This profile will indicate to us which therapy the CSC’s are sensitive or resistant to.
V. Verification of chemosensitivity/chemoresistance in viability assays: The information obtained from the gene expression analysis will be validated in a micro culture where the effect of each indicated substance will be tested. These assays overcome the problem of linearity between gene expression and protein expression levels.