Treatment of a Black American lung cancer cell line harboring KRAS mutations with polyisoprenylated cysteinyl amide inhibitors reveals effects on the MAP kinase signaling pathway, cell migration and apoptosis.

Abstract KRAS mutations are the most common oncogenic mutations in lung adenocarcinoma in Black Americans. Polyisoprenylated cysteinyl amide inhibitors (PCAIs) constitute a group of cancer therapy agents designed to specifically disrupt and suppress hyperactive G protein signaling, such as that triggered by RAS mutations. Here we determine the effects of PCAIs on the viability, G-proteins levels, downstream mediators, and apoptosis-related proteins on the KRAS-mutant, Black American-derived lung adenocarcinoma cell line, NCI-H23. Of the 17 PCAIs tested, compounds NSL-YHJ-2-27 and NSL-YHJ-2-46 showed the most potency with EC50 values of 2.7 and 3.3 µM, respectively. To determine the effect of the PCAIs on the levels of Mitogen-Activated Pathway Kinase (MAPK)-related enzymes, western blot analysis was performed. After 48 hours exposure to 5 μM of the compounds, 57 to 150% increases of BRAF, ERK1/2, MEK1/2, and p90RSK phosphorylation along with a 60 to 78% decrease of pCRAF were observed, indicating significant disruptions in RAS signaling. Furthermore, 5 μM of NSL-YHJ-2-27 depleted the singly polyisoprenylated monomeric G-proteins RAC 1/2/3 and CDC42 by 77 and 76%, respectively. The depletion of these key cytoskeletal proteins may account for the observed 52 and 42% reduction in cell migration after 24 h exposure to NSL-YHJ-2-27 and NSL-YHJ-2-46, respectively. Western blot analysis was used to determine the effect PCAIs on caspase activation necessary for the initiation of apoptosis. After treatment with 5 μM of NSL-YHJ-2-27, full-length inactive caspase 3 and 7 levels dropped by 72 and 60%, while cleaved active caspase 3 and 7 levels increased by 274 and 130%, respectively. Treatment with 5 μM of NSL-YHJ-2-46 depleted full-length caspase 7 by 53% and increased cleaved active caspase 3 levels by 83%. These findings clearly show the direct effects of the PCAIs on the RAS signaling pathway, perhaps through the activation of proapoptotic isoforms of p90RSK. These results support the potential use of the PCAIs to serve as targeted therapies in cancers harboring RAS mutations. Citation Format: Matthew D. Gregory, Pablo E. Puente, Nadine L. Belony, Jassy Mary S. Lazarte, Nada Tawfeeq, Yong Huang, Ite A. Offringa, Nazarius S. Lamango. Treatment of a Black American lung cancer cell line harboring KRAS mutations with polyisoprenylated cysteinyl amide inhibitors reveals effects on the MAP kinase signaling pathway, cell migration and apoptosis [abstract]. In: Proceedings of the 15th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2022 Sep 16-19; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2022;31(1 Suppl):Abstract nr C022.