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Adam Wilson

Adam Wilson

Adam Wilson, a junior at Johns Hopkins University studying mathematics, biophysics, and philosophy, investigated a new immortalization method for pediatric low-grade glioma cell lines.

Pediatric low-grade gliomas (pLGGs) are a type of brain cancer that, along with its treatments, are life-threatening, and can cause lifelong disability. pLGGs, for unknown reasons, are difficult to grow in vitro. A challenge has been to find a way to grow pLGG cells to enable further research of pediatric low-grade gliomas. While a method using SV40 has been proposed to immortalize pLGG cells, it is not widespread and requires further investigation. This summer has been one of the most transformative of my life. Whether it be learning from some of the most brilliant scientists in the world, kayaking down the Charles River, laughing with my BSRP cohort friends, or embracing failure and pivoting to new scientific opportunities, Broad has been invaluable to my development as a scientist, leader, and lifelong learner.Moreover, the cell cycle arrest observed in pLGG cells when extracted from the body is unlikely to arise from the telomeres of the cell not meeting a threshold when in culture—the problem SV40 immortalized pLGG cells attempts to solve. Rather, we suggest it is the result of the tumor-suppressor proteins p53 and RB1 causing cell cycle arrest. When p53 is activated, RB1 forms complexes with E2F transcription factors that then downregulates many cell cycle genes. Thus, the cell cycle arrest brought by p53 and RB1 may be the mechanism preventing pLGG growth in vitro.

We hypothesize a mechanism for bypassing the cell cycle arrest checkpoint via doxycycline-inducible E6/E7 immortalization. E6/E7 genes produce oncoproteins which in turn inhibit the tumor-suppressor proteins p53 and RB1 found in pLGG cells, thereby upregulating the large number of cell cycle genes previously downregulated by p53 and RB1. Upon creation of our E6/E7 pLGG cell model via DNA cloning and viral transduction, we aim to grow a large quantity of cells, remove doxycycline—thus making the E6/E7 genes no longer expressed—and proceed to conduct experiments on the cells. Because we suppose p53/RB1 induced cell cycle arrest to be the cause of low pLGG growth in vitro, our E6/E7 immortalized pLGG cells should be more comparable to the originals than previously proposed methods. We then aim to conduct CRISPR screens using our cell model to attempt to discover the genes responsible for pLGG growth in the brain.

 

Project: Immortalizing pediatric low-grade glioma cell lines via E6/E7 genes

Mentors: Sean Misek and Simona Dalin, Beroukhim Lab