Vivian Nguyen Rose Hills
Engineering Novel Cas9 PAM Targeting Variants for Therapeutic Use
Over the past decade, CRISPR-Cas9 has emerged as an unparalleled tool for single-gene disease therapy. Yet, the vast majority of the genome is currently untargetable by Cas9 due to its strict requirement for a specific protospacer adjacent motif (PAM). While there have been attempts to engineer Cas9 with broader PAM recognition, these variants exhibit significantly lower on-target specificity and higher off-target activity compared to wild type Streptococcus pyogenes Cas9. To expand Cas9’s targetable genomic space without decreased on-target activity and increased off target editing, this research aims to engineer novel SpCas9 variants capable of specifically recognizing non-NGG PAMs through directed evolution in a yeast selection system. I will focus on engineering a SpCas9 variant that can specifically recognize the NAC PAM to treat retinitis pigmentosa, a dominant genetic disorder that causes blindness. Ultimately, this project will help expand and diversify the therapeutic potential of CRISPR-Cas9.
Message To Sponsor
Thank you for your generous sponsorship! Your support has made it possible for me to immerse myself in this summer project and to continue developing my skills as an aspiring researcher. I am immensely grateful for this opportunity and I am so excited to start working on engineering therapeutically relevant Cas9 variants!