Eddie Polanco 2

The second week of the program was really good. I have gotten more experience with the techniques and more involved with my project. There are two main projects that I am working on one involving making a vaccine for the various serotypes of Dengue and the other one working on developing a small molecule cancer therapy based on the T-cell surface protein programmed death 1 (PD-1). PD-1 is a negative regulator of T-cells inducing apoptosis when it binds to its substrate PD-L1/-L2.  PD-L1/-L2 is a negative regulator of the immune system, expressed on healthy cells in order to prevent autoimmune diseases.

The primary technique used in this lab is called phage display. Phage display works by expressing proteins on bacteriophage and then through a series of selections you force evolution to favor proteins with specific properties such as high binding affinity to a target substrate. Using this technique we are trying to find a PD-1 variant that binds with high affinity to PD-L1 and PD-L2. We believe that this will be an effective cancer therapy by binding to PD-L1 before wild type variants of PD-1 are able to bind to the proteins expressed on the T-cells.

 

Picture Captions:

Picture 1: Phage display is a cycle through which a protein can be engineered through directed evolution to have specific properties. (A) Input phage expressing protein of interest are selected for binding affinity against a target of interest in a process called biopanning. (B) Unbound phage are washed away before the bound phage are eluted from the target. (C) The bound phage are eluted off the target and (D) then allowed to infect e. coli in order to amplify their genome. (E) The amplified phage are then used as the input phage for the next round of selection. This process is typically repeated 3-5 times until all phage variants have similar binding properties.

 

Picture 2: Our plan for a monoclonal enzyme linked immunosorbant binding assay (ELISA) in order to obtain points to plot a binding curve. The assay is designed to measure binding to PD-L1, PD-L2, and a control experiment in order to see how well various PD-1 variants bind to each substrate.