Abstract 2283: Molecular signaling in multiple myeloma: association of RAS/RAF mutation status and MAPK pathway activation in primary myeloma patient biopsies
Authors: Xu Jing , Pfarr Nicole , Endris Volker , Mai Elias K. , Md Hanafiah Nur Hafzan , Lehners Nicola , Penzel Roland , Weichert Wilko , Ho Anthony D. , Schirmacher Peter , Goldschmidt Hartmut , Andrulis Mindaugas , Raab Marc S. 
Published: July 2016

Abstract
Multiple myeloma (MM) is an incurable plasma cell malignancy. Recent studies have identified a dominant mutation cluster in RAS/BRAF genes, highlighting the MEK/ERK signaling pathway as a potentially promising therapeutic target. However, treatment of RAS/RAF mutant MM with the MEK inhibitor, trametinib, resulted in moderate response rates. There were, however, durable remissions in those patients (pts) who responded. This suggests a variable degree of dependency on MEK/ERK signaling in RAS/RAF mutant MM. To address this issue, we correlated RAS/RAF mutations with the RAF/MEK/ERK cascade activation in primary MM patient biopsies.
Primary material consisted of bone marrow or soft tissue biopsies from 179 pts, including 102 pts with newly diagnosed myeloma (NDMM) and 77 pts with relapsed/refractory myeloma (rrMM). Activation of the RAF/MEK/ERK cascade was assessed using an immunohistochemical (IHC) assay for phospho-ERK. In parallel, KRAS, NRAS, HRAS and BRAF mutation status was assessed by targeted re-sequencing using the Ion Torrent NGS technology.
The mutation frequencies found were KRAS (25%), NRAS (24%), and BRAF (8%) while no HRAS was detected, consistent with previous studies. Overall, RAS/RAF mutations were significantly more frequent in rrMM (p = 0.010), mainly driven by an increase in NRAS mutations (p = 0.017), proving the importance of RAS/RAF-mediated signaling in MM and indicating a potential role for NRAS mutations in drug resistance development. The top nine recurrently detected individual mutations were KRAS Q61H (n = 11), NRAS Q61R (11), NRAS Q61K (10), KRAS G12D/G12V (6 each), BRAF V600E (6), NRAS G13D (5), NRAS G13R/Q61H (4 each).
When correlated with ERK activation status, KRAS but not NRAS mutations were associated with pathway activation compared to RAS/RAF-wild type (wt) (p = 0.003). However, these were not mutually exclusive, suggesting that ERK activation is dependent on the type of mutation rather than the fact that the RAS/RAF gene is mutated. Therefore, each recurrent RAS/RAF mutation was tested against all RAS/RAF-wt samples: Only KRAS G12D and BRAF V600E were consistently associated with ERK activation (p&lt; 0.001 each).
In summary, we found that RAS/RAF mutations are not generally associated with RAF/MEK/ERK pathway activation in MM. However, we did identify specific amino acid changes in KRAS and BRAF which are more likely to be associated with pathway activation and two mutations which were consistently associated with MEK/ERK signaling. These findings indicate that confirmation of protein-level pathway activation is needed when considering targeted therapy. In this regard, IHC is a cost-effective and reliable method to assess pathway activation and should therefore be considered in future. The clinical relevance of RAS/RAF mutations in MM should be further elucidated in prospective cohort studies.
Citation Format: Jing Xu, Nicole Pfarr, Volker Endris, Elias K. Mai, Nur Hafzan Md Hanafiah, Nicola Lehners, Roland Penzel, Wilko Weichert, Anthony D. Ho, Peter Schirmacher, Hartmut Goldschmidt, Mindaugas Andrulis, Marc S. Raab. Molecular signaling in multiple myeloma: association of RAS/RAF mutation status and MAPK pathway activation in primary myeloma patient biopsies. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2283.