Fatty acid-binding protein-5 (FABP5) or epidermal FABP belongs to a low molecular weight lipid binding protein family. FABP5 is involved in binding, storing, and transporting hydrophobic ligands to the proper cellar compartment. Recent studies have suggested that FABP5 play important roles in regulation of gene expression associated with with cell growth and differentiation. FABP5 expression level was closely related to malignency in several types of cancers. The functions of FABP5 in modulation of cellular signaling have been studies and suggested that FABP5 is involved in EGFR, NFkB and PPAR pathways. Using IMTACTM (Isobaric Mass Tagged Affinity Characterization) to screen small molecule library in live cancer cells, ligands covalently bound to FABP5 were identified. Our initial characterization of compounds showed that they specifically bind to FABP5 and inhibit cancer cell growth. These FABP5-specific covalent small molecule ligands will provide a valuable pharmacological tool to dissect FABP5 functions and shed light on the therapeutic potential of targeting FABP5 for the treatment of cancer and metabolic disorders. Citation Format: Hang Chen, Heather Ha, Robert Stanley, Cindy Huang, Qian Cai, Irene Yuan, Ping Cao. Discovery of BGS1954, a covalent small molecule ligand of FABP5 using IMTACTM platform [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 A135. doi:10.1158/1535-7163.TARG-19-A135

Guanosine monophosphate synthetase (GMPS) is a glutamine amidotransferase catalyzing amination of xanthosine monophosphate (XMP) to form GMP. It is a enzyme involved in de novo purine biosynthesis. T and B lymphocytes, and certain cancer cells, such as malignant melanoma are dependent on de novo synthesis of purine nucleotide for activation and proliferation. GMPS is an attractive target for drug discovery and development to treat autoimmune diseases, organ transplant rejection, and cancers. Mycophenolate acid (MPA), an active ingredient of Mycophenolate Mofetil (Cellcept) has been used for organ transplantation and autoimmune diseases. MPA is a natural product which inhibits inosine monophosphate dehydrogenase (IMPDH), an enzyme upstream of GMPS. Using IMTACTM (Isobaric Mass Tagged Affinity Characterization) platform to perform global proteome ligand screen in live cancer cells, a highly potent and highly selective irreversible inhibitor against GMPS was discovered. Here we report the identification, in vitro characterization and in vivo evaluation of this molecule BGS2019. Target engagement of BGS2019 was determined by In-Gel-Fluorescent assay in cell with an EC50 of 15.6nM. It inhibited GMPS enzymatic activity with an IC50 of 6nM. The mechanism of BGS2019 in cell-based assays was tested, which showed the accumulation of IMP (the substrate for GMPS) and the reduction of GMP, GDP and GTP, upon BGS2019 treatment. BGS2019 was evaluated for its cell growth inhibition activity in several cancer cell lines and it suppressed cancer cell growth with IC50 values from 30nM to 100nM. In vivo administration of BGS2019 was well tolerated in mice and exhibited tumor suppression in animal model. Citation Format: Hang Chen, Heather Ha, Robert Stanley, Cindy Huang, Qian Cai, Irene Yuan, Ping Cao. Discovery of BGS2019, a highly potent and selective covalent inhibitor of GMPS using IMTACTM platform [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 B135. doi:10.1158/1535-7163.TARG-19-B135

There is currently a resurgence of interest in developing covalent drugs due to the high potency, good selectivity, and prolonged effects which may results in less-frequent drug dosing and wider therapeutic windows for patients. BridGene Biosciences established a novel IMTACTM (Isobaric Mass Tagged Affinity Characterization) platform for covalent drug discovery. Using IMTACTM platform, an intelligently designed drug-like probe with alkyne tag is able to “fish out” and identify proteins covalently bound to probes from live cells which enables us to discover druggable opportunities across the entire proteome. There are several advantages of IMTACTM : 1. Discover ligands for unliganded or undruggable targets; 2. Reduce the time and cost of lead discovery; 3. Identify additional targets of drugs or clinical candidates; 4. Build ligand-target interaction database and guide drug design. Using IMTACTM platform, our covalent library is screened in various live cells including cancer immune, and neuronal cell lines. The hit rate of IMTAC screening is about 20%, and the targets discovered by IMTACTM includes GMPS, Rictor, BTK, FABP5, ALDH, FUS, MIF and etc. Given the drug-like probe design, the probes can have nanomolar potency on the targets. And the drug like probe design minimizes the SAR efforts of developing a hit to a drug. IMTACTM platform allows us to expand the therapeutic applications of small molecule drugs to a new horizon. Citation Format: Heather Ha, Robert Stanley, Qian Cai, Wendy Zhou, Hang Chen, Irene Yuan, Ping Cao. Small molecule covalent drug discovery by IMTACTM [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 A136. doi:10.1158/1535-7163.TARG-19-A136

Hepatocellular carcinoma (HCC) accounts for 65% of all cases of liver cancers, ranking the 6th most common cancer and the 2nd cause of cancer mortality. HCC has more than 700,000 new cases/year and 600,000 deaths/year. Sorafenib is currently the standard of care for advanced disease, provides a response rate of ~2% and median survival <11 months. The fibroblast growth factor 19 (FGF19)-fibroblast growth factor receptor 4 (FGFR4)-Klotho β (KLB) signaling pathway regulates bile acid synthesis and is also a key driver in certain subtypes of HCC (around 30%). We generated BGS2219 and BGS2223 through FGFR4 structure-based design. BGS2219 and BGS223 are potent, selective and irreversible FGFR4 inhibitors. BGS2219 inhibits FGFR4 with an IC50 of 4nM and has greater than 2500 fold selectivity over FGFR1, 2 and 3. BG2219 took effects in both HCC and RMS tumor cell lines. In vivo administration of BG2219 was well tolerated in mice at all dosages with no body weight loss and and inhibited tumor growth in a dose -dependent fashion in HCC xenograft model. Complete tumor regression was observed at 40mg/kg. BGS2219 was tested in a combo therapy regimen with R1507 (anti-IGFR1 mab) in RMS (rhabdomyosarcoma) xenograft model in mice and showed that BG2219 plus R1507 significantly reduce the tumor volume in comparison with R1507 alone. BGS2223 is derived from BG2219, with improved oral bioavailability. BGS2223 has no hit in Cerep Screen safety panel and no activity detected in hERG assay at 10mM. In vivo oral administration of BG2223 in mice was well tolerated with no body weight loss and showed dose-dependent tumor growth inhibition in HCC CDX model at all dosages. Tumor regression was achieved at high dose treatment. BGS22 treatment in vivo in HCC tumor bearing mice caused alteration of CYP7A1 and FGF19 expression, which were utilized as biomarkers for FGFR4 inhibition. Citation Format: Hang Chen, Katherine Slemmons, Heather Ha, Robert Stanley, Qian Cai, Irene Yuan, Lee Helman, Ping Cao. Discovery of highly potent and selective covalent inhibitors of FGFR4 [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 C072. doi:10.1158/1535-7163.TARG-19-C072