Efficacy of a Covalent ERK1/2 Inhibitor, CC-90003, in KRAS Mutant Cancer Models Reveals Novel Mechanisms of Response and Resistance



Ida Aronchik1, Yumin Dai1, Matt Labenski2, Carmen Barnes2, Terri Jones2, Lixin QIao2, Lisa Beebe2, Mehnaz Malek1, Winfried Elis3, Tao Shi4, Konstantinos Mavrommatis1, Gordon L. Bray1 and Ellen H. Filvaroff1.


  1. Celgene Corporation, San Francisco, CA
  2. Celgene Corporation, Cambridge, MA
  3. Oncotest GMBH, Charles River Discovery, Freiburg,Germany
  4. Celgene Corporation, San Diego, CA


As a critical signaling node, ERK1/2 are attractive drug targets, particularly in tumors driven by activation of the MAPK pathway. Utility of targeting the MAPK pathway has been demonstrated by clinical responses to inhibitors of MEK1/2 or RAF kinases in some mutant BRAF-activated malignancies. Unlike tumors with mutations in BRAF, those with mutations in KRAS (>30% of all cancers and >90% of certain cancer types) are generally not responsive to inhibitors of MEK1/2 or RAF. Here, a covalent ERK1/2 inhibitor, CC-90003, was characterized and shown to be active in preclinical models of KRAS-mutant tumors. A unique occupancy assay was used to understand the mechanism of resistance in a KRAS-mutant patient-derived xenograft (PDX) model of colorectal cancer. Finally, combination of CC-90003 with docetaxel achieved full tumor regression and prevented tumor regrowth after cessation of treatment in a PDX model of lung cancer. This effect corresponded to changes in a stemness gene network, revealing a potential effect on tumor stem cell reprograming.

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Mass spectrometry assays of intact protein kinases were performed. The protein kinases were incubated for 1 hour at room temperature in an excess of CC-90003 in comparison to the quantity of protein. After incubation, the samples were each diluted with 10 μL of 0.2% TFA before being desalted using micro pipette tips that contain 0.6 μL of C4 resin fixed at the end and then spotted onto a MALDI target plate. Sinapic acid was used as the desorption matrix and analyzed on a MALDI TOF mass spectrometer modified with a CovalX HM2 detection system. The HM2 was used with the settings at HV1 – 2.7 kV, HV2 – 20 kV, 20 ns bin size and 6000 units of laser intensity at a constant. The use of a mass spectrometer modified with a CovalX HM2 system allowed researchers to confirm that a targeted cysteine covalently bound to CC-90003 (Cys183 of ERK1 and Cys164 of ERK2) was located in the A-loop of the ATP binding site. The analysis also helped to discover that CC-90003 is a strong inhibitor of kinase activities of ERK1 and ERK2 with IC50s in the 10-20 nM range as well as having good kinase selectivity.



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