Of the three RAS oncoproteins, only HRAS is delocalized and inactivated by farnesyltransferase inhibitors (FTI), an approach yet to be exploited clinically. In this study, we treat mice bearing Hras-driven poorly differentiated and anaplastic thyroid cancers ( ) with the FTI tipifarnib. Treatment caused sustained tumor regression and increased survival; however, early and late resistance was observed. Adaptive reactivation of RAS-MAPK signaling was abrogated by selective RTK (i.e., EGFR, FGFR) inhibitors, but responses were ineffective, whereas combination of tipifarnib with the MEK inhibitor AZD6244 improved outcomes. A subset of tumor-bearing mice treated with tipifarnib developed acquired resistance. Whole-exome sequencing of resistant tumors identified a nonsense mutation and an activating mutation in at high allelic frequency, supporting the on-target effects of the drug. Cell lines modified with these genetic lesions recapitulated tipifarnib resistance This study demonstrates the feasibility of targeting Ras membrane association in cancers and predicts combination therapies that confer additional benefit. Tipifarnib effectively inhibits oncogenic HRAS-driven tumorigenesis and abrogating adaptive signaling improves responses. NF1 and GNAS mutations drive acquired resistance to Hras inhibition, supporting the on-target effects of the drug. .

Of the three RAS oncoproteins, only HRAS is delocalized and functionally inactivated by farnesyltransferase inhibition (FTI), an approach that has yet to be exploited clinically. We treated a murine model of poorly differentiated and anaplastic thyroid cancer (Tpo-cre/HrasG12V/p53flox/flox; Hras;p53) with the FTI tipifarnib, and observed sustained tumor regression and increased survival; however, tumors eventually recurred. Following HRAS delocalization by tipifarnib in vitro, ERK phosphorylation was only inhibited transiently in HRAS-mutant cell lines, which was associated with increased GTP loading of wild-type RAS proteins in the setting of RTK ligand stimulation. This adaptive reactivation of RAS-MAPK signaling was abrogated by selective RTK (i.e. EGFR, FGFR) inhibitors, or by MEK inhibitors. Importantly, tipifarnib combined with the MEK inhibitor AZD6244 led to improved responses in Hras;p53 mouse tumors, whereas combination with the EGFR/FGFR inhibitors erlotinib and ponatinib did not, suggesting heterogeneity of upstream inputs. In order to identify acquired resistance mechanisms, tumor-bearing Hras;p53 mice were treated with tipifarnib until resistance developed (6 months). Whole exome sequencing of resistant tumors identified a truncating NF1 mutation and an activating mutation in GNAS at high allelic frequency. Upon NF1 knockdown in the human HRAS- mutant cell line C643, tipifarnib failed to inhibit pERK in vitro and caused resistance to tipifarnib in xenografts. By contrast, activating GNAS mutations transduced into Hras;p53 mouse tumor cell lines produced tipifarnib-resistant xenografts that activated the GNAS-cAMP-CREB pathway and demonstrated signs of redifferentiation. These data show that pharmacological targeting of RAS in a genetically accurate, mouse model of a RAS-driven virulent cancer leads to objective major responses and improved survival. We identified adaptive and acquired resistance mechanisms, and show that combined treatment with selective MEK inhibitors are beneficial. These data should also inform a currently enrolling clinical trial of tipifarnib for HRAS-mutant malignancies. Citation Format: Brian R. Untch, Vanessa Dos Anjos, Maria ER Garcia-Rendueles, Jeff A. Knauf, Alan L. Ho, James A. Fagin. The farnesyltransferase inhibitor tipifarnib causes dramatic tumor regression and increases survival in murine HrasG12V driven aggressive thyroid cancers: Consequent adaptive and acquired resistance mechanisms inform combination treatments with improved responses [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2063. doi:10.1158/1538-7445.AM2017-2063