CRISPR-Cas9 Mediated PIK3CA Modification as A Novel Treatment Modality in Cancer. Doctoral Thesis Study

Background: CRISPR-Cas9 is the leading technology among all genome modification methods. It’s been utilized in cancer etiology, tumor pathogenesis, tumor microenvironment, drug efficiency studies, and developing new strategies via novel biomarker discoveries. PI3K/Akt/mTOR signaling pathways has a great impact on cancer cell growth and proliferation as well as cancer cells’ escape from the immune system and apoptosis. Thus, it’s especially promising for targeted anti-cancer therapies. Methods: In this study, CRISPR-Cas9 induced knockdown was performed on PIK3CA which is located on this important cellular pathway, and the impacts of this gene modification on both the behavior of cancer cells and the whole PI3K pathway were investigated using Cukurova University AGENTEM and Cukurova Technopolis InfoGenom infrastructure. Experiments were carried out on MCF-7 cell-line, and electroporation was used for all transfection steps. CRISPR-Cas9 mediated modification of PIK3CA was shown using Sanger and next-generation sequencing methods, and its its effects were observed through inverted microscope, flow cytometry, and real-time PCR expression assay. Results: This study indicates knockdown of PIK3CA gene via CRISPR-Cas9 induces cell death and has the potential for new therapeutic approaches. Conclusions: The stand-alone PIK3CA inhibitor treatment for wider range of breast cancers together with CRISPR-Cas9 strategy as a novel combinatorial therapy should be considered according to our findings.