Most solid tumors exhibit limited responsiveness to immunotherapy. Patients with mismatch repair (MMR)-deficient (MMRd) colorectal cancer—characterized by absent or low expression of key DNA mismatch repair proteins such as MLH1 and MSH2—often respond to immune checkpoint blockade with complete tumor eradication. The Lieberman Lab therefore postulates that tumor-selective nanoparticle delivery of Cas9 mRNA and sgRNAs to knockout MMR genes in tumor cells could convert mismatch repair–proficient, immunotherapy-resistant solid tumors into MMRd, immunogenic tumors that could respond to immune checkpoint therapy. EpCAM is highly expressed on all epithelial cancer cells but not on normal, untransformed cells. My research project will seek to develop EpCAM-aptamer-conjugated lipid nanoparticles (LNPs) encapsulating Cas9 mRNA and sgRNAs targeting MMR genes to induce gene knockdown in vivo in epithelial cancer cells. My first goal will be to determine whether these EpCAM-aptamer-conjugated LNPs cause selective gene knockdown in vitro and in vivo. If successful, I will next evaluate the effect of knocking out MMR genes on DNA repair, microsatellite instability, chromosomal instability, and tumor growth in colorectal cancer cell lines in vitro. If these LNPs cause MMR deficiency in the tumor cell lines in vitro, I will administer them to mice implanted with MMR-proficient cancer cells to determine their effect on tumor growth, their impact on the properties of the tumor and tumor-infiltrating immune cells, and whether they augment the anti-tumor effectiveness of checkpoint inhibitor antibody therapy using anti-PD-1. I will apply techniques such as LNP transfection, flow cytometry, PCR and Western Blots for assessment of gene knockout efficiency, RiboGreen and DLS for evaluation of LNP quality, and tumor-infiltrating immune cell isolation. My project will help elucidate the mechanisms underlying why MMRd tumors respond to immune therapy and also lay the groundwork for developing a platform for tumor-selective gene editing.
Northeastern University
