Inhibiting Oncogenic RAS in Multiple Myeloma By Targeting Scaffold-ERK Interactions
Authors: Gocke Christian B. , Wang Qiuju , McMillan Ross , Penchev Vesselin , Sage Julien , Khavari Paul A , Matsui William 
Published: October 2019

Abstract
The introduction of novel therapeutic agents over the past decade has significantly improved the outcomes of patients with newly diagnosed multiple myeloma (MM). Despite these advances, virtually all patients relapse and develop drug-resistant disease that carries a dismal prognosis. In addition to chemoresistance, relapse and tumor regrowth are dependent on self-renewal to maintain clonogenic growth potential over time. A better understanding of the factors responsible for tumor self-renewal and regrowth may lead to novel therapies and improved survival rates. RAS mutations are common in MM and have been demonstrated to increase in frequency with relapse and disease progression. These findings implicate RAS in chemoresistance, self-renewal, and tumor regrowth upon relapse. We have begun to directly examine the functional role of mutant RAS signaling in MM by stably transducing KRASmut MM cells with a tetracycline-inducible shRNA and found that KRAS knockdown decreases MM clonogenic growth in vitro. Although inhibiting oncogenic RAS may be beneficial for treating relapsed disease it remains an elusive drug target. Recently, our collaborators have developed a novel small peptide RAS signaling inhibitor that mimics the WW domain of IQGAP1 which competes for ERK binding to scaffolds. The WW peptide specifically targets solid tumors containing RAS (or BRAF) mutations and is well tolerated in mouse models. Thus, by targeting only tumorigenic cells it may demonstrate decreased toxicity compared to existing small molecule inhibitors of MAPK signaling.
We analyzed gene expression profiling data from the University of Arkansas (GSE2658) and found that elevated IQGAP1 expression levels (top 12.5%) are associated with increased disease-related mortality in MM (log-rank p = 0.0002, HR 4.3, 95% CI 2.0-9.1). This suggests that IQGAP1 may play an important biological role in MM disease progression and that inhibition of IQGAP1 and RAS signaling with the WW peptide may have efficacy and specificity in MM patients. We have treated RAS mutated MM cell lines with the WW peptide or a scrambled (SCR) control peptide and found that the WW peptide effectively decreases pERK levels after 24 hours at doses as low as 20 nM. The WW peptide also decreases cell growth in RAS mutated but not wild-type MM cells by inducing a cell cycle arrest (by BrdU/7-AAD staining). Importantly, treatment of RAS mutated MM cell lines with the WW peptide significantly decreased colony formation in vitro and led to a reduction in the frequency of highly clonogenic CD138neg MM cells. Finally, an immunodeficient mouse xenograft model was used to determine the effect of the WW peptide on engraftment potential. We treated NCI-H929NRAS(G13D) cells with SCR or WW peptide and injected them into NOD/SCID/IL2γchainKO (NSG) mice. Engraftment as detected by kappa light chain ELISA was significantly reduced in the WW peptide treated group. These data further implicate RAS signaling as well as MAPK signaling in MM self-renewal and identify a potential novel therapy for treatment of relapsed/refractory MM.

Disclosures
No relevant conflicts of interest to declare.