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Supplemental Figure 1. Glesatinib is a potent and selective MET inhibitor. Supplemental Figure 2. Waterfall plot of glesatinib activity across pre-clinical tumor models. Supplemental Figure 3. SNU-638 Criz-res line is homogeneous and harbors the F1200L and Y1230H resistance mutations on the same allele. Supplemental Figure 4. Long term treatment of a METex14 del mutant and amplified PDX model (PULM-039) leads to capmatinib and crizotinib-resistant tumors but not glesatinib-resistant tumors. Supplemental Figure 5. Molecular basis for resistance that arises from the G1163R mutation. Supplemental Figure 6. Glesatinib is active against type I MET inhibitor-resistance mutations in a spheroid growth assay.

Purpose: MET exon 14 deletion (METex14 del) mutations represent a novel class of non–small cell lung cancer (NSCLC) driver mutations. We evaluated glesatinib, a spectrum-selective MET inhibitor exhibiting a type II binding mode, in METex14 del–positive nonclinical models and NSCLC patients and assessed its ability to overcome resistance to type I MET inhibitors.Experimental Design: As most MET inhibitors in clinical development bind the active site with a type I binding mode, we investigated mechanisms of acquired resistance to each MET inhibitor class utilizing in vitro and in vivo models and in glesatinib clinical trials.Results: Glesatinib inhibited MET signaling, demonstrated marked regression of METex14 del-driven patient-derived xenografts, and demonstrated a durable RECIST partial response in a METex14 del mutation-positive patient enrolled on a glesatinib clinical trial. Prolonged treatment of nonclinical models with selected MET inhibitors resulted in differences in resistance kinetics and mutations within the MET activation loop (i.e., D1228N, Y1230C/H) that conferred resistance to type I MET inhibitors, but remained sensitive to glesatinib. In vivo models exhibiting METex14 del/A-loop double mutations and resistance to type I inhibitors exhibited a marked response to glesatinib. Finally, a METex14 del mutation-positive NSCLC patient who responded to crizotinib but later relapsed, demonstrated a mixed response to glesatinib including reduction in size of a MET Y1230H mutation-positive liver metastasis and concurrent loss of detection of this mutation in plasma DNA.Conclusions: Together, these data demonstrate that glesatinib exhibits a distinct mechanism of target inhibition and can overcome resistance to type I MET inhibitors. Clin Cancer Res; 23(21); 6661–72. ©2017 AACR.

Cutaneous neurofibromas (CNF) are benign tumors that occur in the dermis of individuals with neurofibromatosis type 1 (NF), an inherited tumor predisposition syndrome. CNFs cause disfigurement, pain, burning, and itching, resulting in reduced quality of life in NF patients. However, due to their benign nature there are few in vitro or in vivo models of CNFs, which has limited the research of CNF biology and drug discovery efforts. To address this, we developed a patient derived explant (PDE) ex vivo culture model of CNF tumors and normal skin from NF patients. CNF PDEs remain viable in culture for over 9 days and recapitulate the cellular composition and molecular signaling of CNFs. We identified reciprocal inflammatory signaling in CNF PDEs, in which tumors rely on either prostaglandin or leukotriene mediated signaling pathways. Ex vivo glucocorticoid treatment reduced expression of pro-inflammatory genes, confirming CNF PDEs are a useful model for mechanistic studies and preclinical drug testing.

Malignant peripheral nerve sheath tumors (MPNSTs) are a rare and deadly sarcoma with few therapeutic options that are the leading cause of death for patients with Neurofibromatosis Type 1 (NF1). MPNSTs are characterized by a large burden of genomic alterations, chemoresistance, and a 5-year survival rate of 25-50%. NF1 is the major negative regulator of RAS, therefore a hallmark of MPNSTs is deregulated RAS/MAPK signaling. To date, no targeted therapies have been approved for MPNST treatment, highlighting the need for an understanding of adaptive signaling mechanisms that drive resistance. The HIPPO pathway is a key regulator of organ growth and cellular differentiation and is a central driver of resistance to RAS pathway inhibition in other cancers. The key HIPPO effector, YAP1, interacts with RAS and AKT signaling pathways, creating alternate routes for resistance in multiple cancers. In our studies, we identified strong YAP activation in response to MEK and AKT inhibition in MPNST cell lines. Since YAP acts as a transcriptional co-activator through interaction with TEAD transcription factors, we proposed that targeting the Hippo pathway in MPNST will abrogate MAPK inhibitor resistance. To investigate tumor signaling responses and efficacy in vivo, we utilized 3 genomically distinct MPNST patient-derived xenograft (PDX) models. To assess drivers of MPNST resistance, we have developed a preclinical model of drug resistance that simulates clinical treatment schedules. Using a cross-over and a drug holiday design, we are able to evaluate patterns of response and resistance to resumed treatment. We observed distinctive responses to MEK and AKT inhibitors in each PDX line; however, the impact on MPNST growth was minimal with single agent treatment. Comparison of pathway activation between the “drug holiday” biopsies and the treatment endpoint, demonstrated distinct YAP and RAS (pERK) activation levels in resistant tumors. For example, strong YAP activation is observed in correlation with decreased pERK in resistant tumors. YAP activation was strongest at the invasive edge of viable tumor regions. Currently, we are evaluating the efficacy and signaling adaptations to investigate the underlying molecular mechanisms in these MPNST models. Overall, we have demonstrated the development of a clinically relevant model of MPNST resistance and a potential switch between RAS and YAP signaling that promotes resistance to MEK or AKT inhibition. Citation Format: Lauren McGee, Curt Essenburg, Lisa Turner, Angela Hirbe, Anwesha Dey, Jason Zbieg, Carrie Graveel, Matt Steensma. YAP signaling promotes resistance to MEK and AKT inhibition in NF1-related MPNSTs [abstract]. In: Proceedings of the AACR Special Conference: Sarcomas; 2022 May 9-12; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(18_Suppl):Abstract nr A018.

Next-generation sequencing (NGS) capabilities can affect therapeutic decisions in patients with complex, advanced, or refractory cancer. We report the feasibility of a tumor sequencing advisory board at a regional cancer center. Specimens were analyzed for approximately 2800 mutations in 50 genes. Outcomes of interest included tumor sequencing advisory board function and processes, timely discussion of results, and proportion of reports having potentially actionable mutations. NGS results were successfully generated for 15 patients, with median time from tissue processing to reporting of 11.6 days (range, 5 to 21 days), and presented at a biweekly multidisciplinary tumor sequencing advisory board. Attendance averaged 19 participants (range, 12 to 24) at 20 days after patient enrollment (range, 10 to 30 days). Twenty-seven (range, 1 to 4 per patient) potentially actionable mutations were detected in 11 of 15 patients: TP53 (n = 6), KRAS (n = 4), MET (n = 3), APC (n = 3), CDKN2A (n = 2), PTEN (n = 2), PIK3CA, FLT3, NRAS, VHL, BRAF, SMAD4, and ATM. The Hotspot Panel is now offered as a clinically available test at our institution. NGS results can be obtained by in-house high-throughput sequencing and reviewed in a multidisciplinary tumor sequencing advisory board in a clinically relevant manner. The essential components of a center for personalized cancer care can support clinical decisions outside the university.