Characterising Drug Resistance in the Colorectal Cancer Cell Line RKO

Colorectal cancer (CRC) is the second largest cause of cancer-related death in the UK and resistance formation to treatment is inevitable. In vitro models of acquired resistance can be used to characterise drug resistance. RKO, a CRC cell line, was used in this thesis along with three drug-adapted sublines made resistant to the most common chemotherapeutic agents used to treat CRC: 5-Fluorouracil, Irinotecan and Oxaliplatin. RKO and the drug-adapted sublines were characterised by their doubling time via the use of growth curve assays using MTT reagent to determine their rate of growth. They were further characterised by the establishment of an IC50 using the compounds 5-FU, Irinotecan, Oxaliplatin, Miyabeacin, I-BET, JQ1, Rucaparib and Olaparib in a cell viability assay, also using MTT. Verapamil was added in combination with the last five of the compounds mentioned. The doubling time of RKO was 14.5 ± 0.4 hrs, RKOr5-FU was 18.0 ± 1.6 hrs, RKOrIRINO was 24.0 ± 4.7 hrs and RKOrOXALI was 22.8 ± 4.5 hrs. A resistance factor (RF) which states that a greater than two-fold increase in IC50 concentration means a drug-adapted subline has become resistant to a given compound. 5-FU and Irinotecan were determined to have an RF ≥ 2.0 in RKOr5-FU. 5-FU, Irinotecan, I-BET, JQ1 and Rucaparib had an RF ≥ 2.0 in RKOrIRINO. Oxaliplatin, I-BET and JQ1 had an RF ≥ 2.0 in RKOrOXALI. The addition of Verapamil sensitised RKOrIRINO to JQ1, lowering the RF below 2.0. An IC50 for Olaparib could not be determined in RKOr5-FU and RKOrIRINO. In conclusion, this thesis characterised and introduced novel acquired resistance models for the colorectal cancer cell line RKO.