And so it began. Week 2 of SC-ACE. First week in my new lab. We started off with an absolutely brain-frying lab meeting from 9 am to 2 pm. I got my introduction to Acute Myeloid Leukemia (AML) and the different research projects going on in this lab. It was my first experience of a lab meeting, and I thought it was very special. Everybody gave a presentation of their work the week before, and the other people in the lab would give suggestions or critique.
In short, AML is a fast moving blood and bone marrow cancer. In this disease, immature white blood cells pile up in the bone marrow and crowd out the healthy cells. They eat up all the body’s nutrients but provide nothing in return. A common genetic mutation in AML is called FLT3-ITD. Found in about 30% of patients, FLT3 is linked to poor patient outcomes. The lab focuses heavily on genetic therapies that can target this mutation. One technique they are currently working on is T-cell engineering, which allows researchers to design immune cells to specifically target AML cells with FLT3 mutation.
The picture on the right shows engineered T cells attacking and destroying a tumor.
The second day was all about my project briefing. It was another heap of information but super exciting. Our goal is to create retroviral plasmids that express the mutation. These plasmids will eventually be inserted into a cell and used to study how engineered T cells recognize AML cells with FLT3 mutation.
Here is a high-level overview of the process:
We start by getting the DNA. DNA is extracted from patient blood. PCR, a technique used to make many copies of a DNA segment is used to amplify a 400 base pair (think about it as length) piece of FLT3 gene.
Next, we will clone the DNA.
The product from the previous step will inserted into a TOPO vector, which is basically a small circular DNA designed to be able to store genes long term and make them easy to move around. Then the FLT3 sequence will be cut out of the TOPO vector (what we just inserted) using restriction enzymes that act as molecular scissors. Then what we cut out will be inserted into another plasmid called pMIG-II. We insert it into pMIG-II because pMIG-II has a protein that will make it easy to track in future steps.
The plasmid is put into E. coli bacteria through a process called transformation. Think of the bacteria as tiny factories copying the plasmid DNA. These factories are then selected, verified to make sure we have the correct sequence, then grown into larger cultures. After we have enough, we can extract the DNA, purify it and insert it into our AML cells for testing.
This workflow is intense. There are enzymes, bacterial transformations, plasmid extractions, restriction enzymes, sequencing, virus generation. The briefing also highlighted why TCR-T cells are exciting. Unlike CAR T-cells, which target proteins on the cell surface, TCR-T cells can detect antigens presented on MHC molecules, which lets them selectively go after cancer cells while sparing healthy blood cells.
My brain is overloaded. I’ll be splitting this week into two blog posts so it’s not as boring to read. Part 2 will cover my first days actually running experiments in the lab.
