Lucian Dipietro

Discovery of a potent, metabolically stabilized resorcylic lactone as an anti-inflammatory lead

With bioactivity-guided phenotype screenings, a potent anti-inflammatory compound f152A1 has been isolated, characterized and identified as the known natural product LL-Z1640-2. Metabolic instability precluded its use for the study on animal disease models. Via total synthesis, a potent, metabolically stabilized analog ER-803064 has been created; addition of the (S)-Me group at C4 onto f152A1 has resulted in a dramatic improvement on its metabolic stability, while preserving the anti-inflammatory activities.

Abstract 2641: The development of potent, selective RET inhibitors that target both wild-type RET and prospectively identified resistance mutations to multi-kinase inhibitors

Introduction: RET/PTC1 was one of the first gene fusions identified from a solid tumor. In the past 3 years, oncogenic RET fusions have been identified in additional cancer types, most notably NSCLC and colon carcinomas. Activating germline RET mutations are also well established drivers of multiple endocrine neoplasia while somatic RET mutations are the most prevalent alterations in sporadic medullary thyroid cancers. In light of these findings, a number of approved multi-kinase inhibitors (mKIs) with in vitro activity against wild-type (WT) RET, such as cabozantinib, vandetanib, and lenvatinib, have been repurposed for treating RET-driven diseases. We have designed a next-generation inhibitor specifically tailored to target RET, while sparing other closely-related kinases, such as KDR/VEGFR2. Given that secondary mutations are a common resistance mechanism to kinase inhibitors, we prospectively identified resistance mutations that abrogate mKI activity and crafted our RET-selective inhibitors to also target these mutations. The structures of several mKIs bound to RET were analyzed and amino acid substitutions that would disrupt the protein-inhibitor interactions were predicted. In vitro resistance screens with cabozantinib, ponatinib, and vandetanib in a Ba/F3 KIF5B-RET cell line were conducted and confirmed these predictions. The proprietary Blueprint Medicines’ kinase inhibitor library was used to identify inhibitors of WT and resistance mutant RET as starting points for lead optimization. Results: Both structural analysis and in vitro screening revealed that only a handful of positions within the RET kinase domain enable resistance mutations to mKIs, suggesting a narrow mutational spectrum. Using our library of kinase inhibitors, we identified potent, orally bioavailable inhibitors of RET that target both the WT and resistance mutants in KIF5B-RET-driven cell lines while sparing the majority of the kinome. These inhibitors also suppressed the proliferation of thyroid cancer cell lines harboring RET fusions or activating RET mutations and demonstrated in vivo activity in xenograft models. Our analysis of the PK-PD-efficacy relationship revealed that over 70% target suppression is required for maximal efficacy. Finally, our RET-selective inhibitors induced dose-dependent tumor growth inhibition in a KIF5B-RET fusion positive lung adenocarcinoma PDX model at well-tolerated doses, further validating RET fusions as oncogenic drivers in NSCLC. Conclusion: This work describes the identification of potent inhibitors that specifically target WT RET and resistance mutations predicted to arise upon mKI treatment. By sparing kinases with known toxicity profiles, these molecules are predicted to robustly inhibit RET at tolerated doses and may provide patients with RET-driven diseases an opportunity for more durable and effective therapies. Citation Format: Rami Rahal, Erica K. Evans, Wei Hu, Michelle Maynard, Paul Fleming, Lucian DiPietro, Joseph L. Kim, Michael P. Sheets, Doug P. Wilson, Kevin J. Wilson, Nicolas Stransky, Jason D. Brubaker, Timothy Guzi, Nancy E. Kohl, Christoph Lengauer. The development of potent, selective RET inhibitors that target both wild-type RET and prospectively identified resistance mutations to multi-kinase inhibitors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2641.

Abstract LB-324: First isoform selective inhibitor of FGFR4 for the treatment of genomically defined patients with hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is the third leading cause of cancer deaths worldwide. Chemotherapy has proven ineffective, and Sorafenib remains the only approved targeted drug with no second or third line treatment options. Sorafenib slows the growth of advanced liver cancers and helps some patients live longer - by an average of about three months. There is a pressing need for more effective therapies. FGF19 is a highly controlled hormone normally expressed in the intestine, that acts in the liver to regulate bile acid synthesis and hepatocyte proliferation via activation of FGFR4. In 7% of patients with HCC, FGF19 is contained within a focal amplification on chromosome 11q13.3. Overexpression of FGF19 in transgenic mice produces liver tumors which are sensitive to treatment with a FGFR4 tool antibody. Additionally, the growth of tumor cells in xenograft models with FGF19 amplification is dependent on FGFR4 signaling. Thus, selective inhibition of FGFR4 might represent a viable strategy for treating this genetically defined subgroup of HCC patients. Herein, we describe our efforts to identify an ultraselective small molecule inhibitor of FGFR4 which spares the other FGFR isoforms with the intent to avoid FGFR1-3 driven, dose limiting toxicities like soft tissue mineralization. Utilizing structure based drug design, we prepared a series of inhibitor templates designed to covalently modify a target cysteine residue present in FGFR4, but not the other FGFR isoforms. Optimization of one of these templates led to the identification of BLU9931, a highly potent and exquisitely selective, covalent inhibitor of FGFR4. BLU9931 persistently inhibits FGFR4 mediated signaling in cancer cells as evidenced by decreased phosphorylation of FRS2 and ERK. Importantly, BLU9931 does not block signaling driven by FGFR1. Upon oral dosing in mice, BLU9931 is well tolerated and demonstrates robust and dose dependent induction of the FGFR4 target gene CYP7a1 in Hep3B cells, a FGF19 amplified HCC xenograft model. Upon extended dosing, BLU9931 causes sustained regression of tumors, including complete responses. We then explored if HCCs with alterations other than FGF19 amplification are also dependent on FGFR4 signaling. Dosing of molecularly annotated patient derived HCC xenografts with BLU9931 suggests that selective targeting of FGFR4 represents a viable option for the treatment of an expanded population segment of genomically defined HCC patients, much larger than originally anticipated. Citation Format: Margit Hagel, Chandra Miduturu, Mike Sheets, Weifan Weng, Nooreen Rubin, Neil Bifulco, Lucian DiPietro, Joseph Kim, Natasja Brooijmans, Nicolas Stransky, Christopher Winter, Christoph Lengauer, Timothy Guzi. First isoform selective inhibitor of FGFR4 for the treatment of genomically defined patients with hepatocellular carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-324. doi:10.1158/1538-7445.AM2014-LB-324