Emely Morales Colon, MS
Lilia Kucheryavykh, PhD
Universidad Central del Caribe, Bayamón, Puerto Rico
Introduction
Tumor cell proliferation pathways denote a promising target for recurring glioblastoma. On glioblastoma (GBM) it is uncommon to view alterations on the Raf/MEK/ERK signaling pathway. Unfortunately, the inhibition of NF1 gene produces the expression of phosphorylated focal adhesion kinase (p-FAK) and leads to the activation of the Raf/MEK/ERK pathway, causing an uncontrolled cell division. Combinatorial targeting of FAK and RAF/MEK represents a promising opportunity for a therapeutic intervention in GBM. We hypothesize that combinatorial targeting of FAK and RAF/MEK will prevail the resistance to RAF/MEK inhibitors. The purpose of the study was to evaluate a novel MEK/ERK dual clamp inhibitor (Avutometinib) and a FAK inhibitor (Defactinib) on GBM cell proliferation and regulation.
Methods
Primary human glioblastoma cell lines were used. Dose response of Avutometinib (500nM, 5µM, 1µM) and Defactinib (13nM, 5µM, 10µM) were evaluated at 24 and 72 hours, compared to control. In addition, live and dead assay was performed. Western blot analysis of FAK, ERK, MEK, and cyclins was employed to divulge the inhibitory effect on cell proliferation and apoptosis.
Results
At a 72-hour treatment with both inhibitors, a reduction of 35% in cell viability was observed. The protein expression of p-ERK was reduced to 50% on both cell lines with the combinatorial treatment of Defactinib (13nM and 5µM) and Avutometinib (500nM and 1µM) compared to control. Flow cytometry data indicated a cell cycle arrest at G1. Cyclins protein expression demonstrated a 50% reduction with Avutometinib 1µM at 72 hours.
Conclusion
RAF/MEK/ERK signaling with combinatorial treatments overcomes GBM resistance to inhibition. The effective concentration that leads to more than 50% of protein expression was 5µM for Defactinib and 1µM for Avutometinib within a 72-hour treatment.
Funding
PRSTRT2022 and NIH Grant 1R15CA287203
IRB Protocol Number
2024-31
