Akademik Veri Yönetim Sistemi
Dükel M., Pavlopoulou A., Meuwissen R. L. J., Ayar Kayali H.
TÜBİTAK Projesi, 2022 - 2025
Colorectal cancer is the third most common type of cancer and is second cause of mortality throughout America and Europe. One million new colorectal cancer cases occur each year worldwide and more than half of these cases die. Most forms of colorectal cancer are slowly grow and step occurred symptoms is usually advanced stage of disease. Only 40% of patients with colorectal cancer can be diagnosed early, and in much of late-diagnosed patients, cancerous cells migrated to other tissues. Therefore, understanding metastasis is a vital for cancer studies and researches. Cancer metastasis is a complex process with multi-steps. During metastasis, cancer cells improve their ability to invade to surrounded tissues, resulting in loss of cell-cell, cell matrix connectivity, and acquire consequently anoikis resistance. Anoikis is a specific type apoptosis that occurs in epithelial cells when they lost their connection with the basement membrane. Cancer cells acquire anoikis resistance to survive and migrate to other tissues. Several studies had been done for anoikis but the exact mechanism of this is not yet known. Crispr/Cas9 genome-wide sequencing is a widely used method in recent years to understand drug resistance and metastatic processes. 15,000-20000 genes are mutated using Crispr/Cas9 library containing thousands of sgRNAs for the genome wide sequencing, and the effects of the function loss for these genes during processes are investigated.
In this study, it is planned to investigate the genes that play a vital role in the anoikis resistance by Crispr/Cas9 genome wide sequencing in colon cancer cells. To the best of our knowledge, there is no study that aimed to test on anoikis resistance properties in colon cancer cells via Crispr/Cas9 genome wide sequencing.
In present study, first we are planning to test anoikis resistance properties and Crispr/Cas9 activity in CCD-18Co, CCD-18Co, SW620 and HCT-116 cell lines. Second, we will make viruses using the Crispr sgRNAs Library. Next, we aimed to infect the CCD-18Co, SW620 or HCT-116 with sgRNAs viruses. In addition, the cells will be subjected to the anoikis process and DNA isolation will be perform from all groups. Moreover, we will conduct 2 step PCR and samples will be sent for next generation sequencing. After bioinformatic analysis, we aimed to determine at least 3 genes that will be effective for anoikis resistance. We are planning to validate target genes for anoikis resistance in vitro. Moreover, we will test those genes affects for other metastatic characters (invasion, migration and colony formation). Finally, we are planning to find out if those genes are effective during tumor formation and growth in vivo.
For present work, we are planning to employ 1 doctoral and 1 master students. All students will work all experimental studies under the control of the executive, researchers and the consultant. One of the most important steps of Crispr/Cas9 genome sequencing is bioinformatical analysis of sequence data. Hence, two bioinformatics specialized in this field is included in the project. The consultant will ensure communication and organization between the project team and will take part in the project's publication.
Crispr genome sequencing is widely used today and studies can be published in Journals with high impact factor. This technique is widely used by many other countries for cancer studies. In particular, a great progress has been made in uncovering drug resistance mechanisms and contributes greatly to the new drug development or personalized cancer treatment process. If our study is supported, it will be contributed to better understanding of the anoikis resistance feature and in the light of these data that will help to the development of drugs to target metastatic tumors and to diagnose colon cancer at early stages.