Despite decades of research, efforts to directly target KRAS have been challenging. MRTX849 was identifi ed as a potent, selective, and covalent KRAS G12C inhibitor that exhibits favorable drug-like properties, selectively modifi es mutant cysteine 12 in GDPbound KRAS G12C , and inhibits KRAS-dependent signaling. MRTX849 demonstrated pronounced tumor regression in 17 of 26 (65%) KRAS G12C -positive cell line-and patient-derived xenograft models from multiple tumor types, and objective responses have been observed in patients with KRAS G12C -positive lung and colon adenocarcinomas. Comprehensive pharmacodynamic and pharmacogenomic profi ling in sensitive and partially resistant nonclinical models identifi ed mechanisms implicated in limiting antitumor activity including KRAS nucleotide cycling and pathways that induce feedback reactivation and/or bypass KRAS dependence. These factors included activation of receptor tyrosine kinases (RTK), bypass of KRAS dependence, and genetic dysregulation of cell cycle. Combinations of MRTX849 with agents that target RTKs, mTOR, or cell cycle demonstrated enhanced response and marked tumor regression in several tumor models, including MRTX849-refractory models. SIGNIFICANCE :The discovery of MRTX849 provides a long-awaited opportunity to selectively target KRAS G12C in patients. The in-depth characterization of MRTX849 activity, elucidation of response and resistance mechanisms, and identifi cation of effective combinations provide new insight toward KRAS dependence and the rational development of this class of agents.

KRASG12D, the most common oncogenic KRAS mutation, is a promising target for the treatment of solid tumors. However, when compared to KRASG12C, selective inhibition of KRASG12D presents a significant challenge due to the requirement of inhibitors to bind KRASG12D with high enough affinity to obviate the need for covalent interactions with the mutant KRAS protein. Here, we report the discovery and characterization of the first noncovalent, potent, and selective KRASG12D inhibitor, MRTX1133, which was discovered through an extensive structure-based activity improvement and shown to be efficacious in a KRASG12D mutant xenograft mouse tumor model.

The ability to effectively target mutated KRAS has remained elusive despite decades of research. The recent identification of KRAS G12C inhibitors has provided an effective treatment option for patients harboring this particular mutation and has also provided insight toward targeting other KRAS mutants, including KRAS G12D . MRTX1133 was identified via a structure-based drug design (SBDD) strategy as a potent, selective, and non-covalent KRAS G12D inhibitor directed at the switch II binding pocket. MRTX1133 demonstrated a high-affinity interaction with KRAS G12D with KD or IC50 values each determined at ~0.2 pM or <2 nM using SPR direct binding or HTRF competition assays, respectively. MRTX1133 also demonstrated ~700-fold selectivity for KRAS G12D vs KRAS WT binding utilizing SPR. Interestingly, MRTX1133 demonstrated potent inhibition of active KRAS G12D using an HTRF effector interaction assay with a IC50 value of 9 nM. Insight toward the structural basis of binding of MRTX1133 to both the inactive GDP-bound and active GMPPCP-bound conformations of KRAS G12D is also provided by co-crystal structures. MRTX1133 demonstrated potent inhibition of ERK1/2 phosphorylation and cell viability in KRAS G12D -mutant cell lines with median IC50 values of ~5 nM. Consistent with binding affinity determination in cell-free systems, MRTX1133 demonstrated >1000-fold selectivity for inhibition of ERK1/2 phosphorylation in KRAS G12Dmutant cell lines compared with KRAS WT cell lines. Dose-dependent inhibition of KRASmediated signal transduction was also observed in multiple KRAS G12D -mutant tumor models in vivo. MRTX1133 demonstrated marked tumor regression (>30%) in a subset of KRAS G12Dmutant cell line-and patient-derived xenograft (PDX) models, including 8 out of 11 (73%) pancreatic ductal adenocarcinoma (PDAC) models evaluated. Pharmacological studies and CRISPR-based screens demonstrated co-targeting KRAS G12D in concert with putative feedback or bypass pathways including EGFR and PI3Kα led to enhanced anti-tumor activity relative to targeting each individual protein. Together, these data indicate the feasibility of utilizing SBDD approaches to selectively target alternative KRAS mutant variants with non-covalent, highaffinity small molecules targeting either the active or inactive state of KRAS. In addition, these data illustrate the therapeutic susceptibility and broad dependence of KRAS G12D mutationpositive tumors, including PDAC, on KRAS for tumor cell growth and survival. SignificanceThe development of clinically active KRAS G12C -selective inhibitors represents a milestone achievement for the treatment of cancer; however, the discovery of additional KRAS-mutant selective inhibitors has remained elusive. MRTX1133 is a potent KRAS G12D -selective small molecule inhibitor, is active in vitro and in vivo, induces regression in multiple xenograft tumor models and demonstrates increased anti-tumor activity in rationally designed combinations. These data confirm KRAS G12D functions as an oncogenic driver, including in pancreat...

SOS1 is one of the major guanine nucleotide exchange factors that regulates the ability of KRAS to cycle through its “on” and “off” states. Disrupting the SOS1:KRAS G12C protein–protein interaction (PPI) can increase the proportion of GDP-loaded KRAS G12C , providing a strong mechanistic rationale for combining inhibitors of the SOS1:KRAS complex with inhibitors like MRTX849 that target GDP-loaded KRAS G12C . In this report, we detail the design and discovery of MRTX0902—a potent, selective, brain-penetrant, and orally bioavailable SOS1 binder that disrupts the SOS1:KRAS G12C PPI. Oral administration of MRTX0902 in combination with MRTX849 results in a significant increase in antitumor activity relative to that of either single agent, including tumor regressions in a subset of animals in the MIA PaCa-2 tumor mouse xenograft model.

The ability to effectively target mutated KRAS has remained elusive despite decades of research. MRTX849 was identified via structure-based drug design as a potent, selective, and covalent KRASG12C inhibitor that exhibits favorable drug-like properties. MRTX849 is presently under evaluation in clinical trials and its discovery and evaluation is disclosed here for the first time. MRTX849 demonstrated selective modification of the mutant cysteine residue at amino acid residue 12 in GDP-bound KRASG12C and inhibited KRAS-dependent signaling in vitro and in vivo. In multiple KRASG12C positive cell lines in vitro, MRTX849 treatment demonstrated covalent modification of mutant KRAS and/or inhibition of active KRAS at concentrations as low as 2 nM with near maximal inhibition observed at 15 nM. In vivo, clear evidence of dose-dependent modification of KRASG12C and inhibition of KRAS-dependent signal transduction was observed in multiple KRASG12C mutant tumor models. MRTX849 demonstrated marked tumor regression in 17 out of 26 (65%) of KRASG12C-positive cell line- and patient-derived xenograft (PDX) models but not in tumor models without KRASG12C mutations. While MRTX849 commonly elicited tumor regression, a subset of models was less sensitive to treatment or exhibited tumor stasis after an initial response phase. Comprehensive pharmacodynamic and pharmacogenomic profiling in these models identified mechanisms implicated in limiting anti-tumor response in refractory models including extrinsic factors impacting KRAS nucleotide cycling and/or signaling pathways that induce feedback reactivation and/or bypass KRAS dependence. These factors included upstream activation of ERBB family receptor tyrosine kinases (RTKs) as well as genetic dysregulation of cell cycle transition genes. Based on these observations, combination strategies designed to co-target signaling feedback and bypass pathways were evaluated in multiple tumor models. Afatinib and the SHP2 inhibitor, RMC-4550, in combination with MRTX849 validated these therapeutic hypotheses and the combinations demonstrated marked tumor regression in several tumor models, including models that were refractory to either single agent. MRTX849 in combination with the CDK4/6 inhibitor palbociclib also demonstrated a marked antitumor response; notably in tumor models harboring CDKN2A deletion or other genetic alterations implicated in cell cycle dysregulation. Together, these data indicate the therapeutic susceptibility and broad dependence of KRAS G12C mutation-positive tumors on KRAS for tumor cell growth and survival and provide insight toward the molecular basis of response to single agent and combinatorial therapies. Citation Format: James G Christensen, Jay B Fell, Jill Hallin, Brian Baer, Lars engstrom, James Blake, David Briere, Josh Ballard, Michael Burkhard, John Fischer, Guy Vigers, Ruth Aranda, Vickie Bowcut, Andrew Calinisan, Lauren Hargis, Niranjan Sudhakar, Matt Marx, Peter Olson. The identification of MRTX849, a novel KRASG12C inhibitor under clinical investigation, provides insight toward therapeutic susceptibility of KRAS mutant cancers [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr C069. doi:10.1158/1535-7163.TARG-19-C069