Resident Michigan Medicine Ann Arbor, Michigan, United States
Disclosure: I do not have any relevant financial / non-financial relationships with any proprietary interests.
Participants should be aware of the following financial/non-financial relationships:
Katie K. Spielbauer, MD: I do not have any relevant financial / non-financial relationships with any proprietary interests.
Introduction: Targeted therapies for melanoma such as MEK and BRAF inhibitors eventually fail due to development of chemoresistance. Our studies have shown that C-terminal Hsp90 inhibitors(C-HSP90i) synergize with MEK and BRAF inhibitors in melanoma. Hence, we hypothesized that these C-HSP90i may also overcome therapy resistance by targeting the resistant pathway proteins of BRAF/MEK inhibitors.
Methods: Melanoma cell lines (ATCC) A375 (parent line) and A375R (MEK/BRAF inhibitor resistant daughter line) were utilized. Inhibitory concentrations (IC50) of two C-HSP90i’s (KU757 and KU758), as well as vemurafenib and cobimetinib were measured by Cell-titer Glo. RNA sequencing was performed after treatment of cells with KU757 at IC50. GSEA was utilized to identify targeted pathways. R2 was used to validate gene expression and evaluate survival advantages in the TCGA dataset.
Results: A375R was resistant to both BRAF and MEK inhibitors with a shift in the cobimetinib and vemurafenib IC50 from 3.3nM and 0.14uM in A375 to 109.5nm and 4.0uM in A375R respectively (p< 0.05). But, A375R had similar IC50 values as that of A375 for both of the C-terminal HSP90 inhibitors KU757 and KU758. IC50 for KU757 was 0.59uM vs 0.64uM and 0.89uM vs 0.93uM for KU758 in A375 vs A375R respectively. RNA seq revealed KU757 upregulates cell cycle checkpoint G2/M arrest, ferroptosis and the apoptotic pathway and down regulates genes in involved in AKT/PI3K/MTOR, EIF2, fatty acid metabolism and oxidative phosphorylation. Expression levels of the genes involved in these pathways were further validated in 468 patient samples from the TCGA database. Kaplan Meijer survival analysis of pathway genes modulated by KU757 demonstrated significant survival benefit (Bonferroni p< 0.05) in patients with lower expression of EIF2A, SDBH, VDAC2, LAMA1, COL17A1 that were downregulated by KU757 treatment (as shown in Figure 1).
Conclusions: C-terminal Hsp90 inhibition overcomes BRAF/MEK inhibitor resistant pathways of melanoma in vitro by down regulating oxidative phosphorylation, EIF2, and AKT/PI3K/MTOR pathway which may improve survival outcomes. Future in vivo validation will provide rationale for clinical translation.
Learning Objectives:
List modes of treatment failure in melanoma.
Describe novel therapeutic strategies to overcome resistance to targeted therapies.
Demonstrate understanding of publicly available data sets for validating RNA sequencing data.
