Colorectal cancer on a dish: exploring the 3D-sphere culture of primary colorectal cancer cells from an Indonesian perspective

Abstract

Background : Colorectal Cancer (CRC) is one of the deadliest types of cancer and has emerged as one of Indonesia's most devastating diseases. The growing number of colorectal cancer cases is frequently undiagnosed until the disease has progressed to a metastatic stage. This issue has lasted for years, limiting therapy options and resulting in a bad prognosis for the majority of patients. Thus, the purpose of this work is to develop a CRC detection method for Indonesia and other low-middle income nations that integrates in vitro 3D culture, molecular analysis, and in silico analysis. Methods : Colorectal cancer biopsies were transported to the lab and underwent mechanical disaggregation and centrifuged at 300 x g for five minutes. Approximately 10,000 cells were seeded in each Nunc-Sphera 96-well plate (u-bottom) for the following 7 days in standard culture medium. The 3D-sphere was harvested and RNA was extracted afterwards. Molecular analysis was performed using qPCR and the Human Cancer Pathway Profiler. Protein interaction and pathway analysis were conducted using STRING and Reactome online tools. Results :  Following initial seeding, primary CRC 3D-spheres were grown for 14–16 days. Gene profiling and in silico analyses suggest that CDC20, AURKA, and ACLY are expressed at lower levels than the positive control in the 3D-sphere. These markers have been implicated in metastasis, CRC proliferation, and as a drug target ligand. Conclusion : A combination of 3D culture, gene profiling, and in silico analysis is feasible to detect CRC for Indonesia and other low- and middle-income countries. A future possibility is to use minicolorectal cancer in a dish for ex vivo cancer modeling and pharmacological testing.