This proposal is targeted at the development of a novel research platform for analyzing immune cell gene expression as a result of HIV infection that is based on the on-chip real-time amplification detection (OCRAD) method that came out of the Phase I program. Current methods for performing gene expression analysis require lengthy and complex protocols, involving separate RNA amplification, hybridization, washing and label read-out steps. OCRAD employs a refractometric integrated optic chip (IOC) sensor to perform multiplex on-chip nucleic acid amplification. This combination of on-chip amplification and real-time detection will create a platform with the features necessary to make a significant advancement in the ability of researchers to perform gene expression measurements in a more rapid, simple and less expensive format. Gene expression analysis is an important tool for understanding a cell's differentiation state and may be useful in a clinical setting as a diagnostic or predictive tool. CD8 T cells are critical for the control of many viral infections and are the target of vaccine efforts for HIV. However, during HIV infection immune control by CD8 T cells eventually fails leading to AIDS. Considerable evidence suggests that CD8 T cells in HIV patients undergo an inappropriate differentiation process leading to impaired function, although the molecular signature of this process in humans is not well defined. Using a small gene set profile based on previous gene expression studies using mouse models of viral infection, OCRAD will be applied to analyzing gene expression in CD8 T cell subsets from HIV- healthy patients, HIV+ patients with suppressed viral load and HIV+ patients with viremia. It is believed that this analysis should provide a better understanding of the molecular profile of CD8 T cells during HIV infection. This may ultimately lead to improved diagnostic and/or predictive clinical tools and may uncover important biological pathways that can be used as targets for therapeutic intervention. ? ?
