Novel drug development for KRAS-mutated non-small cell lung cancer
Shakil, Md Salman
Lung cancer is the leading cause of cancer-related death among both men and women and non-small cell lung cancer (NSCLC) accounts for 85 to 88% of all lung cancer cases. In NSCLC, kirsten rat sarcoma (KRAS) viral oncogene homologue mutations are the third most frequent mutation. KRAS-mutant patients show a shorter overall survival as a clinically approved therapeutic agent against KRAS has yet to be developed. Therefore, novel drugs are urgently needed for these patients. The current project examined the cytotoxic effects, selectivity, and anticancer mechanism of novel synthetic compounds towards KRAS-mutated NSCLC in vitro. In this study, A549 (KRAS-mutated NSCLC cells), H522 (p53-mutated NSCLC cells), NIH-3T3 (pre-neoplastic) and PNT1A (normal prostate epithelial cells) cell lines were used to screen out potent drug candidate for KRAS-mutated NSCLC. A549 cells were treated with 11 different metal-based and non-metal-based novel compounds of four different drug class (HDAC inhibitors: JAZZ-90, JAZZ-166, and JAZZ-167; Hydroxythiopyridone derivatives: M1S, M2S, M1S-Ru, and M2S-Ru; Metal-based PCA ligands and complexes: AASH-122 and JAZZ-121; Kinetically inert metal(arene) complexes of PCA: ZR-012 and ZR-014) at 0.015 to 200 µM for 72 h and cell viability was determined using the sulforhodamine B assay. Hydroxythiopyridone derivatives M1S and M2S were the smallest but most potent compounds in A549 cells with EC50 values of 0.36 and 0.32 µM, respectively. Additionally, M1S and M2S were 1.3 and 1.4 times more potent against H522 cells. Furthermore, M1S and M2S exhibited more selectivity towards these NSCLC cells and pre-neoplastic NIH3T3 cells as compared to PNT1A cells. Time-course cytotoxicity studies showed that both the drug candidates had cytostatic and cytotoxic effects at 2×EC50 concentration in A549 and H522 cells, respectively. Western blotting results indicated that the drug candidates were unable to increase the acetylation of histone 3. In A549 cells the drug candidates did not change cyclin D1 levels, after 24 h. However, 2×EC50 concentration of M1S and M2S significantly decreased cyclin D1 expression in H522 cells by 68.1 and 84.9% of control, respectively at 24 h. In both the cell lines, the drug candidates did not produce a significant effect in reducing cyclin B1 expression. Cell cycle analysis showed that the 2×EC50 concentration of M1S and M2S arrested 3.3 and 7.1% of A549 cells at the G2/M phase compared to control at 12 h. While 9.9 and 8.9% of H522 cells were arrested at the G1 phase after M1S and M2S treatment at 2×EC50 concentration. The same treatment also increased the number of sub-G1 apoptotic H522 cells. Thus, our findings provide evidence that M1S and M2S have potent anticancer activity in KRAS-mutated A549 cells and p53 mutated H522 cells, both the compounds warrant further in vitro experimentation to elucidate their specific mechanism of action(s).
Advisor: Rosengren, Rhonda J.
Degree Name: Master of Medical Science
Degree Discipline: Pharmacology & Toxicology
Publisher: University of Otago
Keywords: Novel drug; KRAS; NSCLC; Hydroxythiopyridone derivatives
Research Type: Thesis